Hematology Mod 11, 12, 13, 14

Define the following terms: primary hemostasis, secondary hemostasis, endomitosis, demarcation membrane system

(Cont.) The normal reference range for platelets is 150 to 400 times 10 to the ninth per liter of blood. However, of the platelets that are released into the peripheral blood stream, only two thirds are actively circulating at any one time. The other one third is sequestered in the spleen red pulp. Both pools of platelets are in constant equilibrium, with sequestered platelets being released and circulating platelets entering the spleen for short-term storage.

Describe the three principal mechanisms by which secondary hemostasis is inhibited

(cont.) Anti-thrombin is a protein produced by the liver. Anti-thrombin binds to endothelial cell surfaces via interaction with heparin sulfate proteoglycans (HSPGs). Heparin sulfate is a complex, multi-ringed, organic polymer with associated sulfate moieties. Anti-thrombin subsequently inactivates thrombin (Factor 2a),as well as factor 10a of the common pathway, and factor 9a of the intrinsic pathway. Note that heparin is used as an anti-coagulant drug for treatment of thrombosis. In the proteinC/protein S system, thrombin itself is involved in this negative feedback inhibitory mechanism. Thrombin (2a) binds to an integral, membrane-bound protein called thrombomodulin, TM. And in turn, thrombin is converted from a pro-coagulant into an anti-coagulant factor. When bound to thrombomodulin, thrombin promotes the activation of protein C (PC) to become activated protein C (APC). Inactive protein C binds to endothelial protein C receptor (EPCR). Following thrombin activation, activated protein C is released from EPCR and complexes with protein S. Next, the APC/PS complex inactivates factors 5a and 8a. Recall that factor 5a is the cofactor for factor 10a of the common pathway; and that 8a is the cofactor for 9a of the intrinsic pathway. Tissue factor pathway inhibitor (TFPI) is a protein synthesized by endothelial cells, megakaryocytes, and smooth muscle cells; and it is released from platelet alpha granules during platelet activation. TFPI initially binds10a, and there by inhibits the common pathway. Subsequently, the TPFI/10a complex associates with membranes of cells in the sub-endothelium. TFPI then binds and inactivates the factor 7a/Tissue factor complex, and thereby inhibits the extrinsic pathway.

Bleeding Disorders List various types of anatomical bleeding abnormalities that occur in bleeding disorders

Abnormal bleeding and bruising are classic symptoms of primary or secondary hemostasis disorders. Primary disorders can be due to underlying vascular or platelet abnormalities; and secondary disorders can be due to coagulation factor or fibrinolytic factor defects. Bleeding disorders may beq uantitative or qualitative in nature; and they may b eeither inherited or acquired. Abnormal bleeding characteristics may manifest as prolonged bleeding from wounds, as well as other diverse anatomical bleeding abnormalities. In these disorders, excessive and prolonged bleeding may include: - gastrointestinal bleeding - hematuria (urine in the blood) - hypermenorrhea (heavy menstruation) - gingival bleeding (gum bleeding) - epistaxis (nose bleeds) i- ntracranial bleeding, which can exert life threatening pressure on the brain.

Explain the criteria for lymphoma staging

According to the Ann Arbor scheme commonly used for lymphoma staging, disease progression is considered in stages of one through four. The scheme is named after the city of Ann Arbor, Michigan, where lymphoma specialists gathered in 1971 to design this staging scheme. In stage one, neoplastic cells occur only in a single lymph node or single extra-lymphatic site, designated as one E. In stage two, neoplastic cells occur in two or more lymph node regions on the same side of the diaphragm or with involvement of limited extra-lymphatic sites designated as two E. In stage three, neoplastic cells occur in various lymph node regions on both sides of the diaphragm, which can include the spleen(designated as three S) and/or limited contiguous extra-lymphatic sites, designated as three E. in stage four, neoplastic cells occur in multiple or disseminated foci, with involvement of more extra-lymphatic organs or tissues with or without lymphatic involvement. Patients with lymphoma diagnosis in stages three and four have the worst prognosis, witha n average five-year survival rate of about 60%.

Functional Platelet Disorders Identify various conditions associated with acquired platelet functional abnormalities

Acquired platelet function defects may be caused by various physiological conditions, exposures, and treatments including: chronic kidney failure and hematologic neoplastic disorders. Exposures to aspirin, non-steroidal anti-inflammatory drugs (NSAIDs), and alcohol interfere with platelet activation. In cardiopulmonary bypass surgery, platelets may fragment as they interact with the abnormal surfaces from the surgery, resulting in thrombocytopenia and bleeding episodes. We conclude with a brief evaluation of acquired platelet disorders in chronic kidney failure and hematologic disorders.

Functional Platelet Disorders Identify various conditions associated with acquired platelet functional abnormalities

Acquired platelet function defects: chronic kidney failure In chronic kidney failure, platelet defects occur as a result of waste products that accumulate in the plasma, although the specific damaging metabolites are not clearly established. Platelet aggregation in aggregometry tests is abnormal with standard agonists, suggesting these platelets have a defective secretory or activation response. These platelets also exhibit defective procoagulant activity. Significantly, patients with kidney failure may experience severe bleeding symptoms, including ecchymoses, gastrointestinal bleeding, and heavy bleeding in body cavities. Fortunately, dialysis treatment can effectively decrease these bleeding symptoms. Acquired platelet function defects: hematologic disorders Bleeding and clotting defects are common consequences of hematologic neoplasms. Recall that platelet abnormalities occur in myeloproliferative neoplasms (MPNs), chronic myeloid leukemia(CML), Polycythemia vera (PV), Essential thrombocythemia (ET), and primary myelofibrosis (PMF) due to defects during megakaryocyte development. These defects may manifest as thrombocytopenia, or thrombocytosisas occurs in ET. Although platelet levels would be increased in thrombocytosis, these platelets typically display abnormal function. Patients with myelodysplasticsyndromes (MDS) or acute leukemias typically experience excessive bleeding, usually due to thrombocytopenia, but platelet functional abnormalities may also occur. In multiple myeloma, platelet function is affected by paraproteins that interfere with platelet aggregation and fibrin polymerization.

Describe clinical and neoplastic cell characteristics of ATLL, NNKTL, and PTCL, NOS

Adult T cell leukemia/lymphoma (ATLL) is most prevalent in Southwestern Japan, Central Africa, and the Caribbean. Development of ATLL is closely associated with human T cell leukemia virus one (HTLV-1), which infects about 10 million people worldwide. HTLV-1 shares significant similarities with HIV, the human immune deficiency virus. Like HIV, HTLV- 1 infects CD4 T cells and forms a provirus that is integrated into the chromosome of the infected cell. Additionally, like HIV, HTLV-1 is transmitted through sex, contaminated blood, or breast feeding. However, unlike HIV, HTLV-1 rarely replicates, so it chiefly becomes a latent sub-clinical infection. However, about 2 to 5 percent of HTLV-1 infected individuals will go on to develop ATLL ,in which HTLV-1 genome integration and additional chromosome rearrangements are observed in the cancerous T cells. In ATLL, the neoplastic cells have a distinctive clover-like nucleus. This is an aggressive disease, with a median survival of just 8 to 10 months.

Define the following terms: primary hemostasis, secondary hemostasis, endomitosis, demarcation membrane system

Although megakaryocytes do not undergo true mitosis, where the cell divides into two daughter cells, they do undergo a process called endomitosis. In endomitosis, a cell goes through the S phase, where the DNA content doubles, but there is no nuclear or cytoplasmic division that follows. Thus, the cells become polyploid, and the DNA content ranges from 8N to 64N (whereN is the normal number of haploid chromosomes). Development during these phases is driven by cytokines TPO, IL-6, and IL-11. Importantly, with each round of endomitosis, the developing megakaryocyte's cytoplasmic volume also increases, leading to development of a large, mature megakaryocyte. A mature megakaryocyte ranges from 40 to 100 micrometers. On a stained mature megakaryocyte slide we see a large, purple-stained nucleus with an extensive granular cytoplasm. As ploidy increases, the cytoplasm also becomes more granular, and the nucleus becomes multi lobed during the maturation process. Each megakaryocyte produces 1000 to 3000 platelets, depending on the extent of parent cell ploidy. A higher ploidy increases platelet yield. While the platelets are released into the blood circulation, the megakaryocyte nucleus remains behind, where it is subsequently engulfed by a resident bone marrow macrophage. Here we see a standard peripheral blood smear with arrows pointing to small, purple-stained platelets that appear as small dots scattered amongst the RBCs. ....

Primary Hemostasis Describe the arachidonic acid pathway, and explain the net hemostatic effects of aspirin

Arachidonic Acid (AA) pathway: Agonist activation induces activation of phospholipases that hydrolyze portions of phospholipid membranes to release free arachidonic acid composed chiefly of fatty acid tails. The enzyme cyclooxygenase(COX) modifies arachidonic acid into intermediate compounds PGG2 and PGH2. In platelets, the enzyme thromboxane synthetase converts the intermediates into Thromboxane A2, a platelet agonist that also enhances vasoconstriction. On the other hand, in endothelial cells, prostacyclin synthetase converts the PGG2 and PGH2 intermediates into prostacyclin (PGI2). PGI2 is a platelet antagonist, and it enhances vasodilation. Thus, the prothromboticaction of thromboxane A2 is counterbalanced by PGI2. These pathways are significant to the action of aspirin, which blocks the activity of COX. This inhibitory effect is irreversible in platelets because they have no nucleus and thus are incapable of de novo synthesis of the COX enzymes. On the other hand, endothelial cells can recover through expression of the COX enzyme genes. The net effect of aspirin is a reduced quantity of thromboxane while prostacyclin production continues, and thus, the balance is tipped towards platelet inhibition and vasodilation.

Secondary Hemostasis Define zymogen, tissue factor, contact factor, and thrombus

Beginning on the right side, the extrinsic pathway involves factor 7 and tissue factor within the exposed sub-endothelium. Tissue factor is the key initiator of in vivo coagulation. On the left is the intrinsic pathway, composed of plasma coagulation factors. Note how the intrinsic and extrinsic pathways converge in their activation of common pathway factor 10. Activated factor 10 and factor 5 convert pro-thrombin into thrombin. In turn, thrombin converts fibrinogen into insoluble fibrin. Where do these coagulation reactions take place? The extrinsic pathway is initially activated on sub-endothelial cell surfaces that are exposed when a blood vessel is injured. Recall that the exposed collagen in these areas also provides surfaces for platelet adhesion, activation, and aggregation. Activated clotting factors primarily bind to activated platelet phospholipid membranes at the injury site. This serves to localize the clotting cascade reactions.

Compare and contrast cellular characteristics of CLL/SLL, follicular lymphoma, DLBCL, and Burkitt lymphoma List therapeutic strategies for mature lymphoid leukemia/lymphoma Burkitt lymphoma

Burkitt lymphoma. This B cell neoplasm is named after Dennis Burkitt, who was a British surgeon. In 1957, Burkitt investigated the cause of jaw bone swellings he observed in children from Uganda, and determined that these common swellings were due to malignancies. This is a high-grade non-Hodgkin lymphoma for which there are three subtypes: endemic, sporadic, and immune deficient. The endemic subtype has a high incidence in Africa. The sporadic subtype is responsible for one third of all pediatric lymphomas that occur outside of Africa. The immune deficient subtype accounts for adult cases of Burkitt lymphoma that occur mostly in immune compromised individuals, such as those infected with HIV. Burkitt lymphoma often involves neoplastic growth in extra-nodal sites. In particular, the endemic type often involves neoplastic cell growth in the facial and jaw bones. The sporadic type often involves neoplastic cell growth in the intestines, ovaries, and kidneys. Lymph node biopsy of Burkitt lymphoma characteristically reveals neoplastic cells that appear as a starry sky, such as the example shown here. The sky appearance is due to the abundant blue staining nuclei of the neoplastic cells. The white stars are due to the appearance of scattered tingible body macrophages. In the image, arrows point to tingible body macrophages,f or which we can easily see apoptotic bodies that had been engulfed. Burkitt lymphoma cells also contain numerous intracytoplasmic lipid droplets, which is why they look like they have a white halo around them.

Compare and contrast cellular characteristics of CLL/SLL, follicular lymphoma, DLBCL, and Burkitt lymphoma CLL/SLL

CLL/SLL stands for chronic lymphocytic leukemia / small lymphocytic lymphoma. Although it is described as both a leukemia and a lymphoma, these are considered to be the same disease, but just with different clinical manifestations, depending on the predominant location and organization of the neoplastic cells. CLL presents with peripheral blood lymphocytosis, with lymphocyte counts greater than five times ten to the ninth per liter of blood. The reference range is one to 4.8 times ten to the ninth per liter of blood. The neoplastic cells appear as small, mature lymphocytes. Notably, as indicated in the accompanying blood smear from a CLL patient, smudge cells may be observed. These smudge cells are CLL neoplastic cells that burst open during slide preparation, and their presence is a hallmark of this disease. As CLL progresses, lymph node involvement may occur, at which point the disease would be described as SLL, small lymphocytic lymphoma. SLL presents with lymph adenopathy, swollen lymph nodes, and this may in turn progress to peripheral blood and bone marrow involvement and be described as CLL. SLL is often detected in asymptomatic patients during routine exams, either due to detection of a lymph node mass or during routine CBC analysis. CLL/SLL is typically a low-grade disease, characterized by slow progression to anemia, thrombocytopenia, and neutropenia. On the right is an image of a lymph node biopsy from an SLL patient. Here we see many small lymphocytes with round nuclei and clumped chromatin.

Describe the clinical presentation of patients with mature lymphoid neoplasms

Clinical disease presentation of mature lymphoid neoplasms may be high grade or low grade. High grade lymphoma disease symptoms present rapidly, with an enlarging palpable mass, and larger neoplastic cells that have more numerous mitotic cells, indicative of a faster rate of proliferation. The patient may experience fever, unintentional weight loss, and night sweats. Low grade disease is characterized by a slow progression of symptoms, including painless lymphadenopathy(swollen lymph nodes), and hepatosplenomegaly (enlargement of the liver and spleen). Lymphocytosis is present, but the cells are smaller and they display less mitotic activity, indicative of a lower rate of proliferation. Disease prognosis is a function of both age and stage of the disease progression. Generally, patients who are over 60 have a worse prognosis. Stage of progression indicates the extent and distribution of the neoplastic growth. Patients with widespread lymphoma usually have a worse prognosis. Determination of lymphoma stage involves a combination of radiologic studies, peripheral blood examination, and bone marrow aspirationand core biopsy

hemostasis diagnostic testing: Describe the use of PT, APTT, TT, and D-dimer tests for assessment of secondary hemostasis and fibrinolysis

Clotting factor reflex testing must be performed as a follow up if PT and/or APTT tests are prolonged. First, mixing studies can be performed to confirm that there is an actual deficiency, as opposed to the presence of an inhibitory substance in the patient sample. A portion of the patient sample is combined with normal healthy plasma to see if the normal clotting factors are impaired. If not, then this indicates that the patient sample does not have inhibitory substances, but rather that there is a specific factor defect. Thus, specific factor tests would then follow. For these, there are numerous commercially available test kits to evaluate specific clotting factors. the D-Dimer test. This test is used for assessment of fibrinolysis. This is a sensitive test for measurement of fibrin degradation products. Recall that D-dimers are produced when fibrin is broken down by plasmin. The D-dimer test is mostly used in diagnosis of thrombosis (excess clotting).

clotting factor nomenclature

Clotting factors are traditionally represented as Roman numerals I through XIII, for factors 1 through 13. A Roman numeral followed by the letter "a" indicates the activated form of the factor. Note that the factors were assigned to their numbers in the order of their discovery and not in the order of their respective reaction sequence; and each clotting factor has one or more common names or synonyms. It is important to note some exceptions to the nomenclature. Some factors are usually referred to by name and only occasionally by the Roman numeral or factor number. THESE INCLUDE:prothrombin (factor 2) and its activated form thrombin (factor 2a); fibrinogen (factor1), which is converted to fibrin (factor 1a); tissue factor (factor 3); and calcium, which is factor 4. Factor 6 does not exist Note how the 3 pathways culminate in formation of the stable, cross-linked fibrin clot. Here, clotting factors 12, 11, 9, 8, 7, 10, 5, and 13 are each further labeled with their equivalent Arabic number. Most clotting factors circulate as zymogens, inactive proenzymes, which act as serine proteases following activation. An important exception is factor 13, which functions as a transglutaminase enzyme important for the final step of fibrin cross-linking. As mentioned previously, activated forms of the clotting factors are indicated by a lowercase letter "a". Notably, three factors exhibit no enzyme activity. Factors 8a and 5a are cofactors for factors 9a and 10a respectively, and fibrinogen is a protein substrate that is converted to fibrin. There are three critical enzyme complexes formed during coagulation activation. The extrinsic ten-ase,the intrinsic ten-ase, and prothrombinase. Extrinsic ten-ase forms onthe sub-endothelial cell membrane, whereas intrinsicten-ase and prothrombinase form on platelet membrane surfaces.

Compare and contrast hemostatic characteristics of resting and activated endothelial cells

Consider first resting endothelial cells. There is an intact endothelial cell lining and no injury to the blood vessel. The negatively charged surface structures of the endothelial cells naturally repel proteins and platelets. Further, the cells produce various factors that inhibit platelet activation and promote vasodilation, including prostacyclin (PGI2), nitric oxide (NO), and the enzyme ADPase. The resting endothelial cells of uninjured vessels also inhibit secondary hemostasis by various mechanisms. In these mechanisms, surface proteins interact with soluble factors to inhibit key clotting factors, and there by slow down or block formation of the fibrin-platelet plug. Resting endothelial cells further discourage clot formation by release of fibrinolysis activators tPA and uPA to induce dissolution of the fibrin-platelet clot. Following injury of the endothelial lining, activated endothelial cells secrete a variety of compounds that enhance vasoconstriction and promote platelet activation; including endothelin (ET), platelet activating factor(PAF), thromboxaneA2 (TXA2), and von Willebrand factor (VWF). Activated endothelial cells promote secondary hemostasis through activation of coagulation factors. Secretion of von Willebrand factor (VWF) stabilizes a key coagulation factor, and coagulation is activated by exposure to tissue factor (TF), a membrane protein expressed on the surface of sub-endothelial cells. Tissue factor is not actively secreted, but rather it activates coagulation when there is a breach in the endothelial cell layer. Finally, activated endothelial cells are anti-fibrinolytic. Secretion of fibrinolysis inhibitor such as plasminogen activator inhibitor, PAI-1, favors maintenance of the fibrin-platelet clot.

Compare and contrast cellular characteristics of CLL/SLL, follicular lymphoma, DLBCL, and Burkitt lymphoma List therapeutic strategies for mature lymphoid leukemia/lymphoma DLBCL

Diffuse large B cell lymphoma, DLBCL, is the most common typeof mature B cell neoplasm. It is actually a heterogeneous group of neoplasms that are characteristically composed of large B lymphocytes. These neoplasms arise de novo, or they are transformed by progression from follicular lymphoma or small lymphocytic lymphoma. Included here is an image of a lymph node biopsy from a DLBCL patient. The biopsy reveals many large, abnormal, lymphoid cells and several that are actively undergoing mitosis, as indicated by the arrows. Prognosis: The myc oncogene plays a significant role in DLBCL prognosis. The Myc gene is rearranged in 5 to 15 percent of cases. Over expression of both Myc and Bcl2 oncogenes results in high grade disease. Previous infection with Epstein-Barr Virus (EBV) may also be significant to the susceptibility and prognosis of DLBCL. Recall that EBV, the causative agent of infectious mononucleosis, infects B lymphocytes. Furthermore, the viral DNA may integrate into the B cell genome, resulting in latent infection. The viral DNA integration may induce genetic instability, increasing the risk of lymphoma development. The WHO has designated EBV-positive DLBCL as a distinct sub-category, as patients in this category experience highly aggressive disease with an inferior survival of about 24 months. Treatment: The long-term remission rate of most types of DLBCL is 50 to 60% following treatment with multi-agent chemotherapy and/or anti-CD20 monoclonal antibody therapy (rituximab). CAR-T based therapies forDLBCL are in development.

Compare and contrast cellular characteristics of CLL/SLL, follicular lymphoma, DLBCL, and Burkitt lymphoma List therapeutic strategies for mature lymphoid leukemia/lymphoma Burkitt lymphoma

Etiology: During his original investigations, Dr. Burkitt suspected that a pathogen might be responsible for the rapid-onset tumors he observed in children in Uganda. This suspicion has been supported by decades of research indicating that Epstein-Barr Virus (EBV) may play a significant role in development of not only Burkitt lymphoma, but other B cell cancers as well, including DLBCL, as mentioned previously. How EBV contributes to lymphoma pathogenesis is still not fully understood. However, it is hypothesized that high risk EBV variants in Africa may be responsible for the endemic cases of Burkitt lymphoma, by possibly blocking apoptosis and promoting genetic instability in the EBV infected B cells. Cytogenetic evaluations have revealed that most cases of Burkitt lymphoma share a common translocation that occurs between chromosomes 8 and 14. This translocation results in formation of a fusion gene between the Myc proto-oncogene and the IgH (immunoglobulin heavy chain) gene. As a result of this translocation, Myc oncogene expression is activated by IgH gene regulatory sequences. Thus, hyperexpression of Myc serves as a critical oncogene driver in Burkitt lymphoma cells. Treatment: Fortunately, Burkitt lymphoma is highly curable in children following high dose chemotherapy; and adults respond well to Rituximab anti-CD20 monoclonal antibody immunotherapy.

Explain the concept of hemostatic balance and the physiological consequences of imbalance

Every stage of hemostasis must be finely regulated to assure that clotting is localized and occurs only when needed. This regulation involves complex interactions between all components of hemostasis, including vascular endothelial cells, platelets, coagulation proteins, and fibrinolytic proteins. Accordingly, there are biochemical inhibitors to counteract hemostasis activators at every step of clot formation and dissolution. Hemostatic regulation is represented here as a delicately balanced push and pull with clotting on one side and fibrinolysis on the other. Activators and inhibitors of clotting and fibrinolysis keep the coagulation system in balance. An imbalance in the activation or inhibition of either clotting or fibrinolysis causes thrombosis(too much clotting) or excessive bleeding. In this session we focus on various examples of clotting and fibrinolysis inhibitors. Specifically, we evaluate clotting factor inhibitors: antithrombin (AT), proteinsC & S, tissue factor pathway inhibitor (TFPI), and heparin. We also evaluate fibrinolysis inhibitors: plasminogen activatorinhibitor-1 (PAI-1), alpha2-antiplasmin, alpha2-macroglobulin, and thrombin activatable inhibitor (TAFI). Additionally, we consider several inhibitors of primary hemostasis.

Bleeding Disorders Distinguish between petechiae, purpura, ecchymoses, and hematomas

Excessive bruisability that occurs in bleeding disorders can present in various particular forms. Petechiae, pinpoint-size bruises smaller than 3 millimeters, are a common hallmark manifestation of primary hemostasis disorders. Petechiae result from blood leakage through capillaries. Purpura are intermediate lesions about 3 millimeters to one centimeter in size and are commonly seen in disorders involving blood vessels. Ecchymoses are larger thanone centimeter and result from blood leakage from larger vessels into subcutaneous tissue. Ecchymoses occur in both primary and secondary hemostasis disorders. Hematomas present as a blue or purple raised area due to blood leakage from underlying vessels that collects below intact skin. In the absence of blunt injury, the occurrence of hematom as is indicative of secondary hemostasis disorders.

Describe clinical and neoplastic cell characteristics of ATLL, NNKTL, and PTCL, NOS

Extra-nodal NK/T celllymphoma, nasal type, (NNKTL), is so called because the neoplastic cells exhibit aggressive growth in the nasal cavity that causes destruction of blood vessels and tissue necrosis in the nasal cavity. Extra nasal lesions can also occur in the skin, soft tissue, gastro-intestinal tract, and the testes. The most frequent occurrence of this disease is in East Asia, and in native American populations in Mexico and Central and South America. Most cases of NNKTL are associated with Epstein-Barr Virus(EBV) infections, suggesting that EBV plays a role in lymphoma development. Note that, unlike the B cell lymphomas discussed previously, NNKTL is associated with EBV type 2, which infects T cells and NK cells; whereas EBV type 1 infects B cells. This aggressive lymphoma, which displays tissue damaging cytolytic activity, is derived from either NK cells or cytotoxic T cells. Accordingly, expression of CD56 ,a common NK and CTL marker, is usually expressed by these neoplastic cells.

State the consequences of excessive or inadequate fibrinolysis

Fibrinolysis is the process of digestion and removal of a fibrin clot. The process is highly regulated and limited to the area of fibrin formation. Excessive fibrinolysis results in excessive bleeding inadequate fibrinolysis results in excessive clotting.

hemostasis diagnostic testing: Explain the use of platelet count, platelet function analyzer, and platelet aggregometry for assessment of primary hemostasis

Functional abnormalities can be assessed using a platelet function analyzer (PFA), by platelet aggregationstudies, or by flow cytometry to evaluate platelet surface receptors. The platelet function analyzer, PFA, is an instrument that tests platelets in vitro for their capacity to adhere to collagen. In this assay, whole blood from a patient sample is aspirated at high shear rate through a disposable cartridge. At other end of the cartridge, there is an opening with a membrane that is coated with collagen and a platelet agonist, such as ADP or epinephrine. As platelets encounter the membrane, they adhere to the collagen and are activated by the agonist. Von Willebrand factor (VWF) in the patient blood sample enables strong adhesion of platelets to the membrane in high shear rate flow. A pressure sensor detects the presence of a platelet plug, which blocks the flow of blood through the opening. The readout measurement is called the closure time. A prolonged closure time is indicative of platelet dysfunction, or it can indicate a VWF deficiency.

Compare and contrast NLPHL and cHL

Further, cHL itself includes four sub-types based on surrounding cells and the extent of lymph node fibrosis. These sub-types are nodular sclerosis (NS), lymphocyte rich (LR), lymphocyte depleted (LD), and mixed cellularity (MC). Note that these classic Hodgkin lymphoma sub-types do not seem to have a particular prognostic significance. NLPHL is characterized by nodular proliferation of large neoplastic cells, such as the one indicated by the arrow in the accompanying image from a lymph node biopsy. These odd cells are called popcorn cells, because the large, multi-lobular nucleus resembles a popcorn kernel. Popcorn cells are also referred to as L & H (lymphocyticand histiocytic) cells because the popcorn cells are usually surrounded by lymphocytes and histiocytes (tissue macrophages). These cells display a classic B cell phenotype that includes expression of CD19 and CD20 surface markers. Classic Hodgkin lymphoma (cHL) disease is characterized by a different, odd cell type called the Reed-Sternberg (R-S) cell. Reed-Sternberg cells have two or more nuclear lobes with a clear area around each nucleus, which give the cell an owl's eye appearance. A single, large Reed-Sternberg cell with two nuclear lobes is indicated by the arrow in the accompanying image from a lymph node biopsy. The neoplastic R-S cells are derived from activated post-germinal center B cells which express CD30 in 7 5 to 80 percent of cases, while CD20 expression is evident in only 25 percent of cases. About 50 percent of these neoplastic cells test positive for EBV, suggesting an etiological connection. The exact mechanisms by which EBV contributes to the pathogenesis of cHL and other B cell lymphomas are not understood, and this continues to be an active area of research.

hemostasis diagnostic testing: Explain the use of platelet count, platelet function analyzer, and platelet aggregometry for assessment of primary hemostasis

Hemostasis laboratory testing would typically be initiated based on patient history of either excessive bruising and bleeding episodes, or a tendency to form inappropriate blood clots, called thrombosis. These abnormalities could indicate a defect in primary hemostasis due to abnormal platelet counts or platelet functional abnormalities. Alternatively, the bleeding or clotting defects could implicate secondary hemostasis clotting factor abnormalities. Standard diagnostic testing for evaluation of primary hemostasis would begin with platelet count, a standard CBC parameter. Platelet count (CBC parameter) indicates if platelet levels in the blood are above or below the reference range of 150 to 400 times 10 to the ninth per liter of blood. Note that platelet count does not evaluate platelet function, as it is simply a quantitative measurement. Abnormal platelet morphology revealed in a standard blood smear can indicate an underlying pathology, such as what occurs in some myeloproliferative neoplasms and myelodysplastic syndromes.

Quantitative Platelet Disorders List and explain various causes of thrombocytopenia

Here is a diagram depicting various causes of thrombocytopenia. Starting on the left side: > abnormalities during early hematopoiesis, megakaryopoiesis, and thrombopoiesis in the bone marrow can lead to decreased platelet production. Decreased platelet production indicates a failure of the bone marrow to deliver adequate platelets to the peripheral blood. In such circumstances, there is hypoplasia of megakaryocytes and ineffective thrombopoiesis, due to either hereditary or acquired abnormalities. An example of a hereditary condition is congenital megakaryocytic thrombocytopenia (CAMT), characterized by loss of function mutations in the TPO receptor gene, MPL. Acquired conditions of decreased platelet production include MDS, leukemias, lymphomas, and bone marrow fibrosis. Decreased platelet production is also a consequence of a plastic anemia and megaloblastic anemia, both conditions associated with pancytopenia, a deficiency in all hematopoietic lineages. > Increased splenic sequestration of platelets can decrease their availability in the peripheral blood. Recall that the splenic pool contains about one third of platelets released by the bone marrow, and these are in equilibrium with the circulating platelets. In hypersplenism conditions, the spleen is enlarged and hyperactive, resulting in an increased number of pooled platelets, and the circulating numbers are decreased. The occurrence of hypersplenism is usually complicated by other factors such as hemolytic anemia or infections. > Finally, thrombocytopenia may be attributed to increased platelet destruction due to immune or nonimmune mechanisms We focus on immune mediated mechanisms next.

Identify the major components of the fibrinolytic system

Here is a diagram of the fibrinolytic system. At the starting point is plasminogen, a zymogen produced by the liver. Following activation, plasminogen is converted to plasmin, which in turn mediates the degradation of fibrin into fibrin degradation products (FDP). There are three general categories of plasminogen activators: physiologic, exogenous, and contact. The physiologic activators are tPA and uPA. tPA, which stands for tissue plasminogen activator, is released by endothelial cells and is the principal fibrinolytic activator within the circulatory system. uPA, which stands for urokinase plasminogen activator, is produced by the kidneys and is found in the urine, plasma, and extracellular matrix of many tissues. uPA also plays a role in cell migration, wound healing, and is implicated in cancer cell metastasis. Exogenous plasminogen activators are produced outside of the body by bacterial pathogens. Streptokinase, SK, is produced by streptococcal bacteria, and staphylokinase, SAK, is produced by staphylococcus aureus. Production of these plasminogen activators promotes bacterial dissemination in the host. Contact factor plasminogen activators are associated with initial steps of the intrinsic pathway and include Kallikrein, factor 12a, and factor 11a. These factors account for about 15 percent of plasmin generating activity in vivo.

Compare and contrast NLPHL and cHL

Hodgkin lymphoma (HL) is a lymph node-based neoplasm that is characteristically composed of a small number of large, mononucleated or multinucleated neoplastic cells with very peculiar morphologies Hodgkin lymphoma is classified into two main diseases: Nodular lymphocyte predominant HL (NLPHL), which accounts for 5 percent of cases, and classical Hodgkin lymphoma (cHL), which accounts for 95% of cases.

Quantitative Platelet Disorders Distinguish between autoimmune and alloimmune ITP

ITP autoimmunity is described in terms of three phases: > Newly diagnosed, with a duration of up to three months > Persistent ITP cases, with a duration of 3 months to one year; >Chronic ITP cases (cITP), for which duration is greater than one year. cITP is more often seen in adults. However, in general, ITP autoimmunity is most common in children 5 to 6 years old and often follows from a viral infection. Patients experience mild to moderate bleeding with bruising, petechiae, and mucous membrane bleeding. Spontaneous remission occurs in 85% of children and 15% of adults.

Quantitative Platelet Disorders Distinguish between autoimmune and alloimmune ITP

Immune thrombocytopenia (ITP) is the most common cause of thrombocytopenia. ITP is reminiscent of immune hemolytic anemia, but instead of binding to RBCs, antibodies bind to platelets, which are subsequently removed by splenic macrophages. In autoimmune ITP, the antibodies are auto reactive, meaning they are self antibodies reacting against self platelets. In alloimmune ITP, the antibodies are alloreactive, meaning the antibodies are from another person. Sources of alloreactive antibodies are blood transfusions or maternal antibodies that react against fetal platelets, in a condition called neonatal alloimmune thrombocytopenia (NAIT). As the macrophages phagocytose the antibody-coated platelets, platelet depletion increases the amount of TPO available to drive megakaryocyte development. Bone marrow activity can then compensate by an increased rate of megakaryopoiesis. However, thrombocytopenia develops if the rate of immune destruction exceeds the compensatory capacity of the bone marrow response. Initial diagnosis of ITP would begin with an evaluation of patient history and physical examination. In ITP, most CBC parameters would be normal, except platelet count would be below 100 times 10 to the sixth per liter. Further, an elevated IPF would indicate that there is normal bone marrow megakaryocyte development. Detection of platelet associated antibodies (PAIg) would be a defining diagnosis. Although assays to measure these antibodies have long lacked sensitivity and specificity, there have been significant recent improvements in the detection methods, making this a more reliable approach for confirming ITP.

Explain mechanisms of fibrin and FDP negative feedback

Importantly, fibrin and fibrin degradation products exert negative feedback effects on coagulation, thereby reducing risks of excessive clotting. Fibrin indirectly controls clotting because it has a strong affinity for thrombin, thereby limiting the amount of thrombin that is available to convert more fibrinogen to fibrin. Fibrin degradation products (FDPS) function as inhibitors of fibrin formation, and thus have an anti-coagulant effect. FDPs also interfere with platelet aggregation, thereby exerting negative feedback on primary hemostasis.

Compare and contrast cellular characteristics of CLL/SLL, follicular lymphoma, DLBCL, and Burkitt lymphoma List therapeutic strategies for mature lymphoid leukemia/lymphoma follicular lymphoma

In follicular lymphoma, patients present with generalized, painless lymphadenopathy, and a few have peripheral blood involvement. Involvement of extra-nodal sites is also unusual As indicated in the lymph node diagram, follicles are specialized regions where B cell activation and clonal proliferation normally occur. The darkened area within the follicles is called the germinal center. This is where the most active B cell proliferation occurs. Follicular lymphoma neoplasm cells originate from germinal center B cells. On the right is an image of a lymph node biopsy from a patient with follicular lymphoma. We can see a homogeneous population of small lymphoid cells with angular, twisted nuclei. There is also an absence of "tingible" body macrophages. Tingible body macrophages are so called after they have phagocytosed a dead cell. These would normally be observed in lymph nodes, since normal B cells regularly die during the selective processes that occur in the lymph nodes. This is evidence of the reduced apoptosis that occurs in follicular lymphoma cells. We'll consider why this is so next. Cytogenetic analyses of follicular lymphoma neoplasms reveal a common chromosome translocation causes over-expression of Bcl2, which inhibits apoptosis. Thus, Bcl2 is a common oncogene driver in this disease. Median survival of follicular lymphoma is 7 to 9 years, and local excision of affected lymph nodes extends survival. Patients are typically treated with a combination of anti-CD20 immunotherapy and chemotherapy. Ongoing clinical studies indicate that relapsing patients may benefit from Bcl2 targeted therapy

Functional Platelet Disorders Describe the range of clinical symptoms that occur with functional platelet disorders

In functional platelet disorders, clinical symptoms vary from being asymptomatic, to mild, to severe, life-threatening hemorrhaging. Generally, the types of bleeding symptoms observed in these disorders include petechiae, easy spontaneous bruising, mucosal bleeding, and prolonged bleeding from trauma.

Describe the three principal mechanisms by which secondary hemostasis is inhibited

In non-injured blood vessels, endothelial cells employ three major inhibitory mechanisms to suppress secondary hemostasis. These are: - anti-thrombin - protein C/protein S - tissue factor pathway inhibitor (TFPI) systems .....

Describe general characteristics of mature T/NK cell neoplasms, including frequency, heterogeneity, origin, and symptoms

In this final session on mature lymphoid neoplasms, we begin with a focus on mature T cell and NK cell malignancies, and then conclude with Hodgkin lymphoma. Mature T and NK cell neoplasms are less common than the B cell leukemia/lymphomas described previously, and account for about 10 to 15 percent of mature lymphoid neoplasms. These neoplasms are heterogenous, owing to the numerous mature T cell subtypes with significant differential functions in the immune system. For example, we have previously considered CD4 positive Th cells, CD8 positive Tc cells, and Treg cells; and there are additional subtypes of the Th cells as well. The origin of these neoplasms also varies. Some arise from the cells in lymph nodes, while other types arise in mucosal sites or cutaneous sites affecting the skin. Mature T/NK cell neoplasms mostly occur in adults over the age of 60, and typical symptoms are as described previously for other types of leukemias and lymphomas, including lymphadenopathy, fever, night sweats, and unexplained weight loss.

Indicate various platelet hemostasis functions

In this session we examine platelet development and structure. Platelets, also referred to as thrombocytes, are anucleate cellular fragments of cytoplasm derived from megakaryocyte precursor cells in the bone marrow. Development of megakaryocytes and platelets is regulated by the cytokine thrombopoietin (TPO). Platelets circulate in the peripheral blood for 7 to 10 days. The platelets circulate in a resting state. They are disk shaped and have a smooth, regular surface. Resting platelets repel each other and have minimal interaction with blood vessel walls. Aged or damaged platelets are removed by spleen or liver macrophages. Platelets play various roles in hemostasis. First, platelets are essential components of primary hemostasis, for which they slow down blood loss by formation of the primary platelet plug. Second, platelets contribute to secondary hemostasis by providing a surface for activation of coagulation factors. Furthermore, following activation, platelets secrete a variety of compounds that contribute to secondary hemostasis, vasoconstriction, wound repair, and fibrinolysis. Additionally, some platelet compounds affect immune responses.

Quantitative Platelet Disorders Identify laboratory tests used for evaluation of thrombocytopenia

In this session, the focus is on various causes of thrombocytopenia. A diagnosis of thrombocytopenia is made when the platelet count falls below the reference range of 150 times 10 to the sixth per liter of blood. However, there is usually no increased risk of bleeding episodes unless the platelet count falls below 50 times 10 to the sixth per liter; and the risk of severe and spontaneous bleeding occurs when platelet levels fall below 10 times 10 to the sixth per liter. In such circumstances, patients experience spontaneous bleeding from mucous membranes, including the gastrointestinal tract, urogenital tract, and the nose. Most significantly, platelet depletion may lead to fatal bleeding into the central nervous system. Laboratory evaluation of thrombocytopenia begins with a standard CBC, where platelet count is found to be below the reference range. Platelet function analyzer (PFA) results are consistent with bleeding severity observed in vivo; with increased closure times if platelet counts fall below 50 times 10 to the sixth per liter, and no closure achieved if levels fall below 10 times 10 to the sixth per liter. Measurement of platelet maturity can indicate the basis of the thrombocytopenia. Platelet maturity can be assessed in automated analyzers that measure the immature platelet fraction (IPF) in a blood sample. Newly released platelets are called reticulated platelets. An increase in reticulated platelets is indicative of an increased occurrence of platelet destruction in the circulation. Whereas normal or reduced levels indicate an issue with megakaryopoiesisin the bone marrow.

Functional Platelet Disorders Compare and contrast underlying defects and diagnostic test results for the following inherited platelet abnormalities: Bernard-Soulier syndrome, von Willebrand disease, activation disorders, Glanzmann thrombasthenia, and Scott syndrome

Inherited platelet abnormalities : von Willebrand disease (Adhesion disorder) Von Willebrand disease (VWD) is a deficiency or defect in plasma VWF, which normally mediates platelet binding of GP1b/9 (CD42) to the sub-endothelial collagen. CBC and blood smear evaluations indicate normal platelet counts and normal platelet morphology. Recall that the Von Willebrand factor (VWF) plasma protein also functions in secondary hemostasis through stabilization of factor 8 of the intrinsic pathway. Thus, in hemostasis screening tests, VWD presents with an increased activated partial thromboplastin time (APTT). Aggregation tests can also be useful in diagnosis of von Willebrand disease. Since the platelets have no activation or fibrinogen receptor defects, there is normal platelet fibrinogen aggregation in response to standard agonists: ADP, collagen, and epinephrine, as observed in Bernard-Soulier. This is reflected in the first three aggregation curves below. With ristocetin alone, we see there is no agglutination. However, platelet agglutination can occur with ristocetin, as long as exogenous VWF is provided, as reflected in the last aggregation curve below.

Functional Platelet Disorders Compare and contrast underlying defects and diagnostic test results for the following inherited platelet abnormalities: Bernard-Soulier syndrome, von Willebrand disease, activation disorders, Glanzmann thrombasthenia, and Scott syndrome

Inherited platelet abnormalities: Glanzmann thrombasthenia (GT) Glanzmann Thrombasthenia (GT) is a well-studied platelet aggregation disorder associated with a deficiency or defect in the GP2b/3a (CD41/CD61) fibrinogen receptor. GT is inherited as an autosomal recessive trait,for which bleeding symptoms are only observed in homozygotes Symptoms include purpura, nose bleeds, gum bleeds, and heavy menstrual periods. Bleeding severity ranges from mild bruising to severe, recurrent mucosal and subcutaneous bleeding beginning at birth. CBC and blood smear evaluations indicate normal platelet counts and normal platelet morphology. Reflex testing reveals the functional abnormality, with increased PFA closure time, and decreased expression of GP2b/3a (CD41/Cd61) fibrinogen receptor, as revealed by flow cytometry. Aggregation tests can also be useful in diagnosis of Glanzmann thrombasthenia, as reflected in the aggregation curves below. Aggregation is absent or severely diminished in response to agonists: ADP,collagen, and epinephrine. This is because the platelets cannot aggregate with fibrinogen when the fibrinogen receptor is absent. On the other hand, normal agglutination occurs with ristocetin because the VWF CD42 receptor is normal.

Functional Platelet Disorders Compare and contrast underlying defects and diagnostic test results for the following inherited platelet abnormalities: Bernard-Soulier syndrome, von Willebrand disease, activation disorders, Glanzmann thrombasthenia, and Scott syndrome

Inherited platelet abnormalities: Scott syndrome (procoagulant disorder) Recall that platelets serve an important role in activation of secondary hemostasis by providing appropriate phospholipid surfaces for assembly of coagulation factors. Normally, activated platelets undergo phospholipid rearrangements, referred to as scrambling, which is required for their procoagulant characteristics. In Scott syndrome, a rare recessive disorder, there is abnormal phospholipid scrambling. The platelets secrete and aggregate normally with fibrinogen to form a primary plug, but there is a defective assembly of intrinsic ten-ase and prothrombinase complexes and thus, defective thrombin formation.

Functional Platelet Disorders Compare and contrast underlying defects and diagnostic test results for the following inherited platelet abnormalities: Bernard-Soulier syndrome, von Willebrand disease, activation disorders, Glanzmann thrombasthenia, and Scott syndrome

Inherited platelet abnormalities: activation disorders Platelet activation disorders include granule and secretion abnormalities that affect alpha granules or dense granules. Alternatively, activation disorders can be due to signaling abnormalities. For example, defective thromboxane synthesis can occur if there are abnormalities in the arachidonic acid pathway that emanates from membrane phospholipids, or if there are abnormalities in agonist-signaling receptors. In activation disorders, symptoms are typically mild to moderate bleeding, with easy bruising and excess bleeding after surgery or childbirth. Laboratory diagnostics usually reveal normal platelet counts and normal morphology. An exception is in alpha granule deficiency, where patients present with mild thrombocytopenia and macrothrombocytes. These platelets may appear agranular and stain a gray color, so the condition is called gray platelet syndrome. PFA closure time is variable or increased. Aggregation tests can also be useful in diagnosis of activation disorders, as reflected in the aggregation curves below. While some secondary aggregation can occur with ADP or epinephrine, no aggregation occurs with collagen, indicating an inability of this agonist to activate adequate fibrinogen receptor expression. On the other hand, agglutination occurs with ristocetin because the VWF CD42receptor is normal.

Bleeding Disorders Describe various types of inherited and acquired vascular disorders

Inherited vascular disorders (HTT Cont.) Arteriovenous malformations (AVM) are benign blood vessel tumors (hemangiomas) that result in direct connections between arteries and veins without intermediary capillaries. These malformations form masses that can occur on internal organs, including the lungs, liver, and heart. in childhood and adolescence. Clinical signs of bleeding become more apparent after the age of 40, when patients experience gastrointestinal bleeding and iron deficient anemia episodes. Arteriovenous malformations (AVM) can cause significant complications. These masses can interfere with the functioning of major organs, including the lungs, liver, and heart. On the right we see a CT-scan image of a large arteriovenous malformation in the liver of an HHT patient. HHT disease management varies depending on the severity of the bleeding complications. Skin and mucosal telangiectasia lesions can be treated with laser ablation therapy. Patients who develop iron deficiency anemia can be treated with oral iron supplementation. Anti-VEGF antibody therapy is also being tested as a way to decrease occurrence of angiogenesis and thus prevent further disease progression.

Bleeding Disorders Describe various types of inherited and acquired vascular disorders

Inherited vascular disorders Hereditary hemorrhagictelangiectasia (HHT) is characterized by autosomal mutations in various genes affecting the transforming growth factor-beta signaling pathway. In HHT, blood vessel abnormalities arise through imbalanced regulation of angiogenesis (blood vessel development). Accordingly, HHT patients often present with increased levels of vascular endothelial growth factor (VEGF), which is a major driver of angiogenesis. As a result, HHT patients develop small vascular lesions called telangiectasias, and larger vascular lesions called arteriovenousmal formations (AVMs). Let's examine these further. Telangiectasia lesions develop from small, widened blood vessels in the skin and mucosal linings. ....

Bleeding Disorders Describe various types of inherited and acquired vascular disorders

Inherited vascular disorders Marfan syndrome is characterized as an autosomal dominant inheritance pattern in which 25 percent of cases have mutations in the fibrillin-1 gene. Marfan syndrome is a connective tissue disorder that affects a wide range of tissues and organs including the skin, ligaments, bones, blood vessels, eyes, and the heart. There are various clinical consequences of this condition, including overgrowth of long bones, decreased strength of blood vessels, and most seriously, heart problems that can shorten life span. Individuals with Marfan syndrome have distinctive physical characteristics. They tend to be very tall and slender, with elongated fingers and toes. These individuals have a large arm span that exceeds their body height. They have a narrow face and crowded teeth due to bone abnormalities in the jaw and palate. Interestingly, president Abraham Lincoln, with his towering height, pendulous arms, and thin face, is theorized to have had Marfan syndrome. However, absent a definitive genetic diagnosis, we cannot know this for sure.

Bleeding Disorders Predict lab findings in the various main categories of hemostatic disorders

Initial diagnosis of a bleeding disorder is based on patient history, including age of onset of hemorrhagic symptoms, family history, presence of other diseases, a complete drug history, and toxin exposure. Laboratory screening tests would include quantitative platelet counts, prothrombin time (PT), and activated partial thromboplastin time (APTT). Confirmatory follow-up tests are necessary to narrow down the specific nature of the disorder. Note that initial screening tests may be normal in primary hemostasis disorders, and would thus require platelet function tests to identify the specific abnormality. Recall that platelet function tests include PFA, aggregometry, and flow cytometry for platelet receptors. Secondary hemostasis disorders may be revealed by abnormal PT or APTT tests. These would need to be followed up by confirmatory testing. In order to rule out the presence of clotting factor inhibitors, mixing studies can be carried out by combining a patient plasma sample with a normal control plasma sample to see if fibrin formation is affected in PT or APTT tests. Ultimately, specific factor assays can confirm a particular clotting factor defect.

Functional Platelet Disorders Explain typical laboratory screening test results observed with functional platelet disorders

Laboratory screening test results are similar to thrombocytopenia conditions, except platelet count is usually normal. PT, APTT, and fibrinolysis tests are also usually normal. Platelet function analyzer and aggregometry tests can indicate the specific defect. Functional platelet disorders may be inherited or acquired. The inherited disorders are usually autosomal recessive traits. We will next focus on various examples of inherited disorders and conclude with a brief discussion of acquired disorders. Here is a depiction of two platelets bound together through the interactions of GP2b/3a receptors and fibrinogen. The platelet on the left is adhering to the exposed sub-endothelium through interaction with VWF and the GP1b/9 receptor. We also see a dense granule and an alpha granule represented in the left platelet. Inherited platelet disorders can affect any of the phases of platelet function, leading to bleeding abnormalities. These phases include adhesion tothe sub-endothelium via GP1b/9 binding to VWF (A and B in the figure); activation via dense granules, alpha granules, or membrane activation (D,E and F in the figure); aggregation via GP2b/GP3a and fibrinogen (C in the figure); and platelet membrane pro-coagulant activity (G in the figure).

Quantitative Platelet Disorders Distinguish between autoimmune and alloimmune ITP

Let's now consider alloimmune ITP. Neonatal alloimmune thrombocytopenia (NAIT) is analogous to hemolytic disease of the fetus and new born,in which maternal antibodies attack fetal RBCs. In NAIT, maternal IgG alloantibodies target paternal antigens on fetal platelets. The HPA1 platelet surface antigen is the most significant NAIT target, and can occur if the fetus is HPA1a positive and the mother is homozygous for the HPA1b allele. Thus, the mother may produce anti-HPA1a alloantibodies that attack the fetal platelets. The ITP symptoms range from mild to severe. The most serious risk of NAIT is bleeding in the central nervous system, which can cause intracranial hemorrhage in full-term infants. Treatment includes intravenous immunoglobulins and HPA-1a negative platelet transfusions. Alloimmune ITP is a potential consequence of blood transfusions leading to development of post-transfusion purpura. About 90% of those afflicted are females who are homozygous for the HPA-1b antigen, and produce anti-HPA-1a alloantibody. This could occur in women who had been previously sensitized during pregnancy or prior blood transfusion. Severe thrombocytopenia can occur in rare cases about 5 to 10 days after a transfusion, leading to symptoms of purpura and bleeding from mucosal tissues. The condition is usually self-limiting and resolves in 2 to 6 weeks.

hemostasis diagnostic testing: Describe the use of PT, APTT, TT, and D-dimer tests for assessment of secondary hemostasis and fibrinolysis

Let's now consider secondary hemostasis tests, which assess the activity of clotting factors required for the intrinsic, extrinsic, and common pathways. These tests require the use of platelet poor plasma (PPP). PPP is obtained from centrifuged plasma to remove the platelets. This is important because it removes platelet proteases, platelet phospholipids, and other platelet factors that interfere with the coagulation factors tests. Furthermore, platelet factor 4 (PF4) neutralizes heparin, which is used as a common anti-coagulant agent. Standard screening testsf or secondary hemostasis include: Prothrombintime (PT), Activated partial thromboplastin time(APTT), Thrombin time (TT), and the quantitative fibrinogen test. The prothrombintime (PT) test is used to assess inherited or acquired defects in the extrinsic and common pathways. In this test, purified thromboplastin (tissue factor) is combined with a sample of patient platelet poor plasma. TF + PPP . The time it takes for fibrin formation is the PT time. In this measurement, a clot is detected by optical or electromechanical instruments. The reference interval for PT time is 12 to 15 seconds, and a prolonged PT occurs if there is a deficiency in factor 7, 10,5, prothrombin, or fibrinogen. The prothrombin test is also used to monitor how a patient is responding to anti-coagulant therapy. In clinical settings, prothrombin time is often expressed as an international normalized ratio (INR) value. INR is based on the prothrombin time that has been adjusted with mathematical corrections that account for differences between thromboplastin reagents and instrumentation used by different laboratories. INR is used to monitor patients who are being administered long-term anti-coagulant therapy.

hemostasis diagnostic testing: Explain the use of platelet count, platelet function analyzer, and platelet aggregometry for assessment of primary hemostasis

Let's now examine the platelet aggregometry test. This test uses platelet rich plasma (PRP) obtained by gentle centrifugation of a plasma sample to yield a platelet concentration of 200 times 10 to the third per microliter. The test involves measurement of light transmission through the platelet-rich plasma sample, for which a light detector is used as the readout. A baseline light transmission of unaggregated platelets is registered as having high optical density due to a low level of light transmission picked up by the light detector. Next, an aggregating reagent is added to the platelet-rich sample. For example, the aggregating reagent may be a platelet agonist such as ADP. If the platelets are appropriately activated, then they may be able to aggregate, as indicated in the tube on the right. Here we compare light transmission of non-aggregated and aggregated platelets. As we can see from the sample on the right, platelet aggregation results in lower optical density and increased light transmission.

Outline the stages of megakaryopoiesis & thrombopoiesis

Let's now review how platelets are formed, beginning with megakaryocyte development as it occurs in the bone marrow. We find here the classic depiction of hematopoiesis. Megakaryocytes are part of the myeloid lineage. Beginning with the CMP, the differentiation pathway is similar to that of erythrocytes, including CFU-GEMM, and EMkP, but then diverging to the MkP, megakaryocyte progenitor. The process of megakaryocyte development in the bone marrow is called megakaryopoiesis. The process terminates with thrombopoiesis, in which platelets are formed and released into the circulation. About 1 times 10 to the eleventh thrombocytes are formed each day. We take a closer look at megakaryopoiesis next. Here is a depiction of megakaryocyte and platelet development, beginning with the progenitors BFU-Mk and CFU-Mk, followed by various stages of morphologically recognizable megakaryocytes, and terminating with platelets released into the peripheral blood. The bone marrow transit time from progenitor cell to platelet cell release is 4 to 7 days. The released platelets circulate in the blood, or they may be stored in the spleen. The megakaryocyte progenitor cells, BFU-Mk and CFU-Mk are not morphologically distinguishable. These progenitors undergo mitosis that is driven by the cytokines SCF, FL, GM-CSF, IL-3, and TPO. Given its significance to platelet formation,

Primary Hemostasis List the key platelet receptors involved in platelet adhesion, activation, and aggregation

Let's take a closer look at the process of platelet adhesion that occurs as an initial response to vessel wall injury. When there is a breach in the vascular endothelial cell layer, platelet attachment to exposed sub-endothelial collagen is mediated by the GP1a/2a receptor (CD49b/CD29). This is a moderate interaction that allows adhesion at low shear rates. Maintenance of platelet adhesion under conditions of high shear rates requires both VWF and the GP1b/9 (CD42) receptor. Von Willeb rand Factor(VWF) is a protein produced by endothelial cells and also stored in platelet alpha-granules. VWF binds to collagen, and the GP1b/9 receptor binds to the complex ofVWF bound to collagen. Ultimately, many platelet swill adhere similarly until a monolayer of platelets blankets the exposed xc sub-endothelial area.

Compare and contrast NLPHL and cHL

Lymphoma stage is the most significant prognostic marker in Hodgkin lymphoma. For patients in stage one or two, there is nearly a 90 percent5-year survival rate; whereas for stage four patients, this drops to 60 to 70 percent 5-year survival. Standard treatment for Hodgkin lymphoma is chemotherapy with or without radiation of the affected lymph nodes. Early-stage patients are highly responsive to these modalities, with an 80 percent cure rate. Autologous HSCT may be a second line treatment for patients who relapse. Additionally, several monoclonal antibody immunotherapies have been approved for treatment of Hodgkin lymphoma, including anti-CD30 antibodies for cHL, anti-CD20 antibodies for NLPHL, and various checkpoint inhibitors which enable non-specific enhancement of immune responses.

Describe the various FDPs generated by plasmin degradation of fibrinogen and fibrin

Next, we investigate the process by which plasmin breaks down blood clots. First, let's examine the enzymatic activity of plasmin digestion of fibrinogen proteins in a process called fibrinogenolysis. In this process, a distinct series of protein fragments are produced. These are collectively called fibrinogen degradation products (FDPs). Fibrinogen degradation follows a well-defined sequence of events to yield various fibrinogen cleavage products. First, small peptides are cleaved from the carboxyl end of the alpha chains, releasing X fragments composed of trinodular D-E-D domains Next, one of the D fragments is cleaved from the X fragment, yielding a binodular (D-E) Y fragment. Finally, cleavage ofthe Y fragments yields unimodular D and E fragments. Let's now examine the products of fibrin degradation. Recall that the factor13a transglutaminase catalyzes fibrin crosslinking. Crosslinked fibrin contains D-D dimers that are held together by strong covalent bonds. This schematic depicts the derivatives of plasmin digestion of fibrin. Plasmin does not easily cleave the D-D dimer bonds, so by comparison with fibrinogen-derived products, fibrin degradation products (FDPs) are larger molecules of varying complexity. We see on the far-left degradation products composed of a DD dimer and a single E fragment. In the middle, we see a 5 nodular fragment, referred to as DXD because it is composed of a trinodular X fragment (D-E-D) with covalent bond connections to an additional D domain on each end. A DD dimer is also released. Then, on the far right, we see two types of trinodular fragments, referred to as DY on the top or YD on the bottom. Recall that the Y fragment isa binodal fragment composed of D-E. In these cases, the D is connected to another D by virtue of the covalent bond.

Quantitative Platelet Disorders Identify laboratory tests used for evaluation of thrombocytopenia

Patients with platelet disorders typically present with petechiae, easy bruisability, and excess bleeding from superficial wounds in the skin and mucosal tissues.Platelet disorders maybe inherited or acquired conditions. Quantitative disorders may result in thrombocytopenia, where platelet levels are below the reference range, or may result in thrombocytosis, where platelet levels are above the reference range, as occurs in some myeloproliferative neoplasms and myelodysplastic syndromes. Qualitative disorders may present with morphological abnormalities and typically have functional abnormalities.

Primary Hemostasis List the key platelet receptors involved in platelet adhesion, activation, and aggregation

Platelet activation induced shape change causes the platelet to transform from a flattened disk into a spiny looking sphere. The long projections from the surface are called pseudopods, which form from reorganized cytoskeletal proteins in the platelet structural zone. Reorganization of the cytoskeleton occurs when microtubules contract and there is movement within the actin/myosin network. Membrane lipids are also reorganized. Below are scanning EM images of a smooth, disk-shaped, resting platelet on the left, and on the right is an activated platelet with several spiny pseudopod projections. Platelet Activation : Biochemistry Activation of platelets also induces various internal biochemical changes that trigger secretion of granule contents and activation of the arachidonic acid biochemical pathway. Additionally, activation induces expression of active fibrinogen GP2B/3a (CD41/CD61) receptors, which is critical for platelet aggregation.

Primary Hemostasis List the key platelet receptors involved in platelet adhesion, activation, and aggregation

Platelet activation is induced by agonists, which bind to platelet receptors and trigger a series of changes in platelet shape, biochemistry, and surface receptors. Here is a summary of key platelet agonists. These are agents that bind to platelet surface receptors and induce platelet activation. The list includes both platelet derived and non-platelet derived agents. Platelet derived agonists include ADP, serotonin, platelet activating factor, thromboxane A2, and VWF. Adenosine diphosphate, ADP, carried in platelet dense granules, should not be confused with metabolic ADP in the cytoplasm. Serotonin is a neurotransmitter that is stored in platelet dense granules. Platelet-activating factor (PAF) is a lipid derived compound secreted by activated endothelial cells; and thromboxane A2 and VWF are also produced by activated endothelial cells. Thromboxane A2 is also produced by activated platelets. *Serotonin, PAF, VWF, Thromboxane A2 = produced by CNS, stored by platelets ** PAF, VWF, Thromboxane A2 = also secreted by activated endothelial cells Non-platelet derived agonists include collagen structural proteins in sub-endothelial cell regions; thrombin, the activated version of prothrombin produced by the liver; and the hormone epinephrine, produced by the adrenal gland.

Describe key components of the four platelet structural zones

Platelets can be described in terms of four zones: the peripheral zone, the structural zone, the membrane zone, and the organelle zone. The peripheral zone includes the plasma membrane, including numerous receptors important for hemostasis, and a glycocalyx surface coat. The glycocalyx is composed of glycoproteins and glycolipids that confer a negative charge, which repels other platelets and blood vessels. This prevents platelets from randomly clumping together or from randomly clinging to blood vessel walls. The platelet structural zone is composed of cytoskeleton proteins that stabilize platelets into a discoid shape reminiscent of RBCs The cytoskeleton includes microfilaments composed of actin, microtubules, and myosin motor proteins. These provide structural integrity and mediate shape changes that occur following platelet activation. The platelet membrane systems include the open canicular system and the dense tubular system The open canalicular system is a series of surface-connected channels that originate from invagination of the megakaryocyte plasma membrane. This system provides a route for substances to diffuse in and out of the platelet interior. The dense tubular system originates from residual megakaryocyte endoplasmic reticulum and serves as a storage site for calcium ions. Finally, the platelet organelle zone inncludes standard cellular organelles including mitochondria and lysosomes; as well as two types of specialized, membrane-bound granules containing a range of factors important for mediating hemostasis. Dense granules are so named because they appear dense in electron microscope images. Alpha granules are the most numerous.

Explain the rationale for use of proteosome inhibitors as a therapeutic modality for MM

Proteasomes are multi-protein proteolytic complexes found in the cytoplasm of eukaryotic cells. Each proteasome is a barrel shaped complex that functions in the degradation of damaged or misfolded proteins. Proteasome function is particularly important for cells that produce large quantities of proteins destined for secretion, such as occurs in neoplastic plasma cells. In multiple myeloma, the neoplastic plasma cells exhibit a high production rate of secreted antibody proteins, and they are highly dependent on proteasome activity to prevent overload of the rough endoplasmic reticulum (RER). Recall that in all eukaryotic cells, the RER is the location where proteins destined for secretion are synthesized. Since FDA approval of the first proteasome inhibitor, bortezomib, in 2003, proteasome inhibitors have become an effective therapeutic modality for multiple myeloma. By blocking proteasome function, the inhibitors induce an accumulation of misfolded proteins in the neoplastic cells. This leads to a condition called ER stress as the RER becomes backlogged with antibody proteins, and ultimately, the cell is triggered to undergo apoptosis.

List hemostasis inhibitors produced by resting endothelial cells

Recall that an intact endothelial cell lining is anti-thrombotic and profibrinolytic. These characteristics are attributed to a variety of substances that are either actively secreted or actively expressed on the membrane surfaces. In non-injured blood vessels, endothelial cells secrete inhibitors of primary hemostasis, as indicated here. EC Primary hemostasis inhibitors: Prostacyclin (PGI2) promotes vasodilation and inhibits platelet activation. Nitric oxide (NO) promotes vasodilation and inhibits platelet recruitment. ADPase is an enzyme that breaks down ADP. Recall that non-metabolicADP is an agonist stored in platelet dense granules.

Secondary Hemostasis Define zymogen, tissue factor, contact factor, and thrombus

Secondary hemostasis requires a series of highly coordinated, enzyme-mediated reactions involving clotting factors, also known as coagulation factors. Most clotting factors are synthesized by the liver, and in the absence of blood vessel injury, they circulate as non-active zymogens. A zymogen is a pro-enzyme that requires specific activation in order to assume its particular enzymatic function. Ultimately, secondary hemostasis results in conversion of soluble fibrinogen to insoluble fibrin, providing reinforcement of the platelet plug. The traditional cascade theory of secondary hemostasis describes interaction of three pathways which form enzyme complexes on phospholipid surfaces of platelets or surfaces of exposed sub-endothelium. - The extrinsic pathway is triggered by tissue factor, a membrane surface protein expressed on the surface of sub-endothelial cells. Tissue factor is not present in the blood plasma under normal conditions. - The intrinsic pathway is triggered by factors and co-factors that are all normally present in the blood plasma. - The common pathway follows from a convergence of the extrinsic and intrinsic pathways.

Quantitative Platelet Disorders thrombocytopenia induced by drugs

Some forms of thrombocytopenia are induced by drugs, for which numerous drugs have been implicated. Patients experience symptoms of excessive bleeding that usually appear suddenly and can be severe; and the resulting platelet count can be very low, less than 20 times 10to the sixth per liter. However, simple removal of the drug usually halts the thrombocytopenia and bleeding symptoms. There are various mechanisms by which drugs can cause thrombocytopenia, including generalized hematopoietic suppression, selective suppression of platelet production, or increased platelet destruction by antibody coated platelets. One mechanism for drug induced ITP is through formation of a neoantigen. Drugs such as anti-malaria therapiequinine and quinidine bind to proteins on the platelet surface to formnovel antigenic structures that can be recognized by autoreactive antibodies. Another example of drug induced ITP is heparin induced thrombocytopenia (HIT) Heparin, an anticoagulation drug, binds platelet factor 4, and the heparin/PF4complex attaches to anti-heparin antibodies. The Fc portion of the antibody binds to platelet Fc receptors. About 3 percent of patients on heparin therapy develop ITP. Note that this complex can also induce platelet activation and thrombosis.

hemostasis diagnostic testing: Describe the use of PT, APTT, TT, and D-dimer tests for assessment of secondary hemostasis and fibrinolysis

The activated partial thromboplastin time (APTT) test is used to assess inherited or acquired defects in the intrinsic and common pathways. In this test, a sample of patient platelet poor plasma (PPP) is combined with a reagent called "partial thromboplastin." Note that partial thromboplastin does NOT include thromboplastin at all. Instead, the reagent includes phospholipids and calcium ions. The phospholipids mimic platelet surfaces as required for activation of the intrinsic 10ase complex. The test is conducted in a test tube with a negatively charged surface to activate factor 12. The time it takes for fibrin formation is the APTT time. The reference interval for APTT time is 25 to 35 seconds, and a prolonged APTT occurs if there is a deficiency in factor 12, 11, 9, 8, prothrombin, or fibrinogen. It is also prolonged when there is a VWF deficiency due to the destabilizing effect this would have on factor 8. Thrombin time (TT) is used to measure the conversion of fibrinogen to fibrin, and involves addition of purified thrombin to a patient PPP sample. The reference range for fibrin formation is 15-19 seconds; and a prolonged time is indicative of a fibrinogen deficiency or dysfunction.

hemostasis diagnostic testing: Analyze platelet aggregation curves

The data for platelet aggregometry tests are presented as platelet aggregation curves. These curves indicate the change in optical density (OD) on the Y axis, as a function of time in minutes on the X axis. Note that optical density goes from high to low on the Y axis. On the right is a typical curve generated for normal platelets stimulated by the agonist ADP. -Curve indicates change in OD of the PRP over time (minutes) How can we interpret this curve? Baseline transmission, with an OD value of about 0.9, is reflected in the first part of the curve, labeled (a). Next, we see a dip (labeled b), which coincides with addition of the platelet agonist and the ensuing platelet shape change. The steep decrease in optical density is evident first through a primary wave labeled c, during which small platelet aggregates are forming; and subsequently a secondary wave (labeled d) is representative of complete aggregation, with a final OD reading of about 0.2. This secondary wave is attributed to endogenous ADP release from the activated platelets.

Diagram each of the three classical coagulation pathways

The intrinsic pathway includes factors 12, 11, 9, and 8, and requires calcium ions. *aka Xii, Xi, iX, Viii (Ca++)* These factors are all present in the blood. The reaction cascade culminates in formation of the intrinsic 10-ase (Xase) complex on platelet surfaces. Intrinsic 10-ase is composed of factor 9a and co-factor 8a. In the presence of calcium ions (Ca++), the complex catalyzes conversion of factor 10 to 10a. Note that factor 8 is composed of two sub-units depicted as 8aL and 8aH in the diagram. In the blood, VWF stabilizes non-activated cofactor 8; and thrombin converts 8 to 8a. Note that a factor 8 deficiency underlies the hemophilia A bleeding disorder, and a factor 9 deficiencyunderlies the hemophilia B bleeding disorder, thus underscoring the in vivo significance of these two factors that comprise the intrinsic 10-ase. Factor 8 or FViii deficiency: hemophillia A Factor 9 or FiX deficiency: hemophilia B

Distinguish between leukemic, extranodal, and nodal subtypes

The major WHO subtypes of matureT/NK lymphomas are listed here. These sub-types can be further classified based on their anatomical locations. The leukemic sub-types are comprised of circulating cells that affect the peripheral blood and bone marrow. The extra-nodal types form lymphoma type tumors that are not confined to lymph nodes. These neoplastic cells may form tumors in locations such as the spleen,liver, MALT, or the skin, depending on the particular sub-type. Nodal types are lymphomas that occur in lymph nodes. In this session we will consider one example from each category: adult T cell leukemia/lymphoma extra-nodal NK/T cell lymphoma nasal type; peripheral T cell lymphoma, not otherwise specified.

Quantitative Platelet Disorders Distinguish between autoimmune and alloimmune ITP

The major antigenic targets of antiplatelet antibodies are the GP2b/3a (CD41/CD61) fibrinogen receptor and the GP1b/9(CD42) VWF receptor, which are both abundant platelet surface proteins. ITP Treatment: Treatment of ITP is usually not required for children, as most experience spontaneous remission within 2 to 6 weeks of disease onset. In general, a platelet count of less than 30 times 10 to the sixth per liter is the threshold for treatment. Initial treatment may include administration of intravenous immunoglobulins to block Fc receptors and thus inhibit macrophage uptake of platelets. Persistent and chronic cases may be treated with immunosuppressive therapies such as anti-inflammatory corticosteroids and rituximab anti-CD20 antibodies to destroy B lymphocytes. Finally, if patients do not respond to these therapies, splenectomy can increase the level of circulating platelets.

Describe clinical and neoplastic cell characteristics of ATLL, NNKTL, and PTCL, NOS

The peripheral T cell lymphoma, not otherwise specified (PTCL, NOS) sub-type is a broad category; but it is the most common T cell lymphoma sub-type, accounting for about 30 percent of the T cell lymphomas. Most patients present with lymph node involvement, but neoplastic cells may also occur in the bone marrow, spleen, and liver. Immunophenotyping and cytogenetic analyses reveal that these cells are positive for standard T cell markers such as CD3 and they have a fully rearranged TCR. In nodal cases, the neoplastic cells are usually CD4 positive and CD8 negative. Cytogenetic analyses have revealed that these cells have complex karyotypes, with various chromosomal deletions, translocations, and trisomies. The disease course is usually aggressive, as there is a poor response to standard chemotherapy, and frequent relapses. The five-year overall survivalf or PTCL, NOS is less than 32%. Progress in treatment advances for mature T and NK cell neoplasms has been greatly hindered by the rarity and heterogeneity of these various disease subtypes. Standard chemotherapy continues to be a first line treatment, but there is a high relapse rate in these patients. Hematopoietic stem cell transplantation has prolonged remission, but there is also a high rate of relapse. There are global efforts to develop immunotherapies for treatment of these conditions, with numerous monoclonal antibody therapies in various phases of clinical development.

Describe the structures of arterioles, venules and capillaries

The vasculature provides an extensive distribution system for the blood, and serves as a critical conduit for the transport of oxygen and carbon dioxide, nutrients, waste products, and regulatory agents such as hormones. The structure and connectivity of the vasculature allows for an uninterrupted flow of blood. The vasculature consists of three types of blood vessels: arteries, veins, and capillaries. The arteries enable movement of blood away from the heart and regulate blood pressure via vasodilation. The veins enable movement of blood towards the heart. Capillaries are the smallest type of blood vessel, and they mediate gas and nutrient exchange with the tissues. Hemostasis processes occur primarily in arterioles, the smallest arteries; and in venules, the smallest veins. We examine the structure of these vessels next.

List the three stages and main components of hemostasis

The word hemostasis is derived from the Greek words heme, meaning blood, and stasis, which means stop. Hemostasis is a circulation property that maintains blood as a fluid within the blood vessels and prevents excessive blood loss upon injury. Hemostasis is under control of a complex and highly coordinated circuit of activators and inhibitors that influence the functions of blood vessels, platelets, and plasma clotting factors. Hemostasis can be described in terms of three distinct stages of activity following vascular injury: 1) In primary hemostasis, platelets adhere to the site of vascular injury and connect together with fibrinogen protein, leading to formation of a platelet plug. 2) In secondary hemostasis, activated clotting factors mediate the conversion of soluble protein fibrinogen into insoluble fibrin, to form the more stable fibrin-platelet plug. 3)Finally, after the vessel has healed, the process of fibrinolysis mediates dissolution of the fibrin clot. There are three major categories of hemostasis components involving the vasculature, the cellular elements, and the proteins: Vascular components include arterioles, venules, and capillaries. Cellular element components include vascular endothelial cells and platelets. Protein components include fibrin-forming proteins, fibrinolytic proteins, and inhibitors of all hemostasis stages.

WHO list of mature B cell neoplasm

These categories are based on various criteria, including presentation as either leukemia or lymphoma, location of the growth, immunophenotype, morphology, and genetics. This list includes the plasma cell neoplasms, which secrete abundant amounts of monoclonal immunoglobulin proteins. Lymphomas are further categorized based on location as being nodal, with lymph node involvement; or as extra-nodal, where lymphoma growth does not occur in lymph nodes. For example, in the spleen, thymus, or MALT, as occurs in splenic marginal zone lymphoma, primary mediastinal B cell lymphoma,a nd MALT lymphoma. CLL/SLL = 19% Follicular lymphoma= 12% DLBCL= 28% Burkitt Lymphoma = 1%

Define: monoclonal gammopathy, Bence-Jones proteins, and M spike

This session examines plasma cell derived neoplasms. Plasma cell neoplasms are composed of malignant antibody secreting B lymphocytes. Excessive production of monoclonal antibodies, a condition called monoclonal gammopathy, is a hallmark of plasma cell neoplasms. As a consequence, normal immunoglobulin production by healthy B cells is usually decreased, leading to functional hypogamma globulinemia, and associated immune deficiency. Monoclonal gammopathy can be observed by serum protein electrophoresis, such as in the example provided here. We see a prominent albumin protein band at the top in all three lanes. In normal serum, such as in lanes 1 and 2, the gamma globulins (antibodies)are polyclonal and diverse in structure and thus, do not migrate as a distinct band. In lane 3 we see a distinct monoclonal serum antibody band from a patient with a plasma cell neoplasm. On the right we see results of a densitometry scan of lane 3, which reveals a prominent monoclonal (M) spike that is characteristic of plasma cell neoplasms. Furthermore, in plasma cell neoplasms, monoclonal antibody proteins may be detected in the urine. We see here a urine sample electrophoresis revealing a lower band corresponding to IgG monoclonal antibody proteins containing both heavy and light chains. Above are bands corresponding to free light chain proteins, called Bence-Jones proteins, that are secreted in excess by the neoplastic plasma cells.

Distinguish between MGUS, plasmacytoma, SMM, and MM

This table compares four major sub-types of plasma cell neoplasms: Monoclonal gammopathy of undetermined significance, called MGUS, is a benign, asymptomatic condition in which the M spike levels are below 30 grams per liter of blood, and there are less than 10% bone marrow plasma cells. MGUS incidence increases with age, and occurs in about 3% of people over the age of 70. Plasmacytoma is a malignant, asymptomatic condition characterized by localized plasma cell tumors that form in the bone or in other tissues. An M spike may be detected in this condition. Patients with asymptomatic disease may be stable for many years. Smoldering multiple myeloma (SMM) is also a malignant, asymptomatic plasma cell neoplasm, characterized by a prominent M spike greater that 30 grams per liter of blood and/or the presence of 10% bone marrow plasma cells. However, the presence of these cells does not cause organ or tissue impairment. Multiple myeloma (MM), also referred to as plasma cell myeloma, is a malignant, symptomatic cancer that can cause significant, widespread organ damage and immune deficiency. A hallmark of multiple myeloma is extensive bone marrow infiltration of antibody producing neoplastic plasma cells. A prominent M spike and Bence-Jones proteins are commonly detected in multiple myeloma. Although MGUS, plasmacytoma, and SMM are asymptomatic, each of these conditions may potentially progress to symptomatic multiple myeloma.

Bleeding Disorders Predict lab findings in the various main categories of hemostatic disorders

This table summarizes expected lab findings for various categories of hemostatic disorders. In vascular disorders, platelet counts, PT, and APTT tests re normal. Further tests would be conducted to rule out a platelet function disorder. In platelet functional disorders, platelet counts may be normal or abnormal, while PT and APTT tests are normal. Further testing would include PFA, aggregation tests, and flow cytometry to identify the particular platelet abnormality. In platelet quantitative disorders, platelet counts are abnormal, while PT and APTT tests are normal. Further testing is dependent on the suspected cause of the quantitative defect. In secondary hemostasis disorders, platelet counts are normal, while one or both of the PT and APTT tests are abnormal, depending on the coagulation factor (or factors) involved. Further testing would include thrombin time, mixing studies to rule out inhibitors, and specific factor assays. In a factor 13 disorder, all parameters, platelet counts, PT, and APTT tests are normal. Definitive diagnosis of this condition requires a specific factor 13 assay.

Explain how TAFI inhibits fibrin degradation

To understand fibrinolysis inhibition mediated by TAFI, we need to take a closer look at plasminogen activation, as illustrated here. Plasminogen binds to a lysine (K) residue in the carboxyterminus of fibrin, followed by tPA induced conversion of plasminogen to plasmin; and plasmin then cleaves the fibrin polymer. The thrombinactivatable fibrinolysis inhibitor TAFI is activated by the thrombin/thrombomodulin complex. TAFI inhibits plasmin formation by removing the fibrin C-terminal lysine. Thus, plasminogen is unable to bind to the fibrin substrate. Fibrinolysis is further subject to negative regulation through the action of anti-plasmin inhibitors. Alpha 2 antiplasmin, which is stored in platelet alpha granules, is the principal inhibitor of plasmin. Alpha 2 macroglobulin isa back-up inhibitor when excessive plasmin is generated.

List therapeutic strategies for mature lymphoid leukemia/lymphoma

Treatment of CLL/SLL depends on disease stage and severity of symptoms. Early stage CLL patients are simply monitored for disease progression. Some patients experience what is called Richter's transformation, when they transition to an aggressive, large, B cell lymphoma. This occurs in approximately 2 to 8 percent of patient Symptomatic patients may be treated with traditional chemotherapy or with Venetoclax, the Bcl2 inhibitor. Alternatively, rituximab, ananti-CD20 monoclonal antibody immunotherapy, may be of benefit. We examine this drug more closely next. Rituximab is an anti-CD20 monoclonal antibody that is derived from a mouse antibody that was subsequently humanized by recombinant DNA methods so that the Fc portion of the antibody is not rejected by the human immune system. As indicated by the red arrows on the right, CD20 is a cell marker expressed by pre-B and mature B cells. Following infusionin to CLL patients, the antibody binds to CD20 on B cells and induces ADCC, leading to depletion of neoplastic cells. An unfortunate consequence of this treatment is that healthy B cells are also depleted. However, the B cell population will be replenished after the therapy is discontinued.

Describe clinical presentation of MM in the bone marrow and blood; and describe the basis of CRAB symptoms

Various cytogenetic abnormalities are commonly found in multiple myeloma cells, for which two genetic subtypes have been described based on chromosome content. In the hyperdiploid subtype, chromosomal trisomy may be observed. The non-hyperdiploid subtype is characterized by the occurrence of translocations, deletions, and complex karyotypes. The most frequent translocations involve IgH and cyclin-D1 gene regions, leading to hyper activation of cyclin D1, which then acts as a significant oncogene driver. However, there is not yet a clear-cut association between these cytogenetic characteristics and disease prognosis and treatments. Prognosis and treatment Patients with asymptomatic plasma cell neoplasms may be stable for years, but they do often progress to symptomatic disease. Disease prognosis for symptomatic patients with multiple myeloma is poor, with a median survival time of 3 to 5 years. Symptomatic disease patients under 65 may be treated with high-dose chemotherapy and possibly hematopoietic stem cell transplantation, whereas patients over 65 will more likely be treated with a more standard chemotherapy regimen. Additionally, proteasome inhibitors have also become an important modality for treatment of multiple myeloma, as will be explained next.

Describe the opposing procoagulant and anticoagulant roles of thrombin

We conclude this discussion of hemostasis regulation with a summary of thrombin's various hemostasis functions. Thrombin is a unique hemostatic factor in that it has both pro-and anti-coagulant effects and it plays a central role in hemostatic balance. In its procoagulant capacity, thrombin serves the following functions. Critically, thrombin stabilizes blood clots by catalyzing conversion of fibrinogen to fibrin and by activating factor 13. Thrombin promotes primary hemostasis through activation of platelets, and thrombin functions in positive feedback of secondary hemostasis through activation of factors 5, 8, and 11 and activation of endothelial cell secretion of VWF. Further, thrombin exerts an anti-fibrinolytic, clot promoting function through activation of TAFI. In its anticoagulant capacity, thrombin serves the following functions. Thrombin exerts inhibitory effects on secondary hemostasis through the thrombomodulin/protein C/protein S system, which inactivates clotting factors 5a, 8a, and 10a. Further, thrombin stimulates endothelial cells to secrete tPA, thus exerting a pro-fibrinolytic effect. It also stimulates endothelial cell release of prostacyclin and nitricoxide, both of which are platelet antagonists and vasodilators; thus, exerting inhibition of primary hemostasis.

Quantitative Platelet Disorders Recognize examples of non-immune mediated destruction

We conclude with a brief consideration of conditions that are associated with non-immune mediated destruction of platelets. In some conditions, platelets are over-activated and consumed, leading to symptoms of both thrombosis and bleeding. This occurs in Disseminated intravascular coagulation (DIC), thromboticthrombocytopenic purpura (TTP), and Hemolytic uremicsyndrome (HUS). DIC and TTP will be covered further in a later session in the context of thrombosis; and recall that HUS was covered previously in the context of hemolytic anemia. Thrombocytopenia may also occur by mechanical destruction of platelets if they adhere to artificial heart valves Finally, thrombocytopenia can occur in pregnancy due to platelet distribution changes or various hypertension complications.

Distinguish between precursor and mature lymphoid leukemia/lymphoma

We see here a representation of T cell and B cell development described previously. Recall that precursor B and T cells are derived from primary lymphoid tissues, the bone marrow, and the thymus gland, respectively; and the mature cell types circulate through the peripheral blood and the secondary lymphoid tissues, including the lymph nodes, the spleen, and MALT. Accordingly, as covered previously, in precursor lymphoid neoplasms, ALL/LBL, the predominant neoplastic cells are B or T lymphoblasts. The malignant cell characteristics are similar to immature cells in the bone marrow or thymus gland. By contrast, in the mature lymphoid neoplasms, the predominant neoplastic cells are B or T or NK cells that have mature characteristics similar to those that naturally occur in the peripheral blood or secondary tissues. The genesis of mature lymphoid neoplasms is a multi-step process, resulting in neoplasms that occur mostly in people over the age of 60. As is true for all cancers, these neoplasms develop as a result of acquired accumulation of oncogene and tumor suppressor gene mutations. The precise cancer initiating cell type is not well established, and likely varies from case to case. However, it is understood that cross-over to malignancy occurs at a more mature stage of development as compared to the precursor neoplasms. Diagnosis involves the integration of a range of clinical information, including morphologic appearance assessed from peripheral blood, bone marrow, or tissue biopsy; along with immunophenotype and molecular gene and chromosome analysis.

Describe clinical presentation of MM in the bone marrow and blood; and describe the basis of CRAB symptoms

We will now turn our focus to multiple myeloma. A critical pathological characteristic of multiple myeloma is the effect this disease has on the bone marrow. Normal hematopoietic precursor development is impeded as the bone marrow becomes overrun with neoplastic plasma cells, such as indicated in the biopsy image shown here. Furthermore, multiple myeloma patients commonly present with painful bone lesions due to the infiltration of these cancerous plasma cells. On the right we see a skull X-ray from a patient with multiple myeloma. The image reveals multiple lytic bone lesions giving the skull a "moth-eaten" appearance. Although few malignant plasma cells are observed in the peripheral blood of multiple myeloma patients, rouleaux (stacking of RBCs) may be observed in a blood smear, such as the one shown here. Rouleaux occurs due to high levels of monoclonal antibody proteins in the blood. These are referred to as paraproteins. The plasma itself may also stain blue due to a large quantity of paraproteins. Multiple myeloma symptoms are commonly described by the acronym,CRAB, representing the following disease characteristics. The C is for hypercalcemia, elevated blood calcium levels resulting from damaged bone. The R is for renal insufficiency, as kidneys are damagedby the excessive antibody proteins in the blood. The A is for anemia, as themyeloma cells in the bone marrow disrupt erythropoiesis Note that patients also experience leukocytopenia and with thrombocytopenia since the bone marrow environment is severely compromised. The B is for the painful bone lesions that occur as a result of the cancerous plasma cell infiltration.

Bleeding Disorders Describe various types of inherited and acquired vascular disorders

acquired vascular disorders We conclude this session with a brief consideration of acquired vascular disorders, for which a variety of conditions may affect the integrity of vessel walls or endothelial cells. There are four major categories of acquired vascular disorders as listed in this table: Purpura resulting from decreased connective tissue, Purpura associated with dysproteinemias, Purpuraresulting from vasculitis, and Miscellaneous causes of purpura. Purpura resulting from decreased connective tissue occur as a result of aging in senile purpura, when there is a natural degeneration of the skin matrix. Decreased connective tissue associated purpura also occur in Cushing syndrome, a hormonal abnormality in which patients producetoo much cortisol. Purpura may also occur in patients on glucocorticoid therapy, and in patients with scurvy, caused by a vitamin C deficiency. Purpura associated with dysproteinemias may occur in conditions associated with paraproteins, such as occur in multiple myeloma. Cryoglobulinemia refers to cold-induced clumping of abnormal blood proteins, and cryofibrinogenemia refers to circumstances in which exposure to the cold triggers abnormal fibrinogen precipitation. Amyloidosis is a condition in which misfolded proteins build up in the tissues. Purpura are thought to occur when subendothelial amyloid deposits weaken blood vessels.

Bleeding Disorders Describe various types of inherited and acquired vascular disorders

acquired vascular disorders (cont.) Purpura resulting from vasculitis is associated with inflammation of blood vessels. There are a wide range of conditions that can trigger vasculitis. Immunoglobulin Avasculit is is triggered when IgA antibodies accumulate in small blood vessels. The vessels become inflamed and leak blood, resulting in a red or purple rash. Various infections and drugs may also trigger vasculitis There are various miscellaneous causes of purpura. Mechanical purpura are caused by stressors such as intense exercise, coughing spasms, or seizures. Artificially induced purpura may be caused by physical abuse or self-inflicted trauma. Easy bruising syndrome is a benign condition of vascular fragility that is thought to result from hormonal effects on blood vessels. Purpura fulminans occurs in newborns with clotting factor abnormalities, or it can occur in patients on anti-coagulation therapy. This condition can be serious if thrombi form and blood vessels become occluded, leading to tissue necrosis and possibly death.

Describe the structures of arterioles, venules and capillaries

arteriole structure.: There is a central cavity called the lumen, where blood flow occurs. The lumen is surrounded by a single layer of flat endothelial cells that separate the blood from the tissue below. The vascular endothelial cells provide significant hemostatic functions mediated by various surface structures and secreted compounds. - The innermost layer of the arteriole is called the tunica intima. It includes the endothelial cell layer, a thin basement membrane of connective tissue, and an internal elastic membrane that enables flexibility. - Below the tunica intimais the tunica media. This is a much thicker layer that includes connective tissue composed of collagen, proteoglycans, reticular fibers, and elastin fibers. Importantly, there are smooth muscle cells interspersed through the tunica media. - Finally, the tunica media is surrounded by the tunica adventitia, which is composed of fibroblast, collagen, and elastin fibers. The overall structure of venules is very similar to that of arterioles, except that the tunica media is thinner than that of arterioles, and it contains fewer elastin fibers and fewer smooth muscle cells. The capillaries connect the arterioles and venules. Each capillary has a characteristically small lumen that is just large enough to accommodate single-file movement of RBCs. a single endothelial cell wraps around the perimeter of the small lumen. There is a thin basement membrane of connective tissue that surrounds the endothelial cells along the length of the capillary. Collectively, capillaries makeup the largest surface area of blood vessels, as they must branch out and infiltrate into all of the body's various tissues for efficient exchange of oxygen and CO2, and to facilitate exchangeo f nutrients and waste.

Diagram each of the three classical coagulation pathways

common pathway: Extrinsic 10ase and intrinsic 10ase complexes both contribute to the activation of 10 to 10a. Next, as described earlier, 10a converts factor 5 to 5a; and 10a and 5a form the prothrombinase complex, which converts prothrombin to thrombin = Extrinsic Xase and intrinsic Xase convert X (10) to Xa(10a) Xa converts V (5) to Va (5a) Xa (10a) + Va (5a) = prothombinase Prothrombinase to thrombin

Secondary Hemostasis Diagram each of the three classical coagulation pathways

extrinsic pathway and the role of tissue factor: Also known as thromboplastin or factor 3 or CD142, tissue factor is a transmembrane glycoprotein on sub-endothelial cells including smooth muscle cells and fibroblasts. Tissue factor is not normally present in the blood, and it is not actively secreted. However, micro-vesicles carrying tissue factor are detected in the blood in situations of traumatic disease, such as in cancer, sepsis, or as a consequence. of chemotherapy. Extrinsic pathway is triggered when blood vessel injury enables factor 7 to make contact with tissue factor. The combination of tissue factor, factor 7, and calcium ions (Ca++) brings about activation of factor 7, represented as 7a . The extrinsic Xase (10ase) complex, composed of tissue factor and 7a, activates factor 10 to form factor 10a. Here we again see the extrinsic Xase (10ase) complex, composed of tissue factor and 7a (Viia). The complex assembles where tissue factor is expressed on the surface of sub-endothelial cells such as fibroblasts. The complex mediates conversion of factor 10 (X) into 10a (Xa). Next, factor 10a (Xa) converts factor 5 (V) to 5a (Va), and they team up to form pro-thrombinase. Factor 5 is composed of two sub-units depicted as VaL and VaH in the diagram. Prothrombinase mediates conversion of prothrombin (factor 2)into thrombin (factor 2a). Prothrombinase, composed of factor 10a and its cofactor 5a, requires calcium ions to mediate prothrombin conversion to thrombin. The reaction results in what is called the thrombin "burst" as roughly 1000 molecules of thrombin are formed per one prothrombinase complex. Subsequently, thrombin, which is a serine protease, cleaves fibrinogen to form fibrin.

Primary Hemostasis Explain how the primary platelet plug is formed

final step - platelet aggregation: Aggregation is triggered by a number of agonists, including ADP, thromboxaneA2, and thrombin. The agonists induce platelets to express high affinity fibrinogen receptors, G2b/3a (CD41/CD61). This activation requires calcium ions. Fibrinogen protein is secreted by the liver. It is a trimeric protein with three structural domains, two D domains with an E domain between them, denoted as D-E-D domains. The D domains form non-covalent bonds with the G2b/3a (CD41/CD61) platelet receptors. Thus, fibrinogen acts as a bridge that interconnects the platelets. The interaction of fibrinogen proteins with the many platelets that have gathered at the site of blood vessel injury results in formation of a mesh network of platelets and fibrinogen proteins. This constitutes the primary platelet plug. Included here is a scanning EM image of a platelet plug, with about 20 platelets clumped together. The primary platelet plug slows down the loss of blood, but it is unstable and easily dislodged. Importantly, the activated platelets provide appropriate surfaces on which coagulation factors subsequently bind and become activated to carry out secondary hemostasis. Secondary hemostasis requires a series of enzyme mediated reactions to convert soluble fibrinogen into insoluble fibrin, which reinforces the platelet plug.

Secondary Hemostasis Describe the role of thrombin in conversion of fibrinogen into a fibrin polymer

final steps of secondary hemostasis, where thrombin mediates conversion of fibrinogen to fibrin: Additionally, thrombin activates factor 13 to 13a, and 13a transforms fibrin into cross-linked fibrin. To fully understand these last steps, it is important to first evaluate the structure of fibrinogen, a protein produced by the liver. Fibrinogen is composed of three pairs of proteins, A-alpha, B-beta, and gamma. The overall structure of a fibrinogen protein is described in terms of three nodules, D-E-D. In the conversion of fibrinogen to fibrin, thrombin removes four small alpha/beta peptide extensions protruding from nodule E. Following removal of alpha and beta peptides from nodule E, fibrin monomers spontaneously assemble into a fibrin polymer. Next, factor 13a (Xiiia), which is a transglutaminase enzyme, catalyzes covalent cross-linking between adjacent D nodules to form an insoluble fibrin mesh. The final result of secondary hemostasis is the formation of a stable blood clot, which will remain in place until the underlying vessel and tissue have been repaired. The clot must then be dissolved by the process of fibrinolysis. A thrombus is a blood clot that inappropriately forms within a blood vessel in the absence of injury. Thrombus formation can be dangerous if blood flow is obstructed.Here we see a scanningEM image of a thrombus. The cross-linked fibrin has formed a string-like mesh in which many RBCs have become trapped.

Primary Hemostasis List the key platelet receptors involved in platelet adhesion, activation, and aggregation

formation of the primary hemostatic plug: Platelets have a variety of integral membrane proteins that serve as receptors for VWF, fibrinogen, collagen, and other ligands. These receptors function in adhesion, activation, and aggregation steps of primary hemostasis. This table summarizes three important receptors that bind to the ligands collagen, VWF, and fibrinogen, as indicated in the first column. The collagen receptor is referred to as either GP1a/2a or as CD49b/CD29. This receptor mediates platelet adhesion to the sub-endothelium when blood is flowing slowly, so there is a low shear rate, and platelets are not easily dislodged. The VWF receptor is referred to as either GP1b/9 or as CD42. This receptor mediates platelet adhesion to the sub-endothelium when blood is flowing quickly, with a high shear rate. This stronger interaction is required to prevent platelets from being dislodged. The fibrinogen receptor is referred to as either GP2b/3a or as CD41/CD61. This receptor mediates platelet aggregation. In platelet aggregation, fibrinogen proteins serve as bridges that hold the platelets together.

Explain vasoconstriction and vasodilation

important blood vessel hemostasis functions, beginning with vasoconstriction and vasodilation. Vasoconstriction is an immediate response that occurs following injury of a blood vessel. In vasoconstriction, there is a narrowing of the lumen and a decreased blood flow to minimize blood loss. The process also brings platelets and clotting proteins closer together.Although vasoconstriction occurs immediately after blood vessel injury, it usually lasts for just a short time. Vasoconstriction is controlled by an array of regulatory substances, including serotonin and thromboxaneA2 and endothelin- By contrast, in vasodilation, there is a widening of the lumen and an increased flow of blood; as occurs during an inflammatory response. Let's now focus on the endothelial cell hemostatic functions. These cells serve as a dynamic interface between the vessel wall and the circulating blood, and they are principal regulators of many vascular functions, including the inflammatory response, covered previously, and thromboregulation, to facilitate or inhibit thrombus (blood clot) formation. Generally, a healthy resting endothelium is non-thrombogenic and profibrinolytic, a condition that is unfavorable for blood clot formation; whereas, a damaged endothelium is thrombogenic and anti-fibrinolytic, a condition that is favorable for blood clot formation.

Functional Platelet Disorders Compare and contrast underlying defects and diagnostic test results for the following inherited platelet abnormalities: Bernard-Soulier syndrome, von Willebrand disease, activation disorders, Glanzmann thrombasthenia, and Scott syndrome

inherited platelet abnormalities Bernard-Soulier syndrome (Adhesion disorder) Aggregation tests can also be useful in diagnosis of Bernard-Soulier syndrome. Since the platelets have no activation or fibrinogen receptor defects, there is normal platelet fibrinogen mediated aggregation in response to standard agonists: ADP, collagen, and epinephrine. This is reflected in the three aggregation curves below. Recall, that in aggregometry, aggregation curves reflect the change in optical density on the Y axis, and time in minutes elapsed on the X axis. In contrast to the aggregation observed with agonists, there is no effect in the presence of a compound called ristocetin. Ristocetin binds to GP1b/9(CD42) in combination with VWF and should cause agglutination of the platelets. However, in Bernard-Souliersyndrome, where CD42 is absent, no platelet agglutination occurs with ristocetin alone or with ristocetin and exogenous VWF. This is reflected by the flat line in the last two aggregation curves below.

Functional Platelet Disorders Compare and contrast underlying defects and diagnostic test results for the following inherited platelet abnormalities: Bernard-Soulier syndrome, von Willebrand disease, activation disorders, Glanzmann thrombasthenia, and Scott syndrome

inherited platelet abnormalities Bernard-Soulier syndrome (Adhesion disorder) Bernard-Soulier syndrome (BSS) is a deficiency or defect in the VWF receptorGP1b/9, also called CD42. In von Willebrand disease (VWD), the platelets themselves are normal, but there is a deficiency or defect in the plasma von Willebrand factor, VWF. In Bernard-Soulier syndrome, platelets have decreased levels or functional abnormalities in the GP1b/9(CD42) VWF receptor. Bleeding symptoms in BSS are similar to thrombocytopenia. Depending upon the specific nature of the underlying mutation, severity ranges from mild bruising, to severe, recurrent mucosal and sub-cutaneous bleeding. CBC and blood smear evaluation indicate macrothrombocytopenia, with moderate to severe thrombocytopenia, and large platelets with a heterogenous size distribution. Here we see a bloods mear from a patient with Bernard-Soulier syndrome. The green arrows indicate the presence of numerous abnormally large platelets. Laboratory reflex test results for BSS include increased PFA closure time, and flow cytometry may indicate decreased expression of GP1b/9 (CD42) on platelet surfaces.

List the various fibrinolysis inhibitors

inhibitors of fibrinolysis: The process must be well regulated to prevent systemic proteolysis and bleeding episodes. There are several principal inhibitors that act at the plasminogen activation step. PLG activation step: These are the plasminogen activator inhibitors (PAI1 and PAI2) and thrombinactivatable fibrinolysis inhibitor (TAFI). Inhibitors that act directly on plasmin are alpha 2 anti-plasmin and alpha 2 macroglobulin. Plasminogen activator inhibitor one (PAI1) is produced by activated endothelial cells. PAI1 is the primary physiological inhibitor of tPA and uPA. Note that PAI2 is a minor plasminogen activator inhibitor, found mostly in the placenta. PAI-2 levels in the blood are low, except during pregnancy.

Diagram each of the three classical coagulation pathways

intrinsic pathway. (Cont.) There are several other important points to consider regarding the intrinsic pathway. First, the associated clotting factors have historically been called contact factors because when first evaluated in vitro, these factors were activated by contact with charged surfaces, such as glass in there action test tubes. Factors 12 ,11, and another factor called PK (orpre-kallikrein) are required for in vitro coagulation. Although not designated with a Roman numeral, PK is classified as a component of the intrinsic pathway because it is activated by factor 12a to form Kallikrein, which in turn can induce reciprocalactivation of factor 12. Despite their requirement for in vitro coagulation, deficiencies in 12 or PK cause no bleeding phenotype; and only half of patients with a factor 11 deficiency experience bleeding abnormalities following surgery or injury. Since factor 12and 11 deficiencies are either asymptomatic or just mildly symptomatic, it has been suggested that these early steps are not essential for coagulation in vivo. Note that thrombin can activate factor 11; and the extrinsic 10ase complex can activate factor 9. Recall that the extrinsic 10ase is composed of tissue factor and factor 7a. In addition to activation of factor 10, this enzyme complex can also activate factor 9 to 9a. Thereby, it by passes the requirement for upstream activation by 12 and 11, and it enables assemblyof the intrinsic 10ase. Extrinsic Xase activation of iX (9) to iXa (9a)

Distinguish between precursor and mature lymphoid leukemia/lymphoma

mature lymphoid neoplasms: This session is the first of a two-part series on mature B cell malignancies. Mature lymphoid neoplasms include a heterogenous group of disorders in which the predominant neoplastic cell types are either mature B orT lymphocytes or NK cells. Traditionally, mature lymphoid neoplasms have been grouped according to the anatomical distribution of the disease. As considered previously, in leukemia, neoplastic cells predominate in the bone marrow and/or the peripheral blood; whereas in lymphoma, neoplastic cells form a palpable mass (a tumor) in secondary lymphoid tissues or other sites. Lymphomas are further characterized as Hodgkin type or non-Hodgkin type. We describe Hodgkin lymphoma is a later session; and by definition, all other lymphoma types are in the non-Hodgkin category.

Secondary Hemostasis Explain positive feedback mediated by thrombin & factor Xa

positive feedback occurs at several levels. Notably, thrombin itself activates factors 11 and 8 of the intrinsic pathway, and factor 5 of the common pathway. Thrombin also activates platelets, thereby causing an amplification of primary hemostasis, as well. Positive feedback is also observed with factor 10a as it activates factor 8 from the intrinsic pathway and factor 7 from the extrinsic pathway. Xa (10a) activates Viii (8) and Vii (7)

Primary Hemostasis Outline the sequence of steps involved in formation of the primary hemostatic plug

primary hemostasis: A critical out comeof primary hemostasis is the formation of a primary hemostatic plug, also known as a platelet plug. The platelet plug limits blood loss from the site of an injured blood vessel. There is a well-defined sequence of steps involved in formation of the platelet plug: 1) First, there is adhesion, where platelets bind to sub-endothelial collagen. 2) Subsequently, activation causes platelet shape change and induces secretion of granule contents. 3) Aggregation of the platelets is followed by further activation and further secretions. 4) Additional platelets are drawn to the injured site and join in the aggregation, culminating in formation of the primary hemostatic plug.

Primary Hemostasis Identify key substances stored in platelet dense granules and alpha-granules

various contents carried within platelet granules: Here is a selected list of dense granule components with corresponding roles indicated for each. As mentioned earlier ,non-metabolic ADP functions as a platelet agonist. Polyphosphates serve to accelerate activation of clotting factor five, which plays an important function in secondary hemostasis. Calcium functions in both platelet and clotting factor activation, and serotonin is both a platelet agonist and a vasoconstriction activator. Platelet alpha granules contain more than 300 different types of protein, some that act as promoters of hemostasis, some that have inhibitory functions, and others that have non-hemostatic functions. Here is a selected list of key alpha granule proteins, with corresponding roles indicated for each. Fibrinogen serves as a principal substrate protein for platelet aggregation in primary hemostasis, and as a fibrin substrate for secondary hemostasis. VWF functions in both platelet adhesion for primary hemostasis, and as a carrier for clotting factor 8 as part of secondary hemostasis. Alpha granules package clotting factors 5, 11, and 12, and also package clotting factor inhibitors, Protein S and Tissue FactorPathway Inhibitor (TFPI). Some factors are components of fibrinolysis. Plasminogen is the precursor of plasmin, which breaks down clots; and PAI-1 is a fibrinolysis inhibitor. Platelet factor 4 neutralizes heparin, which is a clotting factor inhibitor. Platelet derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) are both mitogens. They induce mitosis to enable blood vessel healing and growth

Bleeding Disorders Describe various types of inherited and acquired vascular disorders

vascular disorders: These are disorders of primary hemostasis due to blood vessel defects, which may be either inherited or acquired conditions. The most common symptom of vascular disorders is abnormal bleeding into or under the skin. Standard laboratory screening tests are used to exclude platelet or coagulation factor disorders, as most vascular disorders are not associated with platelet or plasma factor defects. Confirmation of a vascular disorder usually requires histologic evaluation of blood vessel or connective tissue. Inherited vascular disorders are commonly characterized by abnormal synthesis of sub endothelial connective tissue or extracellular matrix components. These rare inherited disorders include: Ehlers-Danlos Syndrome,Osteogenesis imperfecta, Pseudoxanthoma elasticum, Marfan syndrome, and Hereditary hemorrhagictelangiectasia (HHT). Ehlers-Danlos syndrome resultsf rom an underlying collagen synthesis disorder. These patients experience blood vessel fragility and joint hypermobility. Osteogenesis imperfectais also associated with a collagen disorder. These patients develop a defective bone matrix and experience many fractures and are at risk of developing intra-cerebral hemorrhage. Pseudoxanthoma elasticum is associated with degeneration of elastic connective tissue fibers. These patients develop retinal and gastrointestinal bleeding episodes.

Outline the stages of megakaryopoiesis & thrombopoiesis

we next consider TPO a bit further. Thrombopoietin ( TPO) is the primary regulator for megakaryocyte and platelet development. TPO binds to the c-Mpl receptor on bone marrow progenitors and megakaryocytes. TPO also plays a role in HSC survival, self-renewal, and expansion. TPO is produced at a constant rate by the liver, and some is also produced by the kidneys, but the level of free TPO is regulated through interaction with platelets. TPO binds to the c-Mpl receptor on circulating peripheral blood platelets and is internalized by the platelets. Thus, the higher the platelet count, the less TPO is available for megakaryocyte development. There is also are recombinant form of TPO that is used as a therapy for thrombocytopenia. Recombinant TPO can increase platelet production by 3 to 20 times above baseline. Here we zoom in on the four stages of morphologically recognizable megakaryocyte development. These are non-proliferating stages of development, but the cells go through notable morphological changes. Each stage is characterized by a particular morphological cell type: beginning with the megakaryoblast, followed by the promegakaryocyte, then the granular megakaryocyte, and finally the mature megakaryocyte which subsequently breaks up into platelet fragments. Notably, during stages two and three, the differentiating megakaryocytes develop granules replete with hemostasis factors, and they form extensive internal membranes, called the demarcation membrane system, the DMS. These highly branched and interconnected membrane channels facilitate communication with the extracellular space by allowing diffusion of extracellular compounds into the cells. Moreover, these membranes mark the boundaries of where platelets will eventually break off from the parent cell.