Platelet Disorders

Quantitative Platelet Disorders

Decreased numbers of platelets (thrombocytopenia) can also cause a bleeding tendency. The evaluation of thrombocytopenia begins with a careful history and physical and review of medications (including over the counter meds). Important issues to touch on would include associated systemic symptoms and any evidence of bleeding or bruising. After the CBC reveals decreased platelet count, the lab evaluation begins. The next step is to review the peripheral smear and confirm that thrombocytopenia is truly present. Pseudothrombocytopenia is a disorder in which the platelets are sticky and tend to clump together in the test tube. The clumped platelets are counted as one platelet or if the clumps are big enough, as a lymphocyte or red cell. This condition is strictly a laboratory artifact; the patients do not exhibit a tendency to thrombose. Once recognized, the blood can be redrawn in a different anticoagulant to try to avoid clumping or manual estimation of platelet count is performed. The important thing is to avoid an extensive evaluation for thrombocytopenia when it is not required. In addition to ruling out pseudothrombocytopenia, the peripheral smear frequently offers additional clues about the cause of thrombocytopenia. Decreased Production vs. Peripheral Consumption The next step in evaluating thrombocytopenia is to perform a bone marrow biopsy. In this test, we evaluate the number of megakaryocytes present to differentiate decreased production versus increased peripheral destruction as a cause of thrombocytopenia. Increased numbers of megakaryocytes suggest peripheral destruction while decreased numbers of megakaryocytes suggests decreased production of platelets. At the same time, the bone marrow sample may offer additional information helpful in making a diagnosis. Immune mediated versus Non-immune mediated peripheral consumption Peripheral consumptive causes of thrombocytopenia are further divided into immune mediated and non-immune mediated causes. Antibody coating of platelets leads to rapid removal from the bloodstream by the reticuloendothelial system. These disorders frequently lead to rapid declines in platelet counts that do not respond to platelet transfusion but rather require immune modulation to correct the low platelet count. Many times, the antibodies directed against platelets are more difficult to assay than those directed against red cells.

Acquired causes of platelet dysfunction--drugs

Drugs There is a large list of medications that can affect platelet function. In this discussion, we will review several commonly used drugs that can have profound effects on platelets. Aspirin acetylates platelet cyclooxygenase. Inhibition of cyclooxygenase blocks the synthesis of prostaglandins that are critical to the release reaction. Thus aspirin inhibits the release reaction and this inhibition is irreversible for the life of the platelet. As platelets live 7 to 10 days, a single aspirin table can alter platelet function for a week. Patients on aspirin usually have only modestly prolonged bleeding times. Aggregation studies show diminished aggregation with collagen, epinephrine, and low doses of ADP. Clinical bleeding with aspirin is unusual unless the patient has an underlying thrombocytopenia, uremia, hemophilia or is anticoagulated. Under those circumstances, aspirin can precipitate a bleeding diathesis. Nonsteroidal anti-inflammatory drugs also inhibit cyclooxygenase and produce a pattern of platelet dysfunction very similar to aspirin. The effect of these meds on platelets is reversible and will go away as the drug is cleared from the blood stream. Eptifibatide (Integrilin) is a drug that blocks the IIbIIIa complex and thus prevents platelet aggregation. Abciximab is a monoclonal antibody directed against the IIbIIIa complex and also prevents platelet aggregation. These drugs are used to prevent coronary artery stenosis, stent reocclusion. Clopidrogel (Plavix) blocks the ADP receptor and prevents platelet aggregation. This drug is commonly used in patients with vascular disease to prevent thrombosis.

Immune mediated peripheral consumption--HIT

HIT: Heparin induced thrombocytopenia is caused by an antibody that interacts with a combination of heparin and a platelet antigen called platelet factor 4. The antibody will only bind in the presence of platelets and heparin. Typically, platelet counts begin to decline after 10 to 14 days of heparin exposure. This delay is due to the lag time in making a new antibody. Platelet counts can become quite low but the real clinical problem with this disorder is a tendency to develop serious thromboses. It is thought that the antibody binding may activate platelets and release prothrombotic substances causing this thrombotic tendency. As mentioned, platelet counts decline after 7 to 14 days of heparin exposure. It is well to note that heparin exposure can be subtle - small amounts of subcutaneous heparin to prevent DVT or minute amounts used to keep arterial lines open may be sufficient to initiate this process. The physical exam reveals only bruising or petechiae and blood smear shows only decreased platelets. There are no lab studies that are specific enough to make the diagnosis. Platelet aggregation studies which combine patient plasma with platelets and heparin looking for aggregation or release of serotonin are difficult and insensitive. Newer ELISA studies looking for antibodies that bind platelet factor 4 in the presence of heparin are too sensitive and plagued by false positives. Thus, the diagnosis of HIT remains clinical and the key to making this diagnosis is appreciating the association between heparin administration and developing thrombocytopenia

Causes of platelet dysfunction

Hereditary There are a series of rare inherited disorders that affect platelet function. These are quite rare. Glanzmann's thrombasthenia is an inherited autosomal recessive disorder in which patients present with mucosal type bleeding (epistaxis, gingival bleeding, and menorrhagia). These patients have deficient expression of the glycoprotein IIbIIIa complex. The lack of this surface receptor leads to a platelet that can adhere to collagen and can release contents but lacks platelet to platelet interactions. Bleeding time is prolonged and platelet aggregation studies demonstrate lack of response to ADP stimulation. Bernard Soulier syndrome is an inherited autosomal recessive or codominant disorder. Patients present with moderate to severe bleeding of the "platelet" type (epistaxis, gingival bleeding, and ecchymoses). Platelets look abnormal and are larger than normal. This is frequently associated with a moderate thrombocytopenia. Bernard Soulier platelets lack expression of glycoprotein Ib. As this is the receptor for Von Willebrand's factor, these platelets do not adhere well to collagen surfaces. Bleeding times are prolonged. Platelets aggregate with ADP but do not respond well to vWF, ristocetin, and thrombin.

Immune mediated peripheral consumption--ITP

ITP: Immune mediated thrombocytopenia purpura is a common disorder in which autoantibodies bind to platelets and cause rapid clearance of platelets. Patients are commonly young females. This autoimmune disorder can be associated with systemic lupus erythematosis, rheumatoid arthritis, CLL, etc. Patients present with petechiae and mucosal bleeding. Physical exam is remarkable for a lack of findings - no splenomegaly, adenopathy, etc. The finding of splenomegaly should cause one to consider other possibilities. The CBC shows markedly low platelet count. Peripheral smear shows only a lack of platelets. There should be no additional findings to make a diagnosis of ITP. Bone marrow demonstrates increased megakaryocytes and no additional findings. There are no accepted anti-platelet antibody assays for ITP thus this is not helpful in making a diagnosis. The diagnosis is established by finding low platelets with increased marrow production and a lack of findings suggesting other diagnoses. Patients respond to steroids or IVIG leading to rapid rises in platelet counts (days). Unfortunately, these treatments are rarely curative and thrombocytopenia frequently recurs when steroids are tapered. Splenectomy resolves thrombocytopenia in two thirds of patients who recur when steroids are tapered. Fortunately, while patients may have very low platelet counts; serious bleeding or death is rare.

Disorders with decreased platelet production

Inherited: there are a series of disorders that present at birth with decreased platelet count. These syndromes may include alterations in other cell lines and sometimes morphologic alterations as well. Some have additional inherited abnormalities such as an absent radius. Treatment is supportive. Drug induced: a variety of medications can cause decreased platelet production. Thiazide diuretics, phenothiazines, and dilantin are common offenders. Chemotherapy agents suppress marrow production in general and sometimes platelets in particular.

Peripheral Consumption: non-immune

Non-immune Mediated •Sequestration -Spleen retains 30% of circulating platelets -Increased retention causes thrombocytopenia -Sequestration can be seen in any disorder causing splenomegaly. Big spleen=drop in platelet count -Liver disease most common offender (b/c leads to big spleen and other things..) -Splenectomy only effective treatment •Disseminated Intravascular Coagulation (DIC) -The coagulation system is activated leading to consumption of coag factors and platelets -Initiated by sepsis, metastatic cancer, obstetric complication, etc -Clinically presents with bleeding and/or thrombosis -Diagnosed by finding decreased coag factors (PT,PTT) and platelets in appropriate clinical setting -Treatment is to relieve the initiating cause The longer DIC occurs the harder it is to correct.

TTP

Non-immune mediated •Thrombotic Thrombocytopenia Purpura (TTP) pathophysiology -Caused by an alteration in vonWillebrand factor processing - decreased protease activity -Large vWF multimers cause platelets to stick and plug small vessels causing ischemia -Protease deficiency is inherited (rare) or autoimmune Don't need the complete pentad to make the diagnosis: fever, neurologic symptoms, renal failure, hemolytic anemia and thrombocytopenia •TTP Clinical -Fever, renal failure, neurologic changes, thrombocytopenia, microangiopathic hemolytic anemia -Anemia from shearing of red cells by platelet clumps •increased LDH, reticulocytes, bilirubin •Blood smear shows schistocytes, polychromasia and decreased platelets. Then diagnosis is made and treatment begins immediately (plasmapheresis). Lots of schistocytes to convince you it might be TPP. Very few platelets--large=working overtime. New large platelets •TTP Treatment -Until the 1980's, TTP was routinely fatal -With plasma exchange, now 90% survive •Blood removed through large IV and plasma is separated •Plasma is replaced with fresh frozen plasma •Antibodies to vWF protease are removed and protease activity is replaced •Prognosis -30% relapse rate. Relapsing and refractory are big problems (refractory is worse). If pheresis didn't work or steroids or Ig then cytoxan, vincristine, then rituxan.

Uremia

Patients with renal failure exhibit a consistent and clinically significant platelet dysfunction. The mechanism of this dysfunction remains somewhat mysterious but involves impairment of the release mechanism. The bleeding time is variably prolonged and platelet aggregation by collagen, epinephrine is abnormal. There is good evidence that these abnormalities are caused by a dialyzable factor in plasma. The specific agent has not been identified and may well be due to several metabolites that accumulate in uremia. The best treatment for bleeding in the uremic patient is dialysis, but this is not always effective. Additionally the use of cryoprecipitate or DDAVP has been shown to shorten the bleeding time and may offer temporary relief of bleeding in uremic patients. Cryoprecipitate and DDAVP will elevate vonWillebrand factor, but it remains unclear how elevating vonWillebrand factor corrects the platelet dysfunction.

Hematopoietic Disorders

Plasma cell dyscrasias have been associated with bleeding tendencies, especially Waldenstrom's macroglobulinemia. Varying abnormalities of platelet aggregation have been described with macroglobulinemia. Interestingly, adding normal immunoglobulins but not IgG fraction from macroglobulinemia patients will restore platelet aggregation. Some have suggested that the large IgM molecule may "stick" to platelets and prevent normal interactions with platelet surface proteins. Treatment of the underlying disorder with diminution of the IgM proteins can lead to improved platelet function. Myelodysplasia is a stem cell disorder characterized by dyserythropoiesis. Megakaryocytes can share in the abnormal morphology seen in hematopoietic precursors and affected platelets demonstrate diminished granules, distorted canalicular systems, and occasionally abnormal fusion of intracytoplasmic granules. These morphologic abnormalities have been associated with prolonged bleeding times and a variety of platelet aggregation abnormalities. Platelet transfusions can partially correct the bleeding tendency in myelodysplasia. On occasion we must recommend platelet transfusions when the platelet count is normal or elevated.

Platelets

Platelet Structure • Derived from megakaryocytes • Storage granules and canalicular system--Canalicular system important to change shape/size and for degranulation - contain epinephrine, ADP, Factors V and VIII, serotonin, fibrinogen and prostaglandins • Microtubules and microfilaments permit shape change •Normal count 150 to 450,000/ul •Decreased platelet count associated with bleeding -20,000/ul or less associated with spontaneous bleeding -100,000/ul fully protective from bleeding Platelet Bleeding •Bleeding associated with thrombocytopenia is distinctive -Petechiae -Mucosal bleeding •Deep tissue bleeding such as hemarthroses is more typical of hemophilia Platelet Function •Platelets circulate as an inert disc •Exposure of subendothelial collagen causes adhesion of platelet through glycoprotein Ib •Platelets undergo shape change to sphere with spines •Platelets release granule contents •Platelets aggregate by way of GpIIbIIIa

Platelet Structure and Function

Platelets are created by budding from megakaryocytes. While small and unassuming, they are surprisingly complex. Platelets do not have nuclei but do possess storage granules and a canalicular system connecting the granules to the membrane surface to permit release of granule contents. Platelets contain microtubules and microfilaments to permit change in shape and release of contents. Platelet storage granules contain a variety of substances including epinephrine, serotonin, ADP, ATP factor V, factor VIII, fibrinogen, prostaglandins. Platelets also express a variety of surface proteins that allow the platelet to interact with its environment. Glycoprotein Ib contains binding sites for Von Willebrands factor and thrombin permitting platelet to platelet interactions. Glycoproteins IIb and IIIa form a complex which serves as a binding site for fibrinogen. Binding of fibrinogen is important in platelet aggregation. The normal platelet count ranges from 150,000 to 450,000 per micro liter of blood. The primary function of platelets is to help seal breaks in blood vessels. Decreases in platelet count can be associated with bleeding. We usually think of spontaneous bleeding occurring with platelet counts of less than 20,000/ul. If a patient is going to have a surgical procedure, a platelet count of 100,000/ul is required to prevent bleeding. While severe thrombocytopenia can present with bleeding anywhere, certain types of bleeding are characteristic of thrombocytopenia. Petechiae look like small, non-blanching red dots under the skin and represent cutaneous bleeding from small blood vessels. They are seen wherever venous pressure is elevated, typically on feet and ankles. Mucosal bleeding such as nosebleeds or oozing from gums is also typical of bleeding with thrombocytopenia. Deep-seated bleeding, such as hemarthroses, is more typical of hemophilia.

Non-immune mediated peripheral consumption

Sequestration: the spleen normally retains approximately 30% of the platelet population. Enlarged spleens from any cause can retain a larger percentage and lead to thrombocytopenia. Thus, this problem can be seen in any disorder with an enlarged spleen, most commonly liver disease with portal hypertension. Platelet sequestration responds to splenectomy, unfortunately patients are frequently not candidates for surgery. DIC: a coagulopathy in which coagulation factors and platelets are consumed in unregulated clotting process. DIC is always initiated by something - sepsis, obstetric complication, metastatic malignancy, etc. This disorder is diagnosed by demonstrating abnormal PT, PTT, low platelet count and elevated D-dimer in the appropriate clinical setting. DIC is best treated by relieving the initiating event. This may be simple with an obstetric complication resolved by delivery. This may be impossible in metastatic cancer for which there is no effective treatment.

cont'd--TTP

TTP: thrombotic thrombocytopenia purpura is an unusual disorder now known to be caused by faulty vonWillebrand factor processing. VonWillebrand factor is usually cleaved by a circulating protease to yield multimers of appropriate size. A lack of this protease leads to large von Willebrand multimers circulating. These large multimers are exceptionally "sticky" leading to plugging of small vessels by platelet clumps. This in turn causes end organ ischemia. The lack of protease can be seen in rare inherited cases or more commonly, an autoimmune disorder in which the protease is inactivated by an autoantibody Clinically TTP presents with fever, renal failure, mental status changes, thrombocytopenia and microangiopathic hemolytic anemia. This classic pentad may not always be complete. The fever, renal failure and neurologic features are due to platelet plugging of small vessels causing ischemia. The anemia in this disorder is caused by shearing of red cells by the platelet clumps in small vessels resulting in microangiopathic hemolysis. On the peripheral smear one will see red cell fragments (schistocytes) and polychromasia along with decreased platelets. Lab studies will confirm hemolysis with elevated LDH, bilirubin, reticulocyte count and decreased haptoglobin. Recognition of decreased platelets associated with microangiopathic hemolytic anemia should distinguish this disorder form the more common ITP. Treatment is with plasma exchange. The prognosis is excellent although there is a 30% chance of recurrence. While this is an uncommon disorder it is important to recognize. TTP is >90% fatal if not treated properly.

Qualitative Platelet Disorders

There are conditions in which a normal number of platelets is still associated with bleeding. While adequate numbers of platelets are available, they do not function normally - a qualitative platelet dysfunction. Platelets recognize a break in lining of a blood vessel by interactions between and specific platelet receptors. Von Willebrand's factor is also important in forming a bridge between collagen and the platelet. Again, specific receptors on the platelet bind Von Willebrand's factor. This adhesion of platelets to a collagen surface triggers additional changes. The platelet changes shape from a disc to a spiny sphere. Expression of additional surface proteins facilitates platelet to platelet interactions known as aggregation. At the same time, granules fuse with the canalicular system and granular contents are released or secreted from the platelet.

Test of Platelet function

To measure platelet function, we have a limited number of tests. The CBC can quantify the number of platelets and some machines can offer platelet size and variation in size. Review of the peripheral blood smear can offer clues about changes in the size or shape of platelets as well as abnormalities of other cell lines. The bleeding time measures the time required to stop bleeding from a "standard" cut in the skin. Despite best efforts to create reproducible bleeding time devices, this test remains very much operator dependent. It is important that you have confidence in the lab doing the test and that they have performed controls and can offer lab specific normal values. Time to stop bleeding from a skin cut is almost entirely dependent on platelet function. Prolonged bleeding times suggest insufficient platelet number or function. Occasionally, vascular abnormalities can also cause prolonged bleeding times but these are rare. The bleeding time is NOT a good screening tool in patients without a history of a bleeding problem. It has been well documented that bleeding times do NOT predict for surgical bleeding and should NOT be part of routine preoperative screens. Platelet aggregation studies can further characterize the type of qualitative platelet defect. In this study, isolated platelets are stimulated with a variety of substances (epinephrine, ADP, ristocetin, collagen, etc). Subsequent platelet aggregation results in altered optical density of the solution providing a readout. There are some recognizable patterns of altered platelet aggregation that can be helpful in understanding the cause of a qualitative platelet defect.

cont'd

Treatment of HIT is simple - stop the heparin and platelet counts rapidly improve. Again, remember that heparin delivery may be subtle and diligence is required to insure that ALL heparin has stopped. In general, once a patient has developed HIT, they should never again receive heparin. Some patients require anticoagulation when they develop HIT. For these patients we substitute other anticoagulant agents like hirudin or danaparoid. Note- low molecular weight heparin can cross react with these antibodies and thus is not acceptable as a substitute for heparin in HIT. A major clinical problem with HIT is not bleeding but rather thrombosis. Patients with HIT have a high risk of thrombosis, which can be venous or arterial. It is recommended that patients be treated anticoagulants for an extended period (6 weeks to 3 months) to prevent these thrombotic complications. This is usually accomplished with one of the above heparin substitutes followed by conversion to oral coumadin.

Thrombocytopenia Evaluation

•As usual, begin with history and physical •Step 2, rule out pseudo-thrombocytopenia -Ex-vivo artifact of platelet clumping -Causes automated counter to mis-count platelets -No clinical consequences •Must review the blood smear to appreciate Further evaluation--If look at smear and there is a lack of platelets! Either not making them or they are being destroyed •Bone marrow biopsy is the next step -Quantitate the number of megakaryocytes -Look for additional clues •Distinguish decreased production from increased peripheral destruction

Tests of Platelet Function

•CBC •Blood smear •Bleeding time--Should not be used anymore -operator dependent -not a good screening tool in healthy patients •Platelet aggregation studies -stimulate platelets with various agents and look for aggregation •PFA100 -Automated test replacing the bleeding time

Hereditary Platelet Dysfunction and Drug induced platelet dysfunction

•Glanzmann's thrombasthenia--Most common--extremely rare tho -autosomal recessive -deficiency of gpIIbIIIa -failure to aggregate with ADP •Bernard Soulier Syndrome -autosomal recessive -Deficiency of gpIb -failure to adhere •Aspirin -acetylation of cyclooxygenase leads to diminished prostaglandins and inhibits release of granule -irreversible--Messed up until the platelets die •Nonsteroidal anti-inflammatory -similar mechanism to aspirin -reversible--Platelets regain function in a few days. •Clopidrogel (Plavix) -targets the platelet ADP receptor, prevents aggregation •Integrilin -targets the gpIIbIIIa complex to prevent aggregation •Abciximab -monoclonal antibody directed against gpIIbIIIa

Peripheral Consumption: Immune mediated--HIT

•Heparin Induced Thrombocytopenia (HIT) -Autoantibody directed against platelet factor 4 and heparin -Causes thrombocytopenia due to rapid clearance of antibody coated platelets -Bleeding is usually not the problem, rather thrombosis -How thrombotic tendency is caused is unknown •HIT Clinical -Platelet counts decline after 7 to 14 days of heparin exposure. Second exposure response within 24 hrs -Heparin exposure can be subtle -Assays for HIT remain problematic •Platelet aggregation studies frequent false negative •ELISA assays for antibody to platelet factor 4 frequent false positive -Diagnosis remains clinical •HIT Treatment -Removal of heparin leads to rapid recovery of platelet counts -If patient needs anticoagulation, must use other agents •Hirudin, danaparoid, argatroban •NOT low molecular weight heparin -No more heparin--If heparin removal doesn't fix it then the diagnosis might be wrong or the patient is still taking heparin •HIT Treatment -Risk of thrombosis is high in HIT •Can be arterial or venous, can be devastating -Recommend treatment with other anticoagulant for three months to prevent thrombosis •Coumadin

Peripheral Consumption: Immune mediated--ITP

•Immune Thrombocytopenia Purpura (ITP) -Autoantibody binds to platelets causing rapid clearance -Most common in women -Can be associated with other autoimmune disorders Or idiopathic •SLE, rheumatoid arthritis, CLL •ITP Clinical -Present with petechiae and mucosal bleeding -CBC and smear show ONLY low platelets -Physical exam shows ONLY bleeding -Bone marrow shows only increased megakaryocytes -No accepted anti-platelet antibody studies •ITP Treatment and Prognosis -Initial treatment steroids and/or IVIG (IVIG binds autoantibodies to prevent binding to platelets. In bleeding patient--can't just give platelets b/c the antibody will bind) •75% response rate -Relapse is common -Splenectomy will correct thrombocytopenia in 70% of steroid failures -Frequently a chronic relapsing problem -Patients rarely die or have serious bleeding

Peripheral Consumption & decreased production

•Immune mediated vs. Non-immune mediated -Distinguish antibody mediated disorders -Helpful in developing differential diagnoses -No single test to help with this distinction -Antibody assays are less reliable with platelets than with red cells ITP--can have negative platelet antibodies. Can have false positives too Decreased Platelet Production •Inherited -Very Rare series of disorders present at birth -Some isolated to platelets, some include alterations in other cell lines -May be associated with other body alterations (absent radius, etc) -Presumed inherited defect in stem cell differentiation •Drug induced -Drugs that rarely suppress megakaryocytic development (thiazide, sulfa, phenothiazines, dilantin) -Drugs that always suppress marrow (chemotherapy) •Physical agents -Radiation, benzene

Uremia & Myelodysplasia

•Prolonged bleeding time •Abnormal platelet aggregation -collagen and epinephrine •Dialyzable plasma factor •Dialysis is correct treatment •Cryoprecipitate and DDAVP may help -?vonWillebrand factor?\ DDAVP=VIII and vWF? Myelodysplasia •Stem cell disorder with dysmorphic hematopoietic cells •Megakaryocytes are also dysmorphic •Prolonged bleeding time and variable platelet aggregation •Treat bleeding with platelet transfusion -even if platelet count is normal (don't function correctly)