9.1 Non-malignant leucocytes disorders
Right shift of neutrophils
A common and important morphologic abnormality of neutrophils is so-called "toxic change". Contrary to what has been written in many books, toxic change in neutrophils is not necessarily associated with "toxemia". The term derives from the fact that these abnormalities were first noticed in human patients with gram negative sepsis and endotoxemia. However, toxic change in neutrophils do not reflect a "toxic effect" of bacteria on neutrophils but are morphologic abnormalities acquired during maturation under conditions that intensely stimulate neutrophil production and shorten the maturation time in the bone marrow. This accelerated maturation occurs secondary to cytokine stimulation, which is usually in reponse to inflammation. Animals recovering from bone marrow injury or who are administered hematopoietic cytokines (e.g. granulocyte-colony stimulating factor or G-CSF) can also show accelerated maturation in neutrophils or toxic change. On the standard hemogram, the presence of toxic change is reported under the WBC exam section and is subjectively graded as mild, moderate and marked. Most of the toxic changes reflect asynchrony of maturation between the nucleus and cytoplasm. During normal granulocytopoiesis, the lengthening and pinching of the nucleus are coordinated with progressive condensation of the chromatin and loss of cytoplasmic protein synthetic machinery (RNA in the form of ribosomes and rough endoplasmic reticulum - which imparts a blue color to the cytoplasm with hematologic stains). With accelerated maturation, nuclear divisions may be skipped (resulting in larger cells than normal) and the cells retain immature features, including increased amounts of rough endoplasmic reticulum or ribosomes in the cytoplasm and lighter chromatin than normal. Specific granules may be less visible than in normally mature cells and, in some species, primary granules (normally inapparent in neutrophils) retain their affinity for the stains used for peripheral blood. The cells can also have frothy or vacuolated cytoplasm. In a blood smear there are five main features of toxic change: Cytoplasmic basophilia: A streaky diffuse irregular blue appearanceto the cytoplasm. It is due to the presence of polyribosomes and rough endoplasmic reticulum. Döhle bodies: These are pale round to linear blue aggregates in the cytoplasm, caused by whorls of rough endoplasmic reticulum (see image to the right). This is often the earliest and first indication of toxic change. However, note that healthy cats can have low numbers of small Döhle bodies and Döhle bodies form in neutrophils with storage (storage-related artifact). Thus, Döhle bodies alone do not always indicate toxic change. Cytoplasmic vacuolation: These are indistinct vacuoles in the cytoplasm, giving it a frothy appearance and is due to degranulation of lysosomes. Note that clear punctate vacuoles are usually not attributed to toxic change, but are frequently another storage-related artifact (these vacuoles can occur within 4 hours of blood collection). Nuclear immaturity: The nuclear chromatin is lighter (finer) and less coarsely clumped than normal. This is the hardest and most subtle change to see. Toxic granulation: These are distinct red granules in the cytoplasm due to the primary granules taking up stain. This is a rarely seen feature in domestic animals, but is most commonly seen in large animals (horses, ruminants, camelids - see image below).
Left shift of neutrophils
A left shift indicates the presence of immature neutrophils in blood and usually, but not always, indicates an inflammatory leukogram (see related links for the historical origin of this term). Immature neutrophils are usually band neutrophils, but earlier forms can be seen. A few to no band neutrophils are seen in the blood of clinically healthy animals we use for establishing our reference intervals. This indicates that low numbers of band neutrophils, particularly in the absence of other features of inflammation, such as toxic change, may not be a clinically relevant finding. Some concepts associated with a left shift: 【】A left shift can be due to release of bone marrow stores. This particularly occurs if the bone marrow reserve of mature neutrophils is low or depleted. Note, that ruminants have lower bone marrow reserve of mature neurophils than other species. Release of the marrow reserve usually occurs in response to acute inflammation. Stores will be rapidly depleted if bone marrow production (resulting in a myeloid or granulocytic hyperplasia) does not kick in - this marrow response usually takes around 3-4 days and occurs due to the release of granulopoietic cytokines, such as granulocyte- colony stimulating factor (G-CSF), granulocyte-monocyte-colony stimulating factor (GM-CSF) or inflammatory cytokines such as interleukin (IL)-1 and IL-6. 【】A left shift can indicate a response by the bone marrow to inflammatory or granulopoietic cytokines, which increase marrow production, i.e. there is a myeloid (granulocytic) hyperplasia. 【】In a single hemogram, we cannot always tell if there is a myeloid hyperplasia. A general rule of thumb is that if there is a persistent left shift with a neutrophilia, there is likely a myeloid hyperplasia. If the neutrophil count is very high (>40,000/uL in a dog or cat, >20,000/uL in a horse or ruminant), then there is also likely a myeloid hyperplasia (even without a left shift). 【】A left shift is usually, but not always, accompanied by toxic change in neutrophils. If there is moderate to severe toxic change in neutrophils and no left shift, then there is a problem with cell identification or cells are not toxic but are dysplastic (this can occur in rare myeloid leukemia). If there is a severe left shift (e.g. degenerative - see below), with no toxic change, then the animal has Pelger-huet anomaly (most common cause in dogs; see canine blood in Atlas), there is a problem with cell identification, the animal may have a leukemia or may be recovering from bone marrow injury (without concurrent peripheral inflammation).
Name non-malignant leucocyte morphologic abnormalities, describe their aetiologies, and explain their impacts on leucocyte functions and the patients' well being where applicable
Classification of leucocyte abnormalities Benign abnormalities - Usually are part of reflexive and beneficial - physiologic responses to external agents such as infections - In non-hereditary cases, are often temporary Malignant abnormalities - Accelerated uncontrolled proliferation of immature cells in the body an/or inhibition of apoptosis - Are monoclonal in nature - Usually associated with a shortened life expectancy of the patient
Define terms relevant to quantitative leucocyte abnormalities including but not limited to leucocytosis, leucopenia, neutrophilia, neutropenia, lymphocytosis, monocytosis, eosinophilia, and basophilia and relate them to the relevant physiologic or pathologic conditions cont. 3
Eosinophils are derived from hematopoietic stem cells initially committed to the myeloid line and then to the basophil-eosinophil granulocyte lineage. Nonpathologic functions of eosinophils and the cationic enzymes of their granules include mediating parasite defense reactions, allergic response, tissue inflammation, and immune modulation. [2, 3] Tissues of the pulmonary and gastrointestinal systems are the normal residence for eosinophils, but peripheral, or blood, eosinophilia (absolute eosinophil count [AEC] >600 cells/µL) indicates an eosinophilic disorder. [4] Untreated, the eosinophilia can be categorized as mild (AEC 600-1500 cells/µL), moderate (AEC 1500-5000 cells/µL) or severe (AEC >5000 cells/µL). An increase in tissue eosinophilia may be seen with or without concurrent peripheral eosinophilia. A secondary or reactive increase in blood eosinophils, tissue eosinophils, or both is associated with a wide variety of conditions, as follows [5, 6] : Infections (especially helminthic parasites) Allergic responses Neoplasms Connective tissue disorders Medications Endocrinopathies Primary eosinophilia is not a reactive phenomenon and can be described as either clonal or idiopathic in nature. If an underlying molecular or cytogenetic abnormality can be identified, the eosinophilia can be designated as a clonal disorder. If reactive causes are ruled out and no underlying clonal origin is proven, the eosinophilia is described as idiopathic. [7] ———————————————————————— Basophilia is defined as an elevated absolute basophil count greater than 200 cells/uL or relative basophil count greater than 2%, though each laboratory should set their own normal ranges based on the local population. The elevation of these cells can often suggest underlying diseases, such as myeloproliferative disease or chronic inflammation. This article does not refer to basophilic staining, the purple "basic" dye that gives these white blood cells (WBCs) their name. There are three main categories when considering the cause of basophilia. First, hematologic causes include myeloproliferative diseases such as chronic myelogenous leukemia (CML), polycythemia vera, primary myelofibrosis (previously called myelofibrosis with myeloid metaplasia), and essential thrombocythemia. Systemic mastocytosis and hypereosinophilic syndrome are also considered myeloproliferative diseases and they too have been associated with basophilia. Other hematologic diseases that are associated with basophilia include other leukemias and myelodysplastic syndrome. Striking basophilia can be seen in the very rare chronic basophilic leukemia. The second category involves chronic inflammation, a broad range of diseases that include tuberculosis, parasitic infections, inflammatory bowel disease, and rheumatoid arthritis. Although these diseases are chronic by nature, a persistent basophilia should not be ignored and hematologic neoplastic syndromes explored. The third category is allergic including food and drug allergies and allergic rhinitis. The degree of basophilia may correlate with symptoms.
Hypersegmentation
Hypersegmentation, the presence of abnormally increased nuclear lobulation, is one of the first hematologic abnormalities seen in megaloblastic anemia. Normal mature circulating neutrophils have an average of 3 lobes and always fewer than 5 lobes. Eosinophils have fewer than 4 lobes and basophils have fewer than 3 lobes. More than 3 cells having 5 lobes or a single cell with 6 lobes found in the course of a 100 cell differential is evidence of hypersegmentation. Hypersegmentation is sometimes referred to as a myeloid "right shift". Hypersegmentation may accompany other disorders in which there is a disturbance of maturation, such as iron defiency.
Describe pathogenesis, clinical findings and laboratory features of infectious mononucleosis
Infectious mononucleosis (IM) is a clinical syndrome. IM represents the immunopathologic expression that occurs under a specific set of circumstances and in response to infection with the Epstein-Barr virus (EBV). 【Pathogenesis】 The most common mode of transmission of EBV is through exposure to infected saliva from asymptomatic individuals, often as a result of kissing. Following exposure, EBV infects epithelial cells of the oropharynx and salivary glands. B lymphocytes may become infected through exposure to these cells or may be directly infected in the tonsillar crypts. B-cell infection allows viral entry into the bloodstream, which systemically spreads the infection. In the immunocompetent patient, the proliferation of infected B cells results in massive activation and proliferation of cytotoxic T lymphocytes, leading to the characteristic lymphoid hyperplasia. Clinically, this is observed as tonsillitis, lymphadenopathy, and hepatosplenomegaly. The T-cell response is largely responsible for the rise in the absolute lymphocyte count and for the finding of atypical lymphocytes. These atypical lymphocytes (ie, Downey cells) actually are CD8 cytotoxic T cells. B-cell infection caused by EBV leads to the transformation of the B cells to immortal plasmacytoid cells, which secrete a wide variety of immunoglobulins (eg, heterophile antibodies). Antibodies against specific EBV antigens and a number of autoantibodies also are produced. 【Clinical findings】 Fever usually does not exceed 102°F in infectious mononucleosis, but it may be as high as 104°F. Pharyngitis often is the most prominent physical finding. Tonsillar edema and erythema with a grayish or greenish exudate are common and are clinically indistinguishable from streptococcal pharyngitis. Affected lymph nodes usually are symmetrically enlarged, firm, mobile, and tender. The nodes usually do not demonstrate warmth or overlying erythema. Splenomegaly is present in most cases of infectious mononucleosis, but it may not be appreciated on physical examination. Hepatomegaly is found in 10-30% of cases. Periorbital edema occurs in 15-35% of patients with infectious mononucleosis. Petechiae of the palate, occurring at the junction of the hard and soft palate, may occur in up to one third of cases. Petechiae are not pathognomonic, but evidence of them is highly suggestive of infectious mononucleosis. Jaundice may occur. 【Laboratory features】 White blood cell count The WBC count and differential can be useful in establishing a diagnosis of infectious mononucleosis. WBC count results usually show a modest elevation, with a peak of 10,000-20,000 during the second or third week of the illness. Findings consistent with infectious mononucleosis include a differential that demonstrates greater than 50% lymphocytes, an absolute lymphocyte count greater than 4500, or an elevated lymphocyte count with greater than 10% atypical lymphocytes. Liver function tests Liver function tests (LFTs) are abnormal in more than 90% of patients with infectious mononucleosis. Serum transaminase and alkaline phosphatase levels usually are modestly elevated. The serum bilirubin may be increased in approximately 40% of patients, but jaundice only occurs in approximately 5% of infectious mononucleosis cases.
Dohle bodies
L Dohle bodies are single or multiple blue cytoplasmic inclusions. They represent remnants of rough endoplasmic reticulum from earlier maturational stages. They are associated with myeloid "left shifts" and are seen in conjunction with toxic granulation.
Define terms relevant to quantitative leucocyte abnormalities including but not limited to leucocytosis, leucopenia, neutrophilia, neutropenia, lymphocytosis, monocytosis, eosinophilia, and basophilia and relate them to the relevant physiologic or pathologic conditions
Leucocytosis The condition in which white cells are present in greater numbers than normal is termed leukocytosis. It is usually caused by an increase in the number of granulocytes (especially neutrophils), some of which may be immature (myelocytes). Most often leukocytosis is the result of the presence of an infection, usually caused by pyogenic (pus-producing) organisms such as Streptococcus, Staphylococcus, Gonococcus, Pneumococcus, or Meningococcus. Leukocyte counts of 12,000 to 20,000 per cubic millimetre during infections are not unusual. As the number of cells increases, the proportion of immature cells usually rises, perhaps because the demands on the leukocyte-producing tissues in the bone marrow have increased to the point at which there is an insufficient number of mature cells for delivery into the circulation. As the infection subsides, the number of younger forms and the total white cell count decrease and ultimately return to normal. During the period of repair following an inflammatory reaction, the monocytes may increase in number, and subsequently the lymphocytes will become more numerous. Certain types of infection are characterized from the beginning by an increase in the number of small lymphocytes unaccompanied by increases in monocytes or granulocytes. Such lymphocytosis is usually of viral origin. Moderate degrees of lymphocytosis are encountered in certain chronic infections such as tuberculosis and brucellosis. ———————————————————————— Leucopenia Leukopenia is characterized by leukocyte counts that are abnormally low (below 4,000 per cubic millimetre). Like leukocytosis, which is usually due to an increase of neutrophils (neutrophilia), leukopenia usually is due to a reduction in the number of neutrophils (neutropenia). Of itself, neutropenia causes no symptoms, but persons with neutropenia of any cause may have frequent and severe bacterial infections. Agranulocytosis is an acute disorder characterized by severe sore throat, fever, and marked fatigue associated with extreme reduction in the number of neutrophilic granulocytes or even their complete disappearance from the blood. Neutropenia may be due to a hypersensitivity mechanism in which a drug provokes the formation of antibodies. If the drug happens to bind to the neutrophil, the antibody destroys the neutrophil when it reacts with the drug. A second important mechanism of neutropenia is toxic damage to the bone marrow. Chemotherapeutic agents used in the treatment of cancer, particularly leukemia, often produce leukopenia (and neutropenia) by damaging the bone marrow. Other drugs that cause neutropenia include pain relievers (analgesics), antihistamines, tranquilizers, anticonvulsants, antimicrobial agents, sulfonamide derivatives, and antithyroid drugs. Neutropenia also is associated with certain types of infections (e.g., typhoid, brucellosis, measles, and HIV infection) and with certain diseases involving the bone marrow (e.g., aplastic anemia) or the spleen. In addition, sufficiently high doses of radiation will cause neutropenia, as will certain chemotherapeutic agents. Treatment is directed toward the cause of the neutropenia.
Define terms relevant to quantitative leucocyte abnormalities including but not limited to leucocytosis, leucopenia, neutrophilia, neutropenia, lymphocytosis, monocytosis, eosinophilia, and basophilia and relate them to the relevant physiologic or pathologic conditions cont. 2
Lymphocytosis or increased blood lymphocyte count is common and occurs at some time in most people, usually in association with viral infections. Lymphocytosis is commonly short lived, and investigation of such cases by immunophenotyping is usually unrewarding. Uncommonly. lymphocytosis may indicate a lymphoproliferative disorder. Infections In viral infection, the elevation in total lymphocyte count is caused by an increase in the absolute count of CD8 T cells. This occurs most dramatically in Epstein-Barr virus infection, but is also associated with other viruses such as acute CMV infection. The cytotoxic CD8 cells are able to kill infected cells, thereby limiting the dissemination of the virus. The CD8 count may remain markedly elevated for many months after the patient recovers. Other infectious diseases that may cause lymphocytosis include toxoplasmosis, in which CD8 T cells are selectively increased, and pertussis, in which all subsets are increased. However, in many infections the blood lymphocyte count remains within normal limits. Disorders of the Spleen Patients who have undergone splenectomy, or whose splenic function is otherwise reduced, may have a lymphocytosis. All lymphocyte subsets may be increased, especially NK cells, and the lymphocytosis itself is not clinically significant. ———————————————————————— Monocytosis Monocytosis is defined as an absolute monocyte count greater than 2SD above the mean for the patient population. Typically, this represents a monocyte count greater than 800 per microliter in adults. Monocyte counts may be significantly higher in children (e.g., up to 3000 per microliter), and age-specific normals should be used to determine if the monocyte count is truly elevated in a younger patient. Monocytosis is most commonly associated with sub-acute or chronic infections (e.g., endocarditis, tuberculosis, syphilis, protozoan, or rickettsial infections), collagen vascular disorders, bone marrow recovery, granulomatous inflammation (e.g., sarcoidosis), and hematologic disorders.
May-Hegglin anomaly
May-Hegglin anomaly is a rare autosomal dominant abnormality characterized by large pale basophilic inclusions resembling Dohle bodies and appear to be altered RNA. Giant platelets, and sometimes thrombocytopenia are associated with this. The anomaly is usually benign but may be associated with bleeding.
Myeloperoxidase deficiency
Myeloperoxidase deficiency (MPO) is a common (1:2,000 individuals) autosomal recessive absence of myeloperoxidase enzyme in neutrophil and monocyte granules. Although people with concommittant MPO deficiency and diabetes mellitus may develop Candidiasis, MPO deficiency in most people is of no clinical consequence. Acquired MPO deficiency may be seen in acute myeloid leukemia, myelodysplastic syndromes, and lead poisoning. P.S. MPO Deficiency: 1)Wright's stained neutrophil; 2)MPO stained neutrophil; 3)MPO stained eosinophil;
Define terms relevant to quantitative leucocyte abnormalities including but not limited to leucocytosis, leucopenia, neutrophilia, neutropenia, lymphocytosis, monocytosis, eosinophilia, and basophilia and relate them to the relevant physiologic or pathologic conditions cont.
Neutropenia It refers to an absolute blood neutrophil count (total leukocyte count per microliter × percent of neutrophils) that is less than 2 SD below the normal mean of the population. Severe neutropenia is a predisposing factor for infections. The organisms normally are found on the skin, in the nasopharynx, and as part of the intestinal flora. The risk of infections is inversely related to the severity of the neutropenia Disorders of Production Cytotoxic drugs given for cancer chemotherapy and as immunosuppressive agents regularly cause neutropenia by decreasing cell production (Chap. 22). These drugs now are probably the most frequent cause of neutropenia in the United States. Neutropenia as a result of impaired production is a common feature of several diseases affecting hematopoietic stem cells, such as acute leukemia (Chaps. 88 and 91), the myelodysplastic syndromes (Chap. 87), and aplastic anemia (Chap. 35). Congenital Immunodeficiency Diseases Neutropenia is a feature of the congenital immunodeficiency diseases and a contributing factor to their susceptibility to infections (Chap. 80). In most of these conditions, neutropenia is attributed to a production disorder based largely on histologic examination of the marrow. In X-linked agammaglobulinemia, which is attributed to defective B-cell development and a mutation in a cytoplasmic (Bruton) tyrosine kinase (BTK), severe neutropenia is present in approximately 25 percent of patients.43 Children with common variable immunodeficiency often have neutropenia associated with thrombocytopenia and hemolytic anemia.43 ———————————————————————— Neutrophlia Several terms are used almost synonymously with neutrophilia, including neutrophilic leukocytosis, polymorphonuclear leukocytosis, and granulocytosis. Leukocytosis is used because an elevation of the number of neutrophils is the most frequent cause for an increase in the total white cell count. Granulocytosis is less specific than neutrophilia, since granulocytes include eosinophils and basophils as well as neutrophils. Extreme neutrophilia is often referred to as a leukemoid reaction because the height of the white cell count may suggest leukemia. This exaggerated reaction may be the result of segmented neutrophils or may be associated with band neutrophils, metamyelocytes, and myelocytes in smaller proportions. In normal individuals, neutrophil counts follow a diurnal pattern of variation, with peak counts in the late afternoon. Neutrophil counts also rise slightly after meals, with erect posture, and with emotional stimuli. Ordinarily these changes are not sufficient to cause neutrophilia.108 Pseudoneutrophilia (Demargination) Vigorous exercise and acute physical and emotional stress can substantially increase the number of blood neutrophils within a few minutes. Chronic neutrophilia follows a prolonged stimulus to proliferation of neutrophil precursors. It can be studied experimentally with repeated doses of endotoxin, glucocorticoids, or colony-stimulating factors. Although the details of the mediators and mechanisms for the development of chronic neutrophilia are not understood fully, a general scheme for this response is now widely accepted (see Fig. 71-2). Expansion of cell production follows stimulation of cell divisions within the mitotic precursor pool, that is, divisions of promyelocytes and myelocytes. Subsequently, the size of the postmitotic pool increases. These changes cause an increase in the marrow granulocytic-to-erythroid ratio.
Pelger-Huet anomaly
Pelger-Huet anomaly is a congenital autosomal dominant disorder in which granulocyte nuclei fail to segment normally. In the homozygote state the nucleus is round. In heterozygotes most granulocytes have bilobed nuclei ("pince-nez" cells) resembling bands. The trait is benign and occurs in 1 in 6,000 people. Cell function is normal. An acquired or pseudo-Pelger-Huet anomaly is seen in myelodysplastic disorders and following drug therapy, and may accompany leukemia and certain infections.
The Alder-Reilly anomaly
The Alder-Reilly anomaly is associated with the genetic mucopoly- saccharidoses. Patients with mucopolysaccharidoses lack the lysozymal enzymes necessary to break down mucopolysaccharides. Dense azurophilic granules, resembling toxic granulation in neutrophils, are seen in all leukocytes. Most characteristic of these disorders are the metachromatic granules surrounded by a clear zone seen in lymphocytes.
The Chediak-Higashi syndrome
The Chediak-Higashi syndrome is a rare autosomal recessive condition associated with abnormally large leukocyte granules resulting from fusion of lysozymes. This disorder may affect granulocytes, leukocytes, and monocytes. Chemotaxis and phagocytosis is defective. Platelets lack dense granules and platelet function is abnormal. Giant melanosomes in occular and skin tissues result in hypopigmentation.
Define leukaemoid reactions, variant lymphocyte, left shit, right shift, and explain their clinical associations with physiologic conditions
The term leukemoid reaction refers to a reactive leukocytosis above 50 3 109/L with neutrophilia and a marked left shift (presence of immature neutrophilic forms). Leukemoid reactions are mostly a result of acute and chronic infection, metabolic disease, inflammation, or response to a malignancy. Leukemoid reaction most often refers to neutrophils, but the increased count may be due to an increase in other types of leukocytes.
Toxic granulation
Toxic granulation is found in severe inflammatory states. The toxic granules are azurophilic, usually found in the promyelocyte, metamyelocyte, band, and segmented stages. The toxic granulation is thought to be due to impaired cytoplasmic maturation,in the effort to rapidly generate large numbers of granulocytes.
