Leukocytes Formation, Function and Pathology Clinical Pathology Kristin M. Canga, RVT

1. Leukocytes Formation, Function and Pathology Clinical Pathology Kristin M. Canga, RVT

2. Leukopoiesis • All WBC production starts out in red bone marrow from the same _____________________ ______ _______ that produces RBCs. • The ____________ that act on the PPSC determine which cell type will be produced. • Each WBC has its own stimulus for production. • All WBCs differentiate and develop in the bone marrow except for some _____________________ which start out in bone marrow but develop elsewhere. • At the beginning of leukopoiesis, all WBCs ___________________ (in the bone marrow)

3. Granulopoiesis • 1. Stem cell 2. Myeloblast • 3. Promyelocyte 4. Myelocyte • 5. Metamyelocyte 6. Band cell • 7. Mature cell (segmented neutrophil, basophil or eosinophil) Leukopoiesis

4. Classification of Leukocytes • Leukocytes may be classified in three different ways: • ____________________________ • ____________________________ • ______________________________________________________________________________

5. Classification by Defense Function • ____________________________ • Phagocytic leukocytes include: • ____________________________ • ____________________________ • ____________________________ • ____________________________ • _________________ production and cellular ______________ • _________________________

6. Classification by Shape of Nucleus • _____________________________ • Means that the nucleus is _________________, or __________________ • Neutrophils • Eosinophils • Basophils • _____________________________ • Means that the nucleus is varying in ___________ and ______________________ • Monocytes • _____________________________ • Means that there is a _____________, rounded nucleus • Lymphocyte

7. Classification by Granules • ____________________(Presence of granules) • ________________– granules not attracted to stain • ________________– granules attracted to acidic stain • ________________– granules attracted to alkaline stain • ____________________(Absence of granules) • ________________ • ________________

8. More on Granulocytes • Originally no granules present • PPSC  _________________, _________________, __________________, __________________, and finally the various granulocytes based on _________________ granules. • _____________________ produces first set of granules as cells mature • First set are called _______________________granules • Golgi Apparatus also produces granules that are attracted to stain • __________________ granules are added as cell matures to determine ____________________ of the cell.

9. Functions of the Specific Granules • Specific Granules contain different substances depending on the cell: • Neutrophil granules contain ____________________ • Aid the cell in killing microorganisms after ___________________ • Eosinophil granules contain __________________________________________ • De-granulate at site of allergic or anaphylactic reaction to reduce swelling • Basophil granules contain ___________ and _________ • Histamines are released to initiate ___________________ at site of reaction • Inflammation draws ____________________ to site • _________________ acts as local anticoagulant to keep blood flowing to injured/damaged area.

10. Details of the Granulocytes • Recall that the granulocytes include: • _____________________ • _____________________ • _____________________

11. Granulocytes - Neutrophils • Also called “polymorphonuclear” cells • Lobes of the ____________ are connected by thin ________________ • As cell ages, the chromatin of the nucleus ______________ (squeezes together) and changes shape. (Think “________ and __________”) • As cell becomes ____________, the segments break apart, losing all evidence of chromatin. (Indicates dead cell)

12. Neutrophil Formation • Adult neutrophils take ~___-___ days to be produced by the bone marrow • Are being produced for release as circulating neutrophils migrate to __________ to fight microorganisms or die of old age. • If high ___________ for neutrophils occurs, they can be produced in less time.

13. Neutrophil Characteristics • Granules are considered to be ________ because they stain neither red, nor blue • Often identified on blood smear based on shape of ______________. • PMN adult cells may contain between 2-4 lobes connected by thin filaments of _____________. • Sometimes the chromatin is so thin that segments look separated. • (your book says 2-5 lobes, either is fine.)

14. Neutrophil Function • Neutrophils only spend ~____ hours in circulation before migrating to __________. • Once in the tissue, neutrophils do not ________________ circulation • For this reason, the entire population of circulating neutrophils needs to be replaced ~________ times daily. • If demand is so high that bone marrow cannot supply enough adults, ________ cells will be released • If demand is still too high, progressively _______________ cells will be released.

15. Neutrophil Function • First line of defense in the blood • Major function is __________________ of invaders • Can respond to foreign microorganisms very quickly. • Use the blood as _________________, squeezing through endothelium cells via ______________. (PLEASE review Figure 9-5 on pg 234 of A&P book)

16. Neutrophil Function • Attracted to site of infection by ______________ • Chemotaxis is the chemical reaction created between the microorganisms and the tissues that ____________cells. • __________________ cells with granules that contain __________________. • Lysosomes contain _______________ enzymes capable of destroying bacteria and viruses upon phagocytosis. • Membrane flows around invader and encases it in vacuole that is created, allowing lysosomes to digest invader. • ____________ is produced through increased _____ absorption, and is toxic to invaders AND ______________.

17. Neutrophil Function • Normally found in tissues that are constantly susceptible to invasion by microorganisms such as the ________ and _____________tract. • Dead or abnormal neutrophils are disposed of by tissue __________________.

18. Normal Neutrophil Count in Blood The neutrophil count in peripheral blood is kept within a specific range (dogs: 3,000 – 11,400/µL; cats: 2,500 – 12,500/µL) in healthy animals and is controlled by three factors: 1) Release of mature neutrophils from the ______________ pool in the bone marrow into ________________________blood. 2) Rate of ______________ from peripheral blood into tissue. 3) Entrance of increase numbers of __________ into the neutrophil _________________ line.

19. Maintaining Normal Neutrophil Pools 1) Release of mature neutrophils from the ____________ pool in the bone marrow into the ________________ blood. • ________________ supply of mature neutrophils are “on call” and ready for __________________ release. • Caused by sudden __________________ movement of neutrophils into _____________.

20. Maintaining Normal Neutrophil Pools 2) Rate of ___________ from peripheral blood into ____________. • Total neutrophil population in peripheral blood can enter the ___________ within a couple of hours • Triggered by massive acute _____________

21. Maintaining Normal Neutrophil Pools 3) Entrance of increased numbers of _________ into the neutrophil ______________ line. • _________ method of control; it takes 3 to 6 days for neutrophils to mature for release

22. Intravascular Pools of Neutrophils • The ______________________________(CNP) • Refers to the blood contained in the ____________________________. • Blood samples obtained for _______________ analysis contain neutrophils from this pool. • The normal range for neutrophils is based on those contained in this pool. • The _____________________________ (MNP) • Composed of neutrophils that line the _________ of small blood vessels (not _______________________). • These neutrophils are not contained in blood sampled for laboratory analysis.

23. Neutrophilia • Defined: An ______________ in neutrophils in blood circulation • If overall neutrophils increase, total ________ count increases causing __________________ • To meet increased demand for neutrophils in tissue, the bone marrow releases its reserve stores of mature, and if necessary, _______________ neutrophils into the blood. • If a blood sample is drawn while these neutrophils are in transit, a ___________ than normal number of neutrophils will be included in the sample (__________________)

24. Neutrophilia in a Periperal Blood Sample

25. Neutrophilia and Stress • _____________________ leukocytosis: • Caused by exercise, ___________, or ________________ • __________________ release results in a shift of neutrophils from the MNP to the CNP (_______________________) • No left shift occurs (no increase in _______ cells) • Count usually not more than ________ the normal value • Should return to normal within ___ minutes of removal of stimulus • Most common in _________ and animals <____mo.

26. Neutrophilia and Stress • ____________________________ • _____________________ -induced • Treatment with exogenous steroids • Steroids inhibit ______________ • Endogenous steroids released in response to major _____________ illnesses, ________________ disturbances, and __________ • Neutrophil count nearly _____________ – mature neutrophilia • No ____________/ Shift from MNP to CNP

27. Neutrophis and Stress • Stress Leukogram – Continued • Expect to see _________________ , and _______________________. • Expect to see _________________ due to lysis of cells. • Expect to see ____________________. • Possible see ___________________ in dogs • Develops over several ________ and may last for several _________.

28. Neutrophilia • Other causes of neutrophilia include: • ____________________ – mild inflammation yields a leukocyte response similar to __________ • severe inflammation yields neutrophilia with a _______________. • Bacterial infections • Conditions associated with extensive ________damage: • Burns, _______________, trauma, extensive ________, neoplasia • Extreme leukocytosis (with neutrophilia) may be associated with _____________________ that produce colony-stimulating factors: • Hepatozoon canis infections, leukemias, and closed cavity infections (i.e. _________________, _______________)

29. Leukemia • Literally means “_______________ blood” • Caused by malignant proliferation of one of the WBC types • In response to some unknown stimulus, stem cells in bone marrow start producing _______________ cells in one cell line at an __________________ rate. • Abnormal cells show up in blood and bone marrow in large numbers, usually before they are _____________ and cause a dramatic _____________in total WBC count. • Leukemias are considered a form of ____________ and can be ___________ or _____________. • Classified by type of _________ involved • Leukemia and leukocytosis may resemble one another; sometimes the distinction is difficult.

30. Left shifts • _______________________ Left Shift - Neutrophilia with some ________ cells present; however, mature, _____________ neutrophils predominate. • ________________________ Left Shift – Neutropenia where ____________ neutrophils outnumber _____________neutrophils • Usually result of extreme migration of cells into tissues and/or detrimental effects of toxins. Degenerative Left Shift

31. Left Shifts • Characterized by an increase in _________ (immature) neutrophils in the blood. • Note: It is normal for 0-300/µL band cells to be present in the blood of a healthy canine/feline. • Usually associated with _________________ conditions • Demand for neutrophils is ______than bone marrow pool. • Left shifts vary from ___________ (slightly increased number of bands) to _____________ (metamyelocytes, myelocytes, and rarely- even promyelocytes present in blood) • ___________ cytoplasm often present during left shifts

32. Toxic Neutrophils Normal Neutrophils Toxic Neutrophils

33. Toxic Neutrophils • Dohle Bodies may be noted in cytoplasm of toxic neutrophils. • Dohle Bodies are __________ and appear similar to _____________ seen in monocytes

34. Left Shift

35. Right shift • ___________________________ – refers to the presence of ___ or more distinct nuclear lobes within neutrophils. • A right shift reflects prolonged ___________time of neutrophils in blood and can occur as a result of: • Resolving chronic ___________________ • Glucocorticoid administration • Hyperadrenocorticism (Cushing’s disease) • ___________________________________ disorders • May develop _____________ when blood film preparation is delayed for more than a few hours.

36. Hyper-segmented Neutrophil

37. Neutropenia • Neutropenia will cause the total ________ count to decrease (_____________________) • If an infection is out of control, all the reserves of ____________________ can be used up faster than the bone marrow can replace them. • Such a condition signifies that the body is _____________ against the invading microorganisms. • Prognosis is ________ for a critically ill animal that has __________________ and ___________________________

38. Neutropenia • Neutropenia can occur due to: • _______________________ of neutrophils (pseudoneutropenia) • Excessive ___________ demand or destruction of neutrophils • Destruction can be _______________-mediated • Reduced or ineffective granulopoiesis • Conditions that cause neutropenia: • Overwhelming _____________ infections (ex: septicemia) • Idiosyncratic drug reactions may result in neutropenia or pancytopenia (e.g. penicillins, cephalosporins, sulfonamides in dogs and chloramphenicol in cats) • Feline ______________ Virus • Canine ____________ Neutropenia (a.k.a. Gray Collie Syndrome)

39. Neutropenia in a Peripheral Blood Smear

40. Eosinophils • Formation: It takes ___ to ___ days to produce an eosinophil from a PPSC. • Population in circulation ~__-__% of total count • _______________ and ____________ pools are also found in peripheral blood (like __________________) • Bone marrow contains good supply of mature cells for release as needed. • Eosinophils migrate into tissue in just a few _______ where they spend the rest of their lives and undergo the same aging process as neutrophils • Granules’ color and size vary depending on the ____________.

41. Eosinophil Granules Feline Canine Equine Bovine

42. Eosinophils • Eosinophils are attracted to, and inhibit local ____________ and hypersensitivity-related reactions. • Their granules contain anti-________________ substances that are released at the site of the allergic reaction. • They ingest substances associated with the _______________ immune response (antigen-antibody reaction complexes) • They have some ________________ and bactericidal functions • They are especially effective in phagocytosis of ______ pathogenic organisms, such as ____________, and some parasitic worms but are not protective against most _______________ infections.

44. Eosinophils • Normal Eosinophil Values: • dogs: 100-750/µL; cats: 0 – 750/µL) • ___________________ is usually a result of: • ___________________: Skin, respiratory, GI tract • ___________________________: Anaphylaxis, Fleas, food, grasses, or non-specific allergens. Feline asthma, Chronic ______________ (skin, GI, respiratory, urogenital), Tumor associated (fibrosarcoma, mast cell tumor, lymphoma, etc.) • ____________________is difficult to detect and evaluate because their numbers are normally ________.

45. Feline eosinophilic leukemia. Two neutrophils (arrows) are adjacent to several eosinophils at various stages of development that include segmented, band, metamyelocyte, and myelocyte forms

46. Basophils • Formation: • Basophils are produced in the bone marrow from the same PPSCs as other cells • Characteristics: • The granules of basophils are ________ soluble and often washed out during the staining procedure • _____________ are not always visible on a stained smear. • Function: • Basophils are the _________ phagocytic of the granulocytes. • Their granules contain ____________ and _____________ which are responsible for at least part of their function (not much is known about basophil production or function).

47. Basophils Above: feline basophil (left); canine basophil (right)

48. Basophil Function • Histamine and heparin granules • _______________ helps initiate __________________ and acute ___________ reactions. • _____________________ are attracted to the site of an allergic reaction by ____________________________________ released from the granules. • _____________ acts as a localized _____________________ to keep blood flowing to an injured or damaged area.

49. Basophilia and Basopenia Normal Basophil Values: ________ in both dogs and cats • ___________________ can be associated with an __________ or _________________ reaction in the tissue. • Sometimes ______________ and _______________________ are seen at the same time. • _______________________: is not clinically significant. Basophils should make up less than ___% of all WBCs in peripheral blood.

50. Agranulocytes • Recall that agranulocytes include • _________________________ • _________________________