MLAB 1415- Hematology Keri Brophy-Martinez

1. MLAB 1415- HematologyKeri Brophy-Martinez Chapter 7: Leukocytes

2. Terms • Myelopoiesis - the production and development of myeloid cells in the bone marrow

3. Leukocytes • Function • Protect the host from infectious agents or pathogens • Innate=Natural Immune response • 1st response to pathogens (nonself) • Always available • Adaptive Immune response • Starts in lymphoid tissue • Slow • Long-lasting immunity (memory) • Lymphocytes become “activated”

4. Differentiation & Maturation • Leukocytes develop from HSC (hematopoietic stem cell) • Common Myeloid progenitor (CMP) cell gives rise to the committed precursor cells for the neutrophilic, eosinophilic, basophilic and monocytic lineages. • Once lineage commitment has occurred, maturation begins.

5. Maturation Changes • Nucleoli disappear • Chromatin condenses • Nuclear mass indents and segments • Cytoplasm becomes granular & pinkens

6. Neutrophil Lifespan • Three areas • Bone marrow • Differentiation, proliferation, maturation • Peripheral blood • Circulate for a few hours • Tissues • Work in host defense

7. Regulation • Neutrophil production primarily regulated by cytokines • Interleukin-3 • Granulocyte -monocyte-colony stimulating factor (GM-CSF) • Granulocyte-colony-stimulating factor (G-CSF)

8. Maturation of the granulocytic series**Least mature to mature** • Myeloblast • Size: 14-20µm • Cytoplasm • Slight amount/blue • No granules ((possible Auer rods to be discussed in another section) • Nucleus • Round/Central or eccentric • Reddish purple • 1-3 nucleoli • Fine meshlike/lacy chromatin with no clumping • N:C ratio = 4:1

9. Maturation of the granulocytic series • Promyelocyte (progranulocyte) • Size: 15-21 µm • Cytoplasm • Increased amount/blue • Fine, azurophilic, nonspecific granules present • Nucleus • Round/Central or eccentric • Reddish-purple • Fine, meshlike/lacy chromatin beginning to clump • 1-2 nucleoli • N:C ratio = 3:1

10. Maturation of the granulocytic series • Myelocyte • Last myeloid cell capable of division • Size: 12-18µm • Cytoplasm • Increased, bluish-pink (“dawning of neutrophilia”) • Fine azurophilic, specific granules • Secretory vesicles • Nucleus • Round or oval/eccentric • Reddish-purple • Chromatin loosely clumped • No nucleolus • N:C ratio = 2:1 or 1:1

11. Maturation of the granulocytic series • Metamyelocyte • Also called “meta” or “juvenile” • Size: 10-18µm • Cytoplasm • Moderate amount • Specific granules (neutrophilic, eosinophilic or basophilic) • Tertiary granules • Secretory vesicles • Nucleus • Indented (kidney-bean shaped) • Central or eccentric • Purple • Clumped chromatin • No nucleolus • N:C ratio = 1:1

12. Maturation of the granulocytic series • Band • 1st stage found in P.B • Size: 9-15µm • Cytoplasm • Moderate amount • Specific granules (see meta) • Nucleus • Elongated, narrow band (sausage) shape with uniform thickness • Central or eccentric • Clumped, coarsely granular chromatin

13. Maturation of the granulocytic series • Segmented neutrophil, eosinophil or basophil • Size: 10-16µm • Cytoplasm • Neutrophil - Granules stain equally with eosin and methylene blue, causing a pinkish appearance • Normal in circulation = 40 - 80% • Eosinophil - Granules stain mainly red, due to reaction with eosin in stain • Normal in circulation = 0-5% • Basophil - Granules stain mainlyblue, due to the reaction with methylene blue in stain • Normal in circulation = 0-1% • Mast cell is a tissue basophil • Nucleus • Neutrophil - purplish-red, clumped granular chromatin, 2-5 distinct nuclear lobes connected by a filament of chromatin • Eosinophil - deep blue purple, coarsely granular chromatin, 2 distinct lobes • Basophil - deep blue purple, coarsely granular chromatin, 2 nuclear lobes (sometimes obscured by cytoplasmic granules)

14. Maturation of the granulocytic series Basophil Eosinophil Neutrophil, segmented

15. Function of the granulocytic series • Neutrophils • Defense against bacterial infections • Eosinophils • Regulate inflammation • Respond to antigenic stimulation in chronic allergies • Interact with larval stages of some helminthic parasites • Basophils • Histamine in granules plays a role in acute, systemic allergic reactions (sudden release of histamine can cause anaphylactic shock) • Granules also contain heparin

16. Granulocytes • Why are they called granulocytes? • The granules are lysosomes created by lipids that contain hydrolytic enzymes such as peroxidase, acid phosphatase, lysozyme and specific esterase.

17. MONOCYTES • Monopoiesis - the production and development of monocytes, macrophages and their precursors. Monocytes are produced in the bone marrow.

18. MONOCYTES • Function • Protect the host from infectious agents or pathogens via innate and adaptive immune response • Phagocytosis • Scavengers

19. MONOCYTES • Differentiation & maturation • Produced from CFU-GM • Can further differentiate into macrophages in the tissues • Lifespan • P.B ~ 8 hours • Tissue • Regulation • Production primarily regulated by cytokines • GM-CSF • IL-3 • M-CSF

20. Maturation of the monocytic series • Monoblast • Size: 12-20µm • Cytoplasm • Blue, moderate amount • No granules • Nucleus • Round, oval or slightly folded • Eccentric • Reddish-purple • Chromatin fine and lacy • Vacuoles may be present • 1-2 nucleoli • N:C ratio = 4:1

21. Maturation of the monocytic series • Promonocyte (difficult to discern) • Size: 12-20µm • Cytoplasm • Paler gray-blue and abundant • “Bleb-like” pseudopods at periphery • May contain fine red “dust-like” particles, azurophilic granules • Vacuoles may be present • Nucleus • Round with chromatin creases or brain-like folding • Central • Reddish-purple with light clumping of chromatin • 0-2 nucleoli • N:C ratio = 3:1 or 2:1

22. Promonocyte

23. Maturation of the monocytic series • Mature monocyte • Normal in circulation: 2-9% • Size: 12-20µm • Cytoplasm • Pale gray-blue and abundant • Bleb-like pseudopods at periphery, variable shape • Numerous fine pale red dustlike granule scattered throughout • Vacuoles common • Nucleus • Increased folding or elongated • Central • Purplish with finer clumped chromatin • No nucleoli • N:C ratio = 2:1 or 1:1

24. Monocyte

25. Macrophages • Once the monocyte enters the tissues, it becomes a macrophage • Size:15-80µm • Cytoplasm • Gray-blue and abundant • Ragged edges • Vacuoles common • Nucleus • Round with netlike appearance • Purplish with finer clumped chromatin • Nucleoli appear • Macrophage alias’ • Histiocytes in the loose connective tissue • Kupffer cells in the liver • Osteoclasts in the bone • Microglial cells in the nervous system • Also scattered throughout the body in all organ tissues (spleen, lung, abdomen, etc.) One monocytes have grown into macrophages, they do not normally reenter the bloodstream except in cases of inflammation

26. Macrophage

27. Lymphocytes

28. Differentiation & Maturation • Develop from HSC (hematopoietic stem cell) • Common Myeloid progenitor (CMP) cell and Common lymphoid progenitor cell (CLP) gives rise to the committed precursor cells: • T lymphocytes (60-80% of lymph population) • thymus • B lymphocytes (10-20%) • Bone marrow • Natural killer cells

29. Lifespan • Primary lymphoid tissue • Bone marrow • Secondary lymphoid tissue • Lymph node, spleen, Peyer’s patches in intestine and mucosal tissues such as tonsils • Active immune response occurs in above tissues where lymphs communicate and interact with antigen-presenting cells, phagocytes and macrophages

30. LYMPHOCYTES: T cells • Function • Cellular immune response (they do not produce antibodies) • Three subsets • Helper T cells • Instrumental in aiding B cells in antibody production • Suppressor T cells • Act as a “thermostat” to shut off the system or keep it under control • Cytotoxic T cells • Destroy by lysis of target cells (such as tumor cells) • Play a role in inflammatory response • Host defense when stimulated by interferon against certain viruses (cytomegalovirus - CM, and hepatitis) by killing the virally infected target cell before the virus replicates • Responsible for chronic organ graft rejection. (T cells have many unique antigens on their cell surfaces, some of which are the HLA - human lymphocyte antigen - markers. HLA markers are instrumental in chronic graft rejection and that is why tissue typing is done to establish a good donor match.)

31. LYMPHOCYTES: B cells • Function • Humoral immune response by transforming into antibody-producing plasma cells • Defense against encapsulated bacteria such as Streptococci • Mediate hyperacute organ rejection

32. Lymphocytes: NK cells • Function • Recognize and kill tumor cells • Recognize and kill cells infected with a virus

33. Maturation sequence of lymphocytes • Lymphoblast • Cell size: 10-18µm • Cytoplasm • Blue/scanty • No granules, Auer rods are never present • Nucleus • Purple, smooth chromatin • Smudged • Round, central or eccentric • 1-2 nucleoli • N:C ratio = 4:1

34. Maturation sequence of lymphocytes • Prolymphocyte (difficult to distinguish) • Size: 9-18µm • Cytoplasm • Blue, scanty • Usually granules are absent, but a few azurophilic granules may be present • Nucleus • Purple, condensed chromatin • Round or indented, eccentric • 0-1 nucleoli • N:C ratio = 3:1

35. Maturation sequence of lymphocytes • Mature lymphocyte • Normal adult value: 25-35% • Cell size: 7-10µm (a typical normal lymph has a nucleus that is the size of a normal RBC) • Cytoplasm • Light blue, scanty to moderate • Few azurophilic granules may be present • Nucleus • Purple, dense, clumped chromatin • Round or indented, eccentric • No nucleoli

36. Large lymphocytes versus monocytes • Size • Large lymph: 12-15µm • Mono: 15-18µm • Nucleus • Large lymph: clumped, condensed • Mono: lacy, brainlike folds • Granules • Large lymph: large azurophilic, easy to count • Mono: red, fine • Cytoplasm • Large lymph: clear, nongranular, may be indented by red cells • Mono: “ground glass” appearance, projection of blunt pseudopod blebs

37. Maturation sequence of lymphocytes • Variant lymphs • Other terms used are reactive, atypical, activated, Downey cell, transformed, virocyte, plasmacytoid • Can be T or B cells • Caused by antigenic stimulants such as viruses, post-transfusion reactions and organ transplants • General characteristics • Increased size due to DNA activity in the nucleus and RNA activity in the cytoplasm • Enlarged and/or monocytoid nucleus • Variation in nuclear chromatin pattern • Nucleus may contain 1-3 nucleoli • Abundant, sometimes foamy or vacuolated cytoplasm • Variation in cytoplasmic color - gray-blue to intense blue • Absence of granules in cytoplasm

38. Reactive lymphocyte

39. Plasma cells • Plasma cells • Function is the synthesis and excretion of antibodies (immunoglobulins) • Normally not present in the peripheral blood; comprise 2% of bone marrow cells. (May be seen in the peripheral blood in the disease called multiple myeloma, a disease of uncontrolled production of immunoglobulins.) • End stage of the B lymphocyte • Appearance • Size: 10-18µm • Cytoplasm is dark blue with perinuclear halo and may contain vacuoles indicating antibody synthesis • Nucleus is round, eccentric, dark purple with dense clumped chromatin • Variant plasma cells • Grape or Mott cell - cytoplasm completely filled with red, pink or colorless globules called Russell bodies • Flame cell - cytoplasm stains bright red-staining proteinaceous material

40. Plasma cells