1. MLAB 1315- Hematology Keri Brophy-Martinez
Unit 3: Leukocytes

2. NEUTROPHILS
Myelopoiesis - the production and development of myeloid cells in the bone marrow
Growth factors (colony-stimulating factors - CSF’s) and interleukins regulate blood cell development by:
Mediating proliferation
Differentiation into cell lines
Maturation of hematopoietic progenitor cells

3. NEUTROPHILS
Three series of granulocytes (named for colors taken up by granules in Wright’s stain)
Neutrophils
Granules stain equally with eosin and methylene blue
Normal in circulation = %
Eosinophils
Granules stain mainly red
Normal in circulation = 0-4%
Basophils
Granules stain mainly with methylene blue
Normal in circulation = 0-2%

4. Maturation of the neutrophilic series
Stem cell - an unspecified cell that gives rise to a specific specialized cell, such as a blood cell
Multipotential and cannot be identified morphologically
Can self-renew and differentiate

5. Maturation of the neutrophilic series
Myeloblast
Size: 10-20µm
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 chromatin with no clumping
N:C ratio = 4:1

6. Maturation of the neutrophilic series
Promyelocyte (progranulocyte)
Size: 10-20µm
Cytoplasm
Increased amount/blue
Fine, azurophilic, nonspecific granules present
Nucleus
Round/Central or eccentric
Reddish-purple
Fine, meshlike chromatin beginning to clump
1-2 nucleoli
N:C ratio = 3:1

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

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

9. Maturation of the neutrophilic series
Band
Size: 10-16µm
Cytoplasm
Moderate amount
Specific granules (see meta)
Nucleus
Elongated, narrow band (sausage) shape with uniform thickness
Central or eccentric
Clumped, coarsely granular chromatin

10. Maturation of the neutrophilic series
Segmented neutrophil, eosinophil or basophil
Size: 10-16µm
Cytoplasm
Neutrophil - specific fine violet pink
Eosinophil - red uniform specific granules
Basophil - coarse violet blue non-uniform specific granules (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)

11. Maturation of the neutrophilic series

12. Function of the neutrophilic 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

13. MONOCYTES
Monopoiesis - the production and development of monocytes, macrophages and their precursors. Monocytes are produced in the bone marrow. Their primary function is phagocytosis.

14. MONOCYTES Other names for macrophages
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

15. 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 thready
Vacuoles may be present
1-2 nucleoli
N:C ratio = 4:1

16. Maturation of the monocytic series
Promonocyte (difficult to discern)
Size: 21-20µm
Cytoplasm
Paler gray-blue and abundant
“Bleb-like” pseudopods at periphery
May contain fine red “dust-like” particles
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

17. Promonocyte

18. Maturation of the monocytic series
Mature monocyte (Normal in circulation: 2-9%)
Size: 15-18µm
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

19. Monocyte

20. LYMPHOCYTES Normal adult value: 21-51%
Produced in the bone marrow (primary lymphoid tissue)

21. LYMPHOCYTES
Growth factors cause lymphs to differentiate. They proliferate and mature in the primary lymphoid organs:

22. LYMPHOCYTES Thymus - T cells Function
Cellular immune response (they do not produce antibodies)
Regulate antibody reactions by helping or suppressing activation of B lymphs. T cells act as “helper cells” (HIV or AIDS virus causes a destruction of these helper cells), which are instrumental in aiding B cells in antibody production, and “suppressor cells”, which act as a “thermostat” to shut off the system or keep it under control. The helper to suppressor ratio is very important and is normally 2:1.
Protect against viral, bacterial, fungal or protozoan infection
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. Cytotoxic and helper T cells act against these HLA markers if they are foreign to the host organism. HLA markers are instrumental in chronic graft rejection and that is why tissue typing is done to establish a good donor match.)

23. LYMPHOCYTES Bone marrow - B cells Function
Humoral (antibody) immune response by transforming into antibody-producing plasma cells
Defense against encapsulated bacteria such as streptococci
Mediate hyperacute organ rejection

24. LYMPHOCYTES Null cells - unknown maturation sequence Function
Destroy by lysis target cells (such as tumor cells) by cytotoxins
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

25. LYMPHOCYTES
T and B cell lymphocytes cannot be differentiated morphologically. They are distinguished functionally and by immunologic marker studies (to be discussed later.) Null cells appear as large lymphocytes with azurophilic granulocytes and are called large granular lymphocytes (LGL).

26. 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

27. Maturation sequence of lymphocytes
Lymphoblast (which is preceded by a lymphoid stem cell)
Cell size: 10-20µm
Cytoplasm
Blue/scanty
No granules, Auer rods are never present
Nucleus
Purple, smooth chromatin
Round, central or eccentric
1-2 nucleoli
N:C ratio = 4:1

28. 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 = 4:1

29. Maturation sequence of lymphocytes
Mature lymphocyte
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

30. Large lymphocytes versus monocytes
Size
Large lymph: µm
Mono: µm
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

31. Maturation sequence of lymphocytes
Variant lymphs
Other terms used are reactive, atypical, Downey cell, transformed, virocyte, plasmacytoid
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

32. Reactive lymphocyte

33. 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

34. Plasma cells