1. Practical Clinical Hematology
Leukemia 11

2. THE LEUKEMIAS
malignancies involving lymphoid or hematopoietic cell lines
increase in circulating leukocytes (not invariable)
bone marrow always involved
replacement/hypoproliferation of normal hematopoietic cells
anemia, infection, hemorrhage
leukemic infiltration in liver, spleen, lymph nodes, and other tissues

3. THE LEUKEMIAS
acute leukemias
chronic leukemias

4. ALL AML Hematopoietic stem cell Neutrophils Eosinophils Basophils
Monocytes
Platelets
Red cells
Myeloid
progenitor
Lymphoid
B-lymphocytes
T-lymphocytes
Plasma
cells
germinal center
naïve
AML

5. THE LEUKEMIAS acute leukemias chronic leukemias
acute lymphoblastic leukemia (ALL)
acute myeloblastic leukemia (AML)
chronic leukemias
chronic lymphoid leukemia (CLL)
chronic myelogenous leukemia (CML)

6. Morphological Features of Blasts in Acute Myeloid and Acute Lymphoblastic Leukemias
ALL
AML
Feature
Variable, small to medium size
Larger, usually uniform
Blast size
Coarse to fine
Usually finely dispersed
Nuclear chromatin
Absent or 1 or 2, often indistinct
1-4, often prominent
Nucleoli
Usually scant, coarse granules sometimes present (~7%)
Moderately abundant, fine granules often present
Cytoplasm
Not present
Present in 60%-- 70% of cases
Auer rods
Usually not dysplastic
Often dyspastic changes in maturing myeloid cells
Other cell types

8. AML

9. The original FAB-Classification system of Acute Leukemias,
Chronic leukemias
Myeloid
Lymphoid L1 L2 L3 M0 M1 M2 M3 M4 M5 M6 M7
Leukemias
The original FAB-Classification system of Acute Leukemias,
heavily based on morphologic findings

10. Maturation and Morphology of Immature Granulocytes
myeloblast
promyelocyte
myelocyte
metamyelocyte
band
neutrophil
MATURATION

11. Maturation and Morphology of Immature Granulocytes
Myeloblast:
The first and earliest granulocyte (15 μm)
High nucleus to cytoplasm (N:C) ratio (5:1)
Round or oval nucleus with loose light staining euchromatin
1-2 nucleoli
Has minimal light blue cytoplasm
Contains no cytoplasmic granules
Begins to produce myeloperoxidase granules (MPO)
Comprises 1% of the nucleated cells in the bone marrow

12. Promyelocyte: larger than a myeloblast (20 μm) High N:C ratio (3:1)
Loose chromatin with nucleoli
Dark blue cytoplasm
Contains large nonspecific cytoplasmic granules
Containing myeloperoxidase (MPO)
Comprises 3-4% of nucleated bone marrow cells

13. Neutrophilic Myelocyte:
medium cell size (12 μm)
High N:C ratio (3:1)
Round, oval, or slightly indented nucleus with darker
blue heterochromatin
Last stage of cell division
Has active RNA, therefore, the cytoplasm is blue
Contains MPO and secondary granules containing leukocyte alkaline phosphatase
Comprises 12% of bone marrow nucleated cells

14. Neutrophilic Metamyelocyte:
size (11 μm)
N:C ratio (2:1)
last mononuclear stage, no mitosis
Nucleus is kidney or horseshoe shaped, and has condensed heterochromatin
Has a prominent Golgi apparatus – clear area located at the indentation site of the nucleus
Cytoplasm is similar to the mature cell
Comprises 18% of bone marrow cells

15. Band Same size as a mature neutrophil (10-12 μm)
N:C ratio has reversed (1:2)
Nucleus is band- or sausage-shaped without segmentation
Cytoplasm is filled with small neutrophilic granules
Last immature stage
Comprises 11% of bone marrow cells and 0-3% of peripheral WBCs Stored in the bone marrow and released when there is an increased demand for neutrophils
• Shift to the left is an increase in immature cells indicating increased demand for WBCs in peripheral blood

16. Neutrophils
Also known as segmented neutrophils, segs, polymorphonuclear cells, polys, and PMNs
N:C ratio is 1:3, and the size is μm
Average nucleus contains 3-5 segments connected by narrow filaments
Hyposegmented is less than 3 segments, and may indicate a shift to the left or an anomaly
Hypersegmented is more than 5 segments and may indicate infection or megaloblastic anemia
Cytoplasm contains very small neutral granules
Makes up 55-75% of all peripheral WBCs
• Granules can become larger upon bacterial infection producing toxic granulation, which are numerous, large, basophilic granules

17. Eosinophils Average size is 13 μm Nucleus is generally bilobed
Cytoplasm is bright red or orange which is due to large specific, secretory granules containing peroxidase, acid phosphatase, aryl sulfatase, beta-glucuronidase, etc. that stain red with the eosin component of Wright’s stain

18. Basophils Is the smallest granulocyte at 10 μm
The nucleus is difficult to see due to heavy granulation
Cytoplasm contains large specific, secondary granules that contain heparin and histamines, which stain purple with Wright’s stain. These granules are water soluble and sometimes appear as holes in the cell if the cells are not fixed well during staining.
Makes up to 0.5% of peripheral WBCs
• Note: Tissue mast cells are similar to basophils but are larger and have no developmental relationship with basophils. Mast cells have a mesenchymal (connective tissue) origin and have granules containing serotonin (basophilic granules contain no serotonin).

19. Acute Non-Lymphoblastic Leukaemia
Class
Alternative
Bone Marrow Appearance M0
AML with minimal differentiation
Identified by ultrastructural myeloperoxidase activity or immunophenotyping. M1
AML without maturation
Monomorphic with one or more distinct nucleoli, occasional auer rod and at least 3% myeloperoxidase positivity. M2
AML with maturation
50% OR > myeloblasts & promyelocytes and common single auer rod. Dysplastic myeloid differentiation may also be present. M3
APL
Dominant cell type is promyelocyte with heavy azurophilic granulation. Bundles of Auer rods confirm diagnosis. Microgranular variant exist (M3v) M4
AMMoL
As M2 but > 20% promonocytes & monocytes. M5
AMoL
> 80% monoblasts is poorly differentiated (M5a)
> 80% monoblasts, promonocytes or monocytes is well differentiated (m5b) M6
AEL
>50% bizzar, dysplastic nucleated red cells with multinucleate forms and cytoplasmic bridging. Myeloblasts usually > 30%. M7
AMegL
Fibrosis, heterogeneous blasts population with cytoplasmic blebs. Platelet peroxidase positive.
myeloblastic leukemia with minimal differentiation; CD13 or CD33 positive

20. AML

21. AML M0 the blasts are undifferentiated by morphology and cytochemistry
The cytoplasm is usually scant, and grey to light blue in color without granules, and Auer rods are not seen.
N:C ratio is high
The nucleus is round to oval or irregular and usually eccentrically placed
The nuclear chromatin ranges from being finely granular and evenly dispersed to being slightly clumped
One or more nucleoli may be visible

22. AML M0 the enzyme MPO demonstrated by immunocytochemistry analysis
Bone marrow smear, May-Giemsa stain, x1000

23. AML M1
The blasts vary in size, but are approximately the size of mature segmented neutrophils
The nuclei are round to slightly oval or irregular, and the nuclear chromatin ranges from finely granular and evenly distributed to slightly clumped.
One to two nucleoli are usually visible
The cytoplasm is scant and grey-blue to light blue in color, and agranular.
N:C ratio is high
Auer rods are seen in a minority of blasts

24. M1

25. M1

26. Immunostain antibody: CD33 Bone marrow smear, Peroxidase stain

27. FAB M1

28. FAB M1
AUER
ROD

29. M2

30. M2

31. M2
with eosinophilia
Peroxidase stain

32. FAB M2

33. FAB M2

34. FAB M3

35. FAB M3

36. FAB M3

37. FAB M4

38. FAB M4
Double esterase
stain

39. FAB M5

40. FAB M5

41. FAB M5

42. FAB M6

43. FAB M6

44. FAB M6

45. FAB M6
PAS Stain

46. ALL

47. ACUTE LYMPHOBLASTIC LEUKEMIA (ALL)
lymphoblast predominant cell type
most frequent in children
better prognosis than AML
FAB classification L1-L3
not used as a diagnosis, just as a descriptor
entirely based on morphology

48. Morphological Features of Acute Lymphoblastic Leukemia
large, homogeneous
large, heterogeneous
small to intermediate
Cell size
uniform
pleiomorphic indentations common
Nuclear shape
often single
large, prominent
small or absent
Nucleoli
moderate (basophilic)
scanty
Cytoplasm
prominent
none to few
Cytoplasmic vacuoles

49. FAB L1 type of ALL

50. FAB L2 type of ALL

51. FAB L2 type of ALL

52. FAB L3 type of ALL

53. FAB L3 type of ALL

54. FAB L3 type of ALL

55. Peripheral blood film in L1 (acute lymphoblastic leukaemia) (ALL) showing lymphoblasts and one nucleated red blood cell (NRBC). The lymphoblasts vary in size but are relatively uniform in morphology. The smaller blast cells show some chromatin condensation, which can be a feature of lymphoblasts but not of myeloblasts. This case was shown on immunophenotyping to be of B lineage.

56. Peripheral blood film in L2 acute lymphoblastic leukaemia (ALL)
Peripheral blood film in L2 acute lymphoblastic leukaemia (ALL). The blast cells are larger and more pleomorphic than in LI ALL and in this case have a more diffuse chromatin pattern; one of the blasts has a hand-mirror conformation. This case was shown on immunophenotyping to be of T lineage

57. Other Lymphoid Leukemia

58. Hairy cell leukemia. Lymphocytes with filamentous cytoplasmic projections

59. Hairy cell leukemia

60. Hairy cell leukemia

61. Hairy cell leukemia (positive TRAP stain)

62. Prolymphocytic leukemia.

63. (A)and (B), Hypergranular acute promyelocytic leukemia, promyelocytes with prominent azurophilic granules.
(C) Hypergranular APL with multiple Auer rods.
(D) Microgranular APLv. These abnormal promyelocytes have lobulated nuclei and absent or fine azurophilic granules.

64. Chronic Leukemia

65. CHRONIC LEUKEMIAS longer clinical course
chronic lymphocytic (lymphatic) leukemia (CLL)
mature-appearing lymphocytes
chronic myelocytic (myeloid, myelogenous, granulocytic) leukemia (CML, CGL)
segs, bands, metamyelocytes, and myelocytes

66. CLL

67. CML

68. CML

69. CML

70. CML

72. Thank you for your attention.