1. Red Blood Cells & Anemias
RBC: ~5 x 106/ml; Generate ~ 8,000,000,000/hour
Andrew D. Leavitt, MD
Departments of Laboratory Medicine & Medicine
March 26, 2012

2. Outline – topics we will cover
Erythropoiesis – names, faces, & numbers (review)
RBC biology - Hemoglobin
O2 dissociation curves
Hemoglobin – fetal, adult
RBC membrane
Hemoglobin switching
Anemias: Production vs. destruction vs. loss
Membrane or contents – Hgb, enzymes
Immune mediated
Physical destruction
Anemia of inflammation
Ribosomopathies

3. CELL SIZE & CYTOPLASM COLOR
Erythropoiesis
Abbott Labs, Morphology
of Human Blood Cells
Pro-
Erythroblast
Basophilic
Polychromatic
Orthochromatic
Reticulocyte
RBC
CHROMATIN STRUCTURE
NUCLEAR SIZE & COLOR
CELL SIZE & CYTOPLASM COLOR
5 stages in the marrow

4. Red Blood Cell (RBC) Delivers O2 from your lungs to all your tissues
Takes CO2 from your tissues to your lungs
Also – binds NO (a vasodilator)
Their size: 8  [capillaries have ~3 M diameter]
Their life-span: 120 days
Mature RBCs have no nucleus
Too few RBCs = anemia

5. What tells your body to make RBCs?
EPO:
Is not stored, but expressed in response to the kidney sensing oxygen in the blood
It -
1. Increases # of E-committed progenitors
2. Increase GATA1 and FOG expression
3. Enhances anti-apoptotic gene expression
4. Increases transferrin receptor expression

6. Iron in a Porphyrin ring
Hemoglobin
Hemoglobin (Hgb) in the RBC carries O2, which is poorly soluble in water, to tissues &
CO2 from tissues to lungs.
~640,000,000 Hemoglobin molecules/RBC
Must coordinate: 1.heme and globin synthesis
2. a (Chromosome 11; 141 aa)
& b (Chromnosome 16; 146) chains
Colors of a bruise are globin breakdown products
b6Glu to Val
Tetramer of two heterodimers (a &b)
Heme
Iron in a Porphyrin ring
Linus Pauling
1949 – molecular diagnosis
2012 – no therapy

7. Hemoglobin oxygen dissociation curve
Arterial
O2 tension
Mean Venous
HbF
; CO2

8. Erythropoiesis: Hemoglobin Switching (ALWAYS: 2 alpha & 2 “beta-like” [3 ‘b’ options])
Embryo
Fetus
Birth
Adult
6 mo.
Globin chain synthesis (%) 50 a b d g z e
Alpha: Chromosome 16
Beta-like: Chromosome 11
HgF a2g2 (75)
HgA a2b2 (25)
HgA a2b2 (96-98)
HgA2 a2d2 (2-3)
HgF a2g2 (< 1)
Hoffman et al. Hematology 2000

9. Schematic of red cell membrane depicting proteins crucial for normal membrane mechanical strength* *
*mutations lead to hereditary spherocytosis

10. Normal Values for Hematology Tests - UCSF

11. Thinking about anemia…a clinical perspective
Framework:
RBC size (MCV) & is the marrow responding (reticulocyte count)
Etiology:
Cell intrinsic:
1. Hgb – too little; abnormal type; unequal a and b synthesis
2. Enzyme
3. Membrane
Cell extrinsic:
Immune mediated
Physical destruction
Blood loss
Marrow does not produce
Primary – leukemia, myelodysplasia
Secondary – vitamin deficiency

12. Reticulocyte Stain – detects the ribonucleoprotein in the young RBC

13. #1.1 Too little Hgb: Iron Deficiency Anemia
Small cells – microcytic (MCV < 80 fL)
Young women – OK
Older people – Worry about gastrointestinal blood loss
normal peripheral blood (40x)
Iron deficiency anemia, peripheral blood (40x)
Lead poisoning can look like Fe deficiency – Lead blocks heme synthesis

14. Iron Deficiency Anemia
peripheral blood, 100X
Hypochromic
RBCs
Elliptocytes
Iron stian of bone marrow – none found
~ 500 million people worldwide

15. The Iron Circuit – mostly a game of recycling
Daily absorption:
Duodenum ~ 1 mg
Daily loss:
urine, feces, skin, hair ~ 1 mg
Transferrin
Plasma (4 mg)
Ineffective erythropoiesis
Macrophage
(0.5 – 1.5 gm)
Bone Marrow
Erythroblasts
( ~ 150 mg)
Circulating RBCs (1.7 – 2.4 gm)
Menstrual loss/hemorrhage

16. Iron Circuit in Disease
Weiss & Goodnough NEJM 2005
Iron Circuit in Disease
Infection/inflammation/malignancy
IL-6/LPS hepcidin (25aa protein)
absorption
macrophage Fe storage
Impairs Erythropoiesis:
Decrease EPO
Directly suppresses marrow
Interferon – g
TNF – a
IL-1, IL-10

17. Iron stain of bone marrow aspirates
Iron deficienyc anemia - iron stain
Anemia of inflammation – iron stain
Storage iron
(blue)

18. #1.2 Abnormal Hgb: Sickle cell disease
Multi-system disease
Cardiovascular/strokes
Kidneys
Skin
Lungs
Immune system
Treatment:
Supportive care
Transfusions
Demethylating agents
Stem cell transplants

19. #1.3 Abnormal a/b Globin ratio: Thalassemia
Small cells – microcytic (MCV < 80 fL)
Imbalance between a and b chains
Name based on which chain is deficient [a or b thal]
Make lots of RBCs, just unstable and lyse, so increased retics
Alpha Thal:
silent carrier -a / aa
trait -- / aa or -a / -a
Hgb H -a / --
Hydrops fetalis -- /--
Beta Thal:
Minor
Intermedia
Major

20. Erythropoiesis: Hemoglobin Switching (ALWAYS: 2 alpha & 2 “beta-like” [3 ‘b’ options])
Embryo
Fetus
Birth
Adult
6 mo.
Globin chain synthesis (%) 50 a b d g z e
Alpha: Chromosome 16
Beta-like: Chromosome 11
HgF a2g2 (75)
HgA a2b2 (25)
HgA a2b2 (96-98)
HgA2 a2d2 (2-3)
HgF a2g2 (< 1)
Hoffman et al. Hematology 2000

21. Normal peripheral smear
Alpha thal trait
Normal peripheral smear
Hemoglobin H, -- / -A
Target cells
Retics
Beta-thalassemia major
Nucleated RBC
Target cells

22. Severe thalassemia Beta-thalassemia major (transfused)
Nucleated RBC
Target cells
Beta-thalassemia major (transfused)
Nucleated
RBC
Normal RBCs
(transfused)

23. #2 Enzyme: G-6-PD deficiency
(Heinz body hemolytic anemia)
Reduced glutathione (GSH)
is critical for RBCs to counter
oxidative stress.
Bite cells
Arrows show Heinz bodies
Sulfa drugs, fava beans, antimalarials..…

24. #3: Membrane: red cell membrane depicting proteins crucial for normal membrane mechanical strength* *
*mutations lead to hereditary spherocytosis

25. Hereditary spherocytosis (HS)
Reticulocytes
spherocytes

26. Thinking about anemia…a clinical perspective
Framework:
RBC size (MCV) & is the marrow responding (reticulocyte count)
Etiology:
Cell intrinsic:
1. Hgb – too little; abnormal type; unequal a and b synthesis
2. Enzyme
3. Membrane
Cell extrinsic:
Immune mediated
Physical destruction
Blood loss
Marrow does not produce
Primary – leukemia, myelodysplasia
Secondary – vitamin deficiency

27. Autoimmune hemolytic anemia
Reticulocytes
spherocytes

28. Testing for anti-RBC antibodies: attached or in circulation
Hemolytic disease of the newborn

29. RBC destruction - extrinsic
Erythroid Hyperplasia, TTP, bone marrow aspirate (100x)
Thrombotic thrombocytopenic purpura (TTP)
schistocytes
(RBC fragments)
Hemolytic uremic syndrome (HUS)
schistocytes
(RBC fragments)
Retics

30. Megaloblastic anemia (MCV > 100)
B12 deficiency
Folate deficiency
Affects DNA synthesis

31. Megaloblastic anemia (MCV > 100)
Anemia yet hypercellular marrow – ineffective erythropoiesis

32. Ribosomopathies and hematopoietic disorders
Our findings indicate that the erythroid lineage has a low threshold for the induction of p53, providing a basis for the
failure of erythro- poiesis in the 5q- syndrome, DBA, and perhaps other bone marrow failure syn- dromes.
Narla & Ebert 2010

33. Thanks for your attention!

34. Blood is good…everyone should have some!
Take home message……
Blood is good…everyone should have some!
Share it…become a blood donor if you can
UCSF Blood donor center:
Millberry Union next to Subway sandwich shop
The best donor center nurses in town