Hematology Flashcards
RBC
They make up the majority of cellular components of blood. They are highly specializesd and have a few uneique features: 1) they lack a nucleus 2) and mitochondria (are anaerobic dependent and therefore need ways to prevent build of O radicals in the cell). They loose these organelles prior to being released from the bone marrow into the periphery.
Hemostatsis
(the arrest of bleeding), which allows blood to clot in response to damage to a blood vessel. Hemostasis results from the complex interactions between the platelet, the endothelial lining of blood vessels, and the blood coagulation factors in response to disruption of the endothelium at sites of injury. This process is counterbalanced by inhibitory factors and the fibrinolytic system, which is responsible for breaking down formed clots. This system has to be finely tuned to allow clotting to take place when necessary while preventing uncontrolled propagation of clots when they do form or formation of pathologic clots (thromboses).
Anemia
decrease in the amount of red blood cells (RBCs) or the amount of hemoglobin in the blood. It can also be defined as a lowered ability of the blood to carry oxygen.
Erythropoiesis
: the process, which produces red blood cells (erythrocytes). It is stimulated by decreased O2 in circulation, which is detected by the kidneys, which then secrete the hormone erythropoietin. This hormone stimulates proliferation and differentiation of red cell precursors, which activates increased erythropoiesis in the hemopoietic tissues, ultimately producing red blood cells.
Lymphocytes
the key players in the adaptive immune response, which involves the development of “memory” following exposure to an infectious agent, providing the ability to respond more vigorously to repeated exposure to the same agent. Diameter: 7–12µm. Mostly small, can be large if reactive. Nucleus round or slightly indented. Condensed chromatin. Usually scanty bluish cytoplasm, may contain a few azurophilic granules
neutrophils
(also known as polymorphonuclear cells or PMNs): the most abundant (40% to 75%) type of white blood cells in mammals and form an essential part of the innate immune system. They are formed from stem cells in the bone marrow. They are short-lived and highly motile. Neutrophils may be subdivided into segmented neutrophils (or segs) and banded neutrophils (or bands). They form part of the polymorphonuclear cell family (PMNs) together with basophils and eosinophils. Diameter: 9–15µm, Cytoplasm slightly acidophilic, Many very fine granules, 2-5 nuclear segments/lobes
monocytes
a type of white blood cells (leukocytes). They are the largest of all leukocytes. They are part of the innate immune system. They are amoeboid in shape, having clear cytoplasm. Monocytes have bean-shaped nuclei that are unilobar. Monocytes constitute 2% to 10% of all leukocytes in the human body. They play multiple roles in immune function: (1) replenishing resident macrophages under normal states, and (2) in response to inflammation signals, monocytes can move quickly to sites of infection in the tissues and divide/differentiate into macrophages and dendritic cells to elicit an immune response. Half of them are stored in the spleen. Monocytes are usually identified in stained smears by their large kidney shaped or notched nucleus. These change into macrophages after entering into the tissue spaces, and in endothelium can transform into foam cells. Diameter: 15–30µm. Large and eccentric nucleus, round, kidney/horseshoe-shaped or lobulated. Chromatin, skein-like or lacy appearance. Abundant cytoplasm, grayish-blue, few to many fine azurophilic granules. May have intracytoplasmic vacuoles
eosinophils
are white blood cells and one of the immune system components responsible for combating multicellular parasites and certain infections in vertebrates. Diameter: 12–17µm. Numerous large, round and red-orange granules. 1-4 nuclear lobes, mostly 2.
Basophils
Basophils contain large cytoplasmic granules which obscure the cell nucleus under the microscope when stained. However, when unstained, the nucleus is visible and it usually has two lobes. Basophils appear in many specific kinds of inflammatory reactions, particularly those that cause allergic symptoms. Basophils contain anticoagulant heparin, which prevents blood from clotting too quickly. They also contain the vasodilator histamine, which promotes blood flow to tissues. They can be found in unusually high numbers at sites of ectoparasite infection, e.g., ticks. Like eosinophils, basophils play a role in both parasitic infections and allergies. Diameter: 12µm. Numerous large round purple-black cytoplasmic granules. Usually two nuclear lobes, but often covered by granules
complete blood count (CBC),
a very commonly used clinical test, will also calculate the hematocrit for you, but it will provide you with much more information as well. It will tell you the hemoglobin concentration in the blood. Hemoglobin is the protein in red blood cells that binds to and carries oxygen, so measuring hemoglobin gives you important information about the oxygen carrying capacity of someone’s blood. If you don’t have enough hemoglobin in your blood (for whatever reason – there are lots of them) you have anemia. A CBC also gives you a precise measurement of the number and size of each of the different blood cell types as well as percentages of the different types of white blood cells (the “differential”).
peripheral smear
A drop of blood can also be smeared on a glass slide, stained, and examined under the microscope to look for any abnormally shaped cells or cellular inclusions.
sickle cell mutation
mutations can lead to a situation where the hemoglobin molecules in certain situations tend to polymerize into long chains or form crystals, leading to abnormally-shaped cells that are fragile and easily destroyed. The most common of these mutations (a substitution of valine for glutamic acid at the 6th position of the beta-globin chain) makes hemoglobin S
thalassemia mutation
mutations in the promoter regions of the globin genes can lead to an imbalance in the number of alpha-globin and beta-globin chains produced in the RBC
porphyria mutation
mutations in the enzymes involved in the synthesis of the heme prosthetic group, leading to a rare disease known as porphyria.
Other mutations in red blood cells
There can be mutations of the hemoglobin molecule that cause it to bind with greater or lesser affinity to the oxygen. Other mutations can make the hemoglobin molecule unstable, leading to premature breakdown and RBC destruction (termed “hemolysis”). Finally, because mature RBCs lack nuclei, they can’t make new RNA, so they have limited ability to respond to changes in the environment. Once they’re released in the periphery, they’re stuck with what they’ve got and, if damaged, have limited ability to repair themselves. Also, since they lack mitochondria, they are dependent on anaerobic metabolism for generation of ATP to maintain critical cellular processes. Thus, mutations in the glycolytic pathway, such as pyruvate kinase deficiency, can lead to another type of hemolytic anemia.
glucose-6-phosphate dehydrogenase (G6PD) deficiency
The most common cause of hemolytic anemia. RBCs must also have the ability to reduce reactive oxygen species which can accumulate in the cell with time and cause cellular damage. Mutations of the genes that encode the enzymes responsible for this function can also be a cause for hemolytic anemia. an X-linked disorder seen in ~15% of the African male population. G6PD is the most common human enzyme defect, being present in more than 400 million people worldwide.
iron deficiency
Anemia can also occur when the bone marrow isn’t making enough RBCs. To make RBCs, the bone marrow needs enough of the necessary substrates. There must be enough iron available to be incorporated into the hemoglobin molecule; one of the most common causes of anemia is iron deficiency. When someone is iron deficient, it is important to know why. It may be due to decreased dietary intake. Often, however, it can be due to blood loss of some form or another that the patient may not even know about, such as occult (not clinically detectable) bleeding from the gastrointestinal tract due to a cancer in the colon. Vitamin B12 and Folic acid are also necessary for the developing RBCs to be able to undergo normal cell division, so when deficiencies of these vitamins occur, anemia results.
erythropoietin
a hormone produced by the kidney called erythropoietin (“red making”), is essential for stimulating the marrow to make red blood cells. Under certain clinical conditions, such as kidney failure, RBC production is decreased due to a lack of erythropoietin production, and anemia results.
“myeloid” cell types
neutrophils (also known as polymorphonuclear cells or PMNs), monocytes, eosinophils, and basophils. are critical components of the innate immune system. Innate immunity provides protection against infection that relies on mechanisms that exist before infection, are capable of a rapid response to microbes, and react in essentially the same way to repeat infections.
Hematologic malignancies
These are all clonal, neoplastic conditions, meaning that the malignant cells have undergone a series of genetic mutations that have altered their differentiation and/or proliferative capacity. Some malignancies have classic mutations associated with them, such as the t(9;22) translocation (also known as the Philadelphia chromosome) associated with chronic myelogenous leukemia (CML). Others are not associated with any characteristic cytogenetic abnormalities.
myeloma
arising from one of the other cell types in the marrow
anemia
is not a diagnosis in of itself. Many causes: nutritional deficiencies (iron defecient, Vit B12, folate) Kidney disease, can also be due to inflammation, hemolysis (red cell destruction).
Hematopoiesis
The processes of making blood. After birth, both white and red blood cells are produced in the bone marrow and released into the peripheral blood when they reach maturity. All blood cells arised from a hematopoietic stem cell, differentiating along different lines of development to produce all the different blood cell types. With RBCs, the nucleus continues to shrink as divisions occurs and becomes red as hemoglobin becomes incorporated. Hematopoiesis is the complex process, usually occurring predominantly in the marrow, which results in the formation of the mature, functional red blood cells, white blood cells, platelets and miscellaneous other cell types (osteoclasts, dendritic cells, etc).
aplastic anemia
a disease in which the bone marrow, and the blood stem cells that reside there, are damaged. This causes a deficiency of all three blood cell types (pancytopenia): red blood cells (anemia), white blood cells (leukopenia), and platelets (thrombocytopenia). Aplastic refers to inability of the stem cells to generate the mature blood cells.
spherocytosis
an auto-hemolytic anemia (a disease of the blood) characterized by the production of red blood cells (RBCs), or erythrocytes, that are sphere-shaped, rather than bi-concave disk shaped. Spherocytes are found in hereditary spherocytosis and autoimmune hemolytic anemia. This occurs because vesicles break off of RBC decreasing the surface area to volume ratio. No cenral pallor due to decreased cell membrance. increased MCHC
Autoimmune hemolytic anemia
occurs when antibodies directed against the person’s own red blood cells (RBCs) cause them to burst (lyse), leading to insufficient plasma concentration. The lifetime of the RBCs is reduced from the normal 100–120 days to just a few days in serious cases. The intracellular components of the RBCs are released into the circulating blood and into tissues, leading to some of the characteristic symptoms of this condition. The antibodies are usually directed against high-incidence antigens, therefore they also commonly act on allogenic RBCs (RBCs originating from outside the person themselves, e.g. in the case of a blood transfusion)[
What COMPLETE BLOOD COUNT (CBC) can tell you
One of the most commonly ordered tests in medicine. Provides information on: Red blood cells: Number, size, hemoglobin content White blood cells: Total count, number and percentage for each type. Platelets: Number, size
MCV
mean corpuscular volume. Mean size of red blood cells. Determined directly using the Coulter principle or manually (MCV = HCT ¸ RBC). Abnormal values due to: Low MCV: Microcytosis, iron deficiency anemia or thalassemia, High MCV: Macrocytosis, megaloblastic anemia, orAnemia can be classified as: Microcytic (MCV100)
RDW
RBC distribution width 11.7-14.2. Measure of the variability in size of red cells. The wider the red cell histogram, the higher the RDW. Increased in anemia and disease with RBC destruction (i.e. schistocytosis)
Normal RBC Morphology
Circular biconcave disc-shaped, Size: 6.7–7.7µm, mean 7.5µm, Lack of nuclei, Eosinophilic cytoplasm, Central area of pallor, <1/3 of diameter, the diameter is about half of a neutrophil
Red Blood Cell Count
(4-6) Obtained using the Coulter principle. Abnormal values due to: Anemia: Decreased due to blood loss, peripheral destruction, or insufficient erythropoiesis in the marrow or Erythrocytosis/ Polycythemia: Increased due to reactive changes (smoking, renal cell carcinoma), thalassemia, or primary marrow neoplasm (polycythemia vera)
HGB
Hemoglobin concentration (14-18) Determined spectrophotometrically after conversion to cyanmethemoglobin. Abnormal values due to: Anemia: Decreased due to blood loss, peripheral destruction, or ineffective erythropoiesis in the bone marrow. Erythrocytosis/Polycythemia: Increased due to reactive changes (smoking, renal cell carcinoma), or primary marrow neoplasm (polycythemia vera)
HCT
hematocrit (M= 39-50, F= 35-47). Volume of red blood cells in whole blood. Obtained directly or by calculation (HCT = RBC × MCV). Rule of thumb: HCT = ~ 3 × HGB. Abnormal values due to: Decreased due to anemia or fluid overload or Increased due to erythrocytosis/polycythemia or dehydration. It is the volume of red blood cells in the whole blood (about 40%)
MCH
Mean cell hemoglobin. Mean quantity of hemoglobin in a single red cell. Parallels MCV: MCV goes up or down, MCH goes up or down. Calculated from directly determined HGB and RBC: MCH = HGB ¸ RBC. Abnormal values due to: Low MCH: Hypochromatic, iron deficiency anemia. High MCH: Hyperchromatic, megaloblastic anemia. Normal value is 27.5-35
MCHC
Mean cell hemoglobin concentration. Average concentration of hemoglobin in a single red cell or “concentration of hemoglobin in packed red cells”. Calculated from the directly determined HGB and the indirectly determined HCT: MCHC = HGB ¸ HCT = MCH ¸ MCV. Abnormal values due to: Decreased in moderate to severe microcytic anemia or Increased in hereditary spherocytosis. Normal= 32-36
Neutropenia
Decreased absolute neutrophil count (ANC), < 0.5×109/L. Causes: Infections: Gram-negative septicemia, typhoid and paratyphoid fevers, CMV, HIV, EBV, HCV, measles, and HIV; Drugs, medication, ionizing radiation; Marrow diseases: leukemia, myelodysplastic syndromes, aplastic anemia; Bone marrow infiltration by tumors; Autoimmune disease: immune neutropenia, SLE, rheumatoid arthritis; Congenital: cyclical neutropenia, familial benign chronic neutropenia, severe congenital neutropenia, congenital aleukia
Neutrophilia
Increased absolute neutrophil count (ANC), >11.1 × 109/L. Causes: Physiologic: neonates, exercise, emotion, pregnancy, parturition, lactation; Acute inflammation caused by infections; Acute inflammation caused by surgery, infarcts, autoimmune, etc.; Endocrine/metabolic: Cushing’s syndrome, thyrotoxicosis, uremia, etc.; Myeloproliferative neoplasms and myelodysplastic/myeloproliferative neoplasms; Malignant diseases: carcinoma, lymphoma, other solid tumors; Drugs: adrenaline, corticosteroids, lithium
Eosinopenia
Decreased eosinophils, <0.01 × 109/L. Causes: Drug induced: administration of corticotropin, corticosteroids, epinephrine or histamine; Acute inflammation or infection. No pathological affect
Eosinophilia
Increased eosinophils, >0.4 × 109/L. Primary stimulating cytokines: IL-5, IL-3 and GM-CSF. Causes: Infections: parasites, fungi; Allergic disorders; Löffler’s syndrome, tropical pulmonary eosinophilia, idiopathic hypereosinophilic syndrome; Leukemias, myeloproliferative neoplasms, myeloid and lymphoid neoplasms with abnormalities of PDGFRA, PDGFRB or FGFR1; Other malignant diseases: Mycosis fungoides, Sézary syndrome, Hodgkin’s disease, T-cell lymphomas metastatic carcinoma; Churg–Strauss syndrome, systemic sclerosis, rheumatoid arthritis
IL-5
an interleukin produced by T helper-2 cells and mast cells. Through binding to the IL-5 receptor, IL-5 stimulates B cell growth and increases immunoglobulin secretion. It is also a key mediator in eosinophil activation.
Basopenia
Decreased basophil count, s syndrome and pregnancy; Administration of progesterone, corticosteroids or corticotrophin
Basophilia
Increased basophil count, >0.2 × 109/L. Causes: Mastocytosis, CML, polycythemia vera, essential thrombocythemia, myelofibrosis, basophilic leukemia, eosinophilic leukemia, and Ph-positive acute leukemia
Monocytopenia
Decreased monocyte count, <0.2 × 109/L. Causes: aplastic anemia, Cyclic neutropenia, hemodialysis, severe thermal injuries, AIDS, hairy cell leukemia.
Monocytosis
Increased monocyte count, >1.0 × 109/L. May be a compensatory event in association with neutropenia. Causes: Physiologic; Infants; Certain infections; Marrow disease: MDS, MDS/MPN, acute immunoblastic and myelomonocytic leukemia; Hodgkin’s disease, carcinoma, multiple myeloma, malignant histiocytosis
Lymphopenia
Decreased lymphocyte count, s disease, aplastic anemia, agranulocytosis, and MDS
Lymphocytosis
Increased lymphocyte count, >5 × 109/L. Causes: Physiologic: infants and young children; Certain viral infections: EBV, CMC, infectious hepatitis, chickenpox, smallpox, measles, rubella, mumps, influenza, primary HIV infection; Pertussis, brucellosis, tuberculosis, secondary and congenital syphilis; Chronic lymphoproliferative disorders, lymphomas; Post-splenectomy
Thrombocytopenia
Decreased platelet count, < 140 × 109/L. Causes: Peripheral destruction: idiopathic thrombocytopenic purpura (ITP), thrombotic thrombocytopenic purpura (TTP), hemolytic-uremic syndrome (HUS), disseminated intravascular coagulation (DIC); Sequestration in spleen; Inadequate production: primary marrow disorders or metastatic tumor in the marrow; Inherited: Wiskott–Aldrich syndrome, May–Hegglin anomaly, Bernard–Soulier syndrome
Thrombocytosis
Increased platelet count, > 400 × 109/L . Causes: Primary marrow neoplasm: essential thrombocythemia, chronic myelogenous leukemia, polycythemia vera, myelofibrosis; Reactive: inflammation, surgery, hyposplenism, splenectomy, asplenia, iron deficiency anemia or hemorrhage
Bite cells
Bite-like detect due removal of Heinz body in spleen. Associated with G6PD deficiency
Schistocytes
Fragmented RBCs, helmet cells. HUS, TTP, DIC, burns, HELLP, mechanical heart valves
Target cells
Central hemoglobin, target shape. Thalassemia, hemoglobin C, iron deficiency, liver disease
Sickle cell
Bipolar spiculated shape. Banana shape. Sickle disease
Basophilic stippling
Morphology: evenly dispersed fine blue granules. Content: aggregated ribosomes (rRNA). Causes: Lead poisoning, porphyria, pyrimidine 5’ nucleotidase deficiency; Hemoglobinopathies, thalassemia; Myelodysplasia, sideroblastic anemia; Infection
Howell-Jolly bodies
Morphology: single, dense, blue dot. Content: nuclear DNA remnant. Causes: Post-splenectomy; Functional asplenia; Megaloblastic anemia; Myelodysplasia
Heinz Body
Not visible on Wright-Giemsa stain. Need to stain with supravital dye (crystal violet). Denatured/oxidized hemoglobin attached to the inner cell membrane. Cause: G6PD deficiency. Associated with bite cells
Dohle Body
Pale blue inclusion at the periphery of the cytoplasm. Infection, inflammation, burns or pregnancy. Contents: condensed RNA.
Toxic Granulation (Hypergranularity)
Increased Numbers and prominence of 10 granules. Due to rapid cell division (not enough time to dilute). Often associated with Döhle bodies and toxic vacuolization. Causes: bacterial infection, marrow recovery, G(M)-CSF
Hypersegmented Neutrophils
More than five lobes. Associated with megaloblastic anemia
hematocrit
also known as packed cell volume (PCV) or erythrocyte volume fraction (EVF), is the volume percentage (%) of red blood cells in blood. It is normally 45% for men and 40% for women.
reticuloyte
young RBC, the count is important for evaulating anemia.
Reticuloendothelial System
The mononuclear phagocyte system (MPS) (also called Reticuloendothelial System or Macrophage System) is a part of the immune system that consists of the phagocytic cells located in reticular connective tissue. The cells are primarily monocytes and macrophages, and they accumulate in lymph nodes and the spleen. The Kupffer cells of the liver and tissue histiocytes are also part of the MPS.
Hepcidin
25 amino acid peptide produced by hepatocyte. Antibacterial peptide. Synthesis is increased by inflammation/infection or iron overload. Negative regulator of iron absorption by the intestinal epithelial cells, transport by the placenta, and release from macrophages (RE system). Modified by a protease associated with turnover. Implications for hemochromatosis (deficiency of hepcidin). During inflammation/infection, inhibition of ferroportin causes increased iron retention in macrophages contributing to anemia. Implication for iron resistant iron deficiency anemia. Hepcidin critical for new approaches for hemochromatosis and anemia of inflammation/infection. This can look like anemia.
Characteristics of Iron Deficiency
Decreased Hgb synthesis. Decreased Cell proliferation (Hemolytic component). Multiple systems besides hematopoietic system involved: Hematopoietic: anemia (see previous slide); Neuromuscular: mild defect muscle performance, neuropsych dysfunction; Epithelial: nails (ridges, koilonychia); tongue (papillary atrophy); Upper GI: dysphagia, esophageal webs, achlorhydria, gastritis; Lower GI: protein losing enteropathy; Immune dysfunction: innate (phagocyte), adaptive (lymphocyte); Pica
Diagnosis of iron deficiency anemia
History and physical exam (epidemiologic and etiologic factors). Routine laboratory tests. Decreased O2 carrying capacity (Hgb, Hct). Decreased production (reticulocyte count and index). Microcytosis (Decreased MCV, smear findings). Hypochromia (Decreased MCHC, smear findings). Wide range in size of cell (increased RDW). Additional studies may be indicated in difficult cases: Decreased serum Fe, increased TIBC, Decreased ferritin, increased FEP
Differential diagnoses of iron defiency anemia
Anemia of chronic inflammation/infection; Anemia of chronic disease; Thalassemia; Sideroblastic anemias
Timeline of normalization of iron levels after supplement is given
Serum iron > Hgb/retic > ferritin > MCV/FEP/RDW
Iron Overload
Etiology: increased iron in diet, increased absorption (abnormality HLA-H gene), Repeat transfusions (chronic anemia). HLA-H gene encodes for a protein in duodenal crypt cells and reticuloendothelial cells which acts as a co-factor for iron absorption resulting in an increase in absorption and accumulation.
Consequences of Iron Overload
Concequences: increased serum iron (sat >50%), increased ferritin, increased liver iron. Organ damage: Cardiac (arrhythmia, failure); liver (dysfunction, failure): endocrine dysfunction (e.g., pancreatic endocrine: diabetes) Treatment: Hemochromatosis → therapeutic phlebotomy; Hemosiderosis → iron chelators: Desferal (IV or subcutaneous infusion) and Exjade (Newer oral preparation)
extramedullary hematopoiesis
Hematopoiesis outside of the bone marrow after birth and is distinctly abnormal
Myeloid
although the root “myel-“ just refers to marrow, the term myeloid most commonly refers to all non-erythroid and non-lymphoid lineages (i.e. the granulocytes, monocytes, megakaryocytes/platelets), though sometimes the term refers only to the granulocytic lineages.
Lymphoid
Refers to T cells, B cells, and NK cells, and their precursors
Bone marrow vasculature.
Nutrient arteries course into and through the bone marrow and branch into capillary-venous sinuses. Capillary-venous sinuses are composed of an endothelial cell layer, basement membrane, and an adventitial layer. Capillary-venous sinus blood eventually flow into a central vein, and from there into the systemic circulation. The passage of blood cells through the capillary-venous sinus endothelial layer is selective: Normally only mature cells are allowed to pass from the marrow into the sinuses and subsequently into the peripheral circulation; immature cells are retained.
stromal elements
The bone marrow environment plays a critical role in hematopoiesis. Stromal elements play important roles in hematopoiesis. These stromal elements include endothelial cells of the capillary-venous sinuses; reticular cells lining the adventitial surface of the capillary-venous sinuses; fibroblasts; lymphocytes; macrophages; adipocytes; and the extracellular matrix produced by stromal cells (e.g. collagen fibers, laminin, fibronectin, chondroitin sulfate, hyaluronic acid, heparan sulfate).
Erythropoietin (EPO)
made by certain kidney cells in response to hypoxia,
Thrombopoietin (TPO)
promotes megakaryopoiesis
Granulocyte-monocyte colony stimulating factor (GM-CSF)
promotes granulopoiesis and monopoiesis
Granulocyte colony stimulating factor (G-CSF)
promotes granulopoiesis
Monocyte colony stimulating factor (M-CSF)
promotes monopoiesis
Interleukin-5 (IL-5)
promotes production of eosinophils
Interleukin-3 (IL-3)
promotes production of basophils
Features characterizing granulopoiesis
1) Nuclear maturation with acquisition of condensed chromatin, loss of nucleoli and progressive nuclear indentation and, ultimately, segmentation. 2) Acquisition of cytoplasmic primary (azurophilic), and then secondary (specific) granules. 3) Loss of the ability to replicate by the metamyelocyte stage.
Eosinophils
The mature eosinophil is around 13 mm in diameter. The cytoplasm is full of large orange-red (eosinophilic) granules. The nucleus contains heavily condensed chromatin and is segmented, usually into two round to oval lobes.
Basophils
The mature basophil is 10 mm in diameter. It contains a lobular but non-segmented nucleus. The nucleus is usually obscure by numerous blue-purple (basophilic) granules.
Sideroblastic anemias
a rare group of congenital or acquired disorders. Vitamin B12 and folate deficiency are also included in underproduction anemias, but they are better characterized as a group of disorders with ineffective erythropoiesis. Sideroblastic anemias are a heterogeneous group of disorders with deposits of iron in mitochondria of erythroid precursors. The sequence below summarizes the pathophysiology. Impaired production of protoporphyrin or incorporation of iron in heme -> Accumulation of iron in mitochondria forming a ring around the nucleus-> Ring sideroblasts (Prussian blue).
causes of folate deficiency
In contrast to B12 deficiency, the most common cause of folate deficiency leading to megaloblastic anemia is inadequate dietary intake. Other causes include malabsorption due to such things as tropical sprue or parasitic infection, which can lead to rapid depletion of folate through interruption of enterohepatic circulation, inborn errors of folate metabolism (very rare), and increased demands (hemolysis, pregnancy/lactation, rapid growth, psoriasis, myeloproliferative disorders). Alcohol consumption also can lead to rapid onset of folate deficiency, not only through decreased dietary intake but also through disruption of cycling from liver stores to tissues.
Clinical and Laboratory Features of folic acid and vitamin B12 deficiency
Both folic acid and vitamin B12 deficiency result in megaloblastic anemia. The onset of folate deficiency can occur quite rapidly (within weeks), particularly in the setting of malabsorption or alcoholism. In someone who is well-nourished, Vitamin B12 deficiency takes several months to develop because of its long half-life within the body and large hepatic stores. Vitamin B12 deficiency develops more slowly and is more likely associated with malabsorption. The symptoms and signs of anemia in both cases are not distinguishable from other causes.
BONE MARROW STRUCTURE
Marrow space is encased by cortical bone, and interspersed by trabecular bone lined by osteoblasts and osteoclasts. Between trabecula is a network of vascular sinusoids with walls of ‘leaky’ endothelial cells. Marrow stromal cells are bound to the non-luminal side of the endothelial cells. These stromal cells: produce the protein framework of the marrow, especially type IV collagen (reticulin); produce regulatory factors and adhesion molecules needed to induce and maintain hematopoiesis. Marrow and blood are interconnected compartments
Erythropoiesis
Rate of erythropoiesis determines the hemoglobin level of normal individuals. Initiated by erythropoietin, a hormone produced by the kidneys. Erythropoietin production stimulated by hypoxia. Erythropoietin acts to: Activate stem cells of bone marrow to differentiate into pronormoblasts; Increases rate of mitosis and maturation process; Increases rate of hemoglobin production; Causes increased rate of reticulocyte release into peripheral blood.
Distinguishing types of granulopoiesis
Granulocyte types are distinguished from each other by the appearance of their secondary (specific) cytoplasmic granules: Neutrophils: pink to rose-violet granules. Eosinophils: reddish-orange granules Basophils: dark purple granules
AUER RODS
ARE SPECIFIC FOR MYELOBLASTS, BUT ARE ONLY SEEN IN ABNORMAL CONDITIONS
Neutrophil Granulocytes
Granules contain destructive enzymes, most famously myeloperoxidase, used to destroy infectious organisms, most commonly bacteria. Also have prominent phagocytic activity. Mature segmented neutrophil granulocytes are also known as PMNs, for ‘polymorphonuclear leukocytes’ (due to their highly and variably segmented nuclei), and also by the abbreviated terms ‘segs’ and ‘polys.’
Eosinophil Granulocytes
Similar maturation stages to neutrophils. Characterized by large, eosinophilic secondary granules. Mature eosinophils usually have 2 nuclear lobes. Lifespan of around 8-12 days. Granules contain destructive enzymes, which are generally used to fight organisms too big to phagocytose (fungi, protozoans, parasites). Also involved in modulation of mast cell activity in hypersensitivity response/allergic disease. Minimal degree of phagocytic activity. Main cytokine initiating eosinophil production: Interleukin-5 (IL-5)
Basophil Granulocytes / Mast Cells
These cell types are though to be related in some manner, though this is a point of contention. Both of these cell types are involved in hypersensitivity/allergic processes, and in innate defenses against microbes. Main cytokines initiating their production are: Interleukin-3 (IL-3) for basophils and Stem Cell Factor (SCF) for mast cells
MATURE BASOPHIL
Prominent large dark blue (basophilic) cytoplasmic granules, which obscure the nucleus. Multilobular but non-segmented nucleus. Found in blood and marrow at low levels
MAST CELLS
Many reddish-purple (metachromatic) cytoplasmic granules. Round nuclei, abundant cytoplasm. Found in marrow at low levels, and at varying levels in different solid tissues
Lymphopoiesis
T LINEAGE: Early T-lymphoid progenitor cells migrate to the thymus, the site of T cell maturation. Some mature T cells migrate back to reside in the marrow. B LINEAGE: B cell maturation takes place in the marrow. A few mature B cells and a fair population of plasma cells reside in the marrow
Normal B-Lymphoblasts (Hematogones)
Size: 10-18 μm in diameter. Nucleus: lacy and fine chromatin, but less fine than myeloblasts 1-2 nucleoli. Cytoplasm: scant, non-granular
Monopoiesis
Initiated by M-CSF (monocyte-colony stimulating factor). Time for monocyte maturation in marrow not well understood. Mature monocytes circulate in peripheral blood an average of 20 days, before entering tissue to become macrophages. Some mature monocytes and macrophages reside in the marrow
Investigation of Cytopenias
caused either by increased distruction or decreased production of marrow lineage. In cytopenias due to increased destruction, growth factors should signal to the marrow to make more of whatever is being destroyed….thus, examination of the marrow shows a compensatory marrow hyperplasia of one or more lineage. In cytopenias due to decreased production, the marrow will not show a compensatory hyperplasia; serum growth factor levels may be increased or decreased. If increased, they are not really getting a response.
PETECHIAE
microhemorrhages within skin (indicative of thrombocytopenia)
GIANT PLATELET
Not an entirely specific finding on blood smear, but usually indicates increased thrombopoiesis, in much the same manner as increased reticulocytes (large RBCs) usually indicates increased erythropoiesis
causes of neutrophilia
acute: infections (bacterial), burns, infarcts, drugs (steriods), stress (pain, extreme temp). Chronic: persistent infections, persistent inflammatory disease (IBD), continued exposure to neutrophilia inducing drugs, tumors producing growth factors, clonal proliferation of neutrophils (CML)
possible causes microcytic and hypochromic anemia
iron deficient anemia, anemia of chronic disease, colon cancer, hemoglobinopathy (thalassemia), primary marrow disease
Specific Testing for folate and vitamin B12 deficiency
Direct measurement of serum cobalamin levels and serum or red cell folate levels is useful in diagnosing deficiencies, although there can be problems with these tests. Cobalamin deficiency in the tissues can exist with a normal serum cobalamin level. Serum folate levels may reflect recent intake and not tissue stores, while red cell folate is a better indicator of tissue folate status but will be low with B12 deficiency as well. Measurement of plasma homocysteine levels has been used as a more sensitive marker of deficiency of B12 and folate in the tissues. As indicated in the metabolic pathways described above, vitamin B12 and folic acid are both required for the synthesis of methionine from homocysteine. So, deficiency of either one of these vitamins should lead to elevated homocysteine levels. Another reaction involving vitamin B12 but not folate is the synthesis of succinyl CoA from methylmalonyl CoA. Thus, in B12 but not folate deficiency, methylmalonic acid levels are increased, making measurement of methylmalonic acid a good way to distinguish the two.
The Schilling Test
Once B12 deficiency is diagnosed, it is important to know the cause. Measurement of serum autoantibodies against intrinsic factor, the cobalamin-intrinsic factor complex, and parietal cells is now commonly used to diagnose pernicious anemia, with positivity in more than 60% of adults with the disease. The Schilling test is an older test used to diagnose defects in B12 absorption that is less commonly used today but still important to know about. In this test, 1 μg of radiolabeled cobalamin is given orally to a fasting individual. The IF produced in the stomach combines with the radiolabeled cobalamin (Cbl) which is absorbed in the terminal ileum. The tagged cobalamin is then bound to transcobalamin II (TcII) and enters the bloodstream. A dose of cold (unlabeled) cobalamin is given intramuscularly 2 hours later, causing some of the labeled cobalamin to be excreted in the urine over the following 24 hours (5-35%). If the patient isn’t absorbing the cobalamin given orally, less radiolabeled cobalamin will be excreted into the urine. If the test is positive, it is then repeated with the patient receiving intrinsic factor in addition to the cobalamin, with correction establishing the diagnosis of pernicious anemia. A variation on this test is the food Schilling test where the cobalamin is incorporated with food, allowing the diagnosis of other malabsorptive problems.
Use of Transfusion and EPO in Chronic Anemia
Only transfuse red cells when the severity of anemia has potential for cardiovascular decompensation. Use EPO to treat anemias for which there is (1) an absolute deficiency (renal disease) or (2) a decrease of this cytokine out of proportion to the hematocrit level and for which a response has been documented. EPO is linked with strokes.
Clinical findings of sideroblastic anemia
Variable anemia; hypochromic, microcytic RBCs; Pappenheimer bodies (precipitated iron in mitochondria), particularly in marrow normoblasts; accumulation of stored iron. Some syndromes are congenital without specific inheritance; others show an X-linked or autosomal recessive pattern. Sporadic cases and secondary sideroblastic anemias (alcoholism, copper deficiency, drug related) are described. The treatment includes vitamin B6 (some respond), supportive care (including transfusions) or treatment of a reversible secondary cause. Iron overload is treated with chelation therapy (desferal or newer oral chelators).
low affinity hemoglobin disease
Hemoglobin mutations with low oxygen affinity are associated with a right-shifted oxyhemoglobin dissociation curve, decreased oxygen affinity, normal tissue oxygen, and mild anemia because of improved oxygen delivery (in some) and/or hemolysis (depending on the specific mutation).
protein/ calorie malnutrition
Protein/calorie malnutrition, to low to support erthropoeisis, is associated with a normochromic, normocytic anemia. There may be associated vitamin and mineral deficiencies which may also play a role in the anemia.
methyltetrahydrofolate
methyl donor critical for synthesis of methionine from homocysteine
tetrahydrofolate
a subratrate for purine an pyrimidine synthesis
entrohepatic circulation (EHC)
Substances are said to undergo an enterohepatic circulation (EHC) when they are excreted into the bile, pass into the lumen of the intestine, are reabsorbed and return to the liver via the circulation
Hemoglobin A1 (a2b2)
also called Hemoglobin A - is the predominant form of hemoglobin in the adult. Because there is no substitute for the α-chain beyond the early gestational period, a total lack of α-globin chains is incompatible with life.
Fetal Hemoglobin
(Hb F, a2g2) is the predominant form in the fetus and newborn. Hemoglobin A2 (α2δ2) makes up a small amount of the total hemoglobin in the adult. Fetal hemoglobin remains elevated in premature babies and infants of mothers with diabetes. It also remains elevated in people with hemolytic anemias and with certain diseases affecting the bone marrow such as myelodysplasia and leukemia. Hemoglobin A2 (α2δ2) comprises about 2% of normal adult hemoglobin. It is evenly distributed in red cells and functions much like hemoglobin A. It has the same Bohr effect, cooperativity and response to 2,3-BPG but is more heat stable and has slightly higher oxygen affinity. As you will learn later in the course, hemoglobin A2 can be elevated in certain clinical situations, such as β-thalassemia, sickle cell trait and disease, hyperthyroidism, and megaloblastic anemias.
oxygen dissociation curve for hemoglobin
Because of cooperativity, when the % saturation of hemoglobin by oxygen is plotted as a function of the partial pressure of oxygen, the resulting curve turns out to be sigmoidal, or S-shaped. hemoglobin is an excellent protein to use for oxygen transport, since oxygen is easily loaded onto the molecule in the lung where the partial pressure of oxygen is ~100 mmHg but then readily unloads in the tissues where the partial pressure of oxygen is ~40 mmHg.
myoglobin dissociation curve
a protein which stores oxygen in muscle cells and is very similar to hemoglobin except that it is a monomer rather than a tetramer and therefore does not undergo allosteric regulation or cooperativity. The myoglobin curve is shaped more like a hyperbola, giving the myoglobin molecule very high oxygen affinity at very low oxygen concentrations. Functionally, myoglobin is a very poor protein to use to transport oxygen from the lungs to the tissues, since it would hold tightly to the oxygen and not release it until the oxygen concentration got very low. On the other hand, myoglobin is a very good protein to use for storage of oxygen in the intracellular environment where oxygen concentration is very low (1-5 mmHg) and where high oxygen affinity is needed to transfer the oxygen from hemoglobin to myoglobin.
P50
A way to quantify this difference in oxygen affinity is by determining the P50, which is defined as the partial pressure of oxygen at which the oxygen carrying protein is 50% saturated. Under normal conditions for temperature (37 C) and pH (7.4), the P50 for hemoglobin is approximately 27 mmHg while the P50 for myoglobin is 2.75 mmHg.
Fetal hemoglobin
Chromosome 16 contains the “a-like” genes, including two copies of the a-globin gene itself along with variants expressed early in embryonic development; therefore, the genome contains a total of 4 copies of the a-globin gene (2 paternal and 2 maternal). The “b-like” genes (genes for the g-, d-, and b-globin chains along with variants produced early in embryonic development) are products of a set of genes on chromosome 11; one copy of the gene set is inherited from each parent.
fetal hemoglobin patterns
Embryos have 3 distinct hemoglobins that are present only between 4 and 14 weeks gestation: Hemoglobin Gower I (z2e2), Hemoglobin Gower II (a2e2) and Hemoglobin Portland (z2g2). Each of these has a higher affinity for oxygen than does hemoglobin A. After week 8 of gestation, fetal hemoglobin or hemoglobin F (a2g2) predominates. The g-chain differs from the b-globin chain by 39 amino acids. Fetal red cells have a higher oxygen affinity than adult red cells, primarily because hemoglobin F binds 2,3-BPG poorly, stabilizing the hemoglobin in the R state and shifting the oxygen dissociation curve to the left. The Bohr effect is also increased by 20% in fetal hemoglobin, so that as fetal blood passes through the intravillous spaces of the placenta, H+ ions are transferred to the maternal circulation and the pH rises, leading to increased oxygen affinity and a further shift of the curve to the left. These changes favor transfer of oxygen from the maternal circulation to the fetal circulation
Adult hemoglobin patterns
At birth, there is 65-95% hemoglobin F and about 20% hemoglobin A. The normal adult level of fetal hemoglobin is approached by 1 year and achieved by 5 years of age. Under normal conditions, adults have 96-97% of their hemoglobin as hemoglobin A (a2b2). In adults, hemoglobin F makes up <1% of the total hemoglobin and is unevenly distributed in red cells.
Cyanosis
is visually perceptible when reduced hemoglobin exceeds 3 g/dL, which generally corresponds to an oxygen saturation level below 85 percent in a patient with a hemoglobin concentration of 15 g/dL. Clinically, it would be very useful to know a patient’s oxygenation status before it reaches this critical level. Traditionally, arterial blood gas analysis was performed to measure oxygen status. Pulse oximetry, a noninvasive alternative, is the norm now.
A pulse oximeter
probe is a photo detector and two light-emitting diodes, one emitting at 660 nm and the other at 940 nm. They are in the red band (660 nm) where deoxyhemoglobin absorbs light maximally and the 940 nm infrared wavelength where oxyhemoglobin absorbs most. The detector and emitter face each other through tissue, so the probe is usually placed on the finger. Only pulsatile flow, representing arterial blood flow, is measured. The photodiodes switch on and off several hundred times/second and light absorption is measured. The displayed value is based on an average of the previous 3-6 seconds.
extravascular hemolysis
the red cell is ingested by macrophages of the RE system. The heme is separated from globin, iron removed and stored in ferritin, and the porphyrin ring converted to bilirubin which is released from the cell. Taken up by a specific transport system in the liver, the bilirubin (lipid soluble) is converted to a water soluble compound by addition (conjugation) of a glucuronic acid. This is completed by the cytochrome P-450 enzyme(s) in liver parenchymal cells. After secretion into the biliary tract and small bowel, the glucuronic acid is removed and bilirubin converted into urobilinogen and other water soluble pigments. Urobilinogen may cycle between the gut and liver (entero-hepatic circulation) or excreted by the kidney into the urine.
Unstable hemoglobins
Tendency to spontaneously denature, >60 variants. Often due to mutations that disrupt the stability of the heme-globin linkage. May not be detected until adulthood. Can also have altered oxygen affinity (higher or lower). May lead to a hemolytic anemia, with jaundice and splenomegaly. Can be referred to as Heinz body anemia. Usually don’t need blood transfusions. Give folic acid regularly. Splenectomy not curative. May not see Heinz bodies until after splenectomy
CYANOSIS
Too much deoxyhemoglobin. >3 g/dL (~1.5 g/dl methemoglobin (8-12% assuming normal hemoglobin level). Sulfhemoglobinemia >0.5 g/dl sulfhemoglobin
Differential Diagnosis of Cyanosis
Inadequate Oxygenation of Hemoglobin (common): Pulmonary disorders, Cardiac right-to-left shunt, Congestive heart failure, Cardiovascular collapse (shock). Low O2 affinity Hb variant (rare). Methemoglobinemia (rare): Congenital-Cytochrome-b5 reductase deficiency, Cytochrome-b5 deficiency, M hemoglobins. Acquired- Drugs, Industrial environmental toxins, etc. Sulfhemoglobinemia (rare): Acquired: drugs, toxins, etc.
Hereditary spherocytosis
Hereditary spherocytosis (HS) is a familial disorder characterized by anemia, intermittent jaundice, splenomegaly and responsiveness to splenectomy. The heterogeneity of clinical features is associated with multiple molecular abnormalities of which spectrin deficiency is the most common. The hallmark of this syndrome is loss of plasma membrane and formation of the microspherocyte. Spherocytes are more susceptible in vitro to osmotic stress, the basis for a common test for the disorder. The basic pathophysiology is that spectrin, ankyrin or band 3 defects weaken the cytoskeleton and destabilize the lipid bilayer. Loss of membrane and formation of the spherocyte leads to decreased deformability and entrapment in the spleen. Conditioning in the red pulp leads to further loss of red cell membrane and, ultimately, removal by the macrophage. Most inherit the condition as an autosomal dominant; 25% are autosomal recessive.
Clinical presentation and treatment of hereditary spherocytosis
Clinically, patients present with a variable degree of anemia as well as jaundice and splenomegaly. One third has hyperbilirubinemia as neonates. Treatment includes supportive care for chronic anemia and intermittent complications and splenectomy which usually resolves the clinical manifestations. Incidence 1/5,000. Anemia (severe 5%, moderate 60-75%, mild 20%), jaundice, splenomegaly. Variable onset: neonatal hyperbilirubinemia (1/3), childhood or adulthood (as one of two major complications). 75% have autosomal dominant pattern, 25% autosomal recessive. Presenting complications: hyperhemolysis, aplastic crisis. Supportive care (including supplementation with folate). Splenectomy usually resolves clinical manifestations
Clinical features of G6PD
this disorder presents as intermittent episodes of acute hemolytic anemia and hyperbilirubinemia associated with oxidant stress (infection, drugs or ingestion of specific foods including fava beans). Rarely it may be characterized as chronic hemolytic anemia punctuated by episodes of acute exacerbation of anemia. This condition may also be a cause for neonatal hyperbilirubinemia. No specific morphologic features are associated with G-6-PD deficiency (originally categorized as congenital non-spherocytic hemolytic anemia) but occasionally the smear will show microspherocytes and “blister” or “bite” cells. Management consists of avoiding oxidant drugs and foods for the most common variants seen in the U.S. For severe cases with chronic anemia, supportive care and folate are included.
neonatal hyperbilirubinemia
Neonatal jaundice can sometimes make the newborn sleepy, and interfere with feeding. Extreme jaundice can cause permanent neurological damage known as kernicterus. Kernicterus is a bilirubin-induced brain dysfunction. Bilirubin is a highly neurotoxic substance that may become elevated in the serum.
aplastic crisis
can occur in anyone with hemolysis. Happens when hemopoeisis drops down occasionaly. Often associated with sickle cell disease. RBC survival is much shorter therefore cannot compensate.
Function of Spleen
Clearance of intravascular particles. Adaptive immune response: origin of IgM agglutinins, especially for encapsulated organisms.
Management of Splenectomy
Problem with splenectomy: overwhelming sepsis (particularly to S. pneumoniae). Risk highest in children 38.5°C (lifelong).
Chronic RBC Adhesion/Vascular Occlusion with sickle cell
Even when not sickled, the red blood cell in sickle cell disease is “sticky” due to membrane injury and retention of adhesion molecules on the its surface, which results in adhesion of sickle RBCs in the microvascular circulation. This adhesion results in transient vaso-occlusion (partial or total blockage of blood flow through the vessel), vessel wall injury and endothelial remodeling, resulting in narrowing of the vessels and chronic organ damage due to slow or absent blood flow through this microcirculation. The most severely affected organs include the spleen, central nervous system vasculature, lung, kidney, retina, and others.
Spleen with sickle cell
The microcirculation of the spleen is especially susceptible to occlusion and injury by sickle RBCs. When large numbers of sickle RBCs become abruptly trapped in microcirculation of the spleen, severe anemia and circulatory shock can occur, a complication called splenic sequestration. Even if overt evidence of sequestration isn’t seen, nearly all patients with sickle cell anemia (HbSS) chronically occlude the spleen’s microcirculation, resulting in “autoinfarction” (destruction) of the spleen by the age of 5. This process begins during the first year of life, and slowly compromises the spleen’s ability to kill encapsulated organisms (e.g. pneumococcus, meningococus, hemophilis). Sepsis (overwhelming blood infection) with these organisms, a common cause of death for infants and young children with sickle cell disease, is significantly reduced by the use of prophylactic penicillin and prompt treatment of fever with additional antibiotic therapy.
Central Nervous System (CNS) Vasculature with sickle cell
The large blood vessels of the central nervous system can be significantly damaged by sickle RBCs. Up to 10% of children with sickle cell anemia (Hb SS) experience an overt large vessel stroke due to this chronic injury, and a larger percentage experience learning disabilities and more subtle neurologic problems. An increase in the velocity of blood flow through the middle cerebral artery, as detected by transcranial Doppler, can predict for an increased risk of stroke in children. This risk can be reduced by prophylactic blood transfusions. Adults with sickle cell disease are more like to have hemorrhages from progressive weakening and rupture of these vessels.
Lung with sickle cell
The microcirculation of the lung is vulnerable to damage from sickle RBCs. Damage to these vessels makes it harder for blood to flow through the lungs, resulting in an increased pressure in the pulmonary arteries, a condition called pulmonary arterial hypertension (PAH). PAH, which puts strain on the right side of the heart (cor pulmonale), may affect 30-40% of patients with sickle cell disease and is now one of the most common chronic causes of death in adults with sickle cell disease.
Kidney with sickle cell
The tubules of the kidney are damaged by chronic vaso-occlusion, resulting in the inability to concentrate the urine to avoid dehydration. Hematuria (blood in the urine) may occur due to ischemia to the collecting system (papillary necrosis), which may also cause severe flank pain. The glomerulus may be affected, most commonly initially manifested by an enlargement of the glomerulus and protein in the urine. Ultimately, up to 10% of adult sickle cell patients will develop renal insufficiency due to permanent damage and scarring of the glomerulus (focal segmental glomerular sclerosis), and some with require dialysis and/or renal transplantation.
Retina with sickle cell
The retinal vessels are susceptible to chronic injury, resulting in the propensity for abnormal vessel formation and hemorrhage, which can lead to retinal detachment and blindness.
Other Areas affected with sickle cell
The femoral and humeral heads may develop avascular necrosis, a source of chronic pain and progressive joint deterioration, leading to hip and shoulder replacement. Skin ulcers, likely due to microvascular ischemia and poor healing, most often form around the ankles.
Acute RBC Adhesion/Occlusion – Sickle Cell “Crisis”
In the setting of hypoxia, dehydration, inflammation, infection, or other stresses, not only are RBCs more likely to sickle, but blood vessels can become acutely damaged and constricted, which may promote significant, sudden vaso-occlusion. This results in a “pain crisis”, in which acute severe pain develops relatively rapidly in a pattern that is unique to each patient and may involve any part of the body, most commonly in the arms, legs, chest or abdomen. The pain is likely due to many factors, including reversible ischemia, and resolves as the inciting factors (e.g. hypoxia, dehydration) are improved. During some pain crises, acute severe vaso-occlusion may occur in critical organs, causing acute end-organ injury. One example is the splenic sequestration described above. Other significant acute vaso-occlusive complications include: hand foot syndrome, Acute Chest Syndrome, Acute Multi-Organ Failure Syndrome, Priapism, Bone Infaction
Hand-Foot Syndrome
Usually seen in infants with sickle cell anemia and Sβothalassemia, self-limited acute severe swelling of the hands and feet can occur, and may be one of the earliest manifestations of sickle cell disease.
Acute Chest Syndrome.
Sickle RBCs can become trapped in the lung circulation, which damages the vessel lining (endothelium), promoting fluid to leak into the lungs, compromising the ability to oxygenate the blood. The patient may develop chest pain, fever, low oxygen saturation, and experience a fall in hemoglobin due to the trapping of RBCs in the lung. This is one of the most common acute causes of death in sickle cell disease, and may be triggered by pneumonia or embolization of fat from the bone marrow.
Priapism
Sickle RBCs can be trapped in the penis, with obstruction of outflow, resulting in sustained painful erections. Unless treated, these can result in impotence.
Bone Infaction
Focal areas of bone may sustain enough ischemia as to become permanently damaged, or necrotic. These areas can remain painful for extended periods of time and may become a source for chronic pain or a site of osteomyelitis (bone infection).
PROGNOSIS of sickle cell
Despite the multiple potential health impacts of sickle cell disease, most people with even the most severe forms of the disease should expect to live into their 50s and 60s, provided they receive comprehensive medical care and appropriate interventions for acute and chronic complications.
Beta globin gene cluster
epsilon, gamma (for hbF), delta (In low capacity), and beta (for HbA)
alpha globin gene cluster
chromosome 26, zeta and alpha (for HbF and HbA)
Polychromasia
Polychromasia (also known as Polychromatophilia) is a disorder where there is an abnormally high number of red blood cells found in the bloodstream as a result of being prematurely released from the bone marrow during blood formation. These cells are often shades of grayish blue. Polychromasia is usually a sign of bone marrow stress as well as immature red blood cells. 3 types are recognized, with types (1) and (2) being referred to as ‘young red blood cells’ and type (3) as ‘old red blood cells’.
HbF
Some of the excess chains form relatively unstable tetramer hemoglobin molecules. The main prenatal hemoglobin is Hb F (α2γ2), so in the fetus/newborn with α-thalassemia, there is an excess of γ-globin chains which forms Hb Barts, made up of 4 γ-chains (γ4). This hemoglobin can be identified by hemoglobin electrophoresis.
HbH
In the first six months of life there is a shift to the production of hemoglobin A1 (α2β2). As this occurs in a person with α-thalassemia, there is an accumulation of β-globin chains, some of which form hemoglobin H, a relatively unstable tetramer of β chains (β4). Hb H can be detected on fresh blood samples using special hemoglobin separation techniques.
Chronic Hemolytic Anemia with thalassemia
The fragile RBC has a very short half-life and is destroyed in the marrow or culled by the spleen from the circulation. This results in characteristic changes: Anemia with some increase in reticulocyte count- The degree of anemia and reticulocytosis will vary with the severity of the thalassemia. For patients with Cooley’s anemia (βothalassemia), severe anemia with hemoglobin values of <7 g/dL develops within the first year of life, and regular transfusions of RBCs are necessary to sustain life beyond the first 2-3 years. In milder forms of thalassemia, transfusion support may become necessary later in life. Abnormal peripheral smear- with microcytosis (small RBCs), target cells, polychromasia (blue-colored cells representing reticulocytes), mild anisocytosis (variation in size of RBCs), though the cells generally appear relatively homogenous without a large variation in size and shape. Thus, the RBC distribution of width (RDW) is often normal or minimally elevated in thalassemia traits. Abnormal chemistry profile- with increased total/indirect bilirubin, lactate dehydrogenase (LDH) and aspartate aminotransferase (AST) as they are released from the lysed RBCs. Splenomegaly-The spleen chronically removes fragile, damaged RBCs from the circulation, and may be enlarged even in persons with thalassemia traits and Hb EE disease. In some cases, the spleen traps increasing numbers of RBCs, resulting in more anemia (fewer circulating RBCs) and an increase in the volume of transfusions needed (hypersplenism), and must be removed.
Expanded Bone Marrow and Extramedullary Hematopoiesis with thalassemia
In an effort to produce an adequate RBC mass, the bone marrow expands and fills with RBC precursors even though they fragile and ultimately destroyed (ineffective erythropoiesis). This may result in characteristic skeletal deformities, including frontal bossing, although gross changes are generally prevented by early transfusion therapy. Osteopenia often develops despite transfusion support, and in some patients is very painful. The liver and spleen may also enlarge as they become sites for erythropoiesis (extramedullary hematopoiesis).
Increased Iron Absorption with thalassemia
Persons with severe forms of thalassemia increase absorption of iron from the diet in response to anemia. These are the same patients who require RBC transfusions, which also add to the iron burden and pathologic accumulation of iron in the heart, liver, and pituitary. Unless iron chelation therapy is used, there is a significant risk of mortality due to heart failure during late childhood or young adulthood.
Delayed Growth and Development with thalassemia
Anemia, increased metabolism associated with ineffective erythropoiesis, and endocrinopathies likely contribute to delayed growth and development, including short stature and delayed-onset of puberty. This can be improved with effective transfusion and iron chelation therapy.
Endocrinopathies with thalassemia
Nearly 2/3 of patients with Cooley’s anemia (β-thalassemia major) have abnormal endocrine function. The pituitary gland is commonly affected, and may lead to hypogonadotrophic hypogonadism (inadequate gonadal/reproductive function). Hypothyroidism (primary or due to pituitary dysfunction) and/or impaired glucose tolerance may occur in 40-60% of patients with β-thalassemia major.
Pulmonary Hypertension with thalassemia
Persons with chronic hemolytic anemia, including thalassemia, appear to be at risk for the development of pulmonary hypertension. This is more common in those who have had a splenectomy.
Hb C
(b6 Glu ->Lys).structural variant with abnormal function.
Hb E
(b26 Glu ->Lys). Structural variant with decreased synthesis.
Normal Adult Hemoglobins
Hb F a2 g 2 (95.0%), Hb A2 a2d2 (<3.5%)
ANTIBODY SPECIFICITY
This can be thought of in terms of the “goodness of fit” (affinity) between an antigenic determinant and a B cell receptor or free antibody. The better the fit, the more that cell or antibody seems to be specific for the determinant. It can be amazingly good: antibodies with association constants (Ka) in the range of 1015 liters/mole have been described. This enormous affinity/specificity makes antibodies not only great protection but unique tools. Such minor differences in antigens can be distinguished that antibodies are essential in hundreds of clinical and research assays, for example, measuring one steroid hormone in the blood in the presence of many others; in drug assays; for Western Blots, and screening libraries of genes in expression vectors; in diagnostic kits, including pregnancy tests; and hundreds of other applications.
steps to activate a B cell to produce antibodies
First, binding of antigen to the B cell’s receptors (membrane-bound versions of the antibody it will eventually secrete) occurs with a particular Ka. If this binding is strong enough, the second step, activation of the B cell, can take place. So an antigen which binds with low affinity may never activate the cell; but if another antigen comes along which not only binds but activates, the product of the cell (the secreted antibody) may combine with the low affinity antigen well enough to be inconvenient.
rheumatic heart disease
The heart valves contain an antigen, laminin, which cross-reacts with Group A streptococci. Obviously, the antigen in the valves does not normally activate the corresponding B cells, or we’d all have an autoimmune disease. When people get a streptococcal infection, the streptococcal antigens do activate these B cells because they bind to them with sufficient affinity. Then the released antibody can react with heart valves; with low affinity, it is true, but occasionally, in some people, with enough affinity to lead to a destructive, complement- mediated, inflammatory process
original theory of antibody design
The original theories were of the instructive type, that is, they said that the antigen told the immune system in some way to make an antibody of appropriate conformation, the way a potter’s mold informs the pot. These theories were Lamarckian; they implied that the outside world could instruct a cell to change its genetic information in some specific way so that a new protein was made.
ANTIBODY GENETICS
The combining site (into which the epitope fits) is made up of the V (variable from one antibody to another) domains of H (heavy) and L (light) chains. If H and L chains are under separate genetic control and any two can associate randomly, then with 1000 L chains and 1000 H chains we can make 1000 x 1000= 1,000,000 antibody combining sites; we could make a million antibodies with 2000 genes.
Receptor editing
Although we just said that a B cell tries to rearrange each allele just once, that isn’t strictly true. In some cells, when a rearrangement is detected as faulty (say a stop codon is generated), or if an anti-self receptor has been displayed, as long as the recombinases (RAG genes) are still active it can “try again.” The process is called receptor editing.
Storage Issues of blood
Each component of blood has a different storage condition which is determined to provide optimum survival (recovery and turnover) and normal function. At day of outdate (the last day the product can be used), recovery of the specific blood component is at least 70% of what would be expected from a freshly collected product, and turnover of the transfused cells in the recipient approximates what should be normal for that blood constituent. For red cells, 2,3-DPG and O2 delivery are adequate for up to 10 days of storage. For products stored longer, 2,3-DPG and O2 dissociation curve for red cells are abnormal, but will return to normal within 12 hours after transfusion. Platelet concentrates infused into patients will achieve maximal peripheral counts and hemostasis 1 hour after the infusion, and platelet turnover approximates a normal rate.
Production and Kinetics of Neutrophils
The production and kinetics of the neutrophil are useful to understand the innate immune response and to provide a conceptual scheme for organizing neutropenic disorders. Under non-stressed conditions, it takes 10 days to two weeks to move from pluripotent stem cells to mature neutrophils. Under stress, this can be shortened to 5 days. Earliest recognized myeloid cells (myeloblast promyelocyte and myelocyte) form the mitotic pool. These cells both divide and mature (by developing azurophilic and specific granules and other cellular constituents). The storage compartment contains metamyelocytes, bands, and segmented neutrophils (segs) which no longer have the capacity to divide but complete the process of maturing granules and their contents as well as cytosolic proteins to provide the mature cells with complete functional activity. The storage pool, made up of mostly bands and segmented neutrophils, provides a reserve of first responding phagocytic cells, easily mobilized into peripheral blood to fight infection. Once released into the circulation, these cells move to tissues to dispatch microbial colonizers and invaders or die (apoptosis) trying.
Myelopoiesis
Regulation of production is provided by the cytokines noted previously to develop progenitors and precursors in combination with cytokines for maturation of specific cell lines (G-CSF, M-CSF, IL-5). Production of these cells can be enhanced during infection, stress, and trauma which release microbial products, cytokines, interleukins and other biologic response modifiers to induce endothelial cells, epithelial cells and lymphocytes to produce SCF, IL-11, IL-3, and other cytokines, causing proliferation of pluripotent stem cells and expansion on maturation of the myeloid/monocyte cell lines.
Innate Immunity Response
Infection or tissue damage results in activation of Toll-like receptors (TLRs) and release of inflammatory mediators including lipids (PAF, LTB4), cytokines and interleukins (IL-1, TNF, IL-6, γ-INF), chemokines (IL-8, GROγ, NAP-2, MCP-1, RANTES), complement, kinins, and coagulation factors which cause vascular dilatation, permeability and emigration of leukocytes into a focus. Movement of neutrophils and monocytes into the area result in release of innate immune responses and inflammation and subsequent emigration of lymphocytes to interact with monocytes to initiate an adaptive immune response.
Biochemistry and cell biology of neutrophil function
Recent studies in patients with recurrent infections and congenital neutrophil dysfunction syndromes have identified the nature of the defects and expanded our understanding of specific aspects of phagocyte function. Included in these are adhesion molecules, β2 integrins (CD11b/18) and selectins, the oxidase enzyme system, and granules and their constituents. Selectins are a family of proteins with a lectin binding domain, an EGF domain, a variable number of short consensus repeats, a transmembrane domain and a cytoplasmic domain. The lectin site binds sialyted, fucosylated carbohydrates. L-selectin and sialy-LeX on neutrophils interacts to induce rolling, the first adhesion function.
β2 integrins
a family of glycosylated heterodimers. The α chain has a cytoplasmic domain, a single membrane spanning segment, and an extracellular portion with an I domain and divalent cation binding sites. The β chain has similar intracellular and membrane domains and a different extracellular portion with an I domain and cysteine-rich repeats. Various combinations produce CR3 and C3bi (CD11b/CD18 and CD11c/CD18, respectively) on the surface of the cell which function as receptors for adherence, chemotaxis, and ingestion.
The oxidase enzyme system
is composed of 6 or more proteins which are distributed in the plasma membrane or specific granule membrane (gp91phox, p22phox) or in the cytosol (p47phox, p67phox, p40phox, and Rac2). With a phagocytic stimulus, assembly of the cytosolic components with the membrane components assembles the system and results in activity with addition of an electron to oxygen to form superoxide anion from which H2O2 and the other reactive oxygen species (ROS) can rapidly be formed.
granule protein classes
Granule proteins and other constituents are contained in the two main granule classes (azurophilic and specific granules) as well as tertiary granules and secretory vesicles. The contents of these organelles contain constituents which aid in the disruption and dissolution of microbes. Stores of receptors of various classes as well as proinflammatory compounds support continued phagocyte function and inflammation. Congenital neutrophil defects may be categorized according to main functional characteristics of the cells.
COMPLEMENT DEVELOPMENT
Newborn C levels are usually around those of adults; preemies are often low. Complement components are mostly made in the liver, though white blood cells also contribute.
the two compartments of the thymus
The lymphocytes (“thymocytes”) whose precursors came in from the marrow, and the supporting structure or stroma, which develops in the neck region and moves down into the fetal chest.
Granulocyte-colony stimulating factor (G-CSF or GCSF)
a glycoprotein that stimulates the bone marrow to produce granulocytes and stem cells and release them into the bloodstream. Functionally, it is a cytokine and hormone, a type of colony-stimulating factor, and is produced by a number of different tissues. The pharmaceutical analogs of naturally occurring G-CSF are called filgrastim and lenograstim. G-CSF also stimulates the survival, proliferation, differentiation, and function of neutrophil precursors and mature neutrophils.
high grade pathogens
live out side of cells (eg staph, strep). Affectively delt with with antibodies.
low grade pathogens
live within the cells. T cell system can deal with this.
Cytokines
Short-range mediators made by any cell, that affect the behavior of the same or another cell. IL-1, TNFα, IL-12
Lymphokines
Short-range mediators made by lymphocytes, which affect the behavior of the same or another cell. A subset of cytokines. IL-2, IFNγ, IL-4, IL-5, IL-10
Chemokines
Small (6-14 kD) short-range mediators made by any cell, that primarily cause inflammation. MIP-1 to -4, RANTES, CCL28, CXCL16, Eotaxin, IL-8
ONTOGENY AND MATURATION OF T CELLS
T cells originate in the thymus, coming out as CTL or Th0 (there is a subtype of Treg that develop in the thymus, too). The thymus consists of epithelial cells, most of which arise from the III and IV pharyngeal pouches in fetal life; macrophages, derived from the bone marrow; and thymocytes (developing thymic lymphocytes), also bone-marrow derived. There are also, of course, supporting cells, fibroblasts, blood vessels, even nerves. There is a dense cortex and a somewhat looser medulla.T cell precursor cells arrive from the bone marrow via the blood, and land in the outer thymic cortex. There they begin to divide rapidly, and can be distinguished from other cells by their large size. At this stage they are ‘double-negative,’ that is, CD4-/CD8-, and have activated Rag-1 and Rag-2 DNA recombinases so they are beginning to rearrange their TCR variable domain genes. These cells will eventually give rise to the mature phenotype ‘single-positives,’ CD8+/CD4- and CD4+/CD8-. The first step is to become double-positive (going from CD4-/CD8- to CD4+/CD8+), and then during selection to turn off one or the other gene. If so, this suggests why the bulk of the cells in the thymus are, in fact, double positive; having failed to be selected for further maturation, they remain ‘stuck’ at the double positive stage until they die. Single-positive T cells acquire other phenotypic refinements as they mature in the thymus, such as recirculation specification molecules and the various molecules with which they interact with APC. Then they are exported from the medulla. ►Fewer than 2% of thymocytes are exported; the rest will die in the thymus. Why? Because the demands on the T cell repertoire are very strict and not many randomly-generated TCR fill the bill.
What are the specifications for a successful T cell?
A T cell must: Not recognize “self”, that is, not bind so firmly to a self structure (MHC alone, or MHC loaded with a “self” peptide) that the T cell becomes activated; this would be autoimmunity. Not recognize free antigen (which is antibody’s job). Recognize antigenic peptide plus self MHC.The repertoire is generated and then selected within the thymus. Imagine a thymocyte that has just rearranged the genes for the alpha (V,J) and beta (V,D,J) chains of its TCR. (Note, these V(D)J are a different set from the ones B cells use.) It puts the receptors on its surface, and begins percolating through the thymus cortex, during which it will brush against the surfaces of thousands of macrophages and epithelial cells.Let us say that the CDRs of TCR a and b genes have been selected during evolution to produce receptors that are roughly complementary to the average shape of an MHC molecule. MHC is very highly polymorphic; there are thousands of alleles in the human species. Since the TCR rearrangements are random, a brand-new thymocyte’s receptors will bind to the particular MHC molecules it encounters on macrophages and epithelial cells on its trip through the thymus with either high, low, or no affinity.
negative selection of T cells
The first possibility is that the immature T cell’s receptor binds to MHC (which will have a “self” peptide in it, derived from a normal protein) with high affinity. By high we mean high enough to result in the activation of the T cell. This is clearly an undesirable cell as its activation would result in autoimmunity. The fate of this immature cell is clear: it dies by the process of apoptosis. The proportion of cells that do this is hard to estimate, but it must be rather small and the phenomenon cannot be directly studied in the normal thymus. But with the development of transgenic techniques, it has become possible to create mice all of whose T cells bear the same receptor. With the appropriate cross-breeding scheme, mice can be generated in which all the developing T cells have high affinity for the MHC they find in the thymus. These mice have double-negative (CD4-/CD8-) thymocytes, but no double- or single- positives, and no T cells; they were all deleted. So negative selection takes place at the transition to the double-positive stage. The process is functionally identical to B cell clonal deletion. This mechanism would delete T cells reactive against the sorts of peptides you’d expect to find expressed in the thymus; but what about liver or thyroid or adrenal-specific gene products? Amazingly, the AIRE (autoimmune regulator) gene, which encodes a transcription factor, causes thymic medullary stromal cells to express a wide variety of otherwise-inexplicable “out- of-place” peptides so that reactive T cells may be removed from the repertoire. In fact, Aire- deficient people develop multiple autoimmunities.
non-selection of T cells
Since the repertoire of T cells is generated by random association of V, (D), and J gene segments, it is reasonable to assume that most of the resultant TCR will have essentially no affinity for the particular MHC molecules they find expressed in the thymus. The cell thus receives no stimulation through its TCR. Under these circumstances it will die in a day or two, again by apoptosis.
positive selection of T cells
If there is low but real affinity of binding between the TCR and the MHC of the thymic stroma (with a “self” peptide in the cleft), the cell binds just enough, not to be deleted, but to be signaled to mature (positive selection). The idea here is that low affinity for self might turn out in the periphery to be high affinity for self + some foreign peptide. This model explains MHC restriction: the T cells that emerge from the thymus of an “A” animal or person see antigen plus “A” MHC, because they were positively selected on “A”. There is plenty of experimental evidence to support this: for example, mice that genetically lack MHC Class I develop normal Th cells but no CTL, because there was nothing in the thymus for their developing CTL to bind to. Although it’s very difficult to do, enough x-ray crystal structures of TCR-peptide-MHC complexes have now been solved that we can say positive selection takes place when the CDR 1s and CDR2s of the TCR α and β chains interact adequately with amino acid residues on the alpha-helical sides of the peptide-binding MHC groove. This is not enough binding energy to be activating, but enough for selection. In the periphery, if the peptide that loads into MHC makes appropriately strong contacts with the CDR3s of the α and β chains, the total binding energy is now sufficient, and the T cell will be stimulated.
extrinsic pathway.
Macrophages phagocytose antigens, digest them in phagolysosomes, load peptides onto MHC Class II, and recycle them to the surface. Of course, like all cells, macrophages also express Class I. B cells use surface Ig to bind an epitope of an antigen, internalize it, digest it to peptides which are loaded onto MHC Class II and recycled to the surface for interaction with Tfh2 cells. Dendritic cells, the best APC, take up antigen and process it for MHC Class II as do macrophages or B cells; but there is also “cross-presentation” by the intrinsic pathway, so some peptides are presented on MHC Class I as well. The result is that DC stimulate both Th and CTL.
intrinsic pathway.
When body cells like this virus-infected hepatocyte make proteins, they shuttle peptides derived from the nascent protein from the cytosol to the endoplasmic reticulum, and thence to the surface, presented in MHC Class I.
MINOR HISTOCOMPATIBILITY ANTIGENS
There are about 30 minors, mismatch at any of which may cause slow (chronic) rejection. One is H-Y, coded for on the Y chromosome. It’s not a transmembrane surface molecule, but an internal protein whose peptides are displayed on MHC Class I (only on male cells, of course). Because of H-Y, male skin grafts will be slowly rejected, even by fully syngeneic inbred females, while males accept female grafts without fuss.
ARCANE TERMINOLOGY
Grafts between genetically identical individuals (e.g., inbred mice, identical twins) are called syngeneic or isografts; between non-identical members of the same species (e.g., people) are allogeneic or allografts; and between members of different species (e.g., baboon hearts into babies) are xenogeneic or xenografts. Grafts from one individual to himself (e.g., hair transplants) are autografts.
autoimmune regulator (AIRE)
a protein that in humans is encoded by the AIRE gene.[1] AIRE is a transcription factor expressed in the medulla of the thymus and controls the mechanism that prevents the immune system from attacking the body itself. In the thymus, the autoimmune regulator causes transcription of a wide selection of organ-specific genes that create proteins that are usually only expressed in peripheral tissues, creating an “immunological self-shadow” in the thymus. It is important that self-reactive T cells that bind strongly to self-antigen are eliminated in the thymus (via the process of negative selection), otherwise they can later bind to their corresponding self-proteins and create an autoimmune reaction. So the expression of non-local proteins by AIRE reduces the threat of the occurrence of autoimmunity later on by allowing for the elimination of auto-reactive T cells that bind antigens not traditionally found in the thymus. Furthermore, it has been found that AIRE is expressed in a population of stromal cells located in secondary lymphoid tissues, however these cells appear to express a distinct set of TSAs compared to mTECs
affects of stress on RBC development
Development from pluripotential hematopoietic stem cells to the most mature red cell in the marrow, the reticulocyte, takes 10-14 days. Reticulocytes may be identified in the peripheral blood for 1 day after release. During stress, the time for red cell production may decrease to 5-7 days, production can increase by 6-8 fold above baseline, and reticulocytes are released early. These “stress” reticulocytes may be detected in the circulation for several days.
RBC turnover
Although the specific changes which limit the lifespan of normal red blood cells is not completely understood, several processes contribute to normal red cell turnover. Age related decreases in red cell enzyme activity, oxidative damage to cell constituents, changes in calcium balance, alterations in membrane carbohydrates and expression of senescent antigens to which there are naturally occurring antibodies all play a role in the 120 day lifespan of the normal red cell. Most of the turnover (90%) occurs in the spleen; the red pulp presents a challenging metabolic environment and the splenic macrophages provide a barrier that culls out old cells.
Etiology
the study of the cause of disease and illness
etiology of hemolysis
Red cells circulate as biconcave discs which have a diameter slightly larger than that of the capillary. Movement through capillary beds requires red cell flexibility which depends on the red cell plasma membrane and associated cytoskeleton. The plasma membrane is a lipid sheath containing phosphatidylserine, phosphatidylethanolamine and phosphatidylinositol fixed to an underlying protein network. Two integral proteins penetrate the lipid bilayer, glycophorin A and component a, and are important for the negative membrane potential of the cell and glucose transport, respectively. Attached to the plasma membrane through an interaction with these integral membrane proteins, the cytoskeleton has two major constituents, spectrin and actin. These proteins, along with other associated proteins, comprise the cytoskeleton critical to the red cell shape and other mechanical properties. Defects in the cytoskeleton play a role in specific hemolytic disorders.
metabolism of RBC
Because the mature red cell lacks a nucleus and other organelles such as mitochondria, its metabolism is simpler than other cells. Energy is generated by the breakdown of glucose through the Embden-Meyerhof pathway. The metabolism of glucose to lactate and pyruvate provides ATP necessary to maintain the plasma membrane and cytoskeleton, and energize metabolic pumps to control intracellular sodium, potassium and calcium. ATP is critical for cell survival. Three associated pathways are also important for cell function. The Rapoport Luebering pathway produces 2,3-diphosphoglycerate which stabilizes the deoxy form of hemoglobin and maximizes transport of O2 to tissues. The hexose monophosphate shunt (phosphogluconate or pentose pathway) produces reduced pyridine nucleotide which reduces glutathione and provides protection from oxidant stress. Finally, the methemoglobin reductase pathway maintains the iron in hemoglobin in the ferrous state required for reversible oxygen binding by hemoglobin. Dysfunction in these pathways may lead to decreased survival and/or altered function.
location of erythropoiesis
The location of red cell production varies in fetal life, childhood and the adult. In the first two months of fetal life, production of red cells occurs in the yolk sac. Beginning at approximately the second month of gestation, production of red cells is transferred to the liver and spleen, peaking at about five months gestation and disappearing from these organs by normal parturition. Beginning somewhere in the second trimester, hematopoiesis moves to the bone marrow in the axial skeleton and distal long bones. At birth, this is the primary site for red cell production. During childhood, active marrow in long bones recedes so that by the adult years most hemapoiesis occurs in the axial skeleton.
Erythropoiesis
Pluripotent stem cells are the source of all immune and hematologic cells. A regenerating pool of pluripotent stem cells retains the marrow hematopoietic capacity while development of lineage specific progenitor cells provides the required leukocytes, erythrocytes and platelets. The earliest identifiable cell in the red cell series is the burst forming unit erythroid (BFUE) and is controlled by growth factors which include IL-3 and GM-CSF derived from stromal cells, lymphocytes and macrophages. The next step (differentiation) is the formation of the colony-forming unit, erythroid (CFUE). This is under the control of the hormone, erythropoietin (EPO). The CFUE progresses (matures) into a series of erythroid precursor cells, easily identified in the marrow and termed normoblasts. Normoblasts progress through distinct steps as basophilic, polychromatophilic, orthochromatic normoblasts. This process takes 7-8 days and is associated with the progressive formation of hemoglobin, change in size of the cell and loss of mitochondria, RNA and the cell’s nucleus. The first mature red cell is termed the marrow reticulocyte which contains some of the remnants of messenger RNA and remains in the marrow for 3-4 days before being released into the peripheral blood. Reticulocytes can be identified in the peripheral blood (stain for the RNA) for an additional day with the mature erythrocyte in the circulation surviving for more than 100 days.
Hemoglobin Structure and Synthesis
The hemoglobin molecule is composed of four subunits: two alpha chains and two beta chains. Each of the globin chains contains a pocket for heme molecule, and therefore, has a capacity to bind oxygen through its interaction with the iron (ferrous form) molecule contained in the heme ring. A single molecule of hemoglobin can bind up to four molecules of oxygen. 2,3 DPG binds to the two beta chains and stabilizes the deoxy form of hemoglobin. The globin molecules are gene products of the globin genes found on chromosomes 16 and 11. The major polypeptide genes include γ, α, δ, and β and hemoglobins include α2β2 (or A, adult hemoglobin); α2γ2 (fetal hemoglobin); α2δ2 (A2 hemoglobin). During human development there is sequential suppression and activation of individual globin genes to provide the predominant hemoglobin during that stage. In fetal life, the major form is fetal hemoglobin with high affinity for O2 (γ chains have no 2,3-DPG binding site, produce right-shifted oxy-hemoglobin dissociation curve, and favor bound oxygen at any oxygen tension) which allows the fetus to develop at the venous O2 saturation levels seen in placental circulation. During the postnatal period, there is a switch from fetal hemoglobin to adult (A1) hemoglobin. The transition takes place somewhere in the first two months of life, and after six months of life A1 is the predominant hemoglobin with a small amount of A2 hemoglobin also produced. Hemoglobin is produced in the mitochondria in maturing normoblasts. Protoporphyrin is synthesized in mitochondria and iron, which is obtained from transferrin and placed temporarily into ferritin stores, is added to the porphyrin ring to make heme. This is bound to the predominant globin chain to make hemoglobin.
