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Flashcards in Clinical Deck (167):
1


Hematopoiesis

Continuous process of developing circulating blood cells from primitive progenetor cells.

2


Daily hematopoiesis requirements

  • Erythrocytes - 120 day lifespan - 10^12/day
  • Granulocytes - 0.5 day lifespan - 10^11/day
  • Platelets - 7-10 days - 10^11/day

3


Development of hematopoiesis


Embryo - made in yolk sac, intravascularly - RBCs nucleated - embryonic hemoglobin

Fetus - 1.5% of body weight - made in liver (mainly), spleen - RBCs anucleate - fetal hemoglobin

Adult - 4.5% body weight - made in marrow (long bones - vertebrae, sternum in adults, femur more through 25) - RBCs anucleate

4


Hematopoiesis requirements


Stem cells - multipotent precursors - can produce all cell lines, are self-renewing, capable of repopulating (e.g. bone marrow transplant), present in very low numbers, morphologically indistinct, dapable of mobility and redistribution via circulation

 

Stroma - provide microenvironment

 

Growth factors - regulate maturation

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Hematopoietic Hierarchy

Once committed to myeloid or lymphoid, cells are locked into that lineage

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Stem cell self-renewal

 

  • Asymmetric - stem cell divides to one stem cell, one progenitor
  • Symmetric - certain stem cells divide to become only stem cells, some become only progenitor cells

7


Bone marrow architecture

 

  • Cords - mixture of cells in various stages of maturation
  • Sinuses
  • Marrow-blood barrier
  • Adipose and fibroblasts and their secretions - provide microenvironment to allow stem cells to survive

8


Important growth factors

 

  • Epo (erythropoietin) - for differentiation of erythrocytes
  • TPO (thrombopoietin) - for differentiation of megakaryocytes
  • G-CSF/GM-CSF - for differentiation of white cells (basophils, neutrophils, eosinophils, monocytes - aka myeloid cells)

9

Hematopoietic GFs

 

  • All except Epo:
    • Redundant - stromal cells can make more than one GF
    • Pleiotrophy - each GF has multiple functions, stimulates multiple progenitors
    • Synergy - most have overlapping functions, combinations more effective than individual GFs
  • GF synthesis highly localized (GF tethering)
  • Myeloid GFs influence both progenitors and mature progeny
    • Maintain viability, initiate cell cycle activity, activate effector functions

10

Non-clonal disorders

  • Stem cell suppression
    1. Multipotent stem cells - marrow aplasia (aplastic anemia\pancytopenia)
    2. Committed precursors - pure red cell aplasia, agranulocytosis, amegakariocytic thrombocytopenia

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Clonal expansions


Proliferation and expansion of a single stem cell and suppression of the normal population

(Normally polyclonal - different stem cells producing progeny.  Cancer is monoclonal)

 

Different phenotypes - acute leukemias, myelodysplastic, myeloproliferative disorders

12


RBC maturation

 

  1. Decreasing cell size, decreasing nuclear-cytoplasmic ratio
  2. Nuclear maturation - chromatin clumping and extrusion
  3. Cytoplasmic maturation - hemoglobin synthesis complete, nucleus gone

RBCs normally about the size of a lymphocyte nucleus, central pallor 1/3 cell diameter, 120 day lifespan (1/120th replaced every day - explains normal 0.8-1% reticulocytes)

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Reticulotyes

Last stage before maturation, released into blood

Gray due to ongoing RNA presence synthesizing hemoglobin

Slightly larger than mature RBCs

1% of circulating RBCs - increased amount reflects increased RBC production - markedly increased in hemolytic states

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Polychromasia


Increased RBC production

Supra-vital staining - shows dark spots on reticulocytes d/t high RNA content - used to see reticulocyte content on blood smear

15


Erythropoietin (epo)


Main regulator of RBC production

Secreted by kidney via O2 sensors

HIF-alpha - hypoxia-induced factor - normally ubiquitinated in presence of sufficient O2 - able to bind and stimulate Epo gene transduction under low O2 conditions

Epo levels normally linearly related to anemia levels (healthy kidney), decreased in polychythemia (bone marrow problem)

Often elevated in smokers - secondary erythrocytosis

Acts on marrow via Jak-State signaling pathway

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Anemia


Decreased total erythrocyte circulating mass

Causes decreased O2 delivery to tissue - main problem

May be a sign of another problem

17

Anemia H&P, labs


History

  • Known bleeding
  • Signs of hypoxia - fatigue, SOB, etc.
  • Signs of hemolysis - dark urine

Physical

  • Jaundice
  • Pallor
  • Tachycardia/tachypnea
  • Lymphadenopathy, splenomegaly

Labs - CBC w/ diff

  • Severity - RBC, Hb, HCT
  • Characteristics - MCV most useful

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Microcytic anemia


Low MCV

Etiology

  • Fe deficiency most common
  • Hemoglobinopathies - thalassemias, sickle cell
  • Membrane defects - hereditary spherocytosis

Workup

  • H&P, CBCD, smear, iron levels, Hb electrophoresis (for hemoglobinopathies)

19


Iron deficiency characteristics

 

  • Bleeding
  • Cigar-shaped cells
  • Hypochromia
  • MCV usually 70s
  • Platelet count may be high
  • May be asymptomatic

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Sickle cell disease characteristics

 

  • Joint pain often
  • Sickle shaped cells
  • MCV 70s or low 80s
  • No hypochromia

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Membrane abnormalities

  • Hereditary spherocytosis - most common
    • Small cells without central pallor
    • MCV <70
    • Large RDW
  • Hereditary elliptocytosis
  • Stomatocytosis

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Thalassemias

Very low MCV

Hypochromic

Wide symptom range - asymptomatic to transfusion dependent

Often normal RBC count

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Macrocytic anemias

Etiologies

  • Nutritional - B12, folate deficiencies
  • Hemolysis usually
  • Myelodysplastic syndromes
  • Medications - hydroxyurea, AZT, phenytoin
  • Toxins - alcohol

Workup

  • Ask about alcohol, medications, supplements
  • B12, folate levels
  • LDH, reticulocyte count, haptoglobin - for hemolysis

 

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B12, folate deficiencies


B12

  • Associated with macroglossia, neuropathies
  • May have very low Hb
  • Usually due to absorption problems
  • May present with multi-lineage cytopenias and hemolysis

Folate

  • Usually due to poor intake
  • Absorption to RBCs slower than into serum - specific RBC folate test

25

Myelodysplasia


Presents in middle age or older

Normal or high B12, folate

Usually slow onset

Requires marrow biopsy to diagnose

 

26


Hemolytic anemia etiologies

Immune

  • Transfusion reaction
  • Drug-induced immune hemolysis
  • Autoimmune

Non-immune

  • Usually drug-induced
  • Infectious
  • DIC
  • HELLP syndrome
  • Sickle cell, thalassemias
  • G6PD deficiency
  • Toxins
  • Cell membrane/enzyme deficiencies

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Hemolytic anemia workup

 

  • Basic labs - LDH, reticulocytes, haptoglobin, smear, bilirubin
  • Physical - hypersplenism, fever, hepatomegaly
  • PNH screen - rare - CD55, CD59 flow cytometry
  • Coomb's test and cold agglutinin - for immune
  • G6PD level
  • Hb electrophoresis
  • DIC panel
  • Other as suspected

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Normocytic anemia


Often underproduction of RBCs

May be mixed with - deficiencies, hemolysis, transfused sickle or thalasemmia patient

Systemic\chronic illnesses - hepatitis, sepsis, renal failure

Other: aplastic anemia, marrow infiltration anemias, toxins, pregnancy, thyroid disorders, normal variant

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Anemia of chronic inflammation or kidney disease

Both cause normocytic, normochromic anemia

Can be macro or microcytic

Determine if treatment is needed or helpful - exogenous erythropoietin (caution of thrombosis risk, do not give with cancers)

A poor prognostic sign in illness - treatment often not needed since symptoms are due to underlying disease

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Marrow infiltration


Hematologic malignancy

  • Leukemia
  • Multiple myeloma
  • Lymphoma

Solid tumor

Scar - myelofibrosis, HIV

Aplastic anemia

Pure red cell aplasia - parvovirus

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Drug-induced anemia

Potentially caused by most drugs

Hemolysis mechanism

  • G6PD deficiency - malarials, sulfas, nitrates
  • Lidocaine
  • IVIG, winrho

Underproduction mechanism

  • Chemotherapy - methotrexate, cyclosporine, hydroxyurea
  • OTC/herbals - may contain lead, mercury, heavy metals
  • Many drugs

 

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Infectious anemia


Parasytes - usually hemolytic

Parvovirus - hypoplastic anemia, pure red cell aplasia

HIV, hepatitis - bone marrow failure of various mechanisms (e.g. other infection, iniltration)

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Anemia based on RBC size

Microcytic - MCV < 80

Normocytic - MCV 80-100

Macrocytic - MCV > 100

34


Microcytic anemia causes

Anything that decreases RBC content

  • Iron deficiency
  • Hemoglobinopathy - thalassemias

Less common:

  • Chronic inflammation - can be normocytic
  • Copper deficiency
  • Lead poisoning
  • Sideroblastic anemia

 

35


Iron deficiency anemia

Most common anemia cause

Microcytosis due to decreased Hb synthesis

Hypochromic usually

Normal iron loss - 1mg/day for men, 1.5 mg/day for menstruating women

Only absorb ~10% of dietary iron - require 10-15 mg/day

Requires acidic stomach environment with intact duodenum

36


Iron deficiency causes

Increases loss

  • GI most common - varices, gastritis, AVM, polyps
  • GU - menorrhagia, birth
  • Donation/phlebotomy

Inadequate intake

  • Achlorhydria
  • Celiac disease
  • IBD
  • Partial gastrectomy
  • Bariatric procedures (gastric bypass)

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Iron deficiency symptoms

General anemia symptoms - fatigue, SOB, dyspnea on exertion (DOE), dizzyness on standing, low exercise tolerance

Pica - craving non-food items in diet - clay (geophagia), paper (amylophagia), ice (pagophagia) - specific for iron deficiency

Signs - tachycardia, tachypnea, orthostasis, pallor

38


Iron lab results

Ferritin most important for diagnosing iron deficiency anemia - acute phase reactant (increased in inflammation, chronic disease) - only decreased in iron deficiency (reflects iron load in blood)

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39


Iron deficiency treatment

PO iron - ferrous sulfate, ferrous gluconate, ferrous fumarate - 325 mg tablets contain ~65 mg available elemental iron - 10% absorbed

Normal total iron stores ~ 3000mg

Takes 3-6 months to increase iron stores in best case

Constipation common, nausea less common

Reticulocytosis often first sign tx - 1g Hb increase 2-4 weeks later

IV iron available, but anaphylactic reactions are common

40


Macrocytic anemia causes


Most common - B12, folate deficiencies - both needed for DNA synthesis

Other - alcohol, hypothyroidism, liver disease, myelodysplastic syndromes, reticulocytosis, medications (especially metabolite inhibitors - hydroxyurea, azathioprine, methotrexate, etc.),

41


Megaloblastic anemia

Only in problems of DNA synthesis - B12, folate deficiencies, myelodysplastic syndromes

Dissociation of nuclear and cytoplasmic maturation of all blood cells - seen by morphological changes on smear:

  • E.g. hypersegmented neutrophils
  • Macrocytic RBCs

42


B12 physiology

Cobalamin in various forms

General properties

  • Water soluble
  • Synthesized by microorganisms
  • Found in animal products
  • Storage pool adequate for 3-4 years - in liver
  • Enterohepatic circulation required for maintaining storage

Absorption

  • Passive - rapid, but inefficient - 1%
  • Active - B12 bound by R-binder in salivary secretions, then released from R-binder by pancreatic proteases, binds to intrinsic factor from stomach parietal cells - IF-B12 complex absorbed in ileum

Pernicious anemia - autoantibodies against IF

B12 needed as a coenzyme for conversion of methyl-malonyl-CoA to succinyl-CoA and homocysteine to methionine

MM-CoA AND HOMOCYSTEINE ELEVATED IN B12 DEFICIENCY

Versus folate deficiency - only homocysteine elevated

43


Folate physiology


Aka folic acid

General properties

  • Water soluble
  • In most foods - highest in liver, yeast, spinach, greens, nuts
  • Easily destroyed by boiling
  • Smaller reserve - 3-4 months - liver, little enterohepatic circulation

Absorption

  • Rapid from entire small intestine
  • 50% absorbed
  • Monoglutamate forms easiest absorbed - converted to methyl-tetrahydrofolate (MTHF) in mucosa
  • No co-transporters required (e.g. IF)

Required for homocysteine to methionine

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B12 deficiency manifestations

Anemia with or without leukopenia/thrombocytopenia

Epithelial - microscopic changes - more multinucleate and dying cells - angular chelitis (lip, mouth cracking), glossitis

Neural tube defects, cleft palate

Vascular disease - hyperhomocysteinemia predispose to thrombi

Neurologic - subacute degeneration of dorsal and lateral white matter of spinal cord - slow progressive weakness, sensory ataxia, paresthesias leading to spasticity, paraplegia, incontinence

  • Not seen with folate deficiency
  • Dementia, psychiatric disturbances

 

45


Folate deficiency manifestations

Any of the same presentations as B12 deficiency, but no neurologic or psychiatric findings

Timing - folate depleted in months, B12 in years

46


B12, folate labs

Interpret direct B12 or folate levels with caution - low normal B12 can cause symptoms, serum folate can be artificially elevated due to a recent meal - measure RBC folate

Low blood count with macrocytosis

Anisocytosis and macro-ovalocytes seen on smear

Hallmark of hypersegmented PMNs - >5% with 5 lobes or >1% with 6 lobes

Other labs may be similar to hemolytic anemia but reticulocyte count will be low

B12 - homocysteine and MMA high

Folate - only homocysteine high

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B12 deficiency treatment


Initial replacement parenteral - 1000ug IM qDay x7, then weekly for 30 days, then monthly

1000-2000ug PO daily - usually only for maintenance

Monitor K+ during initial repletion in severe anemia

48


Folate treatment

1-5 mg/day PO until replete or indefinite

Combined folate/B12 deficiency - give B12 first if neurologic signs present - can be made worse and/or irreversible if folate given alone

49


Hemagglutination


Antibody causing formation of RBC aggregates

The basis for many blood bank tests

IgM - cause hemagglutination - large enough (pentameric) to overcome repellant forces between RBCs

IgG - cannot cause hemagglutination due to small size (monomer)

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Anti-Human Globulin (AHG)


Blood bank reagent - reacts with IgG on RBCs to cause hemagglutination

Allows detection of red cells coated with IgG antibodies

Direct Antiglobulin Test (DAT) - determines IgG or C3 (footprints of former IgM) directly coating RBCs - aka Direct Coombs

DAT used to determine immune hemolysis - autoimmune hemolytic anemia, drug-induced hemolysis, transfusion reactions

Indirect (IAT) - determines IgG in serum - aka Indirect Coombs

IAT used to determine RBC compatability pre-transfusion

51


Intravascular Immune Hemolysis


Usually due to IgM

IgM causes mechanical destruction of RBCs and complement fixation/lysis causing release of free RBC stroma - stimulates vasoactive peptides and clotting

Signs - back pain, hemoglobinemia (brown blood), hemoglobinuria, fever, coagulopathy, hypotension, DIC, vascular collapse, pulmonary compromise, renal failure, death

52


Extravascular Immune Hemolysis


Usually due to IgG

No lysis of RBCs - IgG an inefficient complement activator

IgG-coated RBCs cleared by reticuloendothelial system, broken down within RES cells

Signs/symptoms - low-grade fever, small decrease in RBCs, sometimes asymptomatic

53


Lab signs of hemolysis

Spherocytes

Nucleated RBCs

Reticulocytosis

Elevated LDH

Elevated bilirubin

Low haptoglobin

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ABO Blood group system


ABO antigens - complex carbohydrates on RBC glycoproteins/glycolipids

Glycoconjugate - precursor chain

H-chain - glycoconjugate plus fucose

ABO antigens added/absent based on genetic loci

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ABO antigens

O transferase - non-function, does not modify H-chain

A transferase - adds galactosamine to H-chain

B transferase - adds galactose to H-chain

AB - galactose and galactosamine to H-chain

One gene passed by each parent - multiple genotypes cause each phenotype (e.g. A/A or A/O genes present as A phenotype)

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ABO antibodies

IgM (some IgG) - arise naturally in individuals lacking the corresponding antigen

Seen after infancy following stimulation by cross-reaction with environmental antigens (maybe gut NF)

Cause hemagglutination of antigen-positive RBCs at body temperature - basis of transfusion and solid organ compatability

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Rh blood group system


Unlike ABO antigens, Rh antigens are only on human RBCs

Inherited as a set from each parent - D gene most important

D antigen positive - Rh+

D antigen negative - Rh-

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Rh antibodies

Usually IgG

Do not occur naturally - alloimmunization to Rh antigens occurs by exposure to human RBCs

Rh negative patients - may make IgG anti-D antibodies after transfusion with D positive blood or pregnancy with D positive fetus

Antibody production occurs in 80% of Rh negative patients upon exposure to Rh positive RBCs

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Hemolytic Disease of the Newborn (HDN)


Maternal IgG antibodies cross placenta - IgM too large

Hemolytic disease results if maternal IgG are specific for fetal RBC antigens

Anti-Rh antibodies more severe than other antibodies (e.g. Rh(-) mother with Rh(+) fetus)

Erythroblastosis fetalis - pathophysiologic presence of immature RBCs in fetal blood d/t hemolysis

Hydrops fetalis - clinical outcome of fetus - edema of fetus d/t high output cardiac failure secondary to anemia

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HDN Prevention


HDN usually caused by sensitization of an Rh(-) woman sensitized to Rh(+) during fetal maternal hemorrhage during pregnancy - all subsequent Rh(+) fetuses at risk

HDN prevented by injecting Rh(-) mothers with anti-D from sensitized donor plasma - RhIG - only works on naive women (no previous exposure to Rh(+))

RhIG given in 3rd trimester and again at delivery to prevent alloimmunization during fetal meternal hemorrhage

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ABO HDN

Milder than anti-D HDN, can occur during first pregnancy

Usually seen in O type mother with A or B type fetus

No prevention

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Blood compatability testing


Type - ABO and D antigen type of RBCs

Screen - performed on serum to determine presence of unexptected antibodies - by testing serum with sample RBCs of known type

Errors almost always due to improper patient ID or sample labeling at collection

RBC transfusion - avoid incompatability with ABO hemagglutinins (e.g. don't give type A patient type B RBCs)

Plasma transfusion - avoid incompatability with ABO antigens (e.g. don't give type A patient plasma with anti-A antibodies)

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Rh compatability


Rh+ - can get either Rh+ or Rh-

Rh- - should only get Rh- - can get Rh+ once, but will develop anti-D antibodies

Ignore Rh type for plasma transfusions

Honor Rh type for platelet transfusions - platelet products have some RBCs in them - ABO mismatch of platelets considered safe

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Blood crossmatch


Testing patient's serum with RBCs to be transfused to monitor for agglutination

A double check to type and screen - ensures no agglutination occurs in vitro before actually transfusing

65


Blood donation

All volunteer - participate in confidential deferral process to reduce risk of transfusion not detected by blood testing alone

Usually taken as whole blood, then split into components later - RBCs, platelets, plasma

Blood testing - HIV, Hep B, Hep C, Syphilis, West Nile, HTLV, trypanosomes causing Chagas

CMV only tested when transfusing to immunodeficient patients

Platelets stored at room temperature - screened for bacteria

 

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Hemolytic anemias


Decreased RBC lifespan

Membrane damage of RBCs leads to Hb release

Classified as intrinsic or extrinsic membrane disorders

Extrinsic membrane disorders - cytoplasmic or extracellular

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RBC Membrane structure


Lipid matrix

Structural proteins - spectrin mesh interconnected by proteins, including actin.  Anchored to the lipid membrane by ankyrin

Non-structural proteins - e.g. antigens

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Signs and symptoms of hemolytic anemia


Lab signs of hemolysis

  • Reticulocytosis (polychromasia)
  • Unconjugated hyperbilirubinemia
  • Increased fecal and urine urobilinogen
  • Decreased serum haptoglobin (binds free Hb)
  • Elevated LDH (liberated from RBCs)
  • Mildly elevated AST
  • Hemoglobinemia, hemoglobinuria, hemosiderinuria

Clinical signs

  • Cholelithiasis (brown stones)
  • Leg ulcers
  • Aplastic crises
  • Hyperhemolysis
  • Skeletal abnormalities

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Types of RBCs

Discocyte - 140 u^2 area

Spherocyte - 97 u^2 area

Target cell - 200 u^2 area

Area assuming same cell volume

70

Hereditary Spherocytosis (HS) Etiology, Pathophysiology

  • Spectrin Deficiency
    • Mild-moderate - AD (75%)
    • Severe - AR (25%)

Beta-spectrin to 4.1 protein interaction defect

Pathophys - cytoskeleton abnormality causes membrane instability and loss leading to low surface area to volume ratio, increased osmotic fragility, and less flexibility of RBCs

Above leads to splenic trapping, hemolysis

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HS clinical

Chronic anemia - pallor, jaundice, dark urine, splenomegaly, cholelithiasis

Aplastic and hyperhemolytic crises

Diagnosis

  • Family history often positive
  • Anemia
  • Spherocytosis
  • Reticulocytosis
  • Increased osmotic fragility and low RBC deformability

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Osmotic fragility


RBCs normally can tolerate changes in serum tonicity by swelling (hypotonic solution) or contracting (hypertonic solution)

In hereditary hemolytic anemias, altered cytoskeletons mean RBCs are less able to tolerate changes in tonicity and hemolyze earlier than normal RBCs under changing conditions

73


HS treatment


Symptomatic - transfusion, cholecystectomy, etc.

Splenectomy - curative - no more trapping of RBCs reduces symptoms due to hemolysis - RBCs remain abnormal but function well enough

74


Hereditary Elliptocytosis (HE) types

  1. Mild (90%) - benign, often discovered incidentally
  2. Hereditary Pyropoikilocytosis (HPP)
    1. Rare, severe anemia
    2. RBC fragmentation
    3. Heat sensitivity
    4. AR
  3. Spherocytic HE (10%)
  4. Stomatocytic HE

Dx - morphologic - 20% elliptocytic RBCs

75

Paroxysmal Nocturnal Hemoglobinuria (PNH)


Acquired intrinsic membrane abnormality (only acquired one)

Deficient glycophosphatidylinositol (GPI) - part of complement regulators CD55 and CD59

Causes sensitivity of RBCs to complement, leading to intravascular lysis

PIG-A gene somatic mutation on X chromosome - somatic, so affects both males and females

Associated with aplastic anemia, venous thrombosis, pancytopenia, iron deficiency

Diagnosed by flow cytometry

Presents as chronic hemolytic anemia or venous thrombosis.  Associated with aplastic anemia.

Tx - symptomatic and/or eculizumab (complement inhibitor)

76

Hexose Monophosphate Shunt (HMS) disorder

G6PD deficiency

Sex-linked - most common in african americans

Exposure to oxidant stress leads to hemolysis due to precipitation of Hb within RBCs (Heinz bodies)

Less common variants with chronic hemolysis

Screening - add G6P and NADP to patient serum - if G6PD present, G6P + NADP --> Phosphogluconate + NADPH.  Blue dye + NADPH --> colorless + NADP

Confirm positive screen with quantatative G6PD

Clinical - acute intravascular hemolysis with hemoglobinemia, hemoglobinuria, jaundice - most dramatic case

Often presents with modest decline in Hb without prominent symptoms

 

77


Pyruvate Kinase Deficiency

PK catalyzes change of Phosphoenol Pyruvate to Pyruvate

Also associated with ADP --> ATP

2,3-DPG (precursor to phosphoenol pyruvate) accumulates, leading to increased acidity of blood (right shift of O2-dissociation curve)

Low RBC ATP increases permeability to cations

AR

Chronic hemolysis of varying severity, no specific presentation

Dx by enzyme assay

Splenomegaly, macro-ovalocytosis

Symp's improved with splenectomy, but still require transfusions

78

Autoimmune Hemolytic Anemia (AIHA)

Acquired premature RBC destruction due to antibodies against one or more self RBC antigens

Clinical - anemia of variable severity, splenomegaly, positive direct Coombs

Smear - varying size/shape of cells, WBCs

Dx - evidence of autoantibodies or complement (C3d or C4) attached to RBCs

79


Antiglobulin test interpretation

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80

AIHA types

Warm (70%) - IgG-mediated - splenic clearance, complement amplifies effect

  • Most common in middle-aged women
  • Responds to glucocorticoids or splenectomy
  • Half idiopathic, half due to other factors (often medications)
  • Tx - splenectomy, immunosuppresion, prednisone

Cold - IgM-mediated - hepatic clearance, dependent on complement

  • IgM only attached to complement on RBCs at lower than body temperature
  • Often seen with chronic lymphoproliferative conditions
  • Poor clinical response - keep warm

81


Hemoglobinopathy


Diseases in which one or more globin chains are abnormal

 

82


Thalassemia pathophysiology

Decreased production of one or more globin chain

Beta thalassemia - low beta chain production - poorly soluble, excess alpha globin precipitates rapidly

Alpha thalassemia - low alpha chain production - excess, highly soluble beta globin forms HbH (tetramers of beta chains with heme) - HbH easily oxidized, forming inclusions and leading to hemolytic anemia - useless as O2 carrier

  • In children - hemogloin barts formed (tetramer of gamma chains) - reasonably stable, but releases oxygen poorly

83


Sickle cell pathophysiology

Single point mutation on 6th AA acid

  • HbS (sickle Hb) - glutamic acid to valine
  • HbC - glutamic acid to lyseine

HbS - polymerizes under hypoxic conditions, forming sickle shaped RBCs - form thrombi in small vessels causing pain and organ dysfunction

  • Requires both alleles mutated to cause disease.  HbS/normal genotype rarely forms sickles

HbC - most benign hemoglobinopathy - but SC combo (SC disease) causes symptoms of sickle cell, but without cell sickling - dehydrate to form dense cells - very mild to full sickle cell symptoms

  • CC gene - mild hemolytic anemia with splenomegaly - rarely requires treatment

 

84

Sickle/thalassemia combinations

HbS/B-thalassemia - "sickle-thalassemia"

  • Presents similar to sickle cell anemia
  • Beta thalassemia + - mild disease, may be asymptomatic
  • Beta thalassemia null/HbS - more severe
  • Suggested by low MCV - cells usually smaller than sickle cell alone

HbS/A-thalassemia - less severe than regular HbS

  • Due to lower [Hb] in RBC leading to less risk of sickling

85

Variant hemoglobins

HbD, HbE

Usually asymptomatic if heterozygous, mild/asymptomatic anemia if homozygous

HbS/HbD - severe, similar to sickle cell anemia

86

Hemoglobin genetics

Alpha globin cluster - chromosome 16

Beta globin cluster - chromosome 11

B thalassemia - mainly point mutations - B+ (some B-globin produced), B0 (no B-globin produced)

A-thalassemia - mainly deletions of one (a+) or both (a0) cis alpha-globin genes (a0 rare)

HbC/HbS - point mutations in beta-globin gene

  • HbS - GAG->GTG
  • HbC - GAG->AAG

87

Beta-thalassemia clinical

Thalassemia minor (one gene) - usualy mild or asymptomatic anemia

Thalassemia intermedia (B+/B+, B-thal/HbE, other) -

  • Hb 5-10
  • some skeletal abnormalities
  • hepatosplenomegaly

Cooley's anemia/Thalassemia major - Absence or severe underproduction of both genes

  • Skeletal abnormalities or hepatosplenomegaly
  • Iron overload leads to death in adolescence without treatment
    • Death usually from cardiac iron overload
    • Also liver, pituitary pathology
  • Marrow transplant limited due to toxiity
  • Splenectomy helpful to increase lifetime of transfused RBCs

To differentiate from other hemoglobinopathies - no inflammatory markers

Dx by Hb electrophoresis

88

Iron overload

Due to multiple transfusions for anemia of major hemoglobinopathies

Iron deposition causes multiple organ dysfunction - especially cardiac, liver

E.g. desferol, deferaxirox, deferiprone

All use renal route for iron excretion

89

Alpha thalassemia clinical

Usually asymptomatic decrease in Hb - rarely need treatment

Risk of hydrops fetalis

Dx by genetic testing (smear looks largely normal)

HbH - absence of 3 alpha-hemoglobin genes

  • Useless O2 carrier
  • Moderate anemia, hemolysis, sensitivity to oxidative stress
  • Tx - transfusion, splenectomy, chelation

90

Sickle cell clinical

Spontaneous cell lysis and RBC turnover - chronic anemia

Increased thrombosis/infarct risk - esp stroke, pulmonary infarct

Chronic inflammation

Secondary complications

  • Asplenic state d/t infarcts - increased risk of bacterial infection
  • Joint damage
  • Non-healing skin ulcers
  • Retinopathy
  • Nephropathy
  • Opiate addiction, behavior issues d/t chronic pain

Prognosis

  • Significant mortality in infants and children
  • Median age of death ~40 years

Variable phenotype

  • Hereditary persistance of HbF grants protection (does not sickle)
  • Single alpha hemoglobin mutation - lowers Hb concentration, decreasing sickling

Treatment

  • Hydroxyurea - first line - increased HbF
  • Folic acid
  • Marrow transplant - high mortality, not common
  • Anti-inflammatory and anti-coagulation - ineffective
  • Opiates usually required for pain

91

Acute chest syndrome

Complication of sickle cell disease

Severe chest pain with or without hypoxia, cough, other symptoms

May be preceded by pneumonia, infarct, emboli

Smoking increases risk, but 30-50% idiopathic

Tx - exchange tranfusion

92

Hemostasis

Physiologic process of keeping blood liquid under normal conditions and preventing extravasation after vessel injury

Coagulation - formation of fibrin clot

Coagulation factors - plasma proteins regulating coagulation

Anticoagulation factors - plasma proteins inhibiting coagulation

Fibrinolysis - proteolytic digestion of insoluble fibrin clots

 

93

Vascular injury

Subendothelial factors

  • Collagen initiates recruitment and activation of platelets
  • Tissue factor initiates generation of thrombin - converts fibrinogen to fibrin, activates platelets

94

Venous vs. Arterial hemostasis

Velocity of blood flow is a major mechanism of hemostasis

Venous - low flow, low shear - fibrin-rich, platelet-poor clots

Arterial - high flow, high shear - platelet-rich, fibrin-poor clots

Aspirin is better at arterial thrombus prevention

95

Platelets

Small discoid cells

Stages of platelet thrombus formation

  1. Tethering and rolling - mediated by vWF
  2. Activation and adhesion - mediated by collagen
  3. Aggregation - mediated by fibrinogen and vWF

vWF synthesized in endothelial cells

Activation

  • Mediated by adhesion to collagen and thrombin via tissue factor
  • Causes change in shape, release of activating factors (fibrinogen, vWF, ADP, TXA, calcium), and aggregation of further platelets

Disorders most commonly hemorrhagic, acquired

96


Coagulation

Cascade of enzymatic reactions resulting in fibrin clot generation

Insoluble fibrin network helps stabilize the platelet plug

Properties

  • Circulating proenzymes (zymogens) chemically converted to active enzymes
  • Enzymes, cofactors, substrates assemble for maximum speed and efficiency
  • Positive feedback exists - especially for thrombin generation
  • Negative feedback - activated protein C inactivates Va, VIIIa

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Extrinsic Pathway


Principal initiating pathway of in vivo coagulation

TF depending

TF not normally exposed to blood (extrinsic to vasculature) - expressed on adventitia of vascular and non-vascular cells

TF not on healthy monocytes or endothelium unless induced by cytokines

98


Intrinsic pathway

TENET - 12, 11, 9, 8, 10 (common pathway)

99

Thrombin and Fibrin polymerization

Fibrinogen (soluble, large monomer) dimerizes once converted to fibrin by thrombin

Large fibrin polymers assemble, but are not insoluble until cross-linked by factor XIIIa (also activated by thrombin)

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Hemostatic synthesis

Both coagulation and anti-coagulation factors made in liver - can present as either bleeding or clotting - difficult prophylaxis

Factor VII - shortest half-life of all factors

Vit K dependent - Factors II, VII, IX, X, Protein C, Protein S

Vit K catalyzes carboxylation - required for factors to obtain activity by chelation to Ca (involved in factor binding to lipid membranes)

101


Platelet effects on coagulation


Increase catalytic ability of IXa to activate X by 150x

Platelets with factor VIIIa, and ca increase IXa activation of X by 9 million

102

Mechanisms controlling coagulation

 

  1. Normal circulation
  2. Intact endothelium
  3. Anticoagulants
    1. Antithrombin
    2. Protein C
    3. Protein S
    4. TFPI

103


Antithrombin


Inhibitor of XIIa, XIa, VIIa:TF, IXa, IIa, Xa

Made in liver

Serine protease inhibitor

Fairly high plasma concentration

Inhibitory activity accelerated 2000x by heparin

104


Protein C, S

Protein C - binds EPCR receptor on epithelium, becoming aPC (active protein C)

Protein S acts as a cofactor with aPC to degrade Va, VIIIa

105

TFPI

Tissue Factor Pathway Inhibitor

Inactivates and binds Xa, forming a complex with VIIa:TF to inactivate

Synthesized by endothelial cells (85% remains on the surface)

10% in circulation, 5% on platelets

106

Injury and coagulation

Extrinsic pathway - acts as the initiation phase

  • Quickly forms thrombin
  • Quickly inactivated by TFPI

Intrinsic pathway - acts as the propagation phase

  • Thrombin from extrinsic pathway activates XIa to carry out he intrinsic pathway
  • Makes the majority of thrombin

107


Fibrinolysis

Maintains vascular patency, inhibits excess clotting

  • Plasminogen activated to plasmin - lyses fibrin
  • Occurs via tissue plasminogen activators (t-PA) and urokinase (u-PA)

Inhibited by alpha-2-antiplasmin and plasminogen activator inhibitor 1 (PAI-1)

t-PA used clinicaly to digest new clots

108

Endothelium

Promotes blood fluidity

  • Protects blood from exposure to subendothelial factors (TF, collagen) and holds a negative charge to repel negatively charged platelets
  • Produces inhibitors of platelet activation (NO, PGI2, ADPase), coagulation (TFPI, Protein C receptors), and promotors of fibrinolysis (t-PA)

Consequences of damage

  • TF, PAI-1 synthesis
  • vWF secretion
  • Reduced anticoagulants

109


Overview of coagulation and anticoagulation

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Coagulation tests


Use blue top tube - contain exactly 0.5 mL citrate anticoagulant, hold 4.5 mL blood

Testing done on plasma

111

Clot formation testing (PT, aPTT)


Time-based

Done mechanically or optically

PT - measures extrinsic and common pathway

  • Depends on factors 7, 10, 5, 2, fibrinogen (1)
  • Used for monitoring warfarin therapy, screen for Vit K deficiency, factor deficiencies, liver disease, rare acquired factor inhibitors
  • Process - add recombinant TF and calcium to plasma
  • INR - normalizes values to a WHO standard (removes differences due to reagents).  Goal of therapy - 2.0-3.0

aPTT - measures intrinsic and common pathway

  • Depends on all factors except 7, 13
  • Screen for factor 8, 9, 11, 12, von Willebrand disease, lupus, acquired factor inhibitors

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Prolonged clotting time

First step - mixing studies

  • Suggests whether defect is due to factor deficiency or an inhibitor
  • Inhibitors - will act on both patient and "normal" plasma
  • 50% of any factor will give normal clotting time
  • Mix 1/2 normal serum with 1/2 patient serum
    • Clotting time should normalize if due to deficient factor
    • Clotting time remains roughly the same if due to an inhibitor present

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Thrombin time (TT)

Evaluates conversion of fibrinogen to fibrin (II -> IIa)

Elevations in TT

  • Low/absent fibrinogen
  • Dysfunctional fibrinogen
  • High fibrin split products (act as inhibitors, seen in DIC)
  • Myeloma proteins (act as antithrombins)
  • Inhibitors (e.g. heparin)

Reptilase test - same idea, same causes - except normal in presence of inhibitors

 

114

Quantitative fibrinogen

Based on concept of TT being proportional to [fibrinogen]

Indications

  • Evaluation of prolonged PT or PTT
  • DIC
  • Liver disease
  • Other bleeding disorders

Abnormal low values

  • Liver disease
  • DIC, other consumptive states
  • Thrombolytic tx
  • Congenital low/absent fibrogen
  • Abnormal fibrinogen

Abnormal high values - acute or chronic stable liver disease, inflammatory disease (fibrinogen an acute phase reactant)

115

D-Dimer

Indicates both on-going coagulation and fibrinolysis

Factor 13 - forms cross-linking D-D bonds to stabilize clots (coagulation) - cleaved by plasmin (fibrinolysis)

Uses - evaluation of DVT/PE, DIC

Excreted by kidney, half-life 8 hours

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Specific factor assays

Determines extent to which patient plasma corrects clotting deficit in reagent plasmas with a known factor deficiency

117

Antiphospholipid Antibody testing

Antiphospholipid antibody syndrome (APS) - acquired clotting disorder - increased thrombosis and/or recurrent pregnancy loss

Specific antibodies - anti-cardiolipin, anti-beta-2-glycoprotein antibodies

Done by ELISA

Lupus anticoagulant (LA) test - laboratory phenomenon

  1. Increase of any phospholipid-dependent clotting time (PT, PTT, DRVVT, etc.)
  2. Failure of correction with 1:1 mix of normal plasma
  3. Correction of clotting defect with addition of excess phospholipid

118

Dilute Russel Viper Venom Time (DRVVT)

Screen for lupus anticoagulant

Russel viper venom - a direct factor X activator

Adding venom in dilute phospholipid increases sensitivity to presence of anti-phospholipid antibody

Normalized by addition of excess phospholipid

119

Isolated prolongation of PT causes

  • Liver disease
  • Mild DIC
  • Mild Vit K deficiency
  • Factor VII deficiency (rare)
  • Warfarin

Lab approach

  • Repeat test first
  • See if corrected by mixing
    • No - workup for inhibitor
    • Yes - factor assays

120


Isolated prolongation of PTT

No bleeding

  • Lupus anticoagulant
  • Heparin
  • Prekallikrein deficiency (clinically silent)
  • Factor XII deficiency
  • High MW kininogen deficiency (clinically silent)

Bleeding

  • von Willebrand disease
  • Hemophilia A (F8 deficiency), B (9 deficiency), C (11)
  • Heparin or other inhibitors

Lab approach

  • Repeat to confirm
  • Try mixing with normal serum
    • No - inhibitor workup (esp LA)
    • Yes - base further work on H&P - vWF, factor assays

121

PT and PTT prolonged

  • Severe liver disease
  • DIC
  • Warfarin
  • Severe vit K deficiency
  • Fibrinolysis tx
  • Dilution - e.g. massive transfusion
  • Isolated fibrinogen, F2, V, X deficiency

Mixing study correction

  • No - inhibitor
  • Yes - if no known cause, TT and factor assays

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Platelet Testing

Usually by CBC - automated counter

Limitation due to platelet clumping (pseudothrombocytopenia)

Bleeding time

  • Standardized small incision
  • Screen for platelet function defects, assessment of therapy response
  • Inaccurate for predicting operative bleeding

Platelet function analyzer (PFA)

  • Pinhole in bottom of collagen and epi/ADP coated tube - measure time until blood stops dripping

123

Abnormal bleeding time, abnormal PFA

Thrombocytopenia, anemia

vW disease, afibrinogenemia

Acquired platelet disorders - store pool, uremia, anti-platelet drugs

Congenital platelet disorders

124


Platelet aggregation study

Suspension of platelets - add aggregation stimulating agonist and measure clumping by light abs/transmission

Abnormal aggregation

  • Thrombocytopenia
  • vW disease, afibrinogenemia
  • Acquired platelet function disorders
  • Congenital platelet function disorders

125


TPO

Major megakaryocyte and platelet production hormone

Made in liver

Minor production in kidney, SmM, marrow

Negative feedback regulated by megakaryocyte/platelet mass

126

Platelet physiology

Tethering - vWF - secreted by endothelial cells, spontaneously form dimers

  • Dimers pair to form ultra-large multimers, then proteolyzed to large multimers
  • When deposited on damaged vascular surface, multimers unravel to allow binding to platelet receptors
  • Mediate tethering and rolling of platelets

Activation

  • Agonists stimulate conformational change, allowing platelets to bind fibrinogen
  • Fibrinogen forms bridges with nearby platelets

 

127

Presentation of platelet disorders

  • Mucocutaneous bleeding - gingival, epistaxis, menorrhagia
  • Petechia in lower extremities, dependent regions, buccal mucosa, pressure points
    • Do not blanch by pressure
  • GI, GU, intracranial bleeding - in severe cases

 

128


Hemorrhagic platelet disorder types

Quantitative - thrombocytopenia, thrombocytosis

Qualitative - dysfunction

129

Thrombocytopenia

  • Normal - 150-400,000
  • Normal hemostasis >100,000
  • 50-100,000 - bleeding with trauma, increased bleeding time
  • 20-50,000 - excess bleeding with minor injury
  • <20,000 - spontaneous bleeding, life threatening
  • <10,000 - high risk spontaneous bleeds, require prophylactic transfusion

Causes

  • Sequestration - hypersplenism
  • Decreased production - infection, marrow infiltrates, ineffective production (megaloblastosis), hypoplasia, HIV, congenital
  • Increased destruction
    • Immune (drugs, infection, HIV, hepatitis C, APLS, ITP)
    • Non-immune - sepsis, DIC, type 2 vWD, TTP)
  • Pregnancy
  • Dilution
  • Most common overall - hypersplenism, infection, drug-induced destruction, pregnancy

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Thrombocytopenia evaluation

Urgency depends on severity, presence of bleeding/thrombosis, procedure need, presence of anti-platelet drugs

History components

  • Extent and location of bleeding
  • Prior platelet count
  • Recent viral infection (ITP)
  • New medication
  • Ethanol (some megakaryocyte toxicity)
  • Pregnancy
  • Transfusion hx
  • IV devices
  • Liver/renal function

Labs

  • #1 - smear - number, size, clumping, morphology, RBC abnormalities
  • CBC w/ mean platelet volume
  • Immature platelet fraction (IPF)
  • PT, aPTT
  • D-dimer
  • BUN/creatinine
  • Others as indicated by H&P
  •  

131

Thrombocytopenia Treatment

Treat underlying disease

Platelet transfusions

  • If bleeding - get platelets >40,000 until bleeding stops
  • Prophylaxis
    • Semi-controversial - question of benefit, risk of alloimmunization
    • For procedures - >50,000 for minor, >80,000 for major
    • Leukemia >10,000 (during chemo)

 

132

Drug-mediated platelet destruction

Heparin most common

Antibiotics (vancomycin, etc.)

Sulfonamides

Sedatives

Anticonvulsants

Quinine/quinidine

GP IIb-IIIa inhibitors (anti-platelet antibodies)

Diagnosis

  • Usually not with other blood cell abnormalities or splenomegaly
  • Often not definitive - check recent meds
  • Platelet recovery ~7 days after removal of offending drug

133

Suppressed platelet production

Chemotherapy

Ethanol

Thiazide diuretics

134

Heparin-induced Thrombocytopenia (HIT)

Common - 1-3% of heparin patients

Characteristics

  • 20-100,000 platelet count
  • Life and limb-threatening thrombosis

Pathogenesis

  • PF-4 - constituent of platelet granules, released upon activation
  • HIT antibody - anti-PF-4/heparin complex
  • Platelet Fc receptor - binds HIT-Ab/PF4/heparin complex to cause platelet activation
  • Thrombogenic platelet microparticles released --> paradoxical thrombosis

Diagnosis

  • Ultimately clinical - thrombocytopenia in 20-100,000 range, timing (5-10 days from heparin initiation), thrombosis, lack of other causes
  • Lab - PF-4 ELISA test (high sensitivity, low specificity), serotonin release assay (SRA - tests functional HIT antibodies - labor intensive, but high specificity)

Treatment - discontinue all heparin, give alternate anticoagulants (usually direct thrombin inhibitors)

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Immune\idiopathic Thrombocytopenic Purpura (ITP)

Unknown etiology

Not associated with abnormalities in other blood cells, coagulation defects, or splenomegaly

Have normal/increased marrow megakaryocytes

Diagnosis of exclusion - no specific test

Pathophysiology

  • Autoantibody against platelet antigens - usually fibrinogen receptor (GP IIb-IIIa)
  • Causes autoimmune peripheral platelet destruction (mostly in spleen by Fc-receptor macrophages) and inadequate marrow megakaryocyte response

Idiopathic in adults, associated with viral infection in children

Treatment

  • First line - corticosteroids, IV Ig, IV anti-D, TPO mimetics, splenectomy, H. pylori eradication
  • Second - immunosuppression

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Thrombotic Thrombocytopenic Purpura (TTP)

Systemic microvascular thrombosis

  • Thrombocytopenia
  • Microangiopathic hemolytic anemia
  • Tissue ischemia and infarct

Acute, severe, ~100% fatal without treatment

Preponderance of ultra-large vWF multimers due to absence of vWF-protealyzing ADAMTS-13

  • Usually from autoantibodies
  • Ulta-large vWF binds platelets spontaneously, causing aggregates in microvasculature

Treatment

  • Acquired form (most common) - plasma exchange to clear autoantibodies, restore normal ADAMTS-13
  • Congenital - due to absent ADAMTS-13 - plasma infusion
  • Vigorous supportive care

 

137

Qualitative platelet defects

Prolonged bleeding without thrombocytopenia or coagulation abnormalities

Dx by bleeding time, aggregation studies, PFA, etc.

Acquired

  • Usually drugs - aspirin, NSAIDs, thienopyridines
  • Uremia
  • Myeloproliferative disorders
  • vWF disease
  • Storage pool disease
  • Cardiopulmonary bypass

Congenital forms very rare

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Bleeding history

  • General
    • Spontaneous or with provocation?
    • Frequency and timing
    • Transfusions/re-operations?
    • Location(s)
    • Menstrual/OB in females
  • Acquired vs. inherited
    • Age of first event
    • Family hx
    • Hx of systemic disorders associated with bleeding (liver, kidney)
    • Medications/herbs
  • Typical platelet or vWF disease - skin, mucous bleeding
  • Typical coagulation disorders - deep tissue or joint bleeding

139


Bleeding incidence overview

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Bleeding - labs

Screening

  • CBC w/ smear
  • PT, aPTT (coagulation defects)
  • TT - fibrinogen, heparin, DTI
  • Bleeding time, PFA-100, VWF studies (platelet dysfunction, vW disease)

Diagnostic

  • Platelet aggregation studies
  • D-dimer, fibrinogen
  • Specific factor assays
  • Specific vWF testing

141

Acquired and Congenital bleeding overview

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142


Liver disease bleeding

Hemorrhagic tendancy due to deficiency of any number of coagulation factors

Decreased clearance of fibrin degradation products

Decreased carboxylation of Vit K dependent factors

Thrombocytopenia secondary to splenomegaly and decreased thrombopoietin

Lab and treatment

  • PT - very sensitive to factor 7 deficiency (shorted half-life)
  • Factor 8 produced in vascular endothelium as well - rarely decreased until very severe liver disease
  • Fibrinogen - often maintained until very severe
  • Treatment supportive with FFP and platelet transfusions as needed

143


DIC pathophys

Escape of the clotting process from normal inhibition

Characteristics

  • Acute or chronic
  • Systemic
  • Bleeding and/or thrombosis with organ dysfunction
  • Serious, often lethal

Almost always due to an underlying disease - treat to cause

Excess tissue factor

  • Obstetric complications
  • Promyelocytic leukemia (TF in granulocyte precursors)
  • Malignant tumors
  • Trauma, heat stroke, burns, surgery

Endothelial injury and/or factor XII activation

  • Septicemia - particularly gram negative endotoxins
  • Shock

Bleeding results paradoxically from excess intravascular clotting

  • Continuous thrombin formation consumes substrates
  • Fibrinolysis always present to a degree - degrades fibrin, factor V, VIII
  • Fibrin degradation products accumulate - inhibit clotting and platelet function

144


DIC diagnosis, treatment

Dx

  •     Often suggested by clinical setting
  •     No specific lab - long PT/aPTT/TT, low fibrinogen, moderate thrombocytopenia, D-dimers

Treatment

  • Underlying cause
  • Supportive care
  • Transfusion
  • Heparin (when thrombosis dominates hemorrhage)
  • Antifibrinolytics - carefully
  • Activated protein C - used before, not now (stops clotting, but vastly increases bleeding risk)

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Vitamin K deficiency

Factors 2, 7, 9, 10, protein C, protein S - require gamma carboxylation using vit K

  • Essential for calcium binding - mediates adherence to phospholipid surfaces

Poor intake, malabsorption, antibiotics, medications (e.g. coumadin - inhibits vit K epoxide reductase - needed to recycle vit K)

In newborns, causes hemorhagic disease of the newborn

Lab

  • Long PT
  • aPTT long in severe deficiency
  • Corrected with vit K

Therapy

  • PO or IV vit K
  • FFP (fresh frozen plasma) effective, but not required
  • Prothrombin complex concentrates (PCC) - synthetic 2, 9, 10 - sometimes

 

146

Coumadin


Prevents vitamin K oxide reductase

Overdose a common cause of bleeding

Elevated INR without bleeding - withold coumadin

Elevated INR with bleeding - withould coumadin, give vitamin K, consider FFP or PCCs

147


Other common acquired bleeding

Post-operative - often not intrinsic, due to holes

Renal failure - qualitative platelet dysfunction related to degree of uremia

148


Congenital bleeding disorders

Less common than acquired

Hemophilia A - factor VIII

Hemophilia B - factor IX

Hemophilia C - factor XI

Rare factor deficiencies - II, V, VII, X, XIII

von Willebrand Factor Disease

Platelet Function Defects

149

Hemophilia A

X-linked - 1/5000 live male births

  • Females carriers
  • 20-30% of diagnosed cases are spontaneous mutations

Factor VIII deficiency - critical co-factor in activation of X

  • Normally circulates as a non-covalent complex with vWF - VIII deficiencies possible in vWF disease
  • VIIIa - binds platelets and forms binding site for IXa
  • Enhances conversion of X by IXa by 10,000x
  • Ixa-VIIIa-platelet-calcium complex - major activator of X

Severe

  • Factor VIII <1% of normal
  • Spontaneous bleeds - usually joint, deep tissue
  • Bleeding with circumcision

Moderate

  • Factor VIII 1-5%
  • Less common spontaneous bleeds
  • Excess bleeding with trauma/surgery

Mild

  • Factor VIII >5%
  • Bleed only after trauma or surgery

Screening with aPTT - diagnosis with Factor VIII levels

Treatment - Factor replacement therapy - general goals 30-50% of normal for minor bleeds, 80-100% for severe bleeds

  • DDAVP (synthetic vasopressin) - responders have a robust release of FVIII - not for severe disease

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Hemophilia B

X-linked - 1/30,000 live male births

Factor IX deficiency - normally activated by XIa or FVIIa-TF - serine protease that directly converts X to Xa

Clinically indistinguishable from Hemophilia A

Screening by aPTT - diagnosis by factor assays

Tx

  • Recombinant or plasma-derived factor IX
  • PCCs as an alternate
  • Only use plasma if above not available

 

151


Hemophilia C

Factor XI deficiency

AR

aPTT to screen

Phenotype highly variable

Bleeding often in areas of increased fibrinolysis - mouth, GU

Highest in Ashkinazi jews

Treat with plasma

152

Von Willebrand Disease

vWF made in endothelial cells and megakaryocytes

Stored in Weibel-Palade bodies in endothelium, alpha-granules in platelets

Tethers platelets to collagen after vascular injury

Serves as chaperone protein for factor VIII

1% of all people - equal genotype, but females more commonly show phenotype

Presents as platelet function disorders - mucocutaneous bleeding

Type 1 - 70-80% - partial quantitative defect - bleeding

Type 2 - qualitative deficiency

  • 2A - less high MW multimers
  • 2B - gain of function mutation with excess platelet binding and high clearance
  • 2M - poor function with normal multimers
  • 2N - impaired FVIII binding

Type 3 - near complete absence of vWF

vWF often elevated in pregnancy, OCs, liver disease, inflammation, exercise, stress, post-operative

Treatment

  • DDAVP - usually effective in Type 1, contraindicated in type 2B, ineffective in type 3
  • Plasma-derived vWF-FVIII concentrates
  • Cryoprecipitate - not virally-inactivated, only for emergency bleeding
  • Hormonal medications

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Platelet function disorders

Glanzman - IIb-IIIA fibrinogen receptor abnormalities

Bernard-Soulier - Ib-V-IX complex abnormal - vWF binding abnormal

Storage disease - alpha granule or dense granule

Others

154

Venous vs. arterial thrombosis

Venous - fibrin-rich, platelet poor

Arterial - platelet-rich, fibrin-poor

Guides therapy options

155

DVT

Virchow's triad

  • Stasis
  • Impaired vascular integrity
  • Systemic hypercoagulability

Complex, multifactorial influences determine whether a DVT will occur - no single factor is predictive

Presentation

  • Asymptomatic
  • Extremity pain
  • Extremity swelling
  • 5.5% die if untreated - usually due to PE

PE - asymptomatic, SOB, chest pain, tachycardia, anxiety, hemoptysis, sudden death

DVT dx - doppler ultrasound, venogram

PE dx - chest CT angiogram (gold), CXR, lung V/Q scan

D-dimer - high negative predictive value

156


Hypercoagulability

Increased risk of thrombosis due to ongoing stimulus to thrombosis and/or defects in natural anticoagulant/fibrinolytic mechanisms

Acquired or inherited

Acquired examples

  • Surgery/trauma - TF exposure, stasis
  • Immobilization - stasis
  • Malignancy - TF, stasis
  • Pregnancy - stasis, hypercoag, low anticoag, genetic predisposition
  • Estrogen/OCs - hypercoag, low anticoag
  • Antiphospholipid syndrome, autoimmune disease, homocysteinemia - endothelial cell dysfunction

157

Antiphospholipid Syndrome (APLS)

Acquired autoimmune thrombophilic condition

Characteristics

  • Vascular thrombosis and/or recurrent loss of pregnancy
  • Lab evidence of antiphospholipid antibodies

Primary or secondary (often with another autoimmune condition, e.g. SLE)

Lab - prolonged aPTT with positive lupus anticoagulant

  • But do not bleed!
  • Does not correct with mixing (inhibitor)
  • Specific antibodies - anti-cardiolipin IgG or IgM, anti-beta-2-glycoprotein IgG or IgM

Manifestations

  • DVT/PE
  • Arterial thrombosis - stroke/TIA/MI
  • Recurrent fetal loss
  • Thrombocytopenia

158


Pregnancy and thrombosis

6x increased VTE risk during pregnancy

PE - most common cause of maternal death

Risk greatest during 6 week post-partum period

Due to stasis, venous compression (gravid uterus), altered hemostasis (hormonal)

159


OCs and VTE

Poorly understood mechanisms

Changes in coagulation factors as in pregnancy plus activated Protein C increase

160


Acquired Hyperhomocysteinemia

Due to B6 and B12 deficiency

Causes vessel and endothelial cell damage resulting in hypercoagulability

161


Thrombotic Microangiopathy (TMA)

Features

  • Microvascular thrombosis
  • Microangiopathic hemolytic anemia (schistocytes)
    • Non-immune (negative Coombs)
  • Thrombocytopenia

Elevated plasma Hb and LDH, low haptoglobin

Causes renal and neurologic problems

DDx

  • TTP - more neurologic symptoms
  • HUS - more renal symptoms
  • DIC
  • Malignant HTN
  • Vasculitis
  • SLE
  • APLS
  • HIV
  • Renal allograft

162


TTP

Thrombotic Thrombocytopenic Purpura

Systemic microvascular thrombosis characterized by

  • Thrombocytopenia
  • Microangiopathic hemolytic anemia
  • Tissue ischemia/infarct

Acute, severe

Usually fatal without treatment

  • Acquired anti-ADAMTS-13 antibodies - plasma exchange
  • Congenital - plasma infusion

Inhibited/absent ADAMTS-13 --> excess ultra-large vWF multimers

163

HUS

Hemolytic Uremic Syndrome

Clinically resembles TTP, except renal failure predominates

Sproradic (usually adult) - no precipitating factor

Epidemic (usually childhood) - EHEC (O157:H7) - Shiga toxin damages glomeruli

  • Often in meat

164

Inherited thrombophilias

Manifestations

  • 50% occur without provocation
  • DVT of legs and pelvis
  • Increased incidence of superficial thrombophlebitis
  • No association with arterial thrombosis and APC resistance or prothrombin 20210
  • Association with arterial thrombosis and deficient anti-thrombin, Protein C, Protein S

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165

Activated Protein C Resistance (APCR) - Factor V Leiden

APC - natural anti-coagulant - cleaves Va to inactive form (Vi)

  • A checkpoint to reduce thrombin activation
  • Without, Va is constitutively active
  • Relies on presence of Protein S

Factor V Leiden - 95% of APCR - APC present, but Factor V is mutated to resist inactivation (cleavage site mutation)

  • Thrombosis

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166


Excess procoagulant

Prothrombin 20210A - mutation at nucleotide 20210 causes increased prothrombin levels (higher mRNA stability --> more translation)

  • High prevalence in N. Europeans (up to 20% of familial thrombosis)
  • Heterozygous - 3x increased VT risk

Elevated VIII

  • Inherited and acquired forms
  • Often seen in VTE patients

Elevated XI

167

Antithrombin abnormalities

Antithrombin (AT) - naturally occuring anti-coagulant

  • Inactivates mainly thrombin, Xa
  • Heparin increases activity

AT deficiency - unchecked coagulation/hypercoagulability

  • May also be heparin-resistant

High penetrance

Usually seen in young adults

Often present as thrombosis during pregnancy