Normochromic Anemias

Spleen function

• Storage: 1/3 of body’s platelets and granulocytes (by mass)

• Filtration: Checks RBCs and culls abnormal cells or pits inclusions

• Promotes phagocytosis of capsule bacteria

• Fetal hematopoiesis (with liver)

Hereditary Spherocytosis

Membrane shaved off

• RBC becomes spherical and more compact

• Shortened life span

• Mild to moderate hemolysis

HS gene + population

In 1:2000 Caucasians

• 75% autosomal dominant, 25% autosomal recessive + de novo mutation

HS is a deficiency of…

• Ankyrin

• Band 3

• Band 4.2

This affects the elasticity and flexibility of the RBC

HS RBC elasticity and environment

• Able to stretch 117% normally

• HS only stretches 3% until Lysis

Spleen environment dangerous for spherocytes

• Low pH, low ATP, low glucose

• Splenectomy helps with RBC survival

Issues with membrane diffusion

• Accumulates Na at higher rate in spleen → swell and lyse

Spherocytes indices + smear

• Happens in peripheral circulation, doesn’t exist in bone marrow

• Spherocytes present, lacking central pallor

• Polychromatic macrocytes

• Reticulocytes: 3-10% (normal: 0.5-2.5)

• MCHC: >36% in 50% of patients

  • Moderate anemia

• RDW slightly elevated

• Increased Bilirubin

  • Due to continued hemolysis

• Cholelithiasis (gall stones) in adolescents/ young adults

Spherocytosis mechanisms

1. Thermal injury: denatures RBC membrane proteins and lipid bilayer. Forms vesicles without losing cytoplasm (microsphero)

2. Intrinsic abnormalities: deficiency of spectrin, ankyrin, band 3 (HS)

3. Immune hemolysis: Antibodies attach to RBC membrane, triggering binding of macrophages (microsphero)

HS lab diagnosis

Screening test:

• MCHC, RDW, Spherocytes, bilirubin

Osmotic fragility test (preferred):

• RBC subject to serial dilutions of isotonic water (0.85% NaCl → 0.0%)

• At 0.85% NaCl, RBC has little hemolysis

• Lysis starts when solution becomes more hypotonic (more diluted)

• Data reports level of complete hemolysis

  • Normal: 0.45%,

  • HS Spherocyte: 0.65%

Hereditary Elliptocytosis gene + population

Defective/ deficient Spectrin

Decreased thermal stability

ALL ethnic groups

Subtypes:

1. Common Hereditary Elliptocytosis

2. Southeast Asian Ovalocytosis

3. Spherocytic Hereditary elliptocytosis

4. Hereditary Pyropoikilocytosis

1. Common Hereditary elliptocytosis

1. Mild Common HE: 30-100% RBC are elliptic. Most have no clinical S/S

2. More severe Common HE. Infantile pyropoikilocytosis. Fragmented, bizarre RBCs since birth. Mild hemolysis, jaundice. Converts to mild HE as they age

2. Southeast Asian Ovalocytosis (SAO) peripheral

Due to Band 3 defect

• RBCs spoon-shaped. may have 1-2 slit-like pallor

• Shape may protect from malaria

• Strongly heat-resistant and rigid. Maintains shape under temperature that would normally cause it to burst

Spherocytic hereditary elliptocytosis (SHE)

Cross between elliptocyte & spherocyte

More common in Northern Europe

Gallbladder disease common, splenectomy indicated

Spherocytic hereditary elliptocytosis (SHE) peripheral

Mild hemolysis, increased osmotic fragility

Spherocytes and oval elliptocyte

Hereditary Pyropoikilocytosis

• α and β Spectrin have reduced assembly

• Mutant Spectrin more easily degradable

• More common in African Americans

Hereditary Pyropoikilocytosis (HP) Indices/ Peripheral

• Hgb: 6.5 g/dL

• RBC budding

• MCV 50-75 fL

• Rare elliptocytes, spherocytes

• Sensitivity to heat

  • Fragments at 37-46 °C instead of normal 49 °C

• Severe hemolysis, poor growth, facial abnormalities

Hereditary Stomatocytosis/ Xerocytosis

Deficiency in Stomatin→ in Na/K permeability

• Regulates ions across RBC channels

Overhydrated stomatocytosis:

• Net gain of intracellular Na+→ water follows

Dehydrated stomatocytosis/ xerocytosis:

• Net loss of intracellular K+→ water moves out

Hereditary Stomatocytosis indices/ smear

• 10-30% are stomatocytes

• Elevated MCV

• Decreased MCHC

• Slit-like central pallor

• Splenectomy can help

Hereditary Xerocytosis

Hereditary autosomal dominant

Unknown etiology/ cause

Hereditary Xerocytosis indices/ smear

• mild anemia

• decreased osmotic fragility

• increased surface:volume ratio

• high MCHC

• Loss of K+→ dehydrated RBC

  • Causes xerocytes: RBC where hgb puddled at the end

Glucose 6 Phosphate dehydrogenase deficiency (G6PD) pathway

G6PD keeps glutathione in the reduced state by converting NAD to NAD

• Deficiency can’t restore NADPH and GSH which causes oxidative stress

Usually apparent after oxidative chemicals or drugs, or have infection

G6PD genetics

Most common human enzyme deficiency in the world

• >400m people affected

• X linked recessive disorder

  • Homozygous females have the disease. Heterozygous are carriers

• >400 variants of the G6PD gene from AA substitution

• 5 genotypes:

  • 2 normal, 3 abnormal. Hemolysis vary

• More common in Mediterranean, Africa, China

  • malarial protection due to oxidative stress

G6PD indices/ smear

• In heterozygous women, two populations are present: normal and G6PD cells

• Polychromasia

• Heinz bodies (supravital stain) from denatured/ precipitated Hgb after RBC exposed to oxidizing agents

• Bite cells present due to spleen pitting them

Favism

Ppl with G6PD can develop this after consuming Fava or Broad beans

• Due to chemicals isouramil, divicine

• Triggers hemolytic episodes on 25% of G6PD patients

• Become irritable, lethargic, nausea, fever, ab pain

• Hemoglobinuria after 48 hours

• Normocytic, normochromic, polychromasia

• Decreased haptoglobin, increased bilirubin

  • Indicates RBC breakdown

Neonatal Jaundice

G6PD related. Occurs within 2-3 days after birth

• high bilirubin (kernicterus)

  • can cause neurological complications

• High sensitivity to vitamin K

  • Substitute triple dye used to treat umbilical cords and camphorated powder.

• Treatments

  • Phototherapy, exchange transfusion (pt blood replaced with donated blood to clear out waste/ toxins)

Congenital Non-Spherocytic Hemolytic anemias

• History of neonatal jaundice w/ gallstones, enlarged spleen. Extravascular hemolysis with hyperbilirubinemia

  • Splenectomy may help

• Varies from mild to severe transfusion dependent

Diagnosis of G6PD deficiency

Testing must be done in time. If testing is during an acute hemolytic episode, reticulocyte production increased

• Testing should be done after hemolytic episode resolved and counts return to normal

G6PD enzyme assay of older RBCs

Fluorescent Spot Test- routinely

• Measures NADP and NADPH reduction

Pyruvate Kinase Deficiency + population + indices

Rare enzyme disorder of Embden Meyerhof pathway

• Lack of Pyruvate Kinase= unable to convert ADP to ATP

• Build up of 2,3 DPG

• Rigid, inflexible cells. Sequestered by spleen and hemolyzed

• Occurs in Northern Europe and close Amish populations in Pens

• HCT 18-36%

• Polychromasia

• Few Nucleated RBC

Aplastic Anemia: definition + mechanism

Hypo-proliferative disorder

• cellular depletion

• Pancytopenia: reduced production of all blood cells

Mechanisms:

1. Auto-antibody against antigen on stem cells

2. Immune mechanisms where T lymphocytes suppress stem cell proliferation

Aplastic Anemia: predispositions

These can predispose someone to an aplastic episode

1. Radiation

2. Chemotherapy/ chemicals

3. Benzene exposure

4. Epstein-Barr virus, HepB, HepC

Aplastic anemia: indices + smear + characteristics

• BM has decreased cellularity, pancytopenia, reticulopenia

• Slow progress with S/S reflecting pancytopenia

• When RBCs significantly depleted:

  • Fatigue, heart palpitations, dyspnea (shortness of breath)

• When platelets depleted:

  • Ecchymosis (skin discoloration from blood beneath)

  • Mucosa bleeds

• Lymphocytosis (increased lymphs)

• Normocytic, normochromic anemia

  • Managed by transfusions and steroids

  • Need marrow transplant in severe cases

Fanconi’s Anemia

Autosomal recessive

• Affects physical characteristics and BM development

• Chromosomal abnormalities, defective DNA repair, chromosomal breaks

Fanconi’s Anemia indices + smear + appearance

• BM has macrocytosis, low platelets, low leukocytes, before RBC depletion

• Increased HgbF

• Appearance:

  • Short stature, hyperpigmentation in trunk and neck, microcephaly, broad nose, structural kidney abnormalities

• Lifespan: 16 years

• Treatment: BM transplant

Diamond-Blackfan Anemia + indices + characteristics

• BM lacks erythroids→ pure red cell aplasia

• Leukocytes are mildly decreased

• Hgb 7 g/dL

• Congenital hypoplastic disorder since early infancy

• 80% severely anemic by 6 months

• Abnormalities:

  • Short stature, low birth weight, head and facial abnormalities

Paroxysmal Nocturnal Hemoglobinuria (PNH) genes

Acquired Stem cell disease where RBC membrane abnormality causes Complement-mediated hemolysis

• C proteins attach to cells to mark them for destruction via Abs

Clonal membrane defects (CD55, CD59 markers, Ags, flags):

• Marked cell destruction overnight→ hemoglobinuria

Somatic X linked mutation:

• 9 surface proteins missing→ no protection against Complement lysis

  • Includes: DAF, MIRL, MAC, GPI

PNH lab findings

• The 9 surface proteins also missing from platelets and WBCs

• Intravascular lysis

• Hemoglobinurea: from black tea to tar

• Increased reticulocytes

• Thrombosis: in brain, dermal vessels, liver, abdomen

PNH testing and treatment

Tests:

• Hams Test: rupture in acid solution (activates complement system)

• Sugar water test: hemolyzes in sugar solution

• Flow Cytometry to detect anchoring proteins of the CD55 and CD59 markers

Cold Agglutinin Syndrome (CAS/ CHAD) cause

• IgM-mediated hemolysis via activating complement

• From 0°C - 5°C: Complement attaches to RBC. As temp increases, RBCs agglutinate and hemolyze

CAS/CHAD S/S + sample prep

• Rarely seen, usually in ages 50+

• Acrocyanosis: blue color of fingertips and toes

• Weakness, pallor, weight loss, intravascular hemolysis

• Sample should be warmed 37°C for 15 to 30 minutes

• Pt should move to warmer weather

Paroxysmal Cold Hemoglobinurea (PCH) cause + S/S

• Complement-fixing IgG (D-L antibody) attaches to RBC P antigen at low temps and activates complement system at warmer temps

  • IgG anti P

• Lysis happens at body temp

• Severe intravascular hemolysis

• Hemoglobinemia, hemoglobinurea

• Increased bilirubin

• S/S:

  • Similar to hemolytic transfusion reaction: back pain, fever, chills, abdominal pain