Autoimmune Hemolytic Anemia

What is the difference between primary idiopathic autoimmunity and autoimmunity secondary to disease?

Primary idiopathic = unknown cause

Secondary to disease = some disease or sickness triggers autoantibody production

Drug dependent autoantibodies

Drug binds and lyses RBCs

Dug independent autoantibodies

Drug does not currently have to be in the system to influence autoantibody production

Common drugs that cause autoimmune reactions

Antimicrobials. NSAIDs, Chemo drugs

Common diseases that cause autoimmunity

Epstein-Barr Virus (mono), HIV, Hepatitis, Lupus, Rheumatoid Arthritis, Leukemias, Lymphomas

What is an autoimmune hemolytic anemia?

Immune system antibodies attack RBCs. Depending on if antibodies bind complement or not, it can cause intravascular or extravascular hemolysis

Cold Hemagglutinin Disease antibody + Ab identification

IgM, caused by autoanti-I that binds lower than 37C

What causes CHD?

Primary lymphoproliferative disorder with marrow B cells producing pathogenic autoanti-I (high titre, interrupting in daily life)

Most common way to develop CHD in childhood?

Epstein Barr Virus - often self limiting

Mutational pathway of CHD

Children have i on cells, will start to produce transferase that creates I

• Predisposition to genetic will prohibit transferase production, creating autoanti-I

Difference between pathogenic and non-pathogenuc autoanti-I

Non pathogenic Autoanti-I is polyclonal, low thermal amplitude (does not react at temps closer to 37C)

Pathogenic is monoclonal, high thermal amplitude

Mechanism of CHD

Autoanti-I binds to RBCs when you are in cold environment, allowing for complement to attach to C3b. C3b lines cells, so RBCs are destroyed in spleen, causing intravascular hemolysis. Antibody pops off in warmer temp

Symptoms of CHD

Anemia: Fatigue, SOB, Pallor

Weak hair and nails, chronic headaches

Acrocyanosis (blue nose)

Reynauld’s syndrome = hands/feet pale compared to upper limbs

Severity of CHD depends on…

Thermal amplitude (higher is worse), how much time spent outside in cold

Hematology investigation of CHD

Low Hgb, Diff shows Schistocytes, bite cells, spherocytes, hypochromic cells

Chemistry investigation of CHD

Increased bilirubin, decreased haptoglobin, increased LDH

Unique lab feature of CHD

If slide is made at room temp, you will see agglutination of RBCs on slide

Urinalysis of CHD

Urobilinogen pos, hematuria, Hemoglobinuria

DAT result of CHD

DAT poly pos, C3 pos

Cold agglutinin screen:

Serial dilute patient plasma, add O cells (rich in I antigen) at 4C. Titre of 40 or higher is significant

ABORh discrepancy of CHD

Discrepant reverse group, positive autocontrol. Prewarm to deal with

Testing method to remove cold autoagglutinins to result discrepant ABORh or to allow detection of alloantibodies

Cold autoadsoprtion

Steps of cold autoadsorption

1. Prewarm and wash cells with warm saline (pop autoantibody off)

2. Aliquot patient plasma with patient cells at 4C to bind autoagglutinins

3. Centrifuge and remove plasma

4. Autoadsorbed plasma has no cold autoaggllutinins. Can be used for ABORh or Ab screen. Repeat if necessary

Transfusion lab results for CHD

Antibody Screen positive + Panel

Autocontrol positive

Cord cells neg (cord cells only have i)

DAT: Poly and C3d pos

Cold agglutinin titration steps

1. Prewarm plasma to pop off Abs back into plasma

2. Serial dilute plasma

   1. Test Adult O cells at 4C to determine titre

Treatment of CHD

Rituximab (target CD20 on B cells to destroy)

Lifestyle management (less cold weather)

Plasma exchange in severe cases

How to find compatible blood for CHD patients

Prewarm blood, test with auto adsorbed plasma.. Transfuse with blood warmer

What Antibody causes Paroxysmal cold hemoglobinuria

Biphasic autoanti-P (IgG)

Mechanism of PCH

Autoanti-P adsorbs to P antigen at cold temperatures and begins complement binding. Back at physiological temperature, complement binds to C9, causing intravascular hemolysis

What can PCH be secondary to

EBV, HIV, Hepatitis, Rheumatoid Arthritis, etc.

Unique secondary contributor to PCH

Latent underling Syphillis

Symptoms of CHD

Similar to CHD, fever, chills, lower back pain when cold (kidneys hurt), dark urine

Hematology lab investigation of PCH

Low Hemoglobin, increased schistocytes, reticulocytes

Chemistry lab investigaiton of CHD

Decreased haptoglobin, increased bilirubin, increased LDH, increased free hemoglobin

Urinalysis workup of PCH

Urobilinogen pos, hemoglobinuria, hematuria

DAT result of PCH

Poly pos, C3 pos, IgG can be pos depending on severity

Confirmatory test of PCH

Donath-Landsteiner test

Steps of Donath-Landsteiner test

1. Collect blood, keep warm, allow to clot in red top tube

2. Spin down serum, pour off

3. Incubate serum wirh 3% P pos cells + normal serum at 4C

4. Warm up to 37C

5. Spin and read for hemolysis (if hemolyzed, anti-P present

Theory of donath landsteiner test

Create biphasic environment for PCH to occur in the tube

Why is fresh serum added to Donath Landsteiner test

Act as source if complement if required

Treatment of PCH

Drugs: Rituximab, Aculsimab (anti-complement antibody)

Lifestyle balance: stay warm

Plasma exchange in severe cases

Red cell transfusions for anemia: ensure cells are prewarmed

What causes Warm autoimmune hemolytic anemia

Polyclonal, polyspecific IgG autoimmune antibody reacting at 37C.

Idiopathic or secondary to disease

What antibody type is WAHA

Mainly interacts with Rh antigen

Does WAHA cause intra or extravascular hemolysis

Mainly extravascular - IgG does not fix complement, IgG sensitizes and cells removed in spleen (causing splenomegaly)

Symptoms of WAHA

Splenomegaly, Fatigue, Dyspnea (SOB), discolored urine

Heme lab workup of WAHA

Decreased Hgb, increased reticulocytes, spherocytes

Chemistry workup of WAHA

Increased bilirubin, hemoglobin, decreased haptoglobin, increased LDH

Urinalysis workup of WAHA

Urobilinogen pos, hemoglobinuria, hematuria

Blood bank workup of WAHA

DAT - pos for IgG

ABO Rh normal, but panreacting Ab screen

What can we use to remove WAHA antibodies

Warm autoadsoprtion

Steps of warm autoadsorption

1. Sample with warm autoab eluted with WARM/ZZAP while washing, removing Ab from RBCs

2. DAT neg cells incubated with patient plasma at 37C. Alloantibodies remain in plamsa

3. Spin sample, aliquot plasma. May need to repeat process to adsorb on all autoAb

4. Plasma now ready to Antibody ID or serological crossmatch

Can auto adsorption be performed on recently transfused patients?

No - allo antibodies can be removed from donor cells

How to check if autoadsorption is complete

Check if patient plasma still binds to RBCs, or run the Ab screen

Treatment of WAHA

Corticosteroids - decrease AB production, limit macrophage function

Supplement: iron, folate, B12

Splenectomy (historical)

The types of drug-induced autoimmine hemolytic anemia

Methyldopa hemolytic, Immune Complex, Membrane modification, Drug adsorption

What drug causes a methyldopa hemolytic anemia?

Aldomet - antihypertensive drug used to treat preeclampsia

Methyldopa hemolytic anemia mechanism

Taking drug causes creation of autoantibody against red cell antigen (Not the drug!) Ab is panreacting

Antibody related to dose and time being on drug

What does Methyldopa Hemolyric anemia look like clinically

Looks exactly like WAIHA

• Hematology: Decreased HgB, increased reticulocytes, spherocytes, 

• Urinalysis: Urobilinogen pos, hemoglboin, bood pos

• Chem: Increased Billirubun, decreased haptoglobin, increased LDH

  • DAT pos for IgG (sometimes C3d), panreacting Ab screen, ABORh is normal

Blood bank results of Methyldopa hemolytic anemia

DAT: Pos for IgG (sometimes C3d)

Panreacting AB screen

ABORh normal

Treatment of Methyldopa hemolytic anemia

Stop taking methyldopa. Ab goes away

How to differentiate between Methyldopa hemolytic anemia and WAIHA

Only true difference is by checking drug history

Sometimes Methyldopa DAT can be positive for C3

What drug causes Immune complex drug induced autoimmune hemolytic anemia

Quinidine (anti-arrhythmatic)

Immune complex Autoimmune hemolytic anemia mechanism

Drug creates antibody against the drug, and the drug/antibody complex binds to plamsa protein. Immune complex adsorbs on to RBC membrane and activates complement

DAT results of Immune complex

DAT pos for C3d, neg IgG (as IgG binds to drug, not RBC)

Blood bank results of Immune complex

DAT Pos for C3d

Ab screen negative

Eluate negative

What drug causes membrane modification autoimmune hemolytic anemia

Cephalosporins (broad spectrum antibiotic)

Mechanism of Membranen modification

Drug adsorbs to RBC causing membrane modification. Modificaiton causes non-specific protein binding (IgG, IgM, Complement, IgA, IgE)

Drug makes RBC sticky

DAT result for membrane modification

DAT pos for both igG and C3d (as both are stick on)

Blood bank results of membrane modification

DAT pos for IgG and C3d

Ab screen neg

Eluate is negative (no specific binding to sceen cells)

What drug causes drug adsorption autoimmune hemolytic anemia

IV Peniccilin with massive doses (broad spectrum antimicrobial)

Drug adsorption mechanism

Drug binds to RBCs, immune system sees Penicillin on RBC as foreign, creates Anti-Penicillin antibody

DAT result for drug adsorption

DAT pos for IgG only

Blood bank results of drug adsorption

DAT IgG pos

AB screen negatve

Eluate negative