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Transfusion Associated Graft vs. Host Disease

12 Jul

Mismatched organ transplantation results in rejection that is mediated by T lymphocytes. What would happen if the T-lymphocytes cells were transplanted into an HLA-incompatible host? If the transplanted T-lymphocytes survive they are capable mounting an immune response against the host tissue. This is known as graft vs. host disease (GVHD) and is characterized by widespread tissue damage. Immune competent cells are transplanted under two circumstances, when cellular blood products are transfused and in patients receiving allogeneic stem cells. GVHD associated with transfusions (TA-GVHD) is a rare but almost fatal disease.

Incidence

The exact incidence of TA-GVHD is unknown but the disease is very rare. The annual incidence has been difficult to estimate but probably is in the low double figure. Irradiation of blood products and discontinuation of the practice of directed donation has reduced the incidence to almost zero indicating the preventable nature of disease. It is important to be aware about the disease as the number of predisposed patients is on the rise.

Products Causing TA-GVHD

All blood products stored unfrozen can cause GVHD. Fresh plasma has caused TA-GVHD. The process of freezing destroys T-lymphocytes. Fresh Frozen plasma and thawed erythrocytes are not associated with TA-GVHD.

Pathogenesis

GVHD can occur after transfusion as little as 4 X 103 T-lymphocytes/kg. The average number of leucocytes cellular blood products is as follows; red blood cells 2 X 109, random donor platelets 4 X 107 and apheresis platelets 1 X 108. Patients receiving transfusion of a cellular product are exposed to a dose of T lymphocytes capable of causing GVHD. The risk of TA-GVHD appears to be higher with the use of fresh blood. It is presumed that the number of lymphocytes and the number decreases with storage. The ability to eliminate HLA-incompatible T lymphocytes protects patients from development of TA-GVHD. The immunological mechanisms to eliminate non-HLA incompatible T lymphocytes must be exceedingly efficient because though almost every patient receiving blood products is exposed to a dose of lymphocytes capable of causing TA-GVHD, the disease remains rare even if precautions are not taken. TA-GVHD occurs when the host’s ability to eliminate transfused T lymphocytes is compromised. This may happen when there is T cell dysfunction and when the transfused lymphocytes evade the immune surveillance.

T cell dysfunction may be under the following circumstances:

  1. Severe congenital T cell immunodeficiencies immunodeficiencies
  2. Neonates and infants: Most cases of TA-GVHD have been reported in neonates who have undergone intrauterine transfusion. Infants who have undergone exchange transfusion are also at risk. In both these conditions the allogeneic cells have been shown to persist (6-8 weeks in case of exchange transfusion and 2-4 years in case of in utero transfusion). Maternal lymphocytes regularly cross over into foetal circulation but the foetus can eliminate these lymphocytes. In mixed lymphocyte cultures of foetal and maternal cells, foetal lymphocytes dominate. The protection is specific to the maternal lymphocytes and not other adult lymphocytes making the neonates prone to TA-GVHD
  3. Lymphomas: Virtually all the cases of TA-GVHD associated with lymphomas have been associated with Hodgkin’s lymphoma. The stage or treatment modality does not change the risk. Hodgkin’s lymphoma, unlike non-Hodgkin lymphoma, is associated with T cell dysfunction.
  4. Therapy induced T cell dysfunction: Purine analogues: Cladribine, deoxycoformycin and fludarabine cause a profound long lasting depletion of CD4 positive lymphocytes. The effects of bendamustine, clofarabine are less clear. Alemtuzumab and antithymocyte globulin (horse and rabbit) appear to carry a risk of TA-GVHD.

The most common cause of profound T cell deficiency, AIDS is for some reasons not associated with TA-GVHD.

Homology between donor and recipient HLAs appears to allow the transfused T lymphocyte to escape immunesurveillance and initiate TA-GVHD. HLA homology may be found in close blood relatives and in communities that are relatively homogenous for HLA. Transfusions from a family member increase the risk of TA-GVHD as patents are likely to share HLA epitopes with the host. The incidence of TA-GVHD in Japan, which has limited HAL-hetrogenity, is about 10-20 times higher Caucasians.

Manifestations

The median period between transfusion and onset of symptoms is 10 days but the may be as long as 4 weeks. Fever is usually the first manifestation and is soon followed, maculopapular skin rash, diarrhea and hepatitis. Pancytopenia differentiates TA-GVHD from GVHD following allogeneic stem cell transplant. Pancytopenia is seen in virtually every patient with TA-GVHD but is rare in GVHD associated with allogeneic stem cell transplant. The aim of allogeneic stem cell transplant is haematological reconstitution either for a failing marrow or following a high dose of chemotherapy. The transfused T-lymphocytes and the haemopoietic stem cells are HLA-identical. This spares the haemopoietic stem cells from attack by T lymphocytes and pancytopenia is rare. Unlike patients of GVHD, the haemopoietic stem cells ofpatients with TA-GVHD have an HLA type distinct from the transfused lymphocytes. This makes them susceptible to T-lymphocyte mediated cytotoxicity resulting in pancytopenia.

Diagnosis

TA-GVHD is diagnosed on clinical grounds. Donor lymphocytes may be detected in the blood.

Treatment

TA-GVHD has a very high mortality. The principles of treatment are the same as those of allogeneic stem cell transplant induced GVHD. Immunosuppressive therapies including high-dose steroids, azathioprine, methotrexate, cyclosporine and intravenous immunoglobulin have been used. Spontaneous remissions have been reported. The few successful cases included haploidentical transplant from the mother in an infant of X-linked severe combined immunodeficiency, infusion of patient lymphocytes and the use of anti-CD3 therapy with cyclosporine.

Prevention

TA-GVHS is a preventive disease. Most of the cases reported in the recent times have been in places where anti-GVHD measures are not strictly implemented.

Donor screening: Patients should not receive blood from first or second-degree blood relatives. If this is not avoidable then the blood should be irradiated.

γ-Irradiation: γ-Irradiation with 25Gy kills lymphocytes without affecting platelets or granulocytes. It does however shorten the shelf life of blood. The guidelines for irradiation vary from country to country. These take into account the risk of TA-GVHD, available infrastructure and the cost of irradiation. In Japan, the risk of TA-GVHD is higher and the guidelines more stringent. Generally speaking irradiation is recommended under the following circumstances:

  1. Transfusion from blood relatives
  2. Transfusions to patients with severe congenital T cell deficiencies
  3. Intrauterine transfusion
  4. Neonatal transfusions
  5. Transfusions in cardiac surgery
  6. Transfusions in patients who have received Cladribine, Deoxycoformycin, fludarabine and alemtuzumab
  7. Granulocyte transfusions
  8. Patients of allogeneic stem cell transfusion

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Complications of Blood and Blood Product Transfusion

29 May

IMMUNE COMPLICATION
Haemolytic transfusion reactions: Haemolysis following red cell transfusion occur in two forms. Acute haemolysis occurs within 24 hours, is more serious and usually intravascular. Delayed haemolysis occurs after the first 24 hours is more common, less serous and usually intravascular.
Febrile reaction: A rise of temperature usually with chills by more the 1°C after blood transfusion. Headache nausea and vomiting may be seen in serve reaction. Fever lasting more the 18 hours after transfusion are not likely to be febrile reactions. They are almost always seen with cellular components being most common with platelet transfusion.
Urticarial Reactions complicate about 1% of blood trasnfusions
Anaphylactic reactions occur in about 1:150,000 patinets. They are seen in patients with IgA deficiency. These patients have anti IgA antibodies that react to IgA in the transfused plasma. These patients must be transfused thoroughly washed  RBCs.
Transfusion Induced Acute Lung Injury manifests as sudden deterioration of lung function shortly (2-6 hours) after a blood transfusion. It is treated by respiratory support with oxygenation and positive pressure ventillation.
Graft vs. Host disease is a rare but almost uniformly fatal complication of blood transfusion, usually from a close relative, that manifests as post transfusion fever, skin, liver,  and gastrointestinal manifestations and  pancytopenia.
Immune suppression caused by blood transfusion has been used in the past for decrease renal graft rejection. Today clinical relevance this effect is not clear.
INFECTIONS
Hepatitis (B and C): The incidence of transfusion induced hepatitis has called because of testing of blood and blood products. 
HIV: HIV posed a major risk before testing became mandatory. Testing has substantially reduced but not eliminated the risk of HIV transmission.
Other Viruses: Cytomegalovirus, Epstein-Barre virus, parvovirus b19, HTLV I, HTLV II can be transmitted by blood transfusion. Though some countries mandate testing for some of these viruses the practice, unlike that for HBV, HCV and HIV is not universal.
Malaria and other parasitic diseases: Out of the parasitic diseases that can be transmitted by blood transfusion (malaria, filariasis, bebesiosis, toxiplasmosis, toxoplasmosis and trypanosomiasis (South American, African). Malaria because of it’s widespread distribution is the greatest concern. because of it’s wide spread distribution.
Transfusion induced sepsis: Transfusion induced sepsis occurs as a result on bacterial growth during storage. It is most common with platelet transfusion as platelets are the only component stored at room temperature.
MASSIVE TRANSFUSION
Coagulopathy: Dilutional coagulopathy due to degradation of labile coagulation factors like V and VIII on storage causes coagulopathy.
Citrate toxicity: Citrate used as an anticoagulant binds calcium and causes hypocalcaemia. Clinically significant hypocalcaemia is seen only with very rapid transfusion raters (more than one unit over less than 5 mins) and in patients with liver disease (because of impaired citrate metabolism). Treated with intravenous calcium.
Hypothermia: Blood is refrigerated for storage. In massive transfusion the urgency and the rapid rate of infusion may result in hypothermia that can be prevented by the use of blood warmers.
Acid-base imbalance: Patients needing massive transfusion are likely to be acidotic due to lactic acidosis. Citrate present in the transfused blood may aggregate acidosis. On recovery citrate and lactate are converted to bicarbonate resulting in metabolic alkalosis the commonest
Hyperkalaemia: Stored blood may have unto 80mEq/L of potassium which on transfusion cause lifethreatening hyperkalaemia

Further Reading:
Vein to Vein: An online publication of the Canadian Blood Services

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The ABH Blood Group System

17 Apr

The risk of transfusion reactions prevented widespread use of blood transfusion till the turn of the 20th centaury. In 1900 Karl Landsteiner while mixing red cells and serum from his colleagues discovered that the serum of some individuals reacted against the red cells of other individuals. Based on reactions between sera and red cells he described three blood groups, A, B and C (which was later renamed as O). The fourth group of the ABO system, AB, was described a year later. In 1907 Ruben Ottenberg and Schultz suggested that before transfusion both the donor and the recipient be grouped and the blood mixed in the laboratory to ensure compatibility. Using this process Ottenberg virtually eliminated transfusion reaction.  Landsterner and Weiner discovered the Rh Group in 1940. The use of blood grouping (ABO and Rh) and cross-matching has made blood transfusion safe.

Figure 1. Biochemistry of ABO Groups

The ABO antigens (figure 1)
Antigens of the ABO system are polysaccharides. They are produced by the interaction of genes at two loci, 19q13.2 and 9q34.1 in two steps. The first step involves addition to a precursor of fucose in a α1-2 linkage to a precursor by the action of FUT1 a fucosyltransferase synthesized by the gene encoding for the H antigen located at 19q13.2. The second step is addition of N-acetylgalactosamine or galactose by glycosyltransferases encoded by the genes for A and B group antigens respectively. These genes are located at 9q34.1. The phenotypes resulting from the action of the two genes are listed in table 1.

A and B are codominant and O is recessive. O group is encoded by a gene carrying a mutation resulting in a product that is neither capable of adding GalNAc nor Gal. The commonest O allele has a frame shift mutation resulting in the formation of a stop codon resulting in a premature termination of translation

About 200 alleles ABO genes have been described. Qualitative and quantitative differences between glycosyltransferases encoded by these gene produces a spectrum of antigenicities on the red cell.

Figure 2. Inheritance of ABO and Bombay phenotype


  1. Quantitative Differences in ABH Genes: Quantative differences exist in both A and B group antigens. A1, A2, A3, Ax, Am, Ael are subtypes of A with progressively decreasing Aantigen sites. It has been estimates that A1, which is the commonest A subtype has 800,000 antigenic sites per erythrocyte, A2 has 250,000 sites and Am has only 800 sites. B1 is the most common type of B antigen. Weaker B antigens include B3, Bx, Bel. Individuals with a weak type of antigen can produce antibodies against a strong subtype of the same antigen. Patients with A2 can produce antibody against A1.
  2. Qualitative Differences in ABH Genes
    1. The Cis-AB Group: A rare and interesting type of blood group is cis–AB. Usually individuals with AB have a gene for glycosyltransferase for A group on one chromosome and that for B on the other chromosome. They inherit one gene from each parent. Cases of AB children born to AB group mothers and O group fathers can only be explained if both the A glycsyltransferase and B glucosyltrasnferase activity is carried by the samegene. This is known as cis-AB. Apart from raising paternity disputes cis-AB can also complicate transfusion. The common types of cis-AB express a weak B (B3) and can have anti-B antibodies making only washed O or A cells suitable for transfusion to these patients (Br J Anaesthe 1999; 83:491-2).
    2. The Bombay Phenotype (figure 1 and 2): Blood grouping involves mixing patients cells with anti-A and anti-B sera. Group A cells agglutinate with anti-A, group B with anti-B and group AB with both. If no agglutination is seen the blood group is designated as O. The H antigen is a precursor to A and B antigens.  Patients who have blood group O have inactive glycoryltransferases and nothing is added to H. H is the group O antigen.  Rarely the gene for FUT1 mutated (h) resulting in an inactive product. These individuals do not synthesize H antigen and will not synthesize A or B antigens even if they have genes for the respective glycoryltransferases on chromosome 9q34.1. Their red cells will neither agglutinate with anti-A nor with anti-B sera and will be grouped blood group O. As they lack H they have agglutinating naturally occurring anti-H antibodies. When serum from these patients is mixed with cells from O group individuals the anti-H antibodies in their serum agglutinate O group cells. These individuals are group O on forward matching but not on cross-matching. This is known as the Bombay phenotype. Such individuals can only be transfused red cells from individuals of Bombay phenotype.

Synthesis of Universal Blood Group
Groups A and B are synthesized from H antigen. Bacterial glycosidases can be used to lyse GalNAc and Gal convert A and B group antigens to group O. O blood cells can be given to any individual (except Bombay phenotype). This technology when available for clinical use has the potential to eliminate blood shortages.

Table 1 ABO antigens and antibodies

Antibodies to ABO Antigens (table 1)
The ABO antigens are polysaccharide antigens capable of stimulating T independent B cell responses. Unlike other blood group antigens like the Rh, the serum contains naturally occurring antibodies to the antigens of the ABO and other polysaccharide (Le, Ii and P) blood group systems that are believed to be produced following exposure to environmental antigens like bacteria (J Clin Invest. 1969 July; 48(7): 1280–1291). The antibodies belong to the IgM class and are agglutinating, bind C3. They can cause haemolytic disease of the newborn but the presentations are uncommon and mild.

Physiological Role, Variations and Disease AssociationsThe physiological role of ABO antigens in not known. A relationship between falciparum malaria and ABO antigens has been proposed. Malaria infected cells express a protein PfEMP-1 a protein that can bind to A (and B) on the endothelium. Plasmodium infected cells are less likely to adhere to endothelium of group O individuals reducing the severity of malaria in these individuals (Blood 2007; 110:2250-2258).

The levels of von Willebrand Factor and factor VIII vary with blood group. They are highest in blood group AB and lowest in blood group O. Group O individuals are at risk of bleeding and AB at the risk of thrombosis. About 30% of the variation in vWF can be explained by ABO blood groups. vWF expresses A and/or B antigens and these may be responsible for conferring vWF resistance to degradation by ADAMTS13 resulting in higher vWF levels.

Group O individuals have a lower risk of squamous and basal cell cancer of the skin, pancreatic cancer and gastric cancer. Individuals expressing B antigen are at a higher risk of ovarian cancer and group A individuals are at a higher risk of gastric cancer. Peptic ulcer is more common in O group individuals.

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