Question 26
A patient is being assessed for transplant. He has a panel-reactive antibody (PRA) of 30% by the complement-dependent cytotoxicity (CDC) technique but 0% for Class I and II antigens by flow cytometry (Luminex).
How can you explain the difference between these tests?
Panel reactive antibody refers to the percentage of a panel of antigens representative of the diversity in a given population to which a recipient has antibodies. The goal of the test is to identify anti-HLA antibodies, which can cause hyperacute or acute antibody-mediated rejection. There are different techniques by which these antibodies can be detected.
The complement-dependent cytotoxicity (CDC) assay uses lymphocytes from a variety of individuals. The recipient’s serum and complement is mixed together with the lymphocytes. If the recipient’s serum contains antibodies against the lymphocytes of a particular sample, then those lymphocytes will be killed. For example, if the cells in 20 out of 50 samples are killed, then the PRA is 40%. Advantages of the CDC test are its simplicity, and its ability to detect antibodies that activate complement, which would likely be injurious to graft. Disadvantages include its lower sensitivity to detect antibodies; lack of specificity to identify which anti-HLA antibodies are present; and inability to distinguish between HLA and non-HLA (non-pathogenic) antibodies, IgG and IgM (non-pathogenic) antibodies, or between class I and II anti-HLA antibodies.
The other method to detect antibody is by flow cytometry. Recipient serum is mixed with beads coated with HLA antigens. Anti-HLA antibodies will bind only to beads that contain the antigen to which they have specificity. A fluorescent-conjugated anti-human globulin is then added and binds to the anti-HLA antibodies bound to the beads. The beads are then passed through a Coulter counter, which measures the degree of fluorescence as a measure of the amount of antibody present. This technique is more sensitive than the CDC test; does not require complement activation; will not give false-positive results with non-HLA or IgM antibodies; can distinguish between class I and II anti-HLA antibodies; and can allow for very precise identification of specific antibodies.
In this case, the cause of the 30% PRA by CDC is either non-HLA antibodies or anti-HLA IgM antibodies, neither of which would be detected by flow cytometry, and neither of which are believed to be of clinical significance.
Question 27
A patient is brought in for a deceased-donor kidney transplant. Her PRA is Class I 45% and class II 60% by flow cytometry. Both her T and B cell crossmatch by the CDC technique are negative.
Should she receive this transplant? What is the difference between PRA and a crossmatch? What if you receive a report from the lab that while the crossmatch is negative, “DSA is present?”
The PRA gives the probability that a recipient will have anti-HLA antibodies against the variety of HLA antigens in a selected population. It is determined as part of the pre-transplant assessment when there is no particular donor identified. In contrast, the crossmatch test answers the question of whether a given recipient has anti-HLA antibodies against a particular donor, so-called “donor-specific antibodies” (DSA). A positive T-cell crossmatch, especially by the CDC technique, is associated with a very high risk of hyperacute rejection.
If the crossmatch test is negative, then this patient likely does not have high levels of DSA. Note that the crossmatch test can be done by both complement-dependent cytotoxicity (CDC) and the more sensitive flow cytometry method. This method may detect low levels of DSA that do not cause a positive CDC crossmatch.
Most centers will not transplant deceased-donor organs into a patient with a positive CDC crossmatch. The approach to a positive flow crossmatch or DSA alone varies. Some transplant centers will avoid transplantation while others may use more intensive immunosuppression at the time of transplant. In the case of a living donor, “desensitization protocols” may be used pre-transplant when the crossmatch is positive or DSA is present.
PRA and cross-match do not relevantly affect outcome after liver transplantation and are therefore not routinely performed pre-liver transplant.
Question 28
A kidney/pancreas transplant recipient presents with diarrhea six months post-transplant. Her medications include tacrolimus, MMF, prednisone, Septra, omeprazole, valganciclovir, amlodipine and atorvastatin.
What are the common causes of diarrhea post-transplant and what investigations should be done?
Diarrhea is common in transplant recipients. Common causes include medications, such as mycophenolic acid or sirolimus; the use of antibiotics; post-transplant infections, such as Clostridium Difficile or cytomegalovirus (CMV); and pre-transplant co-morbidities such as inflammatory bowel disease or diabetic gastroparesis.
Appropriate investigations include a complete medication review to identify responsible drugs; stool cultures for routine bacterial infections, C. Diff., ova and parasites; testing for CMV; and endoscopy in patients where the above investigations are negative. A decrease or switch in the immunosuppressive agents mentioned above may be required if no other causes for the diarrhea are found.
Question 29
A lung transplant patient presents with pancytopenia post-transplant. His medications include cyclosporine, azathioprine, prednisone, Septra, valganciclovir, calcium, vitamin D and omeprazole.
What is the differential diagnosis?
Hematologic abnormalities such as anemia, thrombocytopenia and leukopenia are common post-transplant.
Lymphocyte-depleting antibodies, such as thymoglobulin and alemtuzumab may cause leucopenia, lymphopenia and thrombocytopenia. Bone marrow suppression can also result from the antiproliferative agents azathioprine, mycophenolic acid and sirolimus. Other medications that can cause bone marrow suppression include valganciclovir, trimethoprim-sulfamethoxazole, dapsone, and rarely proton pump inhibitors. ACE-inhibitors and angiotensin-II receptor blockers have also been implicated as causes of anemia post-transplant. Rarely, calcineurin inhibitors or sirolimus can cause thrombotic microangiopathy, which may be associated with anemia or thrombocytopenia.
Notable infections include cytomegalovirus and Epstein-Barr virus, which can cause leukopenia; and parvovirus B19, which can cause a severe anemia. In the context of sepsis, bacterial infections may also cause anemia.
Malignancy can also lead to pancytopenia, particularly post-transplant lymphoproliferative disease (PTLD).
Very rare causes include passenger lymphocyte syndrome, infection-induced hemophagocytic syndrome and graft-versus-host disease in a solid-organ transplant recipient.
Question 30
Which cancers are increased in risk post-transplant? What are the risk factors for cancer post-transplant, and what are the recommendations for cancer screening post-transplant?
Transplant recipients are at greater risk of developing cancer compared to the general population. This is particularly true for cancers associated with viral infections (e.g. EBV-associated lymphomas, cervical cancer associated with HPV, and Kaposi’s sarcoma associated with HHV-8). The risk also varies by organ transplanted. For example, cancers associated with chronic kidney disease and end-stage renal disease (e.g. myeloma and renal cell carcinoma) are seen more commonly in kidney transplant recipients, while the incidence of lung cancer is highest in lung transplant recipients compared to other organs. The relative risk for other common cancers, such as breast, colon, thyroid and prostate is either similar to or only moderately increased compared to the general population.
There is good reason to believe that the value of cancer screening tests, harm from interventions and the life-years to be gained by early intervention may be substantially different in transplant recipients compared to that in the general population. Hence, careful individual appraisal needs to be exercised when making recommendations for screening post-transplant.
Finally, it should be noted that the most common type of cancer post-transplant is squamous-cell skin cancer. Because of this, all transplant recipients are advised to use sun protection year-round, and either perform regular self-checks for new skin lesions or have regular examinations by their family doctor or dermatologist.
