Kidney transplantation using elderly non-heart-beating donors: a single-center experience.

Although acceptable outcomes have been reported in both non-heart-beating (NHB) and elderly donors individually, the large pool of elderly NHB donors has not yet been fully utilized. In 1994, we expanded our transplant protocol to include NHB donors aged over 65 years and this study compares the clinical outcomes with regular NHB transplantations. Up to June 2005, 24 patients were transplanted at our center with kidneys from NHB donors aged 65 years or more, whereas 176 patients received grafts from conventional NHB donors during the same period. Grafts from older donors were associated with inferior glomerular filtration rates (29 vs. 44 mL/min after 1 year, p=0.01) and graft survival (52% vs. 68% after 5 years, p=0.19) compared to younger NHB donor grafts, although the difference in graft survival was not statistically significant. Exclusion of older NHB donor kidneys with severe vascular pathology resulted in similar graft survival of older and younger NHB donor kidneys. We conclude that the use of elderly NHB donors in order to expand the donor pool was associated with unacceptable clinical outcomes and cannot be justified without further refinement in their selection, for example, by histological assessment of pretransplant biopsies.

Functional versus structural matching: can the CTLp test be replaced by HLA allele typing?

Human leukocyte antigen (HLA) incompatibilities are the most important immunological barriers to bone marrow transplant success when using unrelated donors. Until recently, standards for donor selection included serological methods for HLA class I antigens and DNA-based typing for HLA class II alleles. In our center cytotoxic T-lymphocyte precursor (CTLp) assays have been an integrated part of the search selection procedure as well. More recently, DNA-based typing for HLA class I became available. This allowed us to determine the correlation of CTLp frequencies directed against incompatibilities at the HLA-A, -B, and -C locus in 211 donor-recipient pairs. HLA class I incompatibilities are significantly (p < 0.001) associated with high CTLp frequencies. Exceptions did occur, high CTLp frequencies are seen in 14% of the HLA-A, -B, and -C allele matched pairs, whereas in 7% of the pairs mismatched for HLA-A or -B a low CTLp frequency occurred. The successful outcome of transplants performed in the latter cases suggest that the CTLp test can be used as a tool to detect permissible mismatches when no fully matched donor is available. The influence of HLA-C mismatches on the CTLp outcome was less clear. Although in the majority of mismatched pairs (64%) the CTLp frequency was high, in 36% of the pairs the CTLp frequency was low. Analysis of HLA amino acid sequences was performed on the HLA-C allele mismatched group. An amino acid difference was always found at five polymorphic positions 97, 99, 113, 114, and 116 situated at the peptide binding groove in the high CTLp frequency group, whereas in the low CTLp frequency group this was observed in only 9 of 17 combinations (p < 0.001). However, this is mainly due to Cw*0303-0304 mismatches. In conclusion, although there is a highly significant correlation between the outcome of the CTLp frequency test and HLA allele class I typing, exceptions occur. It is unclear whether they are all clinically relevant but they certainly provide additional insight in allograft recognition.

Risk factors for cytomegalovirus infection and disease in renal transplant recipients: HLA-DR7 and triple therapy.

In a prospective study, an analysis of risk factors for the development of cytomegalovirus (CMV) infection and disease was performed on 77 renal allograft recipients. Twenty-five out of the 77 recipients (32%) had a CMV infection. Twenty-two of the recipients received triple immunosuppressive therapy (cyclosporin A, prednisolone, and azathioprine) while the remaining 55 received standard therapy (cyclosporin A and prednisolone). In 23 recipients (30%) acute rejection was diagnosed and the first positive parameter of infection occurred 22 days after rejection therapy. Infection occurred in 10 out of 18 HLA-DR7-positive recipients (56%) and in 15 out of 59 HLA-DR7-negative recipients (25%; P < 0.02). In multiple regression analysis, HLA-DR7 was found to be a significant predictor of CMV infection (P < 0.005). CMV disease was diagnosed in only 9 out of 25 recipients with an acute infection. Six recipients (67%) with CMV disease received triple therapy for maintenance immunosuppression; this was significantly correlated to CMV disease (P < 0.05) as compared to three recipients (33%) with CMV disease maintained with standard therapy. Our data suggest that HLA-DR7-positive recipients are more susceptible to CMV infection and that CMV disease is associated with triple immunosuppressive therapy.

The role of the spleen in pancreas transplantation.

Early graft thrombosis and rejection of the graft are the two major causes of graft failure in pancreas transplantation. Inclusion of the spleen in the pancreatic graft has been purported as a possible solution to both complications, but severe graft-versus-host disease led to abolishment of this procedure. By irradiating the donor spleen ex vivo during cold storage, we successfully prevented graft-versus-host disease, allowing us to evaluate the advantages of clinical pancreaticosplenic transplantation. This study reports our experience with 12 pancreaticosplenic transplantations. Using Doppler flow measurements, we have been able to examine the hemodynamic advantages. Our results confirm the purported benefit. Vascular resistance indices in the pancreatic graft are significantly lower when the donor spleen is included. This, however, did not lower the incidence of thrombosis (2 out of 12 cases) in our study. Serial radionuclide studies with 99mTc-hexamethyl propylene amine oxime were performed for further evaluation of graft perfusion. With time the spleen uptake diminishes, compatible with atrophy of the organ. This was confirmed histologically. No indication of an immunologic advantage of transplanting the pancreas together with the spleen was found. All patients went through severe rejection crises. A transient reduction in platelet count (55-88%, mean 71%) of preoperative values was observed. This platelet drop is not seen in patients with a pancreas without spleen transplantation. We conclude that in pancreas transplantation, inclusion of the irradiated spleen has no obvious advantages for early graft thrombosis and rejection of the graft.

Prevention of primary HLA class I allo-immunization with leucocyte-poor blood components produced without the use of platelet filters.

In a prospective study the incidence of allo-immunization and platelet refractoriness was investigated using a consequently leucocyte-poor blood product regime. Twenty-five previously non-transfused patients with acute leukaemia (11 men, 7 women) or autologous bone marrow transplantation for Hodgkin's or non-Hodgkin's lymphoma (2 men, 5 women) received at least 80 donor units of filtered red cells (filtration within 24 h after donation, leucocyte content 8.5 +/- 3.9 x 10(6)/U) and/or of platelet concentrate (produced by the buffy coat method, leucocyte content: 7.8 +/- 4.2 x 10(6)/U). A 1-hour recovery of 20% in three consecutive transfusions, in the absence of clinical factors known to impair increment, was defined as platelet refractoriness. HLA class I antibody screening with a panel of 60 cells was performed before the first transfusion and after 80 U of blood components. Of 25 patients who entered this study, 6 patients developed platelet refractoriness after a mean of 38 units of blood components (range 26-45 U); all 6 were female with a history of multiple pregnancies. In 19 patients regarded as non-refractory, no HLA antibodies were demonstrated (13 men, 6 women). This study, though limited in size, suggests that the use of blood products containing less than 1 x 10(7) leucocytes/donor unit prevents primary HLA class I immunization and platelet refractoriness.

The influence of DR match of blood donor and recipient on the formation of T- and B-cell antibodies and on renal allograft outcome.

It has been shown that patients transfused with one unit of blood mismatched for both HLA DR antigens have an increased rate of formation of cytotoxic leucocyte antibodies compared to patients who received blood which differs in only one DR antigen. In the same study it was found that DR sharing of the blood transfusion donor and patient improved results of kidney and heart transplantation. However, the data were mostly collected in a retrospective manner and came from various centres. Furthermore, no information was available on whether these antibodies were directed to B- or T-cells. Therefore, the influence of DR match of recipient and blood donor on the formation of T- and B-cell antibodies as well as on clinical course after kidney transplantation was studied prospectively in patients transplanted in one centre.

The impact of ischemic lesions in the donor kidney, donor age, recipient age and HLA (A, B, C, DR, DQ) matching on clinical course after kidney grafting.

It is generally accepted that HLA matching improves graft survival. However, there is no consensus on whether this improvement is reflected on daily clinical course. Clinical course after renal transplantation depends on many factors, such as donor age, recipient age, ischemic score in the kidney, and HLA matching. The relative contribution of these factors is unknown. Because management of the recipients in the various centers differs considerably, only a single centre study would reveal the relative contribution of all these factors. Therefore, in our centre we studied the influence of these parameters on the clinical course after renal allografting.

Increasing complexity of HLA-DR2 as detected by serology and oligonucleotide typing.

Serological and oligonucleotide typing was performed on a number of HLA-DR2-positive cells from different ethnic origin, including DR2 haplotypes with various DQ associations. Exons 2 of DRB1 and DRB5 of DR2-positive individuals were locus-specific amplified and hybridized with a number of different oligonucleotides capable of discriminating between the various Dw2, Dw12, Dw21, and Dw22 associated sequences. The linkage of DRB with DQA1 and DQB1 in these haplotypes was analyzed. Among the DR2- positive cells we could define 10 different DR DQ haplotypes by serology and 13 by oligonucleotide typing. The DR2.ES specificity is a serological DRw15 variant which could not be discriminated by oligonucleotide typing from a DRw15 DQw5 haplotype. The DR2.JA variant represents a unique DRB1*1602 DRB5*0101 haplotype. The DR1+2s haplotype consists of a DRB1 DQ region from a Dw1 and a DRB5 gene from a Dw2 haplotype. Its short DR2 serum pattern can be explained by the absence of a DR2 DRB1 gene product. DRB5*0101 sequences were found in association with DRB1*1501, *1502, *1602, and *0101 alleles. Since the DRB5 gene is capable of such different associations it is comparable to the DRB3 and DRB4 genes. This may have implications for the definition of the broad DR2 specificity which is predominantly encoded by the DRB5 gene product. New DR2 haplotypes included the following DQ combinations: DQw2-positive DQA1/B1*0301/0201 and DQw6-positive DQA1/B1*0102/0601 and *0102/0603 haplotypes.

Focal glomerulosclerosis in neonatal kidney grafts.

Two adult patients received both kidneys from neonatal donors. After grafting an accelerated growth of the kidneys was observed. Five weeks after transplantation the craniocaudal diameter had increased from 4 and 3.4 to 8.3 and 6.1 cm, respectively. Histological examination of the kidneys showed segmental glomerulosclerosis 24 and 3 months after grafting, respectively.

HLA antigens in idiopathic thrombocytopenic purpura.

The lymphocytes of 52 patients with the clinical diagnosis of idiopathic thrombocytopenic purpura (ITP) were typed for the HLA-A, -B and -C antigens, and of 27 of those patients also for the DR antigens. ITP proved not to be significantly associated with any of the HLA-A, -C or -DR antigens tested for. On the platelets of 35 of these 52 patients autoantibodies were detected in the direct immunofluorescence test. In these 35 patients with autoimmune thrombocytopenia (AITP), an increased frequency of HLA-Bw38 was found (14.3% versus 2.6% in controls). The frequency of none of the HLA antigens was significantly increased in the group of 17 ITP patients without demonstrable autoantibodies. The difference in association with HLA-Bw38 between AITP and ITP without demonstrable autoantibodies was not significant.

B-cell alloantigens in two recombinant families: prediction of a B/D recombination using B-cell-specific alloantisera.

During the VIIth Histocompatibility Workshop, 44 sera were selected that defined B-cell alloantigens showing correlation with the HLA--D antigenic determinants. The segregation of these antigens, now called DRw (HLA--D-related), was studied in a family that contained one HLA--A/B recombinant and one HLA--B/D recombinant sibling. It could be established that the DRw-1 antigenic determinants segregated with the B-D region in the A/B recombinant sibling and that the DRw-2 antigen segregated with the D region of the B/D recombinant child. In the second family, DRw typing segregated with the D region of the B/D recombinant child. In the second family, DRw typing showed that one of the children was identical with an HLA--ABC non-identical sibling. This suggested that a crossing-over had occurred. This was confirmed by mutual non-stimulation of these HLA--ABC non-identical children in the mixed lymphocyte culture. This crossing-over involved the DRw-3 alloantigen. These data confirm the assumption that serologically defined B-cell alloantigens are coded for by genes located outside the HLA--A/B region on the B-D side of chromosome. Moreover, these data suggest that, within families, serological B-cell genotype identity can predict a mutual unresponsiveness in the lymphocyte culture. However, findings in these two families indicate that discrepancies nevertheless exist between B-cell serology and mixed lymphocyte reactions and that mutual stimulation can occur despite apparent identity of serological B-cell phenotype and that B-cell phenotype identity does not necessarily predict mutual unresponsiveness in the mixed lymphocyte culture.