Role of human leukocyte antigen, donor-specific antibodies, and their impact in renal transplantation.

INTRODUCTION: The clinical significance of the presence of antibody against human leukocyte antigen (HLAb) and donor-specific antibodies (DSAb) prior to renal transplantation remains unclear. This study was done to assess the impact of HLAb and DSAb on graft function, rejection episodes, and graft survival in renal transplantation. METHODS: The Luminex (Luminex, Austin, Texas, United States) is a solid-phase assay using micro-spheres and it is more sensitive at detecting human leukocyte antigen (HLA) antibodies than conventional tests. This retrospective analysis involved 141 consecutive renal transplant recipients between May 2007 and 2009 and with a minimum of 2 years of follow-up. RESULTS: Luminex was positive for HLA class I in 35 and negative in 106; similarly class II positivity was noted in 23 and negative in 118. The DSAb were positive in 33 and negative in 108 recipients. The HLA class I, class II, and DSA-positive groups showed no difference in renal function assessed by estimated glomerular filtration rate (eGFR) at 2 years (52 ± 29 vs 52 ± 22; 56 ± 29 vs 51 ± 29; 48 ± 18 vs 53 ± 19; P = not significant [NS]). But rejection episodes at 1 year were significantly high in HLA class I and DSAb-positive group (17/35 vs 27/106; P = .019 and 16/33 vs 29/108; P = .035). The rejection episodes in the HLA class II-positive group did not show any difference when compared with the negative group (9/23 vs 40/118; P = .63). Graft survival was not affected by positivity to any of these antibodies at 2 years. CONCLUSION: Having HLA class I, class II, and DSAb does not have any influence on early and intermediate graft function. The HLA class I and DSAb positivity increases rejection episodes within 1 year in renal transplantation. Graft survival was not affected by class I, class II, and DSAb at 2 years.

Neutrophil activation occurs in the lower-limbs of patients undergoing elective repair of abdominal aortic aneurysm.

OBJECTIVES: AAA repair is associated with a systemic inflammatory response, mediated in part by neutrophils. The aim of this study was to determine where neutrophil activation occurs. METHODS: Blood was sampled from the femoral vein, portal vein and radial artery of 10 patients undergoing elective AAA repair at four time-points [induction of anaesthesia (systemic sample only), pre-aortic clamp application, pre-clamp removal and after 30min of reperfusion]. Whole blood was analysed for the white cell count, neutrophil count, and for neutrophil CD11b expression. RESULTS: The white cell count and neutrophil counts increased after aortic clamp release. Neutrophil expression of CD11b was significantly higher in the femoral vein than the portal vein and systemic circulation during ischaemia [P=0.001 (FV vs. PV), P=0.017 (FV vs. systemic)] and reperfusion [P=0.001 (FV vs. PV), P=0.013 (FV vs. systemic)]. There were no significant differences in neutrophil CD11b expression between the systemic and portal vein samples at any time. CONCLUSIONS: Ischaemia and reperfusion during abdominal aortic aneurysm repair are associated with a global increase in the white cell count and neutrophil count, but with increased neutrophil CD11b expression only in the femoral vein. This suggests the lower-limbs are sensitive to aortic clamp-related reperfusion injury and may fuel the inflammatory response.

A comparison of methods for determining genotypes at the tumour necrosis factor-alpha-308, interleukin (IL)-1beta+3953, IL-6 -174 and IL-10 -1082/-819/-592 polymorphic loci.

Induced heteroduplex genotyping (IHG) is one of many methods that can be used to determine single nucleotide polymorphisms (SNPs). It is relatively new in comparison to other polymerase chain reaction (PCR)-based techniques. The aim of this study was to compare the results of genotyping using IHG with the results of genotyping using either polymerase chain reaction-sequence-specific primers (PCR-SSP) or polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) for SNPs in the tumour necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-6 and IL-10 genes. Ninety patients who consented to participate in the study had their genotypes determined by IHG and either PCR-SSP (TNF-alpha-308 and IL-10 -1082/-819/-592) or PCR-RFLP (IL-1beta +3953 and IL-6 -174). Results for each locus were compared between techniques by calculating the Kappa statistic as a measure of agreement. The IHG and more traditional genotyping methods produced very similar results at all loci. The Kappa statistics for each locus were as follows: TNF-alpha -308, K = 0.727; IL-1beta +3953, K = 0.886; IL-6 -174, K = 0.909; IL-10 -1082, K = 0.876; IL-10 -592, K = 0.920. IHG is a valid method for the determination of genotypes at the loci examined in this study and produces comparable results to those of more traditional methods of genotyping.

Cytokines, their genetic polymorphisms, and outcome after abdominal aortic aneurysm repair.

BACKGROUND: Excessive cytokine production has been implicated in the development of organ failure. Polymorphic sites in cytokine genes have been shown to affect levels of production in vitro and may influence cytokine production in vivo. The aims of this study were to determine if cytokines or their genetic polymorphisms were related to outcome after abdominal aortic aneurysm (AAA) repair. METHODS: A prospective study of 135 patients undergoing open AAA repair. Plasma levels of TNF-alpha, IL-1beta, IL-6 and IL-10 were measured 24 h post-operatively and genotypes for the TNF-alpha -308, IL-1beta+3953, IL-6 -174, IL-10 -1082 and IL-10 -592 polymorphisms were determined for each patient. RESULTS: After elective AAA high levels of IL-10 were associated with both prolonged critical care (P<0.001) and hospital stay (P=0.001). The presence of a G allele at the IL-6 -174 locus was associated with a higher incidence of organ failure (P=0.04) and an A allele at TNF-alpha -308 with prolonged critical care stay (P=0.03). After ruptured AAA the development of multi-organ failure was associated with high levels of IL-6 (P=0.01) and TNF-alpha (P=0.04). High TNF-alpha levels were also associated with mortality (P=0.01). CONCLUSION: Post-operative cytokine levels are related to outcome after AAA repair. Cytokine gene polymorphisms may provide a method for determining which patients are at high risk of complications.

Cytokine gene polymorphisms and the inflammatory response to abdominal aortic aneurysm repair.

BACKGROUND: Cytokines are key mediators of the inflammatory response to surgery and polymorphic sites in their genes have been shown to affect cytokine production in vitro. The aim of this study was to determine whether cytokine gene polymorphisms affect cytokine production in vivo in patients undergoing abdominal aortic aneurysm (AAA) repair. METHODS: One hundred patients admitted for elective AAA repair had plasma levels of interleukin (IL) 1beta, IL-6, IL-10 and tumour necrosis factor (TNF) alpha measured at induction of anaesthesia and 24 h after operation. Genotypes for each patient were determined using induced heteroduplex genotyping for the following loci: IL-1beta + 3953, IL-6 - 174, IL-10 - 1082/-592 and TNF-alpha - 308. RESULTS: Patients with an IL-10 - 1082 A allele had a significantly higher IL-10 response to surgery than those without an A allele (P = 0.030) and there was also a significant difference in IL-10 response between patients with IL-10 - 1082 AA genotypes and those with GG genotypes (P = 0.030). CONCLUSION: Elective AAA repair results in a measurable cytokine response. In this study the magnitude of this response was not affected by the individual patient's cytokine gene polymorphisms.

The role of cytokine gene polymorphisms in the pathogenesis of abdominal aortic aneurysms: a case-control study.

BACKGROUND: Cytokines are the primary mediators of inflammation and also influence matrix metalloproteinase expression, both of which are important in development of abdominal aortic aneurysm (AAA). A significant, but as yet unknown, familial factor contributes to the pathogenesis of AAA. Many cytokine genes contain polymorphic sites, some of which affect cytokine production in vitro. Cytokine gene polymorphisms may therefore influence the pathogenesis of AAA. The purpose of this study was to determine whether there is any association between cytokine gene polymorphisms and AAA. METHODS AND RESULTS: This case-control study comprised 100 patients with AAA and 100 age-matched and sex-matched control subjects. For each case and control subject in the study, genotypes at the following cytokine gene polymorphic loci were determined: interleukin (IL)-1beta +3953, IL-6 -174, IL-10 -1082, IL-10 -592, and tumor necrosis factors-alpha -308. Allele and genotype frequencies were compared between AAA and control groups, and odds ratios (OR) were calculated for the presence of AAA with each allele at each locus examined as risk factors. The IL-10 -1082 A allele was significantly more common in the AAA group than the control group (P =.03). The OR for the IL-10 -1082 A allele as a risk factor for AAA was 1.8 (95% confidence interval, 0.9-3.6). DISCUSSION: These associations suggest a significant role for IL-10 in the pathogenesis of AAA. This association of AAA with the IL-10 -1082 A allele is also biologically plausible; the IL-10 -1082 A allele is associated with low IL-10 secretion, and it may be that AAA develops in patients who are unable to mount the same anti-inflammatory response as those who do not have AAA.

A comparison of the results of renal transplantation from non-heart-beating, conventional cadaveric, and living donors.

BACKGROUND: In an attempt to address the shortage of conventional kidney donors, a non-heart-beating donor (NHBD) organ retrieval program has been established. We compared the results of kidney transplants from NHBDs (N = 77) with those from heart-beating cadaveric (HBD; N = 224) and living donors (LD; N = 49), performed in the same eight-year period. METHODS: Patients dying after failed attempts at resuscitation in the accident department or after intracerebral hemorrhage/anoxia were considered as potential NHBDs. After death, in situ kidney perfusion and cooling were achieved using an intra-aortic catheter inserted via a femoral artery cut down. Kidney retrieval and transplant operations were performed using standard techniques. RESULTS: The median (range) warm ischemic time for NHBD kidneys was 25 minutes (5 to 53 min). The initial function rates for NHBD, HBD, and LD transplants were 6.5, 76.3, and 93%, respectively. Primary nonfunction occurred in 5 of 75 evaluable NHBD transplants (7%) compared with only 6 out of 224 (2.7%) HBD and 1 out of 49 (2%) LD transplants (P = NS). Eighty-four percent of NHBD kidney recipients required postoperative dialysis for a median of 19 days. The mean (SD) serum creatinine at 12 months was 179 (73) micromol/L in NHBD kidneys compared with 152 (57) micromol/L for HBD kidneys and 138 (44) micromol/L for LD kidneys. The actuarial five-year graft survival rates for NHBD, HBD, and LD transplants were 79, 75, and 78%, respectively. During the period under study, NHBD organs accounted for 22% of the total renal transplant program. CONCLUSIONS: Despite being associated with poor initial graft function, the long-term allograft survival of NHBD kidneys does not differ significantly from the results of HBD and LD transplants.

Workload generated by a living donor programme for renal transplantation.

BACKGROUND: The ethical and medical implications of live kidney donation result in a comprehensive work-up process. The aim of this study was to determine the magnitude of the workload and the yield of renal transplants generated by a live donor programme. METHODS: Referrals to the Leicester live donor programme over the five-year period 1994-1998 were retrospectively assessed. These were initiated by nephrology referral and subsequently investigated in a stepwise manner. Patients were counselled and baseline tests performed prior to consultant surgeon review and assessment of donor renal function/anatomy. RESULTS: One hundred and fifty referrals consisting of 150 recipients with 269 potential donors were originally made. This resulted in 32/120 (27%) related and 3/30 (10%) unrelated recipients (P=0.06) and 32/220 (15%) related and 3/49 (6%) unrelated donors proceeding to live donor transplantation, with a mean work-up time (+/-SD) of 9 (+/-7) months. One hundred and fifteen recipients (77%) and 234 (87%) donors failed to proceed at various stages of assessment, for a variety of immunological, medical and social reasons. A large number of expensive immunological investigations were required for potential donors, the majority of which did not proceed to transplantation. However as a result of performing these in the early stages of assessment the number of more invasive tests is kept to a minimum. CONCLUSIONS: There is a relatively low yield of transplants from live donor referrals, particularly those between unrelated individuals. The vast majority of referrals fail to proceed for legitimate reasons, but as a result, create a significant workload with notable staffing and financial implications.

The application of flow cytometry to histocompatibility testing.

Flow cytometry is a powerful technique that enables the sensitive and quantitative detection of both cellular antigens and bound biological moieties. This article reviews how flow cytometry is increasingly being used as histocompatibility laboratories for the analysis of antibody specificity and HLA antigen expression. A basic description of flow cytometry principles and standardisation is given, together with an outline of clinical application in the areas of pre-transplant cross-matching, antibody screening, post-transplant antibody monitoring and HLA-B27 detection. It is concluded that flow cytometry is a useful multi-parametric analytical tool, yielding clinical benefit especially in the identification of patients at risk of early transplant rejection.

Influence of allograft size to recipient body-weight ratio on the long-term outcome of renal transplantation.

BACKGROUND: The critical nephron mass needed to meet the metabolic demands of an individual depends on the body-weight. This study evaluated the effect of the kidney transplant ultrasonographic size to recipient body-weight ratio (Tx/W) on the outcome of kidney transplantation. METHODS: A consecutive series of 104 cadaveric renal transplants was studied. Transplant cross-sectional area (TXSA) was measured ultrasonographically in the first week after transplantation as an index of renal size. A 'nephron dose' index (Tx/W) was calculated by dividing TXSA by recipient weight and was used to define three groups of patients, with high (more than 0.45), medium (0.3-0.45) or low (less than 0.3) Tx/W ratios. Isotope glomerular filtration rate (GFR) measurements were made at 1, 6 and 12 months after transplantation. RESULTS: The serum creatinine level was significantly lower in the first 5 years after transplantation in patients with a high Tx/W ratio than in those with a medium or low ratio. GFR measurements were marginally higher in the groups with a high and medium Tx/W ratio compared with the low Tx/W group. A statistically significant association between Tx/W ratio and graft survival was not found. CONCLUSION: The renal transplant size to recipient weight ratio was an important determinant of long-term renal allograft function in this study. Extreme mismatching between allograft and recipient size should be avoided where possible, but the findings presented require confirmation in larger studies before clear recommendations can be made about size matching and kidney allocation.

Evidence for the differential expression of a variant EGF receptor protein in human prostate cancer.

Earlier studies have demonstrated an unexplained depletion of the epidermal growth factor receptor (EGFR) protein expression in prostatic cancer. We now attribute this phenomenon to the presence of a variant EGFR (EGFRvIII) that is highly expressed in malignant prostatic neoplasms. In a retrospective study, normal, benign hyperplastic and malignant prostatic tissues were examined at the mRNA and protein levels for the presence of this mutant receptor. The results demonstrated that whilst EGFRvIII was not present in normal prostatic glands, the level of expression of this variant protein increased progressively with the gradual transformation of the tissues to the malignant phenotype. The selective association of high EGFRvIII levels with the cancer phenotype underlines the role that this mutant receptor may maintain in the initiation and progression of malignant prostatic growth, and opens the way for new approaches in the management of this disease including gene therapy.