Update to the study protocol, including statistical analysis plan, for the multicentre, randomised controlled OuTSMART trial: a combined screening/treatment programme to prevent premature failure of renal transplants due to chronic rejection in patients with HLA antibodies.

BACKGROUND: Chronic rejection is the single biggest cause of premature kidney graft failure. HLA antibodies (Ab) are an established prognostic biomarker for premature graft failure so there is a need to test whether treatment decisions based on the presence of the biomarker can alter prognosis. The Optimised TacrolimuS and MMF for HLA Antibodies after Renal Transplantation (OuTSMART) trial combines two elements. Firstly, testing whether a routine screening programme for HLA Ab in all kidney transplant recipients is useful by comparing blinding versus unblinding of HLA Ab status. Secondly, for those found to be HLA Ab+, testing whether the introduction of a standard optimisation treatment protocol can reduce graft failure rates. METHODS: OuTSMART is a prospective, open-labelled, randomised biomarker-based strategy (hybrid) trial, with two arms stratified by biomarker (HLA Ab) status. The primary outcome was amended from graft failure rates at 3 years to time to graft failure to increase power and require fewer participants to be recruited. Length of follow-up subsequently is variable, with all participants followed up for at least 43 months up to a maximum of 89 months. The primary outcome will be analysed using Cox regression adjusting for stratification factors. Analyses will be according to the intention-to-treat using all participants as randomised. Outcomes will be analysed comparing standard care versus biomarker-led care groups within the HLA Ab+ participants (including those who become HLA Ab+ through re-screening) as well as between HLA-Ab-unblinded and HLA-Ab-blinded groups using all participants. DISCUSSION: Changes to the primary outcome permit recruitment of fewer participants to achieve the same statistical power. Pre-stating the statistical analysis plan guards against changes to the analysis methods at the point of analysis that might otherwise introduce bias through knowledge of the data. Any deviations from the analysis plan will be justified in the final report. TRIAL REGISTRATION: ISRCTN registry, ID: ISRCTN46157828 . Registered on 26 March 2013; EudraCT 2012-004308-36 . Registered on 10 December 2012.

Can a combined screening/treatment programme prevent premature failure of renal transplants due to chronic rejection in patients with HLA antibodies: study protocol for the multicentre randomised controlled OuTSMART trial.

BACKGROUND: Renal transplantation is the best treatment for kidney failure, in terms of length and quality of life and cost-effectiveness. However, most transplants fail after 10 to 12 years, consigning patients back onto dialysis. Damage by the immune system accounts for approximately 50% of failing transplants and it is possible to identify patients at risk by screening for the presence of antibodies against human leukocyte antigens. However, it is not clear how best to treat patients with antibodies. This trial will test a combined screening and treatment protocol in renal transplant recipients. METHODS/DESIGN: Recipients>1 year post-transplantation, aged 18 to 70 with an estimated glomerular filtration rate>30 mL/min will be randomly allocated to blinded or unblinded screening arms, before being screened for the presence of antibodies. In the unblinded arm, test results will be revealed. Those with antibodies will have biomarker-led care, consisting of a change in their anti-rejection drugs to prednisone, tacrolimus and mycophenolate mofetil. In the blinded arm, screening results will be double blinded and all recruits will remain on current therapy (standard care). In both arms, those without antibodies will be retested every 8 months for 3 years. The primary outcome is the 3-year kidney failure rate for the antibody-positive recruits, as measured by initiation of long-term dialysis or re-transplantation, predicted to be approximately 20% in the standard care group but <10% in biomarker-led care. The secondary outcomes include the rate of transplant dysfunction, incidence of infection, cancer and diabetes mellitus, an analysis of adherence with medication and a health economic analysis of the combined screening and treatment protocol. Blood samples will be collected and stored every 4 months and will form the basis of separately funded studies to identify new biomarkers associated with the outcomes. DISCUSSION: We have evidence that the biomarker-led care regime will be effective at preventing graft dysfunction and expect this to feed through to graft survival. This trial will confirm the benefit of routine screening and lead to a greater understanding of how to keep kidney transplants working longer. TRIAL REGISTRATION: Current Controlled Trials ISRCTN46157828.

Predictors of renal outcome in HIV-associated nephropathy.

BACKGROUND: Human immunodeficiency virus (HIV)-associated nephropathy (HIVAN) is an important cause of end-stage renal disease among African American patients. This study was performed to study the epidemiology of HIVAN in a predominantly black African population and the impact of highly active antiretroviral therapy and other factors on the development of end-stage renal disease. METHODS: We retrospectively identified all patients with HIVAN, defined by biopsy or strict clinical criteria, in 8 clinics in the United Kingdom. Baseline renal function, HIV parameters, renal pathological index of chronic damage, and responses to highly active antiretroviral therapy were analyzed, and factors associated with adverse renal outcome were identified. RESULTS: From 1998 through 2004, we studied 16,834 patients, 61 of whom had HIVAN. HIVAN prevalence in black patients was 0.93%, and HIVAN incidence in those without renal disease at baseline was 0.61 per 1000 person-years. After a median of 4.2 years, 34 patients (56%) had developed end-stage renal disease. There were no significant differences in renal function and HIV parameters at baseline, time to initiation of highly active antiretroviral therapy, and rates of HIV RNA suppression between the 20 patients who developed end-stage renal disease >3 months after receiving the HIVAN diagnosis and the 23 patients who maintained stable renal function. However, the index of chronic damage score was significantly higher in those who developed end-stage renal disease (P < .001), and an index of chronic damage score >75 was associated with shorter renal survival (P < .001). CONCLUSIONS: Whereas overall patient survival suggested an important benefit of highly active antiretroviral therapy, no additional renal benefit of early initiation of highly active antiretroviral therapy or viral suppression could be demonstrated in this large cohort of patients with established HIVAN. Severity of chronic kidney damage, as quantified by biopsy, was the strongest predictor of renal outcome.

Pretransplant donor-specific antibodies in cytotoxic negative crossmatch kidney transplants: are they relevant?

BACKGROUND: The corresponding antigens of alloantibodies identified in patients awaiting kidney transplantation are often listed as unacceptable for transplantation. The use of solid phase testing, being more sensitive and accurate than conventional complement-dependent cytotoxicity (CDC) assays, has resulted in increased identification of alloantibodies. We aimed to study the clinical importance of alloantibodies defined solely by solid phase techniques. METHODS: All patients transplanted between 1999 and 2001 at our center with available day-of-transplant sera (D0) were included (121 patients). All had negative CDC crossmatches. RESULTS: Thirty-eight patients (31%) had detectable alloantibodies using high-definition assays with 16 having donor-specific antibodies (DSA) and 22 non-DSA. There were no cases of hyperacute rejection in any of the groups. Biopsy-proven acute rejection rates in the DSA and non-DSA were similar to the unsensitized group. Delayed graft function and 1-year graft survival rates were also similar for the three groups as were median 1-year serum creatinine levels. Multivariate analysis, however, showed that DSA were associated with an increased relative risk of longer-term graft failure (relative risk, 6.5; P<0.05). CONCLUSIONS: These data show that in the context of a CDC-negative crossmatch, the presence of D0 DSA has little impact on any early graft parameters. DSA, however, are associated with poorer longer-term graft outcomes in kidney transplantation.

Long-term graft outcome with mycophenolate mofetil and azathioprine: A paired kidney analysis.

BACKGROUND: Randomized controlled trials and U.S. Registry data have demonstrated that mycophenolate mofetil (MMF) reduces acute rejection rates and improves graft survival. We undertook the first paired kidney analysis comparing the effects of MMF and azathioprine on graft outcome in the United Kingdom. METHODS: In all, 238 deceased donors from 1999 to 2002 who donated one kidney to a patient treated with MMF and the other kidney to a patient treated with azathioprine were identified from the national transplant database held by U.K. Transplant. Graft function and rates of change of graft function were compared using multiple linear regression analyses adjusting for covariates on an intention-to-treat basis. Incidence of acute rejection and delayed graft function were studied using logistic regression. Patient and graft survival censored for death were evaluated with Cox regression analyses. RESULTS: The MMF-treated patients exhibited a nonsignificant trend towards improved graft function but with increased rejection rates (44% versus 31%, P < 0.01). Treatment with MMF did not reduce delayed graft function rates. Univariate analysis showed that graft survival was inferior in MMF-treated patients (90% versus 95%, log-rank, P = 0.02) but in multivariate Cox regression models, MMF treatment was not a significant factor. Surprisingly, in the first year 32% of patients achieved daily doses of less than 2 g of MMF compared to 18% of patients who received less than 100 mg of azathioprine (P < 0.01). CONCLUSION: In this real-life study, there was no difference in patient or graft outcome between MMF and azathioprine treated groups despite increased rejection rates in patients receiving MMF therapy.

Homocysteine induced impairment of nitric oxide-dependent vasorelaxation is reversible by the superoxide dismutase mimetic TEMPOL.

BACKGROUND: Elevated plasma homocysteine concentrations in renal patients are associated with accelerated cardiovascular disease. The mechanism(s) by which homocysteine acts remains unclear however, evidence implicates a role involving endothelial dysfunction. METHODS: Rat femoral arteries after acute or 4-h pre-incubation with racemic D,L-homocysteine (100 microM) were mounted on a myograph, pre-constricted with phenylephrine (10 microM) and responses to acetylcholine-dependent vasorelaxation examined. The incubations were repeated in the presence of indomethacin (10 microM), omega-nitro-L-arginine methyl ester (100 microM), L-arginine (100 microM), tetrahydrobiopterin (1 microM), catalase (1200 U/ml), ebselen, a peroxynitrite chelator (20 microM) and TEMPOL, a superoxide dismutase mimetic (1 mM). Results are shown as means+/-standard error, expressed as per cent relaxation to acetylcholine added (nmol/l). RESULTS: Increasing concentrations of homocysteine had no affect when added directly to basally relaxed or pre-constricted freshly isolated vessels. However, 4-h pre-incubation with or without homocysteine significantly shifted the acetylcholine EC(50) (EC(50) was defined as the concentration of acetylcholine that caused relaxation of the phenylephrine contracted tissue by 50%), control((4 h)) = 74.7 nmol/l+/-10.5 vs 100 microM D,L-homocysteine((4 h)) = 159.9 nmol/l+/-20.6; P<0.05) without affecting maximal relaxation. Response to endothelial independent relaxation was unaffected. Indomethacin, indomethacin and omega-nitro-L-arginine methyl ester, l-arginine and tetrahydrobiopterin, catalase and ebselen had no effect on the EC(50) in homocysteine-exposed arteries. However, TEMPOL normalized vasorelaxation in homocysteine-treated arteries (75.2 nmol/l+/-14.6) but had no effect on the 4-h control group. Moreover, washing TEMPOL from the treated vessels restored endothelial dysfunction in D,L-homocysteine-treated vessels (163.9 nmol/l+/-34.1). CONCLUSIONS: We conclude that homocysteine causes endothelial dysfunction by up-regulating a potential superoxide generating system resulting in reduced nitric oxide bio-availability.

Structural remodeling of resistance arteries in uremic hypertension.

BACKGROUND: Structural remodeling of the resistance vasculature is present in many forms of human and experimental hypertension. In particular, an increase in the ratio of wall thickness to lumen diameter develops, and might in itself maintain hypertension by increasing vascular resistance. Because uremia is associated with raised peripheral resistance, hypertension, and histologic changes suggestive of vascular remodeling, we sought to formally examine the structural and mechanical (elastic) properties of isolated pressurized resistance arteries in uremic hypertension. METHODS: Cremaster, cerebral and mesenteric arteries from subtotally nephrectomised Wistar-Kyoto rats, normotensive control Wistar-Kyoto rats, and spontaneously hypertensive rats were mounted on a pressure myograph and relaxed in calcium-free buffer. Wall thickness and lumen diameter were measured at increasing lumen pressures from 10 to 200 mm Hg, and from this wall:lumen ratio, wall cross-sectional area, and an index of elasticity were derived. RESULTS: In uremic hypertensive animals increased wall:lumen ratio and decreased lumen diameter was seen in cremaster and mesenteric arteries, although no significant changes were observed in cerebral arteries, compared to normotensive controls. In spontaneously hypertensive animals increased wall thickness and wall:lumen ratio was seen in cerebral and mesenteric arteries, decreased lumen diameter in cremaster and mesenteric arteries, and increased wall cross-sectional area in cerebral arteries, compared to normotensive controls. Elasticity of the arterial wall in uremic and spontaneously hypertensive animals did not differ from normotensive controls. CONCLUSION: Cremaster and mesenteric resistance arteries undergo predominantly eutrophic inward remodeling in uremic hypertension, broadly similar to that seen in spontaneous hypertension.

Remission of collapsing focal segmental glomerulosclerosis following chemotherapy for myeloma.

Collapsing focal segmental glomerulosclerosis can be idiopathic but has been associated with human immunodeficiency virus infection as well as lymphoproliferative, myeloproliferative, and autoimmune disorders. It has a more rapid progression to end-stage renal failure. Treatment of this condition is poorly defined with no clear evidence of response to any treatment regimens. We report a case of collapsing focal segmental glomerulosclerosis in association with multiple myeloma that underwent partial remission following treatment for the underlying disease.

Altered L-arginine metabolism results in increased nitric oxide release from uraemic endothelial cells.

Results regarding the nitric oxide (NO) system in uraemia are contradictory. L-arginine, the precursor of NO, is also metabolized by arginase to form ornithine and urea. In the present study, endothelial NO production and arginine metabolism in uraemia were assessed. In addition an in vivo model was used to examine excess consumption of NO in uraemia. NO and amino acid measurements were made from basal and stimulated (by bradykinin) uraemic and control endothelial cells. Reverse-transcriptase PCR was used to assess endothelial NO synthase (eNOS) and inducible NOS (iNOS) expression. Finally, aortae of uraemic rats were stained for nitrotyrosine (a marker of peroxynitrite). Basal uraemic cells produced more NO than the control cells. L-arginine levels were greater in uraemic (supernatants/cells), but ornithine levels were higher in control (supernatants/cells). Following stimulation, NO levels in supernatants were similar, but the rise in NO production was greater in control compared with uraemic cells; l-arginine levels still remained higher in uraemic supernatants/cells. Differences in ornithine concentration (supernatants/cells) disappeared following bradykinin stimulation, due to a rise in ornithine levels in the uraemic group. There was no difference in eNOS expression, nor was iNOS detected in either group. Only aortae from uraemic rats showed evidence for nitrotyrosine staining. These studies demonstrated increased basal NO release in uraemic endothelial cells, perhaps by inhibition of arginase and hence diversion of arginine to the NO pathway. The increased NO produced under basal conditions may be inactive due to excessive consumption, resulting in peroxynitrite formation. Interestingly, bradykinin appears to restore arginase activity in uraemia, resulting in normalization of NO production.