G.P. Segoloni

Antiangiogenic and immunomodulatory effects of rapamycin on islet endothelium: Relevance for islet transplantation

Donor intra‐islet endothelial cells contribute to neovascularization after transplantation. Several factors may interfere with this process and ultimately influence islet engraftment. Rapamycin, a central immunosuppressant in islet transplantation, is an mTOR inhibitor that has been shown to inhibit cancer angiogenesis. The aim of this study was to evaluate the effects of rapamycin on islet endothelium. Rapamycin inhibited the outgrowth of endothelial cells from freshly purified human islets and the formation of capillary‐like structures in vitro and in vivo after subcutaneous injection within Matrigel plugs into SCID mice. Rapamycin decreased migration, proliferation and angiogenic properties of human and mouse islet‐derived endothelial cell lines with appearance of apoptosis. The expression of angionesis‐related factors VEGF, αVβ3 integrin and thrombospondin‐1 on islet endothelium was altered in the presence of rapamycin. On the other hand, rapamycin decreased the surface expression of molecules involved in immune processes such as ICAM‐1 and CD40 and reduced the adhesion of T cells to islet endothelium. Our results suggest that rapamycin exerts dual effects on islet endothelium inducing a simultaneous inhibition of angiogenesis and a down‐regulation of receptors involved in lymphocyte adhesion and activation.

Loss of nephrin expression in glomeruli of kidney-transplanted patients under m-TOR inhibitor therapy.

The development of proteinuria has been observed in kidney‐transplanted patients on m‐TOR inhibitor (m‐TORi) treatment. Recent studies suggest that m‐TORi(s) may alter the behavior and integrity of glomerular podocytes. We analyzed renal biopsies from kidney‐transplanted patients and evaluated the expression of nephrin, a critical component of the glomerular slit‐diaphragm. In a group of patients on ‘de novo’ m‐TORi‐treatment, the expression of nephrin within glomeruli was significantly reduced in all cases compared to pretransplant donor biopsies. Biopsies from control transplant patients not treated with m‐TORi(s) failed to present a loss of nephrin. In a group of patients subsequently converted to m‐TORi‐treatment, a protocol biopsy performed before introduction of m‐TORi was also available. The expression of nephrin in the pre‐m‐TORi biopsies was similar to that observed in the pretransplant donor biopsies but was significantly reduced after introduction of m‐TORi(s). Proteinuria increased after the m‐TORi inititiation in this group. However, in some cases proteinuria remained normal despite reduction of nephrin. In vitro, sirolimus downregulated nephrin expression by human podocytes. Our results suggest that m‐TORi(s) may affect nephrin expression in kidney‐transplanted patients, consistently with the observation in vitro on cultured podocytes.

Comparison between 24‐h proteinuria, urinary protein/creatinine ratio and dipstick test in patients with nephropathy: Patterns of proteinuria in dipstick‐negative patients

Objective. Three main tests are commonly employed for the measurement of proteinuria: the dipstick test, the urinary protein/creatinine ratio (P/C) and the 24‐h urine collection. The aim of this study was to evaluate the correlation between these methods, comparing linear regression and ROC curve data. Material and methods. A total of 297 consecutive outpatients with different renal diseases were included in the study. Twenty‐four‐hour proteinuria was considered the reference test. Results. A high degree of correlation was observed between all the tests (p<0.0001), the highest regression coefficient being between 24‐h proteinuria and P/C (R = 0.82), and the lowest between P/C and the dipstick test (R = 0.72). The dipstick test failed to detect pathological proteinuria in 94 patients (31.6 %). Therefore, in these subjects, the patterns of proteinuria were assessed by immunofixation and sodium dodecyl sulphate (SDS) electrophoresis. Conclusions. Our data strongly support the use of urinary P/C for the detection of proteinuria, at least in nephrology units, where the prevalence of proteinuria is likely to be high.

The production of platelet-activating factor during hemodialysis.

Regenerated cellulosic membranes (CU) induced the aggregation of plasma-free human neutrophils when recirculated in a dynamic model of dialysis without the patient on the circuit. Neutrophil aggregation was linked to the production of PAF by these cells. In the absence of detectable PAF production, no neutrophil aggregation occurred, as observed during recirculation with polymethylmethacrylate (PMMA) membranes. With polycarbonate (PC), PAF production and aggregation of neutrophils were both almost half the values with CU. PAF production was studied in ten hemodialysis (HD) patients tested twice with CU and once with PC and PMMA membranes. PAF was extracted in the venous blood during filling of the dialyser for 9/20 of patients with CU (3.1 ± 2.9 ng/ml, mean ± 1 S.D.) a membrane that induced marked leukopenia (> 50% of basal values at 15 min), C3a des Arg generation (> 500% at 5 min), and plasma levels of the elastase-alpha1-proteinase inhibitor complex (> 500% at the end of HD). Membranes such as PC and PMMA showing intermediate or low potential to induce leukopenia and C3a des Arg generation, respectively, did not trigger the production and release of PAF in detectable amounts at any interval. However, with PMMA, plasma neutrophil elastase was significantly higher than baseline at the end of dialysis. These levels were not significantly different (p < 0.05) from those observed with CU and PC membranes

Assessment of cardiovascular risk in waiting-listed renal transplant patients: a single center experience in 558 cases

Abstract:  Cardiac screening is recommended to prevent cardiovascular death after renal transplantation. This retrospective observational study illustrates the results of application of a cardiac assessment algorithm in a series of 558 renal transplant candidates at a single center in Turin, Italy. A dipyridamole‐stress sestamibi myocardial scintiscan (DMS) performed in 302/558 (54.1%) cases was positive in 52 (17.2%), negative in 200 (66.2%), borderline in 16 (5.3%), and with signs of previous necrosis in 34 (11.4%). Coronary lesions detected by angiography in 48.1% of the 52 positives were treated medically (13.5%) or by percutaneous/surgical procedure (34.6%). Coronary lesions were detected in 14.1% of asymptomatic population subgroup. The minor and major cardiovascular event rates and the cardiovascular death rate were 1.9%, 0%, and 0%, respectively, in positive DMS group (high‐cardiological risk) vs. 10%, 4.5%, and 3.5% in the negatives (p > 0.5; n.s.). It is suggested that not increased cardiovascular event or deaths rates in the high‐risk group reflect early coronary lesion detection and correction. Since 55.9% of cardiovascular events or deaths occurred in the negative group more than 24 months after the DMS, its mandatory repetition every two yr after a negative finding is recommended.

Survival probabilities for renal transplant recipients and dialytic patients: a single center prospective study

IN THE EVALUATION of the relative merits of dialysis and transplantation for replacement of renal function, survival of the patient is a basic measure of efficacy. Data from such a study may offer useful information, not only on a clinical ground, but to answer whether the patient who is successfully treated with dialysis can achieve a longer survival with transplantation. Physicians can also obtain suggestions and guidelines for an ethical organ allocation; extremely valuable since we are facing a worldwide widening gap between the number of patients awaiting transplantation and the scarce organ availability. On the other hand, when the relational and psychological levels are considered, results from these studies may provide informed counselling to the patient on the expected benefit of each option and help in removing the disturbing doubts from both doctors and relatives when the death of the recipient occurs early after transplantation. From a survey of previous papers on this issue, three main methodologic approaches are adopted: the first one, a crude comparison between the dialytic population and the grafted patients; the second one, in which prognostic disparities are corrected with statistical methods; the third one, based on the assumption that choosing two populations as similar as possible, one could minimize the various bias connected, for example, with patient selection and the risk of dying during the time spent on the waiting list.

Analysis of Four Scoring Systems and Monocentric Experience to Optimize Criteria for Marginal Kidney Transplantation

There is a strong need among the transplantation community to identify common criteria to utilize the pool of expanded criteria donors (ECD), considering the disparity between organ demand and supply as well as the benefits of transplantation on long-term mortality compared with survival on dialysis, also in patients transplanted with these organs. The purpose of this article was to analyze scoring systems proposed in literature by Nyberg, Anglicheau, Rao (Kidney Donor Risk Index), and Schold, seeking to verify whether our clinical and histological allocation strategy matched the Nyberg score. Herein we have reported the results of a preliminary retrospective study on the 5-year outcomes of organs from 60 marginal donors, who were older than 50 years and histologically evaluated before implantation. The donors matched Nyberg class C and D, that is, marginal donors. We noted a tendency toward an association between global and vascular scores with class D (odds ratio 2.2 and 4.3, respectively). Kaplan-Meier graft survival curves were similar to Nyberg data: 83% for class C versus 73% for class D at 5 years. Without any comparison to the Nyberg score, the only feature that was predictive of renal function at 5 years in our population was hypertension in the donor. Further studies are required to identify which of the scoring systems--clinical and/or histological--is more suitable to allocate ECD kidneys and to predict recipient outcomes.

Study of lymphocyte costimulatory molecules in renal transplantation

RECENT studies have shown that the interaction between the B7-1 and B7-2 receptors, expressed on the surface of antigen presenting cells (APC), and their counter-receptors CD28 and CTLA-4, present on T lymphocytes, triggers a costimulatory signal that is crucial for antigen-dependent lymphocyte activation. In fact, blockade of CTLA-4/CD28-B7 pathway prevents acute rejection of experimental graft, suggesting a new therapeutic strategy. Another critical pathway in lymphocyte activation involves the interaction between a 39 kDa surface glycoprotein named CD40ligand (CD40L) expressed by T lymphocytes and its counter-receptor CD40 present on B cells, dendritic cells, activated macrophages, and endothelial cells. Blockade or genetic deficiency of this costimulatory pathway prevents the activation of both T and B cells. Recently, we have shown the expression of the inducible B7-1, B7-2, CTLA-4, and CD40L/CD40 in the bioptic tissue of patients receiving renal transplantation. We extend these observations and focus on the pathogenic implications of the differential expression of CD40 and CD40L in acute and chronic rejection.

Daily dialysis Kt/V and flexible schedules: is it possible to control efficiency, when and how?

Background Daily hemodialysis is a promising treatment schedule but uniform criteria for defining efficiency are lacking. Methods On our daily dialysis (DD) schedule, duration is flexible (2–3 hours, patients are free to add up to 30min/session), Qb 250–350 mL/min; dialyser 1.6–1.8 m2. Study was performed on 12 pts on DD for ≥2 months, with ≥4 Kt/V on subsequent days, tested in the same laboratory. Goal: To evaluate variability and identify a simple method for weekly calculation, Kt/V was assessed for 133 sessions. Results On flexible DD, variability of Kt/V-session is high (relative error 4.9%-22%). On flexible schedules, within the time range chosen (2–3 hours) variability of average hourly Kt/V is lower (standard deviation: min (0.014; max (0.052 hour, relative error 4.9%-10%) allowing calculation of weekly Kt/V (averaging 3 sessions: relative error <6%) suitable for clinical practice. Conclusions Flexible schedules, allowing patients to increase treatment time, are an interesting clinical option, but a challenge for Kt/V assessment.

Assessment of Platelet Function Analyzer (PFA-100) in Kidney Transplant Patients Before Renal Allograft Biopsy: A Retrospective Single-Center Analysis

BACKGROUND Kidney biopsy (KB) represents the criterion standard to obtain information on diagnosis and prognosis of renal allograft dysfunctions. However, it can be associated with bleeding complications (BCs). Bleeding time test (BTT), the best predictive indicator of post-biopsy BCs, is not a very reproducible test and is invasive. Therefore, the aim of this study was to evaluate whether the platelet function analyzer (PFA-100), a very reliable test to investigate primary hemostasis, could be useful in predicting the risk of bleeding complications in transplant patients undergoing KB. METHODS We carried out a retrospective analysis of PFA-100 collagen-epinephrine (C-EPI) and collagen-adenosine diphosphate (C-ADP) closure times in 119 patients undergoing KB in our center. Data regarding BTT, age, sex, blood pressure, number of renal allograft punctures for each biopsy procedure, thromboplastin time, prothrombin time, complete blood count, and prophylactic therapy with desmopressin were also collected. Major (need for blood transfusion) or minor (no need for any intervention) BCs (hematoma and hematuria) were recorded. RESULTS Indications for KB were: delayed graft function (n=23), allograft dysfunction (n=40), proteinuria (n=27), allograft dysfunction plus proteinuria (n=19), and protocol biopsy (n=10). Nine of the 119 patients (7.5%) developed minor BCs (6 macrohematuria, 3 hematoma), major BCs did not develop. No significant differences were found in any of the clinical and laboratory data, including BTT and PFA-100 (C-EPI and C-ADP) between patients who developed BCs compared with those who did not. In addition, there was no correlation between PFA-100 test (C-EPI and C-ADP) values and BTT data [R2=0.002; P=.6]. CONCLUSIONS The PFA-100 test was not useful in predicting the risk of BCs in kidney transplant patients undergoing renal allograft biopsy.