Alferso C. Abrahams

Differential effects of acute and sustained cyclosporine and tacrolimus on sympathetic nerve activity

Background We studied the effect of acute and sustained cyclosporine and tacrolimus on muscle sympathetic nerve activity (MSNA) in groups of healthy male volunteers. Methods and results Acute cyclosporine in normal dose (2.5 mg/kg) increased MSNA from 11 ± 6 to 19 ± 8 bursts/min (P < 0.05). Acute cyclosporine in high dose (10 mg/kg) increased MSNA from 13 ± 6 to 25 ± 4 bursts/min (P < 0.05) and increased heart rate and mean arterial pressure (heart rate from 64 ± 8 to 74 ± 6 b.p.m., MAP from 92 ± 10 to 105 ± 8 mmHg; both P < 0.05). Sustained cyclosporine (2.5 mg/kg b.i.d. for 2 weeks) suppressed MSNA from 14 ± 6 to 8 ± 7 bursts/min (P < 0.05). Blood pressure increased from 89 ± 6 to 98 ± 6 mmHg (P < 0.05). Body weight increased and plasma renin activity was suppressed. Acute tacrolimus in regular dose (0.05 mg/kg) and high dose (0.20 mg/kg) had no effect on MSNA and blood pressure. Sustained tacrolimus (0.05 mg/kg b.i.d. for 2 weeks) had no effect on blood pressure, body weight and plasma renin activity, but decreased MSNA from 14 ± 6 to 8 ± 5 bursts/min (P < 0.05). Conclusion Sympathetic overactivity plays a role in the acute hypertensive action of cyclosporine. Cyclosporine given during 2 weeks increases blood pressure and suppresses MSNA, possibly by volume retention. Tacrolimus, in the presently applied dosages, does not cause hypertension or sympathetic overactivity. However, sustained tacrolimus also suppresses sympathetic activity, the reason of which is unclear.

A COLLABORATIVE APPROACH TO UNDERSTANDING EPS: THE EUROPEAN PERSPECTIVE

Manchester Royal Infirmary,1 Manchester, UK; University Medical Center Utrecht,2 Utrecht, Netherlands; Robert-Bosch-Hospital,3 Stuttgart, Germany; Erasmus Medical Center,4 Rotterdam, Netherlands; Hans Mak Institute,5 Naarden, Netherlands; University Hospital of North Staffordshire,6 Stoke-on-Trent, UK; Azienda Ospedaliera Universitaria Senese,7 Siena, Italy; Universite catholique de Louvain,8 Brussels, Belgium; Albert Schweitzer Hospital,9 Dordrecht, Netherlands; Karolinska Institutet,10 Stockholm, Sweden; Baxter Healthcare SA,11 Zurich, Switzerland; University of Amsterdam,12 Amsterdam, Netherlands; Leiden University Medical Center,13 Leiden, Netherlands; Rene Dubos Hospital,14 Pontoise, France; and University Medical Center Groningen,15 Groningen, Netherlands

Patients with Encapsulating Peritoneal Sclerosis Have Increased Peritoneal Expression of Connective Tissue Growth Factor (CCN2), Transforming Growth Factor-β1, and Vascular Endothelial Growth Factor

Introduction Encapsulating peritoneal sclerosis (EPS) is a devastating complication of peritoneal dialysis (PD). The pathogenesis is not exactly known and no preventive strategy or targeted medical therapy is available. CCN2 has both pro-fibrotic and pro-angiogenic actions and appears an attractive target. Therefore, we studied peritoneal expression of CCN2, as well as TGFβ1 and VEGF, in different stages of peritoneal fibrosis. Materials and methods Sixteen PD patients were investigated and compared to 12 hemodialysis patients and four pre-emptively transplanted patients. Furthermore, expression was investigated in 12 EPS patients in comparison with 13 PD and 12 non-PD patients without EPS. Peritoneal tissue was taken during kidney transplantation procedure or during EPS surgery. In a subset of patients, CCN2 protein levels in peritoneal effluent and plasma were determined. Samples were examined by qPCR, histology, immunohistochemistry, and ELISA. Results Peritoneal CCN2 expression was 5-fold higher in PD patients compared to pre-emptively transplanted patients (P<0.05), but did not differ from hemodialysis patients. Peritoneal expression of TGFβ1 and VEGF were not different between the three groups; neither was peritoneal thickness. Peritoneum of EPS patients exhibited increased expression of CCN2 (35-fold, P<0.001), TGFβ1 (24-fold, P<0.05), and VEGF (77-fold, P<0.001) compared to PD patients without EPS. In EPS patients, CCN2 protein was mainly localized in peritoneal endothelial cells and fibroblasts. CCN2 protein levels were significantly higher in peritoneal effluent of EPS patients compared to levels in dialysate of PD patients (12.0±4.5 vs. 0.91±0.92 ng/ml, P<0.01), while plasma CCN2 levels were not increased. Conclusions Peritoneal expression of CCN2, TGFβ1, and VEGF are significantly increased in EPS patients. In early stages of peritoneal fibrosis, only CCN2 expression is slightly increased. Peritoneal CCN2 overexpression in EPS patients is a locally driven response. The potential of CCN2 as biomarker and target for CCN2-inhibiting agents to prevent or treat EPS warrants further study.

Effect of GFR on Plasma N-Terminal Connective Tissue Growth Factor (CTGF) Concentrations

BACKGROUND Connective tissue growth factor (CTGF) has a key role in the pathogenesis of renal and cardiac fibrosis. Its amino-terminal fragment (N-CTGF), the predominant form of CTGF detected in plasma, has a molecular weight in the middle molecular range (18 kDa). However, it is unknown whether N-CTGF is a uremic retention solute that accumulates in chronic kidney disease (CKD) due to decreased renal clearance and whether it can be removed by hemodiafiltration. STUDY DESIGN 4 observational studies in patients and 2 pharmacokinetic studies in rodents. SETTING & PARTICIPANTS 4 single-center studies. First study (cross-sectional): 88 patients with CKD not receiving kidney replacement therapy. Second study (cross-sectional): 23 patients with end-stage kidney disease undergoing low-flux hemodialysis. Third study: 9 kidney transplant recipients before and 6 months after transplant. Fourth study: 11 low-flux hemodialysis patients and 12 hemodiafiltration patients before and after one dialysis session. PREDICTOR First, second, and third study: (residual) glomerular filtration rate (GFR). Fourth study: dialysis modality. OUTCOMES & MEASUREMENTS Plasma (N-)CTGF concentrations, measured by enzyme-linked immunosorbent assay. RESULTS In patients with CKD, we observed an independent association between plasma CTGF level and estimated GFR (β = -0.72; P < 0.001). In patients with end-stage kidney disease, plasma CTGF level correlated independently with residual kidney function (β = -0.55; P = 0.046). Successful kidney transplant resulted in a decrease in plasma CTGF level (P = 0.008) proportional to the increase in estimated GFR. Plasma CTGF was not removed by low-flux hemodialysis, whereas it was decreased by 68% by a single hemodiafiltration session (P < 0.001). Pharmacokinetic studies in nonuremic rodents confirmed that renal clearance is the major elimination route of N-CTGF. LIMITATIONS Observational studies with limited number of patients. Fourth study: nonrandomized, evaluation of the effect of one session; randomized longitudinal study is warranted. CONCLUSION Plasma (N-)CTGF is eliminated predominantly by the kidney, accumulates in CKD, and is decreased substantially by a single hemodiafiltration session.

Proteins in Preservation Fluid as Predictors of Delayed Graft Function in Kidneys from Donors after Circulatory Death

BACKGROUND AND OBJECTIVES Kidney transplantation is the preferred treatment for ESRD, and donor kidney shortage urges proper donor-recipient matching. Zero-hour biopsies provide predictive values for short- and long-term transplantation outcomes, but are invasive and may not reflect the entire organ. Alternative, more representative methods to predict transplantation outcome are required. We hypothesized that proteins accumulating in preservation fluid during cold ischemic storage can serve as biomarkers to predict post-transplantation graft function. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS Levels of 158 proteins were measured in preservation fluids from kidneys donated after circulatory death (Maastricht category III) collected in two Dutch centers (University Medical Center Utrecht and Erasmus Medical Center Rotterdam) between 2013 and 2015. Five candidate biomarkers identified in a discovery set of eight kidneys with immediate function (IF) versus eight with delayed graft function (DGF) were subsequently analyzed in a verification set of 40 additional preservation fluids to establish a prediction model. RESULTS Variables tested for their contribution to a prediction model included five proteins (leptin, periostin, GM-CSF, plasminogen activator inhibitor-1, and osteopontin) and two clinical parameters (recipient body mass index [BMI] and dialysis duration) that distinguished between IF and DGF in the discovery set. Stepwise multivariable logistic regression provided a prediction model on the basis of leptin and GM-CSF. Receiver operating characteristic analysis showed an area under the curve (AUC) of 0.87, and addition of recipient BMI generated a model with an AUC of 0.89, outperforming the Kidney Donor Risk Index and the DGF risk calculator, showing AUCs of 0.55 and 0.59, respectively. CONCLUSIONS We demonstrate that donor kidney preservation fluid harbors biomarkers that, together with information on recipient BMI, predict short-term post-transplantation kidney function. Our approach is safe, easy, and performs better than current prediction algorithms, which are only on the basis of clinical parameters.

Significant Decreasing Incidence of Encapsulating Peritoneal Sclerosis in the Dutch Population of Peritoneal Dialysis Patients

The Dutch Encapsulating Peritoneal Sclerosis (EPS) Registry was started in 2009. Cases were identified by contacting all Dutch nephrologists twice yearly. The predefined criteria for EPS allowed for inclusion of patients with diagnosed and suspected EPS. Cases registered between January 2009 and January 2015 were analyzed with follow-up until September 2015. Fifty-three EPS cases were identified, of which 28.3% were post-transplantation EPS cases. Fourteen patients were initially categorized as suspected EPS, of whom 13 developed EPS. A remarkable 6-fold decrease in the yearly incidence of EPS was observed, from 0.85% in 2009 to 0.14% in 2014. This decrease could not be explained by a decrease in the number of PD patients or average duration of PD treatment in this period. Two-year survival of EPS patients was 52%. The use of tamoxifen and surgical interventions increased significantly over the years. Tamoxifen-treated cases showed a trend to better patient survival and post-transplantation EPS had a significantly favorable outcome. In conclusion, the incidence of EPS has declined significantly in the Netherlands from 2009 to 2014.

CD4-positive T cells and M2 macrophages dominate the peritoneal infiltrate of patients with encapsulating peritoneal sclerosis

Background Encapsulating peritoneal sclerosis (EPS) is a severe complication of peritoneal dialysis (PD). Previously, it has been shown that infiltrating CD4-positive T cells and M2 macrophages are associated with several fibrotic conditions. Therefore, the characteristics of the peritoneal cell infiltrate in EPS may be of interest to understand EPS pathogenesis. In this study, we aim to elucidate the composition of the peritoneal cell infiltrate in EPS patients and relate the findings to clinical outcome. Study Design, Setting, and Participants We studied peritoneal membrane biopsies of 23 EPS patients and compared them to biopsies of 15 PD patients without EPS. The cellular infiltrate was characterized by immunohistochemistry to detect T cells(CD3-positive), CD4-positive (CD4+) and CD8-positive T cell subsets, B cells(CD20-positive), granulocytes(CD15-positive), macrophages(CD68-positive), M1(CD80-positive), and M2(CD163-positive) macrophages. Tissues were analysed using digital image analysis. Kaplan-Meier survival analysis was performed to investigate the survival in the different staining groups. Results The cellular infiltrate in EPS biopsies was dominated by mononuclear cells. For both CD3 and CD68, the median percentage of area stained was higher in biopsies of EPS as opposed to non-EPS patients (p<0.001). EPS biopsies showed a higher percentage of area stained for CD4 (1.29%(0.61-3.20)) compared to CD8 (0.71%(0.46-1.01), p = 0.04), while in the non-EPS group these cells were almost equally represented (respectively 0.28%(0.05-0.83) versus 0.22%(0.17-0.43), p = 0.97). The percentage of area stained for both CD80 and CD163 was higher in EPS than in non-EPS biopsies (p<0.001), with CD163+ cells being the most abundant phenotype. Virtually no CD20-positive and CD15-positive cells were present in biopsies of a subgroup of EPS patients. No relation was found between the composition of the mononuclear cell infiltrate and clinical outcome. Conclusions A characteristic mononuclear cell infiltrate consisting of CD4+ and CD163+ cells dominates the peritoneum of EPS patients. These findings suggest a role for both CD4+ T cells and M2 macrophages in the pathogenesis of EPS.

Effect of a Single Intraoperative High-Dose ATG-Fresenius on Delayed Graft Function in Donation After Cardiac-Death Donor Renal Allograft Recipients : A Randomized Study

OBJECTIVES Reducing the incidence of delayed graft function after transplant with donation after cardiac death donor renal allografts would facilitate managing recipients during their first weeks after a transplant. To reduce this incidence, in most studies, induction therapy with depleting anti-T-lymphocyte antibodies is coupled with a reduction of the dosage of the calcineurin inhibitor. The separate effect of anti-T-cell therapy on the incidence and duration of delayed graft function is therefore difficult to assess. PATIENTS AND METHODS We performed a randomized study to evaluate the effect of a single intraoperative high-dose of anti-T-lymphocyte immunoglobulin (ATG)-Fresenius (9 mg/kg body weight) on the incidence of delayed graft function. Eligible adult recipients of a first donation after cardiac death donor renal allograft were randomly assigned to ATG-Fresenius or no induction therapy. Maintenance immunosuppression consisted of tacrolimus, in an unadjusted dose, mycophenolate mofetil, and steroids. RESULTS The study was prematurely terminated because of a lower-than-anticipated inclusion rate. Baseline characteristics were comparable in the ATG-Fresenius group (n=28) and the control group (n=24). Twenty-two patients in the ATG-Fresenius group (79%) had delayed graft function, compared with 13 in the control group (54%; P = .06). Allograft and patient survival were comparable in both groups. Serious adverse events occurred more frequently in the ATG-Fresenius group than they did in the control group (57% vs 29%; P < .05). CONCLUSIONS Intraoperative administration of a single high-dose of ATG-Fresenius in donation after cardiac death donor renal allograft recipients, followed by triple immunosuppression with an unadjusted tacrolimus dose, seems ineffective to reduce the incidence of delayed graft function. Moreover, this was associated with a higher rate of serious adverse events (EudraCT-number, 2007-000210-36.).

IMPROVED OUTCOME OF ENTERIC PERITONITIS IN PERITONEAL DIALYSIS PATIENTS AGED 50 YEARS AND OLDER WITH TEMPORARY DISCONTINUATION OF PERITONEAL DIALYSIS AND INTRAVENOUS MEROPENEM

♦ Background: Peritonitis is a major cause of morbidity, mortality, and technique failure in peritoneal dialysis (PD) patients, especially when caused by enteric microorganisms (EM). We have implemented a treatment protocol specifically aimed at improving the outcome in EM peritonitis. The adapted protocol was applied in all PD patients 50 years and older presenting with peritonitis who were considered to be at risk of EM peritonitis and involves 3 interventions: 1) temporary discontinuation of PD without removing the catheter (peritoneal rest), 2) intravenous meropenem, and 3) meropenem intracatheter as lock (Mero-PerRest protocol). ♦ Methods: In this observational study, we compared the outcome of 203 peritonitis episodes in 71 patients treated with the Mero-PerRest protocol, with 217 episodes in 104 patients treated with a more traditional intraperitoneal gentamicin-rifampicin-based regimen. ♦ Results: In EM peritonitis episodes, the Mero-PerRest protocol resulted in a higher primary cure rate (90.0% vs 65.3%, adjusted odds ratio [OR] 4.54 [95% confidence interval (CI) 1.46 – 14.15]) and better technique survival (90.0% vs 69.4%, adjusted OR 3.41 [95% CI 1.07 – 10.87]). This effect was most distinct in patients with polymicrobial EM peritonitis (cure rate 87.5% vs 34.8%, p = 0.0003). Interestingly, primary cure rate (95.6% vs 84.7%, adjusted OR 3.92 [95% CI 1.37 – 11.19]) and technique survival (95.6% vs 85.6%, adjusted OR 3.60 [95% CI 1.25 – 10.32]) were also excellent in non-EM peritonitis episodes. Patient survival did not differ significantly. ♦ Conclusion: The poor outcome of peritonitis caused by EM in PD patients aged 50 years and older could be improved by applying a treatment protocol involving temporary discontinuation of PD without catheter removal and intravenous and intracatheter meropenem.

Encapsulating peritoneal sclerosis is associated with T-cell activation

BACKGROUND Encapsulating peritoneal sclerosis (EPS) is an excessive fibrotic response of the peritoneum that may occur after long-term peritoneal dialysis (PD). The underlying pathophysiology is poorly understood, but involvement of peritoneal inflammatory T helper 1 cells may be pivotal. METHODS Soluble interleukin-2 receptor alpha (sCD25) concentration was measured as a marker for T-cell activation in serum and ascites from EPS patients and various control patient groups. Peritoneal biopsies were stained for the presence of T cells, and T cells isolated from ascites of EPS patients were characterized in detail for differentiation status and cytokine expression. RESULTS Serum sCD25 concentrations are significantly and specifically increased in EPS patients compared with haemodialysis, PD and predialysis patients. Peritoneal effluent of stable PD patients contains very low levels of sCD25, while sCD25 levels in ascites of EPS patients are high and indicative of local production. In the years preceding the diagnosis of EPS, the serum sCD25 concentrations increased while remaining at stable levels in control PD patients. The peritoneum and ascites of EPS patients showed a significant influx of T cells with relatively increased numbers of CD4(+) T cells. These T cells were fully differentiated and displayed a T helper 1 cell type with a pro-inflammatory cytokine profile. CONCLUSIONS Increased serum sCD25 concentrations and peritoneal lymphocytosis in EPS patients indicate the involvement of activated T cells in the pathophysiology of excessive fibrosis.