A two-hit approach in the development of an experimental peritoneal sclerosis model.

Models of encapsulating peritoneal sclerosis (EPS) are often based on local administration of chemical irritants. Our aim was to develop a clinically relevant "two-hit" model with incorporation of renal failure and exposure to conventional dialysis solutions. We randomly allocated 36 male Wistar rats that had undergone catheter implantation and a 70% nephrectomy to 3 peritoneal infusion groups. The experimental group was exposed to a 3.86% glucose-based conventional dialysis solution for 8 weeks, after which the animals received a second hit of intraperitoneal blood administration. Two weeks later the rats were humanely euthanized The two control groups were exposed to the conventional dialysis solution alone or to a buffer without glucose for 8 weeks. All animals underwent a peritoneal function test at the end of the experiment. Peritoneal adhesions were counted at autopsy, and omental tissue was obtained for morphometrics. The rats that received blood as a second hit developed numerous intraperitoneal adhesions as seen in EPS, but without cocoon formation. Microscopically, no differences in fibrosis scores and vessel counts were observed between the groups. Peritoneal function parameters were also similar in all groups. The short infusion period could be the reason that we found no differences between the groups, with the exception of the large amount of intraperitoneal adhesions in the experimental group. Modifications to the described rat model are required to develop a clinically relevant EPS model. Besides renal failure and long-term exposure to bioincompatibleperitoneal dialysis solutions, a different second hit or several additional hits could be incorporated into an experimental model of EPS.

Are peritoneal calcifications in long-term peritoneal dialysis related to aortic calcifications and disturbances in mineral metabolism?

BACKGROUND: Peritoneal calcifications are associated with long-term peritoneal dialysis (PD). Case reports have suggested a relation with disturbances in mineral metabolism such as the presence of severe hyperparathyroidism. Our aim was to investigate whether relationships are present between peritoneal calcifications and aortic calcifications or disturbances in mineral metabolism in long-term PD patients. METHODS: We included all long-term PD patients (PD ≥ 4 years) in our centre from 1996 to 2008 who had undergone an abdominal computed tomographic (CT) scan. The scans were reviewed by two experienced radiologists in consensus. The presence or absence of peritoneal calcifications was scored, and a severity scoring system for abdominal aortic calcifications was used: 1 = none, 2 = mild, 3 = moderate, 4 = severe and 5 = very severe. For each patient, laboratory data on plasma calcium corrected for albumin, phosphorus and parathyroid hormone (PTH) levels were retrieved every 6 months up to 5 years prior to the CT scan. Individual mean values over 5 years were calculated. RESULTS: We included 31 patients: 12 patients with peritoneal calcifications and 19 patients without. No difference was found in aortic calcification scores (median scores: 3 versus 3). Also, median (range) calcium, 10.7 (9.6-11.5) versus 10.3 (9.4-11.3) mg/dL; phosphorus, 5.2 (3.4-7.0) versus 4.9 (2.9-6.5) mg/dL; and PTH levels, 271 (101-910) versus 263 (40-1197) pg/mL were not different between patients with and without peritoneal calcifications. CONCLUSIONS: The presence of peritoneal calcifications in long-term PD patients could not be related to the presence of aortic calcifications or disturbances in mineral metabolism. Perhaps, local peritoneal factors play a role in the formation of peritoneal calcifications.

Experimental peritoneal sclerosis models should not be based on chlorhexidine gluconate anymore.

BACKGROUND/AIMS: Currently available rodent models of peritoneal sclerosis are not based on clinically relevant factors: renal failure in combination with exposure to bioincompatible fluids. Our aim was to develop a chronic peritoneal infusion model of peritoneal sclerosis in rats with renal failure. METHODS: Male Wistar rats underwent a catheter implantation and a 70% nephrectomy. They were randomly divided into three peritoneal infusion groups: chlorhexidine gluconate/ethanol (CGE) + Dianeal (Baxter Healthcare, Castlebar, Ireland), CGE + buffer (Physioneal without glucose; Baxter, Nivelles, Belgium) and Dianeal alone. After 8 weeks a peritoneal permeability test was performed and omental tissue was obtained for morphometrics. RESULTS: The CGE + Dianeal group (n = 6) and CGE + buffer (n = 6) group showed high peritoneal clearances of small solutes and proteins, ultrafiltration failure, impaired free water transport, severe fibrosis and high vessel counts, but the groups did not differ significantly. The Dianeal group (n = 6) showed significantly lower clearances of small solutes and proteins, normal ultrafiltration and sodium sieving, and significantly lower fibrosis scores and vessel counts. CONCLUSIONS: Abnormalities seen in peritoneal sclerosis can be induced in a peritoneal infusion model in rats with renal failure. However, the addition of a bioincompatible dialysis solution had no contributing role, probably because the effects were overruled by those of CGE.

Does Alport syndrome affect the basement membrane of peritoneal vessels?

Alport syndrome and encapsulating peritoneal sclerosis (EPS) are both rare diseases. Their joint occurrence is highly unlikely. Two patients at our center with Alport syndrome developed EPS. We therefore hypothesized that Alport syndrome might predispose to the development of EPS and that this predisposition might be reflected in a fast peritoneal transport rate at baseline. We compared the mass transfer area coefficient (MTAC) of creatinine and the clearances of albumin, immunoglobulin G, and alpha2-macroglobulin at baseline and for all subsequent available measurements in four patient groups: EPS patients with Alport syndrome, EPS patients without Alport syndrome, Alport patients without EPS, and long-term peritoneal dialysis (PD) patients without EPS. The transport characteristics were obtained during a standard peritoneal permeability analysis. Between July 1995 and December 2008, 5 of 417 PD patients treated at our center had Alport syndrome as their primary kidney disease, and 13 of the 417 developed EPS. Of those 13 EPS patients, 2 had Alport syndrome. We observed no differences in the baseline transport characteristics of the four groups under consideration. Taking all measures of transport characteristics into account, only the MTAC of creatinine was higher in the two EPS groups than in the other two groups (p = 0.01). We could not confirm our hypothesis that Alport syndrome affects peritoneal solute clearances.

Addition of a nitric oxide inhibitor to a more biocompatible peritoneal dialysis solution in a rat model of chronic renal failure.

Biocompatible dialysis solutions have been developed to preserve peritoneal membrane morphology and function. Compared with a conventional solution, a combination of glycerol, amino acids, and dextrose in a bicarbonate/lactate buffer (GLAD) led to less peritoneal fibrosis and fewer vessels in a chronic peritoneal exposure model in the rat. However, no concomitant reduction in small-solute transport was observed. We hypothesized that this result could be attributable to peritoneal vasodilation induced by vasoactive substances such as nitric oxide. The aim of the present study was to investigate whether fast transport of small solutes and proteins induced by exposure to GLAD could be influenced by Ngamma -methyl-L-arginine acetate (L-NMMA), an inhibitor of NO. These investigations used our rat model of long-term peritoneal exposure with chronic renal failure. All rats underwent peritoneal catheter implantation and a 70% nephrectomy. Thereafter, the rats were allocated to 3 groups: 16 weeks of peritoneal exposure to GLAD and L-NMMA, to GLAD only, or to buffer (bicarbonate/lactate without any osmotic agent). Afterward, a standard peritoneal permeability analysis adjusted for the rat was performed. Subsequently, the rats were euthanized, and tissue samples were obtained for morphometric determinations. No effect of L-MNNA on the transport of small solutes and proteins was found. Also, no effect on morphology was found. Our findings make it unlikely that NO is directly involved, being more in favor of a direct effect of amino acids on peritoneal transport.

Computed tomographic findings characteristic for encapsulating peritoneal sclerosis: a case-control study.

BACKGROUND: Computed tomography (CT) is often used to confirm the diagnosis of encapsulating peritoneal sclerosis (EPS) but there is no consensus on specific CT abnormalities. To establish CT findings characteristic for EPS, we compared CT findings between EPS patients and long-term peritoneal dialysis (PD) patients without EPS. METHODS: We included as cases all EPS patients in our center from 1996 to 2008 that underwent a CT scan at the time of diagnosis. Controls were all other long-term PD patients (PD duration > or = 4 years) without EPS that had a CT scan for different reasons. The CT scans were blindly and independently reviewed by 3 radiologists: 2 abdominal radiologists with PD knowledge (Observers 1 and 2) and 1 radiologist without PD experience (Observer 3). RESULTS: We included 15 EPS patients and 16 controls. Observer 1 found 6 CT findings that were significantly more often present in EPS than in controls (p < or = 0.05): peritoneal enhancement, thickening, and calcifications; adhesions of bowel loops; signs of obstruction; and fluid loculation/septation. Observer 2 scored almost identically but Observer 3 scored differently. The sensitivity and specificity of a combination of specific CT findings were, respectively, 100% and 94% for Observers 1 and 2, and 79% and 88% for Observer 3. CONCLUSION: CT scans showed characteristic abnormalities that were significantly more often present in EPS patients compared to long-term PD control patients. CT can be used to confirm the diagnosis of EPS when experienced radiologists apply a combination of specific CT findings.

Experimental study on long-term exposure to a biocompatible, hypertonic, pyruvate-buffered dialysis solution.

BACKGROUND: Chronic exposure to glucose and glucose degradation products (GDPs) in dialysis solutions is involved in the pathogenesis of peritoneal neoangiogenesis and fibrosis, potentially leading to encapsulating peritoneal sclerosis (EPS). High lactate concentrations may contribute to glucose toxicity by creating a state of pseudohypoxia, which stimulates the formation of various growth factors. OBJECTIVE: To study the effects of long-term peritoneal exposure to a filter-sterilized pyruvate-buffered solution with a combination of 3 osmotic agents (amino acids, glycerol, glucose: PYRAGG) on peritoneal function and morphology. METHODS: Rats were exposed daily for a period of 20 weeks to PYRAGG, or to a conventional heat-sterilized solution (LH), or to a filter-sterilized solution (LF), after which a peritoneal function test was done and peritoneal tissue was obtained. RESULTS: Peritoneal solute and fluid transport characteristics at 20 weeks were similar in all groups. Fibrosis was most pronounced in the LH group compared to the others, suggesting an effect of GDPs. A marked reduction in the number of omental vessels was noted in the PYRAGG group (59% reduction compared to LH). A modest reduction (28%) was found in the LF animals. This points to a marked effect of reduced exposure to glucose. CONCLUSIONS: PYRAGG was more biocompatible than a filter-sterilized glucose/lactate solution because it did not induce marked peritoneal abnormalities after long-term exposure. This did not lead to altered peritoneal transport characteristics. It is likely that further development of PYRAGG-like solutions will decrease the incidence of EPS.

Identification of Gene Transcripts Implicated in Peritoneal Membrane Alterations.

♦ BACKGROUND: Permanent stimulation of the peritoneum during peritoneal dialysis (PD) is likely to result in increased expression of genes encoding proteins involved in inflammation and tissue remodeling. Peritoneal fibrosis and neoangiogenesis may develop. ♦ OBJECTIVE: To assess highly expressed genes potentially in volved in peritoneal alterations during PD treatment using an animal model. ♦ METHODS: A PD catheter was implanted in 36 male Wistar rats after 70% nephrectomy. The rats were divided into 3 groups, exposed to dialysis solution for 8 weeks, and sacrificed 2 weeks later. Group B was exposed to a buffer, group D was exposed to a 3.86% glucose-based dialysis solution, and in group D+H, a second hit of intraperitoneal blood on top of the dialysis solution was given to induce the development of peritoneal sclerosis. Before sacrifice, peritoneal function was assessed. Omental tissue was obtained for analysis of gene expression using RT-qPCR. ♦ RESULTS: Fibrosis scores, vessel counts, and peritoneal function parameters were not different between the groups. Genes involved in the transforming growth factor beta signaling pathway, cell proliferation, angiogenesis, and inflammation were more expressed (p < 0.05) in the D+H group. Almost no differences were found between the control groups. We identified 4 genes that were related to peritoneal transport. ♦ CONCLUSION: Already a mid-term peritoneal exposure, when no microscopical and functional alterations are present, provokes activation of gene pathways of cell proliferation, fibrosis, neoangiogenesis, and inflammation.

Imaging in encapsulating peritoneal sclerosis.

Encapsulating peritoneal sclerosis (EPS) is a rare but very severe complication of long-term peritoneal dialysis (PD). Since the first reports on this disease in the eighties, several imaging techniques have been used for its diagnosis. Because of the rarity of this condition, uniformity in modality and protocols for abdominal imaging for diagnosis has been lacking overtime. Nowadays, computed tomography (CT) is most often used. In this review, we provide an overview of all imaging modalities that have been used overtime to diagnose EPS as a late complication of PD. Imaging features characteristic for EPS and advantages as well as shortcomings of all modalities are discussed. We believe that when EPS is suspected, CT with contrast enhancement should be the modality of first choice in clinical practice.

Early diagnostic markers for encapsulating peritoneal sclerosis: a case-control study.

OBJECTIVE: Encapsulating peritoneal sclerosis (EPS) is a severe complication of long-term peritoneal dialysis (PD). The aim of this study was to investigate whether dialysate levels of cancer antigen-125 (CA125), K(+), interleukin (IL)-6, and vascular endothelial growth factor (VEGF) are early diagnostic markers of EPS. Therefore, we analyzed the time courses of the above described dialysate markers in EPS patients and controls. METHODS: Dialysate and serum samples of 11 EPS patients and 31 control patients, all treated with PD for at least 57 months, were longitudinally collected during standard peritoneal permeability analyses. CA125 and IL-6 were measured in dialysate only, K(+) and VEGF were measured in both dialysate and serum. CA125 and IL-6 are expressed as appearance rates (AR). The linear mixed model was used to analyze the time courses. Sensitivity and specificity were calculated based on the results of the last 2 time points. RESULTS: No differences in the time courses of the different markers were present between the groups. For K(+) and VEGF attributed to local production, no differences between the groups were found. However, AR-CA125 was lower during the last 3 years prior to EPS (p < 0.05) and AR-IL-6 tended to be higher 2 years prior to EPS (p = 0.09). The combination of AR-CA125 < 33 U/min and AR-IL-6 > 350 pg/min had a sensitivity of 70% and a specificity of 89% for the development of EPS. CONCLUSIONS: Compared to controls, AR-CA125 showed lower values and AR-IL-6 tended to be higher during the last years prior to the diagnosis of EPS. The sensitivity and specificity of the combination of CA125 and IL-6 indicate their potential use for an early diagnosis of EPS.

The effects of a dialysis solution with a combination of glycerol/amino acids/dextrose on the peritoneal membrane in chronic renal failure.

BACKGROUND: Long-term peritoneal dialysis (PD) with conventional glucose based, lactate-buffered PD fluids may lead to morphological and functional alterations of the peritoneal membrane. It was hypothesized that long-term exposure to a different buffer and a mixture of osmotic agents would cause less peritoneal abnormality. OBJECTIVES: To investigate the effects of long-term exposure to a bicarbonate/lactate-buffered dialysis solution with a mixture of osmotic agents: glycerol 1.4%, amino acids 0.5%, and dextrose 1.1% (= 1% glucose) (GLAD) in a rat model with chronic kidney failure. METHODS: All rats underwent a peritoneal catheter implantation and a 70% nephrectomy. Thereafter, the rats were randomly divided into 3 groups: GLAD, 3.86% Dianeal (Baxter, Nivelles, Belgium), and buffer (Physioneal without glucose, Baxter). All rats were infused daily for 16 weeks with the appropriate PD fluid. Afterwards, a peritoneal permeability analysis (SPARa) was performed using 3.86% Physioneal in all groups. After the SPARa, the rats were sacrificed to obtain tissue samples for morphometric determinations. Omental tissue was stained with picro Sirius red for assessment of fibrosis and with CD31 for vessel density. RESULTS: GLAD and Dianeal showed faster small solute transport compared to the hypotonic buffer. No differences between the groups were present in ultrafiltration. Dianeal had the lowest value for free water transport and the highest protein clearances. Total triglyceride in plasma was not different between GLAD and the buffer. Vessel density after GLAD exposure (20 V/F) was very similar to the value found for the buffer solution (17 V/F); Dianeal caused a significantly higher value (35 V/F, p < 0.01). Also, the amount of fibrosis was higher in the Dianeal-exposed rats (p < 0.01). CONCLUSION: Both hypertonic dialysis solutions increased peritoneal solute transport. GLAD exposure was associated with the best preservation of peritoneal morphology. The results of GLAD were very similar to those of the bicarbonate/lactate-buffered solution without osmotic agents. Studies in humans are needed for further assessment of GLAD.

Biological markers in the peritoneal dialysate effluent: are they useful.

A review is given on biomarkers in peritoneal effluent. It comprises methods to distinguish between diffusion and local production. This is followed by examples of various biomarkers. Their potential use is discussed in 4 situations: inherent fast transporters, longitudinal follow-up of patients, biocompatibility testing of new dialysis solutions, and their potential use in the detection of patients who are likely to develop encapsulating peritoneal sclerosis.