Osmotic agents hamper mesothelial repopulation as seen in the doughnut in vivo model.

BACKGROUND: The problem of mesothelial cell injury derived from the use of peritoneal dialysis solutions has been explored deeply. Conversely, the eventual detrimental effects upon mesothelial cell regeneration have awaked less investigative efforts than those focused on injury. OBJECTIVE: To evaluate in the in vivo and in situ rat "doughnut" model of mesothelial repopulation, the eventual effect of peritoneal lavage with Hank's Balanced Salt Solution (HBSS) as well as that of 4.25% glucose and 7.5% icodextrin dialysis solutions. EXPERIMENTAL ANIMALS: 100 Sprague-Dawley albino rats were included in the study. Animals were divided into five groups of 20 rats each: group 1: control at zero time; group 2: sham-injected rats; group 3: rats exposed to HBSS; group 4: rats treated with 4.25% glucose peritoneal dialysis solution; group 5: rats injected with 7.5% icodextrin. METHODS: Selective exfoliation of a ring of mesothelium (width 0.8 mm, diameter 4 mm) covering the anterior surface of the liver was performed in 80 animals. The control zero-time group was used to evaluate the normal density distribution of the mesothelial cells forming the monolayer. The other groups were treated by means of daily sham injections or intraperitoneal infusion of each experimental solution for a period of 30 consecutive days. After a recovery period of 15 days, imprints and biopsies from the monolayer covering the exfoliated area were taken and processed for light microscopy. RESULTS: Macroscopic observation of the abdominal cavity at the end of the 15-day recovery period showed that the prevalence of fibrotic adhesions between the peritoneal exfoliated area and the neighboring diaphragm was 10% forthe sham-injected group, 5% for the HBSS-exposed animals, 85% for the rats injected with 4.25% glucose, and 95% for the icodextrin-treated group. Prevalence of fibrous adhesions in sham-injected animals and rats exposed to HBSS were devoid of statistically significant differences. Conversely, comparison of these groups with results observed in animals treated with the osmotic agents was significant, at the p < 0.0039 level. Regarding density distribution of mesothelial cells observed in imprints, there were no significant differences between the control zero-time and the sham-injected group. This parameter was marginally lower (p < 0.05) in the HBSS-treated rats. Imprints were not taken from animals exposed to glucose or icodextrin because a dense layer of connective tissue replaced the exfoliated mesothelial area. CONCLUSIONS: Observations made in this study support the contention that both osmotic agents, 4.25% glucose and 7.5% icodextrin, substantially restrain the normal process of mesothelial cell repopulation and induce repair by means of connective tissue. The underlying mechanism is most likely sustained oxidative stress.

Icodextrin-induced lipid peroxidation disrupts the mesothelial cell cycle engine.

Fluids commonly used for peritoneal dialysis hold poor biocompatibility vis a vis the peritoneal membrane, basically due to the presence of osmotic agents. When rat mesothelium was exposed to glucose-enriched dialysis solutions for 2 h in vivo, an early and short-lived acceleration of cell life cycle was observed, which, after 30 d of exposure, resulted in a depopulated monolayer of senescent cells. These changes appear to result from persistent oxidative stress due to continuous exposure to high concentration of glucose and to substances generated by the Maillard reaction. Long-term exposure (30 d) of the peritoneal mesothelium to 7.5% icodextrin resulted in a depopulated monolayer consisting mostly of senescent cells, which, additionally, showed atypical nuclear changes and atypical mitosis suggesting DNA damage. These changes coincided with substantial lipid peroxidation, starting immediately after the introduction of the icodextrin solution into the rat's abdominal cavity. So far, the currently used osmotic agents in peritoneal dialysis fluids induce substantial oxidative injury to the exposed monolayer in vivo. Use of high concentrations of glucose results in premature senescence of the exposed cell population. The 7.5% icodextrin dialysis fluid induces through lipid peroxidation substantial genomic damage, which, in turn, sets the biological mechanisms leading to protective cellular suicide in motion.

Repopulation of the mesothelial monolayer during long-term experimental peritoneal dialysis.

BACKGROUND: Repopulation of the mesothelial monolayer after focal exfoliation, having the monolayer in vivo and in situ exposed to dialysis solutions, has not been thoroughly investigated. This study describes repopulation of a 'doughnut' like mesothelial ring exfoliated from the anterior liver surface of rats. METHODS: Animals were divided into 5 groups of 20 rats each. Group 1 - control unexposed animals: mesothelial cell imprints were taken after 1 (5 rats), 5 (5 rats), and 15 (10 rats) days following the procedure of exfoliation. Group 2 - sham injected animals. Group 3 - rats IP injected once a day, during 30 consecutive days with Hank's balanced salt solution. Groups 4 and 5 - same experimental protocol, but injecting 4.25% glucose single bag or 7.5% Icodextrin PDF. Imprints and/or biopsies were taken after a recovery period of 15 days, counted from the last IP injection. RESULTS: Density distribution of mesothelial cells in group 2 was not significantly different from that seen in unexposed rats, whereas that seen in group 3 Hank's balanced salt solution was marginally but significantly lower (p < 0.05) from that seen in controls. Eighty five percent of rats injected with 4.25% glucose developed fibrous adhesions, binding up together the exfoliated liver surface and the diaphragmatic muscle. For Icodextrin treated rats, the prevalence of fibrous adhesions was 95%. CONCLUSIONS: The 'doughnut' experimental model appears as a promising tool for in vivo and in situ investigation of mesothelial repopulation. Both osmotic agents substantially restrain mesothelial repopulation, leading to repair by connective tissue.

Acute oxidative stress induces peritoneal hyperpermeability, mesothelial loss, and fibrosis.

We explored the acute and long-term effects of short-lived, intense oxidative stress on peritoneal permeability and structure, induced with intraperitoneal injection of the oxidant agent deoxycholate, in rats. Ten minutes after the experimental intervention, peritoneal dialysis, performed over an exposure time of 60 minutes, revealed an increased urea dialysate/plasma ratio, greater glucose absorption, increased albumin losses in the effluent dialysate, and a reduced ultrafiltration rate. Mesothelial-cell imprints taken from the anterior liver surface indicated a substantially decreased density in the cell population. After the recovery period of 30 days, all alterations were still evident. Additionally, macroscopic and histologic observations made at this time interval detected peritoneal fibrosis and sclerosis, characterized by peritoneal adhesions, wrapping of intestinal loops, and the presence of a layer of fibrous tissue dressing the cavitary aspect of the liver peritoneal envelope. This report describes a reproducible experimental model of peritoneal fibrosis induced by acute oxidative injury. On the basis of these findings, it may be speculated that functional and structural alterations observed in patients are related to long-term continuous exposure of the monolayer to oxidative injury resulting from the high concentrations of d-glucose present in peritoneal dialysis solutions.

Peritoneal transport after long-term exposure to Icodextrin in rats.

BACKGROUND: Icodextrin, an effective osmotic substance that has been proposed as an alternative agent for peritoneal dialysis induces ultrafiltration over long dwells. This study examines the peritoneal transport after exposure to Icodextrin in rats. METHODS: Animals were divided in 4 groups and injected daily for 30 days with Icodextrin 7.5 % (n = 14), Glucose 4.25 % (n = 19) or glucose 4.25% plus Icodextrin 7.5 % (n = 13). Rats of the control group (n = 15) were not exposed. A 4-hour permeability study was performed using glucose at days 1, 30 and 60. At days 2, 31 and 61 the same animals were injected with Icodextrin. RESULTS: Slopes of effluent sodium at day 30 were significantly higher (p < 0.001) in the glucose (0.006 +/- 0.016), Icodextrin (0.013 +/- 0.014) and mixed groups (0.012 +/- 0.017) than in the control group (-0.041 +/- 0.021). Urea D/P ratio was not significantly different in the 4 groups. After 30 days, glucose effluent levels were significantly lower (p < 0.001) in the glucose (701 +/- 278 mg/dl), Icodextrin (552 +/- 209 mg/dl) and mixed groups (587 +/- 344 mg/dl) than in control rats (1519 +/- 413 mg/dl). Effluent protein (mg/l) in the mixed group (1,555 +/- 357) was significantly higher (p < 0.001) than control (376 +/- 33), glucose (1,015 +/- 232) and Icodextrin (765 +/- 75) groups at day 30. CONCLUSION: The long-term use of Icodextrin does not affect small molecule transport, but induces changes in the peritoneal protein excretion, especially when Icodextrin and glucose are injected together.