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Increased survival of mesothelial cells from the peritoneum in peritoneal dialysis fluid
1Division of Nephrology, Department of Medicine, Queen Mary Hospital, University of Hong Kong, Pokfulam, Hong Kong. knlai@hkucc.hku.hk
Abstract:
Peritonitis remains the most important factor in patient morbidity and technical failure associated with continuous ambulatory peritoneal dialysis (CAPD). In vitro examination of bacterial infection of cultured human peritoneal mesothelial cells (HPMC) is an attractive approach to the study of peritonitis in CAPD, yet there are few reports on this subject. Previous studies have shown two limitations: (i) cell cultures of HPMC lasted for days only when incubated in culture medium and (ii) short-term studies of <30 min were done in HPMC when incubated with peritoneal dialysis fluid (PDF). Human peritoneal mesothelial cells, maintained in a conventional single chamber culture system with PDF alone, were unable to survive more than 40 min. The present study was designed to prolong the viability of HPMC cultured in PDF, with the object of using cells under different conditions, such as that of simulating CAPD. HPMC were cultured using plastic microtiter plates, where they were grown to confluence and growth was arrested. PDF containing different concentrations of NaHCO3and human serum albumin was added. Cell viability after exposure for up to 24 h was measured by trypan blue, Cell Death Detection ELISA and Annex-V flow cytometry. The data confirmed the 'toxic' effect of PDF, with cell viability being <40% after 2 h incubation in 4.25% glucose in PDF. However, the survival time of HPMC increased significantly in 4.25% glucose PDF at a physiological pH and even further after the addition of human albumin. These experimental conditions simulating CAPD may allow future in vitro studies of mesothelial physiology and peritonitis related to CAPD treatment.
Insights
Peritonitis in continuous ambulatory peritoneal dialysis (CAPD) is a major concern. This study improved human peritoneal mesothelial cell (HPMC) viability in peritoneal dialysis fluid (PDF), enabling better in vitro peritonitis research.
Area of Science:
- Nephrology
- Cell Biology
- Biomedical Engineering
Background:
- Peritonitis is a significant complication in continuous ambulatory peritoneal dialysis (CAPD), leading to patient morbidity and treatment failure.
- Previous in vitro studies of human peritoneal mesothelial cells (HPMC) in peritoneal dialysis fluid (PDF) were limited by short cell survival times.
- Developing robust in vitro models is crucial for understanding CAPD-related peritonitis.
Purpose of the Study:
- To enhance the viability of HPMC cultured in PDF to simulate conditions relevant to CAPD.
- To establish an improved in vitro model for studying mesothelial cell responses to peritonitis in CAPD.
Main Methods:
- Human peritoneal mesothelial cells (HPMC) were cultured to confluence in microtiter plates and growth-arrested.
- Cells were exposed to PDF with varying concentrations of NaHCO3 and human serum albumin.
- Cell viability was assessed using trypan blue, Cell Death Detection ELISA, and Annex-V flow cytometry over 24 hours.
Main Results:
- Peritoneal dialysis fluid (PDF) demonstrated a toxic effect on HPMC, with viability below 40% after 2 hours in 4.25% glucose PDF.
- HPMC survival significantly increased in 4.25% glucose PDF at physiological pH.
- Addition of human serum albumin further prolonged HPMC survival in the simulated CAPD conditions.
Conclusions:
- Optimized PDF formulations and physiological conditions can significantly extend HPMC viability in vitro.
- This improved model allows for extended in vitro investigation of mesothelial cell behavior during CAPD-related peritonitis.
- The findings facilitate future research into CAPD peritonitis mechanisms and therapeutic strategies.