Isolation and Propagation in Vitro of Peritoneal Mesothelial Cells

Abstract

Mesothelial cells lining the peritoneal cavity are the primary site of molecular exchange during peritoneal dialysis, a life support system for over 50 000 patients worldwide. In this study, techniques are described for the isolation and propagation in culture of peritoneal mesothelial cells from rats and rabbits. For comparison, mesothelial cells were also obtained from the serosal surface of human colonic tissue. By electron microscopy the cultured cells were found to exhibit microvilli, a well developed endoplasmic reticulum and golgi apparatus, micropinocytotic vesicles, and lipid-filled intracellular vesicles. Immunochemical probes revealed the expression by these cells in vitro of cytokeratin, fibronectin, vimentin, and keratin, but not von Willebrand factor. Mesothelial cells from rat, rabbit, and human exhibited contact inhibition, but differences in growth rates and dependence on supplements to the growth media. This work provides a multispecies comparison of the behavior of mesothelial cells in vitro for the purpose of developing an experimental system for the study of mesothelial cell biology and the role of these cells in peritoneal dialysis.

Cultured peritoneal mesothelial cells exhibit apical primary cilia

“…Cultured PMC retain a biphasic growth pattern in cell culture, with a reversible transition from a mesenchymal-PMC (M-PMC) phenotype (Fig. 2) to an epithelial-PMC (E-PMC) phenotype as the monolayer is established (Hjelle et al, 1989;Marshall et al, 1993). This transition is characterized by the reorganization of the cytoskeleton and the expression of cytokeratin intermediate filaments as progression of the epithelial phenotype proceeds (Connell and Rheinwald, 1983;Wu et al, 1982).…”

“…This study was approved by the University of Otago ethics committee. Individual specimens were subjected to enzymatic disaggregation using either 0.05% trypsin, 0.02% EDTA (Pronk et al, 1993;Stylianou et al, 1990) or collagenase type II (Hjelle et al, 1989) to isolate hPMC or rPMC. Cells were seeded into 75 cm 2 Falcon plastic flasks (Becton Dickinson, NJ, USA) at 5.0 10 4 cell/cm 2 and incubated with Medium 199 (Gibco BRL, Life Technologies, Auckland, NZ) supplemented with 10% heat inactivated foetal bovine serum (FBS), 2.0 mM GlutaMAX-I (Gibco BRL, Life Technologies, Auckland, NZ), 0.25 mM (30.0 µg/ml) -cysteine, 50 µU/ml penicillin and 50 µg/ml streptomycin, and incubated at 37 (C in a humidified atmosphere of 5% CO 2 :95% air (Bird et al, 1996).…”

LPA1‐induced cytoskeleton reorganization drives fibrosis through CTGF‐dependent fibroblast proliferation

“…Primary PMCs were isolated from mice by enzymatic digestion of the inner surface of the peritoneum, as described previously (30), with minor modifications. Briefly, parietal peritoneal flaps were removed and stretched on a sterile culture dish.…”

Human Prostate DU145 Carcinoma Cells Implanted in Nude Mice Remove the Peritoneal Mesothelium to Invade and Grow as Carcinomas

“…The mesothelial surfaces in the nude mice are similar to those in humans (Leak and Rahil, 1978; Slater et al, 1989; Jonecko, 1990), other animals including mice (Carr et al, 1969; Watters and Buck, 1972; Schwarz, 1974; Baradi and Rao, 1976; Guggenheim et al, 1979; Hjelle et al, 1989; Gaudio et al, 1990; Ettarh and Carr, 1996; Bot et al, 2003; Michailova and Usunoff, 2006; Yung et al, 2006; Yung and Chan, 2007). The epithelial cells show secretory activity with the apical release of lamellar bodies analogous to those of alveolar or pneumocytes type II (Dobbie and Lloyd, 1989; Dobbie and Anderson, 1996a,b) as well as excrescences and expelled material similar to those observed in the gallbladder epithelium (Gilloteaux et al, 1993).…”