Rosiglitazone, a Peroxisome Proliferator-Activated Receptor (PPAR)-γ Agonist, Attenuates Inflammation Via NF-κB Inhibition in Lipopolysaccharide-Induced Peritonitis.

We assessed the anti-inflammatory effect of peroxisome proliferator-activated receptor (PPAR)-γ agonist, rosiglitazone, in a lipopolysaccharide (LPS)-induced peritonitis rat model. LPS was intraperitoneally injected into rats to establish peritonitis model. Male Sprague-Dawley (SD) rats were assigned to normal saline (the solvent of LPS), LPS, rosiglitazone plus LPS, and rosiglitazone alone. A simple peritoneal equilibrium test was performed with 20 ml 4.25 % peritoneal dialysis fluid. We measured the leukocyte count in dialysate and ultrafiltration volume. Peritoneal membrane histochemical staining was performed, and peritoneal thickness was assessed. CD40 and intercellular adhesion molecule-1 messenger RNA (ICAM-1 mRNA) levels in rat visceral peritoneum were detected by reverse transcription (RT)-PCR. IL-6 in rat peritoneal dialysis effluent was measured using enzyme-linked immunosorbent assay. The phosphorylation of NF-κB-p65 and IκBα was analyzed by Western blot. LPS administration resulted in increased peritoneal thickness and decreased ultrafiltration volume. Rosiglitazone pretreatment significantly decreased peritoneal thickness. In addition to CD40 and ICAM-1 mRNA expression, the IL-6, p-p65, and p-IκBα protein expressions were enhanced in LPS-administered animals. Rosiglitazone pretreatment significantly decreased ICAM-1 mRNA upregulation, secretion of IL-6 protein, and phosphorylation of NF-κB-p65 and IκBα without decreasing CD40 mRNA expression. Rosiglitazone has a protective effect in peritonitis, simultaneously decreasing NF-κB phosphorylation, suggesting that NF-κB signaling pathway mediated peritoneal inflammation induced by LPS. PPAR-γ might be considered a potential therapeutic target against peritonitis.

A20 overexpression inhibits lipopolysaccharide-induced NF-κB activation, TRAF6 and CD40 expression in rat peritoneal mesothelial cells.

Zinc finger protein A20 is a key negative regulator of inflammation. However, whether A20 may affect inflammation during peritoneal dialysis (PD)-associated peritonitis is still unclear. This study was aimed to investigate the effect of A20 overexpression on lipopolysaccharide (LPS)-induced inflammatory response in rat peritoneal mesothelial cells (RPMCs). Isolated and cultured RPMCs in vitro. Plasmid pGEM-T easy-A20 was transfected into RPMCs by Lipofectamine™2000. The protein expression of A20, phospho-IκBα, IκBα, TNF receptor-associated factor (TRAF) 6 and CD40 were analyzed by Western blot. The mRNA expression of TRAF6, CD40, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were determined by real time-PCR. NF-κB p65 DNA binding activity, IL-6 and TNF-α levels in cells culture supernatant were determined by ELISA. Our results revealed that RPMCs overexpression of A20 lead to significant decrease of LPS-induced IκBα phosphorylation and NF-κB DNA binding activity (all p<0.01). In addition, A20 also attenuated the expression of TRAF6, CD40, IL-6 and TNF-α as well as levels of IL-6 and TNF-α in cells culture supernatant (all p<0.05). However, A20 only partly inhibited CD40 expression. Our study indicated that A20 overexpression may depress the inflammatory response induced by LPS in cultured RPMCs through negatively regulated the relevant function of adaptors in LPS signaling pathway.

Peroxisome proliferator-activated receptor-gamma is expressed by rat peritoneal mesothelial cells: its potential role in peritoneal cavity local defense.

BACKGROUND/AIMS: Peritoneal mesothelial cells (PMCs) play an important role in peritoneal inflammatory and immune response. It was reported that the peroxisomal proliferator-activated receptor-gamma (PPARgamma) ligand could effectively reduce inflammatory processes. However, the expression and function of PPARgamma in PMCs has not been reported. This study was to investigate the expression of PPARgamma in rat PMCs and the effect of PPARgamma activation on the production of CD40 and ICAM-1 induced by lipopolysaccharide (LPS). METHODS: Rat PMCs (RPMCs) were harvested from the peritoneal cavity of Sprague-Dawley rats and maintained under defined in vitro conditions. The cells were treated separately with LPS, 15d-PGJ(2), and ciglitazone at different time points. The mRNA and protein expression of PPARgamma, CD40 and ICAM-1 were detected by RT-PCR and Western blot, respectively. The intracellular distribution of PPARgamma was detected by immunocytochemistry. RESULTS: RPMCs expressed PPARgamma both at the mRNA and protein level. The specific signals for PPARgamma were mainly localized in the nucleus with weak staining in the cytoplasm. Stimulation of RPMCs with LPS resulted in a time-dependent increase in the expression of PPARgamma with the peak of mRNA at 3 h and protein at 12 h. Thereafter the expression of PPARgamma gradually attenuated. The mRNA expressions for CD40, ICAM-1 and protein expression of ICAM-1 were significantly upregulated following stimulation with LPS. Both 15d-PGJ(2) and ciglitazone decreased the expression of CD40 mRNA and ICAM-1 protein. However, ciglitazone was less effective than 15d-PGJ(2). CONCLUSIONS: There is constitutive expression of PPARgamma in cultured RPMCs and PPARgamma ligands which strongly inhibit LPS-induced CD40 and ICAM-1 production in RPMCs. It suggested that PPARgamma might play a part in the local defense of the peritoneal cavity by downregulating inflammatory mediators, which may play a potential role in preventing peritoneal fibrosis induced by peritonitis. Further in vivo study is needed to demonstrate the long-term effects.