TNF-receptors on human peritoneal mesothelial cells: Regulation of receptor levels and shedding by IL-1α and TNFα

Human peritoneal mesothelial cells (HPMC) respond to tumor necrosis factor α (TNFα) by releasing various cytokines that may activate the endothelium and induce recruitment of leukocytes during peritonitis. We characterized the receptors for TNF on HPMC to elucidate their functions in peritonitis. Scatchard analysis determined the presence of 70 x 10³ TNF receptors/cell with a kDa of 0.44 nM. TNF receptor 1 (TNF-R1, p55) and TNF- R2 (p75) mRNA were demonstrated by reverse-transcriptase-PCR (RT-PCR). TNF- R1 protein was solely detected by flow cytometry (FCM). Interleukin-1α (IL- 1α) induced down-regulation of TNF-R1. This was concomitant with accumulation of soluble TNF-R1 (sTNF-R1) detected by specific ELISA. LPS had a lower TNF-R1-shedding activity while TNFα did not induce shedding. The IL- 1-induced-sTNF-R1-shedding was suppressed by the protein-kinase-A (PKA) inhibitor, H-8, or by H-7, the inhibitor of both PKC and PKA, but not by the specific PKC inhibitor GF. These experiments suggest a role for PKA in the IL-1-shedding signal. No change in TNF-R1 mRNA levels was observed after IL- 1α or TNFα stimulation while TNF-R2 (p75) mRNA basal levels transiently increased three to fivefold, reaching a peak after four hours followed by an accumulation of sTNF-R2 in the supernatant. Our data suggest that the main receptor expressed on HPMC is TNF-R1. Down-regulation and shedding of TNF-R1 induced by IL-1 and the transient expression of TNF-R2 induced by IL-1 and TNF, may regulate the responses to TNF by HPMC. These results may be important in understanding the inflammatory process of peritonitis were TNF plays a major role.