High Glucose Solution and Spent Dialysate Stimulate the Synthesis of Transforming Growth Factor-β1 of Human Peritoneal Mesothelial Cells: Effect of Cytokine Costimulation

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Abstract
To investigate the effect of high glucose and spent peritoneal dialysate on the transforming growth factor-β1 (TGFβ1) synthesis of cultured human peritoneal mesothelial cells (HPMCs) and to examine the effect of costimulation with high glucose or spent dialysate, and cytokines, interleukin-1β (IL-1β), and tumor necrosis factor-α (TNFα) on TGFβ1 synthesis of HPMCs. HPMCs were exposed to different concentrations of glucose (30, 60, and 90 mmol/L) or spent peritoneal dialysate for 48 hours in the absence or presence of IL-1β (1 ng/mL) and TNFα (1 ng/mL). TGFβ1 mRNA expression was assessed by Northern blot analysis and TGFβ1 protein release by Western blot analysis and enzymelinked immunosorbent assay (ELISA). Exposure of HPMCs to high glucose conditions (30, 60, and 90 mmol/L of D-glucose) induced 2.3-, 3.6-, and 4.0-fold increases in TGFβ1 mRNA expression of HPMC with enhanced TGFβ1 protein synthesis and secretion into the media, whereas there were no significant changes in TGFβ1 synthesis with equimolar concentrations of D-mannitol. Incubation with spent dialysate also significantly increased TGFβ1 mRNA expression and protein secretion compared to control media (p < 0.05). Stimulation with IL-1β (1 ng/mL) or TNFα (1 ng/mL) resulted in a significant increase in TGFβ1 mRNA expression after 48 hours: 2.7 and 2.1 times the control level, respectively. However, TNFα-induced increase in TGFβ1 mRNA expression was not translated into TGFβ1 protein secretion, while IL-1β stimulation induced a significant increase in TGFβ1 protein secretion as well as TGFβ1 mRNA expression. Combined stimulation by high glucose or spent dialysate, together with IL-1β or TNFα, showed a greater increase in TGFβ1 mRNA expression and protein secretion compared to stimulation by high glucose or spent dialysate alone. Our results clearly show that high glucose solution and spent dialysate themselves might be sufficient to stimulate the production of TGFβ1 by peritoneal mesothelial cells. In peritoneal dialysis patients, this state of chronic induction of TGFβ1 is further exacerbated in the presence of peritonitis because of the stimulatory effect of proinflammatory cytokines, resulting in augmented TGFβ1 synthesis, thus promoting peritoneal fibrosis.