Total flavones of Abelmoschus manihot improve diabetic nephropathy by inhibiting the iRhom2/TACE signalling pathway activity in rats

Context: Total flavones extracted from Abelmoschus manihot L. (Malvaceae) medic (TFA) have been proven clinically effective at improving renal inflammation and glomerular injury in chronic kidney disease (CKD). Objective: This study evaluated the function of TFA as an inhibitor of iRhom2/TACE (tumour necrosis factor- a converting enzyme) signalling and investigated its anti-DN (diabetic nephropathy) effects in a DN rat model. Materials and methods: In vitro, cells were treated with 200 mu g/mL advanced glycation end products (AGEs), and then co-cultured with 20 mu g/mL TFA for 24 h. Real time PCR, western blotting and co-immunoprecipitation assays were performed. In vivo, DN was induced in 8 week old male Sprague-Dawley rats via unilateral nephrectomy and intraperitoneal injection of streptozotocin, then TFA were administered to rats by gavage for 12 weeks at three different doses (300, 135 and 75mg/kg/d). 4-Phenylbutanoic acid (2.5mg/kg/d) was used as a positive control. Results: IC50 of TFA is 35.6 mu M in HK2 and 39.6 lM in HRMC. TFA treatment (20 lM) inhibited the activation of iRhom2/TACE signalling in cultured cells induced by AGEs. LD50 > 26 g/kg and ED50=67mg/kg of TFA in rat by gavage, TFA dose-dependently downregulated the expression of proinflammatory cytokines and exerted anti-inflammatory effects significantly though inhibiting the activation of iRhom2/TACE signalling. Discussion and conclusions: Our results show that TFA could dose-dependently ameliorate renal inflammation by inhibiting the activation of iRhom2/TACE signalling and attenuating ER stress. These results suggest that TFA has potential therapeutic value for the treatment of DN in humans.