Low-dose of multi-glycoside of Tripterygium wilfordii Hook. f., a natural regulator of TGF-β1/Smad signaling activity improves adriamycin-induced glomerulosclerosis in vivo

Abstract

Ethnopharmacological relevance: Transforming growth factor (TGF)-β1/Smad signaling pathway plays a critical role in the prolonged glomerulosclerosis (GS), which is an important determinant during the progression in chronic kidney disease (CKD). For recent 30 years, multi-glycoside of Tripterygium wilfordii Hook. f. (GTW), an extract from Chinese herbal medicine has been proved clinically effective in improving GS in CKD in China. However, therapeutic mechanisms involved in vivo are still unclear. In this study, we aimed to explain the dose-effects and molecular mechanisms of GTW on GS by regulating TGF-β1/Smad signaling activity in adriamycin (ADR)-induced nephropathy (ADRN).

Materials and methods: Rats with ADRN, created by unilateral nephrectomy and twice adriamycin injections (ADR, 4 mg/kg and 2 mg/kg) within 4 weeks, were divided into four groups, the Sham group, the Vehicle group, the low-dose GTW-treated group, and the high-dose GTW-treated group, and that, sacrificed at the end of the 6th week after administration. Proteinuria, blood biochemical parameters, glomerulosclerotic morphological makers, podocyte shape, and nephrin expression were examined, respectively. Protein expressions of key signaling molecules in TGF-β1/Smad pathway, such as TGF-β1, Smad3, phosphorylated-Smad2/3 (p-Smad2/3), and Smad7, were also evaluated individually.

Results: The results indicated that the characterizations of ADRN involved the typical prolonged GS, a small amount of abnormal proteinuria, and the failing renal function; TGF-β1/Smad signaling molecules, especially Smad3, p-Smad2/3, and Smad7 were activated in vivo, accompanied by the exasperation of glomerulosclerotic lesion; GTW at high-dose (100 mg/kg) and low-dose (50 mg/kg) could slightly ameliorate the prolonged GS and nephrin expression, furthermore, the anti-proliferative action of GTW at high-dose was superior to that at low-dose, but caused the significant liver injury; in ADRN model rats, protein expressions of TGF-β1, p-Smad2/3, and Smad7 in the kidneys could be regulated with the treatment of GTW at low-dose.

Conclusion: This study farther demonstrated that the low-dose of GTW, as a natural regulator in vivo, could effectively and safely ameliorate the prolonged GS in FSGS model, via the potential molecular mechanisms involving the reduction of ECM components and the suppression of TGF-β1 over-expression, as well as the bidirectional regulation of TGF-β1/Smad signaling activity.

Keywords: ADR; ADRN; Adriamycin-induced nephropathy; Alb; BCA; BSA; BUN; BW; CKD; ECM; EDTA; FITC; FSGS; GS; GTW; Glomerulosclerosis; IF; KW; MC; Multi-glycoside of Tripterygium wilfordii Hook. f.; PAS; PB; PBS; PI; PVDF; Phosphorylated-Smad2/3; SDS-PAGE; Scr; Smad7; TCM; TGF-β1; Transforming growth factor-β1; Upro; adriamycin; adriamycin-induced nephropathy; bicinchoninic acid; blood urea nitrogen; body weight; bovine serum albumin; chronic kidney disease; ethylenediaminetetraacetic acid; extracellular matrix; fluoresceine isothiocyanate; focal segmental glomerulosclerosis; glomerulosclerosis; immunofluorescence; kidney weight; mesangial cells; multi-glycoside of Tripterygium wilfordii Hook. f.; p-Smad; periodic acid-Schiff; phosphate buffer; phosphate buffer solution; phosphorylated Smad; polyvinylidene fluoride; protease inhibitors; serum albumin; serum creatinine; sodium dodecyl sulfate-polyacrylamide gel electrophoresis; traditional Chinese medicine; transforming growth factor-β1; urinary protein excretion; α-SMA; α-smooth muscle actin.