Lignans from the bark of Eucommia ulmoides inhibited Ang II-stimulated extracellular matrix biosynthesis in mesangial cells

Abstract

Background: Tree bark of Eucommia ulmoides Oliv., (commonly well-known as "Du-zhong" in China), has been used to treat hypertension, hypercholesterolemia, hyperglycemia, hepatic fibrosis and renal injury. This study aims to investigate the effects of lignans extracted from the bark of Eucommia ulmoides Oliv. on Ang II-induced proliferation and extracellular matrix biosynthesis in rat mesangial cells.

Methods: Rat mesangial cells (RMCs) were cultured in vitro and divided into six groups (control, Ang II, losartan, and low, middle and high concentration lignans groups). RMC proliferation was measured by MTT assay. RT-qPCR and western blotting were used to detect mRNA and protein expression of collagen type I (Col I), collagen type III (Col III), collagen type IV (Col IV), fibronectin and aldose reductase (AR).

Results: Cellular proliferation induced by Ang II was significantly suppressed by Eucommia lignans of different concentrations (P = 0.034, P < 0.001, and P < 0.001). Treatment of cells with Ang II increased Col I, Col III, Col IV, and fibronectin mRNA expression, which was observed at the protein level (P < 0.001, P < 0.001, P = 0.004, and P = 0.004, respectively). The increased mRNA expression and protein levels of Col I, Col III, Col IV, and fibronectin were diminished remarkably with by treatment Eucommia lignans, and elevated AR expression stimulated by Ang II was significantly inhibited by Eucommia lignans.

Conclusions: Eucommia lignans (Du-zhong) inhibited Ang II-stimulated extracellular matrix biosynthesis in mesangial cells.

Figure 1

Effects of Eucommia lignans on RMCs growth. RMCs were treated with various concentrations of Eucommia lignans (10, 20, 30, 40, 50, 60, 70, 80, or 90 mg/L) for 48 h, and cell viability was assessed by MTT method. Results were given in mean ± SD (n = 6). ^##P < 0.01 vs. the control group.

Figure 2

Inhibitory effects of Eucommia lignans on Ang II-induced RMCs proliferation. All values represented mean ± SD (n = 6). Los: losartan; Lig: lignans. ^#P < 0.05 vs. the control group, ^*P < 0.05, ^**P < 0.01 vs. Ang II group.

Figure 3

Inhibitory effects of Eucommia lignans on Ang II-induced ECM biosynthesis in RMCs. (A) depression of Eucommia lignans on Ang II-increased mRNA expression levels; (B) inhibition of Eucommia lignans on Ang II-stimulated protein expression levels. RT-qPCR and western blot was used to detect the mRNA and protein expressions separately. GAPDH was used as an internal loading control gene. Both mRNA and protein relative expression levels were expressed as folds of control. All values were expressed as mean ± SD (n = 3). Los: losartan; Lig: lignans. ^##P < 0.01 vs. the control group, ^**P < 0.01 vs. Ang II group.

Figure 4

Inhibitory effects of Eucommia lignans on Ang II-induced AR expression in RMCs. (A) Inhibition of Eucommia lignans on Ang II-increased AR mRNA levels; (B and C) depression of Eucommia lignans on Ang II-stimulated AR protein expression. Cells were incubated as described in Methods. RT-qPCRand Western blotting was used to detect the mRNA and protein expressions. GAPDH was selected as an internal standard gene. AR relative expression levels were indicated as folds of control. All results were shown in mean ± SD (n = 3). Los: losartan; Lig: lignans. ^##P < 0.01 vs. the control group, ^**P < 0.01 vs. Ang II group.