Aldosterone and TGF-β₁ synergistically increase PAI-1 expression in hepatic stellate cells of rats

Aldosterone is related to the fibrosis of several organs, but the specific mechanism underlying the aldosterone induced hepatic fibrosis is still unclear. Separation, culture and identification of primary hepatic stellate cells (HSCs): The fluids and digestives used in the present study were able to completely remove blood cells, digest hepatocytes and matrix, and effectively separate HSCs. The in situ perfusion was performed at 2 steps: in situ perfusion with pre-perfusion fluid and ex vivo perfusion with enzyme-containing perfusion fluid. Influence of Ald on PAI-1 and Smad expressions in HSCs: cells were divided into control group, Ald group (10(-6) M), spironolactone (SPI) group and Ald+SPI group, and the mRNA and protein expressions of PAI-1 and Smad were detected. Ald induced type I collagen expression in HSCs: Immunohistochemistry was performed to detect type I collagen expression in the supernatant of control group, Ald group (10(-6) M), TGF-β1 group, and Ald+TGF-β1 group. Influence of Ald and TGF-β1 on PAI-1 expression in HSCs: cells were divided into control group, Ald group (10(-6) M), TGF-β1 group, and Ald+TGF-β1 group, and the mRNA and protein expressions of PAI-1 were determined by RT-PCR and Western blot assay, respectively. Synergistic effect of Ald and TGF-β1 on PAI-1 expression in HSCs: cells were divided into control group, Ald group (10(-6)), TGF-β1 group, Ald (10(-6) M)+TGF-β1 group, Ald (10(-7) M)+TGF-β1 group and Ald (10(-8) M)+TGF-β1 group, and the mRNA and protein expressions of PAI-1 were detected by RT-PCR and Western blot assay, respectively. The survival rate, purity, markers and activation of HSCs were determined after separation. Influence of Ald on PAI-1 expression in HSCs: PAI-1 expression increased in HSCs of Ald group, SPI group and Ald+API group, and the PAI-1 expression in Ald group and Ald+SPI group was significantly higher than in control group (P