Inhibitory Effect Of Sirolimus On The Expression Of Type I Collagen And Fibronectin In Myofibroblasts
Keywords
Sirolimus, Fibroblasts, Fibronectins, Collagen Type I
Abstract
Objective: To explore the anti-renal fibrosis effect of sirolimus [sirolimus, also known as rapamycin] at the cellular level. Methods: Primary cultured rat renal cortical myofibroblasts (myofmblast, myof) were used .A control group and a group treated with sirolimus 40 mg/l were set up at each time point of the experiment. The cells and cell culture supernatants were collected after culturing the cells for 12h, 24h, and 48h, and the intracellular ¦Á-smooth muscle was detected using the westernblot method. Expression of kinesin (¦Á-smoothmuscleactin, ¦Á-sma), fibronectin (fn), type ¢¡ collagen (type ¢¡collagen, col-¢¡) and fn, col-¢¡ protein levels in the supernatant; quantified by real-time fluorescence The PCR (real-time quantitative PCR) method was used to detect the changes in the expression of procollagen-¢¡ (procollagen-¢¡) at each time point of 1h, 2h, 4h, and 6h of cell culture; the gelatin zymography method was used to detect matrix metalloproteinase-2 in the cell culture supernatant. (matrixmetalloproteinase-2, mmp-2) and matrix metalloproteinase-9 (matrixmetalloproteinase-9, mmp-9) activities. The experimental results were based on the control group at each time point as the baseline 1, and the ratio to the control group was calculated. Results: ( 1) Sirolimus had no effect on the expression of ¦Á-sma in cells at any time point. (2) After treatment with sirolimus, the expression of col-¢¡ protein in cells at 24 hours was significantly less than that in the control group. And continued until 48h, which were (0.58¡À0.05) times and (0.63¡À0.18) times that of the control group at each time point, respectively. The difference was statistically significant (p<0.05). (3) Compared with the control group at each time point, The expression of intracellular procollagen-¢¡mRNA decreased at 1h and 2h, by (0.38¡À0.05) times and (0.55¡À0.16) times respectively. The difference was statistically significant (p<0.05), and returned to the basic level at 4h. (4 ) There is basal expression of col-¢¡ in the supernatant of cultured cells at 12 hours. After sirolimus treatment for 24 hours, the expression level is significantly reduced compared with the control group, which is (0.59¡À0.25) times that of the control group. The difference is statistically significant (p <0.05), and continued to show a decreasing trend until 48h. It was (0.52¡À0.21) times that of the control group, p<0.05. (5) The expression trend of fn protein in cells was consistent with that in the supernatant, and its expression level at 24h was lower than that of the control group. Significantly reduced, respectively (0.44¡À0.09) times and (0.40¡À0.15) times that of the control group. The differences were statistically significant (p<0.05). The inhibitory effect of sirolimus on fn disappeared at 48h. (6) After After sirolimus treatment for 12h, 24h and 48h, there was no significant change in the activities of mmp-2 and mmp-9 in the supernatant. Conclusion: Sirolimus can exert its effects by inhibiting the expression of col-¢¡ and or fn of myof. Its anti-renal fibrosis effect. For further information of this article and research, feel free to contact our team for asssitance. Original research was done by Zhang Lan, Huang Haichang, Li Jingzi, Liu Ying
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