ABSTRACT
OBJECTIVE@#To investigate the expression of calponin-1 (CNN1) in systemic sclerosis (SSc) and its pathogenic role in fibrosis.@*METHODS@#Skin biopsy samples were collected from 19 patients with SSc and 21 healthy subjects. Real-time PCR was used to detect the expression of and mRNAs in the samples, and the protein expression of CNN1 was detected using immunohistochemistry. In cultured primary human dermal fibroblasts, expression was knocked down RNA interference, and the mRNA expression levels of and the fibrosis-related genes , , , , and were detected using real-time PCR; the proliferation of the cells was assessed using a real-time cell proliferation detection system.@*RESULTS@#Compared with that in samples from normal subjects, the expression of mRNA was significantly increased in the skin tissue of patients with SSc ( < 0.05) with a positive correlation with α-SMA (=0.7219, < 0.0001); the protein expression of CNN1 was also significantly increased in the skin tissue of patients with SSc. In cultured primary skin fibroblasts, the expression of CNN1 mRNA was positively correlated with and mRNA expressions (=0.6547, < 0.05; =0.6438, < 0.05). knockdown in the fibroblasts significantly inhibited the cell proliferation, obviously lowered the expressions of fibrosis-related genes, and reduced the protein expression of collagen.@*CONCLUSIONS@#The expression of is increased in the skin tissues of patients with SSc, and knockdown can reduce the activity of dermal fibroblasts, suggesting the close correlation of CNN1 with fibrosis in SSc.
Subject(s)
Humans , Calcium-Binding Proteins , Cells, Cultured , Fibroblasts , Fibrosis , Microfilament Proteins , Scleroderma, Systemic , SkinABSTRACT
Objective To investigate the effect of Calponin-1 suppression on human myometrium cells through adenovirus mediated siRNA. Methods Human uterine smooth muscle tissues were digested with enzymes, cultured and confirmed with immunocytochemistry. Aadenovirus siRNA-Calponin-1 plasmid was transfected into primary cultured uterine smooth muscle cells in vitro. The expressions of Calponin-1 mRNA and protein were analyzed by RT-PCR and Western blot, respectively.Results The pAdEasy-pShuttle-U6-Calponin-1 siRNA plasmid was successfully constructed, and Calponin-1 siRNA mediated by recombinant adenovirus resulted in markedly reduced expression of Calponin-1 mRNA and protein in human myometrium cells. The gray values of Calponin-1 mRNA in the uterine smooth muscle cells in the experimental, blank control, and empty vector groups were 316.3±39.2, 1048.5±126.4 and 1027.2±127.5, respectively. The gray values of Calponin-1 protein were 323.3±43.2, 1021.5±143.4, and 1019.2±144.5,respectively. The difference between the experimental group and the blank control group as well as the empty vector group was significant (P< 0.05). There was no significant difference between the empty vector group and the blank control group (P>0.05).Conclusion The pAdEasy-pShuttle-U6-Calponin-1 siRNA plasmid can inhibit the expression of Calponin-1 in human myometrium cells in vitro,which may be a useful approach to determine the role of Calponin-1 in delivery.