Abstract:

Chen Yan, Lei Tiechi, Shi Ying, Xu Shizheng. Department of Dermatology, Renmin Hospital of Wuhan University, Wuhan 430060, China Corresponding author: Lei Tiechi, Email: tchlei@whu.edu.cn 【Abstract】 Objective To investigate the effects of ciprofloxacin on dermal collagen synthesis and profibrotic gene expressions in an experimental mouse model of scleroderma induced by bleomycin. Methods Experimental mouse models of scleroderma were established by subcutaneous injection of bleomycin into the dorsal skin of 15 BALB/c mice for 4 consecutive weeks. Then, the mouse models were randomly and equally divided into 3 groups to be topically treated with 1% ciprofloxacin cream （ciprofloxacin group）, 2.5% asiaticoside cream （asiaticoside group） and cream vehicle （model group） respectively for 5 consecutive weeks. Five mice firstly injected with sterile phosphate buffered saline （PBS） for 4 weeks then topically treated with cream vehicle for 5 weeks served as the blank control group. After the 5-week topical treatment, all the mice were sacrificed, skin specimens were resected from the dorsal skin of them, and subjected to HE staining and Masson staining. Further more, an immunohistochemical assay was performed to measure the expressions of type I collagen （COL-1）, matrix metalloproteinase-1 （MMP1） and tissue inhibitor of matrix metalloproteinase-1 （TIMP1）, semi-quantitative reverse transcription PCR to quantify the expressions of connective tissue growth factor （CTGF）, transforming growth factor-β1 （TGFβ1） and Smad3 genes, and alkaline hydrolysis-spectrophotometry to determine the level of hydroxyproline in skin. Statistical analysis was carried out by one-way analysis of variance and the least significant difference （LSD） test with the SPSS 17.0 software. Results Compared with the blank control group, the model group showed increased dermal thickness at injection sites （432.76 ± 93.74 μm vs. 301.69 ± 79.47 μm, P < 0.01）. Masson staining revealed thick and dense collagen bundles in an irregular arrangement in the dermis in the model group, which was consistent with dermal fibrosis in scleroderma. The total content of collagen and staining intensity of COL-1, MMP1 and TIMP1 were all significantly decreased in the ciprofloxacin group and asiaticoside group compared with the model group （F = 1628.54, 33.29, 84.82, 224.81, respectively, all P < 0.01）, while no significant changes were observed in dermal thickness （both P > 0.05）. Moreover, compared with cream vehicle, asiaticoside down-regulated the expressions of the three profibrotic genes （CTGF, TGFβ1 and Smad3） to different extents （all P < 0.05）, while ciprofloxacin only inhibited the expressions of TGFβ1 and Smad3 genes （both P < 0.05） with no significant effect on CTGF gene expression （P > 0.05）. Conclusion Ciprofloxacin may counteract dermal fibrosis by inhibiting the TGFβ1/Smad3 pathway and modulating the unbalanced expressions of MMP1 and TIMP1.