The Journal of Practical Medicine >
Effect of artesunate on autophagy in human tenon′ s capsule fibroblasts
Department of Ophthalmology,the Third Affiliated Hospital of Guangzhou Medical University,Guang⁃ zhou 510150,China
Online published: 2021-08-25
Objective To observe autophagy of human tenon′s capsule fibroblasts(HTFs)induced by arte⁃ sunate(Art)and explore the possible mechanism. Methods The cultured in vitro HTFs were divided into control group and Art(50,100,150,200 μg/mL)groups. After treatment for 24 and 48 h,cell activity was determined by methyl thiazol tetrazolium(MTT)assay. 48 h later,quantitative reverse transcriptase PCR(qRT⁃PCR)and Western Blot analysis were employed to assay the relative expression of microtubule⁃associated protein light chain 3(LC3)⁃ Ⅱ and Becline ⁃1 in HTFs. The autophagy activity of HTFs was evaluated by a Cyto ⁃ ID autophagy detection kit, and then the autophagic flux was evaluated by counting the Cyto ⁃ ID positive cells. Results The results of MTT showed that Art could significantly inhibit the growth of fibroblasts in vitro in a concentration and time dependent manner(P < 0.05),and the inhibition rate increased from 4.7% to 68.6% with increase of concentration and prolongation of time. qRT⁃PCR results showed that LC3⁃Ⅱand beclin⁃l mRNA relative expression increased after Art treatment,from(1.367 ± 0.214)to(4.765 ± 0.371),and(1.235 ± 0.164)to(3.487 ± 0.356),respectively. The difference between the groups had statistical significance(FL = 20.920,P < 0.01;FB = 26.370,P < 0.01). Western Blot results showed that LC3⁃Ⅱand beclin⁃l protein relative expression increased after Art treatment,from(0.101 ± 0.025)to(0.808 ± 0.102),and(0.098 ± 0.036)to(0.784 ± 0.064),respectively. The difference between the groups had statistical significance(FL = 32.852,P < 0.01;FB = 29.874,P < 0.01). Cyto⁃ID staining showed that the percentage of autophagic cell increased dramatically from(13.302 ± 2.301)% to(53.274 ± 7.851)% upon the exposure of HTFs to 150 μg/mL Art(t = 8.804,P < 0.01). Conclusion Artesunate can effectively promote the production of LC3⁃Ⅱand Beclin⁃l,increase the Ievel of autophagy in HTFs,and inhibit the growth of fibroblasts.
Key words: artesunate; human Tenon′s capsule fibroblasts; proliferation; autophagy; glaucoma
CHEN Jing, LUO Gaoquan, CHEN Yi.
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Effect of artesunate on autophagy in human tenon′ s capsule fibroblasts
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