雷帕霉素抑制光老化成纤维细胞周期阻滞的研究
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摘要
目的探讨雷帕霉素(rapamycin,RAPA)对光老化成纤维细胞(fibroblasts,FBs)周期及相关蛋白的影响。方法采用CCK-8法检测不同浓度的RAPA对FBs活性的影响,甄选最佳应用浓度。利用UVB体外反复照射构建FBs的老化模型,流式细胞技术检测各组周期的分布变化,Western Blot技术检测细胞周期相关蛋白p53和p21的表达。结果低浓度RAPA对细胞活性并无影响。与UVB照射组比较,RAPA+UVB组的S期细胞比例下降了6.5%,细胞周期相关蛋白p53和p21的表达分别下降了26.3%和27.8%。结论 RAPA可以缓解光老化FBs的S期阻滞,降低周期蛋白p53和p21的产生,一定程度上改善UVB照射导致的细胞衰老。
Objective To explore the effect of rapamycin(RAPA) on the cell cycle of photoaged fibroblasts(FBs)and on the expressions of associated proteins. Methods Cell counting kit-8(CCK-8) assay was applied in detecting the effect of RAPA of different concentration on the activity of FBs so as to find out the most suitable concentration; the aged model of FBs was built up by repeated UVB radiation; flow cytometry was applied in detecting the changes of cell cycle distribution while Western Blot Analysis in detecting the expressions of proteins related to cell cycle: protein p53 and protein p21. Results RAPA of low concentration was of no effect on cell activity; compared with FBs radiated by UVB, the cell ratio in S phase of FBs pretreated by RAPA and UVB decreased 6.5%, the expressions of protein p53 and protein p21 decreased 26.3% and 27.8% respectively. Conclusions RAPA can relieve the S-phase arrest of photoaged FBs, decrease the expression of protein p53 and protein p21, and thus improve the cell aging induced by UVB radiation to some extent..
Objective To explore the effect of rapamycin(RAPA) on the cell cycle of photoaged fibroblasts(FBs)and on the expressions of associated proteins. Methods Cell counting kit-8(CCK-8) assay was applied in detecting the effect of RAPA of different concentration on the activity of FBs so as to find out the most suitable concentration; the aged model of FBs was built up by repeated UVB radiation; flow cytometry was applied in detecting the changes of cell cycle distribution while Western Blot Analysis in detecting the expressions of proteins related to cell cycle: protein p53 and protein p21. Results RAPA of low concentration was of no effect on cell activity; compared with FBs radiated by UVB, the cell ratio in S phase of FBs pretreated by RAPA and UVB decreased 6.5%, the expressions of protein p53 and protein p21 decreased 26.3% and 27.8% respectively. Conclusions RAPA can relieve the S-phase arrest of photoaged FBs, decrease the expression of protein p53 and protein p21, and thus improve the cell aging induced by UVB radiation to some extent..
引文
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[4]Zeng JP,Bi B,Chen L,et al.Repeated exposure of mouse dermal fibroblasts at a sub-cytotoxic dose of UVB leads to premature senescence:a robust model of cellular photoaging[J].J Dermatol Sci,2014,73(1):49-56.
[5]Chen L,Bi B,Zeng J,et al.Rosiglitazone ameliorates senescencelike phenotypes in a cellular photoaging model[J].J Dermatol Sci,2015,77(3):173-181.
[6]Demidenko ZN,Zubova SG,Bukreeva EI,et al.Rapamycin decelerates cellular senescence[J].Cell Cycle,2009,8(12):1888-1895.
[7]陈亮,毕波,曾继平,等.p H对衰老相关-半乳糖苷酶染色的影响[J].中国美容整形外科杂志,2014,25(12):751-754.
[8]Wang XF,Huang YF,Wang L,et al.Photo-protective activity of pogostone against UV-induced skin premature aging in mice[J].Exp Gerontol,2016,77:76-86.
[9]卢勇舟,陈亮,毕波,等.胶原蛋白碎片对光老化真皮细胞外基质的影响[J].中国美容整形外科杂志,2015,26(4):213-215.
[10]Tobin DJ.Introduction to skin aging[J].J Tissue Viability,2017,26(1):37-46.
[11]Schuch AP,Moreno NC,Schuch NJ,et al.Sunlight damage to cellular DNA:Focus on oxidatively generated lesions[J].Free Radic Biol Med,2017,107(17):110-124.
[12]Greussing R,Hackl M,Charoentong P,et al.Identification of micro RNA-m RNA functional interactions in UVB-induced senescence of human diploid fibroblasts[J].BMC Genomics,2013,14:224.
[13]Bosch R,Philips N,Suárez-Pérez JA,et al.Mechanisms of photoaging and cutaneous photocarcinogenesis,and photoprotective strategies with phytochemicals[J].Antioxidants(Basel),2015,4(2):248-268.
[14]Bhatia-Dey N,Kanherkar RR,Stair SE,et al.Cellular senescence as the causal nexus of aging[J].Front Genet,2016,7:13.
[15]陈亮,毕波,曾继平,等.罗格列酮调控光老化成纤维细胞周期阻滞的研究[J].中国美容整形外科杂志,2015,26(7):436-440.
[16]Karthikeyan R,Kanimozhi G,Prasad NR,et al.7-Hydroxycoumarin prevents UVB-induced activation of NF-B and subsequent overexpression of matrix metalloproteinases and inflammatory markers in human dermal fibroblast cells[J].J Photochem Photobiol B,2016,161:170-176.
[17]Lu J,Guo JH,Tu XL,et al.Tiron inhibits UVB-Induced AP-1binding sites transcriptional activation on MMP-1 and MMP-3promoters by MAPK signaling pathway in human dermal fibroblasts[J].PLo S One,2016,11(8):e0159998.
[18]Rubinsztein DC,Marino G,Kroemer G.Autophagy and aging[J].Cell,2011,146(5):682-695.
[19]Laplante M,Sabatini DM.m TOR signaling in growth control and disease[J].Cell,2012,149(2):274-293.
[20]Finkel T.Relief with rapamycin:m TOR inhibition protects against radiation-induced mucositis[J].Cell Stem Cell,2012,11(3):287-288.
[21]Sonis ST.Oral mucositis[J].Anticancer Drugs,2011,22(7):607-612.