摘要
目的:探讨牙龈间充质干细胞(GMSCs)移植能否抑制辐射诱导的Ⅱ型肺泡上皮细胞(AECⅡ)衰老,及其在预防放射性肺纤维化(RIPF)中的作用。方法:采用6 Gy照射小鼠Ⅱ型肺泡上皮细胞(MLE12)诱导细胞衰老,照后即刻与GMSCs共培养,通过细胞形态、β半乳糖苷酶(β-Gal)染色、衰老分泌相关表型(SASP)检测以评估细胞衰老程度;C57BL/6小鼠单侧右肺17 Gy照射构建RIPF模型,照后1 d移植GMSCs,照后180 d取材,期间记录小鼠生存率,采用肺脏器比、HE染色、Masson染色评估肺内结构及肺间质胶原沉积状况,应用β-Gal免疫组织化学法与AECⅡ免疫荧光共定位评估肺内细胞衰老程度,qRT-PCR检测肺组织SASP表达变化;采用Western blot检测P53-P21、P16通路关键蛋白表达水平,免疫荧光共定位检测AECⅡ中P21表达情况。结果:GMSCs共培养能够有效抑制辐射诱导的MLE12细胞衰老,使照射后升高的β-Gal阳染率降低11.8%(t=6.72,P<0.05),并使SASP(IL-6、IL-8、IL-1β)的表达有效降低(t=28.43、28.43、4.82,P<0.05);GMSCs移植提高了受照小鼠的生存率,改善了照射诱导的肺泡结构塌陷增厚及胶原蛋白沉积情况,照射后肺组织中衰老细胞数目降低23.9%(t=21.83,P<0.05),SASP的表达水平下降(t=8.86、20.63,P<0.05);抑制照射后MLE12及RIPF小鼠肺组织中P53-P21、P16相关蛋白的上调。结论:GMSCs通过下调衰老相关的P53-P21及P16通路蛋白,从而抑制辐射诱导的AECⅡ衰老,达到预防放射性肺纤维化发生发展的目的。
Objective To investigate whether transplantation of gingival mesenchymal stem cells(GMSCs)can inhibit radiation-induced senescence of alveolar epithelial cells typeⅡ(AECⅡ)and its role in the prevention of radiation-induced pulmonary fibrosis(RIPF).Methods Mouse typeⅡalveolar epithelial cells(MLE12)were irradiated with 6 Gy X-rays and then co-cultured with GMSCs.The extent of cellular senescence of MLE12 cells was assessed by cell morphology,β-Gal staining,and senescence secretion-associated phenotype(SASP)assay.RIPF model was constructed by unilaterally irradiating the right chest of C57BL/6 mice with 17 Gy X-rays.GMSCs were transplanted 1 d after irradiation.At 180 d after irradiation,the pulmonary organ ratio,HE staining,and Masson staining were used to assess intra-pulmonary structure and interstitial collagen deposition in the lung.β-Gal immunohistochemistry and immunofluorescence co-localization with AECⅡwere measured to assess the degree of cellular senescence in the lung.The SASP expression changes in lung tissue were detected by qRT-PCR.The protein expressions in P53-P21 and P16 pathways were detected by Western blot assay.P21 expression in AECⅡwas detected by immunofluorescence co-localization assay.Results GMSCs effectively inhibited radiation-induced senescence of MLE12 cells,reduced the ratio of radiation-elevatedβ-Gal positive cells by 11.8%(t=6.72,P<0.05),and decreased the expressions of SASP(IL-6,IL-8,IL-1β)(t=28.43,28.43,4.82,P<0.05).GMSCs transplantation improved the survival rate of irradiated mice,prevented radiation-induced alveolar structural collapse thickening and collagen deposition,reduced the number of senescent cells in the irradiated lung tissues by 23.9%(t=21.83,P<0.05),and inhibited the expressions of SASP(t=8.86,20.63,P<0.05).GMSCs also inhibited the expression of P53-P21,P16-related proteins in MLE12 cells and lung tissues of mice after irradiation.Conclusions GMSCs inhibit senescence-related P53-P21 and P16 pathways,prevent radiation-induced AECⅡsenescence,as well
作者
赵雯月
李娜
李科君
王彦
何宁宁
杜利清
刘强
Zhao Wenyue;Li Na;Li Kejun;Wang Yan;He Ningning;Du Liqing;Liu Qiang(Institute of Radiation Medicine,Chinese Academy of Medical Sciences and Peking Union Medical College,Tianjin 300192 China)
出处
《中华放射医学与防护杂志》
CAS
CSCD
北大核心
2022年第11期830-838,共9页
Chinese Journal of Radiological Medicine and Protection
基金
国家自然科学基金(32071241,31971168)
中国医学科学院医学与健康科技创新工程项目(2021-I2M-1-042)。
关键词
放射性肺纤维化
细胞衰老
干细胞治疗
牙龈间充质干细胞
Radiation-induced pulmonary fibrosis
Cellular senescence
Stem cell therapy
Gingival mesenchymal stem cells
