CAJ | 학술논문

目的观察人脂肪基质血管成分(SVF)对肾小管上皮细胞(HK-2)增殖、迁移和凋亡的影响及其机制.方法取患者肾周脂肪组织,分离SVF并培养脂肪间充质干细胞(AdMSCs),采用流式细胞术鉴定细胞表型.制备SVF和AdMSCs条件培养基,以无血清DMEM培养基作为阴性对照,分别进行细胞增殖、迁移、刮伤愈合以及凋亡等检测,统计学分析HK-2细胞的增殖、迁移细胞数、刮伤愈合面积以及凋亡细胞比率,采用酶联免疫吸附试验(EHSA)测定肝细胞生长因子(HGF)、血管内皮生长因子(VEGF)和基质细胞衍生因子-1α(SDF-1 α)的表达.结果新鲜分离的SVF呈小圆形,第3代AdMSCs呈均一长梭形.流式细胞检测SVF表达CD29、CD34、CD45和CD90阳性率分别为45.3％、29.4％、23.8％和34.9％,AdMSCs表达CD29、CD90和CD34阳性率分别为99.2％、93.5％和3.8％,不表达CD45.SVF组及AdMSCs组HK-2的增殖、迁移和刮伤愈合能力与对照组比较均显著增强,而HK-2的凋亡则显著减轻(P＜0.05).SVF组HK-2的增殖能力显著高于AdMSCs组(P＜0.05),但两组HK-2的迁移、刮伤愈合及凋亡差异无统计学意义(P＞0.05).SVF组HGF、VEGF和SDF-1α的表达量分别为(198.41±12.74)、(1 385.68±321.03)和(374.13±145.11) ng/L,AdMSCs组则分别为(112.77±29.16)、(1005.44±256.87)和(247.71 ±73.21) ng/L,且SVF分泌HGF的量显著高于AdMSCs(P ＜0.05).结论 SVF显著增强肾小管上皮细胞的增殖能力,与AdMSCs比较,在促进迁移和抑制凋亡等特性方面具有相同效果,但SVF具有不需体外培养等优点.
목적 관찰인지방기질혈관성분(SVF)대신소관상피세포(HK-2)증식、천이화조망적영향급기궤제.방법 취환자신주지방조직,분리SVF병배양지방간충질간세포(AdMSCs),채용류식세포술감정세포표형.제비SVF화AdMSCs조건배양기,이무혈청DMEM배양기작위음성대조,분별진행세포증식、천이、괄상유합이급조망등검측,통계학분석HK-2세포적증식、천이세포수、괄상유합면적이급조망세포비솔,채용매련면역흡부시험(EHSA)측정간세포생장인자(HGF)、혈관내피생장인자(VEGF)화기질세포연생인자-1α(SDF-1 α)적표체.결과 신선분리적SVF정소원형,제3대AdMSCs정균일장사형.류식세포검측SVF표체CD29、CD34、CD45화CD90양성솔분별위45.3％、29.4％、23.8％화34.9％,AdMSCs표체CD29、CD90화CD34양성솔분별위99.2％、93.5％화3.8％,불표체CD45.SVF조급AdMSCs조HK-2적증식、천이화괄상유합능력여대조조비교균현저증강,이HK-2적조망칙현저감경(P＜0.05).SVF조HK-2적증식능력현저고우AdMSCs조(P＜0.05),단량조HK-2적천이、괄상유합급조망차이무통계학의의(P＞0.05).SVF조HGF、VEGF화SDF-1α적표체량분별위(198.41±12.74)、(1 385.68±321.03)화(374.13±145.11) ng/L,AdMSCs조칙분별위(112.77±29.16)、(1005.44±256.87)화(247.71 ±73.21) ng/L,차SVF분비HGF적량현저고우AdMSCs(P ＜0.05).결론 SVF현저증강신소관상피세포적증식능력,여AdMSCs비교,재촉진천이화억제조망등특성방면구유상동효과,단SVF구유불수체외배양등우점.
Objective To observe the effects and mechanism of adipose stromal vascular fraction (SVF) on the proliferation,migration and apoptosis of human renal tubular epithelial cells (HK-2).Methods The perinephric adipose tissue was collected for the isolation of SVF and culture of adipose derived mesenchymal stem cells (AdMSCs).Both SVF and AdMSCs were characterized by flow cytometric analysis.SVF conditioned medium was prepared for cell proliferation,migration,scrape wound healing,and apoptosis assay.Simultaneously,AdMSCs conditioned medium served as positive control,while DMEM without fatal bovine serun (FBS) served as negative control.Enzyme linked immunosorbent assay (ELISA) was performed to evaluate the expression levels of hepatocyte growth factor (HGF),vascular endothelial growth factor (VEGF) and stromal cell derived factor-1α (SDF-1α) in the three groups.Results Freshly isolated SVF exhibited small round morphologies,while AdMSCs exhibited typical spindle-shaped morphologies.Flow cytometric analysis demonstrated that SVF expressed CD29 (45.3％),CD34 (29.4％),CD45 (23.8％) and CD90 (34.9％).AdMSCs expressed CD29 (99.2％),CD90 (93.5％),and CD34 (3.8％),but was negative for CD45.The capability of HK-2 proliferation,migration and scrape wound healing was significantly enhanced in both SVF and AdMSCs groups as compared with that in control group (P ＜ 0.05).Simultaneously,both SVF and AdMSCs could significantly attenuate the apoptosis of HK-2 cells (P ＜ 0.05).In addition,the proliferative capability of HK-2 in SVF group was significantly higher than that in AdMSCs group (P ＜ 0.05),while no significant difference in cell migration,scrape wound healing,and apoptosis of HK-2 cells was found between SVF and AdMSCs groups (P＞ 0.05).The expression levels of HGF,VEGF,and SDF-1α were (198.41 ± 12.74),(1 385.68 ± 321.03) and (374.13 ± 145.11) ng/L in SVF group,and those were (112.77 ± 29.16),(1 005.44 ± 256.87) and (247.71 ± 73.21) ng/L in AdMSCs group.However,SVF secreted significantly higher levels of HGF than AdMSCs (P ＜ 0.05).Conclusion SVF showed strong promoting effect on the proliferation of HK-2,and exhibited similar capability with AdMSCs of enhancing HK-2 migration and reducing its apoptosis.These results suggest a potential prospect of SVF in regenerative medicine for the no need of in vitro culture.