BACKGROUND Umbilical cord(UC)mesenchymal stem cell(MSC)transplantation is a potential therapeutic intervention for atherosclerotic vascular disease.Integrin beta 3(ITGB3)promotes cell migration in several cell types.H...BACKGROUND Umbilical cord(UC)mesenchymal stem cell(MSC)transplantation is a potential therapeutic intervention for atherosclerotic vascular disease.Integrin beta 3(ITGB3)promotes cell migration in several cell types.However,whether ITGBmodified MSCs can migrate to plaque sites in vivo and play an anti-atherosclerotic role remains unclear.AIM To investigate whether ITGB3-overexpressing MSCs(MSCs^(ITGB3))would exhibit improved homing efficacy in atherosclerosis.METHODS UC MSCs were isolated and expanded.Lentiviral vectors encoding ITGB3 or green fluorescent protein(GFP)as control were transfected into MSCs.Sixty male apolipoprotein E-/-mice were acquired from Beijing Vital River Lab Animal Technology Co.,Ltd and fed with a high-fat diet(HFD)for 12 wk to induce the formation of atherosclerotic lesions.These HFD-fed mice were randomly separated into three clusters.GFP-labeled MSCs(MSCs^(GFP))or MSCs^(ITGB3)were transplanted into the mice intravenously via the tail vein.Immunofluorescence staining,Oil red O staining,histological analyses,western blotting,enzymelinked immunosorbent assay,and quantitative real-time polymerase chain reaction were used for the analyses.RESULTS ITGB3 modified MSCs successfully differentiated into the“osteocyte”and“adipocyte”phenotypes and were characterized by positive expression(>91.3%)of CD29,CD73,and CD105 and negative expression(<1.35%)of CD34 and Human Leukocyte Antigen-DR.In a transwell assay,MSCs^(ITGB3)showed significantly faster migration than MSCsGFP.ITGB3 overexpression had no effects on MSC viability,differentiation,and secretion.Immunofluorescence staining revealed that ITGB3 overexpression substantially enhanced the homing of MSCs to plaque sites.Oil red O staining and histological analyses further confirmed the therapeutic effects of MSCs^(ITGB3),significantly reducing the plaque area.Enzyme-linked immunosorbent assay and quantitative real-time polymerase chain reaction revealed that MSC^(ITGB3)transplantation considerably decreased the inflammatory response in pathological tissues by improving the dynamic equilibrium of pro-and anti-inflammatory cytokines.CONCLUSION These results showed that ITGB3 overexpression enhanced the MSC homing ability,providing a potential approach for MSC delivery to plaque sites,thereby optimizing their therapeutic effects.展开更多
目的探究柚皮苷对破骨细胞分化和极化功能的影响。方法体外培养RAW264.7细胞,设置对照组(培养基不含柚皮苷)和实验组(培养基中柚皮苷剂量分别为2、20、200μg/L)连续培养7 d,通过抗酒石酸酸性磷酸酶(TRAP)染色对比组间诱导分化结果,通...目的探究柚皮苷对破骨细胞分化和极化功能的影响。方法体外培养RAW264.7细胞,设置对照组(培养基不含柚皮苷)和实验组(培养基中柚皮苷剂量分别为2、20、200μg/L)连续培养7 d,通过抗酒石酸酸性磷酸酶(TRAP)染色对比组间诱导分化结果,通过实时荧光定量逆转录聚合酶链反应(qPCR)对比组间组织蛋白酶K(CK)、基质金属蛋白酶-9(MMP-9)、整合素β3(integrinβ3)和非受体酪氨酸激酶(c-src)mRNA表达水平,通过蛋白免疫印迹法对比组间CK、MMP-9、integrinβ3、磷酸化产物p-src蛋白表达水平。结果与对照组相比,各实验组TRAP染色阳性细胞数显著降低(P<0.05);与对照组相比,20μg/L与200μg/L组CK、MMP-9、integrinβ3及c-src m RNA表达水平显著降低(P<0.05);与对照组相比,20μg/L与200μg/L组CK、MMP-9、integrinβ3及p-src蛋白表达水平显著降低(P<0.05)。结论柚皮苷对破骨细胞的分化和极化有一定的抑制作用,其作用机制可能与下调integrinβ3、c-src及p-src表达有关。展开更多
基金National Natural Science Foundation of China,No.82100301Key Science and Technology Research Program of Hebei Provincial Department of Health,No,20221014.
文摘BACKGROUND Umbilical cord(UC)mesenchymal stem cell(MSC)transplantation is a potential therapeutic intervention for atherosclerotic vascular disease.Integrin beta 3(ITGB3)promotes cell migration in several cell types.However,whether ITGBmodified MSCs can migrate to plaque sites in vivo and play an anti-atherosclerotic role remains unclear.AIM To investigate whether ITGB3-overexpressing MSCs(MSCs^(ITGB3))would exhibit improved homing efficacy in atherosclerosis.METHODS UC MSCs were isolated and expanded.Lentiviral vectors encoding ITGB3 or green fluorescent protein(GFP)as control were transfected into MSCs.Sixty male apolipoprotein E-/-mice were acquired from Beijing Vital River Lab Animal Technology Co.,Ltd and fed with a high-fat diet(HFD)for 12 wk to induce the formation of atherosclerotic lesions.These HFD-fed mice were randomly separated into three clusters.GFP-labeled MSCs(MSCs^(GFP))or MSCs^(ITGB3)were transplanted into the mice intravenously via the tail vein.Immunofluorescence staining,Oil red O staining,histological analyses,western blotting,enzymelinked immunosorbent assay,and quantitative real-time polymerase chain reaction were used for the analyses.RESULTS ITGB3 modified MSCs successfully differentiated into the“osteocyte”and“adipocyte”phenotypes and were characterized by positive expression(>91.3%)of CD29,CD73,and CD105 and negative expression(<1.35%)of CD34 and Human Leukocyte Antigen-DR.In a transwell assay,MSCs^(ITGB3)showed significantly faster migration than MSCsGFP.ITGB3 overexpression had no effects on MSC viability,differentiation,and secretion.Immunofluorescence staining revealed that ITGB3 overexpression substantially enhanced the homing of MSCs to plaque sites.Oil red O staining and histological analyses further confirmed the therapeutic effects of MSCs^(ITGB3),significantly reducing the plaque area.Enzyme-linked immunosorbent assay and quantitative real-time polymerase chain reaction revealed that MSC^(ITGB3)transplantation considerably decreased the inflammatory response in pathological tissues by improving the dynamic equilibrium of pro-and anti-inflammatory cytokines.CONCLUSION These results showed that ITGB3 overexpression enhanced the MSC homing ability,providing a potential approach for MSC delivery to plaque sites,thereby optimizing their therapeutic effects.
文摘目的探究柚皮苷对破骨细胞分化和极化功能的影响。方法体外培养RAW264.7细胞,设置对照组(培养基不含柚皮苷)和实验组(培养基中柚皮苷剂量分别为2、20、200μg/L)连续培养7 d,通过抗酒石酸酸性磷酸酶(TRAP)染色对比组间诱导分化结果,通过实时荧光定量逆转录聚合酶链反应(qPCR)对比组间组织蛋白酶K(CK)、基质金属蛋白酶-9(MMP-9)、整合素β3(integrinβ3)和非受体酪氨酸激酶(c-src)mRNA表达水平,通过蛋白免疫印迹法对比组间CK、MMP-9、integrinβ3、磷酸化产物p-src蛋白表达水平。结果与对照组相比,各实验组TRAP染色阳性细胞数显著降低(P<0.05);与对照组相比,20μg/L与200μg/L组CK、MMP-9、integrinβ3及c-src m RNA表达水平显著降低(P<0.05);与对照组相比,20μg/L与200μg/L组CK、MMP-9、integrinβ3及p-src蛋白表达水平显著降低(P<0.05)。结论柚皮苷对破骨细胞的分化和极化有一定的抑制作用,其作用机制可能与下调integrinβ3、c-src及p-src表达有关。
