细胞外基质增强人脐带干细胞的抗氧化能力

背景:已有研究表明细胞外基质可以促进细胞增殖,降低细胞内活性氧水平,但细胞外基质是否能够增强脐带干细胞的抗氧化能力以提高其在再生医学和组织工程中的应用效果,相关研究较少。目的:探讨细胞外基质对脐带干细胞增殖、抗氧化以及成骨分化潜能的影响。方法:将脐带干细胞分4组:①聚苯乙烯组:用普通聚苯乙烯培养板培养脐带干细胞,无其他特殊处理;②细胞外基质组:用细胞外基质培养脐带干细胞,无其他特殊处理;③聚苯乙烯+过氧化氢组:用聚苯乙烯板培养脐带干细胞,并用200μmol/L过氧化氢预处理细胞2 h;④细胞外基质+过氧化氢组:用细胞外基质培养脐带干细胞,并用200μmol/L过氧化氢预处理细胞2 h。过氧化氢预处理2 h结束后采用CCK-8法检测脐带干细胞的增殖能力;过氧化氢预处理2 h结束后继续培养72 h,流式细胞实验和qRT-PCR实验检测脐带干细胞的抗氧化能力;过氧化氢预处理2 h结束后成骨诱导分化14 d,采用茜素红染色和qRT-PCR实验检测脐带干细胞的成骨能力。结果与结论:①细胞外基质组和细胞外基质+过氧化氢组的吸光度值分别高于聚苯乙烯组和聚苯乙烯+过氧化氢组;②聚苯乙烯+过氧化氢组和聚苯乙烯组活性氧水平分别高于细胞外基质组和细胞外基质+过氧化氢组(P<0.05);抗氧化酶相关基因SOD2和CAT的表达水平在细胞外基质组和细胞外基质+过氧化氢组分别较聚苯乙烯组和聚苯乙烯+过氧化氢组明显上调(P<0.05);③细胞外基质组成骨相关基因COL1A1、RUNX2、OCN、OSTERIX表达量最高,细胞外基质+过氧化氢组次之,聚苯乙烯+过氧化氢组最低,各组间差异有显著性意义(P<0.05);④结果表明,细胞外基质可以提高脐带干细胞的增殖能力、抗氧化能力和成骨分化潜能,是一种有广泛应用前景的细胞体外扩增培养方法。

Extracellular matrix improves antioxidant capacity of human umbilical cord stem cells

BACKGROUND:Extracellular matrix has been shown to improve cell proliferation and reduce intracellular reactive oxygen species levels.However,there is little research on whether extracellular matrix can enhance the antioxidant capacity of umbilical cord stem cells to enhance their application in regenerative medicine and tissue engineering.OBJECTIVE:To investigate the effect of extracellular matrix on umbilical cord stem cell proliferation,antioxidant and osteogenic capacity.METHODS:The umbilical cord stem cells were divided into four groups.In the polystyrene group,the umbilical cord stem cells were cultured with ordinary polystyrene culture plate without other special treatment.In the extracellular matrix group,the umbilical cord stem cells were cultured with extracellular matrix without other special treatment.In the polystyrene+hydrogen peroxide group,the umbilical cord stem cells were cultured with polystyrene plate and pretreated with 200μmol/L hydrogen peroxide for 2 hours.In the extracellular matrix+hydrogen peroxide group,umbilical cord stem cells were cultured with extracellular matrix and pretreated with 200μmol/L hydrogen peroxide for 2 hours.The cells were pretreated with 200μmol/L hydrogen peroxide for 2 hours.Proliferation capacity of umbilical cord stem cells was detected by CCK-8 assay.The cells were cultured for 72 hours after hydrogen peroxide pretreatment for 2 hours.The antioxidant capacity of umbilical cord stem cells was detected by flow cytometry and qRT-PCR.After 2 hours of hydrogen peroxide pretreatment,the cells were induced to differentiate into osteoblasts for 14 days.The osteogenic capacity of umbilical cord stem cells was detected by alizarin red staining and qRT-PCR.RESULTS AND CONCLUSION:The absorbance values of extracellular matrix group and extracellular matrix+hydrogen peroxide group were higher than that of polystyrene group and polystyrene+hydrogen peroxide group,respectively.The levels of reactive oxygen species in the polystyrene+hydrogen peroxide group and the polystyrene group were higher than those in the extracellular matrix group and the extracellular matrix+hydrogen peroxide group,respectively(P<0.05).The expression levels of antioxidant enzyme-related genes SOD2 and CAT in the extracellular matrix group and extracellular matrix+hydrogen peroxide group were significantly higher than those in the polystyrene group and the polystyrene+hydrogen peroxide group,respectively(P<0.05).The expression of bone related genes COL-1,RUNX2,OCN,and OSTERIX was highest in the extracellular matrix group,followed by the extracellular matrix+hydrogen peroxide group,and lowest in the polystyrene+hydrogen peroxide group;there was significant difference between the groups(P<0.05).The results show that extracellular matrix can increase the proliferation capacity,antioxidant capacity and osteogenic differentiation potential of umbilical cord stem cells.It is a method for in vitro amplification and culture of cells with wide application prospects.