低氧环境下PLGA/胶原纤维支架对小鼠骨髓间充质干细胞增殖及成肌腱分化影响

Effects of PLGA/collagen Fiber Scaffold on the Proliferation and Tendon Differentiation of Mouse Bone Marrow Mesenchymal Stem Cells under Hypoxia

目的探讨低氧环境下PLGA/胶原纤维支架对小鼠骨髓间充质干细胞(mBMSCs)增殖及成肌腱分化的影响。方法利用静电纺丝技术制作出30组PLGA/胶原纤维支架,正常条件下培养的mBMSCs为正常对照组;以3%O2,5%CO2,92%N2建立细胞低氧模型,在低氧环境下培养mBMSCs为低氧对照组,mBMSCs种植在PLGA/胶原纤维支架为低氧支架实验组,各组均进行成肌腱分化诱导实验14 d。CCK-8实验检测3 d、5 d、7 d各组mBMSCs增殖的吸光度值,ELISA实验检测各组mBMSCs上清液肌腱细胞标志性蛋白Tenomodulin和Scleraxis含量。结果在3 d、5 d、7 d观察点内,正常对照组、低氧对照组、低氧支架实验组mBMSCs吸光度值差异有统计学意义(P<0.05),第3 d、5 d、7 d观察点内,低氧对照组mBMSCs吸光度值均高于正常对照组,差异有统计学意义(P<0.01);与低氧对照组相比,低氧支架实验组3 d、5 d、7 d吸光度A值均分别增高,差异有统计学意义(P<0.01)。在细胞低氧环境下,低氧支架实验组Tenomodulin蛋白(4.526±0.002)μmol/L和Scleraxis蛋白(3.752±0.004)μmol/L含量高于低氧对照组Tenomodulin蛋白(2.831±0.023)μmol/L和Scleraxis蛋白(2.457±0.032)μmol/L,低氧对照组Tenomodulin蛋白(2.831±0.023)μmol/L和Scleraxis蛋白(2.457±0.032)μmol/L含量高于正常对照组Tenomodulin蛋白(1.542±0.071)μmol/L和Scleraxis蛋白(1.136±0.056)μmol/L,3组对比差异有统计学意义(P<0.01)。结论低氧环境下PLGA/胶原纤维支架可有效促进mBMSCs增殖和成肌腱分化。

Objective To investigate the effect of PLGA/collagen fiber scaffold on the proliferation and tendon differentiation of mouse bone marrow mesenchymal stem cells(m BMSCs)under hypoxic environment.Methods Thirty groups of PLGA/collagen fiber scaffolds were fabricated by electrospinning technology.The m BMSCs cultured under normal conditions served as the normal control group;the hypoxia model of cells was established with 3%O2,5%CO2,and 92%N2,under hypoxia environment cultivated m BMSCs served as the hypoxic control group,and m BMSCs planted on PLGA/collagen fiber scaffolds served as the hypoxic scaffold experimental group.Each group was subjected to tendon differentiation induction experiments for 14 d.CCK-8 experiment was used to detect the absorbance value of m BMSCs proliferation in each group on 3 d,5 d,7 d,and the content of Tenomodulin and Scleraxis in the supernatant of m BMSCs in each group was detected by ELISA.Results Within the observation points of 3,5,and 7 d,the absorbance values of m BMSCs in the normal control group,hypoxia control group,and hypoxia stent experimental group were significantly different(P<0.05).On the 3 rd,5 th,and 7 th day,in the observation point,the absorbance values of m BMSCs in the hypoxic control group were all higher than those of the normal control group,the difference was statistically significant(P<0.01);compared with the hypoxic control group,the absorbance A values of the hypoxic stent experimental group were increased at 3,5,and 7 d respectively,the difference was statistically significant(P<0.01).In the hypoxic environment,the contents of Tenomodulin protein(4.526±0.002)μmol/L and Scleraxis protein(3.752±0.004)μmol/L in the hypoxic stent experimental group were higher than those in the hypoxia control group Tenomodulin protein(2.831±0.023)μmol/L And Scleraxis protein(2.457±0.032)μmol/L,the hypoxic control group Tenomodulin protein(2.831±0.023)μmol/L and Scleraxis protein(2.457±0.032)μmol/L content were higher than the normal control group Tenomodulin protein(1.542±0.071)μmol/L and Scleraxis protein(1.136±0.056)μmol/L,the difference between the three groups was statistically significant(P<0.01).Conclusion The PLGA/collagen fiber scaffold can effectively promote the proliferation and tendon differentiation of m BMSCs under hypoxia.