基底刚度对成纤维细胞生物学行为的影响

目的 了解基底刚度对Fb增殖、迁移和整合素β1表达的影响. 方法 将Fb接种于刚度为(16.2±0.5)、(19.8±1.1)、(200.1±2.6)kPa的硅胶基底上.进行如下检测.(1)分别连续培养5 d或6d,进行细胞计数、噻唑蓝法检测细胞增殖活性(吸光度值表示).(2)培养3d,检测细胞周期,计算增殖指数(PI).(3)划痕实验后培养0(当日)、1、2、3 d,测定Fb迁移率.(4)培养2 d,流式荧光法检测细胞中整合素β1表达.对部分数据进行单因素方差分析. 结果 (1)细胞计数、噻唑蓝法检测均显示,Fb增殖速度及活性均随着硅胶基底刚度的增强而增加.细胞周期检测显示:在刚度为(16.2±0.5)、(19.8±1.1)(200.1±2.6)kPa的硅胶基底上,细胞的PI分别为24.8％、27.4％、32.4％.(2)培养2d,在刚度分别为(19.8±1)、(200.1±2.6) kPa的硅胶基底表面上,Fb迁移率分别为(91.4±5.1)％(100.0±1.3)％,均明显高于刚度为(16.2±0.5)kPa硅胶基底表面的Fb迁移率[(55.8±6.8)％,F值分别为3.5、4.0,P值均小于0.01].(3)刚度为(16.2±0.5)kPa硅胶基底表面的Fb中整合素β1表达率最低,仅43.2％；刚度为(200.1±2.6)kPa硅胶基底表面的Fb中整合素β1表达率最高(81.3％]. 结论 基底刚度对Fb在创面愈合和瘢痕形成过程中的增殖迁移有较大影响,这一效应与其调控Fb整合素β1表达作用相关.

Effect of substrate stiffness on biological behavior of flbroblasts

Objective To study the effect of substrate stiffness on proliferation,migration of fibroblast and integrin β1 expression in fibroblast.Methods Fibroblasts were inoculated on silicon substrate with stiffness of ( 16.2 ±0.5 ),( 19.8 ± 1.1 ),and ( 200.1 ± 2.6) kPa.After being cultured for 5 days or 6 days,cells were counted and cell proliferative activities ( recorded as absorbance value) were assessed with methyl thiazolyl blue (MTT).After being cultured for 3 days.cell cycle was detected and proliferation index (PI) was calculated,The cell scratch test was used for determination of cell migration rate on post scratch day (PSD) 0 ( the day of scratch ),1,2,and 3.After being cultured for 2 days,the expression of integrin β1 was determined by flow cytometry with fluorescence.Data were processed with one-way analysis of variance,Results (1)The proliferaive speed and proliferative activity of fibroblasts were all increased along with the increase in substrate stiffness.PI of fibroblasts inoculated on silicon substrate with stiffness of (16.2 ±0.5),(19.8 ± 1.1),and (200.1 ±2.6) kPa was respectively 24.8％,27.4％,32.4％.On PSD 2,migration rate of fibroblasts inoculated on silicon substrate with stiffness of ( 19.8 ± 1.1 ) and ( 200.1 ± 2.6 ) kPa was respectively ( 91.4 ± 5.1 ) ％,( 100.0 ± 1.3 ) ％,which were higher than that of fibroblasts inoculated on silicon substrate with stiffness of ( 16.2 ± 0.5 ) kpa [ ( 55.8 ± 6.8 ) ％,with F value respectively 3.5,4.0.P values all below 0.01 ].( 3 ) The expression rate of integrin β1 in fibroblasts inoculated on silicon substrate with stiffness of ( 16.2 ±0.5 ) kPa was the lowest (43.22％),and that in fibroblast inoculated on silicon substrata with stiffness of ( 200.1 ± 2.6 ) kPa was the highest ( 81.26％ ).Conclusions Substrate stiffnese may have a great effect on proliferation and migration of fibroblast during the process of wound healing and scar formation,which can be related to regulation of intergrin β1 expression.