摘要
目的研究中波紫外线（UVB）联合纳米碳酸钙处理对HaCaT细胞增殖、凋亡的影响及其机制。方法取对数生长期HaCaT细胞分为对照组、UVB照射组、纳米碳酸钙组和联合处理组4组,UVB照射组和联合处理组接受累计照射剂量为2.97×10^-2J/cm^2的UVB照射,对照组和纳米碳酸钙组给予同等条件下假照射。照射完毕后,对照组和UVB照射组分别用含体积分数为10.00%磷酸盐缓冲液的高糖达尔伯克氏改良伊格尔培养基（DMEM）继续培养,纳米碳酸钙组和联合处理组分别用含质量浓度为250 mg/L纳米碳酸钙的高糖DMEM继续培养。于培养后0、6、12、18、24 h收获细胞,采用噻唑蓝法检测细胞活性,流式细胞术检测细胞凋亡率,免疫印迹法检测凋亡相关蛋白P53和Caspase-3表达情况。结果与同时间点对照组比较,除UVB照射组在6 h时间点差异无统计学意义外（P〉0.05）,其余各处理组在6~24 h各个时间点HaCaT细胞活力均下降（P〈0.05）;联合处理组HaCaT细胞活力在6~24 h各个时间点均低于同时间点UVB照射组（P〈0.05）,在18和24 h均低于同时间点纳米碳酸钙组（P〈0.05）。UVB照射组HaCaT细胞凋亡率高于对照组（P〈0.05）,联合处理组HaCaT细胞凋亡率高于其余3组（P〈0.05）。P53、Caspase-3蛋白表达水平在UVB照射组和纳米碳酸钙组HaCaT细胞均较对照组表达上调;联合处理组HaCaT细胞P53和Caspase-3蛋白的表达水平均高于其余3组。结论 UVB联合纳米碳酸钙处理对HaCaT细胞的损伤效应较单一损伤因素更严重,表现为抑制细胞增殖,诱导细胞凋亡,促进P53及Caspase-3蛋白表达。
Objective To explore the effects of UVB and Nano-CaCO_3 co-treatment in inhibiting cell proliferation and inducing cell apoptosis in human HaCaT keratinocytes,and to explore its possible mechanisms. Methods HaCaT cells（ logarithmic growth phase） were divided into control group,UVB group,Nano-CaCO_3 group and co-treatment group. UVB group and co-treatment group were irradiated with UVB irradiation with cumulative exposure dose of 2. 97 × 10^-2J / cm^2.Control group and Nano-CaCO_3 group were irradiated with UVB irradiation with cumulative exposure dose of zero on equal terms. After that,control group and UVB group were treated with 10. 00% phosphate buffer solution in high-sugar Dulbecco＇s modified Eagle＇s medium（ DMEM） and incubated,Nano-CaCO_3 group and co-treatment group were treated with high-sugar DMEM with Nano-CaCO_3 at 250 mg / L mass concentration and incubated. Subsequently,HaCaT cells were harvested at incubation time of 0,6,12,18 and 24 hours. Then methyl thiazolyl tetrazolium assay was performed to estimate cellular proliferative activity,flow cytometry was used to detect cell apoptosis,Western blot was used to detect the expression of P53 and Caspases-3 protein. Results Contrast to control group at the parallel incubation time points of 6-24 hours,the cell viability of HaCaT cells was significant decreased in the other three groups（ P〈0. 05） except for UVB group at incubation time of 6 hours（ P〉0. 05）. The cell viability of co-treatment group was lower than UVB group at all the incubation time（ P〈0. 05）,and lower than Nano-CaCO_3 group at incubation time of 18 and 24 hours（ P〈0. 05）.The apoptosis rate of HaCaT cells in UVB group was higher than control group（ P〈0. 05）,and which in co-treatment group was higher than the other three groups（ P〈0. 05）. Contrast to control group,the protein expressions of P53 and Caspases-3 in HaCaT cells were upregulated in UVB group and Nano-CaCO_3 group. In co-treatment group,the protein expressions of P53 and Caspases-3 were upregulated contrast to the other three groups. Conclusion Contrast to single damage of UVB or Nano-CaCO_3,co-treatment of UVB with Nano-CaCO_3 increased damage to HaCaT cells,likely by inhibiting proliferative activity,inducing apoptosis,and enhancing protein expressions of P53 and Caspases-3.
出处
《中国职业医学》
CAS
北大核心
2016年第6期657-661,共5页
China Occupational Medicine
基金
广东省职业病防治重点实验室(2012A061400007)
广东省医学科研基金项目(B2014013)
广东省科技计划对外科技合作项目(2013B051000066)
