不同浓度米诺环素刺激骨髓细胞对小鼠体外肝细胞损伤的影响

Effect of different concentrations minocycline combined with bone marrow cells on liver cell in septic mice

(1)目的探讨不同浓度米诺环素刺激的骨髓细胞对体外培养的小鼠肝细胞损伤的保护作用。(2)方法获取C57BL/6小鼠同基因背景下绿色荧光骨髓细胞,分为6组:对照组(b0组)、不同浓度米诺环素刺激组(b_(1~5)组)。b0组为单纯骨髓细胞培养组,b_(1~5)组分别加入不同浓度(5、10、20、50、100μg/mL)米诺环素;将b_(0~5)组置于37℃、5%CO_2培养箱培养培养24小时后,各取出一部分骨髓细胞用MTT法检测其增生率;获取C57BL/6小鼠肝细胞,分为8组(a、a_L、a_(0~5)组),除对照组(a组,为单纯肝细胞培养组)外,其余各组加入终浓度100μg/mL的脂多糖(LPS),建立肝细胞损伤模型。a_(0~5)组分别相应加入b_(0~5)组培养的骨髓细胞2×10~6个,a、aL组加入等容积的PBS液,置于37℃、5%CO_2培养箱培养,1~2天换液,动态观察肝细胞生长情况并检测肝细胞的凋亡率。(3)结果与b_0组相比,不同浓度米诺环素刺激24小时后骨髓细胞的增生率有所不同,b_1组(5μg/mL)最高,b_5组(100μg/mL)最低。与a组相比较,a_L肝细胞受损严重,凋亡率增加;a_(0~5)组肝细胞有不同程度的增殖,其凋亡率有所降低,以低浓度5μg/mL效果最优。(4)结论本次实验显示5μg/mL的米诺环素刺激的骨髓细胞对体外受损肝细胞有保护作用,其机制可能为米诺环素提高骨髓细胞增生率,增加骨髓细胞活性,从而降低肝细胞凋亡和促进肝细胞增殖。

Objective To explore protective effect of different concentrations minocycline com- bined with bone marrow cells on liver cell in septic mice.Methods GFP-l-bone marrow cells were di- vided into 6 groups ： control group （b0 group） 、 minocycline groups （b1-5 groups）, bl- s groups which were added at different concentrations （5.10.20.50. 100gg/mL） of minocycline respectively, All groups were placed in 37℃ 5% CO2 saturated humidity to be cultured for 24h,Then some of the cells were took out for the proliferation rate of bone marrow cells by MTT methods.Hepatocytes were di- vided into 8 groups：control group （a group）.hepatocytes injury groups（ac.a0-5 groups） which were used LPS at the concentration of 100μg/mL to induce injury.GFP＋ bone marrow cells from group b0-5 were added to group a0-5 while a, ac groups were received equal volume of PBS instead. All groups were placed in 37℃ ,5% CO2 saturated humidity to be cultured for 24h,the growth of hepato- cytes was observed and the apoptosis were detected.Results Compared with the b0 group,different concentrations of minocycline stimulated 24 hours after the proliferation rate of bone marrow was dif-ferent,The proliferation rate of bone marrow , proliferation rate of bone marrow cells in b5 group （100μg/mL） was the lowest.Compared with the a group,aL hepatocytes were damaged and the apoptosis rate was increased.The hepatocytes of group a0-a had different degrees of proliferation,and the apoptotic rate was decreased with the lowest con- centration of 5μg/mL.Conclusin In this experiment, 5μg/mL minocycline combined with bone mar- row ceils in vitro damage to liver ceils have protective effect, the mechanism may be minocycline to improve bone marrow cell proliferation rate, increase bone marrow cell activity, thereby reducing liver cell apoptosis death and promote hepatocyte proliferation.