琥珀酸对人外周血中性粒细胞凋亡的影响

目的 了解琥珀酸对人外周血中性粒细胞(PMN)凋亡的影响,探讨其致病机制.方法 体外孵育PMN,将PMN浓度调至5×106个/ml.在细胞中加入琥珀酸刺激,根据所加琥珀酸浓度分为5、10、20、30 mmol/L琥珀酸组;对照组加入常规培养液.取各组细胞培养上清液,检测其活性氧含量.5、20 mmol/L琥珀酸组细胞于处理前及处理后2、4、6、8、10 h分别取细胞悬液1 ml,行半胱氨酸天冬氨酸蛋白酶3(caspase-3)活性及细胞凋亡率检测.结果 5、10、20、30 mmol/L琥珀酸组活性氧含量(0.437±0.056、0.432±0.024、0.395±0.049、0.386±0.010)均低于对照组(0.505±0.028,P＜0.05).随着孵育时间延长,各组细胞caspase-3活性均逐渐增加,但与对照组比较,5 mmol/L琥珀酸组caspase-3活性降低,20 mmol/L琥珀酸组则升高(P＜0.05).对照组细胞处理前凋亡率为(6.1±1.1)%,处理后2 h为(13.2±2.0)%,10 h达(27.7±3.7)%;5 mmol/L琥珀酸组细胞凋亡率除处理后4 h外其余时相点均低于对照组(P＜0.05);20 mmol/L琥珀酸组细胞处理后4～10 h凋亡率均高于对照组(P＜0.05).结论 低浓度琥珀酸抑制PMN凋亡,而高浓度琥珀酸可促进PMN凋亡.细菌感染时,通过代谢产物琥珀酸抑制PMN免疫功能.

The influence of succinic acid on the apoptosis of polymorphonuclear neutrophil in peripheral blood

OBJECTIVE: To investigate the influence of succinic acid on the apoptosis of polymorphonuclear neutrophil (PMN) in human peripheral blood, and to explore its role in infection. METHODS: PMNs were incubated in vitro, and its concentration was adjusted to 5 x 10(6)/mL. Then the cells were divided into normal control group and 5,10, 20, 30 mmol/L succinic acid groups according to different concentrations of succinic acid added into the medium. The supernatant of the cultures in each groups were collected to determine the superoxide content. 1 mL cell suspension was collected from 5, 20 mmol/L succinic acid groups before treatment and at 2, 4, 6, 8, 10 post-treatment hours (PTH) for the determination of caspase-3 activity and the apoptosis rate. RESULTS: The content of superoxide in 5, 10, 20, 30 mmol/L succinic acid groups (0.437 +/- 0.056, 0.432 +/- 0.024, 0.395 +/- 0.049, 0.386 +/- 0.010) was significantly lower than that in control group (0.505 +/- 0.028, P < 0.05). The caspase-3 activity in each group increased along with the incubation time, but was in lower concentration in 5 mmol/L succinic acid group and in higher concentration in 20 mmol/L succinic acid group when compared with that in control group (P < 0.05). The apoptosis rate of PMN in control group was (6.1 +/- 1.1)% before incubation, and it reached (13.2 +/- 2.0)% at 2 PTH, and (27.7 +/- 3.7)% at 10 PTH. The apoptosis rate of PMN in 5 mmol/L succinic acid group was lower than that in control group except that at 4 PTH (P < 0.05). On the other hand, the apoptosis rate in 20 mmol/L succinic acid group (during 4-10 PTH) were obviously higher at each time points compared with the control group (P < 0.05). CONCLUSION: Low concentration of succinic acid can suppress the apoptosis of PMN, while high concentration of succinic acid has an opposite effect. It is known that bacteria can produce succinic acid.