创伤性与非创伤性应激对大鼠内皮祖细胞的影响

The effects of traumatic and non-traumatic stress on endothelial progenitor cells in rats

目的观察大鼠在创伤性和非创伤性应激状态下外周血内皮祖细胞数目的变化,探讨可能影响其变化的相关因素。方法 36只健康雄性Wistar大鼠,随机接受颅脑创伤、电休克和冷水游泳刺激,分别于应激前(0h)及应激后3h、6h、24h、48h、72h和7d采集内眦球后静脉丛血,分离单个核细胞,CD34和CD133双荧光标记内皮祖细胞,流式细胞术计数内皮祖细胞数目。结果颅脑创伤组大鼠外周血内皮祖细胞数目于伤后3h即明显减少(P=0.000);随后迅速增加,至6h达峰值水平(P=0.005);至24h降至正常值范围(P=0.728)。而非创伤性应激(电休克和冷水游泳)后3h,大鼠外周血内皮祖细胞数目即开始增加(P=0.000,0.019);至24h达峰值水平(P=0.000,0.004);随后逐渐减少,至72h恢复至正常值范围(P=0.999,0.055)。对照组大鼠各观察时间点外周血内皮祖细胞数目变化,差异无统计学意义(均P>0.05)。结论应激反应可以促进骨髓内皮祖细胞的动员,创伤性应激后内皮祖细胞数目先减少后增加的特征与非创伤性应激明显不同,其短暂性减少可能与组织损伤修复导致的内皮祖细胞消耗有关。

Objective To investigate the change of endothelial progenitor Cells （EPCs） of rat peripheral blood under traumatic and non-traumatic stress and the possible impact factors on it. Methods Thirty-six healthy adult male Wistar rats were randomly divided into traumatic brain injury （TBI） group （n = 9）, electroconvulsive shock （ECS） group （n = 9）, cold water swim （CWS）group （n = 9） and control group （n = 9）. The rat TBI model was established with the fluid,perfusion instrument at 1.01 × 10^5 Pa, ECS model was established in conscious rats at a 1 s 90 mA electrical pulse, and CWS model was established by putting conscious rats in 4 ℃ water swimming 3 min in a diameter 1.50 m, depth 20 cm cylinder. Then peripheral blood was taken respectively from postbulbar venous plexus of inner canthus on pre-stimulation （0 h） and 3 h, 6 h, 24 h, 48 h, 72 h and 7 d after stimulation. Blood samples were first subjected to a Fieoll gradient centrifugation to isolate mononuclear cells with a commercial kit. The isolated cells suspended in fetal bovine serum （FBS） buffer were incubated with both CD34-phycoerythrin （PE） and CD133-fluorescein isothiocyanate （FITC） for 10 min at room temperature. Then the staining cells were analysed in flow cytometry. Results The number of peripheral blood EPCs decreased to a level lower than the baseline significantly at 3 h after TBI （P = 0.000）, and immediately reversely increased and reached to the peak level at 6 h after TBI （P = 0.005）. It decreased to the basel!he at 24 h after TBI （P = 0.728）. But the number of EPCs after non-traumatic stress （ECS and CWS） both significantly increased at 3 h （P = 0.000, 0.019, respectively） and reached to the peak level at 24 h （P = 0.000, 0.004, respectively）, and gradually decreaSed to the baseline at 72 h （P= 0.999, 0.055, respectively）. However, it was not changed conspicuously in the control group （P 〉 0.05, for all）. Conclusion StreSs is a novel method to enhance the mobilization of EPCs from bone marrow to peripheraI blood. The change of EPCs under traumatic stress is different from non-traumatic stress. The transient low level may due to EPCs consumption by brain tissue injury.