冷却治疗对劳力性热射病大鼠肾脏AQP2 mRNA及蛋白表达的影响

Effects of Cooling Therapy on AQP2 mRNA and AQP2 Protein Expression in Kidney of Rats with Exertional Heat Stroke

目的:研究冷却治疗对劳力性热射病(EHS)大鼠血液浓缩程度及水通道蛋白合成的影响,探讨肾脏水通道蛋白基因转录中AQP2 mRNA表达及蛋白表达在机体水代谢稳态中的作用。方法:将51只雄性SD大鼠随机分为6组,常温对照组(C组,n=8)、高温运动即刻组(E组,n=9)、高温运动休息1组(ER1组,n=9)、高温运动冷浸1组(EC1组,n=8)、高温运动休息2组(ER2组,n=9)和高温运动冷浸2组(EC2组,n=8)。参加高温运动的大鼠在36℃高温、75%高湿环境中运动至力竭,依据热射病经典模型测量直肠温度作为核心温度,当直肠温度上升到约42℃时,建立EHS大鼠模型。采用19℃冷水浸泡5min作为EHS大鼠冷却治疗方法。C组不进行任何处理,E组仅参加高温运动。高温运动后,ER1组常温休息5min;EC1组冷浸5min,ER2组常温休息5min+60min;EC2组冷浸5min,常温休息60min。麻醉取血、取肾脏,采用全自动血细胞分析仪测试红细胞压积(Hct)和血红蛋白(Hb),Real Time-PCR法检测肾脏AQP2 mRNA表达,Western Blot法检测肾脏AQP2蛋白表达。结果:(1)高温运动至力竭后,各组大鼠核心温度较运动前显著性升高(P<0.01,P<0.05)。E组与C组相比,Hct升高(P<0.05),而Hb无明显变化;调节水代谢的AQP2 mRNA表达以及AQP2蛋白表达上调(P<0.01)。(2)冷浸后,EC1组与ER1组相比,大鼠核心温度下降(P<0.01);AQP2 mRNA表达下调(P<0.01)。EC2组与ER2组相比,大鼠核心温度下降(P<0.01);AQP2 mRNA表达下调(P<0.05)。EC1组与E组相比,AQP2 mRNA表达和AQP2蛋白表达均下调(P<0.01,P<0.05)。EC2组与E组相比,Hct和Hb下降不明显;AQP2 mRNA表达和AQP2蛋白表达均下调(P<0.01,P<0.05)。结论:冷浸可以有效降低热射病大鼠机体核心温度;冷浸诱导水通道蛋白AQP2 mRNA表达及AQP2蛋白表达显著性下调,导致水通道蛋白在肾脏的合成减弱;冷浸对红细胞压积Hct及血红蛋白Hb没有明显影响。

Objective To investigate the effect of cooling therapy on blood concentration level and the synthesis of aquaporin in rats with exertional heat stroke,and to explore the role of AQP2 mRNA expression and AQP2 protein expression in the water homeostasis of the kidney during the aquaporin gene transcription.Methods Fifty-one male Sprague-Dawley rats were randomly divided into a normal temperature control group(C,n=8)and a high temperature exercise group which was further divided in to an instant group(E,n=9),a rest group 1(ER1,n=9),a rest group 2(ER2,n=9),a cold water im mersion group 1(EC1,n=8)and a cold water immersion group 2(EC2,n=8).According to the classical model of heat stroke,the rectal temperature was measured as the core temperature.The rat model of EHS was established through exercising to exhaustion and raising the rectal temperature to about 42℃ in the 36℃-high-temperature and 75%-high-humidity environment.EHS rats were immersed in cold water at 19℃ for 5 min for cooling treatment.Group C received no treatment,while group E only par ticipated in exercise in high-temperature environment.After having exercised in high-temperature envi ronment,ER1 group took a rest for 5 minutes in normal temperature,EC1 group accepted a cold-wa ter immersion for 5 minutes,ER2 group took a rest for 65 minutes in the normal temperature,EC2 group accepted a cold-water immersion for 5 minutes followed by a 60-minute rest in the normal tem perature.After anesthesia,both blood and kidney were taken.Hematocrit(Hct)and hemoglobin(Hb)were measured using an automatic blood cell analyzer.Quantitative real time polymerase chain reaction and Western Blotting were used to detect the AQP2 mRNA and AQP2 protein expression in the kid ney.Results After exercising to exhaustion in high-temperature environment,the core temperature of rats in all groups increased significantly compared to before the intervention(P<0.01 or P<0.05).Com pared with group C,a significant increase was observed in the average Hct and expression of AQP 2 mRNA and protein of group E(P<0.05,P<0.01),but not in the Hb.After immersion in the cold water,the core temperature and expression of AQP2 mRNA of group EC1 and EC2 decreased signifi cantly compared with group ER1 and ER2 accordingly.Compared with group E,the AQP2 mRNA and protein expression of group EC1 and EC2 were significantly down-regulated(P<0.01 and P<0.05 for both),while the down-regulation of Hct and Hb was not significant.Conclusion Cold water immer sion can effectively reduce the core temperature of EHS rats and decrease the expression of AQP2 mRNA and protein,but has no significant effect on Hct and Hb.