脓毒症大鼠通过降低肌浆网钙摄取和SERCA1表达损害膈肌舒张功能

目的 探讨脓毒症急性期大鼠膈肌舒张功能障碍的机制.方法 雄性清洁级SD大鼠36只.随机数字表法分为:假手术组(S组)12只、造模组(CLP组)24只.对造模组行盲肠结扎穿孔手术复制脓毒症模型,造模成功后随机分为脓毒症CLP-6 h组和脓毒症CLP-12 h组,各12只,分别于术后6 h和12 h测定单刺激肌颤搐(Pt)、最大强直收缩力(Po)、半舒张时间(1/2RT)、收缩时间(TPT)、最大收缩期上升速率(+dF/dt)和最大舒张期下降速率(-dF/dt);运用Fura-2法测定膈肌肌浆网钙最大摄取率和释放率,Western blotting法检测肌浆网SERCA1、SERCA2和RyR表达.结果 造模后6 h和12 h组大鼠较S组半舒张时间显著延长,-dF/dt显著下降(P<0.01),而仅仅在12 h组大鼠膈肌+dF/dt、Pt、Po和肌浆网钙的峰值释放率较S组显著降低(P<0.01),6 h组大鼠未见明显变化(P>0.05);造模后12 h组大鼠膈肌SERCA1和RyR表达显著下降(P<0.01),SERCA2和Ca2+-ATP酶活性无明显改变(P>0.05).结论 脓毒症大鼠急性期12 h内膈肌收缩和舒张功能显著受损,舒张功能障碍可能与肌浆网钙摄取功能下降及SERCA1低表达有关,肌浆网释放和摄取功能及RyR表达下降共同导致了收缩功能的下降.

Lowered sarcoendoplasmic reticulum calcium uptake and diaphragmatic SERCA1 expression contribute to diaphragmatic contractile and relaxation dysfunction in septic rats

Objective The explore the mechanism responsible for diaphragmatic contractile and relaxation dysfunction in a rat model of sepsis. Methods Thirty-six adult male Sprague-Dawley rats were randomized equally into a sham-operated group and two model groups of sepsis induced by cecal ligation and puncture (CLP) for examination at 6 and 12 h following CLP (CLP-6 h and CLP-12 h groups). The parameters of diaphragm contractile and relaxation were measured, and the calcium uptake and release rates of the diaphragmatic sarcoendoplasmic reticulum (SR) and the protein expressions of SERCA1, SERCA2 and RyR in the diaphragmatic muscles were determined. Results The half-relaxation time of the diaphragm was extended in both the CLP-6 h and CLP-12 h groups with significantly reduced maximum tension declinerate and the peek uptake rate of SERCA (P<0.01). Diaphragmatic maximum twitch force development rate, the maximal twitch, tetanus tensions and the peek release rate of SR decreased only at 12h after CLP (P<0.01). The expression levels of SERCA1 protein decreased significantly in the diaphragmatic muscles at 12h following CLP (P<0.01) while SERCA2 expression level and SERCA activity showed no significant changes. Conclusion In the acute stage of sepsis, both the contractile and relaxation functions of the diaphragm are impaired. Diaphragmatic relaxation dysfunction may result from reduced calcium uptake in the SR and a decreased level of SERCA1 in the diaphragmatic muscles.