AMPK-MLN信号通路在急性腹膜炎腹肌肌浆网钙泵功能障碍中的作用

The abdominal wall muscles couldn’t relax satisfactorily among patients with acute peritonitis (AP) which is usually induced by gastric perforation even given abundant neuromuscular blockades. Relaxation of skeletal muscles depends on uptake of Ca2+ from cytoplasm into the lumen of the SR through sarcoendoplasmic Ca2+-ATPase(SERCA). Our previous researches predicted that the uptake function of SERCA was impaired whereas SERCA protein expression didn’t change. To study the mechanisms of SERCA impairment in AP. Myoregulin(MLN) is a micropeptide containing 46 amino acid encoded by a skeletal muscle-specific RNA annotated as a putative long non-coding RNA. MLN expressed in all skeletal muscle inhibits SERCA and impedes Ca2+ uptake into the SR. Phosphorylation of N-terminal sequence of MLN could diminish its ability to inhibit SERCA activity. We speculate that tetanic contraction of skeletal muscle in AP activates adenosine monophosphate-activated protein kinase(AMPK) pathway decreasing the phosphorylation level of MLN or increasing MLN protein expression, which reinforces the function of SERCA and eventually results in calcium overload and relaxation dysfunction of the abdominal muscle. We will detect the relationship between the change of MLN/p-MLN and SERCA impairment by establishing gastric perforation model in rats. Furthermore, we will test the role of MLN in SERCA impairment of AP through establishing MLN knockout model in rats. Meanwhile we will detect whether the changes of MLN/p-MLN is via AMPK pathway.

急性腹膜炎（胃穿孔为常见原因）患者手术时，腹肌不能很好的松弛。骨骼肌舒张是由ATP依赖的SERCA对细胞质内Ca2+回收完成的。我们前期实验显示，急性腹膜炎时SERCA回收Ca2+功能发生障碍，而蛋白表达量未改变。本课题目的为探讨急性腹膜炎时SERCA功能障碍的机制。MLN是新近发现的由lncRNA编码的含46个氨基酸的短肽，在成年骨骼肌细胞中对SERCA起抑制作用，其N端磷酸化后抑制作用降低。我们推测，急性腹膜炎时，骨骼肌细胞强直收缩通过激活 AMPK通路，降低MLN的磷酸化水平或通过转录调节增加MLN的表达量，使其对SERCA抑制作用增强，造成细胞内Ca2+超载，导致腹肌舒张功能障碍。我们建立胃穿孔大鼠模型，研究腹肌MLN/p-MLN变化与SERCA功能障碍的关系；并用MLN敲除鼠进一步验证MLN在腹肌SERCA 功能障碍中的作用；最后验证 AMPK通路与MLN/p-MLN变化的关系。

全身炎症性疾病（包括胃穿孔引起的腹膜炎、肠坏死，及其进一步引起的脓毒症等）可导致多器官脏器损伤、甚至衰竭。而脓毒症所导致的骨骼肌功能障碍统称为脓毒症肌病。而这类疾病导致的骨骼肌功能障碍的机制及保护措施还未明确。本项目通过建立全身炎症性疾病模型（胃穿孔模型、脓毒症模型），采用形态学、离体肌肉力学、分子生物学等技术手段，从整体、肌肉条等层次，观察到炎症性疾病导致骨骼肌功能下降及神经肌肉接头处AChE活性降低，与其加重骨骼肌氧化应激损伤，导致骨骼肌收缩舒张相关蛋白如SERCA的表达量及活性下降有关。同时我们的研究还发现骨骼肌的重要调节因子NRG-1在脓毒症时显著下调，.我们的研究证实了外源性补充NRG-1β可以减轻脓毒症膈肌的氧化应激损伤并改善了膈肌的收缩功能。另外，本部分研究还发现，NRG-1β部分逆转了AChE 活性降低，可能与其发挥了抗氧化应激作用，降低了 ROS 生成，减轻了对AChE的攻击有关。本课题从新的角度阐明全身炎症性疾病引起的骨骼肌功能障碍的发生机制，为此类疾病的防治提供重要依据。