Heterotrimeric Gα(i) proteins are regulated by lipopolysaccharide and are anti-inflammatory in endotoxemia and polymicrobial sepsis.
Heterotrimeric Gα(i) proteins are regulated by lipopolysaccharide and are anti-inflammatory in endotoxemia and polymicrobial sepsis.
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DOI：
10.1016/j.bbamcr.2011.01.012
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发表时间：
2011-03
期刊：
影响因子：
--
通讯作者：
Cook JA
中科院分区：
文献类型：
--
作者：
Fan H;Li P;Zingarelli B;Borg K;Halushka PV;Birnbaumer L;Cook JA
以前的研究表明异源三聚体Gαi蛋白在脂多糖（LPS）诱导的炎症反应中起作用。我们假设Toll样受体（TLR）信号调节Gαi蛋白，Gαi蛋白在内毒素血症和多微生物脓毒症中具有抗炎作用。用LPS刺激Raw 264.7细胞，用Gαi蛋白活化测定法免疫沉淀Gαi-GTP蛋白复合物。在随后的体内研究中，在内毒素血症之前给予Gαi蛋白抑制剂百日咳毒素（PTx）或Gi蛋白激动剂mastoparan（MP-7）。测定LPS诱导的促炎细胞因子和死亡率。为了检查Gαi2在脓毒症中的作用，对Gαi2（−/−）和野生型（WT）小鼠进行盲肠结扎和穿孔（CLP），并每24小时监测一次，持续120小时。CLP后24小时处死其他小鼠。采集腹膜液、血液和组织样本。检测血浆促炎细胞因子的产生，腹腔液、血液和肺组织中的细菌负荷，肺和肝脏中的髓过氧化物酶（MPO）活性以及脾脏中不同免疫细胞群。我们发现Gαi蛋白被LPS迅速激活，然后迅速失活。这些研究首次提供了Gαi蛋白受TLR信号调节的直接证据。在随后的研究中，PTx增加LPS诱导的血浆TNFα、IL-6，而MP-7抑制LPS诱导的TNFα，并降低LPS诱导的死亡率。在脓毒症研究中，与WT小鼠相比，Gαi2（−/−）小鼠CLP后的存活率显著降低。与WT小鼠相比，Gαi2（−/−）中CLP诱导的血浆TNFα、IL-6、腹腔液、血液和肺组织中的细菌负荷以及肺和肝MPO活性显著增加。与WT小鼠相比，Gαi2（−/−）小鼠也表现出Th 1和Th 2应答增加。综上所述，Gαi蛋白被LPS激活并负性调节内毒素血症和脓毒症。了解Gαi2蛋白在脓毒症炎症反应中的调节作用，可能为脓毒症的治疗提供新的靶点。
Previous studies have implicated a role of heterotrimeric Gαi proteins in lipopolysaccharide (LPS) -induced inflammatory responses. We hypothesized that Toll-like receptor (TLR) signaling regulates Gαi proteins, which are anti-inflammatory in endotoxemia and polymicrobial sepsis. Raw 264.7 cells were stimulated with LPS and the Gαi-GTP protein complex was immunoprecipitated with a Gαi protein activation assay. In subsequent in vivo studies, the Gαi protein inhibitor pertussis toxin (PTx) or Gi protein agonist mastoparan (MP-7) were administrated prior to endotoxemia. LPS-induced pro-inflammatory cytokines and mortality were determined. To examine the role of Gαi2 in sepsis, Gαi2 (−/−) and wildtype (WT) mice were subjected to cecal ligation and puncture (CLP) and monitored every 24 hours for 120 hours. Other mice were sacrificed 24 hours after CLP. Peritoneal fluid, blood, and tissue samples were collected. Plasma pro-inflammatory cytokine production, bacterial load in peritoneal fluid, blood and lung tissue, myeloperoxidase (MPO) activity in lung and liver and different immune cell populations in spleen were studied. We found that Gαi proteins are rapidly activated by LPS followed by rapid inactivation. These studies provide the first direct evidence that Gαi proteins are modulated by TLR signaling. In following studies, PTx augmented LPS-induced plasma TNFα, IL-6, whereas MP-7 suppressed LPS-induced TNFα and decreased LPS-induced mortality. In sepsis studies, the survival rate post-CLP was significantly decreased in the Gαi2 (−/−) mice compared to WT mice. CLP-induced plasma TNFα, IL-6, bacterial load in peritoneal fluid, blood and lung tissue and lung and liver MPO activity were significantly increased in Gαi2 (−/−) compared to WT mice. Gαi2 (−/−) mice also exhibited increased Th1 and Th2 responses compared to WT mice. Taken together, Gαi proteins are activated by LPS and negatively regulate endotoxemia and sepsis. Understanding the role of Gαi2 protein in regulation of the inflammatory response in sepsis may provide novel targets for treatment of sepsis.
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