INTESTINAL PERFUSION AND PERMEABILITY IN SEPSIS

描述：（改编自申请人的摘要）本次竞争 更新应用基于肠道上皮细胞的假设 细胞酸中毒会破坏屏障功能。 在当前 资助的研究中，研究者表明回肠粘膜酸中毒 猪体内的粘膜通透性增加，并且暴露 粘膜细胞 (CaCo-2BBe) 单层对酸的作用也会导致 体外上皮通透性。 研究人员的研究还 表明酸中毒会促进脂质过氧化，增加细胞内 Fe2 ，并部分消耗 CaCo-2BBe 肠细胞中的细胞 ATP。 这些研究还表明，钙蛋白酶和 磷脂酶改善酸诱导的粘膜通透性过高。 这些研究共同支持了细胞内 钙相关紊乱可能在酸中毒的发病机制中很重要 引起粘膜屏障功能障碍。 目前的提案将延长 CaCo-2BBe 细胞和单核细胞的研究：1) 测量 细胞内 [H ], 2) 评估酸中毒和 ATP 的影响 上皮对水溶性大分子的通透性消耗 以及 LPS 和大肠杆菌等颗粒物。 在 此外，该提案将重点关注细胞骨架完整性及其 与酸中毒所致粘膜细胞通透性障碍的关系 在 CaCo-2BBe 单层中。 研究还将评估其效果 [Ca2]i 导致肠上皮细胞酸中毒或中度 ATP 耗竭，以及 A23187、BAPTA 和各种磷脂酶修饰剂的作用 钙蛋白酶和 PKC 对酸性和/或低 ATP 水平诱导的粘膜 通透性功能障碍。 所有上述研究均应 体外 CaCo-2BBe 细胞系中的载体。 最后，实验将 确定回肠粘膜通透性的各种决定因素 失血性损伤大鼠模型的体内研究中的功能障碍。 这些研究将采用体内方法来确定 LPS 的易位和大分子穿过回肠的渗透性 上皮。

DESCRIPTION: (Adapted from the applicant's abstract) This competing renewal application is based on the hypothesis that gut epithelial barrier function is deranged by cellular acidosis. In the currently funded research, the investigator has shown that ileal mucosal acidosis in pigs increases mucosal permeability in vivo, and that exposure of mucosal cell (CaCo-2BBe) monolayers to acid also induce an increase in epithelial permeability in vitro. The investigator's studies have also shown that acidosis promotes lipid peroxidation, increases intracellular Fe2+, and partly depletes cellular ATP in the CaCo-2BBe enterocytes. These studies have demonstrated also that inhibition of calpain and phospholipases ameliorated acid induced mucosal hyperpermeability. These studies collectively support the concept that intracellular calcium related derangements may be important in the pathogenesis of acid induced mucosal barrier dysfunction. The present proposal will extend the studies of CaCo-2BBe cells and monocytes by: 1) measuring intracellular [H+], 2) assessing the effect of acidosis and ATP depletion on epithelial permeability to water soluble macromolecule as well as to particulate matter such as LPS and E.coli bacteria. In addition the proposal will focus on cytoskeletal integrity and its relationship to acidosis induced mucosal cell permeability dysfunction in CaCo-2BBe monolayers. Studies will evaluate also the effect of acidosis or moderate ATP depletion in the enterocytes on [Ca2+]i, and the effects of A23187, BAPTA, and various modifiers of phospholipases calpains and PKC on the acid and/or low ATP level induced mucosal permeability dysfunction. all of the aforementioned studies are to be carrier in the CaCo-2BBe cell line in vitro. Finally, experiments will ascertain the various determinants of ileal mucosal permeability dysfunction in the in vivo studies in a rat model of hemorrhagic injury. These studies will employ in vivo methodology to determine both translocation of LPS and permeability of macromolecules across the ileal epithelium.