A Novel Cytoprotective Peptide for GI Epithelial Cell

描述（由申请方提供）：肠屏障功能破坏可能是介导炎症性肠病（IBD）和肠源性脓毒症（GDS）受试者发病率和死亡率的常见发病机制。如果是这样，通过增加连接粘膜上皮细胞的紧密连接蛋白和粘附连接蛋白的积累来保护粘膜屏障功能和结构的治疗策略可用于治疗这些病症。我们最近的特点是一种新的18 kDa的蛋白质AMP-18，这是在人类和其他6种哺乳动物的胃窦粘膜上皮细胞合成，并具有促有丝分裂和运动的特性。在人结肠上皮细胞中发现了一个来自该蛋白中心结构域的21肽，（Caco-2/bbe）培养物受到用于诱导小鼠结肠炎的氧化剂吲哚美辛或葡聚糖硫酸钠（DSS）的损伤，细胞培养物中的研究表明，这种AMP肽激活p38 MAP激酶，增加特异性紧密连接蛋白的积累，（occludin，ZO-1，claudin-5），粘附连接蛋白（E-cadherin）和热休克蛋白（hsp 25，hsp 72），并保护暴露于细胞松弛素D的细胞中的肌动蛋白微丝网络。当给予小鼠时，AMP肽增加结肠粘膜中过度磷酸化闭合蛋白的含量。为了确定AMP肽的这些生物学作用是否在体内赋予结肠细胞保护，在屏障受损的两种重要动物模型IBD和GDS中测试其有效性。用AMP肽预处理小鼠延迟了血性腹泻的发作，并减少了给予DSS诱导结肠炎的动物的体重减轻。在GDS模型中，该肽还可以防止在部分肝切除术和术后食物剥夺的手术应激后给予盲肠内铜绿假单胞菌的小鼠死亡。我们的目标是在IBD和GDS的细胞培养和体内模型中获得额外的支持，以证明AMP肽通过增加紧密连接蛋白（如occludin）的积累来保护结肠粘膜屏障结构和功能。确认和扩展AMP肽的细胞保护、促有丝分裂和运动发生作用的研究将进一步支持开发这种分子作为治疗剂，不仅治疗IBD和GDS，而且治疗由肠紧密连接破坏和随后增加粘膜对细菌及其产物的渗透性介导的其他GI疾病。

DESCRIPTION (provided by applicant): Disruption of intestinal barrier function may be a common pathogenetic mechanism that mediates morbidity and mortality in subjects with inflammatory bowel disease (IBD) and gut-derived sepsis (GDS). If so, a therapeutic strategy that protects mucosal barrier function and structure by increasing accumulation of tight and adhesion junction proteins that connect mucosal epithelial cells could be used to treat these conditions. We recently characterized a novel 18-kDa protein called AMP-18, that is synthesized in epithelial cells of the gastric antrum mucosa of humans and 6 other mammals, and has mitogenic and motogenic properties. A 21-mer peptide derived from the central domain of the protein was found to be cytoprotective in human colonic epithelial cell (Caco-2/bbe) cultures subjected to injury by an oxidant, indomethacin, or dextran sulfate sodium (DSS) used to induce colitis in mice, Studies in cell culture indicate that this AMP peptide activates p38 MAP kinase, increases accumulation of specific tight junction proteins (occludin, ZO-1, claudin-5), an adherens junction protein (E-cadherin), and heat shock proteins (hsp25, hsp72), and protects the actin microfilament network in cells exposed to cytochalasin D. When given to mice, AMP peptide increases the content of hyperphosphorylated occludin in the colonic mucosa. To determine if these biological effects of AMP peptide confer colonic cytoprotection in vivo, its effectiveness was tested in two important animal models of barrier compromise, IBD and GDS. Pretreatment of mice with AMP peptide delayed the onset of bloody diarrhea and reduced weight loss in animals given DSS to induce colitis. The peptide also prevented the death of mice given intracecal Pseudomonas aeruginosa following the surgical stress of partial hepatectomy and post-operative food deprivation in a model GDS. Our objective is to obtain additional support in cell culture and in vivo models of IBD and GDS to demonstrate that AMP peptide, by increasing accumulation of tight junction proteins such as occludin, protects colonic mucosal barrier structure and function. Studies that confirm and extend the cytoprotective, mitogenic, and motogenic effects of AMP peptide would lend further support to developing this molecule as a therapeutic agent to treat not only IBD and GDS, but other GI diseases mediated by disruption of intestinal tight junctions and the subsequent increase in mucosal permeability to bacteria and their products.