Identification of Lactobacillus Secreted Factors in Regulation of Pgp

性状（由申请方提供）：由ABCB 1基因编码的P-糖蛋白（P-gp/MDR 1）在肠上皮细胞的顶端表面大量表达。Pgp通过介导外源性物质和细菌毒素从粘膜进入肠腔而参与保护肠上皮。Pgp功能和表达的失调与肠道疾病如炎症性肠病（IBD）和结肠炎的发病机制有关。在这方面，人类MDR 1基因多态性与溃疡性结肠炎和克罗恩病患者肠道P-糖蛋白表达减少有关。此外，MDR 1缺陷小鼠自发地发展类似于人类UC的严重结肠炎。此外，在DSS诱导的结肠炎的实验小鼠模型中已经显示出Pgp的功能和表达的降低。因此，它是至关重要的，以描绘的调节机制，增加P-糖蛋白的功能，以保护肠上皮。解开这些机制可能有助于治疗肠道炎症。在这方面，我们最近表明，益生菌，嗜酸乳杆菌（24小时）培养上清液（CS）增加Pgp功能和表达在人肠上皮细胞Caco 2细胞。同样，在小鼠中的平行研究表明，在回肠和结肠中的Pgp表达响应于活的L.嗜酸菌L.嗜酸乳杆菌还阻断了DSS结肠炎小鼠结肠中Pgp mRNA和蛋白表达的降低以及炎症，进一步表明炎症的抑制可能部分归因于嗜酸乳杆菌上调Pgp。嗜酸乳杆菌L.嗜酸乳杆菌CS通过PI 3 K、Erk 1/2 MAPK途径和Pgp启动子活性的刺激发生，表明在基因转录水平上的调节。然而，L.增加Pgp功能、表达和启动子活性的嗜酸乳杆菌是未知的。因此，我们在目的1a-c中提出的研究将集中于基于其物理化学性质鉴定分泌的生物活性因子，并通过蛋白质组学和脂质组学分析进一步表征。目的1d & e将测试所鉴定的蛋白质或脂质对Pgp活性和表达的功能功效以及参与刺激Pgp基因表达的分子机制。因此，我们的研究旨在鉴定L.嗜酸乳杆菌和确定Pgp功能和表达的分子机制的调节所确定的蛋白质或脂质将大大提高我们对肠道Pgp功能的机制的理解，并可能提供新的和更有效的治疗方式，以减轻肠道炎症的基础。自从使用live 细菌可能是治疗肠道疾病的潜在风险，使用分泌的生物活性因子代替活细菌将导致其治疗用途的创新方法。

DESCRIPTION (provided by applicant): P-glycoprotein (P-gp/MDR1), encoded by the ABCB1 gene is abundantly expressed on the apical surface of intestinal epithelial cells. Pgp is involved in the protection of intestinal epithelia by mediating the efflux of xenobiotics and bacterial toxis from the mucosa into the lumen. Dysregulation of Pgp function and expression have been implicated in the pathogenesis of intestinal disorders, such as inflammatory bowel diseases (IBD) and colitis. In this regard, polymorphisms in the human MDR1 gene have been associated with reduced intestinal P-glycoprotein expression in patients with ulcerative colitis and Crohn's disease. Also, MDR1- deficient mice spontaneously develop severe colitis resembling human UC. Further, a decrease in function and expression of Pgp has been shown in experimental mouse model of DSS-induced colitis. Therefore, it is critical to delineate the regulatory mechanisms that increase P-glycoprotein function in order to protect the intestinal epithelium. Unraveling such mechanisms may be beneficial in the treatment of intestinal inflammation. In this regard, we have recently shown that probiotic, Lactobacillus acidophilus (24 h) culture supernatant (CS) increased Pgp function and expression in human intestinal epithelial Caco2 cells. Also, in parallel studies in mice demonstrated a significant increase in Pgp expression in the ileum and colon in response to live L. acidophilus bacteria. L. acidophilus also blocked the reduced expression of Pgp mRNA and protein expression as well as inflammation in the colon of DSS colitis mice further suggesting that suppression of inflammation could be in part due to the up-regulation of Pgp by L. acidophilus. The increase in Pgp function in Caco2 cells by L. acidophilus CS occurred via PI3K, Erk1/2 MAPK pathways and stimulation of Pgp promoter activity indicating modulation at the level of gene transcription. However, the identity of the secreted bioactive factor (s) present in the culture supernatant of L. acidophilus that increase Pgp function, expression and promoter activity are not known. Therefore, our studies proposed in Aims 1 a-c will focus on identifying the secreted bioactive factor (s) on the basis of their physicochemical properties and further characterization by proteomic and lipidomic analysis. Aims 1d & e will test the functional efficacy of the identified protein (s) or lipid (s) on Pgp actvity and expression and the molecular mechanisms involved in the stimulation of Pgp gene expression. Therefore, our studies aimed at identifying the bioactive factor (s) secreted by L. acidophilus and determining the molecular mechanisms underlying the regulation of Pgp function and expression by the identified protein (s) or lipid (s) will greatly enhance our understanding of the mechanisms of intestinal Pgp function and may provide the basis for new and more efficacious treatment modalities for alleviating intestinal inflammation. Since using live bacteria could be a potential risk in the treatment of gut disorders, using secreted bioactive factor (s) instead of live bacteria would lead to an innovative approach for their therapeutic usage.