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 1 d &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.
Disturbances in the intestinal luminal wall health leads to the development of intestinal inflammation associated with various intestinal diseases such as inflammatory bowel diseases (IBD) and colitis. P-glycoprotein (Pgp) plays a critical role in the protection of the intestinal barrier by mediating the removal of drugs/xenobiotics and bacterial toxins from the intestinal wall into the gut cavity. A decrease in function and expression of Pgp has been shown in patients with IBD and in experimental mouse model of colitis. We have recently shown that the probiotic, L. acidophilus culture supernatant increases Pgp function and expression in intestinal epithelial cells, that could contribute to the beneficial effects of probitics in inflammation. The studies proposed in this application will identify the novel secreted bioactive factor (s) that modulate Pgp and could be used as potential therapeutic agents in the prevention and treatment of IBD.
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