The effect of Denosumab, the inhibitor for receptor activator of nuclear factor kappa-B ligand (RANKL), on dinitrobenzensulfonic acid (DNBS)-induced experimental model of crohn's disease.

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Khafipour, Azin
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Background: The pro-inflammatory mediator receptor activator of nuclear factor-kappa B ligand (RANKL) plays a major role in the development of rheumatoid arthritis; however, its role in inflammatory bowel disease is unknown. Genome-wide association meta-analysis for Crohn’s disease (CD) identified a variant, near the TNFSF11 gene that encodes RANKL. Moreover, CD increased expression of RANKL in specific cell lines. This study aims to elucidate if the RANKL inhibitor Denosumab (Prolia™) can reduce the severity of experimental colitis via modifying gut microbiota dysregulation. Methods: CD-like colitis was induced via intrarectal administration of dinitrobenzenesulfonic acid (DNBS, 4mg/kg) dissolved in Ethanol (30%) to C57Bl/6 mice (n=12). One day before colitis induction, daily injection of Prolia (10mg/kg/d, intraperitonealy) was initiated and continued over four days. Vehicle mice received PBS1%. On the sacrifice day, inflammatory status was evaluated clinically. DNA was extracted from colonic mucosa and fecal samples, and V4 region of bacterial 16S rRNA gene was amplified and subjected to Illumina sequencing for microbiome analysis. Alpha- and beta-diversities were calculated in QIIME and subjected to SAS and PERMANOVA, respectively. Differences between were considered significant at P<0.05. Results: Disease severity, macroscopic score and pro-inflammatory cytokines (Il-6, IL-1β and TNF-α) were increased in DNBS/Ethanol-treated vs. vehicle mice. Prolia treatment decreased (P<0.05) only the pro-inflammatory cytokines. Prolia treatment also modified the alpha- and beta-diversity of colonic mucosa and fecal microbiota. DNBS/Ethanol group clustered separately (P<0.05) compared to the vehicle group, Prolia treatment attenuated the negative effects of DNBS/Ethanol. Conclusions: The development of colitis in DNBS/Ethanol model was accompanied by disruption of gut microbiota. Preventative treatment with Prolia modulated intestinal inflammation and gut microbiota dysbiosis in a murine model of colitis. Our results provide a rationale for considering Prolia as a future potential therapy in CD, but more experimental and clinical studies need to be performed.
Inflammatory Bowel Disease (IBD), Denosumab (Prolia), gut microbiota, inflammation, Dinitrobenzensulfonic Acid (DNBS), experimental model of colitis, Crohn's disease, Qiime, Next generation sequencing, Clustering analysis