Orally administered extract from \u3ci\u3ePrunella vulgaris\u3c/i\u3e attenuates spontaneous colitis in mdr1a\u3csup\u3e-/-\u3c/sup\u3e mice

AIM: To investigate the ability of a Prunella vulgaris (P. vulgaris) ethanolic extract to attenuate spontaneous typhlocolitis in mdr1a-/- mice. METHODS: Vehicle (5% ethanol) or P. vulgaris ethanolic extract (2.4 mg/d) were administered daily by oral gavage to mdr1a-/- or wild type FVBWT mice from 6 wk of age up to 20 wk of age. Clinical signs of disease were noted by monitoring weight loss. Mice experiencing weight loss in excess of 15% were removed from the study. At the time mice were removed from the study, blood and colon tissue were collected for analyses that included histological evaluation of lesions, inflammatory cytokine levels, and myeloperoxidase activity. RESULTS: Administration of P. vulgaris extracts to mdr1a-/- mice delayed onset of colitis and reduced severity of mucosal inflammation when compared to vehicle-treated mdr1a-/- mice. Oral administration of the P. vulgaris extract resulted in reduced (P \u3c 0.05) serum levels of IL-10 (4.6 ± 2 vs 19.4 ± 4), CXCL9 (1319.0 ± 277 vs 3901.0 ± 858), and TNFα (9.9 ± 3 vs 14.8 ± 1) as well as reduced gene expression by more than two-fold for Ccl2, Ccl20, Cxcl1, Cxcl9, IL-1 α, Mmp10, VCAM-1, ICAM, IL-2, and TNFα in the colonic mucosa of mdr1a-/- mice compared to vehicle-treated mdr1a-/- mice. Histologically, several microscopic parameters were reduced (P \u3c 0.05) in P. vulgaris -treated mdr1a-/- mice, as was myeloperoxidase activity in the colon (2.49 ± 0.16 vs 3.36 ± 0.06, P \u3c 0.05). The numbers of CD4+ T cells (2031.9 ± 412.1 vs 5054.5 ± 809.5) and germinal center B cells (2749.6 ± 473.7 vs 4934.0 ± 645.9) observed in the cecal tonsils of P. vulgaris - treated mdr1a-/- were significantly reduced (P \u3c 0.05) from vehicle-treated mdr1a-/- mice. Vehicle-treated mdr1a-/- mice were found to produce serum antibodies to antigens derived from members of the intestinal microbiota, indicative of severe colitis and a loss of adaptive tolerance to the members of the microbiota. These serum antibodies were greatly reduced or absent in P. vulgaris -treated mdr1a-/- mice. CONCLUSION: The anti-inflammatory activity of P. vulgaris ethanolic extract effectively attenuated the severity of intestinal inflammation in mdr1a-/- mice

Age-related impact of proteobacteria colonization on mucosal homeostasis and the microbial community in gastrointestinal health and disease

Inflammatory bowel disease is a group of chronic intestinal inflammatory disorders with a complex etiology, and has been associated with a microbial dysbiosis and presence of GI pathogens including adherent and invasive Escherichia coli (AIEC) and Campylobacter. A gnotobiotic murine model (altered Schaedler flora, ASF) devoid of proteobacteria was utilized to evaluate microbial and host responses after colonization with AIEC LF82, E. coli Nissle 1917, and C. jejuni alone or treated with dextran sodium sulfate (DSS). The ultimate goal of this research was to understand the effects on mucosal homeostasis after disturbance of the resident microbiota by proteobacterial pathobionts. As few studies have evaluated the stability of the microbiota over multiple generations, Chapter 3 evaluates the impact of AIEC LF82 colonization on the heritability/transmissibility of the ASF over multiple generations. Results indicated minimal changes in ASF abundance within a generation, but there was a significant shift in the microbial population when comparing early and later generations. As early-life microbial exposures have been demonstrated to influence mucosal homeostasis later in life, Chapter 4 describes changes in the sensitivity to colitis following E. coli LF82 colonization of ASF mice as an adult or neonate. We demonstrate neonatal colonization with LF82 increases the susceptibility to colitis, with increased inflammatory responses and minimal microbial changes, with neonatal colonization inducing subclinical, chronic inflammation that may predispose to more severe inflammation. In Chapter 5 of this dissertation, the benefits associated with probiotic Escherichia coli Nissle 1917 (EcN) colonization of neonatal mice to prevent or attenuate colitis as described in Chapter 4 were evaluated. Results indicated that co-colonization of neonatal mice with EcN did not impact the severity of colitis, but did reduce the host’s proinflammatory cytokine response. Finally, in Chapter 6, both conventional and gnotobiotic mice were used to demonstrate that the pathogenesis of C. jejuni is differentially influenced by the complexity of the microbiota. By investigating host-microbial relationships and their effects on intestinal homeostasis, this work provided insights into the underlying mechanisms contributing to the onset of inflammatory bowel disease, as well as what constitutes a ‘healthy’ state in the relationship with our gut microbiota.</p

Orally administered extract from \u3ci\u3ePrunella vulgaris\u3c/i\u3e attenuates spontaneous colitis in mdr1a\u3csup\u3e-/-\u3c/sup\u3e mice

AIM: To investigate the ability of a Prunella vulgaris (P. vulgaris) ethanolic extract to attenuate spontaneous typhlocolitis in mdr1a-/- mice. METHODS: Vehicle (5% ethanol) or P. vulgaris ethanolic extract (2.4 mg/d) were administered daily by oral gavage to mdr1a-/- or wild type FVBWT mice from 6 wk of age up to 20 wk of age. Clinical signs of disease were noted by monitoring weight loss. Mice experiencing weight loss in excess of 15% were removed from the study. At the time mice were removed from the study, blood and colon tissue were collected for analyses that included histological evaluation of lesions, inflammatory cytokine levels, and myeloperoxidase activity. RESULTS: Administration of P. vulgaris extracts to mdr1a-/- mice delayed onset of colitis and reduced severity of mucosal inflammation when compared to vehicle-treated mdr1a-/- mice. Oral administration of the P. vulgaris extract resulted in reduced (P \u3c 0.05) serum levels of IL-10 (4.6 ± 2 vs 19.4 ± 4), CXCL9 (1319.0 ± 277 vs 3901.0 ± 858), and TNFα (9.9 ± 3 vs 14.8 ± 1) as well as reduced gene expression by more than two-fold for Ccl2, Ccl20, Cxcl1, Cxcl9, IL-1 α, Mmp10, VCAM-1, ICAM, IL-2, and TNFα in the colonic mucosa of mdr1a-/- mice compared to vehicle-treated mdr1a-/- mice. Histologically, several microscopic parameters were reduced (P \u3c 0.05) in P. vulgaris -treated mdr1a-/- mice, as was myeloperoxidase activity in the colon (2.49 ± 0.16 vs 3.36 ± 0.06, P \u3c 0.05). The numbers of CD4+ T cells (2031.9 ± 412.1 vs 5054.5 ± 809.5) and germinal center B cells (2749.6 ± 473.7 vs 4934.0 ± 645.9) observed in the cecal tonsils of P. vulgaris - treated mdr1a-/- were significantly reduced (P \u3c 0.05) from vehicle-treated mdr1a-/- mice. Vehicle-treated mdr1a-/- mice were found to produce serum antibodies to antigens derived from members of the intestinal microbiota, indicative of severe colitis and a loss of adaptive tolerance to the members of the microbiota. These serum antibodies were greatly reduced or absent in P. vulgaris -treated mdr1a-/- mice. CONCLUSION: The anti-inflammatory activity of P. vulgaris ethanolic extract effectively attenuated the severity of intestinal inflammation in mdr1a-/- mice

Pathogenic and non-pathogenic Escherichia coli colonization and host inflammatory response in a defined microbiota mouse model

Most Escherichia coli strains in the human intestine are harmless. However, enterohemorrhagic E. coli (EHEC) is a foodborne pathogen that causes intestinal disease in humans. Conventionally reared (CONV) mice are inconsistent models for human infections with EHEC because they are often resistant to E. coli colonization, in part due to their gastrointestinal (GI) microbiota. Although antibiotic manipulation of the mouse microbiota has been a common means to overcome colonization resistance, these models have limitations. Currently, there are no licensed treatments for clinical EHEC infections and, thus, new tools to study EHEC colonization need to be developed. Here, we used a defined microbiota mouse model, consisting of the altered Schaedler flora (ASF), to characterize intestinal colonization and compare host responses following colonization with EHEC strain 278F2 or non-pathogenic E. coli strain MG1655. Significantly higher (P<0.05) levels of both strains were found in feces and cecal and colonic contents of C3H/HeN ASF compared to C3H/HeN CONV mice. GI inflammation was significantly elevated (P<0.05) in the cecum of EHEC 278F2-colonized compared to E. coli MG1655-colonized C3H/HeN ASF mice. In addition, EHEC 278F2 differentially modulated inflammatory-associated genes in colonic tissue of C3H/HeN ASF mice compared to E. coli MG1655-colonized mice. This approach allowed for prolonged colonization of the murine GI tract by pathogenic and non-pathogenic E. coli strains, and for evaluation of host inflammatory processes. Overall, this system can be used as a powerful tool for future studies to assess therapeutics, microbe-microbe interactions, and strategies for preventing EHEC infections