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Increased production of BDNF in colonic epithelial cells induced by fecal supernatants from diarrheic IBS patients.

Wang P, Chen FX, Du C, Li CQ, Yu YB, Zuo XL, Li YQ - Sci Rep (2015)

Bottom Line: Using human Caco-2 cells, we found that IBS-D FSN significantly increased BDNF mRNA and protein levels compared to control FSN, which were remarkably suppressed by the serine protease inhibitor.We found a significant increase in PAR-2 expression in Caco-2 after IBS-D FSN stimulation.Moreover, we found that phosphorylation of p38 MAPK, not NF-κB p65, contributed to PAR-2-mediated BDNF overexpression.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Gastroenterology, Qilu Hospital, Shandong University, Jinan 250012, P.R. China [2] Laboratory of Translational Gastroenterology, Qilu Hospital, Shandong University, Jinan 250012, P.R. China.

ABSTRACT
Colonic brain-derived neurotrophic factor (BDNF) plays an essential role in pathogenesis of abdominal pain in diarrhea-predominant irritable bowel syndrome (IBS-D), but regulation on its expression remains unclear. We investigated the role of fecal supernatants (FSN) from IBS-D patients on regulating BDNF expression in colonic epithelial cells of human and mice. Using human Caco-2 cells, we found that IBS-D FSN significantly increased BDNF mRNA and protein levels compared to control FSN, which were remarkably suppressed by the serine protease inhibitor. To further explore the potential mechanisms, we investigated the impact of protease-activated receptor-2 (PAR-2) on BDNF expression. We found a significant increase in PAR-2 expression in Caco-2 after IBS-D FSN stimulation. Knockdown of PAR-2 significantly inhibited IBS-D FSN-induced upregulation of BDNF. Moreover, we found that phosphorylation of p38 MAPK, not NF-κB p65, contributed to PAR-2-mediated BDNF overexpression. To confirm these results, we intracolonically infused IBS-D or control FSN in mice and found that IBS-D FSN significantly elevated colonic BDNF and visceral hypersensitivity in mice, which were both suppressed by the inhibitor of serine protease or antagonist of PAR-2. Together, our data indicate that activation of PAR-2 signaling by IBS-D FSN promotes expression of colonic BDNF, thereby contributing to IBS-like visceral hypersensitivity.

No MeSH data available.


Related in: MedlinePlus

IBS-D fecal supernatants (FSN) induced colonic BDNF overexpression and visceral hypersensitivity in mice.(a and b) Western blotting of BDNF expression in colon of mice (***P < 0.001). (c) Immunohistochemical staining of BDNF in epithelium (Black arrowheads) and mucosa in colon of mice (Bars: 50 μm). (d) Measurement of visceral sensitivity to colorectal distension in mice by using electromyography recording (*P < 0.05, ***P < 0.001 IBS-D FSN vs. control FSN; †††P < 0.001, IBS-D FSN + FUT-175 or ENMD-1068 or TrkB/Fc vs. IBS-D FSN; n = 8 per group; AUC: area under curve). The gels were run under the same experimental conditions. Cropped gels/blots are presented (full-length gels/blots are shown in suppl. Fig. S1 with indicated cropping lines).
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f5: IBS-D fecal supernatants (FSN) induced colonic BDNF overexpression and visceral hypersensitivity in mice.(a and b) Western blotting of BDNF expression in colon of mice (***P < 0.001). (c) Immunohistochemical staining of BDNF in epithelium (Black arrowheads) and mucosa in colon of mice (Bars: 50 μm). (d) Measurement of visceral sensitivity to colorectal distension in mice by using electromyography recording (*P < 0.05, ***P < 0.001 IBS-D FSN vs. control FSN; †††P < 0.001, IBS-D FSN + FUT-175 or ENMD-1068 or TrkB/Fc vs. IBS-D FSN; n = 8 per group; AUC: area under curve). The gels were run under the same experimental conditions. Cropped gels/blots are presented (full-length gels/blots are shown in suppl. Fig. S1 with indicated cropping lines).

Mentions: Next we aimed to further confirm the effect of IBS-D FSN on colonic BDNF expression in vivo. We stimulated colonic epithelium of mice by intracolonic administration of selected IBS-D FSN (a serine-protease activity greater than 1093 U/mg protein (the largest activity in HC group)), control FSN or saline for 1 h. Control FSN had no specific effect on BDNF expression as compared with saline (Fig. 5a,b). BDNF levels in colon of mice nearly doubled after stimulation by IBS-D FSN compared to control FSN. IBS-D FSN failed to upregulate BDNF expression in colon of mice when preincubated with FUT-175. In mice pretreated with ENMD-1068, IBS-D FSN could not induce elevation of colonic BDNF either. Immunochemistry staining confirmed the western blotting results and showed that the increased BDNF was mainly observed in colonic epithelium and mucosa (Fig. 5c).


Increased production of BDNF in colonic epithelial cells induced by fecal supernatants from diarrheic IBS patients.

Wang P, Chen FX, Du C, Li CQ, Yu YB, Zuo XL, Li YQ - Sci Rep (2015)

IBS-D fecal supernatants (FSN) induced colonic BDNF overexpression and visceral hypersensitivity in mice.(a and b) Western blotting of BDNF expression in colon of mice (***P < 0.001). (c) Immunohistochemical staining of BDNF in epithelium (Black arrowheads) and mucosa in colon of mice (Bars: 50 μm). (d) Measurement of visceral sensitivity to colorectal distension in mice by using electromyography recording (*P < 0.05, ***P < 0.001 IBS-D FSN vs. control FSN; †††P < 0.001, IBS-D FSN + FUT-175 or ENMD-1068 or TrkB/Fc vs. IBS-D FSN; n = 8 per group; AUC: area under curve). The gels were run under the same experimental conditions. Cropped gels/blots are presented (full-length gels/blots are shown in suppl. Fig. S1 with indicated cropping lines).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4441152&req=5

f5: IBS-D fecal supernatants (FSN) induced colonic BDNF overexpression and visceral hypersensitivity in mice.(a and b) Western blotting of BDNF expression in colon of mice (***P < 0.001). (c) Immunohistochemical staining of BDNF in epithelium (Black arrowheads) and mucosa in colon of mice (Bars: 50 μm). (d) Measurement of visceral sensitivity to colorectal distension in mice by using electromyography recording (*P < 0.05, ***P < 0.001 IBS-D FSN vs. control FSN; †††P < 0.001, IBS-D FSN + FUT-175 or ENMD-1068 or TrkB/Fc vs. IBS-D FSN; n = 8 per group; AUC: area under curve). The gels were run under the same experimental conditions. Cropped gels/blots are presented (full-length gels/blots are shown in suppl. Fig. S1 with indicated cropping lines).
Mentions: Next we aimed to further confirm the effect of IBS-D FSN on colonic BDNF expression in vivo. We stimulated colonic epithelium of mice by intracolonic administration of selected IBS-D FSN (a serine-protease activity greater than 1093 U/mg protein (the largest activity in HC group)), control FSN or saline for 1 h. Control FSN had no specific effect on BDNF expression as compared with saline (Fig. 5a,b). BDNF levels in colon of mice nearly doubled after stimulation by IBS-D FSN compared to control FSN. IBS-D FSN failed to upregulate BDNF expression in colon of mice when preincubated with FUT-175. In mice pretreated with ENMD-1068, IBS-D FSN could not induce elevation of colonic BDNF either. Immunochemistry staining confirmed the western blotting results and showed that the increased BDNF was mainly observed in colonic epithelium and mucosa (Fig. 5c).

Bottom Line: Using human Caco-2 cells, we found that IBS-D FSN significantly increased BDNF mRNA and protein levels compared to control FSN, which were remarkably suppressed by the serine protease inhibitor.We found a significant increase in PAR-2 expression in Caco-2 after IBS-D FSN stimulation.Moreover, we found that phosphorylation of p38 MAPK, not NF-κB p65, contributed to PAR-2-mediated BDNF overexpression.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Gastroenterology, Qilu Hospital, Shandong University, Jinan 250012, P.R. China [2] Laboratory of Translational Gastroenterology, Qilu Hospital, Shandong University, Jinan 250012, P.R. China.

ABSTRACT
Colonic brain-derived neurotrophic factor (BDNF) plays an essential role in pathogenesis of abdominal pain in diarrhea-predominant irritable bowel syndrome (IBS-D), but regulation on its expression remains unclear. We investigated the role of fecal supernatants (FSN) from IBS-D patients on regulating BDNF expression in colonic epithelial cells of human and mice. Using human Caco-2 cells, we found that IBS-D FSN significantly increased BDNF mRNA and protein levels compared to control FSN, which were remarkably suppressed by the serine protease inhibitor. To further explore the potential mechanisms, we investigated the impact of protease-activated receptor-2 (PAR-2) on BDNF expression. We found a significant increase in PAR-2 expression in Caco-2 after IBS-D FSN stimulation. Knockdown of PAR-2 significantly inhibited IBS-D FSN-induced upregulation of BDNF. Moreover, we found that phosphorylation of p38 MAPK, not NF-κB p65, contributed to PAR-2-mediated BDNF overexpression. To confirm these results, we intracolonically infused IBS-D or control FSN in mice and found that IBS-D FSN significantly elevated colonic BDNF and visceral hypersensitivity in mice, which were both suppressed by the inhibitor of serine protease or antagonist of PAR-2. Together, our data indicate that activation of PAR-2 signaling by IBS-D FSN promotes expression of colonic BDNF, thereby contributing to IBS-like visceral hypersensitivity.

No MeSH data available.


Related in: MedlinePlus