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, 12 (10), e0185999
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Nanoparticle Curcumin Ameliorates Experimental Colitis via Modulation of Gut Microbiota and Induction of Regulatory T Cells

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Nanoparticle Curcumin Ameliorates Experimental Colitis via Modulation of Gut Microbiota and Induction of Regulatory T Cells

Masashi Ohno et al. PLoS One.

Abstract

Background and aims: Curcumin is a hydrophobic polyphenol derived from turmeric, a traditional Indian spice. Curcumin exhibits various biological functions, but its clinical application is limited due to its poor absorbability after oral administration. A newly developed nanoparticle curcumin shows improved absorbability in vivo. In this study, we examined the effects of nanoparticle curcumin (named Theracurmin) on experimental colitis in mice.

Methods: BALB/c mice were fed with 3% dextran sulfate sodium (DSS) in water. Mucosal cytokine expression and lymphocyte subpopulation were analyzed by real-time PCR and flow cytometry, respectively. The profile of the gut microbiota was analyzed by real-time PCR.

Results: Treatment with nanoparticle curcumin significantly attenuated body weight loss, disease activity index, histological colitis score and significantly improved mucosal permeability. Immunoblot analysis showed that NF-κB activation in colonic epithelial cells was significantly suppressed by treatment with nanoparticle curcumin. Mucosal mRNA expression of inflammatory mediators was significantly suppressed by treatment with nanoparticle curcumin. Treatment with nanoparticle curcumin increased the abundance of butyrate-producing bacteria and fecal butyrate level. This was accompanied by increased expansion of CD4+ Foxp3+ regulatory T cells and CD103+ CD8α- regulatory dendritic cells in the colonic mucosa.

Conclusions: Treatment with nanoparticle curcumin suppressed the development of DSS-induced colitis potentially via modulation of gut microbial structure. These responses were associated with induction of mucosal immune cells with regulatory properties. Nanoparticle curcumin is one of the promising candidates as a therapeutic option for the treatment of IBD.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Effect of nanoparticle curcumin on the development of DSS colitis.
BALB/cAJcl mice were treated with nanoparticle curcumin (Theracurmin) for 7 days prior to the start of 3% DSS treatment. The mice were sacrificed on day 18. (A) Body weight. (B) Disease activity index. (C) Representative photographs of the colon. (D) Colonic weight/length on day 18. The data are expressed as means ± SEM (n = 6 mice/group). The data are representative of four independent experiments. Values not sharing a letter are significantly different (P<0.05).
Fig 2
Fig 2. Histological evaluation of colitis.
(A) Histological picture of the colonic tissue on day 18. (original magnification ×200.) (B) Histological sore. The data are expressed as means ± SEM (n = 6 mice/group). (C) Epithelial permeability. Mice were orally administrated with FITC-labeled dextran (44 mg/100 g body weight), (MW 4000; FD4, Sigma-Aldrich Co.). Serum was collected 5 h later and fluorescence intensity was determined. Values not sharing a letter are significantly different (P<0.05).
Fig 3
Fig 3. The effect of nanoparticle curcumin on NF-κB activation.
(A) Immunoblot for NF-κBp65 in the nuclear protein of colonic epithelium. Lamin A/C was used as a loading control. The picture is representative of four independent experiments. (B) Immunohistochemical staining for NF-κBp65 in the tissues. (original magnification ×200). NF-κBp65 was detected in the nucleus of the epithelial cells in the DSS group, but this was completely blocked in the DSS plus nanoparticle curcumin group. (C) Immunostaining of NF-κBp65 in HT-29 cells. HT-29 cells were stimulated with TNF-α (100ng/ml) in the presence or absence of nanoparticle curcumin (10μM) for 15 minutes. NF-κB p65, green fluorescence; nucleus, DAPI (blue). (D) The effect of nanoparticle curcumin on IκBα phosphorylation in response to TNF-α. HT-29 cells were stimulated with TNF-α (100 ng/ml) in the presence or absence of nanoparticle curcumin (0μM, 10μM, or 50μM) for 15 minutes, and then lysed with lysis buffer. Lysates were subjected to immunoblot analysis. GAPDH were used as loading control. The data represent four independent experiments.
Fig 4
Fig 4. The effect of nanoparticle curcumin on the expression of proinflammatory mediators and neutrophil infiltration.
(A) Real-time PCR analysis for the mucosal mRNA expression of TNF-α, IL-1β, IL-6, CXCL1 and CXCL2. The cytokine mRNA expression was converted to a value relative to β-actin mRNA expression, and presented as an increase relative to the control mice. The data are expressed as means ± SEM (n = 6 mice/group). Values not sharing a letter are significantly different (P<0.05). (B) Proportion of Gr-1+ neutrophils in the lamina propria of the colon. Mucosa Gr-1+ neutrophils were analyzed by flow cytometry. (C) The number of Gr-1+ neutrophils. The data are expressed as means ± SEM (n = 6 mice/group). Values not sharing a letter are significantly different (P<0.05).
Fig 5
Fig 5. The effect of nanoparticle curcumin on the gut microbial structure.
(A) T-RFLP analysis of the gut microbiota. The value indicates the percentage of the predicted bacteria. (B) Real-time PCR analysis for Clostridium cluster IV. (C) Real-time PCR analysis for Clostridium subcluster XIVa. The values were normalized to the amount of total bacteria, and presented as relative amount to the control group. The data were expressed as means ± SEM (n = 4 mice/group). Values not sharing a letter are significantly different (P<0.05).
Fig 6
Fig 6. Effect of nanoparticle curcumin on the fecal short-chain fatty acid (SCFA) levels.
The concentrations of fecal SCFAs were measured by high-performance liquid chromatography. The data were expressed as means ± SEM (n = 6 mice/group). Values not sharing a letter are significantly different (P<0.05).
Fig 7
Fig 7. The effects of nanoparticle curcumin on the induction of Tregs and regulatory DCs in the lamina propria of the colon.
(A) Flow cytometry analysis for CD4+ Foxp3+ Tregs in the lamina propria of the colon. Representative picture from two independent experiments. (B) Proportion of CD4+ Foxp3+ Treg cells in CD4+ cells in the lamina propria. The data are expressed as means ± SEM (n = 6 mice/group). Values not sharing a letter are significantly different (P<0.05). (C) Flow cytometry analysis for CD103+ CD8α DCs in the lamina propria of the colon. Representative picture from two independent experiments. (D) Proportion of CD103+ CD8α DCs in CD11c+ cells in the lamina propria. The data are expressed as means ± SEM (n = 6 mice/group). Values not sharing a letter are significantly different (P<0.05).

Cited by 12 PubMed Central articles

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Grant support

This work was supported by a Grant-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan; https://www.jsps.go.jp/english/e-grants/index.html, 15K08967 to A.A and 15K19322 to A.N., the Intractable Diseases from the Ministry of Health, Labor and Welfare of Japan http://www.nanbyou.or.jp/english/index.htm, 067 to A.A., the Practical Research Project for Rare/Intractable Diseases from the Japan Agency for Medical Research and Development, AMED http://www.amed.go.jp/en/program/list/01/05/016.html, 15AeK0109047h0002 to A.A., and the Smoking Research Foundation; http://www.srf.or.jp/english/index.html, 1848 to A.A.
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