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Hyaluronic acid–bilirubin nanomedicine for targeted modulation of dysregulated intestinal barrier, microbiome and immune responses in colitis

Abstract

While conventional approaches for inflammatory bowel diseases mainly focus on suppressing hyperactive immune responses, it remains unclear how to address disrupted intestinal barriers, dysbiosis of the gut commensal microbiota and dysregulated mucosal immune responses in inflammatory bowel diseases. Moreover, immunosuppressive agents can cause off-target systemic side effects and complications. Here, we report the development of hyaluronic acid–bilirubin nanomedicine (HABN) that accumulates in inflamed colonic epithelium and restores the epithelium barriers in a murine model of acute colitis. Surprisingly, HABN also modulates the gut microbiota, increasing the overall richness and diversity and markedly augmenting the abundance of Akkermansia muciniphila and Clostridium XIVα, which are microorganisms with crucial roles in gut homeostasis. Importantly, HABN associated with pro-inflammatory macrophages, regulated innate immune responses and exerted potent therapeutic efficacy against colitis. Our work sheds light on the impact of nanotherapeutics on gut homeostasis, microbiome and innate immune responses for the treatment of inflammatory diseases.

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Fig. 1: HABN localizes in inflamed colon in DSS-treated mice.
Fig. 2: HABN exerts strong efficacy in a murine model of colitis.
Fig. 3: HABN protects colonic epithelium.
Fig. 4: HABN alters the composition of gut microbiome.
Fig. 5: HABN alleviates colitis in a delayed therapeutic setting.

Data availability

The data supporting the findings of this study are available within the article and its Supplementary Information files. All relevant data can be provided by the authors upon reasonable request.

References

  1. 1.

    Turner, J. R. Intestinal mucosal barrier function in health and disease. Nat. Rev. Immunol. 9, 799–809 (2009).

    CAS  Google Scholar 

  2. 2.

    Citi, S. Intestinal barriers protect against disease. Science 359, 1097–1098 (2018).

    CAS  Google Scholar 

  3. 3.

    Round, J. L. & Mazmanian, S. K. The gut microbiota shapes intestinal immune responses during health and disease. Nat. Rev. Immunol. 9, 313–323 (2009).

    CAS  Google Scholar 

  4. 4.

    Kostic, A. D., Xavier, R. J. & Gevers, D. The microbiome in inflammatory bowel disease: current status and the future ahead. Gastroenterology 146, 1489–1499 (2014).

    CAS  Google Scholar 

  5. 5.

    Halfvarson, J. et al. Dynamics of the human gut microbiome in inflammatory bowel disease. Nat. Microbiol. 2, 17004 (2017).

    CAS  Google Scholar 

  6. 6.

    Bernstein, C. N. et al. World gastroenterology organization practice guidelines for the diagnosis and management of IBD in 2010. Inflamm. Bowel. Dis. 16, 112–124 (2010).

    Google Scholar 

  7. 7.

    Lautenschlager, C., Schmidt, C., Fischer, D. & Stallmach, A. Drug delivery strategies in the therapy of inflammatory bowel disease. Adv. Drug Deliv. Rev. 71, 58–76 (2014).

    Google Scholar 

  8. 8.

    Wilson, D. S. et al. Orally delivered thioketal nanoparticles loaded with TNF-alpha-siRNA target inflammation and inhibit gene expression in the intestines. Nat. Mater. 9, 923–928 (2010).

    CAS  Google Scholar 

  9. 9.

    Zhang, S. et al. An inflammation-targeting hydrogel for local drug delivery in inflammatory bowel disease. Sci. Transl. Med. 7, 300ra128 (2015).

    Google Scholar 

  10. 10.

    Stallmach, A., Hagel, S. & Bruns, T. Adverse effects of biologics used for treating IBD. Best Pract. Res. Clin. Gastroenterol. 24, 167–182 (2010).

    CAS  Google Scholar 

  11. 11.

    Rayahin, J. E., Buhrman, J. S., Zhang, Y., Koh, T. J. & Gemeinhart, R. A. High and low molecular weight hyaluronic acid differentially influence macrophage activation. ACS Biomater. Sci. Eng. 1, 481–493 (2015).

    CAS  Google Scholar 

  12. 12.

    Hill, D. R., Kessler, S. P., Rho, H. K., Cowman, M. K. & de la Motte, C. A. Specific-sized hyaluronan fragments promote expression of human beta-defensin 2 in intestinal epithelium. J. Biol. Chem. 287, 30610–30624 (2012).

    CAS  Google Scholar 

  13. 13.

    Bollyky, P. L. et al. Intact extracellular matrix and the maintenance of immune tolerance: high molecular weight hyaluronan promotes persistence of induced CD4+CD25+ regulatory T cells. J. Leukoc. Biol. 86, 567–572 (2009).

    CAS  Google Scholar 

  14. 14.

    Zheng, L., Riehl, T. E. & Stenson, W. F. Regulation of colonic epithelial repair in mice by Toll-like receptors and hyaluronic acid. Gastroenterology 137, 2041–2051 (2009).

    CAS  Google Scholar 

  15. 15.

    Xiao, B. et al. Combination therapy for ulcerative colitis: orally targeted nanoparticles prevent mucosal damage and relieve inflammation. Theranostics 6, 2250–2266 (2016).

    CAS  Google Scholar 

  16. 16.

    Petrey, A. C. & de la Motte, C. A. Hyaluronan, a crucial regulator of inflammation. Front. Immunol. 5, 101 (2014).

    Google Scholar 

  17. 17.

    Kapitulnik, J. Bilirubin: an endogenous product of heme degradation with both cytotoxic and cytoprotective properties. Mol. Pharmacol. 66, 773–779 (2004).

    CAS  Google Scholar 

  18. 18.

    Sedlak, T. W. et al. Bilirubin and glutathione have complementary antioxidant and cytoprotective roles. Proc. Natl Acad. Sci. USA 106, 5171–5176 (2009).

    CAS  Google Scholar 

  19. 19.

    Chassaing, B., Aitken, J. D., Malleshappa, M. & Vijay-Kumar, M. Dextran sulfate sodium (DSS)-induced colitis in mice. Curr. Protoc. Immunol. 104, 15.25.1–15.25.14 (2014).

    Google Scholar 

  20. 20.

    Hansberry, D. R., Shah, K., Agarwal, P. & Agarwal, N. Fecal Myeloperoxidase as a Biomarker for Inflammatory Bowel Disease. Cureus 9, e1004 (2017).

    Google Scholar 

  21. 21.

    Hall, E. D., McCall, J. M., Chase, R. L., Yonkers, P. A. & Braughler, J. M. A nonglucocorticoid steroid analog of methylprednisolone duplicates its high-dose pharmacology in models of central nervous system trauma and neuronal membrane damage. J. Pharmacol. Exp. Ther. 242, 137–142 (1987).

    CAS  Google Scholar 

  22. 22.

    Li, B., Alli, R., Vogel, P. & Geiger, T. L. IL-10 modulates DSS-induced colitis through a macrophage-ROS-NO axis. Mucosal Immunol. 7, 869–878 (2014).

    CAS  Google Scholar 

  23. 23.

    Gibson, G. R. et al. Expert consensus document: the international scientific association for probiotics and prebiotics (ISAPP) consensus statement on the definition and scope of prebiotics. Nat. Rev. Gastroenterol. Hepatol. 14, 491–502 (2017).

    Google Scholar 

  24. 24.

    Ma, C. et al. Gut microbiome-mediated bile acid metabolism regulates liver cancer via NKT cells. Science 360, eaan5931 (2018).

    Google Scholar 

  25. 25.

    Cani, P. D. & de Vos, W. M. Next-generation beneficial microbes: the case of Akkermansia muciniphila. Front. Microbiol. 8, 1765 (2017).

    Google Scholar 

  26. 26.

    Everard, A. et al. Cross-talk between Akkermansia muciniphila and intestinal epithelium controls diet-induced obesity. Proc. Natl Acad. Sci. USA 110, 9066–9071 (2013).

    CAS  Google Scholar 

  27. 27.

    Zhang, Z. et al. Chlorogenic acid ameliorates experimental colitis by promoting growth of Akkermansia in mice. Nutrients 9, 677 (2017).

    Google Scholar 

  28. 28.

    Furusawa, Y. et al. Commensal microbe-derived butyrate induces the differentiation of colonic regulatory T cells. Nature 504, 446–450 (2013).

    CAS  Google Scholar 

  29. 29.

    Madsen, K. L., Doyle, J. S., Jewell, L. D., Tavernini, M. M. & Fedorak, R. N. Lactobacillus species prevents colitis in interleukin 10 gene-deficient mice. Gastroenterology 116, 1107–1114 (1999).

    CAS  Google Scholar 

  30. 30.

    Galdeano, C. M. & Perdigon, G. The probiotic bacterium Lactobacillus casei induces activation of the gut mucosal immune system through innate immunity. Clin. Vaccine Immunol. 13, 219–226 (2006).

    CAS  Google Scholar 

  31. 31.

    Geier, M. S., Butler, R. N., Giffard, P. M. & Howarth, G. S. Lactobacillus fermentum BR11, a potential new probiotic, alleviates symptoms of colitis induced by dextran sulfate sodium (DSS) in rats. Int. J. Food Microbiol. 114, 267–274 (2007).

    Google Scholar 

  32. 32.

    Sartor, R. B. Therapeutic manipulation of the enteric microflora in inflammatory bowel diseases: antibiotics, probiotics, and prebiotics. Gastroenterology 126, 1620–1633 (2004).

    Google Scholar 

  33. 33.

    Guandalini, S. Use of Lactobacillus-GG in paediatric Crohn’s disease. Dig. Liver Dis. 34, S63–S65 (2002). Suppl 2.

    Google Scholar 

  34. 34.

    Zocco, M. A. et al. Efficacy of Lactobacillus GG in maintaining remission of ulcerative colitis. Aliment Pharmacol. Ther. 23, 1567–1574 (2006).

    CAS  Google Scholar 

  35. 35.

    Oliva, S. et al. Randomised clinical trial: the effectiveness of Lactobacillus reuteri ATCC 55730 rectal enema in children with active distal ulcerative colitis. Aliment Pharmacol. Ther. 35, 327–334 (2012).

    CAS  Google Scholar 

  36. 36.

    Png, C. W. et al. Mucolytic bacteria with increased prevalence in IBD mucosa augment in vitro utilization of mucin by other bacteria. Am. J. Gastroenterol. 105, 2420–2428 (2010).

    CAS  Google Scholar 

  37. 37.

    Fabia, R. et al. Impairment of bacterial flora in human ulcerative colitis and experimental colitis in the rat. Digestion 54, 248–255 (1993).

    CAS  Google Scholar 

  38. 38.

    Rakoff-Nahoum, S., Paglino, J., Eslami-Varzaneh, F., Edberg, S. & Medzhitov, R. Recognition of commensal microflora by toll-like receptors is required for intestinal homeostasis. Cell 118, 229–241 (2004).

    CAS  Google Scholar 

  39. 39.

    Lee, Y. et al. Bilirubin nanoparticles as a nanomedicine for anti-inflammation therapy. Angew Chem. Int. Ed. Engl. 55, 7460–7463 (2016).

    CAS  Google Scholar 

  40. 40.

    Kim, D. E. et al. Bilirubin nanoparticles ameliorate allergic lung inflammation in a mouse model of asthma. Biomaterials 140, 37–44 (2017).

    CAS  Google Scholar 

  41. 41.

    Lee, S., Lee, Y., Kim, H., Lee, D. Y. & Jon, S. Bilirubin nanoparticle-assisted delivery of a small molecule-drug conjugate for targeted cancer therapy. Biomacromolecules 19, 2270–2277 (2018).

    CAS  Google Scholar 

  42. 42.

    Cho, I. & Blaser, M. J. The human microbiome: at the interface of health and disease. Nat. Rev. Genet. 13, 260–270 (2012).

    CAS  Google Scholar 

  43. 43.

    Lynch, S. V. & Pedersen, O. The human intestinal microbiome in health and disease. N. Engl. J. Med. 375, 2369–2379 (2016).

    CAS  Google Scholar 

  44. 44.

    Pietroiusti, A., Magrini, A. & Campagnolo, L. New frontiers in nanotoxicology: gut microbiota/microbiome-mediated effects of engineered nanomaterials. Toxicol. Appl. Pharmacol. 299, 90–95 (2016).

    CAS  Google Scholar 

  45. 45.

    Javurek, A. B. et al. Gut dysbiosis and neurobehavioral alterations in rats exposed to silver nanoparticles. Sci. Rep. 7, 2822 (2017).

    Google Scholar 

  46. 46.

    Qiu, K., Durham, P. G. & Anselmo, A. C. Inorganic nanoparticles and the microbiome. Nano Res. 11, 4936–4954 (2018).

    CAS  Google Scholar 

  47. 47.

    Mosquera, M. J. et al. Immunomodulatory nanogels overcome restricted immunity in a murine model of gut microbiome-mediated metabolic syndrome. Sci. Adv. 5, eaav9788 (2019).

    CAS  Google Scholar 

  48. 48.

    Lee, Y., Lee, S. & Jon, S. Biotinylated Bilirubin nanoparticles as a tumor microenvironment-responsive drug delivery system for targeted cancer therapy. Adv. Sci. (Weinh.) 5, 1800017 (2018).

    Google Scholar 

  49. 49.

    Reissig, J. L., Storminger, J. L. & Leloir, L. F. A modified colorimetric method for the estimation of N-acetylamino sugars. J. Biol. Chem. 217, 959–966 (1955).

    CAS  Google Scholar 

  50. 50.

    McCoy, K. D., Geuking, M. B. & Ronchi, F. Gut microbiome standardization in control and experimental mice. Curr. Protoc. Immunol. 117, 23.1.1–23.1.13 (2017).

    Google Scholar 

  51. 51.

    Seekatz, A. M. et al. Fecal microbiota transplantation eliminates Clostridium difficile in a murine model of relapsing disease. Infect. Immun. 83, 3838–3846 (2015).

    CAS  Google Scholar 

  52. 52.

    Schloss, P. D. et al. Introducing mothur: open-source, platform-independent, community-supported software for describing and comparing microbial communities. Appl. Environ. Microbiol. 75, 7537–7541 (2009).

    CAS  Google Scholar 

  53. 53.

    Kozich, J. J., Westcott, S. L., Baxter, N. T., Highlander, S. K. & Schloss, P. D. Development of a dual-index sequencing strategy and curation pipeline for analyzing amplicon sequence data on the MiSeq Illumina sequencing platform. Appl. Environ. Microbiol. 79, 5112–5120 (2013).

    CAS  Google Scholar 

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Acknowledgements

This work was supported in part by the NIH (grant nos. R01EB022563, R01AI127070, R01CA210273, R01CA223804, U01CA210152, R01DK108901), the MTRAC for Life Sciences Hub and the Emerald Foundation. J.J.M. is a Young Investigator supported by the Melanoma Research Alliance (grant no. 348774), the DoD/CDMRP Peer Reviewed Cancer Research Program (grant no. W81XWH-16-1-0369) and a NSF CAREER Award (no. 1553831). Opinions interpretations, conclusions, and recommendations are those of the author and are not necessarily endorsed by the Department of Defense. The authors thank H. Atsushi for his technical help with LPMC isolation and flow cytometric analysis; the University of Michigan Medical School Host Microbiome Initiative for microbial community analysis; the University of Michigan Cancer Center Immunology Core for ELISA analysis; the ULAM In Vivo Animal Core for tissue sectioning and histological analysis of colon samples; the ULAM Pathology Core for blood analysis; and the College of Pharmacy Biochemical NMR Core at the University of Michigan.

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Y.L. and J.M. designed the experiments. Y.L. performed all experiments. K.S. and N.K. contributed technical expertise, including qPCR analysis, LPMC isolation and flow cytometry analysis. Y.L. and J.M. analysed the data. M.G. aided with interpretation of data on gut microbiome analysis. S.J. contributed the initial design of bilirubin conjugates. Y.L. and J.M. wrote the paper.

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Correspondence to James J. Moon.

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Lee, Y., Sugihara, K., Gillilland, M.G. et al. Hyaluronic acid–bilirubin nanomedicine for targeted modulation of dysregulated intestinal barrier, microbiome and immune responses in colitis. Nat. Mater. 19, 118–126 (2020). https://doi.org/10.1038/s41563-019-0462-9

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