Prophylactic efficacy of orally administered Bacillus poly-γ-glutamic acid, a non-LPS TLR4 ligand, against norovirus infection in mice

Wooseong Lee; Minwoo Kim; Seung Hoon Lee; Hae Gwang Jung; Jong Won Oh

doi:10.1038/s41598-018-26935-y

Prophylactic efficacy of orally administered Bacillus poly-γ-glutamic acid, a non-LPS TLR4 ligand, against norovirus infection in mice

Wooseong Lee, Minwoo Kim, Seung Hoon Lee, Hae Gwang Jung, Jong Won Oh

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19 Citations (Scopus)

Abstract

Poly-gamma-glutamic acid (γ-PGA), an extracellular biopolymer produced by Bacillus sp., is a non-canonical toll-like receptor 4 (TLR4) agonist. Here we show its antiviral efficacy against noroviruses. γ-PGA with a molecular mass of 2,000-kDa limited murine norovirus (MNV) replication in the macrophage cell line RAW264.7 by inducing interferon (IFN)-β and conferred resistance to viral infection-induced cell death. Additionally, γ-PGA interfered with viral entry into cells. The potent antiviral state mounted by γ-PGA was not attributed to the upregulation of TLR4 or TLR3, a sensor known to recognize norovirus RNA. γ-PGA sensing by TLR4 required the two TLR4-associated accessory factors MD2 and CD14. In ex vivo cultures of mouse ileum, γ-PGA selectively increased the expression of IFN-β in villi. In contrast, IFN-β induction was negligible in the ileal Peyer's patches (PPs) where its expression was primarily induced by the replication of MNV. Oral administration of γ-PGA, which increased serum IFN-β levels without inducing proinflammatory cytokines, reduced MNV loads in the ileum with PPs and mesenteric lymph nodes in mice. Our results disclose a γ-PGA-mediated non-conventional TLR4 signaling in the ileum, highlighting the potential use of γ-PGA as a prophylactic antiviral agent against noroviruses.

Original language	English
Article number	8667
Journal	Scientific reports
Volume	8
Issue number	1
DOIs	https://doi.org/10.1038/s41598-018-26935-y
Publication status	Published - 2018 Dec 1

Bibliographical note

Funding Information:
We thank Dr. Herbert W. Virgin (Washington University School of Medicine, St. Louis, MO, USA) and Dr. Kyeong-Ok Chang (Kansas State University, Manhattan, Kansas, USA) for providing the MNV1. CW1 strain and the HG23 cell line, respectively. We thank Dr. Moon-Hee Sung (Kookmin University, Seoul, Korea) for helpful discussions. This work was supported by a grant from the National Research Foundation of Korea funded by the Korean government (MSIP) (2016R1A5A1004694) and in part by the Yonsei University Future-leading Research Initiative grant 2015 RMS2-2014-22-0193. W.L. and S.-H.L were the recipients of a postdoctoral fellowship (2017-12-0035) and a graduate school research scholarship (2017), respectively, from Yonsei University.

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© 2018 The Author(s).

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title = "Prophylactic efficacy of orally administered Bacillus poly-γ-glutamic acid, a non-LPS TLR4 ligand, against norovirus infection in mice",

abstract = "Poly-gamma-glutamic acid (γ-PGA), an extracellular biopolymer produced by Bacillus sp., is a non-canonical toll-like receptor 4 (TLR4) agonist. Here we show its antiviral efficacy against noroviruses. γ-PGA with a molecular mass of 2,000-kDa limited murine norovirus (MNV) replication in the macrophage cell line RAW264.7 by inducing interferon (IFN)-β and conferred resistance to viral infection-induced cell death. Additionally, γ-PGA interfered with viral entry into cells. The potent antiviral state mounted by γ-PGA was not attributed to the upregulation of TLR4 or TLR3, a sensor known to recognize norovirus RNA. γ-PGA sensing by TLR4 required the two TLR4-associated accessory factors MD2 and CD14. In ex vivo cultures of mouse ileum, γ-PGA selectively increased the expression of IFN-β in villi. In contrast, IFN-β induction was negligible in the ileal Peyer's patches (PPs) where its expression was primarily induced by the replication of MNV. Oral administration of γ-PGA, which increased serum IFN-β levels without inducing proinflammatory cytokines, reduced MNV loads in the ileum with PPs and mesenteric lymph nodes in mice. Our results disclose a γ-PGA-mediated non-conventional TLR4 signaling in the ileum, highlighting the potential use of γ-PGA as a prophylactic antiviral agent against noroviruses.",

author = "Wooseong Lee and Minwoo Kim and Lee, {Seung Hoon} and Jung, {Hae Gwang} and Oh, {Jong Won}",

note = "Funding Information: We thank Dr. Herbert W. Virgin (Washington University School of Medicine, St. Louis, MO, USA) and Dr. Kyeong-Ok Chang (Kansas State University, Manhattan, Kansas, USA) for providing the MNV1. CW1 strain and the HG23 cell line, respectively. We thank Dr. Moon-Hee Sung (Kookmin University, Seoul, Korea) for helpful discussions. This work was supported by a grant from the National Research Foundation of Korea funded by the Korean government (MSIP) (2016R1A5A1004694) and in part by the Yonsei University Future-leading Research Initiative grant 2015 RMS2-2014-22-0193. W.L. and S.-H.L were the recipients of a postdoctoral fellowship (2017-12-0035) and a graduate school research scholarship (2017), respectively, from Yonsei University. Publisher Copyright: {\textcopyright} 2018 The Author(s).",

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doi = "10.1038/s41598-018-26935-y",

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AU - Lee, Seung Hoon

AU - Jung, Hae Gwang

AU - Oh, Jong Won

N1 - Funding Information: We thank Dr. Herbert W. Virgin (Washington University School of Medicine, St. Louis, MO, USA) and Dr. Kyeong-Ok Chang (Kansas State University, Manhattan, Kansas, USA) for providing the MNV1. CW1 strain and the HG23 cell line, respectively. We thank Dr. Moon-Hee Sung (Kookmin University, Seoul, Korea) for helpful discussions. This work was supported by a grant from the National Research Foundation of Korea funded by the Korean government (MSIP) (2016R1A5A1004694) and in part by the Yonsei University Future-leading Research Initiative grant 2015 RMS2-2014-22-0193. W.L. and S.-H.L were the recipients of a postdoctoral fellowship (2017-12-0035) and a graduate school research scholarship (2017), respectively, from Yonsei University. Publisher Copyright: © 2018 The Author(s).

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N2 - Poly-gamma-glutamic acid (γ-PGA), an extracellular biopolymer produced by Bacillus sp., is a non-canonical toll-like receptor 4 (TLR4) agonist. Here we show its antiviral efficacy against noroviruses. γ-PGA with a molecular mass of 2,000-kDa limited murine norovirus (MNV) replication in the macrophage cell line RAW264.7 by inducing interferon (IFN)-β and conferred resistance to viral infection-induced cell death. Additionally, γ-PGA interfered with viral entry into cells. The potent antiviral state mounted by γ-PGA was not attributed to the upregulation of TLR4 or TLR3, a sensor known to recognize norovirus RNA. γ-PGA sensing by TLR4 required the two TLR4-associated accessory factors MD2 and CD14. In ex vivo cultures of mouse ileum, γ-PGA selectively increased the expression of IFN-β in villi. In contrast, IFN-β induction was negligible in the ileal Peyer's patches (PPs) where its expression was primarily induced by the replication of MNV. Oral administration of γ-PGA, which increased serum IFN-β levels without inducing proinflammatory cytokines, reduced MNV loads in the ileum with PPs and mesenteric lymph nodes in mice. Our results disclose a γ-PGA-mediated non-conventional TLR4 signaling in the ileum, highlighting the potential use of γ-PGA as a prophylactic antiviral agent against noroviruses.

AB - Poly-gamma-glutamic acid (γ-PGA), an extracellular biopolymer produced by Bacillus sp., is a non-canonical toll-like receptor 4 (TLR4) agonist. Here we show its antiviral efficacy against noroviruses. γ-PGA with a molecular mass of 2,000-kDa limited murine norovirus (MNV) replication in the macrophage cell line RAW264.7 by inducing interferon (IFN)-β and conferred resistance to viral infection-induced cell death. Additionally, γ-PGA interfered with viral entry into cells. The potent antiviral state mounted by γ-PGA was not attributed to the upregulation of TLR4 or TLR3, a sensor known to recognize norovirus RNA. γ-PGA sensing by TLR4 required the two TLR4-associated accessory factors MD2 and CD14. In ex vivo cultures of mouse ileum, γ-PGA selectively increased the expression of IFN-β in villi. In contrast, IFN-β induction was negligible in the ileal Peyer's patches (PPs) where its expression was primarily induced by the replication of MNV. Oral administration of γ-PGA, which increased serum IFN-β levels without inducing proinflammatory cytokines, reduced MNV loads in the ileum with PPs and mesenteric lymph nodes in mice. Our results disclose a γ-PGA-mediated non-conventional TLR4 signaling in the ileum, highlighting the potential use of γ-PGA as a prophylactic antiviral agent against noroviruses.

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Prophylactic efficacy of orally administered Bacillus poly-γ-glutamic acid, a non-LPS TLR4 ligand, against norovirus infection in mice

Abstract

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