Dysbiosis in infants and children is associated with increased susceptibility to inflammatory bowel disease (IBD) in adults. However, the mechanisms whereby the gut microbiota colonization in early life confers health outcomes throughout the lifespan remain unclear. Studies from the previous funding period isolated and cloned a Lactobacillus rhamnosus GG (LGG)-derived secretory protein, p40, and demonstrated that neonatal p40 supplementation prevents colitis in adult mice. Our preliminary studies discovered that p40 interacts with two transcriptional factors, Mga and Max, to regulate expression of Setd1? gene, which encodes a methyltransferase for catalyzing mono and trimethylation of histone 3-lysine 4 (H3K4me1/3). We identified TGF? as a potential target of p40-up-regulated Setd1?. IECs are rapidly renewed and continuously regenerated from intestinal stem cells (ISCs). We found that p40 modulates H3K4m31/3 in ISCs in early stage, and p40 treatment in neonates, but not adult mice, stimulates sustained increase in TGF? gene expression in IECs. TGF? has been shown to have multiple functions against inflammation. Thus, we hypothesize that p40 up-regulates Setd1? gene expression in IECs through increasing Mga:Max dimerization. Supplementation with p40 in early life stimulates Setd1?-dependent H3K4me1/3 deposition at the TGF? locus in ISCs, which is inherited by IECs to enable the sustained increase in TGF? production, and subsequently prevention of intestinal inflammation in adulthood.
In Aim 1, we will determine whether p40-stimulated Setd1? gene expression and H3K4me1/3 are required to drive the increase in expressing TGF? gene in IECs, and elucidate whether the interaction between p40 and the Mga:Max dimers mediates up-regulation of Setd1? production.
In Aim 2, we will identify the temporal window of p40 exposure in early life that causes the sustained increase in TGF? production in IECs, and define whether p40-regulated sustained increase in TGF? gene expression in IECs requires the increase of Setd1? gene expression inISCs in early life. Human and mouse enteroids and colonoids and 2D cultures and cell lines with silencing Setd1? or Mga genes, and mouse models of constitutive and inducible Setd1? gene knock down in IECs or ISCs will be utilized for these two aims.
In Aim 3, we will determine whether the sustained increase in TGF? production after p40 supplementation in early life is required to prevent colitis in adult mice. We will use neutralizing antibodies and inhibitors to block TGF? function in mice with induced and spontaneously developed colitis. We will also determine whether sustained increase in TGF? production enhances Tregs induction in the intestine and protective epithelial responses for the prevention of colitis in adulthood.
This aim will be tested in mice with TGF? receptor II deletion in CD4+ T cells and Smad4 deletion in IECs. Together, our proposed research will elucidate a novel mechanism underlying the consequences of long-lasting effects of p40 on prevention of colitis, and lay the foundation for developing early intervention with p40 as a novel therapy for prevention of IBD in adults.

Public Health Relevance

This proposal will not only provide a novel mechanism whereby p40, a functional factor of Lactobacillus rhamnosus GG, regulates histone modifications in intestinal epithelial cells, but also elucidate the contribution of p40-regulated epigenetic programming in intestinal stem cells in early life to enabling persistent effects of p40 on stimulating TGF? production to prevent intestinal inflammation. Thus, these studies have significant relevance to translational research by supporting early p40 intervention as a novel therapeutic strategy for prevention of intestinal inflammatory diseases in adulthood.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK081134-10A1
Application #
9972333
Study Section
Gastrointestinal Mucosal Pathobiology Study Section (GMPB)
Program Officer
Greenwel, Patricia
Project Start
2009-04-01
Project End
2024-03-31
Budget Start
2020-05-15
Budget End
2021-03-31
Support Year
10
Fiscal Year
2020
Total Cost
Indirect Cost
Name
Vanderbilt University Medical Center
Department
Type
DUNS #
079917897
City
Nashville
State
TN
Country
United States
Zip Code
37232
Shen, Xi; Liu, Liping; Peek, Richard M et al. (2018) Supplementation of p40, a Lactobacillus rhamnosus GG-derived protein, in early life promotes epidermal growth factor receptor-dependent intestinal development and long-term health outcomes. Mucosal Immunol 11:1316-1328
Yan, F; Liu, L; Cao, H et al. (2017) Neonatal colonization of mice with LGG promotes intestinal development and decreases susceptibility to colitis in adulthood. Mucosal Immunol 10:117-127
Wang, Y; Liu, L; Moore, D J et al. (2017) An LGG-derived protein promotes IgA production through upregulation of APRIL expression in intestinal epithelial cells. Mucosal Immunol 10:373-384
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Zhao, Gang; Liu, Liping; Peek Jr, Richard M et al. (2016) Activation of Epidermal Growth Factor Receptor in Macrophages Mediates Feedback Inhibition of M2 Polarization and Gastrointestinal Tumor Cell Growth. J Biol Chem 291:20462-72
Wang, Lihong; Cao, Hailong; Liu, Liping et al. (2014) Activation of epidermal growth factor receptor mediates mucin production stimulated by p40, a Lactobacillus rhamnosus GG-derived protein. J Biol Chem 289:20234-44
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Yu, Jinpu; Wang, Yue; Yan, Fang et al. (2014) Noncanonical NF-?B activation mediates STAT3-stimulated IDO upregulation in myeloid-derived suppressor cells in breast cancer. J Immunol 193:2574-86
Wang, Lihong; Cao, Hailong; Lu, Ning et al. (2013) Berberine inhibits proliferation and down-regulates epidermal growth factor receptor through activation of Cbl in colon tumor cells. PLoS One 8:e56666

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