Dysbiosis is a hallmark of inflammatory bowel disease (IBD), but it is unclear which specific intestinal bacteria predispose to and which protect from IBD and how they are regulated. Peptidoglycan recognition proteins (Pglyrps) are antibacterial, participate in maintaining intestinal microflora, and modulate inflammatory responses. Mice deficient in any one of the four Pglyrp genes are more sensitive to dextran sulfate sodium (DSS)-induced colitis, and stools from Pglyrp-deficient mice transferred to wild type (WT) germ-free mice predispose them to much more severe colitis than stools from WT mice. However, the identities of these Pglyrp-regulated bacteria that predispose Pglyrp-deficient mice to colitis or protect WT mice from colitis are not known. Here we identified significant changes in β-diversity of stool bacteria in Pglyrp-deficient mice compared with WT mice. The most consistent changes in microbiome in all Pglyrp-deficient mice were in Bacteroidales, from which we selected four species, two with increased abundance (Prevotella falsenii and Parabacteroides distasonis) and two with decreased abundance (Bacteroides eggerthii and Alistipes finegoldii). We then gavaged WT mice with stock type strains of these species to test the hypothesis that they predispose to or protect from DSS-induced colitis. P. falsenii, P. distasonis, and B. eggerthii all enhanced DSS-induced colitis in both WT mice with otherwise undisturbed intestinal microflora and in WT mice with antibiotic-depleted intestinal microflora. By contrast, A. finegoldii (which is the most abundant species in WT mice) attenuated DSS-induced colitis both in WT mice with otherwise undisturbed intestinal microflora and in WT mice with antibiotic-depleted intestinal microflora, similar to the colitis protective effect of the entire normal microflora. These results identify P. falsenii, P. distasonis, and B. eggerthii as colitis-promoting species and A. finegoldii as colitis-protective species.

SUMMARY There are multiple mechanisms of maintaining tolerance in the gut that protect the intestine from excessive inflammatory response to intestinal microorganisms. We report here that all four mammalian Peptidoglycan Recognition Proteins (PGRPs or Pglyrps) protect the host from colitis induced by dextran sulfate sodium (DSS). Pglyrp1−/−, Pglyrp2−/−, Pglyrp3−/−, and Pglyrp4−/− mice are all more sensitive than wild type (WT) mice to DSS-induced colitis due to changes to more inflammatory gut microflora, higher production of interferon-γ and interferon-inducible genes, and increase in NK cells in the colon upon initial exposure to DSS, which leads to severe hyperplasia of the lamina propria, loss of epithelial cells, and ulceration in the colon. Thus in WT mice PGRPs protect the colon from early inflammatory response and loss of the barrier function of intestinal epithelium by promoting normal bacterial flora and by preventing damaging production of interferon-γ by NK cells in response to injury.

Skin protects the body from the environment and is an important component of the innate and adaptive immune systems. Atopic dermatitis and contact dermatitis are among the most frequent inflammatory skin diseases and are both determined by multigenic predisposition, environmental factors, and aberrant immune response. Peptidoglycan Recognition Proteins (Pglyrps) are expressed in the skin and we report here that they modulate sensitivity to experimentally-induced atopic dermatitis and contact dermatitis. Pglyrp3 −/− and Pglyrp4 −/− mice (but not Pglyrp2 −/− mice) develop more severe oxazolone-induced atopic dermatitis than wild type (WT) mice. The common mechanism underlying this increased sensitivity of Pglyrp3 −/− and Pglyrp4 −/− mice to atopic dermatitis is reduced recruitment of Treg cells to the skin and enhanced production and activation Th17 cells in Pglyrp3 −/− and Pglyrp4 −/− mice, which results in more severe inflammation and keratinocyte proliferation. This mechanism is supported by decreased inflammation in Pglyrp3 −/− mice following in vivo induction of Treg cells by vitamin D or after neutralization of IL-17. By contrast, Pglyrp1 −/− mice develop less severe oxazolone-induced atopic dermatitis and also oxazolone-induced contact dermatitis than WT mice. Thus, Pglyrp3 and Pglyrp4 limit over-activation of Th17 cells by promoting accumulation of Treg cells at the site of chronic inflammation, which protects the skin from exaggerated inflammatory response to cell activators and allergens, whereas Pglyrp1 has an opposite pro-inflammatory effect in the skin.

Asthma is a common inflammatory disease involving crosstalk between innate and adaptive immunity. We reveal that antibacterial innate immunity protein, peptidoglycan recognition protein 1 (Pglyrp1), is involved in the development of allergic asthma. Pglyrp1−/− mice developed less severe asthma than wild type (WT) mice following sensitization with house dust mite (HDM) allergen. HDM-sensitized Pglyrp1−/− mice, compared with WT mice, had diminished: bronchial hyper-responsiveness (lung airway resistance); numbers of eosinophils, neutrophils, lymphocytes, and macrophages in bronchoalveolar lavage fluid and lungs; inflammatory cell infiltrates in the lungs around bronchi, bronchioles, and pulmonary arteries and veins; lung remodeling (mucin-producing goblet cell hyperplasia and metaplasia and smooth muscle hypertrophy and fibrosis); levels of IgE, eotaxins, IL-4, IL-5, and IL-17 in the lungs; and numbers of Th2 and Th17 cells and expression of their marker genes in the lungs. The mechanism underlying this decreased sensitivity of Pglyrp1−/− mice to asthma was increased generation and activation of CD8α+β+ and CD8α+β− plasmacytoid dendritic cells (pDC) and increased recruitment and activity of regulatory T (Treg) cells in the lungs. In vivo depletion of pDC in HDM-sensitized Pglyrp1−/− mice reversed the low responsive asthma phenotype of Pglyrp1−/− mice to resemble the more severe WT phenotype. Thus, Pglyrp1−/− mice efficiently control allergic asthma by up-regulating pDC and Treg cells in the lungs, whereas in WT mice Pglyrp1 is proinflammatory and decreases pDC and Treg, and increases pro-asthmatic Th2 and Th17 responses. Blocking Pglyrp1 or enhancing pDC in the lungs may be beneficial for prevention and treatment of asthma.

Skin protects the body from the environment and is an important component of the innate and adaptive immune systems. Psoriasis is a frequent inflammatory skin disease of unknown etiology determined by multigenic predisposition, environmental factors, and aberrant immune response. Peptidoglycan Recognition Proteins (Pglyrps) are expressed in the skin and we report here that they modulate sensitivity in an experimentally-induced mouse model of psoriasis. We demonstrate that Pglyrp2−/− mice (but not Pglyrp3−/− and Pglyrp4−/− mice) are more sensitive to the development of 12-O-tetradecanoylphorbol 13-acetate (TPA)-induced psoriasis-like inflammation, whereas Pglyrp1−/− mice are less sensitive. The mechanism underlying this increased sensitivity of Pglyrp2−/− mice to TPA-induced psoriasis-like inflammation is reduced recruitment of Treg cells to the skin and enhanced production and activation of Th17 cells in the skin in Pglyrp2−/− mice, which results in more severe inflammation and keratinocyte proliferation. Thus, in wild type mice, Pglyrp2 limits over-activation of Th17 cells by promoting accumulation of Treg cells at the site of inflammation, which protects the skin from the exaggerated inflammatory response.

Aberrant immune response and changes in the gut microflora are the main causes of inflammatory bowel disease (IBD). Peptidoglycan recognition proteins (Pglyrp1, Pglyrp2, Pglyrp3, and Pglyrp4) are bactericidal innate immunity proteins that maintain normal gut microbiome, protect against experimental colitis, and are associated with inflammatory bowel disease in humans. Nod2 is an intracellular bacterial sensor and may be required for maintaining normal gut microbiome. Mutations in Nod2 are strongly associated with Crohn's disease, but the causative mechanism is not understood, and Nod2 role in ulcerative colitis is not known. Because IBD is likely caused by variable multiple mutations in different individuals, in this study we examined the combined role of Pglyrp3 and Nod2 in the development of experimental colitis in mice. We demonstrate that a combined deficiency of Pglyrp3 and Nod2 results in higher sensitivity to dextran sodium sulfate (DSS)-induced colitis compared with a single deficiency. Pglyrp3−/−Nod2−/− mice had decreased survival and higher loss of body weight, increased intestinal bleeding, higher apoptosis of colonic mucosa, elevated expression of cytokines and chemokines, altered gut microbiome, and increased levels of ATP in the colon. Increased sensitivity to DSS-induced colitis in Pglyrp3−/−Nod2−/− mice depended on increased apoptosis of intestinal epithelium, changed gut microflora, and elevated ATP. Pglyrp3 deficiency contributed colitispredisposing intestinal microflora and increased intestinal ATP, whereas Nod2 deficiency contributed higher apoptosis and responsiveness to increased level of ATP. In summary, Pglyrp3 and Nod2 are both required for maintaining gut homeostasis and protection against colitis, but their protective mechanisms differ.