Germ-free Environment Research Articles

TMIC-15. GUT MICROBIOTA CONTROLS NEUROFIBROMATOSIS-1 (NF1) OPTIC GLIOMA GROWTH THROUGH IMMUNE CIRCUIT MODULATION

Abstract Changes in the gut microbiome can have profound effects on the nervous system through modulation of T cell and microglia function. Since our prior studies demonstrated that T cells and microglia positively regulate low-grade glioma growth through the establishment of T cell-microglia immune circuit, we sought to explore the impact of changes in the gut microbiota on tumor biology. For these studies, genetically engineered Neurofibromatosis type 1 (NF1) optic glioma (Nf1-OPG) mice were raised in a germ-free environment or treated with antibiotics to deplete gut bacteria. First, we demonstrated that Nf1-OPG mice raised in a germ-free (gnotobiotic) environment or treated with specific antibiotics lacked optic gliomas and had improved OPG-induced retinal pathology (increased retinal nerve fiber layer thickness). Second, germ-free and antibiotic-treated Nf1-OPG mice gavaged with fecal microbiota from Nf1-OPG mice raised in a standard barrier facility restored optic glioma growth. Third, we showed that both germ-free and antibiotic-treated Nf1-OPG mice exhibited reduced intra-tumoral CD8+ T cell content resulting from decreased microglia chemokine production. Fourth, we discovered a gut-brain paracrine axis whose interruption reduced optic glioma growth and attenuated retinal nerve fiber layer thinning. Collectively, these findings establish a mechanistic relationship between the gut microbiota and brain tumor growth relevant to potential therapeutic interventions for pediatric low-grade gliomas.

The microbiome in HLA-B27-associated disease: implications for acute anterior uveitis and recommendations for future studies.

The pathogenesis of human leukocyte antigen (HLA)-B27-associated diseases such as acute anterior uveitis (AAU) and ankylosing spondylitis (AS) remains poorly understood, though Gram-negative bacteria and subclinical bowel inflammation are strongly implicated. Accumulating evidence from animal models and clinical studies supports several hypotheses, including HLA-B27-dependent dysbiosis, altered intestinal permeability, and molecular mimicry. However, the existing literature is hampered by inadequate studies designed to establish causation or uncover the role of viruses and fungi. Moreover, the unique disease model afforded by AAU to study the gut microbiota has been neglected. This review critically evaluates the current literature and prevailing hypotheses on the link between the gut microbiota and HLA-B27-associated disease. We propose a new potential role for HLA-B27-driven altered antibody responses to gut microbiota in disease pathogenesis and outline recommendations for future well-controlled human studies, focusing on AAU.

Microbiome and gut-brain axis affecting stress behavior

In context to the neurological system, the function and evolution of host physiology as influenced by intestinal microbiota are of great interest. It has long been understood how crucial the gut-brain axis is in controlling reactions to stress. More recently, the microbiota has become a crucial component in this gut-brain regulation, particularly under stressful circumstances brought on by actual or perceived homeostatic strain. The gut microbiota seems to have an impact on the growth of emotional behavior, stress- and pain-modulation systems, and brain neurotransmitter systems, according to studies employing mice raised in a germ-free environment. Current evidence suggests that multiple mechanisms, including neural pathways and immune signaling, may be involved in gut microbiota–to–brain signaling and that the brain can in turn alter microbial composition and behavior via the autonomic nervous system. The gut microbiota has been implicated in a variety of stress-related conditions including anxiety, depression, and irritable bowel syndrome, although this is largely based on animal studies or correlative analysis in patient populations. Additional research in humans is sorely needed to reveal the relative impact and causal contribution of the microbiome to stress-related disorders. We will briefly explore the crucial aspect of this axis in this review, as well as the methodological issues that have been raised in previous attempts to define normal microbiota and chronicle its temporal development.

Gut microbiome and type 2 diabetes.

Dietary patterns with excess caloric have shaped a complex metabolic disorders like type 2 diabetes (T2D). T2D involves complications in the metabolism of glucose, lipid, cholesterol and their storage. Along with the metabolic dysregulation, systemic inflammation is also the reason for Insulin Resistance and T2D. The importance of gut microbiota has recently been highlighted. It establishes a link between dietary patterns and the types of bacteria that overgrow and modify fermentation bi-products such as SCFA, secondary bile acids, and mucosal immune cells. These changes have a direct impact on the liver's metabolism and immune system. As a result, using Pre-Pro-biotics to manage microbiota can assist overcome or lessening disease symptoms. Antibiotics are currently employed to produce a germ-free environment or to eradicate specific types of bacteria in order to better understand the role of microflora. This chapter covers the basics of good bacteria, as well as the mechanisms that they work on.

Estimation for Akshaya Failure Model with Competing Risks under Progressive Censoring Scheme with Analyzing of Thymic Lymphoma of Mice Application

In several experiments of survival analysis, the cause of death or failure of any subject may be characterized by more than one cause. Since the cause of failure may be dependent or independent, in this work, we discuss the competing risk lifetime model under progressive type‐II censored where the removal follows a binomial distribution. We consider the Akshaya lifetime failure model under independent causes and the number of subjects removed at every failure time when the removal follows the binomial distribution with known parameters. The classical and Bayesian approaches are used to account for the point and interval estimation procedures for parameters and parametric functions. The Bayes estimate is obtained by using the Markov Chain Monte Carlo (MCMC) method under symmetric and asymmetric loss functions. We apply the Metropolis–Hasting algorithm to generate MCMC samples from the posterior density function. A simulated data set is applied to diagnose the performance of the two techniques applied here. The data represented the survival times of mice kept in a conventional germ‐free environment, all of which were exposed to a fixed dose of radiation at the age of 5 to 6 weeks, which was used as a practice for the model discussed. There are 3 causes of death. In group 1, we considered thymic lymphoma to be the first cause and other causes to be the second. On the base of mice data, the survival mean time (cumulative incidence function) of mice of the second cause is higher than the first cause.

Abstract P103: The introduction of a single strain of Bacillus into a germ-free environment did not impact the anti-PD-1 efficacy in a MC38 syngeneic model

Abstract Background. Although immunotherapy has led to exceptional and durable clinical response, the majority of patients respond poorly to the current immunotherapies. Growing evidence has linked some of the poor responsiveness to the gut microbiota, and the modulation of gut microbiome composition is becoming a promising new strategy to enhance immune checkpoint inhibitor (ICI) treatment outcome. Mouse tumor modelling under germ-free (GF) conditions combined with introduction of defined bacterial strains could be a useful approach to investigate the impact of microbiota on ICI efficacy, as well as understanding the underlying mechanisms. We previously demonstrated that GF mice exhibited a significantly poor response to anti-PD-1 therapy when compared to the specific pathogen free (SPF) mice in a subcutaneous MC38 colorectal cancer model, which is consistent with other reports. Methods. Introduction of a single strain of Bacillus in the GF-environment was assessed for its impact on the anti-mouse PD-1 monoclonal antibody (mAb) therapy in GF-mice and SPF mice, both for efficacy and pharmacodynamics tumor infiltrating lymphocytes (TILs) profiling. C57BL/6 mice were inoculated subcutaneously with MC38 tumor cells and when the tumors were reached 80-120mm3, the mice were randomized for isotype or anti-PD-1 mAb treatment. Fecal sample 16S rRNA analysis (NGS) was used to confirm the gut bacteria status. Results. The MC38 tumor in GF mice has significantly fast baseline growth kinetics, even with the introduction of Bacillus under GF conditions compared to SPF mice, suggesting tumor immunity was not enhanced by Bacillus. Despite the introduction of Bacillus, GF mice also showed reduced response to anti-PD-1 treatment when compared to the SPF mice as previously reported, further confirming that introduction of Bacillus had minimal effects on the efficacy of anti-PD-1 therapy. Moreover, the SPF mice and GF mice with Bacillus exhibited distinct profiles of TILs, consistent with distinct efficacies as observed. GF free mice showed a lower frequency of CD45+ TILs in comparison to SPF mice. In addition, GF mice exhibited lower frequency of CD8+ TILs and TIL- NKT when compared to the SPF mice, both of which are consistent with the stronger efficacy seen in SPF mice. Meanwhile, GF mice also exhibited higher granulocytic myeloid derived suppressor cells (gMDSC) and lower M1/M2 ratio, both of which imply a more suppressive tumor microenvironment in GF mice. Fecal sample analysis using 16S rRNA analysis confirmed a single strain of Bacillus was indeed introduced into the guts of all the GF mice. Conclusions. The GF conditions provide a useful environment for the investigation of specific microbiota strains on the impact on ICI treatment outcome. In summary, the introduction of Bacillus in GF mice did not impact the efficacy of anti-PD-1, thus suggesting that other strain(s) of gut microbiota in SPF mice may impact this and need to be investigated. Citation Format: Tao Yang, Bonnie Xiaobo Chen, Rongfei Lu, Xiaoyu An, Mingfa Zang, Jingjun Li, Sheng Guo, Wubin Qian, Jian Fei, Tongyang Hao, Edward Xu, Henry Li. The introduction of a single strain of Bacillus into a germ-free environment did not impact the anti-PD-1 efficacy in a MC38 syngeneic model [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P103.

HI (HARLEQUIN ICHTHYOSIS) BABY

HI is a very rare type of genetical abnormality but can be seen and scared by seeing various social media footages. It is associated with deletion and truncation mutations of a keratinocyte lipid transporter. Harlequin disorder is categorized by diffuse epidermal hyperkeratinization along with defective desquamation. During birth, the HI phenotype is conspicuous with thick hyperkeratotic plate-like scales with deep dermal ssures, severe ectropion and eclabium, among other ndings. In this ichthyosis marked eclabium and ectropion are present secondary to the taut as well as in the unyielding skin. The ears may be inattentive, absent or poorly developed. Even the arms, feet, and digits have exion contractures and may be hypoplastic. The skin wall is harshly compromised, leading to undue water loss, causes electrolyte abnormalities, temperature alteration and an increased risk of deadly infection. Some of the primary treatments are smearing retinoids application for shedding the hard and scaly skin, topical antibiotics application can prevent infection, insertion an ET tube in the airway to assistance with breathing, applying lubricating eye drops or protective devices on the eyes. Following ongoing treatment like humidied incubator (for premature infants), continuous monitoring of TPR and SpO2, early intubation(optional), frequent cultures of the skin should be taken for lab testing, monitoring serum electrolyte levels, maintaining a germ-free environment to evade infection etc.

Comparative Efficacy of Two Types of Antibiotic Mixtures in Gut Flora Depletion in Female C57BL/6 Mice.

Over the last decade, interest in the role of the microbiome in health and disease has increased. The use of germ-free animals and depletion of the microbial flora using antimicrobials are 2 methods commonly used to study the microbiome in laboratory mice. Germ-free mice are born, raised, and studied in isolators in the absence of any known microbes; however, the equipment, supplies, and training required for the use of these mice can be costly and time-consuming. The use of antibiotics to decrease the microbial flora does not require special equipment, can be used for any mouse strain, and is relatively inexpensive; however, mice treated in this manner still retain microbes and they do not live in a germ-free environment. One commonly used antibiotic cocktail regimen uses ampicillin, neomycin, metronidazole, and vancomycin in the drinking water for 2 to 4 wk. We found that the palatability of this mixture is low, resulting in weight loss and leading to removal of mice from the study. The addition of sucralose to the medicated water and making wet food (mash) with the medicated water improved intake; however, the low palatability still resulted in a high number of mice requiring removal. The current study evaluated a new combination of antibiotics designed to reduce the gut microbiota while maintaining body weights. C57BL/6NCrl mice were placed on one of the following drinking water regimens: ampicillin/neomycin/metronidazole/vancomycin water (n = 16), enrofloxacin/ampicillin water ( n = 12), or standard reverse osmosis deionized water (RODI) ( n = 11). During an 8 day regimen, mice were weighed and water consumption was measured. Feces were collected before and after 8 d of treatment. Quantitative real-time PCR (real-time qPCR) for 16S bacterial ribosome was performed on each sample, and values were compared among groups. The combination of enrofloxacin and ampicillin improved water intake, together with a greater reduction in gut flora.

Dynamic assessment of the impact of gut microbiota transfer in a mouse model of chronic intestinal dysbiosis

Abstract Objective There is growing evidence supporting that the gut microbiota is a major driver of human health and disease. While gut microbiota transfer (GMT) is commonly used as an approach to restore "eubiosis", there is a surprising lack of data on whether the transferred microbiota efficiently and durably repopulate the gut of the transplanted subject. Moreover, little is known on the effects of GMT on non-alcoholic fatty liver disease (NAFLD). Methods Chronic dysbiosis and NAFLD-like liver injury were induced by feeding C57Bl/6j mice for 16 weeks with a high-fat diet. For GMT, dysbiotic mice underwent preliminary gut cleansing, followed by oral gavage with a suspension of fresh fecal matter procured from a pool of lean mice (1 dose, or 10 doses). We next characterized microbiota composition and we measured the relative abundance of specific pathobionts in recipient mice, using high-throughput shotgun analysis in a dynamic manner, over time. All experiments took place in a specific germ-free environment. Results After 4 months on a high-fat diet, mice displayed fatty liver infiltration with moderate parenchymal inflammatory changes. Dysbiosis was evidenced by a reduced bacterial diversity, as well as a dramatically increased abundance of Firmicutes, and lower Verrucomicrobia and Actinobacteria. Gut microbiota transfer was associated with a transitory reduction in NAFLD-induced hepatocellular injury. While dysbiotic mice displayed a shift in their microbiota composition towards that of lean donors after GMT, this effect rapidly faded after one week, and mice recovered their initial, dysbiotic microbiota. Conclusion The current study indicates that, when used in mice with chronically established dysbiosis, GMT is merely associated with transitory changes in gut microbiota composition, as well as significant but moderate reduction in hepatocellular injury.

Putative Pathobionts in HLA-B27-Associated Spondyloarthropathy.

Spondyloarthritis (SpA) is a group of immune mediated inflammatory diseases with a strong association to the major histocompatibility (MHC) class I molecule, HLA-B27. Although the association between HLA-B27 and AS has been known for almost 50 years, the mechanisms underlying disease pathogenesis are elusive. Over the years, three hypotheses have been proposed to explain HLA-B27 and disease association: 1) HLA B27 presents arthritogenic peptides and thus creates a pathological immune response; 2) HLA-B27 misfolding causes endoplasmic reticulum (ER) stress which activates the unfolded protein response (UPR); 3) HLA-B27 dimerizes on the cell surface and acts as a target for natural killer (NK) cells. None of these hypotheses explains SpA pathogenesis completely. Evidence supports the hypothesis that HLA-B27-related diseases have a microbial pathogenesis. In animal models of various SpAs, a germ-free environment abrogates disease development and colonizing these animals with gut commensal microbes can restore disease manifestations. The depth of microbial influence on SpA development has been realized due to our ability to characterize microbial communities in the gut using next-generation sequencing approaches. In this review, we will discuss various putative pathobionts in the pathogenesis of HLA-B27-associated diseases. We pursue whether a single pathobiont or a disruption of microbial community and function is associated with HLA-B27-related diseases. Furthermore, rather than a specific pathobiont, metabolic functions of various disease-associated microbes might be key. While the use of germ-free models of SpA have facilitated understanding the role of microbes in disease development, future studies with animal models that mimic diverse microbial communities instead of mono-colonization are indispensable. We discuss the causal mechanisms underlying disease pathogenesis including the role of these pathobionts on mucin degradation, mucosal adherence, and gut epithelial barrier disruption and inflammation. Finally, we review the various uses of microbes as therapeutic modalities including pre/probiotics, diet, microbial metabolites and fecal microbiota transplant. Unravelling these complex host-microbe interactions will lead to the development of new targets/therapies for alleviation of SpA and other HLA-B27 associated diseases.

Evaluation of cognitive function in diabetic germ-free mice

It is widely known that diabetes is a risk factor for dementia. It has been reported that the cognitive function is impaired in the streptozotocin (STZ)-induced diabetic mouse model. Recently, it has also been reported that alteration of the intestinal flora is a risk factor for dementia. However, effects of alteration of the intestinal flora due to dementia on the cognitive function have not been adequately assessed. One of the reasons is that germ-free mice, which have no intestinal flora, need to be housed in a germ-free environment. Therefore, it is difficult to subject to behavioral tests in a germ-free environment to assess the cognitive function.

Dietary simple sugars alter microbial ecology in the gut and promote colitis in mice.

The higher prevalence of inflammatory bowel disease (IBD) in Western countries points to Western diet as a possible IBD risk factor. High sugar, which is linked to many noncommunicable diseases, is a hallmark of the Western diet, but its role in IBD remains unknown. Here, we studied the effects of simple sugars such as glucose and fructose on colitis pathogenesis in wild-type and Il10-/- mice. Wild-type mice fed 10% glucose in drinking water or high-glucose diet developed severe colitis induced by dextran sulfate sodium. High-glucose-fed Il10-/- mice also developed a worsened colitis compared to glucose-untreated Il10-/- mice. Short-term intake of high glucose or fructose did not trigger inflammatory responses in healthy gut but markedly altered gut microbiota composition. In particular, the abundance of the mucus-degrading bacteria Akkermansia muciniphila and Bacteroides fragilis was increased. Consistently, bacteria-derived mucolytic enzymes were enriched leading to erosion of the colonic mucus layer of sugar-fed wild-type and Il10-/- mice. Sugar-induced exacerbation of colitis was not observed when mice were treated with antibiotics or maintained in a germ-free environment, suggesting that altered microbiota played a critical role in sugar-induced colitis pathogenesis. Furthermore, germ-free mice colonized with microbiota from sugar-treated mice showed increased colitis susceptibility. Together, these data suggest that intake of simple sugars predisposes to colitis and enhances its pathogenesis via modulation of gut microbiota in mice.

Early Exposure to Gut Microbiome Reduces Hepatocellular Carcinoma Risks in Mice

Aims. Liver cancer is a multietiological disease that has multiple factors contributing to the hepatocarcinogenic process, e.g., hepatitis viruses, carcinogens, male sex, or metabolic factors. Notably, emerging evidence reported that gut microbiota is crucial to the pathogenesis of hepatocellular carcinoma (HCC) via activation of innate immunity. However, the effect of time to gut microbiota exposure after birth is unknown. Using a germ-free animal housing environment, instead of antibiotics, we examined the effects of various time-to-exposure (TTE) to gut microbiota durations on HCC risk. Methods. HBV or carcinogen-mediated spontaneous HCC models were implemented in this study. The HCC incidence rates in mice either kept germ-free (GF; that is, with no exposure to gut microbiota) or exposed to gut microbiota after being moved to a specific pathogen-free (SPF) housing environment and with various time-to-exposure (TTE) durations, namely, 5 weeks after birth, 10 weeks after birth, or since conception (that is, 5-week TTE group, 10-week TTE group, and SPF group, respectively), were recorded. The mice were sacrificed at 30 or 40 weeks after birth, and macro-/microscopic observations and pathological diagnosis were performed. Results. The incidence of liver tumors among the male mice was higher than that among the female mice in the carcinogen-induced HCC mice sacrificed at 40 weeks after birth (with P=0.011, 0.035, 0.0003, and 0.012, respectively, in the GF group, 5-week TTE group, 10-week TTE group, and SPF group). Similarly, in the HBV-HCC model, the incidence of liver tumors among the male mice was significantly higher than that among the female mice (with P=0.013, 0.020, 0.012, and 0.002, respectively, in the GF group, 5-week TTE group, 10-week TTE group, and SPF group). These results suggest that gut microbiota exposure is irrelevant to the male sex preference of HCC. Surprisingly, when comparing carcinogen-induced HCC male mice in the 10-week TTE group (90%; n=10), 5-week TTE group (56%; n=9), and SPF group (30%; n=10) (P=0.020), we found that the incidence of liver tumors was higher in the mice with later exposure to gut microbiome. Similarly, when comparing HBV-HCC male mice in the 10-week TTE group (100%; n=11), 5-week TTE group (70%; n=10), and SPF group (33%; n=9) (P=0.080), we also found that the incidence of liver tumors was higher in the mice with later exposure to gut microbiome. Conclusions. Early (prepubertal) exposure to gut microbiome reduces the risk of HCC development, indicating a potentially important factor for cancer surveillance. Exploring the mechanisms by which such exposure affects HCC risk might lead to novel cancer vaccines.

The gut microbiome-joint connection: implications in osteoarthritis.

Osteoarthritis is a debilitating disease leading to joint degeneration, inflammation, pain, and disability. Despite efforts to develop a disease modifying treatment, the only accepted and available clinical approaches involve palliation. Although many factors contribute to the development of osteoarthritis, the gut microbiome has recently emerged as an important pathogenic factor in osteoarthritis initiation and progression. This review examines the literature to date regarding the link between the gut microbiome and osteoarthritis. Studies showing correlations between serum levels of bacterial metabolites and joint degeneration were the first links connecting a dysbiosis of the gut microbiome with osteoarthritis. Further investigations have demonstrated that microbial community shifts induced by antibiotics, a germ-free environment or high-fat are important underlying factors in joint homeostasis and osteoarthritis. It follows that strategies to manipulate the microbiome have demonstrated efficacy in mitigating joint degeneration in osteoarthritis. Moreover, we have observed that dietary supplementation with nutraceuticals that are joint protective may exert their influence via shifts in the gut microbiome. Although role of the microbiome in osteoarthritis is an area of intense study, no clear mechanism of action has been determined. Increased understanding of how the two factors interact may provide mechanistic insight into osteoarthritis and lead to disease modifying treatments.

Microbial evolution and ecological opportunity in the gut environment.

Recent genomic and metagenomic studies have highlighted the presence of rapidly evolving microbial populations in the human gut. However, despite the fundamental implications of this intuitive finding for both basic and applied gut microbiome research, very little is known about the mode, tempo and potential functional consequences of microbial evolution in the guts of individual human hosts over a lifetime. Here I assess the potential relevance of ecological opportunity to bacterial adaptation, colonization and persistence in the neonate and germ-free mammalian gut environment as well as over the course of an individual lifetime using data emerging from mouse models as well as human studies to provide examples where possible. I then briefly outline how the continued development and application of experimental evolution approaches coupled to genomic and metagenomic analysis is essential to disentangling drift from selection and identifying specific drivers of evolution in the gut microbiome within and between individual human hosts and populations.

Dysbiosis Modulates Ocular Surface Inflammatory Response to Liposaccharide.

PurposeThe purpose of this study was to investigate the inflammatory response of cornea and conjunctiva to topically applied lipopolysaccharide (LPS) in mice with and without antibiotic (antibiotic cocktail, ABX) induced dysbiosis.MethodsDysbiosis was induced by oral antibiotics for 14 days in a group of conventional female C57BL/6J (B6) mice. 16S rRNA sequencing investigated microbiome composition. Intestinal microbiome differences were assessed using 16S rRNA sequencing of fecal pellet DNA. Blood was collected after euthanasia. CD86 expression in draining nodes was examined by flow cytometry. At day 15, a single dose of LPS or vehicle was topically applied to ABX and naïve mice. Corneal epithelium and conjunctiva were obtained after 4 hours and processed for gene expression analysis. A separate group of germ-free (GF) B6 mice was also topically challenged with LPS.ResultsAntibiotic treatment significantly decreased intestinal diversity and increased serum levels of LPS. This was accompanied by a significant increase in CD86+MHC II+CD11c+CD11b+ cells in draining nodes. Compared to vehicle, topically applied LPS increased IL-1β, TNF-α, and CXCL10 mRNA transcripts in cornea and IL-1β, TNF-α, and CXCL10 in the conjunctiva in conventional and antibiotic-treated groups. However, there was higher TNF-α, CXCL10, and IL-12 expression in the cornea of LPS-treated ABX mice compared to LPS-treated mice with intact microbiota. LPS stimulation on GF conjunctiva mirrored the results in ABX mice, although greater IL-12 and IFN-γ expression was observed in GF conjunctiva compared to conventional LPS-treated mice.ConclusionsAcute depletion of commensals through antibiotics or germ-free environment worsens the inflammatory response to LPS.

Interleukin 22 disrupts pancreatic function in newborn mice expressing IL-23

Neonatal inflammatory diseases are associated with severe morbidity, but the inflammatory factors underlying them and their potential effector mechanisms are poorly defined. Here we show that necrotizing enterocolitis in neonate mice is accompanied by elevation of IL-23 and IL-22 and decreased production of pancreatic enzymes. These phenotypes are mirrored in neonate mice overexpressing IL-23 in CX3CR1+ myeloid cells or in keratinocytes. The mice fail to grow and die prematurely, displaying systemic inflammation, nutrient malabsorption and decreased expression of intestinal and pancreatic genes mediating digestion and absorption of carbohydrates, proteins, and lipids. Germ-free environment improves, and genetic ablation of IL-22 restores normal growth in mice overexpressing IL-23. Mechanistically, IL-22 acts directly at the level of pancreatic acinar cells to decrease expression of the pancreas associated transcription factor 1a (PTF1a). These results show that augmented production of IL-23 and IL-22 in early life has a negative impact on pancreatic enzyme secretion and food absorption.

Host-Encoded Sensors of Bacteria: Our Windows into the Microbial World.

Bacterial pathogens can be very efficient at causing disease and are the cause of some of the worst epidemics that have affected humanity. However, most infections are prevented by the actions of our immune system. Immune activation depends on the rapid detection of bacteria by a diverse family of sensory proteins known as pattern recognition receptors. These receptors detect conserved features of bacteria that are not found in humans but are often necessary for survival within the host or environment. In this review, we discuss the strategies used by pattern recognition receptors to detect bacteria and their products. We also discuss emerging evidence that some pattern recognition receptors can be activated by bacterial pathogens specifically, through the surveillance of host activities that are commonly targeted by virulence factors. This collection of surveillance mechanisms provides an interconnected network of defense, which is important to maintain the germ-free environment of the inner organs of humans and other multicellular organisms.

Transcriptional Response to Intermittent Hypoxia in the Heart of Germ‐Free Mice

Accumulating evidence has demonstrated that the gut microbiota plays a critical role in health and disease and regulates the organismal response to environmental stresses. For example, we have found that intermittent hypoxia/hypercapnia (IHH), a condition mimicking obstructive sleep apnea in humans, induced significant alterations in gut microbiome/metabolism and enhanced atherogenesis and progression in two atherosclerosis mouse models (i.e., the LDLR−/− and ApoE−/− mice). Indeed, with a joint microbiome and metabolome analysis, we have determined that IHH‐induced remarkable compositional changes in both microbial (>10%, most remarkably in Clostridia) and metabolites (>22%) in the gut of LDLR−/− mice, which are tightly correlated with a significantly enhanced atherosclerotic lesion formation in pulmonary arteries and aorta. In the current project, in order to further understand the role of the microbiome in the cardiovascular response to hypoxia and/or hypercapnia, we developed a set up that allows the treatment of experimental mice with different levels of O2 and/or CO2 in a germ‐free environment. Using this experimental set up, we studied the impact of the microbiome on transcriptional response to intermittent hypoxia (IH) in the left ventricle of mouse heart. We identified modest but significant changes in gene expression in both conventionally reared and germ‐free mice under IH condition. We found 52 significantly altered genes (19 up‐regulated and 33 down‐regulated) in the conventional mice and 76 significantly altered genes (24 upregulated and 52 down‐regulated) in the germ‐free mice. In addition, we found that these significantly altered genes play important roles in the development and function of cardiomyocytes and endothelial cells. Interestingly, there were only 15 genes (2 up‐regulated and 13 down‐regulated) that were commonly altered in both conventional and germ‐free mice, representing 28% changes in conventional mice and 20% changes in germ‐free mice, respectively. In summary, we discovered that IH induces distinct transcriptional alterations between conventional and germ‐free mice, which demonstrates that microbiome plays a critical role in the transcriptional response to IH in the left ventricle of mouse heart.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Gut microbiota in liver disease: too much is harmful, nothing at all is not helpful either.

The intestinal microbiome plays a major role in the pathogenesis of liver disease, with a hallmark event being dysbiosis, or an imbalance of pathobionts and beneficial bacteria with the associated deleterious effects on their host. Reducing the number of intestinal bacteria with antibiotic treatment is generally advantageous in experimental liver diseases. Complete absence of intestinal microbiota as in germ-free rodents can be protective in autoimmune hepatitis and hepatic tumors induced by chemicals, or it can exacerbate disease as in acute toxic liver injury and liver fibrosis/cirrhosis. In alcoholic liver disease, nonalcoholic fatty liver disease, and autoimmune cholangiopathies, germ-free status can be associated with worsened or improved hepatic phenotype depending on the experimental model and type of rodent. Some of the unexpected outcomes can be explained by the limitations of rodents raised in a germ-free environment including a deficient immune system and an altered metabolism of lipids, cholesterol, xenobiotics/toxins, and bile acids. Given these limitations and to advance understanding of the interactions between host and intestinal microbiota, simplified model systems such as humanized gnotobiotic mice, or gnotobiotic mice monoassociated with a single bacterial strain or colonized with a defined set of microbes, are unique and useful models for investigation of liver disease in a complex ecosystem.