Immune homeostasis,dysbiosis and therapeutic modulation of the gut microbiota

The distal gut harbours ∼10¹³ bacteria, representing the most densely populated ecosystem known. The functional diversity expressed by these communities is enormous and relatively unexplored. The past decade of research has unveiled the profound influence that the resident microbial populations bestow to host immunity and metabolism. The evolution of these communities from birth generates a highly adapted and highly personalized microbiota that is stable in healthy individuals. Immune homeostasis is achieved and maintained due in part to the extensive interplay between the gut microbiota and host mucosal immune system. Imbalances of gut microbiota may lead to a number of pathologies such as obesity, type I and type II diabetes, inflammatory bowel disease (IBD), colorectal cancer (CRC) and inflammaging/immunosenscence in the elderly. In-depth understanding of the underlying mechanisms that control homeostasis and dysbiosis of the gut microbiota represents an important step in our ability to reliably modulate the gut microbiota with positive clinical outcomes. The potential of microbiome-based therapeutics to treat epidemic human disease is of great interest. New therapeutic paradigms, including second-generation personalized probiotics, prebiotics, narrow spectrum antibiotic treatment and faecal microbiome transplantation, may provide safer and natural alternatives to traditional clinical interventions for chronic diseases. This review discusses host–microbiota homeostasis, consequences of its perturbation and the associated challenges in therapeutic developments that lie ahead.     

Age-associated inflammation inhibits epidermal stem cell function

Altered stem cell homeostasis is linked to organismal aging. However, the mechanisms involved remain poorly understood. Here we report novel alterations in hair follicle stem cells during skin aging, including increased numbers, decreased function, and an inability to tolerate stress. Performing high-throughput RNA sequencing on aging stem cells, cytokine arrays, and functional assays, we identify an age-associated imbalance in epidermal Jak–Stat signaling that inhibits stem cell function. Collectively, this study reveals a role for the aging epidermis in the disruption of cytokine and stem cell homeostasis, suggesting that stem cell decline during aging may be part of broader tumor-suppressive mechanisms.     

From pathogens to microbiota: How Drosophila intestinal stem cells react to gut microbes

The intestine acts as one of the interfaces between an organism and its external environment. As the primary digestive organ, it is constantly exposed to a multitude of stresses as it processes and absorbs nutrients. Among these is the recurring damage induced by ingested pathogenic and commensal microorganisms. Both the bacterial activity and immune response itself can result in the loss of epithelial cells, which subsequently requires replacement. In the Drosophila midgut, this regenerative role is fulfilled by intestinal stem cells (ISCs). Microbes not only trigger cell loss and replacement, but also modify intestinal and whole organism physiology, thus modulating ISC activity. Regulation of ISCs is integrated through a complex network of signaling pathways initiated by other gut cell populations, including enterocytes, enteroblasts, enteroendocrine and visceral muscles cells. The gut also receives signals from circulating immune cells, the hemocytes, to properly respond against infection. This review summarizes the types of gut microbes found in Drosophila, mechanisms for their elimination, and provides an integrated view of the signaling pathways that regulate tissue renewal in the midgut.     

Pre-treatment with IL-1β enhances the efficacy of MSC transplantation in DSS-induced colitis

Mesenchymal stem cells (MSCs) have been used experimentally for treating inflammatory disorders, partly due to their immunosuppressive properties. Although interleukin-1β (IL-1β) is one of the most important inflammatory mediators, growing evidence indicates that IL-1β signaling elicits the immunosuppressive properties of MSCs. However, it remains unclear how IL-1β signaling accomplishes this activity. Here, we focus on the therapeutic efficacy of IL-1β-primed MSCs in the dextran sulfate sodium (DSS)-induced colitis model, in addition to the underlining mechanisms. We first found that IL-1β-primed MSCs, without any observable phenotype change in vitro, significantly attenuated the development of DSS-induced murine colitis. Moreover, IL-1β-primed MSCs modulated the balance of immune cells in the spleen and the mesenteric lymph nodes (MLNs) through elevating cyclooxygenase-2 (COX-2), IL-6 and IL-8 expression and influencing the polarization of peritoneal macrophages. Importantly, IL-1β-primed MSCs possessed an enhanced ability to migrate to the inflammatory site of the gut via upregulation of chemokine receptor type 4 (CXCR4) expression. In summary, IL-1β-primed MSCs have improved efficacy in treating DSS-induced colitis, which at least partly depends on their increased immunosuppressive capacities and enhanced migration ability.     

Probiotics in inflammatory bowel disease: Pathophysiological background and clinical applications

Ulcerative colitis and Crohn’s disease, collectively termed the inflammatory bowel diseases (IBD), are chronic inflammatory disorders of the gastrointestinal tract. A “dysbiotic” relationship between the commensal gut flora and the intestinal mucosa-associated immune system has been at the core of the pathogenesis of these conditions. Probiotics are “good bacteria” with the ability to benefit the health of the host and their therapeutic application has been studied in IBD. The theoretical basis for such utilization relies upon the ability of probiotic microorganisms to interfere with the dysregulated homeostasis that takes place in IBD and restore the immune-bacterial interaction at the intestinal mucosa. Proposed mechanisms of action include the reconstitution of altered flora composition, enhancement of the integrity of the epithelial barrier, promotion of tolerogenic action by dendritic cells, strengthening of the defensive mechanisms of the innate immunity, and the suppression of pro-inflammatory adaptive immune responses. Despite this abundance of supporting experimental evidence, clinical application of probiotics in IBD has been disappointing. Possible explanations for such discrepancy include the great diversity of microorganisms that fall under the definition of probiotics, the lack of standardization of dosages and administration schemes, the heterogeneity between clinical trials, and the inclusion in the treatment arms of patients with a large variety of clinical phenotypes. Addressing these important issues will be critical for the optimal usage of probiotic-based therapies for patients with IBD.     

Cytoskeletal Regulation of Epithelial Barrier Function During Inflammation

Increased epithelial permeability is a common and important consequence of mucosal inflammation that results in perturbed body homeostasis and enhanced exposure to external pathogens. The integrity and barrier properties of epithelial layers are regulated by specialized adhesive plasma membrane structures known as intercellular junctions. It is generally believed that inflammatory stimuli increase transepithelial permeability by inducing junctional disassembly. This review highlights molecular events that lead to disruption of epithelial junctions during inflammation. We specifically focus on key mechanisms of junctional regulation that are dependent on reorganization of the perijunctional F-actin cytoskeleton. We discuss critical roles of myosin-II–dependent contractility and actin filament turnover in remodeling of the F-actin cytoskeleton that drive disruption of epithelial barriers under different inflammatory conditions. Finally, we highlight signaling pathways induced by inflammatory mediators that regulate reorganization of actin filaments and junctional disassembly in mucosal epithelia.The epithelium plays an important role in inflammation by serving as an interface between invading pathogens and the immune system of the host. Under physiological conditions, polarized epithelia form a protective barrier that allows regulated paracellular fluxes of solutes and nutrients as well as antigen sampling and surveillance by mucosal immune cells. However, during inflammation, this protective mechanism becomes compromised by various stimuli that originate on both sides of the epithelial barrier. On the apical (luminal) side, invading pathogenic microorganisms increase epithelial permeability to gain access into host tissue. The pathogens release a variety of epithelial barrier-disrupting agents that include pore-forming toxins, cytoskeleton-modifying proteins, and bacterial lipopolysaccharide (LPS). On the basal (tissue) side of the epithelial layer, activated immune cells also induce barrier disruption to facilitate their movement to sites of pathogen invasion. Mucosal immune cells increase epithelial permeability by secreting proinflammatory cytokines such as interferon (IFN) γ, tumor necrosis (TNF) α, and interleukin (IL)−1β or by releasing proteases and reactive oxygen species (ROS). As a result, mucosal inflammation commonly leads to sustained epithelial barrier compromise, which increases body exposure to external noxious agents, thereby further exaggerating the inflammatory response. Consequently it is believed that decreasing epithelial permeability may have beneficial effects by limiting inflammatory responses. Thus, understanding mechanisms that control the epithelial barrier disruption is important in identifying novel molecular targets for pharmacological modulation of mucosal inflammation.Properties of the epithelial barrier are regulated by specialized plasma membrane structures referred to as apical junctions. These structures are composed of adhesive and scaffolding proteins that are anchored into different cytoskeletal structures such as actin filaments, intermediate filaments, and microtubules. During inflammation, it is known that reorganizations of apical junctions mediate epithelial barrier dysfunction. Mounting evidence suggests that the actin cytoskeleton plays a pivotal role in regulating junctional integrity and remodeling under physiological and pathological states.In this review, we will discuss the role of actin filaments in the regulation of epithelial barrier integrity and its breakdown during inflammation. How inflammatory processes in different mucosal tissues induce remodeling of junction-associated filaments (F) actin and alter structure and barrier properties of epithelial cell-cell adhesions will be discussed. While epithelial junctions will be the major focus of this review, we will occasionally refer to examples of junctional regulation in the vascular endothelium. Likewise, we limit the discussion to simple columnar epithelia and exclude complex stratified epithelia such as the epidermis. Finally, we specifically focus on the role of actin filaments in maintenance and disassembly of epithelial junctions without discussing junctional reassembly during epithelial wound healing.     

Pulmonary Aspergillus fumigatus infection in rats affects gastrointestinal homeostasis

Microbiota inhabiting mucosal tissues is involved in maintenance of their immune homeostasis. Growing body of evidence indicate that dysbiosis in gut influence immune responses at distal sites including lungs. There are also reports concerning gut involvement with pulmonary injury/inflammation in settings of respiratory viral and bacterial infections. The impact of infections with other microorganisms on gut homeostasis is not explored. In this study, the rat model of sublethal pulmonary infection with Aspergillus fumigatus was used to investigate the effect of fungal respiratory infection on gut immune-mediated homeostasis. Signs of intestinal damage, intestinal and gut-draining lymphoid tissue cytokine responses and gut bacterial microbiota diversity were examined. Intestinal injury, inflammatory cell infiltration, as well as increased levels of intestinal interferon-γ (IFN-γ) and interleukin-17 (IL-17) (as opposed to unchanged levels of anti-inflammatory cytokine IL-10) during the two-week period depict intestinal inflammation in rats with pulmonary A. fumigatus infection. It could not be ascribed to the fungus as it was not detected in the intestine of infected rats. Increased production of pro-inflammatory cytokines by major gut-draining mesenteric lymph nodes point to these lymphoid organs as places of generation of cytokine-producing cells. No changes in spleen or systemic cytokine responses was observed, showing lack of the effects of pulmonary A. fumigatus infection outside mucosal immune system. Drop of intestinal bacterial microbiota diversity (disappearance of several bacterial bands) was noted early in infection with normalization starting from day seven. From day three, appearance of new bacterial bands (unique to infected individuals, not present in controls) was seen, and some of them are pathogens. Alterations in intestinal bacterial community might have affected intestinal immune tolerance contributing to inflammation. Disruption of gut homeostasis during pulmonary infection might render gastrointestinal tract more susceptible to variety of physiological and pathological stimuli. Data which showed for the first time gut involvement with pulmonary infection with A. fumigatus provide the baseline for future studies of the impact of fungal lung infections to gut homeostasis, particularly in individuals susceptible to these infections.     

Gut microbiota: Role in pathogen colonization,immune responses,and inflammatory disease

The intestinal tract of mammals is colonized by a large number of microorganisms including trillions of bacteria that are referred to collectively as the gut microbiota. These indigenous microorganisms have co-evolved with the host in a symbiotic relationship. In addition to metabolic benefits, symbiotic bacteria provide the host with several functions that promote immune homeostasis, immune responses, and protection against pathogen colonization. The ability of symbiotic bacteria to inhibit pathogen colonization is mediated via several mechanisms including direct killing, competition for limited nutrients, and enhancement of immune responses. Pathogens have evolved strategies to promote their replication in the presence of the gut microbiota. Perturbation of the gut microbiota structure by environmental and genetic factors increases the risk of pathogen infection, promotes the overgrowth of harmful pathobionts, and the development of inflammatory disease. Understanding the interaction of the microbiota with pathogens and the immune system will provide critical insight into the pathogenesis of disease and the development of strategies to prevent and treat inflammatory disease.     

Roles of B-1 and B-2 cells in innate and acquired IgA-mediated immunity

Summary: The gut harbors an extremely dense and complex community of microorganisms that are in constant dialog with our immune cells. The gut bacteria provide strong selective pressure to the host to evolve innate and adaptive immune responses required for the maintenance of local and systemic homeostasis. One of the most conspicuous responses of the gut immune system following microbial colonization is the production of immunoglobulin A (IgA). In this review, we discuss the roles of B-1 and B-2 cells in IgA-mediated immunity and present an updated view for the sites and mechanisms of IgA synthesis in the gut. We summarize the role of secretory IgAs for regulation of microbial communities and provide clues as to how the gut microbiota contributes to the development of the gut-associated lymphoid tissues.     

Characteristics of gut microbiota in representative mice strains: Implications for biological research

BackgroundExperimental animals are used to study physiological phenomena, pathological mechanisms, and disease prevention. The gut microbiome is known as a potential confounding factor for inconsistent data from preclinical studies. Although many gut microbiome studies have been conducted in recent decades, few have focused on gut microbiota fluctuation among representative mouse strains.MethodsA range of frequently used mouse strains were selected from 34 isolation packages representing disease‐related animal (DRA), immunity defect animal (IDA), or gene‐editing animal (GEA) from the BALB/c and C57BL/6J backgrounds together with normal mice, and their microbial genomic DNA were isolated from mouse feces to sequence for the exploration of gut microbiota.ResultsMouse background strain, classification, introduced source, introduced year, and reproduction type significantly affected the gut microbiota structure (p < 0.001 for all parameters), with background strain contributing the greatest influence (R ² = 0.237). In normal groups, distinct gut microbiota types existed in different mouse strains. Sixty‐four core operational taxonomic units were obtained from normal mice, and 12 belonged to Lactobacillus. Interestingly, the gut microbiota in C57BL/6J was more stable than that in BALB/c mice. Furthermore, the gut microbiota in the IDA, GEA, and DRA groups significantly differed from that in normal groups (p < 0.001 for all). Compared with the normal group, there was a significantly higher Chao1 and Shannon index (p < 0.001 for all) in the IDA, GEA, and DRA groups. Markedly changed classes occurred with Firmicutes and Bacteroidetes. The abundances of Helicobacter, Blautia, Enterobacter, Bacillus, Clostridioides, Paenibacillus, and Clostridiales all significantly decreased in the IDA, GEA, and DRA groups, whereas those of Saccharimonas, Rikenella, and Odoribacter all significantly increased.     

Targeting Mitochondria-Derived Reactive Oxygen Species to Reduce Epithelial Barrier Dysfunction and Colitis

Epithelial permeability is often increased in inflammatory bowel diseases. We hypothesized that perturbed mitochondrial function would cause barrier dysfunction and hence epithelial mitochondria could be targeted to treat intestinal inflammation. Mitochondrial dysfunction was induced in human colon-derived epithelial cell lines or colonic biopsy specimens using dinitrophenol, and barrier function was assessed by transepithelial flux of Escherichia coli with or without mitochondria-targeted antioxidant (MTA) cotreatment. The impact of mitochondria-targeted antioxidants on gut permeability and dextran sodium sulfate (DSS)–induced colitis in mice was tested. Mitochondrial superoxide evoked by dinitrophenol elicited significant internalization and translocation of E. coli across epithelia and control colonic biopsy specimens, which was more striking in Crohn’s disease biopsy specimens; the mitochondria-targeted antioxidant, MitoTEMPO, inhibited these barrier defects. Increased gut permeability and reduced epithelial mitochondrial voltage-dependent anion channel expression were observed 3 days after DSS. These changes and the severity of DSS-colitis were reduced by MitoTEMPO treatment. In vitro DSS-stimulated IL-8 production by epithelia was reduced by MitoTEMPO. Metabolic stress evokes significant penetration of commensal bacteria across the epithelium, which is mediated by mitochondria-derived superoxide acting as a signaling, not a cytotoxic, molecule. MitoTEMPO inhibited this barrier dysfunction and suppressed colitis in DSS-colitis, likely via enhancing barrier function and inhibiting proinflammatory cytokine production. These novel findings support consideration of MTAs in the maintenance of epithelial barrier function and the management of inflammatory bowel diseases.The mammalian gut harbors an immense and diverse microbiota, and host-bacteria interactions are key determinants of digestive health and general well-being.^1,2 While providing distinct benefits to the host (eg, vitamin synthesis), commensal bacteria that cross the epithelium and enter the mucosa have the potential to provoke inflammation, and movement into the circulation can result in sepsis and death. Consequently, the barrier function of the epithelium, both intrinsic (eg, epithelial tight junctions and cell membranes) and extrinsic (eg, mucus) elements, is a critical component of innate defense. Thus, the commensal bacteria-host interaction pivots on the gut epithelium as a point of first contact and knowledge of the dynamic nature of this interface is important to understanding normal intestinal function and disease.³The epithelial barrier is not static; rather, it is highly dynamic and tightly regulated to maintain gut homeostasis. However, uncontrolled or prolonged increases in gut permeability have the potential to initiate or exaggerate enteric inflammatory disease. For example, the consensus on the etiology of inflammatory bowel disease (IBD; Crohn’s disease or ulcerative colitis) is that disease develops because of an inappropriate immune response to the gut microbiota in a genetically susceptible individual.⁴ This suggests a barrier defect because microbes, or their products, in the gut lumen must access the mucosal immune system by crossing the epithelial cell layer. Although increased epithelial permeability as a primary cause of IBD remains unproved,³ the leaky gut hypothesis⁵ is supported by observations in experimental models of colitis^3,6,7 and increased epithelial permeability has been repeatedly demonstrated in active IBD.^8,9 Therefore, the ability to enhance the barrier property of the epithelium would be of value in ameliorating enteric inflammatory disease.Given that control of epithelial permeability (apical junction complex formation and transcellular permeation) is energy dependent, mitochondria should be essential for appropriate regulation of barrier function. Indeed, factors such as infection, nonsteroidal anti-inflammatory drugs, and smoking, which can contribute to the pathophysiological characteristics of Crohn’s disease, also perturb mitochondrial function.^10–12 Furthermore, structurally abnormal mitochondria have been observed in tissue from patients with gut inflammation,¹³ in animal models of gut disease,¹⁴ and in epithelial monolayers treated with bacterial toxins or low-grade pathogens.¹⁵ However, despite these findings and the keen interest in mitochondria in the pathophysiological characteristics of neuromuscular disease, diabetes, and obesity,^16–18 there are limited data on the role of mitochondria in colitis.We have previously shown that model epithelia treated with dinitrophenol (DNP) to uncouple oxidative phosphorylation display decreased barrier function, as characterized by lower transepithelial resistance (TER) indicative of increased paracellular permeability, and the internalization and translocation of noninvasive, nontoxigenic commensal E. coli.^13,19 The latter is particularly intriguing for two reasons: entry of commensal bacteria into the enterocyte likely represents a threat and the enterocytes’ response (eg, IL-8 synthesis) could promote inflammation; and the contribution of transcellular permeability to a barrier defect is not well understood and needs to be comprehensively assessed and fully integrated to any consideration of the epithelial barrier and innate defense. Thus, the current study was designed to uncover the mechanism(s) underlying the increased internalization and translocation of commensal Escherichia coli across gut epithelia, which occurs as a consequence of metabolic stress evoked by targeted perturbation of mitochondrial function.     

Apoptosis of leukocytes triggered by acute DNA damage promotes lymphoma formation

Apoptosis triggered by p53 upon DNA damage secures removal of cells with compromised genomes, and is thought to prevent tumorigenesis. In contrast, we provide evidence that p53-induced apoptosis can actively drive tumor formation. Mice defective in p53-induced apoptosis due to loss of its proapoptotic target gene, puma, resist γ-irradiation (IR)-induced lymphomagenesis. In wild-type animals, repeated irradiation injury-induced expansion of hematopoietic stem/progenitor cells (HSCs) leads to lymphoma formation. Puma^−/− HSCs, protected from IR-induced cell death, show reduced compensatory proliferation and replication stress-associated DNA damage, and fail to form thymic lymphomas, demonstrating that the maintenance of stem/progenitor cell homeostasis is critical to prevent IR-induced tumorigenesis.     

Vγ2Vδ2 T-cell co-stimulation increases natural killer cell killing of monocyte-derived dendritic cells

Interactions between natural killer (NK) cells and dendritic cells (DC) affect maturation and function of both cell populations, including NK cell killing of DC (editing), which is important for controlling the quality of immune responses. We also know that antigen-stimulated Vγ2Vδ2 T cells co-stimulate NK cells via 4-1BB to enhance the killing of tumour cell lines but we do not know what regulates 4-1BB expression or whether other NK effector functions including DC killing, might also be influenced by NK–γδ T-cell cross-talk. Here we show that antigen-stimulated γδ T cells co-stimulate NK cells through inducible T-cell co-stimulator (ICOS)– ICOS ligand (ICOSL) and this signal increases NK cell killing of autologous DC. Effects of NK–γδ T-cell co-culture, which could be reproduced with soluble ICOS-Fc fusion protein, included increased CD69 and 4-1BB expression, interferon-γ, tumour necrosis factor-α, macrophage inflammatory protein-1β, I-309, RANTES and sFas ligand production, as well as elevated mRNA levels for co-stimulatory receptors OX40 (TNFRSF4) and GITR (TNFRSF18). Hence, ICOS–ICOSL co-stimulation of NK by Vγ2Vδ2 T cells had broad effects on NK phenotype and effector functions. The NK–γδ T-cell cross-talk links innate and antigen-specific lymphocyte responses in the control of cytotoxic effector function and DC killing.     

Gut-microbiota interactions in non-mammals: What can we learn from Drosophila?

Millions of people suffer from inflammatory diseases of the intestine, some of them potentiating gastrointestinal cancer. These gut-associated pathologies arise from imbalanced interactions between the host gut epithelia and resident or ingested microbes, interactions that are still poorly understood at the molecular level. Drosophila has been a very powerful model to study development and diseases. Its relatively simple tissue organization and sophisticated genetics are some of the advantages of using it as an experimental model to dissect gut-microbe interactions. Recent progress made in various research fields such as Drosophila microbiota composition, gut epithelium structure or gut immune reactions led us to believe that Drosophila is becoming an ad hoc model system to dissect the mechanisms that cooperate to maintain intestinal homeostasis in higher eukaryotes. It further may help us understand how an alteration of these finely tuned processes precipitates the inflammatory processes found in some inflammatory bowel diseases.     

Amino acids 89–96 of Salmonella typhimurium flagellin represent the major domain responsible for TLR5-independent adjuvanticity in the humoral immune response

Toll-like receptor 5 (TLR5) signaling in response to flagellin is dispensable for inducing humoral immunity, but alterations of aa 89–96, the TLR5 binding site, significantly reduced the adjuvanticity of flagellin. These observations indicate that the underlying mechanism remains incompletely understood. Here, we found that the native form of Salmonella typhimurium aa 89–96-mutant flagellin extracted from flagella retains some TLR5 recognition activity, indicating that aa 89–96 is the primary, but not the only site that imparts TLR5 activity. Additionally, this mutation impaired the production of IL-1β and IL-18. Using TLR5KO mice, we found that aa 89–96 is critical for the humoral adjuvant effect, but this effect was independent of TLR5 activation triggered by this region of flagellin. In summary, our findings suggest that aa 89–96 of flagellin is not only the crucial site responsible for TLR5 recognition, but is also important for humoral immune adjuvanticity through a TLR5-independent pathway.     

Dietary β-carotene and ultraviolet-induced immunosuppression

Ultraviolet (UV)-induced immunosuppression is a critical step in UV carcinogenesis, permitting tumour outgrowth. We investigated the effect of dietary β-carotene on UV suppression of contact hypersensitivity (CHS) to trinitrochlorobenzene (TNCB) in BALB/c mice. Mice were fed for 10–16 weeks chow alone or supplemented with 1% β-carotene or placebo as beadlets. Serum β-carotene was detectable by high performance liquid chromatography (HPLC) analysis only in β-carotene-fed mice (2.06 ± 0.15 μg/ml). Serum retinol was 0.22–0.27 μg/ml in all three groups. Mice (n = 41/dietary group) were irradiated with 0, 4.5, 9 or 18 kJ/m² of UVB and the CHS response was measured. Decreased CHS responses were observed in all UV-irradiated groups compared with unirradiated controls. UV dose–responses for suppression of CHS derived by first-order regression analyses of plots of percentage suppression of CHS as a function of log₁₀UV dose showed significant slopes (P < 0.02) for all three dietary groups and similar residual variances between groups, P > 0.05. The UV dose for 50% suppression of CHS was 6.3 kJ/m² for control, 6.4 kJ/m² for placebo, and 5.5 kJ/m² for β-carotene-fed mice. No significant differences in slopes or elevations between UV dose–responses were observed, P > 0.05. Skin levels of the initiator of UV-induced immunosuppression, cis urocanic acid, were determined by HPLC in mice given 0 or 9 kJ/m² of UV (n = 28/dietary group). No significant differences were observed between dietary groups (range 35.2–41.1 ng/mg skin, P > 0.15) We conclude feeding β-carotene to BALB/c mice does not alter susceptibility to UV immune suppression, in contrast to human studies.     

Impact of peroxisome proliferator-activated receptor γ on angiotensin II type 1 receptor-mediated insulin sensitivity,vascular inflammation and atherogenesis in hypercholesterolemic mice

Introduction

The angiotensin II type 1 receptor (AT1R) and the peroxisome proliferator-activated receptor γ (PPARγ) have been implicated in the pathogenesis of atherosclerosis. A number of studies have reported that AT1R inhibition or genetic AT1R disruption and PPARγ activation inhibit vascular inflammation and improve glucose and lipid metabolism, underscoring a molecular interaction of AT1R and PPARγ. We here analyzed the hypothesis that vasculoprotective anti-inflammatory and metabolic effects of AT1R inhibition are mediated by PPARγ.

Material and methods

Female ApoE^–/–/AT1R^–/– mice were fedwith a high-fat and cholesterol-rich diet and received continuous treatment with the selective PPARγ antagonist GW9662 or vehicle at a rate of 700 ng/kg/min for 4 weeks using subcutaneously implanted osmotic mini-pumps. Additionally, one group of female ApoE^–/– mice served as a control group. After treatment for 4 weeks mice were sacrificed and read-outs (plaque development, vascular inflammation and insulinsensitivity) were performed.

Results

Using AT1R deficient ApoE^–/– mice (ApoE^–/–/AT1R^–/– mice) we found decreased cholesterol-induced endothelial dysfunction and atherogenesis compared to ApoE^–/– mice. Inhibition of PPARγ by application of the specific PPARγ antagonist GW9662 significantly abolished the anti-atherogenic effects of AT1R deficiency in ApoE^–/–/AT1R^–/– mice (plaque area as % of control: ApoE^–/–: 39 ±5%; ApoE^–/–/AT1R^–/–: 17 ±7%, p = 0.044 vs. ApoE^–/–; ApoE^–/–/AT1R^–/– + GW9662: 31 ±8%, p = 0.047 vs. ApoE^–/–/AT1R^–/–). Focusing on IL6 as a pro-inflammatory humoral marker we detected significantly increased IL-6 levels in GW9662-treated animals (IL-6 in pg/ml: ApoE^–/–: 230 ±16; ApoE^–/–/AT1R^–/–: 117 ±20, p = 0.01 vs. ApoE^–/–; ApoE^–/–/AT1R^–/– + GW9662: 199 ±20, p = 0.01 vs. ApoE^–/–/AT1R^–/–), while the anti-inflammatory marker IL-10 was significantly reduced after PPARγ inhibition in GW9662 animals (IL-10 in pg/ml: ApoE^–/–: 18 ±4; ApoE^–/–/AT1R^–/–: 55 ±12, p = 0.03 vs. ApoE^–/–; ApoE^–/–/AT1R^–/– + GW9662: 19 ±4, p = 0.03 vs. ApoE^–/–/AT1R^–/–). Metabolic parameters of glucose homeostasis (glucose and insulin tolerance test) were significantly deteriorated in ApoE^–/–/AT1R^–/– mice treated with GW9662 as compared to vehicle-treated ApoE^–/–/AT1R^–/– mice. Systolic blood pressure and plasma cholesterol levels were similar in all groups.

Conclusions

Genetic disruption of the AT1R attenuates atherosclerosis and improves endothelial function in an ApoE^–/– mouse model of hypercholesterolemia-induced atherosclerosis via PPARγ, indicating a significant role of PPARγ in reduced vascular inflammation, improvement of insulin sensitivity and atheroprotection of AT1R deficiency.     

Control of hair follicle cell fate by underlying mesenchyme through a CSL–Wnt5a–FoxN1 regulatory axis

Epithelial–mesenchymal interactions are key to skin morphogenesis and homeostasis. We report that maintenance of the hair follicle keratinocyte cell fate is defective in mice with mesenchymal deletion of the CSL/RBP-Jκ gene, the effector of “canonical” Notch signaling. Hair follicle reconstitution assays demonstrate that this can be attributed to an intrinsic defect of dermal papilla cells. Similar consequences on hair follicle differentiation result from deletion of Wnt5a, a specific dermal papilla signature gene that we found to be under direct Notch/CSL control in these cells. Functional rescue experiments establish Wnt5a as an essential downstream mediator of Notch–CSL signaling, impinging on expression in the keratinocyte compartment of FoxN1, a gene with a key hair follicle regulatory function. Thus, Notch/CSL signaling plays a unique function in control of hair follicle differentiation by the underlying mesenchyme, with Wnt5a signaling and FoxN1 as mediators.