Reprogramming of connexin landscape fosters fast gap junction intercellular communication in human papillomavirus-infected epithelia

Human papillomaviruses (HPVs) are highly prevalent commensal viruses that require epithelial stratification to complete their replicative cycle. While HPV infections are most often asymptomatic, certain HPV types can cause lesions, that are usually benign. In rare cases, these infections may progress to non-replicative viral cycles associated with high HPV oncogene expression promoting cell transformation, and eventually cancer when not cleared by host responses. While the consequences of HPV-induced transformation on keratinocytes have been extensively explored, the impact of viral replication on epithelial homeostasis remains largely unexplored. Gap junction intercellular communication (GJIC) is critical for stratified epithelium integrity and function. This process is ensured by a family of proteins named connexins (Cxs), including 8 isoforms that are expressed in stratified squamous epithelia. GJIC was reported to be impaired in HPV-transformed cells, which was attributed to the decreased expression of the Cx43 isoform. However, it remains unknown whether and how HPV replication might impact on the expression of Cx isoforms and GJIC in stratified squamous epithelia. To address this question, we have used 3D-epithelial cell cultures (3D-EpCs), the only model supporting the productive HPV life cycle. We report a transcriptional downregulation of most epithelial Cx isoforms except Cx45 in HPV-replicating epithelia. At the protein level, HPV replication results in a reduction of Cx43 expression while that of Cx45 increases and displays a topological shift toward the cell membrane. To quantify GJIC, we pioneered quantitative gap-fluorescence loss in photobleaching (FLIP) assay in 3D-EpCs, which allowed us to show that the reprogramming of Cx landscape in response to HPV replication translates into accelerated GJIC in living epithelia. Supporting the pathophysiological relevance of our observations, the HPV-associated Cx43 and Cx45 expression pattern was confirmed in human cervical biopsies harboring HPV. In conclusion, the reprogramming of Cx expression and distribution in HPV-replicating epithelia fosters accelerated GJIC, which may participate in epithelial homeostasis and host immunosurveillance

Multi-Tissue Characterization of GILZ Expression in Dendritic Cell Subsets at Steady State and in Inflammatory Contexts

Dendritic cells (DCs) are key players in the control of tolerance and immunity. Glucocorticoids (GCs) are known to regulate DC function by promoting their tolerogenic differentiation through the induction of inhibitory ligands, cytokines, and enzymes. The GC-induced effects in DCs were shown to critically depend on increased expression of the Glucocorticoid-Induced Leucine Zipper protein (GILZ). GILZ expression levels were further shown to control antigen-presenting cell function, as well as T-cell priming capacity of DCs. However, the pattern of GILZ expression in DC subsets across tissues remains poorly described, as well as the modulation of its expression levels in different pathological settings. To fill in this knowledge gap, we conducted an exhaustive analysis of GILZ relative expression levels in DC subsets from various tissues using multiparametric flow cytometry. This study was performed at steady state, in the context of acute as well as chronic skin inflammation, and in a model of cancer. Our results show the heterogeneity of GILZ expression among DC subsets as well as the complexity of its modulation, that varies in a cell subset- and context-specific manner. Considering the contribution of GILZ in the control of DC functions and its potential as an immune checkpoint in cancer settings, these results are of high relevance for optimal GILZ targeting in therapeutic strategies

Gut Microbiota Reshaped by Pectin Treatment Improves Liver Steatosis in Obese Mice

Pectin, a soluble fiber, improves non-alcoholic fatty-liver disease (NAFLD), but its mechanisms are unclear. We aimed to investigate the role of pectin-induced changes in intestinal microbiota (IM) in NAFLD. We recovered the IM from mice fed a high-fat diet, treated or not with pectin, to perform a fecal microbiota transfer (FMT). Mice fed a high-fat diet, which induces NAFLD, were treated with pectin or received a fecal microbiota transfer (FMT) from mice treated with pectin before (preventive FMT) or after (curative FMT) being fed a high-fat diet. Pectin prevented the development of NAFLD, induced browning of adipose tissue, and modified the IM without increasing the abundance of proteobacteria. Preventive FMT also induced browning of white adipose tissue but did not improve liver steatosis, in contrast to curative FMT, which induced an improvement in steatosis. This was associated with an increase in the concentration of short-chain fatty acids (SCFAs), in contrast to preventive FMT, which induced an increase in the concentration of branched SCFAs. Overall, we show that the effect of pectin may be partially mediated by gut bacteria

A new hemostatic agent composed of Zn2+-enriched Ca2+ alginate activates vascular endothelial cells in vitro and promotes tissue repair in vivo

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Modulation of the Bile Acid Enterohepatic Cycle by Intestinal Microbiota Alleviates Alcohol Liver Disease

Reshaping the intestinal microbiota by the ingestion of fiber, such as pectin, improves alcohol-induced liver lesions in mice by modulating bacterial metabolites, including indoles, as well as bile acids (BAs). In this context, we aimed to elucidate how oral supplementation of pectin affects BA metabolism in alcohol-challenged mice receiving feces from patients with alcoholic hepatitis. Pectin reduced alcohol liver disease. This beneficial effect correlated with lower BA levels in the plasma and liver but higher levels in the caecum, suggesting that pectin stimulated BA excretion. Pectin modified the overall BA composition, favoring an augmentation in the proportion of hydrophilic forms in the liver, plasma, and gut. This effect was linked to an imbalance between hydrophobic and hydrophilic (less toxic) BAs in the gut. Pectin induced the enrichment of intestinal bacteria harboring genes that encode BA-metabolizing enzymes. The modulation of BA content by pectin inhibited farnesoid X receptor signaling in the ileum and the subsequent upregulation of Cyp7a1 in the liver. Despite an increase in BA synthesis, pectin reduced BA serum levels by promoting their intestinal excretion. In conclusion, pectin alleviates alcohol liver disease by modifying the BA cycle through effects on the intestinal microbiota and enhanced BA excretion

DataSheet_1_P2X7 purinergic receptor plays a critical role in maintaining T-cell homeostasis and preventing lupus pathogenesis.docx

The severe lymphoproliferative and lupus diseases developed by MRL/lpr mice depend on interactions between the Faslpr mutation and MRL genetic background. Thus, the Faslpr mutation causes limited disease in C57BL/6 mice. We previously found that accumulating B220+ CD4–CD8– double negative (DN) T cells in MRL/lpr mice show defective P2X7 receptor ( P2X7)-induced cellular functions, suggesting that P2X7 contributes to T-cell homeostasis, along with Fas. Therefore, we generated a B6/lpr mouse strain (called B6/lpr-p2x7KO) carrying homozygous P2X7 knockout alleles. B6/lpr-p2x7KO mice accumulated high numbers of FasL-expressing B220+ DN T cells of CD45RBhighCD44high effector/memory CD8+ T-cell origin and developed severe lupus, characterized by leukocyte infiltration into the tissues, high levels of IgG anti-dsDNA and rheumatoid factor autoantibodies, and marked cytokine network dysregulation. B6/lpr-p2x7KO mice also exhibited a considerably reduced lifespan. P2X7 is therefore a novel regulator of T-cell homeostasis, of which cooperation with Fas is critical to prevent lymphoaccumulation and autoimmunity.</p

Dual effect of cardiac FKBP12.6 overexpression on excitation-contraction coupling and the incidence of ventricular arrhythmia depending on its expression level

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Bile acid-receptor TGR5 deficiency worsens liver injury in alcohol-fed mice by inducing intestinal microbiota dysbiosis

International audienceBackground & aims: Bile-acid metabolism and the intestinal microbiota are impaired in alcohol-related liver disease. Activation of the bile-acid receptor TGR5 (or GPBAR1) controls both biliary homeostasis and inflammatory processes. We examined the role of TGR5 in alcohol-induced liver injury in mice.Methods: We used TGR5-deficient (TGR5-KO) and wild-type (WT) female mice, fed alcohol or not, to study the involvement of liver macrophages, the intestinal microbiota (16S sequencing), and bile-acid profiles (high-performance liquid chromatography coupled to tandem mass spectrometry). Hepatic triglyceride accumulation and inflammatory processes were assessed in parallel.Results: TGR5 deficiency worsened liver injury, as shown by greater steatosis and inflammation than in WT mice. Isolation of liver macrophages from WT and TGR5-KO alcohol-fed mice showed that TGR5 deficiency did not increase the pro-inflammatory phenotype of liver macrophages but increased their recruitment to the liver. TGR5 deficiency induced dysbiosis, independently of alcohol intake, and transplantation of the TGR5-KO intestinal microbiota to WT mice was sufficient to worsen alcohol-induced liver inflammation. Secondary bile-acid levels were markedly lower in alcohol-fed TGR5-KO than normally fed WT and TGR5-KO mice. Consistent with these results, predictive analysis showed the abundance of bacterial genes involved in bile-acid transformation to be lower in alcohol-fed TGR5-KO than WT mice. This altered bile-acid profile may explain, in particular, why bile-acid synthesis was not repressed and inflammatory processes were exacerbated.Conclusions: A lack of TGR5 was associated with worsening of alcohol-induced liver injury, a phenotype mainly related to intestinal microbiota dysbiosis and an altered bile-acid profile, following the consumption of alcohol

Pivotal Role for Cxcr2 in Regulating Tumor-Associated Neutrophil in Breast Cancer.

Chemokines present in the tumor microenvironment are essential for the control of tumor progression. We show here that several ligands of the chemokine receptor Cxcr2 were up-regulated in the PyMT (polyoma middle T oncogene) model of breast cancer. Interestingly, the knock-down of Cxcr2 in PyMT animals led to an increased growth of the primary tumor and lung metastasis. The analysis of tumor content of PyMT-Cxcr2-/- animals highlighted an increased infiltration of tumor associated neutrophils (TANs), mirrored by a decreased recruitment of tumor associated macrophages (TAMs) compared to PyMT animals. Analysis of PyMT-Cxcr2-/- TANs revealed that they lost their killing ability compared to PyMT-Cxcr2+/+ TANs. The transcriptomic analysis of PyMT-Cxcr2-/- TANs showed that they had a more pronounced pro-tumor TAN2 profile compared to PyMT TANs. In particular, PyMT-Cxcr2-/- TANs displayed an up-regulation of the pathways involved in reactive oxygen species (ROS) production and angiogenesis and factors favoring metastasis, but reduced apoptosis. In summary, our data reveal that a lack of Cxcr2 provides TANs with pro-tumor effects

Microbiota tryptophan metabolism induces aryl hydrocarbon receptor activation and improves alcohol-induced liver injury

International audienceObjective: Chronic alcohol consumption is an important cause of liver-related deaths. Specific intestinal microbiota profiles are associated with susceptibility or resistance to alcoholic liver disease in both mice and humans. We aimed to identify the mechanisms by which targeting intestinal microbiota can improve alcohol-induced liver lesions.Design: We used human associated mice, a mouse model of alcoholic liver disease transplanted with the intestinal microbiota of alcoholic patients and used the prebiotic, pectin, to modulate the intestinal microbiota. Based on metabolomic analyses, we focused on microbiota tryptophan metabolites, which are ligands of the aryl hydrocarbon receptor (AhR). Involvement of the AhR pathway was assessed using both a pharmacological approach and AhR-deficient mice.Results: Pectin treatment modified the microbiome and metabolome in human microbiota-associated alcohol-fed mice, leading to a specific faecal signature. High production of bacterial tryptophan metabolites was associated with an improvement of liver injury. The AhR agonist Ficz (6-formylindolo (3,2-b) carbazole) reduced liver lesions, similarly to prebiotic treatment. Conversely, inactivation of the ahr gene in alcohol-fed AhR knock-out mice abrogated the beneficial effects of the prebiotic. Importantly, patients with severe alcoholic hepatitis have low levels of bacterial tryptophan derivatives that are AhR agonists.Conclusions: Improvement of alcoholic liver disease by targeting the intestinal microbiota involves the AhR pathway, which should be considered as a new therapeutic target