Pierre Lapaquette

Ileal lesions in Crohn's disease (CD) patients are colonized by pathogenic adherent-invasive Escherichia coli (AIEC) able to invade and to replicate within intestinal epithelial cells. Recent genome-wide association studies have highlighted the autophagy pathway as being associated with CD risk. In the present study we investigated whether defects in autophagy enhance replication of commensal and pathogenic Escherichia coli and CD-associated AIEC. We show that functional autophagy limits intracellular AIEC replication and that a subpopulation of the intracellular bacteria is located within LC3-positive autophagosomes. In IRGM and ATG16L1 deficient cells intracellular AIEC LF82 bacteria have enhanced replication. Surprisingly autophagy deficiency did not interfere with the ability of intracellular bacteria to survive and/or replicate for any other E. coli strains tested, including non-pathogenic, environmental, commensal, or pathogenic strains involved in gastro enteritis. Together these findings demonstrate a central role for autophagy restraining Adherent-Invasive E. coli strains associated with ileal CD. AIEC infection in patients with polymorphisms in autophagy genes may have a significant impact on the outcome of intestinal inflammation.

Ileal lesions in Crohn's disease (CD) patients are abnormally colonized by pathogenic adherentinvasive Escherichia coli (AIEC).AIEC bacteria are able to replicate within epithelial cells after lysis of the endocytic vacuole and within macrophages in a large vacuole.CD-associated polymorphisms in NOD2, ATG16L1 and IRGM affect bacterial autophagy, a crucial innate immunity mechanism.We previously determined that defects in autophagy impaired the ability of epithelial cells to control AIEC replication.AIEC behave differently within epithelial cells and macrophages and so we investigated the impact of defects in autophagy on AIEC intramacrophagic replication and proinflammatory cytokine response.AIEC bacteria induced the recruitment of the autophagy machinery at the site of phagocytosis, and functional autophagy limited AIEC intramacrophagic replication.Impaired ATG16L1, IRGM or NOD2 expression induced increased intramacrophagic AIEC and increased secretion of IL-6 and TNF-a in response to AIEC infection.In contrast, forced induction of autophagy decreased the numbers of intramacrophagic AIEC and pro-inflammatory cytokine release, even in a NOD2-deficient context.On the basis of our findings, we speculate that stimulating autophagy in CD patients would be a powerful therapeutic strategy to concomitantly restrain intracellular AIEC replication and slow down the inflammatory response.

Autophagy is an intracellular catabolic pathway essential for the recycling of proteins and larger substrates such as aggregates, apoptotic corpses, or long-lived and superfluous organelles whose accumulation could be toxic for cells. Because of its unique feature to engulf part of cytoplasm in double-membrane cup-shaped structures, which further fuses with lysosomes, autophagy is also involved in the elimination of host cell invaders and takes an active part of the innate and adaptive immune response. Its pivotal role in maintenance of the inflammatory balance makes dysfunctions of the autophagy process having important pathological consequences. Indeed, defects in autophagy are associated with a wide range of human diseases including metabolic disorders (diabetes and obesity), inflammatory bowel disease (IBD), and cancer. In this review, we will focus on interrelations that exist between inflammation and autophagy. We will discuss in particular how mediators of inflammation can regulate autophagy activity and, conversely, how autophagy shapes the inflammatory response. Impact of genetic polymorphisms in autophagy-related gene on inflammatory bowel disease will be also discussed.

Innate sensing of pathogens initiates inflammatory cytokine responses that need to be tightly controlled. We found here that after engagement of Toll-like receptors (TLRs) in myeloid cells, deficient sumoylation caused increased secretion of transcription factor NF-κB-dependent inflammatory cytokines and a massive type I interferon signature. In mice, diminished sumoylation conferred susceptibility to endotoxin shock and resistance to viral infection. Overproduction of several NF-κB-dependent inflammatory cytokines required expression of the type I interferon receptor, which identified type I interferon as a central sumoylation-controlled hub for inflammation. Mechanistically, the small ubiquitin-like modifier SUMO operated from a distal enhancer of the gene encoding interferon-β (Ifnb1) to silence both basal and stimulus-induced activity of the Ifnb1 promoter. Therefore, sumoylation restrained inflammation by silencing Ifnb1 expression and by strictly suppressing an unanticipated priming by type I interferons of the TLR-induced production of inflammatory cytokines.

Despite numerous studies on specific sumoylated transcriptional regulators, the global role of SUMO on chromatin in relation to transcription regulation remains largely unknown. Here, we determined the genome-wide localization of SUMO1 and SUMO2/3, as well as of UBC9 (encoded by UBE2I) and PIASY (encoded by PIAS4), two markers for active sumoylation, along with Pol II and histone marks in proliferating versus senescent human fibroblasts together with gene expression profiling. We found that, whereas SUMO alone is widely distributed over the genome with strong association at active promoters, active sumoylation occurs most prominently at promoters of histone and protein biogenesis genes, as well as Pol I rRNAs and Pol III tRNAs. Remarkably, these four classes of genes are up-regulated by inhibition of sumoylation, indicating that SUMO normally acts to restrain their expression. In line with this finding, sumoylation-deficient cells show an increase in both cell size and global protein levels. Strikingly, we found that in senescent cells, the SUMO machinery is selectively retained at histone and tRNA gene clusters, whereas it is massively released from all other unique chromatin regions. These data, which reveal the highly dynamic nature of the SUMO landscape, suggest that maintenance of a repressive environment at histone and tRNA loci is a hallmark of the senescent state. The approach taken in our study thus permitted the identification of a common biological output and uncovered hitherto unknown functions for active sumoylation at chromatin as a key mechanism that, in dynamically marking chromatin by a simple modifier, orchestrates concerted transcriptional regulation of a network of genes essential for cell growth and proliferation.

Advances in genetics have shed light on the molecular basis of Crohn's disease (CD) predisposition and pathogenesis, via linkage disequilibrium analysis to genome-wide association studies. The discovery of genetic variants of NOD2, an intracellular pathogen molecular sensor, as risk factors for CD has paved the way for further research on innate immunity in this disease. Remarkably, polymorphisms in autophagy genes, such as ATG16L1 and IRGM, have been identified, allowing the pivotal role of autophagy in innate immunity to be uncovered. In this review, we summarize recent studies on the CD-associated NOD2, ATG16L1 and IRGM risk variants and their contribution to the autophagy functions that have most influenced our understanding of CD pathophysiology.

Formerly a commensal organism of the mucosal surfaces of most healthy individuals, Candida albicans is an opportunistic pathogen that causes infections ranging from superficial to the more life-threatening disseminated infections, especially in the ever-growing population of vulnerable patients in the hospital setting. In these situations, the fungus takes advantage of its host following a disturbance in the host defense system and/or the mucosal microbiota. Overwhelming evidence suggests that the gastrointestinal tract is the main source of disseminated C. albicans infections. Major risk factors for disseminated candidiasis include damage to the mucosal intestinal barrier, immune dysfunction, and dysbiosis of the resident microbiota. A better understanding of C. albicans' interaction with the intestinal epithelial barrier will be useful for designing future therapies to avoid systemic candidiasis. In this review, we provide an overview of the current knowledge regarding the mechanisms of pathogenicity that allow the fungus to reach and translocate the gut barrier.

Abstract In this study, we show that calcium pectinate beads (CPB) allow the formation of 20 µm spherical microcolonies of the probiotic bacteria Lacticaseibacillus paracasei (formerly designated as Lactobacillus paracasei ) ATCC334 with a high cell density, reaching more than 10 log (CFU/g). The bacteria within these microcolonies are well structured and adhere to a three-dimensional network made of calcium-pectinate through the synthesis of extracellular polymeric substances (EPS) and thus display a biofilm-like phenotype, an attractive property for their use as probiotics. During bacterial development in the CPB, a coalescence phenomenon arises between neighboring microcolonies accompanied by their peripheral spatialization within the bead. Moreover, the cells of L. paracasei ATCC334 encased in these pectinate beads exhibit increased resistance to acidic stress (pH 1.5), osmotic stress (4.5 M NaCl), the freeze-drying process and combined stresses, simulating the harsh conditions encountered in the gastrointestinal (GI) tract. In vivo, the oral administration of CPB-formulated L. paracasei ATCC334 in mice demonstrated that biofilm-like microcolonies are successfully released from the CPB matrix in the colonic environment. In addition, these CPB-formulated probiotic bacteria display the ability to reduce the severity of a DSS-induced colitis mouse model, with a decrease in colonic mucosal injuries, less inflammation, and reduced weight loss compared to DSS control mice. To conclude, this work paves the way for a new form of probiotic administration in the form of biofilm-like microcolonies with enhanced functionalities.

The formation of membrane vesicles (MVs) by Gram-positive bacteria has gained increasing attention over the last decade. Recently, models of vesicle formation have been proposed and involve the digestion of the cell wall by prophage-encoded or stress-induced peptidoglycan (PG) hydrolases and the inhibition of PG synthesis by β-lactam antibiotics. The impact of these mechanisms on vesicle formation is largely dependent on the strain and growth conditions. To date, no information on the production of vesicles by the lactobacilli family has been reported. Here, we aimed to characterize the MVs released by the Gram-positive bacteria Lacticaseibacillus casei BL23 and also investigated the mechanisms involved in vesicle formation. Using electron microscopy, we established that the size of the majority of L. casei BL23 vesicles ranged from 50 to 100 nm. Furthermore, we showed that the vesicles were released consistently throughout the growth of the bacteria in standard culture conditions. The protein composition of the vesicles released in the supernatant was identified and a significant number of prophage proteins was detected. Moreover, using a mutant strain harboring a defective PLE2 prophage, we were able to show that the spontaneous and mitomycin-triggered induction of the prophage PLE2 contribute to the production of MVs by L. casei BL23. Finally, we also demonstrated the influence of prophages on the membrane integrity of bacteria. Overall, our results suggest a key role of the prophage PLE2 in the production of MVs by L. casei BL23 in the absence or presence of genotoxic stress. IMPORTANCE The last few decades have demonstrated that membrane vesicles (MVs) produced by microorganisms can have a wide variety of functions. This diversity places MVs at the crossroads of major research topics in current microbiology such as antibiotic resistance, horizontal gene transfer, cell communication, biofilm development, bacteriophage resistance, and pathogenesis. In particular, vesicles produced by probiotic strains have been shown to play a significant role in their beneficial effects. Thus, the study of vesicle biogenesis is a key element for promoting and improving their release. Overall, our results suggest a key role of spontaneous and mitomycin-triggered prophage induction in MV production by the Gram-positive bacteria Lacticaseibacillus casei BL23. This phenomenon is of great interest as prophage-induced MVs could potentially influence bacterial behavior, stress resistance, and vesicle functions.

Gene silencing by small RNAs has emerged as a powerful post-transcriptional regulator of gene expression, however processes underlying regulation of the small RNA pathway in vivo are still largely elusive. Here, we identified sumoylation as a novel post-translational modification acting on Ago2, the main effector of small RNA-mediated gene silencing. We demonstrate that Ago2 can be modified by SUMO1 and SUMO2/3 and identified Lys402 as the major Ago2 sumoylation site in vivo. Ago2 physically interacts with the SUMO E2 conjugating enzyme Ubc9 and the E3 ligase RanBP2 facilitates Ago2 sumoylation in vitro. Mutation of Lys402 enhances the stability of Ago2 protein and impairment of cellular sumoylation by siRNA- or shRNA-mediated extinction of Ubc9 or in Ubc9 knockout mouse tissues results in increased steady-state levels and enhanced stability of Ago2. Similarly, knockdown of RanBP2 or of the SAE2 E1 enzyme enhances Ago2 protein levels. Lys402 is located in the L2g1 loop linking the PAZ and PIWI domains of Ago2, in the immediate vicinity of Tyr393 which can be phosphorylated, implying that the L2g1 linker represents an easily accessible hot spot for post-translational modifications. Altogether, our results show that sumoylation of Ago2 at Lys402 negatively regulates its stability, thereby establishing a first link between SUMO and the small RNA machinery.

Autophagy is a lysosomal degradation process that contributes to host immunity by eliminating invasive pathogens and the modulating inflammatory response. Several infectious and immune disorders are associated with autophagy defects, suggesting that stimulation of autophagy in these diseases should be beneficial. Here, we show that resveratrol is able to boost xenophagy, a selective form of autophagy that target invasive bacteria. We demonstrated that resveratrol promotes in vitro autophagy-dependent clearance of intracellular bacteria in intestinal epithelial cells and macrophages. These results were validated in vivo using infection in a GFP-LC3 zebrafish model. We also compared the ability of resveratrol derivatives, designed to improve the bioavailability of the parent molecule, to stimulate autophagy and to induce intracellular bacteria clearance. Together, our data demonstrate the ability of resveratrol to stimulate xenophagy, and thereby enhance the clearance of two invasive bacteria involved life-threatening diseases, Salmonella Typhimurium and Crohn’s disease-associated Adherent-Invasive Escherichia coli. These findings encourage the further development of pro-autophagic nutrients to strengthen intestinal homeostasis in basal and infectious states.

Bacterial strains of the Lactobacillaceae family are widely used as probiotics for their multifaceted potential beneficial properties. However, no official recommendations for their clinical use exist since, in many cases, oral administrations of these bacteria displayed limited beneficial effects in human. Additional research is thus needed to improve the efficiency of existing strains with strong potential. In this context, we assess in vitro the effects of nine polyphenols to stimulate biofilm formation by lactobacilli, a feature enhancing their functionalities. Among these polyphenols, we identify trans-Resveratrol (referred to hereafter as Resveratrol) as a potent inducer of biofilm formation by Lacticaseibacillus paracasei (formerly designated as Lactobacillus paracasei) ATCC334 strain. This effect is strain-dependent and relies on the enhancement of L. paracasei adhesion to abiotic and biotic surfaces, including intestinal epithelial cells. Mechanistically, Resveratrol modify physico-chemical properties of the bacterial surface and thereby enhances L. paracasei aggregation, subsequently facilitating adhesion and biofilm development. Together, our in vitro data demonstrate that Resveratrol might be used to modulate the behavior of Lactobacilli with probiotic properties. Combination of probiotics and polyphenols could be considered to enhance the probiotic functionalities in further in vivo studies.

Candida albicans (C. albicans) is an opportunistic pathogen causing infections ranging from superficial to life-threatening disseminated infections. In a susceptible host, C. albicans is able to translocate through the gut barrier, promoting its dissemination into deeper organs. C. albicans hyphae can invade human epithelial cells by two well-documented mechanisms: epithelial-driven endocytosis and C. albicans-driven active penetration. One mechanism by which host cells protect themselves against intracellular C. albicans is termed autophagy. The protective role of autophagy during C. albicans infection has been investigated in myeloid cells; however, far less is known regarding the role of this process during the infection of epithelial cells. In the present study, we investigated the role of autophagy-related proteins during the infection of epithelial cells, including intestinal epithelial cells and gut explants, by C. albicans. Using cell imaging, we show that key molecular players of the autophagy machinery (LC3-II, PI3P, ATG16L1, and WIPI2) were recruited at Candida invasion sites. We deepened these observations by electron microscopy analyses that reveal the presence of autophagosomes in the vicinity of invading hyphae. Importantly, these events occur during active penetration of C. albicans into host cells and are associated with plasma membrane damage. In this context, we show that the autophagy-related key proteins ATG5 and ATG16L1 contribute to plasma membrane repair mediated by lysosomal exocytosis and participate in protecting epithelial cells against C. albicans-induced cell death. Our findings provide a novel mechanism by which epithelial cells, forming the first line of defense against C. albicans in the gut, can react to limit C. albicans invasion.

Susceptibility to Crohn disease (CD), an inflammatory bowel disease, is influenced by common variants at many loci like the exonic synonymous IRGM SNP (rs10065172, NM_001145805.1, c.313C>T). We recently showed that miR-196 is overexpressed in the inflammatory intestinal epithelia of individuals with CD and downregulates the IRGM protective (c.313C) but not the risk-associated (c.313T) allele. Eventually, loss ofIRGM/miRNA regulation compromises xenophagy. These results highlight a critical “ménage à trois” in risk susceptibility combining IRGM allele, miRNA and xenophagy.

Disruption of the sumoylation/desumoylation equilibrium is associated with several disease states such as cancer and infections, however the mechanisms regulating the global SUMO balance remain poorly defined. Here, we show that infection by Shigella flexneri, the causative agent of human bacillary dysentery, switches off host sumoylation during epithelial cell infection in vitro and in vivo and that this effect is mainly mediated by a calcium/calpain-induced cleavage of the SUMO E1 enzyme SAE2, thus leading to sumoylation inhibition. Furthermore, we describe a mechanism by which Shigella promotes its own invasion by altering the sumoylation state of RhoGDIα, a master negative regulator of RhoGTPase activity and actin polymerization. Together, our data suggest that SUMO modification is essential to restrain pathogenic bacterial entry by limiting cytoskeletal rearrangement induced by bacterial effectors. Moreover, these findings identify calcium-activated calpains as powerful modulators of cellular sumoylation levels with potentially broad implications in several physiological and pathological situations.

A symbiotic relationship has set up between the gut microbiota and its host in the course of evolution, forming an interkingdom consortium. The gut offers a favorable ecological niche for microbial communities, with the whole body and external factors (e.g., diet or medications) contributing to modulating this microenvironment. Reciprocally, the gut microbiota is important for maintaining health by acting not only on the gut mucosa but also on other organs. However, failure in one or another of these two partners can lead to the breakdown in their symbiotic equilibrium and contribute to disease onset and/or progression. Several microbial and host processes are devoted to facing up the stress that could alter the symbiosis, ensuring the resilience of the ecosystem. Among these processes, autophagy is a host catabolic process integrating a wide range of stress in order to maintain cell survival and homeostasis. This cytoprotective mechanism, which is ubiquitous and operates at basal level in all tissues, can be rapidly down- or up-regulated at the transcriptional, post-transcriptional, or post-translational levels, to respond to various stress conditions. Because of its sensitivity to all, metabolic-, immune-, and microbial-derived stimuli, autophagy is at the crossroad of the dialogue between changes occurring in the gut microbiota and the host responses. In this review, we first delineate the modulation of host autophagy by the gut microbiota locally in the gut and in peripheral organs. Then, we describe the autophagy-related mechanisms affecting the gut microbiota. We conclude this review with the current challenges and an outlook toward the future interventions aiming at modulating host autophagy by targeting the gut microbiota.

Abstract Omega-3 (n-3) polyunsaturated fatty acids (PUFAs), particularly docosahexaenoic acid (DHA), are required for the structure and function of the retina. Several observational studies indicate that consumption of a diet with relatively high levels of n-3 PUFAs, such as those provided by fish oils, has a protective effect against the development of age-related macular degeneration. Given the accumulating evidence showing the role of gut microbiota in regulating retinal physiology and host lipid metabolism, we evaluated the potential of long-term dietary supplementation with the Gram-positive bacterium Lactobacillus helveticus strain VEL12193 to modulate the retinal n-3 PUFA content. A set of complementary approaches was used to study the impact of such a supplementation on the gut microbiota and host lipid/fatty acid (FA) metabolism. L. helveticus -supplementation was associated with a decrease in retinal saturated FAs (SFAs) and monounsaturated FAs (MUFAs) as well as an increase in retinal n-3 and omega-6 (n-6) PUFAs. Interestingly, supplementation with L. helveticus enriched the retina in C22:5n-3 (docosapentaenoic acid, DPA), C22:6n-3 (DHA), C18:2n-6 (linoleic acid, LA) and C20:3n-6 (dihomo gamma-linolenic acid, DGLA). Long-term consumption of L. helveticus also modulated gut microbiota composition and some changes in OTUs abundance correlated with the retinal FA content. This study provides a proof of concept that targeting the gut microbiota could be an effective strategy to modulate the retinal FA content, including that of protective n-3 PUFAs, thus opening paths for the design of novel preventive and/or therapeutical strategies for retinopathies.

Ileal lesions in Crohn's disease (CD) patients are colonized by pathogenic adherent-invasive Escherichia coli (AIEC) able to invade and to replicate within intestinal epithelial cells. Recent advances have highlighted the importance of the innate immune system and the critical relationship between the gut flora and the intestinal mucosa. Several combinations of genetic predisposing factors to CD have been described, with the most significant replicable associations including genes for intracellular receptor of bacterial cell walls (NOD2/CARD15), and for bacterial clearance and antigen processing through autophagy (ATG16L1 and IRGM). We recently reported that in IRGM and ATG16L1 deficient cells, intracellular AIEC LF82 bacteria have enhanced replication and that autophagy deficiency surprisingly did not interfere with the ability of intracellular bacteria to survive and/or replicate for any other E. coli strains tested, including nonpathogenic, environmental, commensal, or pathogenic strains involved in gastroenteritis. As autophagy is an innate defense mechanism acting as a cell-autonomous system for elimination of intracellular pathogens, these findings lead weight to the notion that intracellular bacteria including AIEC might play a role in CD pathogenesis.

Several old and new observations suggest the existence in Crohn's disease of a phagocytic disorder of macrophages related to impaired bactericidal activity of host cells or to the presence of invasive bacteria that have developed strategies to counteract macrophage killing. It was recently reported that disordered macrophage cytokine secretion underlies impaired acute inflammation and bacterial clearance in Crohn's disease. Secretion of proinflammatory cytokines by CD macrophages was impaired in response to E. coli or specific Toll-like receptor agonists. In addition, major advances in the etiology of Crohn's disease came from the existence of polymorphism in NOD2 and autophagy-related susceptibility genes (ATG16L1 and IRGM) in patients and from the identification of the presence of adherent-invasive E. coli (AIEC) colonizing the CD ileal mucosa and able to resist to macrophage killing. The role of impaired autophagy in Crohn's disease patients has been recently reinforced by the observation that the peptidoglycan receptor NOD2, in addition to sense intracellular bacteria, can induce autophagy by recruiting the critical autophagy protein ATG16L1 to the plasma membrane during bacterial internalization. Defects in autophagy might be the key element of the pathogenic pathway that lead to defective microbial killing, increased exposure to commensal and pathogenic intestinal bacteria and T cell activation. Defects in Paneth cells secreting lysozyme and antimicrobial peptides are observed in patients with ATG16L1 risk allele. Thus, the induction of autophagy or administration of preparations that mirrors the secretion of Paneth cells or both may be regarded as new therapeutic avenues for the treatment of Crohn's disease.

Autophagy is a lysosomal degradation mechanism which helps to control intracellular infections and contributes to the regulation of innate and adaptive immune responses. Defects in autophagy lead to exacerbated proliferation of microorganisms and/or to excessive immune responses which are both highly deleterious. Thus, infectious and chronic inflammatory human diseases, such as Crohn's disease, are often associated with inappropriate modulation of autophagy, which is mainly linked to autophagy-associated gene polymorphisms. In this review, we highlight the current understanding of role of autophagy in infections and immunity.

Candida albicans (C. albicans) is an opportunistic pathogen causing infections ranging from superficial to life-threatening dissemination, in which C. albicans is able to translocate through the gut barrier into deeper organs. In its filamentous form (hyphae), C. albicans can invade epithelial cells by two mechanisms: epithelial cell-driven endocytosis and C. albicans-driven active penetration of host cell plasma membrane (PM). Autophagic machinery is known to be involved in the epithelial barrier maintenance, especially the intestinal barrier that is continuously challenged by exposure to the gut microbiota or to xenobiotics. The protective role of autophagy during C. albicans infection has been investigated in myeloid cells, however, far less was known regarding its role during infection of epithelial cells. Here, we demonstrated that key proteins of the autophagic machinery and vesicles presenting features of autophagosomes are recruited at C. albicans invasion sites. These events are associated with host PM damage caused by the active penetration of C. albicans. We showed that ATG5 and ATG16L1 proteins contribute to PM repair mediated by lysosomal membrane exocytosis and participate in protection of epithelial cells’ integrity against C. albicans-induced cell death. Our findings extend the knowledge on emerging roles of the autophagic machinery in stress-related membrane dynamics.

Arlette Darfeuille-Michaud: Researcher, Lecturer, Leader, Mentor, and FriendArlette lived her early years in the tiny French village of Beyssenac (in the beautiful Corrèze region) where her family owned and operated a fruit orchard. Arlette earned her PhD in 1987 at Auvergne University in the lab of Dr Bernard Joly, studying Klebsiella pneumoniae strains involved in nosocomial infections and Escherichia coli pathogenesis. Arlette then joined the faculty at Auvergne University as a lecturer in 1989, and then was promoted to professor in microbiology and molecular biology in 1994. Arlette then initiated a line of experimentation that would ultimately help contribute to our understanding of the pathogenesis of inflammatory bowel disease (IBD). Briefly, working with gastroenterologist Dr Jean-Frédéric Colombel and Dr Christel Neut, Arlette made the discovery, reported in Gastroenterology in 1998, that many ileal samples of Crohn’s patients were colonized by a previously unappreciated class of E coli strains. As Dr Colombel recently recalled,“Arlette called us, very excited and enthusiastic, exclaiming that she had discovered that these Escherichia coli belonged to a new pathovar she named AIEC (for adherent-invasive E coli). We developed, then, an intense collaboration that extended beyond science to very close friendship.”Arlette then pursued her research on characterizing disease-promoting determinants of AIEC, which included its type 1 pili, flagella, outer membrane vesicles, and long polar fimbriae. Arlette subsequently focused on identifying host molecules that permitted AIEC colonization, resulting in her discovery in 2007 that Crohn’s patients exhibit elevated expression of glycoprotein CEACAM6 that mediate AIEC adhesion. A few years later, in collaboration with Dr Paul Hofman, Arlette reported in Gut an abnormal expression of the Gp96 ER stress response chaperone on the apical side of ileal intestinal cells in Crohn’s patients, favoring AIEC invasion. In 2010, Arlette reported the complete genome of a Crohn's disease-associated E coli strain, specifically AIEC strain LF82, which is now studied as a reference strain in labs around the world. Arlette demonstrated that AIEC survive and replicate in mature phagolysosomes in macrophages, possibly as a result of defective autophagy that is thought to play a role in Crohn's disease. More recently, Arlette, with Dr Richard Bonnet and Dr Denis Pezet, revealed potential mechanisms by which tumor-associated E coli could lead to cancer progression. These studies highlighted Arlette’s extensive body of research, which included over 100 peer-reviewed manuscripts, many in the highest-ranked journals including Gastroenterology, Gut, and Journal of Clinical Investigation, and has been instrumental in leading to the emerging paradigm that inability to manage opportunistic bacteria is a pivotal event in numerous intestinal diseases, particularly IBD and cancer. The importance of Arlette’s research was also recognized by numerous invited lectures and awards including the Charles Debray Research Award in Gastroenterology (1994), Jacques Piraud Research Award (2011), and Chevalier de La Légion d'Honneur (2012). Arlette firmly believed in the importance of translating discoveries in basic research to prevent and treat IBD and, consequently, attained 3 patents on her work aimed at development of IBD therapeutics.A key ingredient in Arlette’s success, in many areas of her life, was her boundless energy and her unbridled optimism. She used these qualities to inspire collaborators, students and administrators to build an INSERM research unit in Clermont-Ferrand, which Arlette often referred to as “the middle of nowhere.” Founded in 2012, with Arlette as Director, and named “Microbe, Intestine, Inflammation and Host Susceptibility,” the unit includes 21 full-time research personnel, 4 postdoctoral fellows and 7 PhD students. Arlette’s irrepressible enthusiasm attracted a talented cadre of basic scientists and clinicians while her visionary leadership forged a multidisciplinary approach involving an array of scientific fields including microbiology, cellular and molecular biology, immunology, gastroenterology, surgery, and internal medicine. Arlette was especially passionate about developing scientific talent. She was a spirited lecturer who taught over 3000 students in 30 years of teaching. She inspired many of her students to pursue careers in research and medicine. She was an extraordinary mentor to the numerous PhD students and postdoctoral fellows whom were fortunate enough to join her lab. She provided mentorship at all levels, helping her mentees in training and in career development. She made it her utmost responsibility to ensure that her students were always well prepared for presentations, especially their thesis defense. Arlette’s students’ thesis defenses were especially important days to her, on which she viewed herself as a proud mother taking great pride in her student’s achievement, but yet being a bit melancholic to see her fledglings leave the nest. But Arlette’s mentorship did not end at graduation. She continued to provide support and encouragement to her trainees up until the end of her life. Many of Arlette’s trainees are now emerging scholars around the world (including several in Clermont-Ferrand) in areas related to IBD and/or on bacterial colonization and pathogenesis, and will undoubtedly continue to benefit from what they’ve learned from Arlette throughout their careers.Despite her demanding career, Arlette was a wonderful wife to Jean-Eric with whom they lovingly raised 2 sons, Pierre-Yohan, who earned a PhD in acoustics and Vincent, who will soon complete medical school. The respite for Arlette’s family was an old mill house that she and Jean-Eric beautifully restored. Arlette loved to host her scientific family at her home, especially when it was time to celebrate the numerous accomplishments of her trainees and colleagues. At such gatherings, Arlette was always thrilled to personally prepare homemade specialties from her native region.Arlette cared deeply for all her co-workers and colleagues, in good and challenging times. We remember the loving care she gave to her friend and co-worker, Dr Anne-Lise Glasser, during her long fight against cancer while never wavering in her optimism for the best possible outcome. While the news of Arlette’s illness was a stunning blow to all who knew her, it seemed to only rally her inner strength and incredible energy to fight her illness to the end and lead a productive life as long as possible. She continued to engage in numerous local and international research activities, including grant and manuscript reviews, and helping colleagues design experiments, write manuscripts, and plan conferences, while always thinking about how such activities could benefit patients with Crohn’s disease. Fittingly, one of Arlette's last activities was her overseeing the successful thesis defense of her final PhD student, which took place the day before she left us.Arlette will be dearly and forever missed by her family, her many friends and colleagues across the world. On behalf of all the trainees and students who received Arlette’s guidance during their lifetime, we, her former PhD students, view ourselves as incredibly fortunate and honored to have received her mentorship and we will continue, every day, to pay tribute to her legacy.Editor’s Note. This article is co-published with Gut. Arlette Darfeuille-Michaud: Researcher, Lecturer, Leader, Mentor, and FriendArlette lived her early years in the tiny French village of Beyssenac (in the beautiful Corrèze region) where her family owned and operated a fruit orchard. Arlette earned her PhD in 1987 at Auvergne University in the lab of Dr Bernard Joly, studying Klebsiella pneumoniae strains involved in nosocomial infections and Escherichia coli pathogenesis. Arlette then joined the faculty at Auvergne University as a lecturer in 1989, and then was promoted to professor in microbiology and molecular biology in 1994. Arlette then initiated a line of experimentation that would ultimately help contribute to our understanding of the pathogenesis of inflammatory bowel disease (IBD). Briefly, working with gastroenterologist Dr Jean-Frédéric Colombel and Dr Christel Neut, Arlette made the discovery, reported in Gastroenterology in 1998, that many ileal samples of Crohn’s patients were colonized by a previously unappreciated class of E coli strains. As Dr Colombel recently recalled,“Arlette called us, very excited and enthusiastic, exclaiming that she had discovered that these Escherichia coli belonged to a new pathovar she named AIEC (for adherent-invasive E coli). We developed, then, an intense collaboration that extended beyond science to very close friendship.”Arlette then pursued her research on characterizing disease-promoting determinants of AIEC, which included its type 1 pili, flagella, outer membrane vesicles, and long polar fimbriae. Arlette subsequently focused on identifying host molecules that permitted AIEC colonization, resulting in her discovery in 2007 that Crohn’s patients exhibit elevated expression of glycoprotein CEACAM6 that mediate AIEC adhesion. A few years later, in collaboration with Dr Paul Hofman, Arlette reported in Gut an abnormal expression of the Gp96 ER stress response chaperone on the apical side of ileal intestinal cells in Crohn’s patients, favoring AIEC invasion. In 2010, Arlette reported the complete genome of a Crohn's disease-associated E coli strain, specifically AIEC strain LF82, which is now studied as a reference strain in labs around the world. Arlette demonstrated that AIEC survive and replicate in mature phagolysosomes in macrophages, possibly as a result of defective autophagy that is thought to play a role in Crohn's disease. More recently, Arlette, with Dr Richard Bonnet and Dr Denis Pezet, revealed potential mechanisms by which tumor-associated E coli could lead to cancer progression. These studies highlighted Arlette’s extensive body of research, which included over 100 peer-reviewed manuscripts, many in the highest-ranked journals including Gastroenterology, Gut, and Journal of Clinical Investigation, and has been instrumental in leading to the emerging paradigm that inability to manage opportunistic bacteria is a pivotal event in numerous intestinal diseases, particularly IBD and cancer. The importance of Arlette’s research was also recognized by numerous invited lectures and awards including the Charles Debray Research Award in Gastroenterology (1994), Jacques Piraud Research Award (2011), and Chevalier de La Légion d'Honneur (2012). Arlette firmly believed in the importance of translating discoveries in basic research to prevent and treat IBD and, consequently, attained 3 patents on her work aimed at development of IBD therapeutics.A key ingredient in Arlette’s success, in many areas of her life, was her boundless energy and her unbridled optimism. She used these qualities to inspire collaborators, students and administrators to build an INSERM research unit in Clermont-Ferrand, which Arlette often referred to as “the middle of nowhere.” Founded in 2012, with Arlette as Director, and named “Microbe, Intestine, Inflammation and Host Susceptibility,” the unit includes 21 full-time research personnel, 4 postdoctoral fellows and 7 PhD students. Arlette’s irrepressible enthusiasm attracted a talented cadre of basic scientists and clinicians while her visionary leadership forged a multidisciplinary approach involving an array of scientific fields including microbiology, cellular and molecular biology, immunology, gastroenterology, surgery, and internal medicine. Arlette was especially passionate about developing scientific talent. She was a spirited lecturer who taught over 3000 students in 30 years of teaching. She inspired many of her students to pursue careers in research and medicine. She was an extraordinary mentor to the numerous PhD students and postdoctoral fellows whom were fortunate enough to join her lab. She provided mentorship at all levels, helping her mentees in training and in career development. She made it her utmost responsibility to ensure that her students were always well prepared for presentations, especially their thesis defense. Arlette’s students’ thesis defenses were especially important days to her, on which she viewed herself as a proud mother taking great pride in her student’s achievement, but yet being a bit melancholic to see her fledglings leave the nest. But Arlette’s mentorship did not end at graduation. She continued to provide support and encouragement to her trainees up until the end of her life. Many of Arlette’s trainees are now emerging scholars around the world (including several in Clermont-Ferrand) in areas related to IBD and/or on bacterial colonization and pathogenesis, and will undoubtedly continue to benefit from what they’ve learned from Arlette throughout their careers.Despite her demanding career, Arlette was a wonderful wife to Jean-Eric with whom they lovingly raised 2 sons, Pierre-Yohan, who earned a PhD in acoustics and Vincent, who will soon complete medical school. The respite for Arlette’s family was an old mill house that she and Jean-Eric beautifully restored. Arlette loved to host her scientific family at her home, especially when it was time to celebrate the numerous accomplishments of her trainees and colleagues. At such gatherings, Arlette was always thrilled to personally prepare homemade specialties from her native region.Arlette cared deeply for all her co-workers and colleagues, in good and challenging times. We remember the loving care she gave to her friend and co-worker, Dr Anne-Lise Glasser, during her long fight against cancer while never wavering in her optimism for the best possible outcome. While the news of Arlette’s illness was a stunning blow to all who knew her, it seemed to only rally her inner strength and incredible energy to fight her illness to the end and lead a productive life as long as possible. She continued to engage in numerous local and international research activities, including grant and manuscript reviews, and helping colleagues design experiments, write manuscripts, and plan conferences, while always thinking about how such activities could benefit patients with Crohn’s disease. Fittingly, one of Arlette's last activities was her overseeing the successful thesis defense of her final PhD student, which took place the day before she left us.Arlette will be dearly and forever missed by her family, her many friends and colleagues across the world. On behalf of all the trainees and students who received Arlette’s guidance during their lifetime, we, her former PhD students, view ourselves as incredibly fortunate and honored to have received her mentorship and we will continue, every day, to pay tribute to her legacy.Editor’s Note. This article is co-published with Gut. Arlette lived her early years in the tiny French village of Beyssenac (in the beautiful Corrèze region) where her family owned and operated a fruit orchard. Arlette earned her PhD in 1987 at Auvergne University in the lab of Dr Bernard Joly, studying Klebsiella pneumoniae strains involved in nosocomial infections and Escherichia coli pathogenesis. Arlette then joined the faculty at Auvergne University as a lecturer in 1989, and then was promoted to professor in microbiology and molecular biology in 1994. Arlette then initiated a line of experimentation that would ultimately help contribute to our understanding of the pathogenesis of inflammatory bowel disease (IBD). Briefly, working with gastroenterologist Dr Jean-Frédéric Colombel and Dr Christel Neut, Arlette made the discovery, reported in Gastroenterology in 1998, that many ileal samples of Crohn’s patients were colonized by a previously unappreciated class of E coli strains. As Dr Colombel recently recalled,“Arlette called us, very excited and enthusiastic, exclaiming that she had discovered that these Escherichia coli belonged to a new pathovar she named AIEC (for adherent-invasive E coli). We developed, then, an intense collaboration that extended beyond science to very close friendship.” Arlette then pursued her research on characterizing disease-promoting determinants of AIEC, which included its type 1 pili, flagella, outer membrane vesicles, and long polar fimbriae. Arlette subsequently focused on identifying host molecules that permitted AIEC colonization, resulting in her discovery in 2007 that Crohn’s patients exhibit elevated expression of glycoprotein CEACAM6 that mediate AIEC adhesion. A few years later, in collaboration with Dr Paul Hofman, Arlette reported in Gut an abnormal expression of the Gp96 ER stress response chaperone on the apical side of ileal intestinal cells in Crohn’s patients, favoring AIEC invasion. In 2010, Arlette reported the complete genome of a Crohn's disease-associated E coli strain, specifically AIEC strain LF82, which is now studied as a reference strain in labs around the world. Arlette demonstrated that AIEC survive and replicate in mature phagolysosomes in macrophages, possibly as a result of defective autophagy that is thought to play a role in Crohn's disease. More recently, Arlette, with Dr Richard Bonnet and Dr Denis Pezet, revealed potential mechanisms by which tumor-associated E coli could lead to cancer progression. These studies highlighted Arlette’s extensive body of research, which included over 100 peer-reviewed manuscripts, many in the highest-ranked journals including Gastroenterology, Gut, and Journal of Clinical Investigation, and has been instrumental in leading to the emerging paradigm that inability to manage opportunistic bacteria is a pivotal event in numerous intestinal diseases, particularly IBD and cancer. The importance of Arlette’s research was also recognized by numerous invited lectures and awards including the Charles Debray Research Award in Gastroenterology (1994), Jacques Piraud Research Award (2011), and Chevalier de La Légion d'Honneur (2012). Arlette firmly believed in the importance of translating discoveries in basic research to prevent and treat IBD and, consequently, attained 3 patents on her work aimed at development of IBD therapeutics. A key ingredient in Arlette’s success, in many areas of her life, was her boundless energy and her unbridled optimism. She used these qualities to inspire collaborators, students and administrators to build an INSERM research unit in Clermont-Ferrand, which Arlette often referred to as “the middle of nowhere.” Founded in 2012, with Arlette as Director, and named “Microbe, Intestine, Inflammation and Host Susceptibility,” the unit includes 21 full-time research personnel, 4 postdoctoral fellows and 7 PhD students. Arlette’s irrepressible enthusiasm attracted a talented cadre of basic scientists and clinicians while her visionary leadership forged a multidisciplinary approach involving an array of scientific fields including microbiology, cellular and molecular biology, immunology, gastroenterology, surgery, and internal medicine. Arlette was especially passionate about developing scientific talent. She was a spirited lecturer who taught over 3000 students in 30 years of teaching. She inspired many of her students to pursue careers in research and medicine. She was an extraordinary mentor to the numerous PhD students and postdoctoral fellows whom were fortunate enough to join her lab. She provided mentorship at all levels, helping her mentees in training and in career development. She made it her utmost responsibility to ensure that her students were always well prepared for presentations, especially their thesis defense. Arlette’s students’ thesis defenses were especially important days to her, on which she viewed herself as a proud mother taking great pride in her student’s achievement, but yet being a bit melancholic to see her fledglings leave the nest. But Arlette’s mentorship did not end at graduation. She continued to provide support and encouragement to her trainees up until the end of her life. Many of Arlette’s trainees are now emerging scholars around the world (including several in Clermont-Ferrand) in areas related to IBD and/or on bacterial colonization and pathogenesis, and will undoubtedly continue to benefit from what they’ve learned from Arlette throughout their careers. Despite her demanding career, Arlette was a wonderful wife to Jean-Eric with whom they lovingly raised 2 sons, Pierre-Yohan, who earned a PhD in acoustics and Vincent, who will soon complete medical school. The respite for Arlette’s family was an old mill house that she and Jean-Eric beautifully restored. Arlette loved to host her scientific family at her home, especially when it was time to celebrate the numerous accomplishments of her trainees and colleagues. At such gatherings, Arlette was always thrilled to personally prepare homemade specialties from her native region. Arlette cared deeply for all her co-workers and colleagues, in good and challenging times. We remember the loving care she gave to her friend and co-worker, Dr Anne-Lise Glasser, during her long fight against cancer while never wavering in her optimism for the best possible outcome. While the news of Arlette’s illness was a stunning blow to all who knew her, it seemed to only rally her inner strength and incredible energy to fight her illness to the end and lead a productive life as long as possible. She continued to engage in numerous local and international research activities, including grant and manuscript reviews, and helping colleagues design experiments, write manuscripts, and plan conferences, while always thinking about how such activities could benefit patients with Crohn’s disease. Fittingly, one of Arlette's last activities was her overseeing the successful thesis defense of her final PhD student, which took place the day before she left us. Arlette will be dearly and forever missed by her family, her many friends and colleagues across the world. On behalf of all the trainees and students who received Arlette’s guidance during their lifetime, we, her former PhD students, view ourselves as incredibly fortunate and honored to have received her mentorship and we will continue, every day, to pay tribute to her legacy. Editor’s Note. This article is co-published with Gut. Editor’s Note. This article is co-published with Gut. Editor’s Note. This article is co-published with Gut.

Abstract Omega-3 (n-3) polyunsaturated fatty acids (PUFAs), particularly docosahexaenoic acid (DHA), are required for the structure and function of the retina. They could also help to prevent or delay the development of retinopathies. Given the accumulating evidence showing the role of gut microbiota in regulating retinal physiology and host lipid metabolism, we evaluated the potential of long-term dietary supplementation with the Gram-positive bacterium Lacticaseibacillus helveticus strain VEL12193 to modulate the retinal n-3 PUFA content. A set of complementary approaches was used to study the impact of such a supplementation on the gut microbiota and host lipid/fatty acid (FA) metabolism. L. helveticus-supplementation was associated with a decrease in retinal saturated FAs (SFAs) and monounsaturated FAs (MUFAs) as well as an increase in retinal n-3 and omega-6 (n-6) PUFAs. Interestingly, supplementation with L. helveticus enriched the retina in C22:5n-3 (docosapentaenoic acid, DPA), C22:6n-3 (DHA), C18:2n-6 (linoleic acid, LA) and C20:3n-6 (dihomo gamma-linolenic acid, DGLA). Long-term consumption of L. helveticus also modulated gut microbiota composition and some changes in OTUs abundance correlated with the retinal FA content. This study provides a proof of concept that targeting the gut microbiota could be an effective strategy to modulate the retinal FA content, including that of protective n-3 PUFAs, thus opening paths for the design of novel preventive and/or therapeutical strategies for retinopathies.

The abnormal inflammatory response observed in inflammatory bowel disease (IBD), Crohn’s disease (CD), and ulcerative colitis (UC), involves interplay between environmental factors, the intestinal microbiota, and host genetic factors. Several genome-wide association studies have indicated the association between CD and UC and variants in genes encoding proteins involved in gut homeostasis, immunity, and host defense against microbes, reinforcing the hypothesis that commensal microbiota or infectious agents play a central role in IBD onset. In this chapter, we will talk about susceptibility genes related to autophagy, their role in the etiology of CD, and their complex interplay with the gut microbiota with a focus on adherent-invasive Escherichia coli (AIEC), which abnormally colonize the ileal mucosa of CD patients.

Mo1740 Defects in autophagy favour adherent-invasive E. coli persistence within macrophages leading to increased pro-inflammatory response. Digestive Disease Week (DDW)