Empéripolèse des cellules de lymphomes humains Ramos par les fibroblastes

Le fichier vidéo en format .avi accompagne mon document et correspond à l'annexe II. C'est une vidéomicroscopie dont la légende est mise en annexe II.Les fibroblastes (Fbs) constituent le type cellulaire dominant du stroma tumoral. Une vision "cafocentrique" du microenvironnement tumoral (MET) soutient que les interactions dynamiques et réciproques entre les cellules tumorales et les "cancer-associated fibroblasts" (CAFs) favoriseraient le potentiel métastasique des cellules tumorales. Notre principale hypothèse soutient qu'une interaction physique directe entre les cellules tumorales et les CAFs au sein du stroma tumoral est non seulement possible grâce au couple α4β1/VCAM-1 mais qu'elle pourrait éventuellement constituer une cible thérapeutique de choix. Grâce à la cytofluorométrie et aux anticorps bloquants, nous avons montré que des cellules de lymphome humain Ramos sont capables d'interagir avec les Fbs, que cette interaction est modulable et qu'elle implique le couple α4β1/VCAM-1. Une augmentation dose-dépendante de cette interaction ainsi que du niveau d'expression de VCAM-1 après une stimulation préalable des Fbs par le TNF-α ont été observés. Ce résultat a été confirmé par la surexpression de VCAM-1 par transfection. Des analyses par microscopie confocale ont révélé que les Ramos sont internalisés dans le cytoplasme des Fbs, phénomène connu sous le nom d'empéripolèse et qui correspond en partie à de la migration transcellulaire. Une colocalisation de la sous-unité α4 de α4β1 et de VCAM-1 à la jonction d'adhésion des Ramos aux Fbs ainsi que l'implication de la vimentine au niveau des cônes de migration ont été observées. La vidéomicroscopie a confirmé ce processus cellulaire actif. Finalement, un modèle mimétique de billes fluorescentes de polystyrène tapissées de α4β1 a confirmé les précédents résultats. Cette étude révèle que les Ramos peuvent interagir puis transmigrer à travers les Fbs grâce au couple α4β1/VCAM-1 de façon modulable. Tel que stipulé dans l'hypothèse du "Seed and Soil", nos résultats supportent l'hypothèse d'un adressage cellulaire et tissulaire dans la dissémination métastasique des cellules tumorales qui s'implanteraient préférentiellement là où l'expression de VCAM-1 a été induite. Le couple α4β1/VCAM-1 pourrait servir de cible thérapeutique de choix dans le développement de nouveaux antagonistes anticancéreux.Fibroblasts (Fbs) often represent the majority of stromal cells within tumors. A “cafocentric” view of the tumor microenvironment (MET) hypothesizes that the dynamic and reciprocal interactions between tumor cells and cancer-associated fibroblasts (CAFs) may enhance the metastatic potency of tumor cells. Very few studies investigated a physical interaction between Fbs and tumor cells within the tumor stroma. Using flow cytometry and blocking antibodies, we demonstrated that a human lymphoma cell line Ramos could interact with Fbs through the couple α4β1/VCAM-1. We observed a dose-dependent increased interaction after tumor necrosis factor-α (TNF-α) stimulation which correlated with increased VCAM-1 expression. A similar observation was made after VCAM-1 overexpression by transfection. Further investigations by confocal microscopy revealed the engulfment of Ramos into the cytoplasm of Fbs, a process called emperipolesis which mostly corresponds to transcellular migration and involves the clustering of the α4 sub-unit and VCAM-1 and the involvement of vimentin intermediate filaments at the transmigration cups. We visualized this cell-in-cell penetration by real time-lapse video-microscopy, which confirmed an active cellular process. Finally, we used α4β1 polystyrene coated-beads as a mimetic model to further confirm the precedent results. Our results showed that Ramos lymphoma cells can interact with and further transmigrate through Fbs by an active cellular process, the emperipolesis which involves the couple of receptors α4β1/VCAM-1. This crosstalk between Fbs and tumor cells through emperipolesis may modulate tumor cells escape from the primary tumor and support the "seed and soil" hypothesis as well as a possible involvement in tumor drug resistance. This fibroblastic permeability can be modulated by inflammation and the CAFs may react as endothelial cells and should be considered as a future target in the MET. Tumor cells may selectively search for new metastatic niches where VCAM-1 expression was already induced. Our results strengthen the growing idea that Fbs and tumor cell integrins may serve as suitable targets in the development of antagonist-based cancer therapy

Adhesion and transcellular migration of neutrophils and B lymphocytes on fibroblasts

During tissue inflammation, infiltrated leukocytes may have physical contacts with fibroblasts. We observed that neutrophils and B lymphocytes adhered in a larger proportion than T cells on cultured fibroblasts. Microscopy showed that adhesion was also characterized by leukocyte engulfment by the fibroblasts. In migration assays, only neutrophils and B lymphocytes were selectively able to migrate through a fibroblast barrier. Adhesion and migration were increased by stimulation with tumor necrosis factor-alpha (TNF-alpha) and phorbol-12-myristate-13-acetate (PMA). Antibodies against ICAM-1/beta2 integrin blocked the interaction of neutrophils to fibroblasts. For B lymphocytes the couple VCAM-1/alpha4 integrin was also involved in this interaction. Human skin fibroblasts presented similar adhesion characteristics as rat cardiac fibroblasts. By measuring the distance between the border of migration holes and cadherin-positive adherens junctions, more than 65% of the holes correspond to the transcellular route over the paracellular route. Furthermore, vimentin staining revealed that the migration holes were highly nested by intermediate filaments in accordance with the transcellular route. Our results demonstrated that engulfment of neutrophils and B lymphocytes by fibroblasts resulted in selective passage by a transcellular route.This work was supported by grants from the Canadian Institutes of Health Research and the Natural Sciences and Engineering Research Counci

Adaptive Fuzzy-PI Control for Speed’s Tracking in a Leisure Electrical Vehicle

This paper deals with a comparison study between different kinds of controllers based on a speed tracking problem inside an electrical vehicle using an axial-flux permanent-magnet traction motor. In fact, the framework of these works is the control of the powertrain of electric vehicles which is a great challenge to ensure driving comfort. To do so, the vehicle’s performances are carried out using an established simulation platform implemented on MatlabSimulink environment. Then, obtained results are compared for the three synthesized controllers. It has been found that required speed levels are reached with a clear advantage of the adaptive Fuzzy-PI controller versus the PI and the fuzzy counterparts, from the points of view of rise and selling times

Active Surveillance Program to Increase Awareness on Invasive Fungal Diseases: the French RESSIF Network (2012 to 2018)

International audienceThe French National Reference Center for Invasive Mycoses and Antifungals leads an active and sustained nationwide surveillance program on probable and proven invasive fungal diseases (IFDs) to determine their epidemiology in France. Between 2012 and 2018, a total of 10,886 IFDs were recorded. The incidence increased slightly over time (2.16 to 2.36/10,000 hospitalization days, P = 0.0562) in relation with an increase of fungemia incidence (1.03 to 1.19/10,000, P = 0.0023), while that of other IFDs remained stable. The proportion of ≥65-year-old patients increased from 38.4% to 45.3% (P 60% of the cases) with a global mortality rate of 42.5% and 59.3%, respectively, at 3 months and significant changes in diagnosis procedure over time. More concurrent infections were also diagnosed over time (from 5.4% to 9.4% for mold IFDs, P = 0.0115). In conclusion, we observed an aging of patients with IFD with a significant increase in incidence only for yeast fungemia, a trend toward more concurrent infections, which raises diagnostic and therapeutic issues. Overall, global survival associated with IFDs has not improved despite updated guidelines and new diagnostic tools.IMPORTANCE The epidemiology of invasive fungal diseases (IFDs) is hard to delineate given the difficulties in ascertaining the diagnosis that is often based on the confrontation of clinical and microbiological criteria. The present report underlines the interest of active surveillance involving mycologists and clinicians to describe the global incidence and that of the main IFDs. Globally, although the incidence of Pneumocystis pneumonia, invasive aspergillosis, and mucormycosis remained stable over the study period (2012 to 2018), that of yeast fungemia increased slightly. We also show here that IFDs seem to affect older people more frequently. The most worrisome observation is the lack of improvement in the global survival rate associated with IFDs despite the increasing use of more sensitive diagnostic tools, the availability of new antifungal drugs very active in clinical trials, and a still low/marginal rate of acquired in vitro resistance in France. Therefore, other tracks of improvement should be investigated actively

Rare predicted loss-of-function variants of type I IFN immunity genes are associated with life-threatening COVID-19

BackgroundWe previously reported that impaired type I IFN activity, due to inborn errors of TLR3- and TLR7-dependent type I interferon (IFN) immunity or to autoantibodies against type I IFN, account for 15-20% of cases of life-threatening COVID-19 in unvaccinated patients. Therefore, the determinants of life-threatening COVID-19 remain to be identified in similar to 80% of cases.MethodsWe report here a genome-wide rare variant burden association analysis in 3269 unvaccinated patients with life-threatening COVID-19, and 1373 unvaccinated SARS-CoV-2-infected individuals without pneumonia. Among the 928 patients tested for autoantibodies against type I IFN, a quarter (234) were positive and were excluded.ResultsNo gene reached genome-wide significance. Under a recessive model, the most significant gene with at-risk variants was TLR7, with an OR of 27.68 (95%CI 1.5-528.7, P=1.1x10(-4)) for biochemically loss-of-function (bLOF) variants. We replicated the enrichment in rare predicted LOF (pLOF) variants at 13 influenza susceptibility loci involved in TLR3-dependent type I IFN immunity (OR=3.70[95%CI 1.3-8.2], P=2.1x10(-4)). This enrichment was further strengthened by (1) adding the recently reported TYK2 and TLR7 COVID-19 loci, particularly under a recessive model (OR=19.65[95%CI 2.1-2635.4], P=3.4x10(-3)), and (2) considering as pLOF branchpoint variants with potentially strong impacts on splicing among the 15 loci (OR=4.40[9%CI 2.3-8.4], P=7.7x10(-8)). Finally, the patients with pLOF/bLOF variants at these 15 loci were significantly younger (mean age [SD]=43.3 [20.3] years) than the other patients (56.0 [17.3] years; P=1.68x10(-5)).ConclusionsRare variants of TLR3- and TLR7-dependent type I IFN immunity genes can underlie life-threatening COVID-19, particularly with recessive inheritance, in patients under 60 years old

Correction: Rare predicted loss-of-function variants of type I IFN immunity genes are associated with life-threatening COVID-19

International audienc

The risk of COVID-19 death is much greater and age dependent with type I IFN autoantibodies

International audienceSignificance There is growing evidence that preexisting autoantibodies neutralizing type I interferons (IFNs) are strong determinants of life-threatening COVID-19 pneumonia. It is important to estimate their quantitative impact on COVID-19 mortality upon SARS-CoV-2 infection, by age and sex, as both the prevalence of these autoantibodies and the risk of COVID-19 death increase with age and are higher in men. Using an unvaccinated sample of 1,261 deceased patients and 34,159 individuals from the general population, we found that autoantibodies against type I IFNs strongly increased the SARS-CoV-2 infection fatality rate at all ages, in both men and women. Autoantibodies against type I IFNs are strong and common predictors of life-threatening COVID-19. Testing for these autoantibodies should be considered in the general population

The risk of COVID-19 death is much greater and age dependent with type I IFN autoantibodies

International audienceSignificance There is growing evidence that preexisting autoantibodies neutralizing type I interferons (IFNs) are strong determinants of life-threatening COVID-19 pneumonia. It is important to estimate their quantitative impact on COVID-19 mortality upon SARS-CoV-2 infection, by age and sex, as both the prevalence of these autoantibodies and the risk of COVID-19 death increase with age and are higher in men. Using an unvaccinated sample of 1,261 deceased patients and 34,159 individuals from the general population, we found that autoantibodies against type I IFNs strongly increased the SARS-CoV-2 infection fatality rate at all ages, in both men and women. Autoantibodies against type I IFNs are strong and common predictors of life-threatening COVID-19. Testing for these autoantibodies should be considered in the general population