Analyse des lymphocytes T non conventionnels à TCRyo, MAIT et NKT comme réservoir potentiel du VIH

The persistence of the human immunodeficiency virus (HIV) in infected individuals undergoing treatment is a major characteristic of the infection and the main obstacle to achieving a cure. Therefore, a better understanding of the lymphocyte populations forming the cellular reservoir of HIV is still highly needed. The participation of conventional T cells with TCRαβ is already well described in the literature. This thesis aims to explore non-conventional invariant or semi-invariant TCR cells as potential HIV targets. To this end we: i) analyzed their concentration in blood according to the clinical profiles of people living with HIV (PLHIV), ii) assessed the proportion of unconventional lymphocytes expressing CD4, CCR5, and CXCR4 receptors, as well as the density of these HIV entry receptors, iii) validated the digital PCR (dPCR) approach and we have showed that dPCR is more accurate than real-time qPCR (qPCR) for HIV DNA quantification.The expression of the CD4 receptor and the HIV co-receptors, CCR5 and CXCR4, was studied in TCRVα7.2 cells described according to their co-expression of the CD161 marker into CD161-, CD161+ or MAIT-like and CD161bright or MAIT cells, NKT cells and TCRγδ cells divided into three subpopulations: Vδ1, Vδ2, and Vδ* (representing neither δ1 nor δ2 cells, describing the Vδ3 and Vδ5 cells). This comparative study demonstrated preferential depletion of the Vδ2 LTγδ populations and CD4 MAIT cells, while other subpopulations were found to be more frequent under effective treatment: no-Vδ1 and no-Vδ2 LTγδ and TCRVα7.2+ CD161-. These TCRVα7.2+ CD161- cells and Vδ* LTγδ are two populations with a high potential of participation in the HIV reservoir due to their expression of CD4 and the co-receptors CCR5 and CXCR4.The relative quantification of the HIV reservoir in these cells requires a more accurate technique than qPCR. A dPCR technique was developed and compared analytically to the qPCR technique used routinely in the laboratory. Because of its low sample volume requirement and better reproducibility, dPCR will allow us to more accurately quantify HIV DNA in TCRVα7.2+ CD161- and LTγδ Vδ* cells to determine their participation in the HIV reservoir.La persistance du virus de l’immunodéficience humaine (VIH) chez des sujets infectés sous traitement est une caractéristique majeure de l’infection et l’obstacle principal à la guérison. Une meilleure connaissance des populations lymphocytaires formant le réservoir cellulaire du VIH demeure nécessaire. La participation des lymphocytes T conventionnels à TCRαβ est déjà bien décrite dans la littérature. Le but de ce travail de thèse était d’explorer les cellules non conventionnelles à TCR invariant ou semi-invariants, comme cible potentielle du VIH. Dans cet objectif nous avons : i) analysé leur concentration dans le sang suivant les profils cliniques des personnes vivant avec le VIH (PVVIH), ii) évalué la part des lymphocytes non conventionnels qui expriment les récepteurs CD4, CCR5 et CXCR4, ainsi que la densité de ces récepteurs d’entrée du VIH, iii) validé une approche de PCR digitale (dPCR) plus précise que la PCR en temps réel (qPCR) pour la quantification de l’ADN VIH.L’expression du récepteur CD4 et des co-récepteurs du VIH, CCR5 et CXCR4, a été étudié dans les cellules à TCRVα7.2 décrites selon leur co-expression du marqueur CD161 en : CD161-, CD161+ ou MAIT-like et CD161bright ou MAIT, les cellules NKT et les cellules à TCRγδ divisées en sous-populations Vδ1, Vδ2 et Vδ* représentant les cellules ni δ1 ni δ2, décrivant les cellules Vδ3 et Vδ5. Cette étude comparative a mis en évidence la déplétion préférentielle des populations de LTγδ Vδ2, de cellules MAIT CD4, tandis que d’autres sous populations se trouvaient plus fréquentes sous traitement efficace : LTγδ non Vδ1 et non Vδ2 et TCRVα7.2+ CD161-. Ces cellules TCRVα7.2+ CD161- et les LTγδ Vδ* constituent des populations à fort potentiel de participation au réservoir du VIH en raison de leur expression du CD4 et des corécepteurs CCR5 et CXCR4.La quantification relative du réservoir VIH dans ces cellules nécessite une technique plus précise que la qPCR. Une technique de dPCR a été mise au point et comparée analytiquement à la technique de qPCR utilisée en routine au laboratoire. La dPCR par son besoin d’un faible volume d’échantillon et sa meilleure reproductibilité, nous permettra une quantification plus juste de l’ADN VIH dans les cellules TCRVα7.2+ CD161- et les LTγδ Vδ* afin de pouvoir déterminer leur participation au réservoir du VIH

Characterization of the HIV reservoir in cells with an invariant or semi-invariant TCR

La persistance du virus de l’immunodéficience humaine (VIH) chez des sujets infectés sous traitement est une caractéristique majeure de l’infection et l’obstacle principal à la guérison. Une meilleure connaissance des populations lymphocytaires formant le réservoir cellulaire du VIH demeure nécessaire. La participation des lymphocytes T conventionnels à TCRαβ est déjà bien décrite dans la littérature. Le but de ce travail de thèse était d’explorer les cellules non conventionnelles à TCR invariant ou semi-invariants, comme cible potentielle du VIH. Dans cet objectif nous avons : i) analysé leur concentration dans le sang suivant les profils cliniques des personnes vivant avec le VIH (PVVIH), ii) évalué la part des lymphocytes non conventionnels qui expriment les récepteurs CD4, CCR5 et CXCR4, ainsi que la densité de ces récepteurs d’entrée du VIH, iii) validé une approche de PCR digitale (dPCR) plus précise que la PCR en temps réel (qPCR) pour la quantification de l’ADN VIH.L’expression du récepteur CD4 et des co-récepteurs du VIH, CCR5 et CXCR4, a été étudié dans les cellules à TCRVα7.2 décrites selon leur co-expression du marqueur CD161 en : CD161-, CD161+ ou MAIT-like et CD161bright ou MAIT, les cellules NKT et les cellules à TCRγδ divisées en sous-populations Vδ1, Vδ2 et Vδ* représentant les cellules ni δ1 ni δ2, décrivant les cellules Vδ3 et Vδ5. Cette étude comparative a mis en évidence la déplétion préférentielle des populations de LTγδ Vδ2, de cellules MAIT CD4, tandis que d’autres sous populations se trouvaient plus fréquentes sous traitement efficace : LTγδ non Vδ1 et non Vδ2 et TCRVα7.2+ CD161-. Ces cellules TCRVα7.2+ CD161- et les LTγδ Vδ* constituent des populations à fort potentiel de participation au réservoir du VIH en raison de leur expression du CD4 et des corécepteurs CCR5 et CXCR4.La quantification relative du réservoir VIH dans ces cellules nécessite une technique plus précise que la qPCR. Une technique de dPCR a été mise au point et comparée analytiquement à la technique de qPCR utilisée en routine au laboratoire. La dPCR par son besoin d’un faible volume d’échantillon et sa meilleure reproductibilité, nous permettra une quantification plus juste de l’ADN VIH dans les cellules TCRVα7.2+ CD161- et les LTγδ Vδ* afin de pouvoir déterminer leur participation au réservoir du VIH.The persistence of the human immunodeficiency virus (HIV) in infected individuals undergoing treatment is a major characteristic of the infection and the main obstacle to achieving a cure. Therefore, a better understanding of the lymphocyte populations forming the cellular reservoir of HIV is still highly needed. The participation of conventional T cells with TCRαβ is already well described in the literature. This thesis aims to explore non-conventional invariant or semi-invariant TCR cells as potential HIV targets. To this end we: i) analyzed their concentration in blood according to the clinical profiles of people living with HIV (PLHIV), ii) assessed the proportion of unconventional lymphocytes expressing CD4, CCR5, and CXCR4 receptors, as well as the density of these HIV entry receptors, iii) validated the digital PCR (dPCR) approach and we have showed that dPCR is more accurate than real-time qPCR (qPCR) for HIV DNA quantification.The expression of the CD4 receptor and the HIV co-receptors, CCR5 and CXCR4, was studied in TCRVα7.2 cells described according to their co-expression of the CD161 marker into CD161-, CD161+ or MAIT-like and CD161bright or MAIT cells, NKT cells and TCRγδ cells divided into three subpopulations: Vδ1, Vδ2, and Vδ* (representing neither δ1 nor δ2 cells, describing the Vδ3 and Vδ5 cells). This comparative study demonstrated preferential depletion of the Vδ2 LTγδ populations and CD4 MAIT cells, while other subpopulations were found to be more frequent under effective treatment: no-Vδ1 and no-Vδ2 LTγδ and TCRVα7.2+ CD161-. These TCRVα7.2+ CD161- cells and Vδ* LTγδ are two populations with a high potential of participation in the HIV reservoir due to their expression of CD4 and the co-receptors CCR5 and CXCR4.The relative quantification of the HIV reservoir in these cells requires a more accurate technique than qPCR. A dPCR technique was developed and compared analytically to the qPCR technique used routinely in the laboratory. Because of its low sample volume requirement and better reproducibility, dPCR will allow us to more accurately quantify HIV DNA in TCRVα7.2+ CD161- and LTγδ Vδ* cells to determine their participation in the HIV reservoir

Accuracy of real-time PCR and digital PCR for the monitoring of total HIV DNA under prolonged antiretroviral therapy

International audienceAbstract Total HIV DNA is a standard marker to monitor the HIV reservoir in people living with HIV. We investigated HIV DNA quantification accuracy by a real-time PCR kit (qPCR) and digital PCR (dPCR) method within the same set of primers and probes. Among 48 aviremic patients followed for up to 7 years with qPCR, the mean coefficient of variation of total HIV DNA between two successive measurements was 77% (± 0.42log 10 HIVDNA copies/10 6 PBMC). The total HIV DNA quantified by the two PCR methods has a high correlation (0.99 and 0.83, for 8E5 and PLHIV samples, respectively), but we observed better repeatability and reproducibility of the dPCR compared to the qPCR (CV of 11.9% vs. 24.7% for qPCR, p-value = 0.024). Furthermore, we highlighted a decay of the number of HIV copies in the 8E5 cell line qPCR standard over time (from 0.73 to 0.43 copies per cell), contributing to variations of HIV DNA results in patients whose HIV reservoir should be theoretically stabilized. Our study highlighted that absolute quantification of total HIV DNA by dPCR allows more accurate monitoring of the HIV reservoir than qPCR in patients under prolonged antiretroviral therapy

Th17 CD4+ T-Cell as a Preferential Target for HIV Reservoirs

International audienceAmong CD4+ T-cells, T helper 17 (Th17) cells play a sentinel role in the defense against bacterial/fungal pathogens at mucosal barriers. However, Th17 cells are also highly susceptible to HIV-1 infection and are rapidly depleted from gut mucosal sites, causing an imbalance of the Th17/Treg ratio and impairing cytokines production. Consequently, damage to the gut mucosal barrier leads to an enhanced microbial translocation and systemic inflammation, a hallmark of HIV-1 disease progression. Th17 cells’ expression of mucosal homing receptors (CCR6 and α4β7), as well as HIV receptors and co-receptors (CD4, α4β7, CCR5, and CXCR4), contributes to susceptibility to HIV infection. The up-regulation of numerous intracellular factors facilitating HIV production, alongside the downregulation of factors inhibiting HIV, helps to explain the frequency of HIV DNA within Th17 cells. Th17 cells harbor long-lived viral reservoirs in people living with HIV (PLWH) receiving antiretroviral therapy (ART). Moreover, cell longevity and the proliferation of a fraction of Th17 CD4 T cells allow HIV reservoirs to be maintained in ART patients

Unexpected effects of sublethal doses of insecticide on the peripheral olfactory response and sexual behavior in a pest insect

International audiencePesticides have long been used as the main solution to limit agricultural pests, but their widespread use resulted in chronic or diffuse environmental pollutions, development of insect resistances, and biodiversity reduction. The effects of low residual doses of these chemical products on organisms that affect both targeted species (crop pests) but also beneficial insects became a major concern, particularly because low doses of pesticides can induce unexpected positive-also called hormetic-effects on insects, leading to surges in pest population growth at greater rate than what would have been observed without pesticide application. The present study aimed to examine the effects of sublethal doses of deltamethrin, one of the most used synthetic pyrethroids, known to present a residual activity and persistence in the environment, on the peripheral olfactory system and sexual behavior of a major pest insect, the cotton leafworm Spodoptera littoralis. We highlighted here a hormetic effect of sublethal dose of deltamethrin on the male responses to sex pheromone, without any modification of their response to host-plant odorants. We also identified several antennal actors potentially involved in this hormetic effect and in the antennal detoxification or antennal stress response of/to deltamethrin exposur

Interglaciaires pléistocènes en domaine karstique méditerranéen. Interprétation de la sédimentation et de la biodiversité fossile à la Caune de l’Arago (Tautavel, Pyrénées-Orientales)

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Interglaciaires pléistocènes en domaine karstique méditerranéen. Interprétation de la sédimentation et de la biodiversité fossile à la Caune de l’Arago (Tautavel, Pyrénées-Orientales)

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Monocyte CD169 expression as a biomarker in the early diagnosis of COVID-19

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Erratum to: Monocyte CD169 Expression as a Biomarker in the Early Diagnosis of Coronavirus Disease 2019

International audienceAbstract We assessed the expression of CD169, a type I interferon-inducible receptor, on monocytes (monocyte CD169 [mCD169]) in 53 adult patients admitted to the hospital during the coronavirus disease 2019 (COVID-19) outbreak for a suspicion of severe acute respiratory syndrome coronavirus 2 infection. Monocyte CD169 was strongly overexpressed in 30 of 32 (93.7%) confirmed COVID-19 cases, compared with 3 of 21 (14.3%) patients in whom the diagnosis of COVID-19 was finally ruled out. Monocyte CD169 was associated with the plasma interferon-alpha level and thrombocytopenia. Monocyte CD169 testing may be helpful for the rapid triage of suspected COVID-19 patients during an outbreak