CCR7 Is Recruited to the Immunological Synapse, Acts as Co-stimulatory Molecule and Drives LFA-1 Clustering for Efficient T Cell Adhesion Through ZAP70

The chemokine receptor CCR7 guides T cells and dendritic cells to and within lymph nodes to launch the onset of adaptive immunity. Here, we demonstrate that CCR7 in addition acts as a potent co-stimulatory molecule in T cell activation. We found that antigen recognition and engagement of the TCR results in CCR7 accumulation at the immunological synapse where CCR7 and the TCR co-localize within sub-synaptic vesicles. We demonstrate that CCR7 triggering alone is sufficient to recruit and activate ZAP70, a critical kinase for T cell activation, through Src kinase, whereas TCR CCR7 co-stimulation results in increased and prolonged ZAP70 kinase activity. Finally, we show that ZAP70, acting as adapter molecule, is critical for CCR7-mediated inside-out signaling to integrins, thereby modulating LFA-1 valency regulation to promote cell adhesion, a key step in immunological synapse formation and efficient T cell activation

Early skewed differentiation and PD-1 expression in CD4+ cells relate to immune dysfunction and viral persistence in individuals living with HIV 1 year post-cART initiation

Achieving HIV functional cure is a priority. Strategies such as adoptive cell transfer have been assayed, without success yet mainly due to immune dysfunctions observed among individuals. Samples from 25 HIV+ subjects were collected at diagnosis (baseline sample, BSL) and one year post-cART initiation (post- cART ). At BSL, bulk and HIV-specific CD4 phenotype (CD45RO , CCR 7, CD95 and PD1 expression) was assessed by flow cytometry after a short stimulation with HIV peptides. Also, proportion of CD4+/HLA-DR+/ CD38+ cells was measured. At post-cART , HIV-specific CD8TC s were obtained after 2-week expansion with peptides. Phenotype and antiviral activity (VIA and VITA L assays) were evaluated post-expansion. Plasma CXCL10 (IP-10) was assessed by ELISA. Cell-associated HIV DNA (total and integrated) and unspliced (US) and multiply-spliced (MS) RNA were quantified by real-time PCR. Non-parametric statistics were applied. Early CD4TC exhaustion, elevated activation and inadequate differentiation seem to be associated with viral persistence, inflammation, as well as with the phenotype and antiviral capacity of HIV-specific CD8TC s that persist one year after cART is initiated. These parameters could serve as predictors of CD8TC function on treated subjects.Fil: Salido, Jimena Patricia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Czernikier, Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Trifone, César Ariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Figueroa, M. I.. Fundación Huésped; ArgentinaFil: Salomon, Horacio Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Cahn, P.. Fundación Huésped; ArgentinaFil: Sued, O.. Fundación Huésped; ArgentinaFil: Laufer, Natalia Lorna. Gobierno de la Ciudad de Buenos Aires. Hospital General de Agudos "Juan A. Fernández"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Ghiglione, Yanina Alexandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Turk, Gabriela Julia Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; Argentin

Epithelial chemokine CXCL14 synergizes with CXCL12 via allosteric modulation of CXCR4

The chemokine receptor, CXC chemokine receptor 4 (CXCR4), is selective for CXC chemokine ligand 12 (CXCL12), is broadly expressed in blood and tissue cells, and is essential during embryogenesis and hematopoiesis. CXCL14 is a homeostatic chemokine with unknown receptor selectivity and preferential expression in peripheral tissues. Here, we demonstrate that CXCL14 synergized with CXCL12 in the induction of chemokine responses in primary human lymphoid cells and cell lines that express CXCR4. Combining subactive concentrations of CXCL12 with 100–300 nM CXCL14 resulted in chemotaxis responses that exceeded maximal responses that were obtained with CXCL12 alone. CXCL14 did not activate CXCR4-expressing cells (i.e., failed to trigger chemotaxis and Ca2+ mobilization, as well as signaling via ERK1/2 and the small GTPase Rac1); however, CXCL14 bound to CXCR4 with high affinity, induced redistribution of cell-surface CXCR4, and enhanced HIV-1 infection by >3-fold. We postulate that CXCL14 is a positive allosteric modulator of CXCR4 that enhances the potency of CXCR4 ligands. Our findings provide new insights that will inform the development of novel therapeutics that target CXCR4 in a range of diseases, including cancer, autoimmunity, and HIV.—Collins, P. J., McCully, M. L., Mart´ınez-Muñoz, L., Santiago, C.,Wheeldon, J., Caucheteux, S., Thelen, S., Cecchinato, V., Laufer, J.M., Purvanov, V.,Monneau, Y. R., Lortat-Jacob, H., Legler, D. F., Uguccioni, M., Thelen, M., Piguet, V., Mellado, M., Moser, B. Epithelial chemokine CXCL14 synergizes with CXCL12 via allosteric modulation of CXCR4. FASEB J. 31, 000–000 (2017). www.fasebj.or

A Possible Sterilizing Cure of HIV-1 Infection Without Stem Cell Transplantation

Background: A sterilizing cure of HIV-1 infection has been reported in 2 persons living with HIV-1 who underwent allogeneic hematopoietic stem cell transplantations from donors who were homozygous for the CCR5D32 gene polymorphism. However, this has been considered elusive during natural infection. Objective: To evaluate persistent HIV-1 reservoir cells in an elite controller with undetectable HIV-1 viremia for more than 8 years in the absence of antiretroviral therapy. Design: Detailed investigation of virologic and immunologic characteristics. Setting: Tertiary care centers in Buenos Aires, Argentina, and Boston, Massachusetts. Patient: A patient with HIV-1 infection and durable drug-free suppression of HIV-1 replication. Measurements: Analysis of genome-intact and replication-competent HIV-1 using near-full-length individual proviral sequencing and viral outgrowth assays, respectively; analysis of HIV-1 plasma RNA by ultrasensitive HIV-1 viral load testing. Results: No genome-intact HIV-1 proviruses were detected in analysis of a total of 1.188 billion peripheral blood mononuclear cells and 503 million mononuclear cells from placental tissues. Seven defective proviruses, some of them derived from clonally expanded cells, were detected. A viral outgrowth assay failed to retrieve replication-competent HIV-1 from 150 million resting CD4+ T cells. No HIV-1 RNA was detected in 4.5 mL of plasma. Limitations: Absence of evidence for intact HIV-1 proviruses in large numbers of cells is not evidence of absence of intact HIV-1 proviruses. A sterilizing cure of HIV-1 can never be empirically proved. Conclusion: Genome-intact and replication-competent HIV-1 were not detected in an elite controller despite analysis of massive numbers of cells from blood and tissues, suggesting that this patient may have naturally achieved a sterilizing cure of HIV-1 infection. These observations raise the possibility that a sterilizing cure may be an extremely rare but possible outcome of HIV-1 infection.Fil: Turk, Gabriela Julia Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Seiger, Kyra. Massachusetts Institute of Technology; Estados UnidosFil: Lian, Xiaodong. Massachusetts Institute of Technology; Estados UnidosFil: Sun, Weiwei. Massachusetts Institute of Technology; Estados UnidosFil: Parsons, Elizabeth M.. Massachusetts Institute of Technology; Estados UnidosFil: Gao, Ce. Massachusetts Institute of Technology; Estados UnidosFil: Rassadkina, Yelizaveta. Massachusetts Institute of Technology; Estados UnidosFil: Polo, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Czernikier, Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Ghiglione, Yanina Alexandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Vellicce, Alejandra Fabiana. Universidad de Buenos Aires. Facultad de Medicina. Hospital de Clínicas General San Martín; ArgentinaFil: Varriale, Joseph. No especifíca;Fil: Lai, Jun. No especifíca;Fil: Yuki, Yuko. No especifíca;Fil: Martin, Maureen. No especifíca;Fil: Rhodes, Ajantha. University of Melbourne; AustraliaFil: Lewin, Sharon R.. University of Melbourne; Australia. Monash University; AustraliaFil: Walker, Bruce D.. Massachusetts Institute of Technology; Estados UnidosFil: Carrington, Mary. Massachusetts Institute of Technology; Estados UnidosFil: Siliciano, Robert. No especifíca;Fil: Siliciano, Janet. No especifíca;Fil: Lichterfeld, Mathias. Massachusetts Institute of Technology; Estados UnidosFil: Laufer, Natalia Lorna. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Yu, Xu G.. Massachusetts Institute of Technology; Estados Unido

Cross-species oncogenomics offers insight into human muscle-invasive bladder cancer

Background In humans, muscle-invasive bladder cancer (MIBC) is highly aggressive and associated with a poor prognosis. With a high mutation load and large number of altered genes, strategies to delineate key driver events are necessary. Dogs and cats develop urothelial carcinoma (UC) with histological and clinical similarities to human MIBC. Cattle that graze on bracken fern also develop UC, associated with exposure to the carcinogen ptaquiloside. These species may represent relevant animal models of spontaneous and carcinogen-induced UC that can provide insight into human MIBC. Results Whole-exome sequencing of domestic canine (n = 87) and feline (n = 23) UC, and comparative analysis with human MIBC reveals a lower mutation rate in animal cases and the absence of APOBEC mutational signatures. A convergence of driver genes (ARID1A, KDM6A, TP53, FAT1, and NRAS) is discovered, along with common focally amplified and deleted genes involved in regulation of the cell cycle and chromatin remodelling. We identify mismatch repair deficiency in a subset of canine and feline UCs with biallelic inactivation of MSH2. Bovine UC (n = 8) is distinctly different; we identify novel mutational signatures which are recapitulated in vitro in human urinary bladder UC cells treated with bracken fern extracts or purified ptaquiloside. Conclusion Canine and feline urinary bladder UC represent relevant models of MIBC in humans, and cross-species analysis can identify evolutionarily conserved driver genes. We characterize mutational signatures in bovine UC associated with bracken fern and ptaquiloside exposure, a human-linked cancer exposure. Our work demonstrates the relevance of cross-species comparative analysis in understanding both human and animal UC

Evaluating the Benefits of Aphasia Intervention Delivered in Virtual Reality: Results of a Quasi-Randomised Study

Introduction This study evaluated an intervention for people with aphasia delivered in a novel virtual reality platform called EVA Park. EVA Park contains a number of functional and fantastic locations and allows for interactive communication between multiple users. Twenty people with aphasia had 5 weeks’ intervention, during which they received daily language stimulation sessions in EVA Park from a support worker. The study employed a quasi randomised design, which compared a group that received immediate intervention with a waitlist control group. Outcome measures explored the effects of intervention on communication and language skills, communicative confidence and feelings of social isolation. Compliance with the intervention was also explored through attrition and usage data. Results There was excellent compliance with the intervention, with no participants lost to follow up and most (18/20) receiving at least 88% of the intended treatment dose. Intervention brought about significant gains on a measure of functional communication. Gains were achieved by both groups of participants, once intervention was received, and were well maintained. Changes on the measures of communicative confidence and feelings of social isolation were not achieved. Results are discussed with reference to previous aphasia therapy findings

Immune Response to SARS-CoV-2 Third Vaccine in Patients With Rheumatoid Arthritis Who Had No Seroconversion After Primary 2-Dose Regimen With Inactivated or Vector-Based Vaccines

Objective. The aim of this study was to assess the immune response after a third dose of SARS-CoV-2 vaccine in patients with rheumatoid arthritis (RA) with undetectable antibody titers after the primary regimen of 2 doses. Methods. Patients with RA with no seroconversion after 2 doses of SARS-CoV-2 vaccine and who received a third dose of either an mRNA or vector-based vaccine were included. Anti-SARS-CoV-2 IgG antibodies, neutralizing activity, and T cell responses were assessed after the third dose. Results. A total of 21 nonresponder patients were included. At the time of vaccination, 29% were receiving glucocorticoids and 85% biologic disease-modifying antirheumatic drugs (including 6 taking abatacept [ABA] and 4 taking rituximab [RTX]). The majority (95%) received the BNT162b2 vaccine and only one of them received the ChAdOx1 nCoV-19 vaccine. After the third dose, 91% of the patients presented detectable anti-SARS-CoV-2 IgG and 76% showed neutralizing activity. Compared to other treatments, ABA and RTX were associated with the absence of neutralizing activity in 4 out of 5 (80%) patients and lower titers of neutralizing antibodies (median 3, IQR 0-20 vs 8, IQR 4-128; P = 0.20). Specific T cell response was detected in 41% of all patients after the second dose, increasing to 71% after the third dose. The use of ABA was associated with a lower frequency of T cell response (33% vs 87%, P = 0.03). Conclusion. In this RA cohort, 91% of patients who failed to seroconvert after 2 doses of SARS-CoV-2 vaccine presented detectable anti-SARS-CoV-2 IgG after a third dose. The use of ABA was associated with a lower frequency of specific T cell response.Fil: Isnardi, Carolina A.. No especifíca;Fil: Cerda, Osvaldo L.. No especifíca;Fil: Landi, Margarita. Austral University Hospital; LiberiaFil: Cruces, Leonel Hernán. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Schneeberger, Emilce E.. No especifíca;Fil: Montoro, Claudia Calle. Austral University Hospital; LiberiaFil: Alfaro, María Agustina. No especifíca;Fil: Roldán, Brian M.. No especifíca;Fil: Gómez Vara, Andrea B.. No especifíca;Fil: Giorgis, Pamela. No especifíca;Fil: Ezquer, Roberto Alejandro. No especifíca;Fil: Crespo Rocha, María G. No especifíca;Fil: Reyes Gómez, Camila R.. No especifíca;Fil: de Los Ángeles Correa, Mária. No especifíca;Fil: Rosemffet, Marcos G.. No especifíca;Fil: Abarza, Virginia Carrizo. No especifíca;Fil: Pellet, Santiago Catalan. Austral University Hospital; LiberiaFil: Perandones, Miguel. No especifíca;Fil: Reimundes, Cecilia. Austral University Hospital; LiberiaFil: Longueira, Yesica Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Turk, Gabriela Julia Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Quiroga, María Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Laufer, Natalia Lorna. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Quintana, Rosana Maris. No especifíca;Fil: de la Vega, María Celina. No especifíca;Fil: Kreplak, Nicolás. No especifíca;Fil: Pifano, Marina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Maid, Pablo. Austral University Hospital; LiberiaFil: Pons Estel, Guillermo J.. No especifíca;Fil: Citera, Gustavo. No especifíca

Beyond migration : Chemokines in lymphocyte priming, differentiation, and modulating effector functions

Chemokines and their receptors coordinate the positioning of leukocytes, and lymphocytes in particular, in space and time. Discrete lymphocyte subsets, depending on their activation and differentiation status, express various sets of chemokine receptors to be recruited to distinct tissues. Thus, the network of chemokines and their receptors ensures the correct localization of specialized lymphocyte subsets within the appropriate microenvironment enabling them to search for cognate antigens, to become activated, and to fulfill their effector functions. The chemokine system therefore is vital for the initiation as well as the regulation of immune responses to protect the body from pathogens while maintaining tolerance towards self. Besides the well investigated function of orchestrating directed cell migration, chemokines additionally act on lymphocytes in multiple ways to shape immune responses. In this review, we highlight and discuss the role of chemokines and chemokine receptors in controlling cell-to-cell contacts required for lymphocyte arrest on endothelial cells and immunological synapse formation, in lymphocyte priming and differentiation, survival, as well as in modulating effector functions.publishe

ZAP70 expression enhances chemokine-driven chronic lymphocytic leukemia cell migration and arrest by valency regulation of integrins.

The ζ-associated protein of 70 kDa (ZAP70) is expressed in the aggressive form of B-cell chronic lymphocytic leukemia (CLL). Moreover, the integrin very late antigen (VLA)-1 is highly expressed on subtypes of CLL that are associated with high proliferation rates in the lymph node context. We herein identify a critical role for ZAP70 in chemokine-mediated, inside-out signaling to integrins in trisomy 12 carrying Ohio State University-CLL cell lines derived from a patient with previously treated CLL. We found that ZAP70-positive CLL cells migrated significantly better toward ligands of the lymph node homing chemokine receptors CCR7 and CXCR4 compared with ZAP70-negative cells. In addition, ZAP70-expressing CLL cells adhered more efficiently to integrin ligands under static conditions. We discovered that ZAP70 expression controls chemokine-driven clustering of the integrins VLA-4 and lymphocyte function-associated antigen-1. More precisely, chemokine stimulation resulted in a ZAP70-dependent integrin valency regulation on CLL cells, whereas high-affinity regulation of integrins was independent of ZAP70. Consequently, ZAP70-expressing CLL cells show increased chemokine-driven arrest on immobilized integrin ligands and on chemokine-presenting endothelial cells under physiologic flow conditions. Hence, we describe a novel mechanism showing how ZAP70 controls chemokine-driven valency regulation of integrins and arrest of CLL cells on endothelial cells, a process that might contribute to CLL disease progression.-Laufer, J. M., Lyck, R., Legler, D. F. ZAP70 expression enhances chemokine-driven chronic lymphocytic leukemia cell migration and arrest by valency regulation of integrins