RESUMEN
Integrins are cell surface receptors that mediate the interactions of cells with their surroundings and play essential roles in cell adhesion, migration, and homeostasis. Eight of the 24 integrins bind to the tripeptide Arg-Gly-Asp (RGD) motif in their extracellular ligands, comprising the RGD-binding integrin subfamily. Despite similarity in recognizing the RGD motif and some redundancy, these integrins can selectively recognize RGD-containing ligands to fulfill specific functions in cellular processes. Antibodies against individual RGD-binding integrins are desirable for investigating their specific functions, and were selected here from a synthetic yeast-displayed Fab library. We discovered 11 antibodies that exhibit high specificity and affinity toward their target integrins, i.e. αVß3, αVß5, αVß6, αVß8, and α5ß1. Of these, six are function-blocking antibodies and contain a ligand-mimetic R(G/L/T)D motif in their CDR3 sequences. We report antibody-binding specificity, kinetics, and binding affinity for purified integrin ectodomains, as well as intact integrins on the cell surface. We further used these antibodies to reveal binding preferences of the αV subunit for its 5 ß-subunit partners: ß6 = ß8 > ß3 > ß1 = ß5.
Asunto(s)
Saccharomyces cerevisiae , Humanos , Saccharomyces cerevisiae/genética , Cadenas beta de Integrinas/inmunología , Cadenas beta de Integrinas/química , Cadenas beta de Integrinas/metabolismo , Cadenas beta de Integrinas/genética , Integrina alfaV/inmunología , Integrina alfaV/metabolismo , Integrinas/inmunología , Integrinas/metabolismo , Biblioteca de Péptidos , Técnicas de Visualización de Superficie Celular , Unión Proteica , Especificidad de AnticuerposRESUMEN
Sarcospan (SSPN) is a 25-kDa transmembrane protein that is broadly expressed at the cell surface of many tissues, including, but not limited to, the myofibers from skeletal and smooth muscles, cardiomyocytes, adipocytes, kidney epithelial cells, and neurons. SSPN is a core component of the dystrophin-glycoprotein complex (DGC) that links the intracellular actin cytoskeleton with the extracellular matrix. It is also associated with integrin α7ß1, the predominant integrin expressed in skeletal muscle. As a tetraspanin-like protein with four transmembrane spanning domains, SSPN functions as a scaffold to facilitate protein-protein interactions at the cell membrane. Duchenne muscular dystrophy, Becker muscular dystrophy, and X-linked dilated cardiomyopathy are caused by the loss of dystrophin at the muscle cell surface and a concomitant loss of the entire DGC, including SSPN. SSPN overexpression ameliorates Duchenne muscular dystrophy in the mdx murine model, which supports SSPN being a viable therapeutic target. Other rescue studies support SSPN as a biomarker for the proper assembly and membrane expression of the DGC. Highly specific and robust antibodies to SSPN are needed for basic research on the molecular mechanisms of SSPN rescue, pre-clinical studies, and biomarker evaluations in human samples. The development of SSPN antibodies is challenged by the presence of its four transmembrane domains and limited antigenic epitopes. To address the significant barrier presented by limited commercially available antibodies, we aimed to generate a panel of robust SSPN-specific antibodies that can serve as a resource for the research community. We created antibodies to three SSPN protein epitopes, including the intracellular N- and C-termini as well as the large extracellular loop (LEL) between transmembrane domains 3 and 4. We developed a panel of rabbit antibodies (poly- and monoclonal) against an N-terminal peptide fragment of SSPN. We used several assays to show that the rabbit antibodies recognize mouse SSPN with a high functional affinity and specificity. We developed mouse monoclonal antibodies against the C-terminal peptide and the large extracellular loop of human SSPN. These antibodies are superior to commercially available antibodies and outperform them in various applications, including immunoblotting, indirect immunofluorescence analysis, immunoprecipitation, and an ELISA. These newly developed antibodies will significantly improve the quality and ease of SSPN detection for basic and translational research.
Asunto(s)
Proteínas de la Membrana , Investigación Biomédica Traslacional , Animales , Humanos , Ratones , Distrofina/metabolismo , Distrofina/inmunología , Distrofina/genética , Integrinas/metabolismo , Integrinas/inmunología , Proteínas de la Membrana/inmunología , Proteínas de la Membrana/metabolismo , Distrofia Muscular de Duchenne/inmunología , Distrofia Muscular de Duchenne/metabolismoRESUMEN
Immune system recognizes invading microbes at both pathogen and antigen levels. Toll-like receptors (TLRs) play a key role in the first-line defense against pathogens. Major functions of TLRs include cytokine and chemokine production. TLRs share common downstream signaling pathways with other receptors. The crosstalk revolving around TLRs is rather significant and complex, underscoring the intricate nature of immune system. The profiles of produced cytokines and chemokines via TLRs can be affected by other receptors. Integrins are critical heterodimeric adhesion molecules expressed on many different cells. There are studies describing synergetic or inhibitory interplay between TLRs and integrins. Thus, we reviewed the crosstalk between TLRs and integrins. Understanding the nature of the crosstalk could allow us to modulate TLR functions via integrins.
Asunto(s)
Integrinas , Receptor Cross-Talk , Transducción de Señal , Receptores Toll-Like , Humanos , Receptores Toll-Like/metabolismo , Integrinas/metabolismo , Integrinas/inmunología , Animales , Citocinas/metabolismo , Inmunidad InnataRESUMEN
Background: It has been well documented that Takayasu arteritis (TAK) and ulcerative colitis (UC) coexist in the same patients. HLA-B*52 characterizes the co-occurrence, which is one of the common genetic features between these two diseases, indicating shared underlying pathologic mechanisms. Anti-integrin αvß6 antibody (Ab) is present in sera of UC patients in a highly specific manner. We investigated if there were any associations between anti-integrin αvß6 Ab and TAK, considering the risk HLA alleles. Methods: A total of 227 Japanese TAK patients were recruited in the current study and their serum samples were subjected to measurement of anti-integrin αvß6 Ab by ELISA. The clinical information, including the co-occurrence of UC, was collected. The HLA allele carrier status was determined by Luminex or genotype imputation. Results: The information about the presence of UC was available for 165 patients, among which eight (4.84%) patients had UC. Anti-integrin αvß6 antibody was identified in 7 out of 8 TAK subjects with UC (87.5%) while only 5 out of 157 (3.18%) TAK subjects without UC had the antibody (OR 121, p=7.46×10-8). A total of 99 out of 218 (45.4%) patients were HLA-B*52 carriers. There was no significant association between the presence of anti-integrin αvß6 Ab and HLA-B*52 carrier status in those without UC (OR 2.01, 95% CI 0.33-12.4, p = 0.189). Conclusions: The prevalence of anti-integrin αvß6 Ab was high in TAK patients with UC, but not in the absence of concomitant UC. The effect of HLA-B*52 on anti-integrin αvß6 Ab production would be minimal.
Asunto(s)
Antígenos de Neoplasias , Colitis Ulcerosa , Integrinas , Arteritis de Takayasu , Humanos , Colitis Ulcerosa/inmunología , Colitis Ulcerosa/genética , Arteritis de Takayasu/inmunología , Arteritis de Takayasu/genética , Femenino , Integrinas/inmunología , Masculino , Adulto , Persona de Mediana Edad , Antígenos de Neoplasias/inmunología , Antígenos de Neoplasias/genética , Antígeno HLA-B52/inmunología , Antígeno HLA-B52/genética , Alelos , Adulto Joven , Japón/epidemiología , Genotipo , Autoanticuerpos/sangre , Autoanticuerpos/inmunologíaRESUMEN
BACKGROUND: No specific biomarker for immune checkpoint inhibitor (ICI)-induced colitis has been established. Previously, we identified anti-integrin αvß6 autoantibodies in >90% of patients with ulcerative colitis (UC). Given that a subset of ICI-induced colitis is similar to UC, we aimed to clarify the relationship between such autoantibodies and ICI-induced colitis. METHODS: Serum anti-integrin αvß6 autoantibody levels were compared between 26 patients with ICI-induced colitis and 157 controls. Endoscopic images of ICI-induced colitis were centrally reviewed. Characteristics of anti-integrin αvß6 autoantibodies in the ICI-induced colitis patients were compared with those of UC patients. RESULTS: Anti-integrin αvß6 autoantibodies were found in 8/26 (30.8%) patients with ICI-induced colitis and 3/157 (1.9%) controls (P < 0.001). Patients with anti-integrin αvß6 autoantibodies had significantly more typical UC endoscopic features than those without the autoantibodies (P < 0.001). Anti-integrin αvß6 autoantibodies in ICI-induced colitis patients were associated with grade ≥3 colitis (P = 0.001) and steroid resistance (P = 0.005). Anti-integrin αvß6 autoantibody titers correlated with ICI-induced colitis disease activity. Anti-integrin αvß6 autoantibodies of ICI-induced colitis exhibited similar characteristics to those of UC. CONCLUSIONS: Anti-integrin αvß6 autoantibodies may serve as potential biomarkers for the diagnosis, classification, risk management, and monitoring the disease activity, of ICI-induced colitis.
Asunto(s)
Autoanticuerpos , Biomarcadores , Colitis Ulcerosa , Inhibidores de Puntos de Control Inmunológico , Integrinas , Humanos , Masculino , Femenino , Autoanticuerpos/sangre , Autoanticuerpos/inmunología , Colitis Ulcerosa/inmunología , Colitis Ulcerosa/tratamiento farmacológico , Colitis Ulcerosa/sangre , Persona de Mediana Edad , Integrinas/inmunología , Integrinas/antagonistas & inhibidores , Anciano , Inhibidores de Puntos de Control Inmunológico/efectos adversos , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Biomarcadores/sangre , Adulto , Antígenos de Neoplasias/inmunología , Colitis/inducido químicamente , Colitis/inmunologíaRESUMEN
BACKGROUND: Closing mucosal defects to reach mucosal healing is an important goal of therapy in inflammatory bowel disease (IBD). Among other cells, monocyte-derived macrophages are centrally involved in such intestinal wound healing. We had previously demonstrated that the anti-α4ß7 integrin antibody vedolizumab blocks the recruitment of non-classical monocytes as biased progenitors of wound healing macrophages to the gut and delays wound healing. However, although important for the interpretation of disappointing results in recent phase III trials in IBD, the effects of the anti-ß7 antibody etrolizumab on wound healing are unclear so far. METHODS: We analyzed the expression of etrolizumab targets on human and mouse monocyte subsets by flow cytometry and assessed their function in adhesion and homing assays. We explored wound-associated monocyte recruitment dynamics with multi-photon microscopy and compared the effects of etrolizumab and vedolizumab surrogate (-s) antibodies on experimental wound healing and wound-associated macrophage abundance. Finally, we investigated wound healing macrophage signatures in the large intestinal transcriptome of patients with Crohn's disease treated with etrolizumab. RESULTS: Human and mouse non-classical monocytes expressed more αEß7 integrin than classical monocytes and were a target of etrolizumab-s, which blocked non-classical monocyte adhesion to MAdCAM-1 and E-Cadherin as well as gut homing in vivo. Intestinal wound healing was delayed on treatment with etrolizumab-s along with a reduction of peri-lesional wound healing macrophages. Wound healing macrophage signatures in the colon of patients with Crohn's disease were substantially down-regulated on treatment with etrolizumab, but not with placebo. CONCLUSIONS: Combined blockade of αEß7 and α4ß7 with etrolizumab seems to exceed the effect of anti-α4ß7 treatment on intestinal wound healing, which might help to inform further investigations to understand the recent observations in the etrolizumab phase III trial program.
Asunto(s)
Fármacos Gastrointestinales , Enfermedades Inflamatorias del Intestino , Integrinas , Macrófagos , Cicatrización de Heridas , Animales , Humanos , Ratones , Enfermedad de Crohn/tratamiento farmacológico , Enfermedad de Crohn/inmunología , Enfermedad de Crohn/patología , Fármacos Gastrointestinales/inmunología , Fármacos Gastrointestinales/farmacología , Fármacos Gastrointestinales/uso terapéutico , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Enfermedades Inflamatorias del Intestino/inmunología , Enfermedades Inflamatorias del Intestino/patología , Integrinas/antagonistas & inhibidores , Integrinas/inmunología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/patología , Monocitos/efectos de los fármacos , Monocitos/inmunología , Monocitos/patología , Cicatrización de Heridas/efectos de los fármacos , Cicatrización de Heridas/inmunologíaRESUMEN
Rap1 GTPase drives assembly of the Mig-10/RIAM/Lamellipodin (MRL protein)-integrin-talin (MIT) complex that enables integrin-dependent lymphocyte functions. Here we used tandem affinity tag-based proteomics to isolate and analyze the MIT complex and reveal that Phostensin (Ptsn), a regulatory subunit of protein phosphatase 1, is a component of the complex. Ptsn mediates dephosphorylation of Rap1, thereby preserving the activity and membrane localization of Rap1 to stabilize the MIT complex. CRISPR/Cas9-induced deletion of PPP1R18, which encodes Ptsn, markedly suppresses integrin activation in Jurkat human T cells. We generated apparently healthy Ppp1r18-/- mice that manifest lymphocytosis and reduced population of peripheral lymphoid tissues ascribable, in part, to defective activation of integrins αLß2 and α4ß7. Ppp1r18-/- T cells exhibit reduced capacity to induce colitis in a murine adoptive transfer model. Thus, Ptsn enables lymphocyte integrin-mediated functions by dephosphorylating Rap1 to stabilize the MIT complex. As a consequence, loss of Ptsn ameliorates T cell-mediated colitis.
Asunto(s)
Integrinas , Tejido Linfoide , Proteína Fosfatasa 1 , Linfocitos T , Proteínas Adaptadoras Transductoras de Señales/inmunología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Adhesión Celular/fisiología , Colitis/inmunología , Colitis/metabolismo , Integrinas/inmunología , Integrinas/metabolismo , Tejido Linfoide/inmunología , Tejido Linfoide/metabolismo , Proteínas de la Membrana/metabolismo , Ratones , Proteína Fosfatasa 1/inmunología , Proteína Fosfatasa 1/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Talina/metabolismo , Proteínas de Unión al GTP rap1/inmunología , Proteínas de Unión al GTP rap1/metabolismoRESUMEN
Integrins are transmembrane receptor heterodimers composed of α and ß subunits. They are known to mediate extracellular signals to promote cell adhesion and spreading, and are therefore essential for cellular immunity. However, proteins that bind to integrin cytoplasmic domains and mediate intracellular signaling to promote cell adhesion require identification. Calcium and integrin-binding protein 1 (CIB1) that binds to the integrin α-cytoplasmic domain has rarely been examined in insects. In this study, we found that 20-hydroxyecdysone promoted cell phagocytosis and spreading in Helicoverpa armigera. Transcriptomic analyses of hemocytes identified an integrin α gene (HaINTα-PS1) whose expression could be induced by either 20-hydroxyecdysone injection or bead challenge. Isothermal titration calorimetry assays showed that H. armigera CIB1-like (HaCIB1-like) weakly bound to the cytoplasmic domain of HaINTα-PS1 in the presence of calcium. HaINTα-PS1 or HaCIB1-like knockdown inhibited hemocytic encapsulation and phagocytosis, and plasmatocyte spreading. Moreover, HaCIB1-like overexpression in a H. armigera epidermal cell line overexpanded cells and impaired cell phagocytosis. Thus, insect CIB1-like potentially interacted with integrin α-cytoplasmic domain and facilitated cell adhesion. This study enriches our understanding of the molecular mechanism underlying integrin-mediated cellular immunity in insects.
Asunto(s)
Proteínas de Unión al Calcio , Integrinas , Mariposas Nocturnas , Animales , Proteínas de Unión al Calcio/inmunología , Proteínas de Unión al Calcio/metabolismo , Ecdisterona/metabolismo , Inmunidad Celular , Integrinas/inmunología , Integrinas/metabolismo , Mariposas Nocturnas/inmunología , Mariposas Nocturnas/metabolismoRESUMEN
Altering T cell trafficking to mucosal regions can enhance immune responses towards pathogenic infections and cancers at these sites, leading to better outcomes. All-trans-retinoic acid (ATRA) promotes T cell migration to mucosal surfaces by inducing transcription of the mucosal-homing receptors CCR9 and α4ß7 via binding to retinoic acid receptors (RARs), which heterodimerize with retinoid X receptors (RXRs) to function. However, the unstable nature and toxicity of ATRA limit its use as a widespread treatment modality for mucosal diseases. Therefore, identifying alternatives that could reduce or eliminate the use of ATRA are needed. Rexinoids are synthetically derived compounds structurally similar to ATRA. Originally named for their ability to bind RXRs, rexinoids can enhance RAR-mediated gene transcription. Furthermore, rexinoids are more stable than ATRA and possess an improved safety profile, making them attractive candidates for use in clinical settings. Here we show that select novel rexinoids act as ATRA mimics, as they cause increased CCR9 and α4ß7 expression and enhanced migration to the CCR9 ligand, CCL25 in vitro, even in the absence of ATRA. Conversely, other rexinoids act synergistically with ATRA, as culturing cells with suboptimal doses of both compounds resulted in CCR9 expression and migration to CCL25. Overall, our findings show that rexinoids can be used independently or synergistically with ATRA to promote mucosal homing of T cells in vitro, and lends support for the prospective clinical use of these compounds in immunotherapeutic approaches for pathogenic infections or cancers at mucosal surfaces.
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Movimiento Celular/efectos de los fármacos , Integrinas/genética , Receptores CCR/genética , Linfocitos T/efectos de los fármacos , Tretinoina/farmacología , Animales , Femenino , Integrinas/inmunología , Ratones , Ratones Endogámicos BALB C , Membrana Mucosa/metabolismo , Receptores CCR/inmunología , Linfocitos T/inmunologíaRESUMEN
Monocytes and macrophages participate in both pro- and anti-inflammatory responses during sepsis. Integrins are the cell adhesion receptors that mediate leukocyte migration and functions. To date, it is not known whether integrin profiles correlate with their trafficking, differentiation, and polarization during sepsis. In this study, using endotoxemia and cecal ligation and puncture model of murine sepsis, we have analyzed the role of surface integrins in tissue-specific infiltration, distribution of monocytes and macrophages, and their association with inflammation-induced phenotypic and functional alterations postinduction (p.i.) of sepsis. Our data show that Ly-6Chi inflammatory monocytes infiltrated into the peritoneum from blood and bone marrow within a few hours p.i. of sepsis, with differential distribution of small (Ly-6CloCD11bloF4/80lo) and large peritoneal macrophages (Ly-6CloCD11bhiF4/80hi) in both models. The results from flow cytometry studies demonstrated a higher expression of integrin α4ß1 on the Ly-6Chi monocytes in different tissues, whereas macrophages in the peritoneum and lungs expressed higher levels of integrin α5ß1 and αvß3 in both models. Additionally, F4/80+ cells with CD206hiMHCIIlo phenotype increased in the lungs of both models by six hours p.i. and expressed higher levels of integrin αvß3 in both lungs and peritoneum. The presence of such cells correlated with higher levels of IL-10 and lower levels of IL-6 and IL-1ß transcripts within six hours p.i. in the lungs compared with the mesentery. Furthermore, bioinformatic analysis with its experimental validation revealed an association of integrin α4 and α5 with inflammatory (e.g., p-SRC) and integrin αv with regulatory molecules (e.g., TGFBR1) in macrophages during sepsis.
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Inflamación/inmunología , Integrinas/genética , Macrófagos/inmunología , Monocitos/inmunología , Sepsis/inmunología , Animales , Perfilación de la Expresión Génica , Integrinas/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Fenotipo , Propiedades de SuperficieRESUMEN
Rapid recruitment of neutrophils to an inflamed site is one of the hallmarks of an effective host defense mechanism. The main pathway through which this happens is by the innate immune response. Neutrophils, which play an important part in innate immune defense, migrate into lungs through the modulation actions of chemokines to execute a variety of pro-inflammatory functions. Despite the importance of chemokines in host immunity, little has been discussed on their roles in host immunity. A holistic understanding of neutrophil recruitment, pattern recognition pathways, the roles of chemokines and the pathophysiological roles of neutrophils in host immunity may allow for new approaches in the treatment of infectious and inflammatory disease of the lung. Herein, this review aims at highlighting some of the developments in lung neutrophil-immunity by focusing on the functions and roles of CXC/CC chemokines and pattern recognition receptors in neutrophil immunity during pulmonary inflammations. The pathophysiological roles of neutrophils in COVID-19 and thromboembolism have also been summarized. We finally summarized various neutrophil biomarkers that can be utilized as prognostic molecules in pulmonary inflammations and discussed various neutrophil-targeted therapies for neutrophil-driven pulmonary inflammatory diseases.
Asunto(s)
Inmunidad Innata/inmunología , Neutrófilos/inmunología , Neumonía/inmunología , Biomarcadores/sangre , COVID-19/inmunología , Degranulación de la Célula/inmunología , Quimiocinas/inmunología , Ensayos Clínicos como Asunto , Trampas Extracelulares/inmunología , Humanos , Integrinas/inmunología , Pulmón/inmunología , Pulmón/patología , Neutrófilos/efectos de los fármacos , Neumonía/diagnóstico , Neumonía/tratamiento farmacológico , Receptores de Reconocimiento de Patrones/inmunología , Estallido Respiratorio/inmunología , SARS-CoV-2 , Tromboembolia/inmunologíaRESUMEN
Presence of TGFß in the tumor microenvironment is one of the most relevant cancer immune-escape mechanisms. TGFß is secreted in an inactive form, and its activation within the tumor may depend on different cell types and mechanisms than its production. Here we show in mouse melanoma and breast cancer models that regulatory T (Treg) cells expressing the ß8 chain of αvß8 integrin (Itgß8) are the main cell type in the tumors that activates TGFß, produced by the cancer cells and stored in the tumor micro-environment. Itgß8 ablation in Treg cells impairs TGFß signalling in intra-tumoral T lymphocytes but not in the tumor draining lymph nodes. Successively, the effector function of tumor infiltrating CD8+ T lymphocytes strengthens, leading to efficient control of tumor growth. In cancer patients, anti-Itgß8 antibody treatment elicits similar improved cytotoxic T cell activation. Thus, this study reveals that Treg cells work in concert with cancer cells to produce bioactive-TGFß and to create an immunosuppressive micro-environment.
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Integrinas/inmunología , Neoplasias/inmunología , Linfocitos T Reguladores/inmunología , Factor de Crecimiento Transformador beta/inmunología , Animales , Femenino , Humanos , Integrinas/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neoplasias/genética , Microambiente TumoralRESUMEN
Inflammatory Bowel Disease (IBD) comprises a series of chronic and relapsing intestinal diseases, with Crohn's disease and ulcerative colitis being the most common. The abundant and uncontrolled deposition of extracellular matrix, namely fibrosis, is one of the major hallmarks of IBD and is responsible for the progressive narrowing and closure of the intestine, defined as stenosis. Although fibrosis is usually considered the product of chronic inflammation, the substantial failure of anti-inflammatory therapies to target and reduce fibrosis in IBD suggests that fibrosis might be sustained in an inflammation-independent manner. Pharmacological therapies targeting integrins have recently shown great promise in the treatment of IBD. The efficacy of these therapies mainly relies on their capacity to target the integrin-mediated recruitment and functionality of the immune cells at the damage site. However, by nature, integrins also act as mechanosensitive molecules involved in the intracellular transduction of signals and modifications originating from the extracellular matrix. Therefore, understanding integrin signaling in the context of IBD may offer important insights into mechanisms of matrix remodeling, which are uncoupled from inflammation and could underlie the onset and persistency of intestinal fibrosis. In this review, we present the currently available knowledge on the role of integrins in the etiopathogenesis of IBD, highlighting their role in the context of immune-dependent and independent mechanisms.
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Matriz Extracelular/inmunología , Enfermedades Inflamatorias del Intestino/inmunología , Integrinas/inmunología , Mecanotransducción Celular/inmunología , Animales , Matriz Extracelular/patología , Fibrosis , Humanos , Enfermedades Inflamatorias del Intestino/patologíaRESUMEN
Cold atmospheric plasma (CAP) reduces bacteria and interacts with tissues and cells, thus improving wound healing. The CAP-related induction of neutrophils was recently described in stained sections of wound tissue in mice. Consequently, this study aimed to examine the functionality of human polymorphonuclear cells (PMN)/granulocytes through either a plasma-treated solution (PTS) or the direct CAP treatment with different plasma modes and treatment durations. PTS analysis yielded mode-dependent differences in the production of reactive oxygen species (ROS) and reactive nitrogen species (RNS) after CAP treatment. Live-cell imaging did not show any chemo-attractive or NETosis-inducing effect on PMNs treated with PTS. The time to maximum ROS production (TmaxROS) in PMNs was reduced by PTS and direct CAP treatment. PMNs directly treated with CAP showed an altered cell migration dependent on the treatment duration as well as decreased TmaxROS without inducing apoptosis. Additionally, flow cytometry showed enhanced integrin and selectin expression, as a marker of activation, on PMN surfaces. In conclusion, the modification of PMN immunoreactivity may be a main supporting mechanism for CAP-induced improvement in wound healing.
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Granulocitos/inmunología , Integrinas/inmunología , Gases em Plasma/farmacología , Especies de Nitrógeno Reactivo/inmunología , Especies Reactivas de Oxígeno/inmunología , Selectinas/inmunología , Apoptosis/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/inmunología , Granulocitos/citología , HumanosRESUMEN
Polymorphonuclear neutrophils (PMNs), the most abundant white blood cells, are recruited rapidly to sites of infection to exert potent anti-microbial activity. Information regarding their role in infection with human immunodeficiency virus (HIV) is limited. Here we report that addition of PMNs to HIV-infected cultures of human tonsil tissue or peripheral blood mononuclear cells causes immediate and long-lasting suppression of HIV-1 spread and virus-induced depletion of CD4 T cells. This inhibition of HIV-1 spread strictly requires PMN contact with infected cells and is not mediated by soluble factors. 2-Photon (2PM) imaging visualized contacts of PMNs with HIV-1-infected CD4 T cells in tonsil tissue that do not result in lysis or uptake of infected cells. The anti-HIV activity of PMNs also does not involve degranulation, formation of neutrophil extracellular traps, or integrin-dependent cell communication. These results reveal that PMNs efficiently blunt HIV-1 replication in primary target cells and tissue by an unconventional mechanism.
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Linfocitos T CD4-Positivos/inmunología , VIH-1/genética , Leucocitos Mononucleares/inmunología , Neutrófilos/inmunología , Tonsila Palatina/inmunología , Linfocitos T CD4-Positivos/virología , Comunicación Celular , Trampas Extracelulares , Infecciones por VIH/inmunología , Infecciones por VIH/virología , VIH-1/crecimiento & desarrollo , VIH-1/patogenicidad , Humanos , Integrinas/genética , Integrinas/inmunología , Leucocitos Mononucleares/virología , Neutrófilos/virología , Tonsila Palatina/citología , Cultivo Primario de Células , Carga Viral , Replicación ViralRESUMEN
Glioblastoma multiforme (GBM), the most aggressive brain cancer, recurs because glioblastoma stem cells (GSCs) are resistant to all standard therapies. We showed that GSCs, but not normal astrocytes, are sensitive to lysis by healthy allogeneic natural killer (NK) cells in vitro. Mass cytometry and single-cell RNA sequencing of primary tumor samples revealed that GBM tumor-infiltrating NK cells acquired an altered phenotype associated with impaired lytic function relative to matched peripheral blood NK cells from patients with GBM or healthy donors. We attributed this immune evasion tactic to direct cell-to-cell contact between GSCs and NK cells via αv integrin-mediated TGF-ß activation. Treatment of GSC-engrafted mice with allogeneic NK cells in combination with inhibitors of integrin or TGF-ß signaling or with TGFBR2 gene-edited allogeneic NK cells prevented GSC-induced NK cell dysfunction and tumor growth. These findings reveal an important mechanism of NK cell immune evasion by GSCs and suggest the αv integrin/TGF-ß axis as a potentially useful therapeutic target in GBM.
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Glioblastoma/inmunología , Integrinas/inmunología , Células Asesinas Naturales/inmunología , Proteínas de Neoplasias/inmunología , Células Madre Neoplásicas/inmunología , Factor de Crecimiento Transformador beta/inmunología , Animales , Femenino , Glioblastoma/genética , Glioblastoma/patología , Glioblastoma/terapia , Xenoinjertos , Humanos , Integrinas/genética , Células Asesinas Naturales/patología , Masculino , Ratones , Proteínas de Neoplasias/genética , Trasplante de Neoplasias , Células Madre Neoplásicas/patología , Receptor Tipo II de Factor de Crecimiento Transformador beta/genética , Receptor Tipo II de Factor de Crecimiento Transformador beta/inmunología , Factor de Crecimiento Transformador beta/genéticaRESUMEN
Signal-regulatory protein alpha (SIRPA) is a well-known inhibitor of phagocytosis when it complexes with CD47 expressed on target cells. Here we show that SIRPA decreased in vitro infection by a number of pathogenic viruses, including New World and Old World arenaviruses, Zika virus, vesicular stomatitis virus and pseudoviruses bearing the Machupo virus, Ebola virus and SARS-CoV-2 glycoproteins, but not HSV-1, MLV or mNoV. Moreover, mice with targeted mutation of the Sirpa gene that renders it non-functional were more susceptible to infection with the New World arenaviruses Junín virus vaccine strain Candid 1 and Tacaribe virus, but not MLV or mNoV. All SIRPA-inhibited viruses have in common the requirement for trafficking to a low pH endosomal compartment. This was clearly demonstrated with SARS-CoV-2 pseudovirus, which was only inhibited by SIRPA in cells in which it required trafficking to the endosome. Similar to its role in phagocytosis inhibition, SIRPA decreased virus internalization but not binding to cell surface receptors. We also found that increasing SIRPA levels via treatment with IL-4 led to even greater anti-viral activity. These data suggest that enhancing SIRPA's activity could be a target for anti-viral therapies.
Asunto(s)
Endocitosis , Virus ARN/inmunología , Receptores Inmunológicos/fisiología , Internalización del Virus , Animales , Antivirales/farmacología , Línea Celular , Membrana Celular/virología , Chlorocebus aethiops , Sistemas de Liberación de Medicamentos , Integrinas/inmunología , Interleucina-4/farmacología , Ratones , Ratones Noqueados , Dominios Proteicos , Receptores Inmunológicos/genética , Células VeroRESUMEN
Treating macaques with an anti-α4ß7 antibody under the umbrella of combination antiretroviral therapy (cART) during early SIV infection can lead to viral remission, with viral loads maintained at < 50 SIV RNA copies/ml after removal of all treatment in a subset of animals. Depletion of CD8+ lymphocytes in controllers resulted in transient recrudescence of viremia, suggesting that the combination of cART and anti-α4ß7 antibody treatment led to a state where ongoing immune responses kept the virus undetectable in the absence of treatment. A previous mathematical model of HIV infection and cART incorporates immune effector cell responses and exhibits the property of two different viral load set-points. While the lower set-point could correspond to the attainment of long-term viral remission, attaining the higher set-point may be the result of viral rebound. Here we expand that model to include possible mechanisms of action of an anti-α4ß7 antibody operating in these treated animals. We show that the model can fit the longitudinal viral load data from both IgG control and anti-α4ß7 antibody treated macaques, suggesting explanations for the viral control associated with cART and an anti-α4ß7 antibody treatment. This effective perturbation to the virus-host interaction can also explain observations in other nonhuman primate experiments in which cART and immunotherapy have led to post-treatment control or resetting of the viral load set-point. Interestingly, because the viral kinetics in the various treated animals differed-some animals exhibited large fluctuations in viral load after cART cessation-the model suggests that anti-α4ß7 treatment could act by different primary mechanisms in different animals and still lead to post-treatment viral control. This outcome is nonetheless in accordance with a model with two stable viral load set-points, in which therapy can perturb the system from one set-point to a lower one through different biological mechanisms.