RESUMO
INTRODUCTION: The etiology of brain aneurysms remains poorly understood. Finnish research suggests that oral bacteria might contribute to the development and rupture of brain aneurysms. Previous studies by our team have not confirmed these findings, likely due to methodological differences. We aimed to replicate the Finnish study with a French population, using the same primers and comparing the results to internal controls. METHODS: We used RT-qPCR to retrospectively analyze the expression of oral bacterial genes in eight patients. During surgical procedures, four tissue types were consistently sampled from each patient: the aneurysmal wall, the superficial temporal artery (STA), the middle meningeal artery (MMA), and the dura mater (DM). Results were expressed as fold differences employing the 2-∆∆Ct method, and statistical analyses were performed accordingly. RESULTS: Our cohort included eight patients, evenly split into ruptured and unruptured groups. The sex distribution was balanced (4 females, 4 males). We observed DNA expression from oral bacteria in all sampled tissues; however, there were no significant differences between the ruptured and unruptured groups. CONCLUSION: We detected oral bacterial gene expression in the aneurysmal wall, STA, MMA, and DM in a sample of French patients. Although limited by the small sample size, our results suggest a potential role for bacterial involvement in vascular invasiveness related to bacteremia. These findings do not definitively link oral bacteria to the pathogenesis of aneurysm development and rupture.
Assuntos
Aneurisma Intracraniano , Humanos , Feminino , Masculino , Aneurisma Intracraniano/microbiologia , Pessoa de Meia-Idade , França , Idoso , Estudos Retrospectivos , Boca/microbiologia , Adulto , Aneurisma Roto/microbiologia , Artérias Temporais , Dura-Máter , Bactérias/genética , Bactérias/isolamento & purificação , Artérias MeníngeasRESUMO
Photodynamic therapy (PDT) is a two-stage treatment relying on cytotoxicity induced by photoexcitation of a nontoxic dye, called photosensitizer (PS). Using 5-aminolevulinic acid (5-ALA), the pro-drug of PS protoporphyrin IX, we investigated the impact of PDT on hepatocellular carcinoma (HCC). Optimal 5-ALA PDT dose was determined on three HCC cell lines by analyzing cell death after treatment with varying doses. HCC-patient-derived tumor hepatocytes and healthy donor liver myofibroblasts were treated with optimal 5-ALA PDT doses. The proliferation of cancer cells and healthy donor immune cells cultured with 5-ALA-PDT-treated conditioned media was analyzed. Finally, therapy efficacy on humanized SCID mice model of HCC was investigated. 5-ALA PDT induced a dose-dependent decrease in viability, with an up-to-four-fold reduction in viability of patient tumor hepatocytes. The 5-ALA PDT treated conditioned media induced immune cell clonal expansion. 5-ALA PDT has no impact on myofibroblasts in terms of viability, while their activation decreased cancer cell proliferation and reduced the tumor growth rate of the in vivo model. For the first time, 5-ALA PDT has been validated on primary patient tumor hepatocytes and donor healthy liver myofibroblasts. 5-ALA PDT may be an effective anti-HCC therapy, which might induce an anti-tumor immune response.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Transplante de Fígado , Fotoquimioterapia , Camundongos , Animais , Humanos , Ácido Aminolevulínico/farmacologia , Ácido Aminolevulínico/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Meios de Cultivo Condicionados/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Camundongos SCID , Doadores Vivos , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêutico , Protoporfirinas/metabolismo , Linhagem Celular TumoralRESUMO
Photodynamic therapy (PDT) has shown improvements in cancer treatment and in the induction of a proper anti-tumor immune response. However, current photosensitizers (PS) lack tumor specificity, resulting in reduced efficacy and side effects in patients with intraperitoneal ovarian cancer (OC). In order to target peritoneal metastases of OC, which overexpress folate receptor (FRα) in 80% of cases, we proposed a targeted PDT using a PS coupled with folic acid. Herein, we applied this targeted PDT in an in vivo mouse model of peritoneal ovarian carcinomatosis. The efficacy of the treatment was evaluated in mice without and with human peripheral blood mononuclear cell (PBMC) reconstitution. When mice were reconstituted, using a fractionized PDT protocol led to a significantly higher decrease in the tumor growth than that obtained in the non-reconstituted mice (p = 0.0469). Simultaneously, an immune response was reflected by an increase in NK cells, and both CD4+ and CD8+ T cells were activated. A promotion in cytokines IFNγ and TNFα and an inhibition in cytokines TGFß, IL-8, and IL-10 was also noticed. Our work showed that a fractionized FRα-targeted PDT protocol is effective for the treatment of OC and goes beyond local induction of tumor cell death, with the promotion of a subsequent anti-tumor response.
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Neoplasias Ovarianas , Neoplasias Peritoneais , Fotoquimioterapia , Humanos , Camundongos , Animais , Feminino , Leucócitos Mononucleares/metabolismo , Fotoquimioterapia/métodos , Neoplasias Ovarianas/metabolismo , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêutico , Neoplasias Peritoneais/secundário , Citocinas/metabolismo , Ácido Fólico/uso terapêutico , Linhagem Celular Tumoral , Receptor 1 de Folato/metabolismo , Receptor 1 de Folato/uso terapêuticoRESUMO
Nasopharyngeal carcinoma (NPC) is a malignant tumour of the head and neck affecting localised regions of the world, with the highest rates described in Southeast Asia, Northern Africa, and Greenland. Its high morbidity rate is linked to both late-stage diagnosis and unresponsiveness to conventional anti-cancer treatments. Multiple aetiological factors have been described including environmental factors, genetics, and viral factors (Epstein Barr Virus, EBV), making NPC treatment that much more complex. The most common forms of NPCs are those that originate from the epithelial tissue lining the nasopharynx and are often linked to EBV infection. Indeed, they represent 75-95% of NPCs in the low-risk populations and almost 100% of NPCs in high-risk populations. Although conventional surgery has been improved with nasopharyngectomy's being carried out using more sophisticated surgical equipment for better tumour resection, recent findings in the tumour microenvironment have led to novel treatment options including immunotherapies and photodynamic therapy, able to target the tumour and improve the immune system. This review provides an update on the disease's aetiology and the future of NPC treatments with a focus on therapies activating T cell immunity.
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Infecções por Vírus Epstein-Barr/terapia , Carcinoma Nasofaríngeo/virologia , Neoplasias Nasofaríngeas/virologia , Linfócitos T/metabolismo , Terapia Combinada , Infecções por Vírus Epstein-Barr/imunologia , Herpesvirus Humano 4/imunologia , Humanos , Imunoterapia , Carcinoma Nasofaríngeo/imunologia , Carcinoma Nasofaríngeo/terapia , Neoplasias Nasofaríngeas/imunologia , Neoplasias Nasofaríngeas/terapia , Faringectomia , Fotoquimioterapia , Microambiente TumoralRESUMO
The development of immunotherapy has recently modified the anti-tumor therapeutic arsenal; particularly, immune checkpoint inhibitors have led to a significant increase in overall survival. The current challenge is now to select good responder patients by identifying early biomarkers to propose therapeutic combinations that potentiate the efficacy of the therapy. Here we report the case of a 60-year-old man with superficial melanoma treated with high-dose hypo fractionated radiotherapy (H-SRT) combined with a single dose of anti-PD1 immunotherapy (Nivolumab) for a metastatic lymph node recurrence due to cancer progression. In this study, we present the results obtained regarding the activation of the Th1 immune response after H-SRT treatment followed by anti PD-1 therapeutic protocol. These results were correlated with clinical data to identify potential immunological biomarkers of treatment efficacy. This exceptional case report shows that a combination of H-SRT with a single dose of anti-PD1 immunotherapy may allow a better activation of the immune response in favor of a complete clinical response.
Assuntos
Antineoplásicos Imunológicos/uso terapêutico , Melanoma/radioterapia , Melanoma/terapia , Nivolumabe/uso terapêutico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Células Th1/efeitos dos fármacos , Humanos , Imunoterapia/métodos , Masculino , Melanoma/imunologia , Pessoa de Meia-IdadeRESUMO
Colorectal cancer (CRC) remains the most cancer type related to chronic inflammation; however, the mechanisms that link inflammation to CRC development and progression are still poorly understood. Our study aimed to investigate one of the prominent inflammatory response in cancers, iNOS/NO system. In this regard, we evaluated the link between the iNOS/NO system and CRC progression, its relation with the host immune responses and its response to cetuximab combined with chemotherapy. We found that the nitrite levels were nearly twice as high in metastatic CRC plasma and culture supernatants from PBMCs and tumor explants compared with those without metastases and healthy controls. Interestingly, we showed that the highest iNOS expression and NO levels are present in the damaged CRC tissues that have highest leukocyte infiltration. Our findings highlight the implication of iNOS/NO system in tissue alteration and leukocyte invasion. Thus, we observed imbalance between effector/memory T cell markers and Treg transcription factor (Foxp3). Accordingly, we detected higher IFNγ and T-bet expression levels in colorectal tumor tissues at early stage. In contrast, consistent with iNOS and Foxp3 expression, TGFß, CTLA-4 and IL-10 were significantly related to the tumor stage progression. Furthermore, our study revealed that Cetuximab combined with chemotherapy treatment markedly down-regulates iNOS/NO system as well as IL-10 and TGFß levels. Altogether, we conclude that cetuximab can potentiate the efficacy of chemotherapy, particularly by iNOS/NO system and immunosuppressive cytokines modulation. Thus, we suggest that iNOS/NO system may represent an attractive candidate biomarker for monitoring CRC progression, malignity and response to therapy.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Fatores Imunológicos/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico/metabolismo , Regulação para Cima/fisiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Cetuximab/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Linfócitos T Reguladores/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacosRESUMO
Colitis associated cancer (CAC) is the colorectal cancer (CRC) subtype that is associated with bowel disease such as ulcerative colitis (UC). The data on role of NF-κB signaling in development and progression of CAC were derived from preclinical studies, whereas data from human are rare. The aim of this work was to study the contribution of NF-κB pathway during UC and CAC, as well as the immunomodulatory effect of all-trans retinoic acid (AtRA). We analyzed the expression of NOS2, TNF-α, TLR4, and NF-κB, in colonic mucosa. We also studied NO/TNF-α modulation by LPS in colonic mucosa pretreated with AtRA. A marked increase in TLR4, NF-κB, TNF-α, and NOS2 expression was reported in colonic mucosa. The relationship between LPS/TLR4 and TNF-α/NO production, as well as the role of NF-κB signaling, was confirmed by ex vivo experiments and the role of LPS/TLR4 in NOS2/TNF-α induction through NF-κB pathway was suggested. AtRA downregulates NOS2 and TNF-α expression. Collectively, our study indicates that AtRA modulates in situ LPS/TLR4/NF-κB signaling pathway targeting NOS2 and TNF-α expression. Therefore, we suggest that AtRA has a potential value in new strategies to improve the current therapy, as well as in the clinical prevention of CAC development and progression.
Assuntos
Colite Ulcerativa/sangue , Colite/sangue , Neoplasias Colorretais/sangue , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico/sangue , Fator de Necrose Tumoral alfa/sangue , Idoso , Western Blotting , Colite Ulcerativa/metabolismo , Neoplasias Colorretais/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imuno-Histoquímica , Mucosa Intestinal/metabolismo , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Galectin-9 (gal-9) is a multifunctional ß-galactoside-binding lectin, frequently released in the extracellular medium, where it acts as a pleiotropic immune modulator. Despite its overall immunosuppressive effects, a recent study has reported bimodal action of gal-9 on human resting blood T cells with apoptosis occurring in the majority of them, followed by a wave of activation and expansion of Th1 cells in the surviving population. Our knowledge of the signaling events triggered by exogenous gal-9 in T cells remains limited. One of these events is cytosolic calcium (Ca(2+)) release reported in some murine and human T cells. The aim of this study was to investigate the contribution of Ca(2+) mobilization to apoptotic and nonapoptotic effects of exogenous gal-9 in human T cells. We found that the T cell receptor (TCR)-CD3 complex and the Lck kinase were required for Ca(2+) mobilization but not for apoptosis induction in Jurkat cells. These data were confirmed in human CD4(+) T cells from peripheral blood as follows: a specific Lck chemical inhibitor abrogated Ca(2+) mobilization but not apoptosis induction. Moreover, Lck activity was also required for the production of Th1-type cytokines, i.e. interleukin-2 and interferon-γ, which resulted from gal-9 stimulation in peripheral CD4(+) T cells. These findings indicate that gal-9 acts on T cells by two distinct pathways as follows: one mimicking antigen-specific activation of the TCR with a mandatory contribution of proximal elements of the TCR complex, especially Lck, and another resulting in apoptosis that is independent of this complex.
Assuntos
Apoptose , Complexo CD3/metabolismo , Galectinas/metabolismo , Proteína Tirosina Quinase p56(lck) Linfócito-Específica/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/metabolismo , Complexo CD3/genética , Cálcio/metabolismo , Citocinas/genética , Citocinas/metabolismo , Galectinas/genética , Humanos , Proteína Tirosina Quinase p56(lck) Linfócito-Específica/genética , Receptores de Antígenos de Linfócitos T/genética , Linfócitos T/citologiaRESUMO
Epstein-Barr Virus (EBV) is present in the neoplastic cells of around 20-30% of patients with Hodgkin Lymphoma (HL). Although, an immunosuppressive environment is currently described in HL patients, little is known concerning the regulatory mechanism induced by EBV proteins expression in tumour cells. This study aimed to investigate an association between regulatory Type 1 cells (Tr1) and EBV tissue positivity in HL patients. Transcriptomic analysis of both EBV-positive and EBV-negative tumours showed that EBV infection increased gene expression of Tr1-related markers (ITGA2, ITGB2, LAG3) and associated-immunosuppressive cytokines (IL10). This up-regulation was associated with an over-expression of several chemokine markers known to attract T-helper type 2 (Th2) and regulatory T cells thus contributing to immune suppression. This Tr1 cells recruitment in EBV-positive HL was confirmed by immunohistochemical analysis of frozen nodes biopsies and by flow cytometric analysis of peripheral blood mononuclear cells of EBV-positive patients. Additionally, we showed that IL10 production was significantly enhanced in tumours and blood of EBV-positive HL patients. Our results propose a new model in which EBV can recruit Tr1 cells to the nodes' microenvironment, suggesting that the expression of EBV proteins in tumour cells could enable the escape of EBV-infected tumour cells from the virus-specific CTL response.
Assuntos
Infecções por Vírus Epstein-Barr/imunologia , Doença de Hodgkin/imunologia , Linfócitos T Reguladores/imunologia , Adolescente , Adulto , Idoso , Biomarcadores/metabolismo , Quimiocinas/metabolismo , Criança , Citocinas/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Doença de Hodgkin/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Linfócitos T Reguladores/virologia , Células Th2/imunologia , Células Th2/virologia , Regulação para Cima , Adulto JovemRESUMO
CCL18 is both a constitutively expressed and an inducible chemokine, whose role in the inflammatory reaction is poorly known. The aim of this study was to evaluate whether CCL18 has the capacity to attract human T cells with a regulatory function (regulatory T cells [Treg]). Results from chemotaxis assays performed on different types of Treg showed that CD4(+)CD25(+)CD127(low) cells, but neither T regulatory type 1 clones nor Treg differentiated in vitro with anti-CD3/CD46 mAbs, were recruited by CCL18 in a dose-dependent manner. CCL18-recruited memory CD4(+) T cells were enriched in CD25(high), CD25(+)CD127(low), latency-associated peptide/TGF-ß1, and CCR4-expressing T cells, whereas there was no enrichment in Foxp3(+) cells as compared with controls. Stimulated CCL18-recruited memory T cells produced significantly increased amounts of the regulatory cytokines IL-10 and TGF-ß1, as well as IL-4, but not IFN-γ and IL-17. Cell surface CCL18 binding was found predominantly on IL-10(+) (26.3 ± 5.8%) and on a few latency-associated peptide/TGF-ß1(+) (18.1 ± 1.9%) and IL-4(+) (14.5 ± 2.9%) memory T cells. In an in vivo model of SCID mice grafted with human skin and reconstituted with autologous PBMCs, the intradermal injection of CCL18 led to the cutaneous recruitment of CD4(+), CD25(+), and IL-10(+) cells, but not Foxp3(+) cells. Furthermore, CCL18-recruited memory T cells inhibited the proliferation of CD4(+)CD25(-) effector T cells through an IL-10-dependent mechanism. These data suggest that CCL18 may contribute to maintaining tolerance and/or suppressing deleterious inflammation by attracting memory Tregs into tissues, particularly in the lung, where it is highly and constitutively expressed.
Assuntos
Movimento Celular/imunologia , Quimiocinas CC/fisiologia , Pulmão/citologia , Pulmão/imunologia , Linfócitos T Reguladores/imunologia , Animais , Células Cultivadas , Quimiotaxia de Leucócito/imunologia , Células Clonais , Humanos , Interleucina-10/biossíntese , Pulmão/metabolismo , Camundongos , Camundongos SCID , Transplante de Pele/imunologia , Transplante de Pele/patologia , Linfócitos T Reguladores/metabolismoRESUMO
Virus infections are involved in chronic inflammation and, in some cases, cancer development. Although a viral infection activates the immune system's response that eradicates the pathogen mainly through inflammatory mechanisms, it is now recognized that this inflammatory condition is also favorable to the development of tumors. Indeed, it is well described that viruses, such as hepatitis C virus (HCV), Epstein Barr virus (EBV), human papillomavirus (HPV) or human T-cell lymphotropic virus type-1 (HTLV-1), are important risk factors for tumor malignancies. The inflammatory response is a fundamental immune mechanism which involves several molecular and cellular components consisting of cytokines and chemokines that are released by various proinflammatory cells. In parallel to this process, some endogenous recruited components release anti-inflammatory mediators to restore homeostasis. The development of tools and strategies using viruses to hijack the immune response is mostly linked to the presence of regulatory T-cells (Treg) that can inhibit inflammation and antiviral responses of other effector cells. In this review, we will focus on current understanding of the role of natural and induced Treg in the control and the resolution of inflammatory response in HCV-, HTLV-1-, and EBV-associated cancers.
Assuntos
Hepacivirus/metabolismo , Herpesvirus Humano 4/metabolismo , Vírus Linfotrópico T Tipo 1 Humano/metabolismo , Inflamação/metabolismo , Neoplasias/virologia , Linfócitos T Reguladores/citologia , Animais , Carcinoma/virologia , Carcinoma Hepatocelular/virologia , Doença de Hodgkin/virologia , Homeostase , Humanos , Imunidade Inata , Leucemia de Células T/virologia , Neoplasias Hepáticas/virologia , Linfoma de Células T/virologia , Carcinoma Nasofaríngeo , Neoplasias Nasofaríngeas/virologia , Fatores de RiscoRESUMO
Ovarian cancer (OC) is one of the most lethal cancers among women. Frequent recurrence in the peritoneum due to the presence of microscopic tumor residues justifies the development of new therapies. Indeed, our main objective is to develop a targeted photodynamic therapy (PDT) treatment of peritoneal carcinomatosis from OC to improve the life expectancy of cancer patients. Herein, we propose a targeted-PDT using a vectorized photosensitizer (PS) coupled with a newly folic acid analog (FAA), named PSFAA, in order to target folate receptor alpha (FRα) overexpressed on peritoneal metastasis. This PSFAA was the result of the coupling of pyropheophorbide-a (Pyro-a), as the PS, to a newly synthesized FAA via a polyethylene glycol (PEG) spacer. The selectivity and the PDT efficacy of PSFAA was evaluated on two human OC cell lines overexpressing FRα compared to fibrosarcoma cells underexpressing FRα. Final PSFAA, including the synthesis of a newly FAA and its conjugation to Pyro-a, was obtained after 10 synthesis steps, with an overall yield of 19%. Photophysical properties of PSFAA in EtOH were performed and showed similarity with those of free Pyro-a, such as the fluorescence and singlet oxygen quantum yields (Φf = 0.39 and ΦΔ = 0.53 for free Pyro-a, and Φf = 0.26 and ΦΔ = 0.41 for PSFAA). Any toxicity of PSFAA was noticed. After light illumination, a dose-dependent effect on PS concentration and light dose was shown. Furthermore, a PDT efficacy of PSFAA on OC cell secretome was detected inducing a decrease of a pro-inflammatory cytokine secretion (IL-6). This new PSFAA has shown promising biological properties highlighting the selectivity of the therapy opening new perspectives in the treatment of a cancer in a therapeutic impasse.
Assuntos
Clorofila , Ácido Fólico , Interleucina-6 , Neoplasias Ovarianas , Fotoquimioterapia , Fármacos Fotossensibilizantes , Fotoquimioterapia/métodos , Humanos , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/patologia , Feminino , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/administração & dosagem , Ácido Fólico/química , Linhagem Celular Tumoral , Clorofila/análogos & derivados , Clorofila/farmacologia , Clorofila/administração & dosagem , Clorofila/uso terapêutico , Clorofila/química , Interleucina-6/metabolismo , Morte Celular/efeitos dos fármacos , Receptor 1 de Folato/metabolismo , Inflamação/tratamento farmacológico , Sobrevivência Celular/efeitos dos fármacosRESUMO
Epstein-Barr virus (EBV) is a common human herpesvirus. Infection with EBV is associated with several human malignancies in which the virus expresses a set of latent proteins, among which is latent membrane protein 1 (LMP1). LMP1 is able to transform numerous cell types and is considered the main oncogenic protein of EBV. The mechanism of action is based on mimicry of activated members of the tumor necrosis factor (TNF) receptor superfamily, through the ability of LMP1 to bind similar adapters and to activate signaling pathways. We previously generated two unique models: a monocytic cell line and a lymphocytic (NC5) cell line immortalized by EBV that expresses the type II latency program. Here we generated LMP1 dominant negative forms (DNs), based on fusion between green fluorescent protein (GFP) and transformation effector site 1 (TES1) or TES2 of LMP1. Then we generated cell lines conditionally expressing these DNs. These DNs inhibit NF-κB and Akt pathways, resulting in the impairment of survival processes and increased apoptosis in these cell lines. This proapoptotic effect is due to reduced interaction of LMP1 with specific adapters and the recruitment of these adapters to DNs, which enable the generation of an apoptotic complex involving TRADD, FADD, and caspase 8. Similar results were obtained with cell lines displaying a latency III program in which LMP1-DNs decrease cell viability. Finally, we prove that synthetic peptides display similar inhibitory effects in EBV-infected cells. DNs derived from LMP1 could be used to develop therapeutic approaches for malignant diseases associated with EBV.
Assuntos
Transformação Celular Viral , Regulação para Baixo , Infecções por Vírus Epstein-Barr/virologia , Herpesvirus Humano 4/fisiologia , Linfócitos T/virologia , Proteínas da Matriz Viral/metabolismo , Apoptose , Linhagem Celular , Infecções por Vírus Epstein-Barr/metabolismo , Infecções por Vírus Epstein-Barr/fisiopatologia , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/crescimento & desenvolvimento , Humanos , Transdução de Sinais , Linfócitos T/citologia , Linfócitos T/metabolismo , Proteínas da Matriz Viral/genética , Latência ViralRESUMO
The aim of this study was to evaluate the nonchemotactic function of CCL18 on human dendritic cells (DCs). In different protocols of DC differentiation, CCL18 was highly produced, suggesting that it may constitute a mandatory mediator of the differentiation process. Differentiation of monocytes from healthy subjects in the presence of granulocyte-macrophage colony-stimulating factor and CCL18 led to the development of DCs with a semimature phenotype, with intermediate levels of costimulatory and MHC class II molecules, increased CCR7 expression, which induced, in coculture with allogenic naive T cells, an increase in IL-10 production. The generated T cells were able to suppress the proliferation of effector CD4(+)CD25(-) cells, through a cytokine-dependent mechanism, and exhibited characteristics of type 1 T regulatory cells. The generation of tolerogenic DCs by CCL18 was dependent on the production of indoleamine 2,3-dioxigenase through an interleukin-10-mediated mechanism. Surprisingly, when DCs originated from allergic patients, the tolerogenic effect of CCL18 was lost in relation with a decreased binding of CCL18 to its putative receptor. This study is the first to define a chemokine able to generate tolerogenic DCs. However, this function was absent in allergic donors and may participate to the decreased tolerance observed in allergic diseases.
Assuntos
Apresentação de Antígeno/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Quimiocinas CC/farmacologia , Células Dendríticas/efeitos dos fármacos , Tolerância Imunológica/efeitos dos fármacos , Linfócitos T Reguladores/efeitos dos fármacos , Apresentação de Antígeno/imunologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/fisiologia , Diferenciação Celular/genética , Células Cultivadas , Quimiocinas CC/genética , Quimiocinas CC/metabolismo , Quimiocinas CC/fisiologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Células Dendríticas/fisiologia , Fatores de Transcrição Forkhead/metabolismo , Saúde , Humanos , Hipersensibilidade/imunologia , Hipersensibilidade/metabolismo , Tolerância Imunológica/genética , Tolerância Imunológica/imunologia , Interleucina-10/metabolismo , Cultura Primária de Células , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismoRESUMO
Background: Pancreatic adenocarcinoma (PDAC) is a devastating disease with an urgent need for therapeutic innovation. Immune checkpoint inhibition has shown promise in a variety of solid tumors, but most clinical trials have failed to demonstrate clinical efficacy in PDAC. This low efficacy is partly explained by a highly immunosuppressive microenvironment, which dampens anti-tumor immunity through the recruitment or induction of immunosuppressive cells, particularly regulatory T cells (Tregs). In this context, our laboratory has developed a novel immunotherapeutic strategy aimed at inhibiting the suppressive activity of Tregs, based on a patented (EP3152234B1) monoclonal antibody (mAb) targeting galectin-9 (LGALS9). Materials and methods: CD4+ conventional T cells (TCD4 or Tconv), Treg ratio, and LGALS9 expression were analyzed by immunohistochemistry (IHC) and cytometry in blood and pancreas of K-rasLSL.G12D/+;Pdx-1-Cre (KC) and K-rasWildType (WT);Pdx1-Cre (WT) mice aged 4-13 months. Pancreatic intraepithelial neoplasm (PanIN) progression and grade were quantified using FIJI software and validated by pathologists. The anti-galectin-9 mAb was validated for its use in mice on isolated murine C57BL/6 Treg by immunofluorescence staining and cytometry. Its specificity and functionality were validated in proliferation assays on rLGALS9-immunosuppressed murine Tconv and in suppression assays between murine Treg and Tconv. Finally, 2-month-old KC mice were treated with anti-LGALS9 and compared to WT mice for peripheral and infiltrating TCD4, Treg, and PanIN progression. Results: IHC and cytometry revealed a significant increase in LGALS9 expression and Treg levels in the blood and pancreas of KC mice proportional to the stages of precancerous lesions. Although present in WT mice, LGALS9 is expressed at a basal level with low and restricted expression that increases slightly over time, while Treg cells are few in number in their circulation and even absent from the pancreas over time. Using our anti-LGALS9 mAb in mice, it is shown that (i) murine Treg express LGALS9, (ii) the mAb could target and inhibit recombinant murine LGALS9, and (iii) neutralize murine Treg suppressive activity. Finally, the anti-LGALS9 mAb in KC mice reduced (i) LGALS9 expression in pancreatic cancer cells, (ii) the Treg ratio, and (iii) the total surface area and grade of PanIN. Conclusion: We demonstrate for the first time that an anti-LGALS9 antibody, by specifically targeting endogenous LGALS9 tumor and exogenous LGALS9 produced by Treg, was able to limit the progression of pancreatic neoplastic lesions in mice, opening up new prospects for its use as an immunotherapeutic tool in PDAC.
Assuntos
Adenocarcinoma , Carcinoma in Situ , Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Camundongos , Animais , Camundongos Transgênicos , Camundongos Endogâmicos C57BL , Carcinoma in Situ/metabolismo , Carcinoma in Situ/patologia , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Galectinas , Imunoterapia , Microambiente TumoralRESUMO
Nasopharyngeal carcinoma-derived small extracellular vesicles (NPCSEVs) have an immunosuppressive impact on the tumour microenvironment. In this study, we investigated their influence on the generation of tolerogenic dendritic cells and the potential involvement of the galectin-9 (Gal9) they carry in this process. We analysed the phenotype and immunosuppressive properties of NPCSEVs and explored the ability of DCs exposed to NPCSEVs (NPCSEV-DCs) to regulate T cell proliferation. To assess their impact at the pathophysiological level, we performed real-time fluorescent chemoattraction assays. Finally, we analysed phenotype and immunosuppressive functions of NPCSEV-DCs using a proprietary anti-Gal9 neutralising antibody to assess the role of Gal9 in this effect. We described that NPCSEV-DCs were able to inhibit T cell proliferation despite their mature phenotype. These mature regulatory DCs (mregDCs) have a specific oxidative metabolism and secrete high levels of IL-4. Chemoattraction assays revealed that NPCSEVs could preferentially recruit NPCSEV-DCs. Finally, and very interestingly, the reduction of the immunosuppressive function of NPCSEV-DCs using an anti-Gal9 antibody clearly suggested an important role for vesicular Gal9 in the induction of mregDCs. These results revealed for the first time that NPCSEVs promote the emergence of mregDCs using a galectin-9 dependent mechanism and open new perspectives for antitumour immunotherapy targeting NPCSEVs.
Assuntos
Vesículas Extracelulares , Neoplasias Nasofaríngeas , Humanos , Carcinoma Nasofaríngeo , Células Dendríticas , Galectinas/metabolismo , Neoplasias Nasofaríngeas/metabolismo , Microambiente TumoralRESUMO
The purpose of this study was to assess the direct effect of CCL18, a chemokine elevated in allergic diseases and induced by Th2 cytokines, on the polarization of human CD4(+) T cells. Purified human T cells from healthy subjects were pretreated or not with CCL18, and evaluated for cytokine production. CCL18-pretreated memory but not naive CD4(+) T cells exhibited an increased production of IL-10 (12.3 ± 2.6 vs. 5.6 ± 0.9 ng/ml for medium) and TGF-ß1 but not IL-4, IFN-γ, and IL-17 compared with control cells. Pretreatment of highly purified CD4(+)CD25(-) memory T cells with CCL18 led to their conversion to CD4(+)CD25(+)Foxp3(+) regulatory T cells able to inhibit the proliferation of CD4(+)CD25(-) effector T cells by both cytokine and cell contact-dependent mechanisms. However, this regulatory effect of CCL18 was lost when T cells originated from allergic subjects in relation with a decreased binding of CCL18 to these cells [0.7 ± 0.3 mean fluorescence intensity (MFI)] as compared to those from healthy subjects (6.0 ± 1.7 MFI). This study is the first to define a chemokine that generates adaptive regulatory T cells from CD4(+)CD25(-) memory T cells. This mechanism appears defective in allergic patients and may underlie the decreased tolerance observed in allergic diseases.
Assuntos
Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/metabolismo , Quimiocinas CC/farmacologia , Hipersensibilidade/imunologia , Hipersensibilidade/metabolismo , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/metabolismo , Linfócitos T CD4-Positivos/efeitos dos fármacos , Humanos , Interleucina-10/metabolismo , Interleucina-13/metabolismo , Linfócitos T Reguladores/efeitos dos fármacos , Fator de Crescimento Transformador beta1/metabolismoRESUMO
Photodynamic Therapy (PDT) relies on local or systemic administration of a light-sensitive dye, called photosensitizer, to accumulate into the target site followed by excitation with light of appropriate wavelength and fluence. This photo-activated molecule reacts with the intracellular oxygen to induce selective cytotoxicity of targeted cells by the generation of reactive oxygen species. Hepatocellular carcinoma (HCC), one of the leading causes of cancer-associated mortality worldwide, has insufficient treatment options available. In this review, we discuss the mechanism and merits of PDT along with its recent developments as an anti-cancerous therapy. We also highlight the application of this novel therapy for diagnosis, visualization, and treatment of HCC. We examine the underlying challenges, some pre-clinical and clinical studies, and possibilities of future studies associated with PDT. Finally, we discuss the mechanism of an active immune response by PDT and thereafter explored the role of PDT in the generation of anti-tumor immune response in the context of HCC, with an emphasis on checkpoint inhibitor-based immunotherapy. The objective of this review is to propose PDT as a plausible adjuvant to existing therapies for HCC, highlighting a feasible combinatorial approach for HCC treatment.
RESUMO
There has been very limited use of computer assisted semen analysis (CASA) to evaluate reptile sperm. The aim of this study was to examine sperm kinematic variables in American crocodile (Crocodylus acutus) semen samples and to assess whether sperm subpopulations could be characterized. Eight ejaculates (two ejaculates/male) from four sexually mature captive crocodiles were obtained. An ISAS®v1 CASA-Mot system, with an image acquisition rate of 50 Hz, and ISAS®D4C20 counting chambers were used for sperm analyses. The percentages of motile and progressively motile spermatozoa did not differ among animals (P > 0.05) but there was a significant animal effect with regards to kinematic variables (P < 0.05). Principal component (PC) analysis revealed that kinematic variables grouped into three components: PC1, related to velocity; PC2 to progressiveness and PC3 to oscillation. Subpopulation structure analysis identified four groups (P < 0.05), which represented, on average, 9.8%, 32.1%, 26.8%, and 31.3% of the total sperm population. Males differed in the proportion of sperm in each of the kinematic subpopulations. This new approach for the analysis of reptile sperm kinematic subpopulations, reflecting quantifiable parameters generated by CASA system technology, opens up possibilities for future assessments of crocodile sperm and will be useful in the future development of assisted reproduction for these species.
Assuntos
Jacarés e Crocodilos/genética , Linhagem da Célula/genética , Reprodução/genética , Espermatozoides/citologia , Animais , Fenômenos Biomecânicos , Masculino , Sêmen/citologia , Sêmen/fisiologia , Análise do Sêmen , Preservação do Sêmen , Motilidade dos Espermatozoides/genética , Espermatozoides/fisiologia , Estados UnidosRESUMO
To date, pancreatic adenocarcinoma (ADKP) is a devastating disease for which the incidence rate is close to the mortality rate. The survival rate has evolved only 2-5% in 45 years, highlighting the failure of current therapies. Otherwise, the use of photodynamic therapy (PDT), based on the use of an adapted photosensitizer (PS) has already proved its worth and has prompted a growing interest in the field of oncology. We have developed a new photosensitizer (PS-FOL/PS2), protected by a recently published patent (WO2019 016397-A1, 24 January 2019). This photosensitizer is associated with an addressing molecule (folic acid) targeting the folate receptor 1 (FOLR1) with a high affinity. Folate binds to FOLR1, in a specific way, expressed in 100% of ADKP or over-expressed in 30% of cases. The first objective of this study is to evaluate the effectiveness of this PS2-PDT in four ADKP cell lines: Capan-1, Capan-2, MiapaCa-2, and Panc-1. For this purpose, we first evaluated the gene and protein expression of FOLR1 on four ADKP cell lines. Subsequently, we evaluated PS2's efficacy in our cell lines and we assessed the impact of PDT on the secretome of cancer cells and its impact on the immune system. Finally, we evaluate the PDT efficacy on a humanized SCID mouse model of pancreatic cancer. In a very interesting way, we observed a significant increase in the proliferation of activated-human PBMC when cultured with conditioned media of ADKP cancer cells subjected to PDT. Furthermore, to evaluate in vivo the impact of this new PS, we analyzed the tumor growth in a humanized SCID mice model of pancreatic cancer. Four conditions were tested: Untreated, mice (nontreated), mice with PS (PS2), mice subjected to illumination (Light only), and mice subjected to illumination in the presence of PS (PDT). We noticed that the mice subjected to PDT presented a strong decrease in the growth of the tumor over time after illumination. Our investigations have not only suggested that PS2-PDT is an effective therapy in the treatment of PDAC but also that it activates the immune system and could be considered as a real adjuvant for anti-cancer vaccination. Thus, this new study provides new treatment options for patients in a therapeutic impasse and will provide a new arsenal in the fight against PDAC.