RESUMO
Gut bacteria modulate the response to immune checkpoint blockade (ICB) treatment in cancer, but the effect of diet and supplements on this interaction is not well studied. We assessed fecal microbiota profiles, dietary habits, and commercially available probiotic supplement use in melanoma patients and performed parallel preclinical studies. Higher dietary fiber was associated with significantly improved progression-free survival in 128 patients on ICB, with the most pronounced benefit observed in patients with sufficient dietary fiber intake and no probiotic use. Findings were recapitulated in preclinical models, which demonstrated impaired treatment response to antiprogrammed cell death 1 (antiPD-1)based therapy in mice receiving a low-fiber diet or probiotics, with a lower frequency of interferon-γpositive cytotoxic T cells in the tumor microenvironment. Together, these data have clinical implications for patients receiving ICB for cancer.
Assuntos
Fibras na Dieta , Microbioma Gastrointestinal , Inibidores de Checkpoint Imunológico/uso terapêutico , Melanoma/terapia , Probióticos , Animais , Estudos de Coortes , Ácidos Graxos Voláteis/análise , Transplante de Microbiota Fecal , Fezes/química , Fezes/microbiologia , Feminino , Humanos , Imunoterapia , Masculino , Melanoma/imunologia , Melanoma/microbiologia , Melanoma Experimental/imunologia , Melanoma Experimental/microbiologia , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Intervalo Livre de Progressão , Linfócitos TRESUMO
Antibiotic-induced microbial imbalance, or dysbiosis, has systemic and long-lasting effects on the host and response to cancer therapies. However, the effects on tumor endothelial cells are largely unknown. Therefore, the goal of the current study was to generate matched B16-F10 melanoma associated endothelial cell lines isolated from mice with and without antibiotic-induced dysbiosis. After validating endothelial cell markers on a genomic and proteomic level, functional angiogenesis assays (i.e., migration and tube formation) also confirmed their vasculature origin. Subsequently, we found that tumor endothelial cells derived from dysbiotic mice (TEC-Dys) were more sensitive to ionizing radiotherapy in the range of clinically-relevant hypofractionated doses, as compared to tumor endothelial cells derived from orthobiotic mice (TEC-Ortho). In order to identify tumor vasculature-associated drug targets during dysbiosis, we used tandem mass tag mass spectroscopy and focused on the statistically significant cellular membrane proteins overexpressed in TEC-Dys. By these criteria c-Met was the most differentially expressed protein, which was validated histologically by comparing tumors with or without dysbiosis. Moreover, in vitro, c-Met inhibitors Foretinib, Crizotinib and Cabozantinib were significantly more effective against TEC-Dys than TEC-Ortho. In vivo, Foretinib inhibited tumor growth to a greater extent during dysbiosis as compared to orthobiotic conditions. Thus, we surmise that tumor response in dysbiotic patients may be greatly improved by targeting dysbiosis-induced pathways, such as c-Met, distinct from the many targets suppressed due to dysbiosis.
Assuntos
Disbiose , Células Endoteliais/enzimologia , Melanoma Experimental , Neovascularização Patológica , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-met , Animais , Disbiose/enzimologia , Disbiose/microbiologia , Melanoma Experimental/irrigação sanguínea , Melanoma Experimental/enzimologia , Melanoma Experimental/microbiologia , Melanoma Experimental/terapia , Camundongos , Neovascularização Patológica/enzimologia , Neovascularização Patológica/microbiologia , Neovascularização Patológica/terapia , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-met/metabolismo , RadioterapiaRESUMO
VNP20009, an attenuated mutant of Salmonella, is potentially applied for tumor therapy due to its specific accumulation and proliferation in the hypoxic zone of tumor. However, studies have shown that human immunity system and the associated toxicities of attenuated Salmonella evidently alleviated the anti-tumor effect when tumor is reduced. As apoptosis-inducing factor (AIF) can directly induce nuclear apoptosis in the absence of caspases to avoid unwished apoptosis in normal cells, therefore, a eukaryotic expressing VNP20009-AbVec-Igκ-AIF (V-A-AIF) strain was constructed in the present study, and its anti-melanoma effects were evaluated in vitro and in vivo. The results showed that AIF expressed by the V-A-AIF strain significantly enhanced the apoptosis of B16F10 cells in vitro, seen as remarkable decrease of tumor volume, formation of larger necrotic areas, and prolongation of the lifespan in a melanoma-bearing mouse model. Furthermore, we observed that the colonization of the V-A-AIF strain and the massive expression of AIF in tumors significantly promoted apoptosis of tumor cells by upregulating the expression ratio of Bcl-2-associated X protein/B cell lymphoma-2 (Bax/Bcl-2), suppressed the inflammatory response by downregulating toll-like receptor-4/nuclear factor kappa-B (TLR-4/NFκB) signaling pathway, seen as reduction of the expressions of phosphorylated phosphoinositide 3 kinase (PI3K) and protein kinase B (AKT), and decrease of the production of interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α). Our study demonstrated that the colonization of the V-A-AIF strain in tumor triggers a decent anti-tumor effect in vivo and in vitro, suggesting that the engineered strain may provide a potential reagent for cancer therapy.
Assuntos
Anticarcinógenos/uso terapêutico , Fator de Indução de Apoptose/genética , Apoptose , Melanoma Experimental/microbiologia , Melanoma Experimental/terapia , Salmonella/genética , Animais , Linhagem Celular Tumoral , Proliferação de Células , Modelos Animais de Doenças , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Plasmídeos/genética , Salmonella/fisiologiaRESUMO
Loss of target antigens in tumor cells has become one of the major hurdles limiting the efficacy of adoptive cell therapy (ACT)-based immunotherapies. The optimal approach to overcome this challenge includes broadening the immune response from the initially targeted tumor-associated antigen (TAA) to other TAAs expressed in the tumor. To induce a more broadly targeted antitumor response, we utilized our previously developed Re-energized ACT (ReACT), which capitalizes on the synergistic effect of pathogen-based immunotherapy and ACT. In this study, we showed that ReACT induced a sufficient endogenous CD8+ T-cell response beyond the initial target to prevent the outgrowth of antigen loss variants in a B16-F10 melanoma model. Sequentially, selective depletion experiments revealed that Batf3-driven cDC1s were essential for the activation of endogenous tumor-specific CD8+ T cells. In ReACT-treated mice that eradicated tumors, we observed that endogenous CD8+ T cells differentiated into memory cells and facilitated the rejection of local and distal tumor rechallenge. By targeting one TAA with ReACT, we provided broader TAA coverage to counter antigen escape and generate a durable memory response against local relapse and metastasis.See related Spotlight on p. 2.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Imunoterapia Adotiva/métodos , Listeria monocytogenes/patogenicidade , Listeriose/complicações , Melanoma Experimental/imunologia , Neoplasias Cutâneas/imunologia , Antígeno gp100 de Melanoma/imunologia , Animais , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Linhagem Celular Tumoral , Feminino , Memória Imunológica , Listeriose/imunologia , Listeriose/microbiologia , Melanoma Experimental/metabolismo , Melanoma Experimental/microbiologia , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/microbiologia , Neoplasias Cutâneas/terapiaRESUMO
OBJECTIVES: Mycobacterium indicus pranii (MIP) is an atypical mycobacterium species with potent antitumor efficacy. Macrophages and dendritic cells (DCs) are antigen-presenting cells, playing key roles in the activation of antitumor immunity. We have previously shown the potent activation of macrophages and DCs by MIP, which is mediated by MyD88-TLR2 signaling axis. In the present study, we further examined the role of MyD88 and TLR2 in MIP-mediated tumor regression. RESULTS: Wild-type and MyD88-/- mice were implanted with B16F10 tumor cells, treated with MIP or phosphate-buffered saline (PBS) and monitored for tumor growth. As expected, MIP therapy led to significant tumor regression in wild-type mice. However, antitumor efficacy of MIP was lost in MyD88-/- animals. Both PBS-treated (control) and MIP-treated MyD88-/- mice developed tumors with comparable volume. Since MyD88 relays TLR engagement signals, we analyzed the antitumor efficacy of MIP in TLR2-/- and TLR4-/- mice. It was observed that MIP therapy reduced tumor burden in wild-type and TLR4-/- mice but not in TLR2-/- mice. Tumor volume in MIP-treated TLR2-/- mice were comparable with those in PBS-treated wild-type animals. These results implicated the MyD88-TLR2 signaling axis in the antitumor efficacy of MIP.
Assuntos
Melanoma Experimental/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Micobactérias não Tuberculosas/imunologia , Receptor 2 Toll-Like/imunologia , Carga Tumoral/imunologia , Animais , Linhagem Celular Tumoral , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Humanos , Macrófagos/imunologia , Macrófagos/metabolismo , Melanoma Experimental/genética , Melanoma Experimental/microbiologia , Melanoma Experimental/terapia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Micobactérias não Tuberculosas/fisiologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia , Receptor 4 Toll-Like/metabolismo , Carga Tumoral/genéticaRESUMO
The overall use of antibiotics has increased significantly in recent years. Besides fighting infections, antibiotics also alter the gut microbiota. Commensal bacteria in the gastrointestinal tract are crucial to maintain immune homeostasis, and microbial imbalance or dysbiosis affects disease susceptibility and progression. We hypothesized that antibiotic-induced dysbiosis of the gut microbiota would suppress cytokine profiles in the host, thereby leading to changes in the tumor microenvironment. The induced dysbiosis was characterized by alterations in bacterial abundance, composition, and diversity in our animal models. On the host side, antibiotic-induced dysbiosis caused elongated small intestines and ceca, and B16-F10 melanoma and Lewis lung carcinoma progressed more quickly than in control mice. Mechanistic studies revealed that this progression was mediated by suppressed TNFα levels, both locally and systemically, resulting in reduced expression of tumor endothelial adhesion molecules, particularly intercellular adhesion molecule-1 (ICAM-1) and a subsequent decrease in the number of activated and effector CD8+ T cells in the tumor. However, suppression of ICAM-1 or its binding site, the alpha subunit of lymphocyte function-associated antigen-1, was not seen in the spleen or thymus during dysbiosis. TNFα supplementation in dysbiotic mice was able to increase ICAM-1 expression and leukocyte trafficking into the tumor. Overall, these results demonstrate the importance of commensal bacteria in supporting anticancer immune surveillance, define an important role of tumor endothelial cells within this process, and suggest adverse consequences of antibiotics on cancer control. SIGNIFICANCE: Antibiotic-induced dysbiosis enhances distal tumor progression by altering host cytokine levels, resulting in suppression of tumor endothelial adhesion molecules and activated and effector CD8+ T cells in the tumor.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Carcinoma Pulmonar de Lewis/imunologia , Disbiose/imunologia , Linfócitos do Interstício Tumoral/imunologia , Melanoma Experimental/imunologia , Animais , Antibacterianos/efeitos adversos , Carcinoma Pulmonar de Lewis/microbiologia , Carcinoma Pulmonar de Lewis/patologia , Progressão da Doença , Disbiose/induzido quimicamente , Endotélio/imunologia , Endotélio/metabolismo , Endotélio/patologia , Microbioma Gastrointestinal/efeitos dos fármacos , Microbioma Gastrointestinal/imunologia , Molécula 1 de Adesão Intercelular/imunologia , Molécula 1 de Adesão Intercelular/metabolismo , Melanoma Experimental/microbiologia , Melanoma Experimental/patologia , Camundongos , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Chemotherapy is one of effective methods for the treatment of tumor. Patients often develop drug resistance after chemotherapic cycles. Salmonella has potential as antitumor agent. Salmonella used in tandem with chemotherapy had additive effects, providing a rationale for using tumor-targeting Salmonella in combination with conventional chemotherapy. To improve the efficacy and safety of Salmonella, a further understanding of Salmonella interactions with the tumor microenvironment is required. The presence of plasma membrane multidrug resistance protein P-glycoprotein (P-gp) is highly relevant for the success of chemotherapy. Following Salmonella infection, dose-dependent downregulation of P-gp expressions were examined. Salmonella significantly decreased the efflux capabilities of P-gp, as based on the influx of Rhodamine 123 assay. In addition, Salmonella significant reduced the protein express the expression levels of phosph-protein kinase B (P-AKT), phosph-mammalian targets of rapamycin (P-mTOR), and phosph-p70 ribosomal s6 kinase (P-p70s6K) in tumor cells. The Salmonella-induced downregulation of P-gp was rescued by transfection of cells with active P-AKT. Our results demonstrate that Salmonella in tumor sites leads to decrease the expression of P-gp and enhances the combination of Salmonella and 5-Fluorouracil therapeutic effects.
Assuntos
Fluoruracila/administração & dosagem , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/microbiologia , Infecções por Salmonella/genética , Salmonella/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Animais , Membrana Celular/efeitos dos fármacos , Membrana Celular/genética , Terapia Combinada , Resistência a Múltiplos Medicamentos/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Melanoma Experimental/patologia , Camundongos , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Quinases S6 Ribossômicas 70-kDa/genética , Salmonella/patogenicidade , Infecções por Salmonella/microbiologia , Infecções por Salmonella/patologia , Serina-Treonina Quinases TOR/genética , Microambiente Tumoral/efeitos dos fármacosRESUMO
AIM: We evaluated a novel approach combining the use of attenuated Salmonella immunotherapy with a Toll-like receptor agonist, imiquimod, in B16F1 melanoma-bearing mice. MATERIALS & METHODS: B16F1 melanoma-bearing mice were daily treated with topical imiquimod in combination with one intratumoral injection of attenuated Salmonella enterica serovar Typhimurium LVR01. RESULTS: The combined therapy resulted in retarded tumor growth and prolonged survival. Combination treatment led to an enhancement in the expression of pro-inflammatory cytokines and chemokines in the tumor microenvironment, with a Th1-skewed profile, resulting in a broad antitumor response. The induced immunity was effective in controlling the occurrence of metastasis. CONCLUSION: Salmonella LVR01 immunotherapy in combination with imiquimod is a novel approach that could be considered as an effective antimelanoma therapy.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Imunoterapia/métodos , Melanoma Experimental/terapia , Salmonella typhi/imunologia , Receptor 7 Toll-Like/agonistas , Animais , Antineoplásicos/uso terapêutico , Morte Celular , Linhagem Celular Tumoral , Feminino , Imiquimode/uso terapêutico , Melanoma Experimental/imunologia , Melanoma Experimental/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Viabilidade Microbiana , Metástase Neoplásica/prevenção & controle , Salmonella typhi/fisiologia , Análise de Sobrevida , Resultado do Tratamento , Carga Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologiaRESUMO
Microbiota maintains host tissue homeostasis and influences tissue-resident macrophages. However, the mechanisms by which commensal bacteria in regulating the alveolar macrophages remain unclear. Here, by using an antibiotic-treated (Abt) mouse model, we found commensal bacteria depletion induced lower frequencies and numbers of alveolar macrophages. This effect was accompanied by the altered levels of genes involved in several biological pathways, including M2 macrophage polarization, as determined by gene expression analysis. Alveolar macrophages from the Abt mice had higher protein and gene levels of Arg1, CCL24, IL-13, IL-10, IL-6 and IL-1ß, which could be recovered to normal levels by reconstructing commensal bacteria in the upper respiratory of Abt mice. Moreover, alveolar macrophages performed significant enhancement of M2 functions, especially CCL24 secretion, in the Abt mice challenged with B16/F10 melanoma. Adoptive transfer of normal alveolar macrophages or antibody neutralization of CCL24 significantly recovered the decrease of γδT17 cells and rescued the defect anti-tumor response of Abt mice, indicating the elevated amount of alveolar macrophage-derived CCL24 inhibited γδT cell mediated anti-tumor response. In conclusion, we demonstrated the ability of commensal bacteria to maintain alveolar macrophages with a low level of CCL24 production, which was necessary for the normal anti-tumor response in the lung.
Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Quimiocina CCL24/metabolismo , Macrófagos Alveolares/imunologia , Melanoma Experimental/terapia , Animais , Bactérias/classificação , Bactérias/imunologia , Fenômenos Fisiológicos Bacterianos , Linhagem Celular Tumoral , Citocinas/genética , Citocinas/metabolismo , Feminino , Linfócitos Intraepiteliais/efeitos dos fármacos , Linfócitos Intraepiteliais/imunologia , Macrófagos Alveolares/efeitos dos fármacos , Melanoma Experimental/imunologia , Melanoma Experimental/microbiologia , Camundongos , Microbiota , Análise de Sequência de DNA , Transdução de Sinais , SimbioseRESUMO
VNP20009 is a very effective anti-cancer agent and can specifically target tumors and inhibit tumor growth. It was assumed that the tumor targeting ability of VNP20009 correlated to its anticancer capacity. However, our observation contradicted to this assumption. Three VNP20009 mutant strains (ΔslyA, ΔSTM3120 and ΔhtrA) with reduced fitness in normal tissues and unchanged fitness in tumors partially or completely lost their anti-cancer capacities. The genes slyA, STM3120 and htrA were required for survival within macrophages and were indispensable for tumor microenvironment remodeling by VNP20009. The infiltration of immune cells occurred less in the tumors of mice infected with the mutant strains. In addition, the mRNA levels of TNF-α and IL-1ß were significantly decreased in the tumors of mice treated with the mutant strains. Our results indicate that the immune responses elicited by bacteria rather than the bacterial titer in tumors play a "decisive" role in VNP20009-mediated bacterial cancer therapy, which provides a novel perspective for the underlying mechanism of bacterial cancer therapy.
Assuntos
Proteínas de Bactérias/genética , Vacinas Bacterianas/farmacologia , Proteínas de Choque Térmico/genética , Melanoma Experimental/terapia , Proteínas Periplásmicas/genética , Salmonella enterica/genética , Serina Endopeptidases/genética , Neoplasias Cutâneas/terapia , Fatores de Transcrição/genética , Animais , Proteínas de Bactérias/imunologia , Proteínas de Choque Térmico/imunologia , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Macrófagos/imunologia , Macrófagos/microbiologia , Macrófagos/patologia , Melanoma Experimental/imunologia , Melanoma Experimental/microbiologia , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Proteínas Periplásmicas/imunologia , Células RAW 264.7 , Salmonella enterica/imunologia , Serina Endopeptidases/imunologia , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/microbiologia , Neoplasias Cutâneas/patologia , Fatores de Tempo , Fatores de Transcrição/imunologia , Carga Tumoral , Microambiente Tumoral , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Bacteria-based living anticancer agents have emerged as promising therapeutics. However, the functional role of autophagy in bacterial cancer therapy has been little investigated. In this study, Salmonella VNP20009 induced autophagy in B16F10 cells, which is an unfavorable factor in bacterial cancer therapy. Inhibiting the induction of autophagy by chloroquine or siRNA in bacterial cancer therapy dose- and time-dependently promoted cell death. The combined therapy of VNP20009 and chloroquine not only enhanced the bacterial tumor targeting ability but also facilitated the infiltration of immune cells into the tumor. Our results showed that the combined therapy of VNP20009 and chloroquine could significantly inhibit tumor growth and prolong mouse survival time. This study provides a novel strategy for improving the anti-cancer efficacy of bacterial cancer therapy.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Autofagia/efeitos dos fármacos , Melanoma Experimental/tratamento farmacológico , Infecções por Salmonella/tratamento farmacológico , Salmonella typhimurium/efeitos dos fármacos , Animais , Vacinas Bacterianas/administração & dosagem , Linhagem Celular Tumoral , Cloroquina/administração & dosagem , Sinergismo Farmacológico , Humanos , Melanoma Experimental/microbiologia , Melanoma Experimental/patologia , Camundongos , Infecções por Salmonella/microbiologia , Infecções por Salmonella/patologia , Salmonella typhimurium/imunologia , Salmonella typhimurium/fisiologia , Análise de Sobrevida , Carga Tumoral/efeitos dos fármacosRESUMO
Salmonella is a Gram-negative, facultative anaerobe that is a common cause of host intestinal infections. Salmonella grows under aerobic and anaerobic conditions, and it has been proven capable of inhibiting tumor growth. However, the molecular mechanism by which Salmonella inhibits tumor growth is still unclear. Angiogenesis plays an important role in the development and progression of tumors. We investigated the antitumor effect of Salmonella in a syngeneic murine tumor model. Hypoxia-inducible factor-1 (HIF-1)α plays a significant role in tumor angiogenesis. We examined the molecular mechanism by which Salmonella regulated vascular endothelial growth factor (VEGF), which is an important angiogenic factor. The expression of VEGF in tumor cells was decreased by treatment with Salmonella. The conditioned medium from Salmonella-treated cells inhibited the proliferation of endothelial cells. Salmonella inhibited the expression of HIF-1α as well as downregulated its upstream signal mediator protein kinase B (AKT). Salmonella significantly inhibited tumor growth in vivo, and immunohistochemical studies of the tumors revealed decreased intratumoral microvessel density. These results suggest that Salmonella therapy, which exerts anti-angiogenic activities, represents a promising strategy for the treatment of tumors.
Assuntos
Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias Mamárias Experimentais/irrigação sanguínea , Neoplasias Mamárias Experimentais/microbiologia , Melanoma Experimental/irrigação sanguínea , Melanoma Experimental/microbiologia , Salmonella enterica/fisiologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Meios de Cultivo Condicionados , Regulação para Baixo , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Células Endoteliais/microbiologia , Feminino , Humanos , Neoplasias Mamárias Experimentais/metabolismo , Melanoma Experimental/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Neovascularização Patológica/metabolismo , Neovascularização Patológica/microbiologia , Neovascularização Patológica/patologia , Salmonella enterica/metabolismo , Transdução de Sinais , TransfecçãoRESUMO
In multicellular organisms, cell motility is central in all morphogenetic processes, tissue maintenance, wound healing and immune surveillance. Hence, failures in its regulation potentiates numerous diseases. Here, cell migration assays on plastic 2D surfaces were performed using normal (Melan A) and tumoral (B16F10) murine melanocytes in random motility conditions. The trajectories of the centroids of the cell perimeters were tracked through time-lapse microscopy. The statistics of these trajectories was analyzed by building velocity and turn angle distributions, as well as velocity autocorrelations and the scaling of mean-squared displacements. We find that these cells exhibit a crossover from a normal to a super-diffusive motion without angular persistence at long time scales. Moreover, these melanocytes move with non-Gaussian velocity distributions. This major finding indicates that amongst those animal cells supposedly migrating through Lévy walks, some of them can instead perform q-Gaussian walks. Furthermore, our results reveal that B16F10 cells infected by mycoplasmas exhibit essentially the same diffusivity than their healthy counterparts. Finally, a q-Gaussian random walk model was proposed to account for these melanocytic migratory traits. Simulations based on this model correctly describe the crossover to super-diffusivity in the cell migration tracks.
Assuntos
Melanócitos/citologia , Melanócitos/patologia , Melanoma Experimental/patologia , Modelos Biológicos , Animais , Movimento Celular , Melanoma Experimental/complicações , Melanoma Experimental/microbiologia , Camundongos , Infecções por Mycoplasma/complicações , Invasividade Neoplásica , Distribuição NormalRESUMO
Commensal bacteria are crucial to maintain immune homeostasis in mucosal tissues and disturbances in their ecology can affect disease susceptibility. Here, we report evidence that commensal bacteria shape the efficiency of immune surveillance in mucosal tissues. Antibiotic-treated (Abt) mice were more susceptible to development of engrafted B16/F10 melanoma and Lewis lung carcinoma, exhibiting a shortened mean survival time with more numerous and larger tumor foci in the lungs. The defective antitumor response of Abt mice was independent of dehydration caused by antibiotics. Host defenses relied upon intact commensal bacteria with no class specificity. Mechanistic investigations revealed a defective induction of the γδT17 cell response in lungs of Abt mice; here, more aggressive tumor development was observed, possibly related to a reduction in IL6 and IL23 expression there. Adding normal γδT cells or supplementing IL17 restored the impaired immune surveillance phenotype in Abt mice. Overall, our results demonstrated the importance of commensal bacteria in supporting the host immune response against cancer, defined an important role for γδT17 responses in the mechanism, and suggested deleterious effects of antibiotic treatment on cancer susceptibility and progression.
Assuntos
Carcinoma Pulmonar de Lewis/imunologia , Carcinoma Pulmonar de Lewis/microbiologia , Interleucina-17/imunologia , Melanoma Experimental/imunologia , Melanoma Experimental/microbiologia , Microbiota/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Animais , Feminino , Vigilância Imunológica , Camundongos , Camundongos Endogâmicos C57BL , Linfócitos T/imunologiaRESUMO
BACKGROUND: Melanocytes are cells located in epidermis and mucous membranes that synthesize melanin and cytokines. It is known that melanin has antimicrobial activity and that melanocytes are melanized in presence of microbial molecules. OBJECTIVE: To study the antifungal activity of melanin on Candida spp. METHODOLOGY: The minimum inhibitory concentration (MIC) to melanin was determined in 4 Candida ATCC strains (C. albicans SC5314, C. parapsilosis 22019, C. glabrata 2001, C. krusei 6258) and 56 clinical isolates of Candida spp. (33 C. albicans, 12 C. glabrata, 3 C. famata, 3 C. krusei, 3 C. parapsilosis, 2 C. tropicalis) using a broth microdilution method. In addition, the antifungal activity of melanocytes and mice melanoma cells was tested against C. albicans. RESULTS: Melanin inhibited the tested isolates, including the susceptible dose-dependent and fluconazole-resistant strains; MIC range and MIC50 were 0.09-50 µg/mL and 6.25 µg/mL, respectively. Pigmented cells lysates inhibited C. albicans. CONCLUSIONS: Melanin is able to inhibit clinical isolates of Candida spp. Melanization could be an important protective mechanism of melanocytes.
Assuntos
Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Fluconazol/farmacologia , Melaninas/farmacologia , Melanócitos/imunologia , Animais , Candida albicans/classificação , Candida albicans/crescimento & desenvolvimento , Farmacorresistência Fúngica , Melaninas/metabolismo , Melanócitos/metabolismo , Melanoma Experimental/metabolismo , Melanoma Experimental/microbiologia , Camundongos , Pigmentação da PeleRESUMO
Background: Melanocytes are cells located in epidermis and mucous membranes that synthesize melanin and cytokines. It is known that melanin has antimicrobial activity and that melanocytes are melanized in presence of microbial molecules. Objective: To study the antifungal activity of melanin on Candida spp. Methodology: The minimum inhibitory concentration (MIC) to melanin was determined in 4 Candida ATCC strains (C. albicans SC5314, C. parapsilosis 22019, C. glabrata 2001, C. krusei 6258) and 56 clinical isolates of Candida spp. (33 C. albicans, 12 C. glabrata, 3 C. famata, 3 C. krusei, 3 C. parapsilosis, 2 C. tropicalis) using a broth microdilution method. In addition, the antifungal activity of melanocytes and mice melanoma cells was tested against C. albicans. Results: Melanin inhibited the tested isolates, including the susceptible dose-dependent and fluconazole-resistant strains; MIC range and MIC50 were 0.09-50 μg/mL and 6.25 μg/mL, respectively. Pigmented cells lysates inhibited C. albicans. Conclusions: Melanin is able to inhibit clinical isolates of Candida spp. Melanization could be an important protective mechanism of melanocytes.
Introducción: Los melanocitos son células presentes en piel y en mucosas que sintetizan melanina, además de citoquinas. Es sabido que melanina presenta actividad antimicrobiana y que los melanocitos se melanizan al ser expuestos a moléculas microbianas. Objetivo: Estudiar la actividad antifúngica de melanina en cepas clínicas de Candida spp. Metodología: Se midió la concentración inhibitoria mínima (CIM) a melanina, de 4 cepas de Candida ATCC (C. albicans SC5314, C. parapsilosis 22019, C. glabrata 2001 y C. krusei 6258) y 56 aislados clínicos de Candida spp. (33 C. albicans, 12 C. glabrata, 3 C. famata, 3 C. krusei, 3 C. parapsilosis, 2 C. tropicalis) mediante un método de microdilución en caldo. Además se estudió el efecto antifúngico de lisados de melanocitos y células de melanoma de ratón en C. albicans. Resultados: Melanina inhibió las cepas analizadas, incluso cepas susceptibles dosis-dependiente y resistentes a fluconazol, siendo los rangos de CIM y CIM50 de 0,09-50 μg/mL y 6,25 μg/ mL, respectivamente. Los lisados de células pigmentadas inhibieron C. albicans. Conclusiones: Melanina es capaz de inhibir cepas clínicas de Candida spp. La melanización podría ser un importante mecanismo protector de los melanocitos.
Assuntos
Animais , Camundongos , Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Fluconazol/farmacologia , Melaninas/farmacologia , Melanócitos/imunologia , Candida albicans/classificação , Candida albicans/crescimento & desenvolvimento , Farmacorresistência Fúngica , Melaninas/metabolismo , Melanócitos/metabolismo , Melanoma Experimental/metabolismo , Melanoma Experimental/microbiologia , Pigmentação da PeleRESUMO
Many studies have demonstrated that intravenously administered bacteria can target and proliferate in solid tumors and then quickly be released from other organs. Here, we employed the tumor-targeting property of Escherichia coli Nissle 1917 to inhibit mouse B16 melanoma and 4T1 breast tumors through the expression of azurin protein. For this purpose, recombinant azurin-expressing E. coli Nissle 1917 was developed. The levels of in vitro and in vivo azurin secretion in the engineered bacterium were determined by immunochemistry. Our results demonstrated that B16 melanoma and orthotopic 4T1 breast tumor growth were remarkably restrained and pulmonary metastasis was prevented in immunocompetent mice. It is worth noting that this therapeutic effect partially resulted from the antitumor activity of neutrophils and lymphocytes due to inflammatory responses caused by bacterial infections. No toxicity was observed in the animal during the experiments. This study indicates that E. coli Nissle 1917 could be a potential carrier to deliver antitumor drugs effectively for cancer therapy.
Assuntos
Azurina/biossíntese , Sistemas de Liberação de Medicamentos , Escherichia coli , Neoplasias Mamárias Experimentais/terapia , Melanoma Experimental/terapia , Animais , Antineoplásicos/uso terapêutico , Azurina/genética , Azurina/uso terapêutico , Linhagem Celular Tumoral , Modelos Animais de Doenças , Escherichia coli/classificação , Escherichia coli/genética , Escherichia coli/metabolismo , Feminino , Engenharia Genética , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Experimentais/metabolismo , Neoplasias Mamárias Experimentais/microbiologia , Melanoma Experimental/metabolismo , Melanoma Experimental/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , PlasmídeosRESUMO
Corynebacterium parvum (CP), with their potent anti-tumor activities, has been well documented. Non-cell Corynebacterium Parvum (NCPP) is a neotype of biological preparation, which based on manipulating CP with nanotechnology. The present study was designed to investigate the effect of NCPP/CP on bone marrow derived dendritic cells (BMDCs) in tumor-bearing mice, especially focused on the differentiation and maturation of these BMDCs. BM cells from tumor-bearing mice administrated with NCPP/CP were analyzed by flow cytometry, which exhibit enhanced numbers of DCs and macrophages. In the meanwhile, flow cytometry analysis showed mild but significant difference for CD80 expression on these LPS- treated BMDCs between NCPP/CP administrated mice and the control animals. Furthermore, antigen presenting assay for these LPS-treated BMDCs showed significant difference for cytolytic assay of CD8+T cells against B16 melanoma cells, which indicate that NCPP treatments have enhanced the cytolytic rates of CD8+T cells from 47.9%±2.3% to 54.2%±2.4%. The data suggest that NCPP/CP treatment can efficiently facilitate the generation of BMDCs in vivo and enhance the maturation of these BMDCs in vitro.
Assuntos
Antígenos de Bactérias/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Dendríticas/imunologia , Melanoma Experimental/imunologia , Propionibacterium acnes/imunologia , Animais , Apresentação de Antígeno/efeitos dos fármacos , Antígenos de Bactérias/administração & dosagem , Antígeno B7-1/metabolismo , Células da Medula Óssea/imunologia , Diferenciação Celular/efeitos dos fármacos , Separação Celular , Células Cultivadas , Citotoxicidade Imunológica/efeitos dos fármacos , Células Dendríticas/efeitos dos fármacos , Citometria de Fluxo , Masculino , Melanoma Experimental/microbiologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
OBJECTIVE: To study the effection of suppression murine melanoma growth by Intratumor injection of recombinant attenuated salmonella carrying heat shock protein 70 and herpes simplex virus thymidine kinase genes. METHODS: Plasmids PCMV-mtHSP70-IRES-TK were electro-transferred into salmonella typhimurium SL7207 to construct recombinant salmonella typhimurium. In vivo, Recombinant bacteria were injected into the mouse melanoma and the antitumor effection was observed. The survival period was recorded and safety analysis for this vaccine in each group. RESULTS: In vivo, the mtHSP70/HSV-tk recombinant bacteria can suppress tumor growth significantly and extend survival. After recombinant Salmonella, 10(9) CFU/mL, was administered as an intratumoral injection, No diarrhea were observed. During therapy, body weight did not change markedly. CONCLUSION: Results of the animal experiment suggests intratumor injection of recombinant attenuated salmonella typhimurium containing mtHSP70 and HSV-tk genes, has targeting ability against B16 tumor cell and could significantly inhibit tumor growth .
Assuntos
Vacinas Anticâncer/farmacologia , Proteínas de Choque Térmico HSP70/imunologia , Melanoma Experimental/terapia , Simplexvirus/enzimologia , Timidina Quinase/imunologia , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Vacinas Anticâncer/genética , Vacinas Anticâncer/imunologia , Terapia Genética/métodos , Proteínas de Choque Térmico HSP70/genética , Melanoma Experimental/microbiologia , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis/genética , Salmonella typhimurium/genética , Salmonella typhimurium/imunologia , Simplexvirus/genética , Neoplasias Cutâneas/terapia , Timidina Quinase/genética , Vacinas Atenuadas/genética , Vacinas Atenuadas/imunologia , Vacinas Atenuadas/farmacologia , Vacinas de DNA/genética , Vacinas de DNA/imunologia , Vacinas de DNA/farmacologiaRESUMO
Attenuated Salmonella can invade tumor cells and acts as a eukaryotic expression vector for gene propagation. We constructed a bi-gene, eukaryotic co-expression DNA vaccine of Mycobacterium tuberculosis heat shock protein 70 (mtHSP70) and Herpes simplex virus-thymidine kinase (HSV-tk) and used attenuated Salmonella as a vector to treat murine melanoma. In vitro, recombinant Salmonella can carry plasmid stably and can invade into the cytoplasm of B16 tumor cells expressing the protein of the mtHSP70/HSV-tk gene by Western blot assay. In vivo, after the recombinant Salmonella was injected into tumors, the HSV-tk precursor drug ganciclovir (GCV) was administered to start the HSV-tk killing of tumor cells. We found that the mtHSP70/HSV-tk recombinant bacteria can raise CD8+ T lymphocytes in peripheral blood by flow cytometry and in tumor tissues by immunofluorescence detection, increase IFNγ contents in tumor tissue by ELISA and significantly suppress tumor growth.