RESUMO
Stimulator of IFN genes (STING) is a critical component of the innate immune system, playing an essential role in defending against DNA virus infections. However, the mechanisms governing basal STING regulation remain poorly understood. In this study, we demonstrate that the basal level of STING is critically maintained by hypoxia-inducible factor 1 (HIF-1)α through transcription. Under normal conditions, HIF-1α binds constitutively to the promoter region of STING, actively promoting its transcription. Knocking down HIF-1α results in a decrease in STING expression in multiple cell lines and zebrafish, which in turn reduces cellular responses to synthetic dsDNAs, including cell signaling and IFN production. Moreover, this decrease in STING levels leads to an increase in cellular susceptibility to DNA viruses HSV-1 and pseudorabies virus. These findings unveil a (to our knowledge) novel role of HIF-1α in maintaining basal STING levels and provide valuable insights into STING-mediated antiviral activities and associated diseases.
Assuntos
Herpesvirus Humano 1 , Subunidade alfa do Fator 1 Induzível por Hipóxia , Imunidade Inata , Proteínas de Membrana , Peixe-Zebra , Animais , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Peixe-Zebra/imunologia , Herpesvirus Humano 1/imunologia , Herpesvirus Suídeo 1/imunologia , Imunidade Celular , Regulação da Expressão Gênica/imunologia , Transdução de Sinais/imunologia , Transcrição Gênica/imunologia , Regiões Promotoras Genéticas , Células HEK293 , Linhagem Celular , Herpes Simples/imunologia , Pseudorraiva/imunologiaRESUMO
G-protein coupled receptor (GPCR) kinases (GRKs) and hypoxia-inducible factor-1α (HIF-1α) play key roles in rheumatoid arthritis (RA). Several studies have demonstrated that HIF-1α expression is positively regulated by GRK2, suggesting its posttranscriptional effects on HIF-1α. In this study, we review the role of HIF-1α and GRK2 in RA pathophysiology, focusing on their proinflammatory roles in immune cells and fibroblast-like synoviocytes (FLS).We then introduce several drugs that inhibit GRK2 and HIF-1α, and briefly outline their molecular mechanisms. We conclude by presenting gaps in knowledge and our prospects for the pharmacological potential of targeting these proteins and the relevant downstream signaling pathways.Future research is warranted and paramount for untangling these novel and promising roles for GRK2 and HIF-1α in RA.
Assuntos
Artrite Reumatoide , Quinase 2 de Receptor Acoplado a Proteína G , Subunidade alfa do Fator 1 Induzível por Hipóxia , Sinoviócitos , Humanos , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/genética , Artrite Reumatoide/imunologia , Hipóxia/genética , Hipóxia/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Inflamação/tratamento farmacológico , Inflamação/genética , Inflamação/imunologia , Sinoviócitos/imunologia , Quinase 2 de Receptor Acoplado a Proteína G/antagonistas & inibidores , Quinase 2 de Receptor Acoplado a Proteína G/genética , Quinase 2 de Receptor Acoplado a Proteína G/imunologiaRESUMO
The hypoxia-inducible factors (HIF)-1α and HIF-2α are central regulators of transcriptional programmes in settings such as development and tumour expansion. HIF-2α moonlights as a cap-dependent translation factor. We provide new insights into how the interferon-stimulated gene 15 (ISG15), a ubiquitin-like modifier, and the HIFs regulate one another in hypoxia and interferon-induced cells. We show that upon ISGylation induction and HIF-α stabilization, both HIFs promote protein ISGylates through transcriptional and/or post-transcriptional pathways. We show the first evidence of HIF-2α modification by ISG15. ISGylation induces system-level alterations to the HIF transcriptional programme and increases the cytoplasmic/nuclear fraction and translation activity of HIF-2α. This work identifies ISG15 as a regulator of hypoxic mRNA translation, which has implications for immune processes and disease progression.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos , Hipóxia , Polirribossomos , Humanos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/imunologia , Hipóxia Celular/genética , Hipóxia Celular/imunologia , Hipóxia/genética , Hipóxia/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Interferons/genética , Interferons/imunologia , Polirribossomos/genética , Polirribossomos/imunologiaRESUMO
The hypoxia-inducible factors (HIFs) regulate the main transcriptional pathway of response to hypoxia in T cells and are negatively regulated by von Hippel-Lindau factor (VHL). But the role of HIFs in the regulation of CD4 T cell responses during infection with M. tuberculosis isn't well understood. Here we show that mice lacking VHL in T cells (Vhl cKO) are highly susceptible to infection with M. tuberculosis, which is associated with a low accumulation of mycobacteria-specific T cells in the lungs that display reduced proliferation, altered differentiation and enhanced expression of inhibitory receptors. In contrast, HIF-1 deficiency in T cells is redundant for M. tuberculosis control. Vhl cKO mice also show reduced responses to vaccination. Further, VHL promotes proper MYC-activation, cell-growth responses, DNA synthesis, proliferation and survival of CD4 T cells after TCR activation. The VHL-deficient T cell responses are rescued by the loss of HIF-1α, indicating that the increased susceptibility to M. tuberculosis infection and the impaired responses of Vhl-deficient T cells are HIF-1-dependent.
Assuntos
Subunidade alfa do Fator 1 Induzível por Hipóxia , Tuberculose , Proteína Supressora de Tumor Von Hippel-Lindau , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Hipóxia , Fator 1 Induzível por Hipóxia , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Camundongos , Linfócitos T/imunologia , Tuberculose/genética , Tuberculose/imunologia , Tuberculose/prevenção & controle , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Proteína Supressora de Tumor Von Hippel-Lindau/imunologiaRESUMO
The morbidity and mortality of malaria are still high. Programmed cell death-1(PD-1) is an important co-inhibitory factor and CD8 T cells with PD-1 were reported to be exhausted cells. It remains unknown what the role of CD4 T cells expressing PD-1 is and what the upstream regulating molecules of PD-1 in CD4 T cells are. The C57BL/6 mice were injected with Plasmodium yoelii (P. yoelii) in this study. Expressions of PD-1, activation markers, and cytokines were tested. The differentially expressed genes between PD-1+/- CD4 T cells were detected by microarray sequencing. Western blot, chromatin immunoprecipitation (ChIP), siRNA, hypoxia inducible factor-1α (HIF-1α) inducer and inhibitor were used to explore PD-1's upstream molecules, respectively. The proportions of PD-1+ CD4 T cells increased post P. yoelii infection. PD-1+ CD4 T cells expressed more activated surface markers and could produce more cytokines. Nuclear factor of activated T cells 1 (NFATc1) was found to be a key transcription factor to induce PD-1 expression after infection. Both the inducer and the inhibitor of HIF-1α could change the expressions of NFATc1 and PD-1 in vivo and in vitro, respectively. Taken together, P. yoelii infection induced NFATc1 expression by HIF-1α. The highly expressed NFATc1 entered the nucleus and initiated PD-1 expression. PD-1+ CD4 T cells appeared to be more activated and could secrete more cytokines to regulate the host's immune responses against malaria.
Assuntos
Linfócitos T CD4-Positivos , Subunidade alfa do Fator 1 Induzível por Hipóxia , Malária , Fatores de Transcrição NFATC , Plasmodium yoelii , Receptor de Morte Celular Programada 1 , Animais , Linfócitos T CD4-Positivos/imunologia , Citocinas/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Malária/genética , Malária/imunologia , Malária/parasitologia , Camundongos , Camundongos Endogâmicos C57BL , Fatores de Transcrição NFATC/genética , Fatores de Transcrição NFATC/imunologia , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/imunologia , Transdução de SinaisRESUMO
Patients with periodontitis undergoing orthodontic therapy may suffer from undesired dental root resorption. The purpose of this in vitro study was to investigate the molecular mechanisms resulting in PD-L1 expression of cementoblasts in response to infection with Porphyromonas gingivalis (P. gingivalis) peptidoglycan (PGN) and compressive force (CF), and its interaction with hypoxia-inducible factor (HIF)-1α molecule: The cementoblast (OCCM-30) cells were kinetically infected with various concentrations of P. gingivalis PGN in the presence and absence of CF. Western blotting and RT-qPCR were performed to examine the protein expression of PD-L1 and HIF-1α as well as their gene expression. Immunofluorescence was applied to visualize the localization of these proteins within cells. An HIF-1α inhibitor was added for further investigation of necroptosis by flow cytometry analysis. Releases of soluble GAS-6 were measured by ELISA. P. gingivalis PGN dose dependently stimulated PD-L1 upregulation in cementoblasts at protein and mRNA levels. CF combined with P. gingivalis PGN had synergistic effects on the induction of PD-L1. Blockade of HIF-1α inhibited the P. gingivalis PGN-inducible PD-L1 protein expression under compression, indicating an HIF-1α dependent regulation of PD-L1 induction. Concomitantly, an HIF-1α inhibitor decreased the GAS-6 release in the presence of CF and P. gingivalis PGN co-stimulation. The data suggest that PGN of P. gingivalis participates in PD-L1 up-regulation in cementoblasts. Additionally, the influence of compressive force on P. gingivalis PGN-induced PD-L1 expression occurs in HIF-1α dependently. In this regard, HIF-1α may play roles in the immune response of cementoblasts via immune-inhibitory PD-L1. Our results underline the importance of molecular mechanisms involved in bacteria-induced periodontics and root resorption.
Assuntos
Antígeno B7-H1 , Reabsorção da Raiz , Antígeno B7-H1/biossíntese , Antígeno B7-H1/genética , Antígeno B7-H1/imunologia , Cemento Dentário/imunologia , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Peptidoglicano/imunologia , Porphyromonas gingivalis/metabolismo , Reabsorção da Raiz/genética , Reabsorção da Raiz/imunologiaRESUMO
BACKGROUND: Distant metastasis to vital organs is the major contributor to breast cancer mortality, and regional lymph node metastasis is an important facilitator of distant metastasis and recurrence in this cancer. The early diagnosis and precise treatment of lymph node metastasis are crucial for staging and prognosis in breast cancer. Herein, we report a visualized precision medicine nanoplatform of metastatic lymph nodes for ultrasonic/photoacoustic (US/PA) dual modal imaging-guided in situ targeted hyperthermia-combined chemotherapy. RESULTS: Carbon nanoparticles (CNs), approved by the China Food and Drug Administration, were loaded with docetaxel and rationally combined with anti-hypoxia-inducible factor 1α antibody-modified poly (lactic-co-glycolic acid) (PLGA) nanoparticles to achieve the combination of passive targeting at the lymph nodes and intracellular targeting at HIF 1α factor. The accumulation and retention of nanoparticles in metastatic lymph nodes via lymphatic delivery were enhanced. Docetaxel could be effectively offloaded by CNs that have active carbon nanoparticles, and the PLGA membrane prevented drug leakage. The nanoparticles exhibited excellent photothermal performance with a photothermal conversion efficiency of 28.9%, killing tumor cells in metastatic lymph nodes through hyperthermia. In vitro and in vivo systematic evaluations revealed that hyperpyrexia triggered the rupture of nanoparticles caused by the phase transition of perfluorohexane, resulting in docetaxel release for achieving in situ hyperthermia-combined chemotherapy. CONCLUSIONS: The laser-triggered highly efficient in situ chemotherapy nanosystem achieves targeted synergistic chemo-hyperthermia treatment of metastatic lymph nodes, and lymphatic delivery represents a strategy to avoid additional injury caused by drugs entering the blood circulation.
Assuntos
Antineoplásicos/uso terapêutico , Hipertermia Induzida/métodos , Linfonodos/metabolismo , Nanopartículas/química , Neoplasias/tratamento farmacológico , Animais , Anticorpos/química , Anticorpos/imunologia , Antineoplásicos/química , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Carbono/química , Linhagem Celular Tumoral , Docetaxel/química , Docetaxel/metabolismo , Docetaxel/farmacologia , Docetaxel/uso terapêutico , Feminino , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Raios Infravermelhos , Metástase Linfática , Nanomedicina , Nanopartículas/metabolismo , Neoplasias/patologia , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Ratos , Transplante HeterólogoRESUMO
T helper 17 (TH17) cells are involved in several autoimmune diseases such as multiple sclerosis (MS) and rheumatoid arthritis (RA). In addition to retinoic acid receptor-related orphan nuclear receptor gamma t (ROR-γt), hypoxia-inducible factor-1α (HIF-1α) is essential for the differentiation and inflammatory function of TH17 cells. To investigate the roles of HIF-1α in the functional regulation of TH17 cells under the normal physiological condition without genetic modification, the nucleus-transducible form of transcription modulation domain (TMD) of HIF-1α (ntHIF-1α-TMD) was generated by conjugating HIF-1α-TMD to Hph-1 protein transduction domain (PTD). ntHIF-1α-TMD was effectively delivered into the nucleus of T cells without cellular cytotoxicity. ntHIF-1α-TMD significantly blocked the differentiation of naïve T cells into TH17 cells in a dose-dependent manner via IL-17A and ROR-γt expression inhibition. However, T-cell activation events such as induction of CD69, CD25, and IL-2 and the differentiation potential of naïve T cells into TH1, TH2, or Treg cells were not affected by ntHIF-1α-TMD. Interestingly, TH17 cells differentiated from naïve T cells in the presence of ntHIF-1α-TMD showed a substantial level of suppressive activity toward the activated T cells, and the increase of Foxp3 and IL-10 expression was detected in these TH17 cells. When mRNA expression pattern was compared between TH17 cells and ntHIF-1α-TMD-treated TH17 cells, the expression of the genes involved in the differentiation and functions of TH17 cells was downregulated, and that of the genes necessary for immune-suppressive functions of Treg cells was upregulated. When the mice with experimental autoimmune encephalomyelitis (EAE) were treated with ntHIF-1α-TMD with anti-IL-17A mAb as a positive control, the therapeutic efficacy of ntHIF-1α-TMD in vivo was comparable with that of anti-IL-17A mAb, and ntHIF-1α-TMD-mediated therapeutic effect was contributed by the functional conversion of TH17 cells into immune-suppressive T cells. The results in this study demonstrate that ntHIF-1α-TMD can be a new therapeutic reagent for the treatment of various autoimmune diseases in which TH17 cells are dominant and pathogenic T cells.
Assuntos
Encefalomielite Autoimune Experimental/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Células Th17/imunologia , Animais , Diferenciação Celular/imunologia , Feminino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Uncontrolled growth of solid tumors will result in a hallmark hypoxic condition, whereby the key transcriptional regulator of hypoxia inducible factor-1α (HIF-1α) will be stabilized to activate the transcription of target genes that are responsible for the metabolism, proliferation, and metastasis of tumor cells. Targeting and inhibiting the transcriptional activity of HIF-1 may provide an interesting strategy for cancer therapy. In the present study, an immune library and a synthetic library were constructed for the phage display selection of Nbs against recombinant PAS B domain protein (rPasB) of HIF-1α. After panning and screening, seven different nanobodies (Nbs) were selected, of which five were confirmed via immunoprecipitation to target the native HIF-1α subunit. The inhibitory effect of the selected Nbs on HIF-1 induced activation of target genes has been evaluated after intracellular expression of these Nbs in HeLa cells. The dramatic inhibition of both intrabody formats on the expression of HIF-1-related target genes has been confirmed, which indicated the inhibitory efficacy of selected Nbs on the transcriptional activity of HIF-1.
Assuntos
Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Anticorpos de Domínio Único/farmacologia , Transcrição Gênica/efeitos dos fármacos , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/metabolismo , Hipóxia Celular/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Células HeLa , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Domínios Proteicos/genética , Domínios Proteicos/imunologia , Anticorpos de Domínio Único/genética , Anticorpos de Domínio Único/imunologia , Anticorpos de Domínio Único/metabolismo , Transfecção , Neoplasias do Colo do Útero/patologiaRESUMO
Hypoxia-inducible factor-1α (HIF-1α) is an important regulator of glucose metabolism and inflammatory cytokine production in innate immune responses. Viruses modulate HIF-1α to support viral replication and the survival of infected cells, but it is unclear if this transcription factor also plays an important role in regulating antiviral immune responses. In this study, we found that short and long dsRNA differentially engage TLR3, inducing distinct levels of proinflammatory cytokine production (TNF-α and IL-6) in bone marrow-derived macrophages from C57BL/6 mice. These responses are associated with differential accumulation of HIF-1α, which augments NF-κB activation. Unlike TLR4 responses, increased HIF-1α following TLR3 engagement is not associated with significant alterations in glycolytic activity and was more pronounced in low glucose conditions. We also show that the mechanisms supporting HIF-1α stabilization may differ following stimulation with short versus long dsRNA and that pyruvate kinase M2 and mitochondrial reactive oxygen species play a central role in these processes. Collectively, this work suggests that HIF-1α may fine-tune proinflammatory cytokine production during early antiviral immune responses, particularly when there is limited glucose availability or under other conditions of stress. Our findings also suggest we may be able to regulate the magnitude of proinflammatory cytokine production during antiviral responses by targeting proteins or molecules that contribute to HIF-1α stabilization.
Assuntos
Citocinas/biossíntese , Glucose/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Macrófagos/imunologia , Ácidos Nucleicos/imunologia , Receptor 3 Toll-Like/imunologia , Animais , Células Cultivadas , Camundongos , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio/imunologiaRESUMO
TNF is a central cytokine in the pathogenesis of rheumatoid arthritis (RA). Elevated level of TNF causes local inflammation that affects immune cells and fibroblast-like synoviocytes (FLS). Nowadays, only 20-30% of patients experience remission after the standard of care therapy-antibodies against TNF. Interestingly, responders show reduced levels of GLUT1 and GAPDH, highlighting a potential link to cellular metabolism. The aim of the study was to investigate whether TNF directly affects the metabolic phenotype of FLS. Real-time respirometry displayed TNF-induced upregulation of glycolysis along with a modest increase of oxidative phosphorylation in FLS from healthy donors. In addition, TNF stimulation enhanced HIF1A and GLUT1 expression. The upregulation of HIF1A and GLUT1 reflects their enriched level in FLS from RA patients (RA-FLS). The inhibition of TAK1, HIF1a and hexokinase deciphered the importance of TNF/TAK1/HIF1A/glycolysis signaling axis. To prove that inhibition of glycolysis reduced the pathogenic phenotype, we showed that 2-deoxyglucose, a hexokinase inhibitor, partially decreased secretion of RA biomarkers. In summary, we identified a direct role of TNF on glycolytic reprogramming of FLS and confirmed the potency of immunometabolism for RA. Further studies are needed to evaluate the therapeutic impact especially regarding non-responder data.
Assuntos
Artrite Reumatoide/imunologia , Transportador de Glucose Tipo 1/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Sinoviócitos/imunologia , Fator de Necrose Tumoral alfa/imunologia , Células Cultivadas , Humanos , Sinoviócitos/citologiaAssuntos
Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 1/patologia , Modelos Animais de Doenças , Coração/fisiopatologia , Pâncreas/patologia , Animais , Glicemia/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Feminino , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/imunologia , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Insulina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pâncreas/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/imunologia , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Receptor Muscarínico M2/imunologia , Receptor Muscarínico M2/metabolismo , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Hypoxia is an important phenomenon in solid tumors that contributes to metastasis, tumor microenvironment (TME) deregulation, and resistance to therapies. The receptor tyrosine kinase AXL is an HIF target, but its roles during hypoxic stress leading to the TME deregulation are not well defined. We report here that the mammary gland-specific deletion of Axl in a HER2+ mouse model of breast cancer leads to a normalization of the blood vessels, a proinflammatory TME, and a reduction of lung metastases by dampening the hypoxic response in tumor cells. During hypoxia, interfering with AXL reduces HIF-1α levels altering the hypoxic response leading to a reduction of hypoxia-induced epithelial-to-mesenchymal transition (EMT), invasion, and production of key cytokines for macrophages behaviors. These observations suggest that inhibition of Axl generates a suitable setting to increase immunotherapy. Accordingly, combining pharmacological inhibition of Axl with anti-PD-1 in a preclinical model of HER2+ breast cancer reduces the primary tumor and metastatic burdens, suggesting a potential therapeutic approach to manage HER2+ patients whose tumors present high hypoxic features.
Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Imunoterapia , Proteínas Proto-Oncogênicas/genética , Receptores Proteína Tirosina Quinases/genética , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/fisiopatologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Feminino , Deleção de Genes , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Marcação de Genes , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Inibidores de Checkpoint Imunológico/administração & dosagem , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Metástase Neoplásica/tratamento farmacológico , Metástase Neoplásica/genética , Metástase Neoplásica/imunologia , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/imunologia , Proteínas Proto-Oncogênicas/imunologia , Receptores Proteína Tirosina Quinases/imunologia , Microambiente Tumoral/efeitos dos fármacos , Receptor Tirosina Quinase AxlRESUMO
Patients with severe COVID-19 infection exhibit a low level of oxygen in affected tissue and blood. To understand the pathophysiology of COVID-19 infection, it is therefore necessary to understand cell function during hypoxia. We investigated aspects of human monocyte activation under hypoxic conditions. HMGB1 is an alarmin released by stressed cells. Under normoxic conditions, HMGB1 activates interferon regulatory factor (IRF)5 and nuclear factor-κB in monocytes, leading to expression of type I interferon (IFN) and inflammatory cytokines including tumor necrosis factor α, and interleukin 1ß, respectively. When hypoxic monocytes are activated by HMGB1, they produce proinflammatory cytokines but fail to produce type I IFN. Hypoxia-inducible factor-1α, induced by hypoxia, functions as a direct transcriptional repressor of IRF5 and IRF3. As hypoxia is a stressor that induces secretion of HMGB1 by epithelial cells, hypoxia establishes a microenvironment that favors monocyte production of inflammatory cytokines but not IFN. These findings have implications for the pathogenesis of COVID-19.
Assuntos
Hipóxia Celular/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Monócitos/imunologia , COVID-19/imunologia , Células Cultivadas , Citocinas/imunologia , Humanos , Fatores Reguladores de Interferon/metabolismo , Interferon Tipo I/imunologia , Interferon Tipo I/metabolismo , Interleucina-1beta/metabolismo , Monócitos/metabolismo , NF-kappa B/imunologia , NF-kappa B/metabolismo , Oxigênio/metabolismo , SARS-CoV-2/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Adoptive T cell therapies (ACTs) hold great promise in cancer treatment, but low overall response rates in patients with solid tumors underscore remaining challenges in realizing the potential of this cellular immunotherapy approach. Promoting CD8+ T cell adaptation to tissue residency represents an underutilized but promising strategy to improve tumor-infiltrating lymphocyte (TIL) function. Here, we report that deletion of the HIF negative regulator von Hippel-Lindau (VHL) in CD8+ T cells induced HIF-1α/HIF-2α-dependent differentiation of tissue-resident memory-like (Trm-like) TILs in mouse models of malignancy. VHL-deficient TILs accumulated in tumors and exhibited a core Trm signature despite an exhaustion-associated phenotype, which led to retained polyfunctionality and response to αPD-1 immunotherapy, resulting in tumor eradication and protective tissue-resident memory. VHL deficiency similarly facilitated enhanced accumulation of chimeric antigen receptor (CAR) T cells with a Trm-like phenotype in tumors. Thus, HIF activity in CD8+ TILs promotes accumulation and antitumor activity, providing a new strategy to enhance the efficacy of ACTs.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/imunologia , Linfócitos T CD8-Positivos/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Imunidade Celular , Memória Imunológica , Linfócitos do Interstício Tumoral/imunologia , Neoplasias Experimentais/imunologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Linfócitos T CD8-Positivos/patologia , Linhagem Celular Tumoral , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Linfócitos do Interstício Tumoral/patologia , Camundongos , Camundongos Knockout , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Proteína Supressora de Tumor Von Hippel-Lindau/imunologiaRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Psoriasis is a chronic inflammatory skin disease mediated by immunity. Our pre-clinical studies have proved that QZLX mixture can improve patients' clinical symptoms with psoriasis without noticeable adverse reactions. In a psoriasis-like mouse model induced by imiquimod, QZLX mixture has been shown to alleviate epidermal inflammation and inhibit the hyperproliferation of keratinocytes. However, its related molecular mechanism remains to be elucidated. AIM OF THE STUDY: To assess the mechanism of QZLX mixture against psoriasis. MATERIALS AND METHODS: This study combines network pharmacology and experiments to study the mechanism of QZLX against psoriasis. First, construct the active compound-target network and PPI network. Secondly, determine possible drug targets through Molecular docking and KEGG. Thirdly, high-performance liquid chromatography (HPLC) was used for the quality control of QZLX. Finally, use a mouse model of psoriasis to further confirm the role of QZLX. RESULTS: (1) Network pharmacology analysis shows that QZLX alleviates psoriasis's epidermal inflammation, and neovascularization may be achieved by inhibiting the IL6/STAT3 signaling pathway. (2) QZLX improves the pathological characteristics of IMQ-induced skin damage in psoriasis-like mice. (3) QZLX inhibits the IL6/STAT3 signaling pathway and reduces the expression of IL-17, IL-23, and TNF-α related to inflammation in peripheral blood, as well as the expression of S100A7 in the lesion area. QZLX is better than MTX in inhibiting neovascularization by down-regulating the expression of HIF-1 and CD31 in the lesion area. Finally, inhibition of Ki67 alleviates the excessive proliferation of keratinocytes. CONCLUSION: In sum, this study clarifies the mechanism of QZLX against psoriasis and provides evidence to support its clinical use.
Assuntos
Anti-Inflamatórios/uso terapêutico , Medicamentos de Ervas Chinesas/uso terapêutico , Psoríase/tratamento farmacológico , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Proliferação de Células/efeitos dos fármacos , Citocinas/genética , Citocinas/imunologia , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/farmacologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Imiquimode , Queratinócitos/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Simulação de Acoplamento Molecular , Compostos Fitoquímicos/análise , Compostos Fitoquímicos/farmacologia , Compostos Fitoquímicos/uso terapêutico , Mapas de Interação de Proteínas , Psoríase/induzido quimicamente , Psoríase/imunologia , Psoríase/patologia , Fator de Transcrição STAT3/imunologia , Pele/efeitos dos fármacos , Pele/imunologia , Pele/patologiaRESUMO
B cells play a major role in regulating disease incidence through various factors, including spleen tyrosine kinase (Syk), which transmits signals to all hematopoietic lineage cells. Hypoxia-inducible factor (HIF)-1α accumulates under hypoxic conditions, which is also oxidative stress to induce nuclear factor (erythroid-derived 2)-like 2 (Nrf2) responsible for gene expression of antioxidant enzymes. In the present study, we investigated whether B cells are regulated by crosstalk of HIF-1α and Nrf2 via reactive oxygen species (ROS)-mediated Syk activation. When B cells were incubated under hypoxic conditions, Syk phosphorylation, HIF-1α, and Nrf2 levels were increased. Hypoxia-inducible results were consistent with CoCl2 treatment, which mimics hypoxic conditions. Cell viability was reduced by the Syk inhibitor BAY 61-3606. Increased Nrf2 levels due to hypoxia or CoCl2 were inhibited by treatment with a HIF inhibitor. Hypoxia- or CoCl2-induced ROS increased HIF-1α and Nrf2 levels, which were attenuated by treatment with N-acetyl-L-cysteine (NAC), a ROS scavenger. HIF-1α levels were reduced in doxycycline-treated shNrf2 cells. Clobetasol propionate, a Nrf2 inhibitor, also inhibited HIF-1α levels induced by hypoxia or CoCl2. ROS-mediated Syk phosphorylation at tyrosine 525/526 was confirmed by treatment with H2O2, hypoxia, and CoCl2, and attenuated with NAC treatment. Inhibition of Syk phosphorylation by BAY 61-3606 is consistent with a decrease in protein HIF-1α and Nrf2 levels. Taken together, HIF-1α levels might control Nrf2 levels and vice versa, and could be associated with Syk phosphorylation in B cells. The results indicate that B cells could be regulated by crosstalk of HIF-1α and Nrf2 through ROS-mediated Syk activation.
Assuntos
Linfócitos B/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Fator 2 Relacionado a NF-E2/imunologia , Quinase Syk/imunologia , Animais , Hipóxia Celular/imunologia , Linhagem Celular , Sobrevivência Celular , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Masculino , Camundongos Endogâmicos C57BL , RNA Interferente Pequeno/genética , Espécies Reativas de Oxigênio/imunologiaRESUMO
Adoptive transfer of antitumor cytotoxic T cells is an emerging form of cancer immunotherapy. A key challenge to expanding the utility of adoptive cell therapies is how to enhance the survival and function of the transferred T cells. Immune-cell survival requires adaptation to different microenvironments and particularly to the hypoxic milieu of solid tumors. The hypoxia-inducible factor (HIF) transcription factors are an essential aspect of this adaptation. In this study, we undertook experiments to define structural determinants of HIF that potentiate antitumor efficacy in cytotoxic T cells. We first created retroviral vectors to deliver ectopic expression of HIF1α and HIF2α in mouse CD8+ T cells, together or individually and with or without sensitivity to the oxygen-dependent HIFα inhibitors Von Hippel-Lindau and factor-inhibiting HIF (FIH). HIF2α, but not HIF1α, drove broad transcriptional changes in CD8+ T cells, resulting in increased cytotoxic differentiation and cytolytic function against tumor targets. A specific mutation replacing the hydroxyl group-acceptor site for FIH in HIF2α gave rise to the most effective antitumor T cells after adoptive transfer in vivo In addition, codelivering an FIH-insensitive form of HIF2α with an anti-CD19 chimeric antigen receptor greatly enhanced cytolytic function of human CD8+ T cells against lymphoma cells both in vitro and in a xenograft adoptive transfer model. These experiments point to a means to increase the antitumor efficacy of therapeutic CD8+ T cells via ectopic expression of the HIF transcription factor.See related Spotlight on p. 364.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/imunologia , Linfócitos T CD8-Positivos/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Hipóxia/imunologia , Imunoterapia Adotiva , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/antagonistas & inibidores , Linhagem Celular Tumoral , Feminino , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fatores de Transcrição , Microambiente TumoralRESUMO
Signaling via TNF-R1 mediates pleiotropic biological outcomes ranging from inflammation and proliferation to cell death. Previous reports demonstrated that pro-survival signaling emanates from membrane resident TNF-R1 complexes (complex I) while only internalized TNF-R1 complexes are capable for DISC formation (complex II) and thus, apoptosis induction. Internalized TNF-R1 containing endosomes undergo intracellular maturation towards lysosomes, resulting in activation and release of Cathepsin D (CtsD) into the cytoplasm. We recently revealed HSP90 as target for proteolytic cleavage by CtsD, resulting in cell death amplification. In this study, we show that extrinsic cell death activation via TNF or TRAIL results in HSP90ß degradation. Co-incubation of cells with either TNF or TRAIL in combination with the HSP90ß inhibitor KUNB105 but not HSP90α selective inhibition promotes apoptosis induction. In an attempt to reveal further downstream targets of combined TNF-R1 or TRAIL-R1/-R2 activation with HSP90ß inhibition, we identify HIF1α and validate its ligand:inhibitor triggered degradation. Together, these findings suggest that selective inhibition of HSP90 isoforms together with death ligand stimulation may provide novel strategies for therapy of inflammatory diseases or cancer, in future.
Assuntos
Proteínas de Choque Térmico HSP90/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Ligante Indutor de Apoptose Relacionado a TNF/imunologia , Fator de Necrose Tumoral alfa/imunologia , Apoptose , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Células HeLa , Humanos , Proteoma , Células U937RESUMO
OBJECTIVE: To investigate the effects of hypoxia and Porphyromonas gingivalis- lipopolysaccharide (P. gingivalis-LPS) on activation of the NACHT leucine-rich repeat protein 3 (NLRP3) inflammasome in human gingival fibroblasts (HGFs). DESIGN: Periodontitis was optimally simulated using a hypoxic concentration of 1%. HGFs were stimulated using P. gingivalis-LPS (1.0 µg/ml) in normoxia and hypoxia for 3 h and 6 h, respectively. The expression levels of genes and proteins of hypoxia-inducible factor-1α (HIF-1α), interleukin-1ß, gasdermin D (GSDMD) and the NLRP3 inflammasome, including NLRP3, apoptosis-associated speck-like protein containing CARD (ASC), caspase-1 and its activated forms, were measured using quantitative real-time polymerase chain reaction and western blot. ELISA was used to detect and determine levels of the inflammatory factor interleukin-1ß in cell supernatants. Lactate dehydrogenase (LDH) release assay, caspase-1 activity assay and Hoechst 33342/Propidium Iodide (PI) staining were performed to further verify the presence of pyroptosis. RESULTS: The NLRP3 inflammasome (i.e., NLRP3, ASC, caspase-1) was not affected by individual stimulation using P. gingivalis-LPS or hypoxia. However, the combination of both hypoxia and P. gingivalis-LPS stimulation significantly enhanced inflammasome activation and promoted the expression of interleukin-1ß, gasdermin D and HIF-1α at gene and protein levels; PI positive cells and the release of LDH were also elevated. CONCLUSION: Hypoxia and P. gingivalis-LPS synergistically induced NLRP3 inflammasome activation in HGFs, and subsequently high levels of interleukin-1ß and GSDMD-mediated pyroptosis can cause an HGF inflammatory response, which plays an important role in the pathogenesis of periodontitis.