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1.
Sci Transl Med ; 13(601)2021 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-34233950

RESUMO

Inflammation is a well-known driver of lung tumorigenesis. One strategy by which tumor cells escape tight homeostatic control is by decreasing the expression of the potent anti-inflammatory protein tumor necrosis factor alpha-induced protein 3 (TNFAIP3), also known as A20. We observed that tumor cell intrinsic loss of A20 markedly enhanced lung tumorigenesis and was associated with reduced CD8+ T cell-mediated immune surveillance in patients with lung cancer and in mouse models. In mice, we observed that this effect was completely dependent on increased cellular sensitivity to interferon-γ (IFN-γ) signaling by aberrant activation of TANK-binding kinase 1 (TBK1) and increased downstream expression and activation of signal transducer and activator of transcription 1 (STAT1). Interrupting this autocrine feed forward loop by knocking out IFN-α/ß receptor completely restored infiltration of cytotoxic T cells and rescued loss of A20 depending tumorigenesis. Downstream of STAT1, programmed death ligand 1 (PD-L1) was highly expressed in A20 knockout lung tumors. Accordingly, immune checkpoint blockade (ICB) treatment was highly efficient in mice harboring A20-deficient lung tumors. Furthermore, an A20 loss-of-function gene expression signature positively correlated with survival of melanoma patients treated with anti-programmed cell death protein 1. Together, we have identified A20 as a master immune checkpoint regulating the TBK1-STAT1-PD-L1 axis that may be exploited to improve ICB therapy in patients with lung adenocarcinoma.


Assuntos
Adenocarcinoma de Pulmão , Neoplasias Pulmonares , Proteína 3 Induzida por Fator de Necrose Tumoral alfa/genética , Adenocarcinoma de Pulmão/genética , Animais , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Regulação para Baixo , Humanos , Interferon gama/metabolismo , Neoplasias Pulmonares/genética , Camundongos , Transdução de Sinais
2.
Molecules ; 26(11)2021 Jun 02.
Artigo em Inglês | MEDLINE | ID: mdl-34199417

RESUMO

Blockade of the programmed cell death 1 (PD-1)/programmed cell death-ligand 1 (PD-L1) interaction is currently the focus in the field of cancer immunotherapy, and so far, several monoclonal antibodies (mAbs) have achieved encouraging outcomes in cancer treatment. Despite this achievement, mAbs-based therapies are struggling with limitations including poor tissue and tumor penetration, long half-life time, poor oral bioavailability, and expensive production costs, which prompted a shift towards the development of the small-molecule inhibitors of PD-1/PD-L1 pathways. Even though many small-molecule inhibitors targeting PD-1/PD-L1 interaction have been reported, their development lags behind the corresponding mAb, partly due to the challenges of developing drug-like small molecules. Herein, we report the discovery of a series of novel inhibitors targeting PD-1/PD-L1 interaction via structural simplification strategy by using BMS-1058 as a starting point. Among them, compound A9 stands out as the most promising candidate with excellent PD-L1 inhibitory activity (IC50 = 0.93 nM, LE = 0.43) and high binding affinity to hPD-L1 (KD = 3.64 nM, LE = 0.40). Furthermore, A9 can significantly promote the production of IFN-γ in a dose-dependent manner by rescuing PD-L1 mediated T-cell inhibition in Hep3B/OS-8/hPD-L1 and CD3-positive T cells co-culture assay. Taken together, these results suggest that A9 is a promising inhibitor of PD-1/PD-L1 interaction and is worthy for further study.


Assuntos
Antígeno B7-H1/metabolismo , Receptor de Morte Celular Programada 1/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Linfócitos T/citologia , Antígeno B7-H1/química , Linhagem Celular , Cristalografia por Raios X , Humanos , Interferon gama/metabolismo , Modelos Moleculares , Simulação de Acoplamento Molecular , Estrutura Molecular , Cultura Primária de Células , Receptor de Morte Celular Programada 1/química , Ligação Proteica/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-Atividade , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo
3.
Int J Mol Sci ; 22(13)2021 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-34206740

RESUMO

N-Glycosylations are an important post-translational modification of proteins that can significantly impact cell function. Terminal sialic acid in hybrid or complex N-glycans has been shown to be relevant in various types of cancer, but its role in non-malignant cells remains poorly understood. We have previously shown that the motility of human bone marrow derived mesenchymal stromal cells (MSCs) can be modified by altering N-glycoforms. The goal of this study was to determine the role of sialylated N-glycans in MSCs. Here, we show that IFN-gamma or exposure to culture media low in fetal bovine serum (FBS) increases sialylated N-glycans, while PDGF-BB reduces them. These stimuli alter mRNA levels of sialyltransferases such as ST3Gal1, ST6Gal1, or ST3Gal4, suggesting that sialylation of N-glycans is regulated by transcriptional control of sialyltransferases. We next show that 2,4,7,8,9-pentaacetyl-3Fax-Neu5Ac-CO2Me (3F-Neu5Ac) effectively inhibits sialylations in MSCs. Supplementation with 3F-Neu5Ac increases adhesion and migration of MSCs, as assessed by both videomicroscopy and wound/scratch assays. Interestingly, pre-treatment with 3F-Neu5Ac also increases the survival of MSCs in an in vitro ischemia model. We also show that pre-treatment or continuous treatment with 3F-Neu5Ac inhibits both osteogenic and adipogenic differentiation of MSCs. Finally, secretion of key trophic factors by MSCs is variably affected upon exposure to 3F-Neu5Ac. Altogether, our experiments suggest that sialylation of N-glycans is tightly regulated in response to environmental cues and that glycoengineering MSCs to reduce sialylated N-glycans could be beneficial to increase both cell migration and survival, which may positively impact the therapeutic potential of the cells.


Assuntos
Movimento Celular , Células-Tronco Mesenquimais/metabolismo , Ácido N-Acetilneuramínico/metabolismo , Polissacarídeos/metabolismo , Sialiltransferases/metabolismo , Adipócitos/metabolismo , Diferenciação Celular , Sobrevivência Celular , Células Cultivadas , Inibidores Enzimáticos/farmacologia , Humanos , Interferon gama/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/fisiologia , Osteoblastos/citologia , Sialiltransferases/antagonistas & inibidores
4.
Int J Mol Sci ; 22(13)2021 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-34206968

RESUMO

The search for and analysis of new ligands for innate immunity receptors are of special significance for understanding the regulatory mechanisms of immune response. Here we show that the major heat shock protein 70 (Hsp70) can bind to and activate TREM-1, the innate immunity receptor expressed on monocytes. The Hsp70-TREM-1 interaction activates expression of TNFα and IFNγ mRNAs in monocytes and stimulates IL-2 secretion by PBMCs. Moreover, incubation of PBMCs with Hsp70 leads to an appearance of cytotoxic lymphocyte subpopulations active against the MHC-negative tumor cells. In addition, both the CD4+ T-lymphocytes and CD14+ monocytes are necessary for the Hsp70 signal transduction and a consequent activation of the cytotoxic lymphocytes. We believe that data presented in this study will broaden the views on the involvement of Hsp70 in the antitumor immunity.


Assuntos
Proteínas de Choque Térmico HSP70/metabolismo , Neoplasias/metabolismo , Receptor Gatilho 1 Expresso em Células Mieloides/metabolismo , Células HEK293 , Antígenos HLA/genética , Antígenos HLA/metabolismo , Células HeLa , Humanos , Interferon gama/metabolismo , Células K562 , Monócitos/metabolismo , Linfócitos T Citotóxicos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
5.
Int J Mol Sci ; 22(13)2021 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-34206987

RESUMO

Hepatitis C virus (HCV) is one of the main triggers of chronic liver disease. Despite tremendous progress in the HCV field, there is still no vaccine against this virus. Potential vaccines can be based on its recombinant proteins. To increase the humoral and, especially, cellular immune response to them, more effective adjuvants are needed. Here, we evaluated a panel of compounds as potential adjuvants using the HCV NS5B protein as an immunogen. These compounds included inhibitors of polyamine biosynthesis and urea cycle, the mTOR pathway, antioxidants, and cellular receptors. A pronounced stimulation of cell proliferation and interferon-γ (IFN-γ) secretion in response to concanavalin A was shown for antioxidant N-acetylcysteine (NAC), polyamine biosynthesis inhibitor 2-difluoromethylornithine (DFMO), and TLR9 agonist CpG ODN 1826 (CpG). Their usage during the immunization of mice with the recombinant NS5B protein significantly increased antibody titers, enhanced lymphocyte proliferation and IFN-γ production. NAC and CpG decreased relative Treg numbers; CpG increased the number of myeloid-derived suppressor cells (MDSCs), whereas neither NAC nor DFMO affected MDSC counts. NAC and DFMO suppressed NO and interleukin 10 (IL-10) production by splenocytes, while DFMO increased the levels of IL-12. This is the first evidence of immunomodulatory activity of NAC and DFMO during prophylactic immunization against infectious diseases.


Assuntos
Acetilcisteína/farmacologia , Adjuvantes Imunológicos/farmacologia , Eflornitina/farmacologia , Hepatite C/imunologia , Imunidade Ativa/efeitos dos fármacos , Proteínas não Estruturais Virais/imunologia , Animais , Proliferação de Células , Células Cultivadas , Feminino , Imunogenicidade da Vacina/efeitos dos fármacos , Interferon gama/metabolismo , Interleucina-10/metabolismo , Interleucina-12/metabolismo , Camundongos , Camundongos Endogâmicos DBA , Células Supressoras Mieloides/efeitos dos fármacos , Células Supressoras Mieloides/imunologia , Óxido Nítrico/metabolismo , Oligodesoxirribonucleotídeos/farmacologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Vacinas contra Hepatite Viral/imunologia
6.
Int J Mol Sci ; 22(10)2021 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-34063554

RESUMO

Acute lung injury (ALI) afflicts approximately 200,000 patients annually and has a 40% mortality rate. The COVID-19 pandemic has massively increased the rate of ALI incidence. The pathogenesis of ALI involves tissue damage from invading microbes and, in severe cases, the overexpression of inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß). This study aimed to develop a therapy to normalize the excess production of inflammatory cytokines and promote tissue repair in the lipopolysaccharide (LPS)-induced ALI. Based on our previous studies, we tested the insulin-like growth factor I (IGF-I) and BTP-2 therapies. IGF-I was selected, because we and others have shown that elevated inflammatory cytokines suppress the expression of growth hormone receptors in the liver, leading to a decrease in the circulating IGF-I. IGF-I is a growth factor that increases vascular protection, enhances tissue repair, and decreases pro-inflammatory cytokines. It is also required to produce anti-inflammatory 1,25-dihydroxyvitamin D. BTP-2, an inhibitor of cytosolic calcium, was used to suppress the LPS-induced increase in cytosolic calcium, which otherwise leads to an increase in proinflammatory cytokines. We showed that LPS increased the expression of the primary inflammatory mediators such as toll like receptor-4 (TLR-4), IL-1ß, interleukin-17 (IL-17), TNF-α, and interferon-γ (IFN-γ), which were normalized by the IGF-I + BTP-2 dual therapy in the lungs, along with improved vascular gene expression markers. The histologic lung injury score was markedly elevated by LPS and reduced to normal by the combination therapy. In conclusion, the LPS-induced increases in inflammatory cytokines, vascular injuries, and lung injuries were all improved by IGF-I + BTP-2 combination therapy.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/metabolismo , Anilidas/farmacologia , Citocinas/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Fator de Crescimento Insulin-Like I/farmacologia , Tiadiazóis/farmacologia , Lesão Pulmonar Aguda/patologia , Lesão Pulmonar Aguda/virologia , Anilidas/uso terapêutico , Animais , COVID-19/complicações , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Citocinas/genética , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica/genética , Imuno-Histoquímica , Fator de Crescimento Insulin-Like I/metabolismo , Fator de Crescimento Insulin-Like I/uso terapêutico , Interferon gama/genética , Interferon gama/metabolismo , Interleucina-17/genética , Interleucina-17/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Lipopolissacarídeos/toxicidade , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Tiadiazóis/uso terapêutico , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
7.
J Med Microbiol ; 70(6)2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34115583

RESUMO

Introduction. Leishmaniasis is a neglected tropical and subtropical disease caused by over 20 protozoan species.Hypothesis. Treatment of this complex disease with traditional synthetic drugs is a major challenge worldwide. Natural constituents are unique candidates for future therapeutic development.Aim. This study aimed to assess the in vivo anti-leishmanial effect of the Gossypium hirsutum extract, and its fractions compared to the standard drug (Glucantime, MA) in a murine model and explore the mechanism of action.Methodology. Footpads of BALB/c mice were infected with stationary phase promastigotes and treated topically and intraperitoneally with G. hirsutum extract, its fractions, or Glucantime, 4 weeks post-infection. The extract and fractions were prepared using the Soxhlet apparatus with chloroform followed by the column procedure.Results. The crude extract significantly decreased the footpad parasite load and lesion size compared to the untreated control group (P<0.05), as revealed by dilution assay, quantitative real-time PCR, and histopathological analyses. The primary mode of action involved an immunomodulatory role towards the Th1 response in the up-regulation of IFN-γ and IL-12 and the suppression of IL-10 gene expression profiling against cutaneous leishmaniasis caused by Leishmania major.Conclusion. This finding suggests that the extract possesses multiple combinatory effects of diverse bioactive phytochemical compositions that exert its mechanisms of action through agonistic-synergistic interactions. The topical extract formulation could be a suitable and unique candidate for future investigation and pharmacological development. Further studies are crucial to evaluate the therapeutic potentials of the extract alone and in combination with conventional drugs using clinical settings.


Assuntos
Antiprotozoários/uso terapêutico , Gossypium , Leishmania major/efeitos dos fármacos , Leishmaniose Cutânea/tratamento farmacológico , Fitoterapia , Extratos Vegetais/uso terapêutico , Administração Tópica , Animais , Antiprotozoários/farmacologia , Feminino , Injeções Intraperitoneais , Interferon gama/genética , Interferon gama/metabolismo , Interleucina-10/genética , Subunidade p40 da Interleucina-12/genética , Subunidade p40 da Interleucina-12/metabolismo , Leishmania major/fisiologia , Leishmaniose Cutânea/parasitologia , Leishmaniose Cutânea/patologia , Leishmaniose Cutânea/fisiopatologia , Linfonodos/patologia , Antimoniato de Meglumina/administração & dosagem , Antimoniato de Meglumina/uso terapêutico , Camundongos , Camundongos Endogâmicos BALB C , Carga Parasitária , Extratos Vegetais/administração & dosagem , Extratos Vegetais/farmacologia , Baço/parasitologia , Baço/patologia , Células Th1/imunologia , Transcriptoma
8.
Nat Commun ; 12(1): 3931, 2021 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-34168143

RESUMO

STAT1α is a key transcription factor driving pro-inflammatory responses in macrophages. We found that the interferon gamma (IFNγ)-regulated transcriptional program in macrophages is controlled by ADP-ribosylation (ADPRylation) of STAT1α, a post-translational modification resulting in the site-specific covalent attachment of ADP-ribose moieties. PARP-1, the major nuclear poly(ADP-ribose) polymerase (PARP), supports IFNγ-stimulated enhancer formation by regulating the genome-wide binding and IFNγ-dependent transcriptional activation of STAT1α. It does so by ADPRylating STAT1α on specific residues in its DNA-binding domain (DBD) and transcription activation (TA) domain. ADPRylation of the DBD controls STAT1α binding to its cognate DNA elements, whereas ADPRylation of the TA domain regulates enhancer activation by modulating STAT1α phosphorylation and p300 acetyltransferase activity. Loss of ADPRylation at either site leads to diminished IFNγ-dependent transcription and downstream pro-inflammatory responses. We conclude that PARP-1-mediated ADPRylation of STAT1α drives distinct enhancer activation mechanisms and is a critical regulator of inflammatory responses in macrophages.


Assuntos
ADP-Ribosilação , Interferon gama/metabolismo , Macrófagos/fisiologia , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/metabolismo , Animais , Sítios de Ligação , DNA/metabolismo , Elementos Facilitadores Genéticos , Feminino , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Inflamação/metabolismo , Inflamação/patologia , Macrófagos/patologia , Masculino , Camundongos Endogâmicos C57BL , Fosforilação , Poli(ADP-Ribose) Polimerase-1/genética , Poli(ADP-Ribose) Polimerase-1/metabolismo , Fator de Transcrição STAT1/química , Ativação Transcricional
9.
Aging (Albany NY) ; 13(11): 15620-15637, 2021 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-34106880

RESUMO

Amyloid ß (Aß) plays a major role in the neurodegeneration of Alzheimer's disease (AD). The accumulation of misfolded Aß causes oxidative stress and inflammatory damage leading to apoptotic cell death. Traditional Chinese herbal medicine (CHM) has been widely used in treating neurodegenerative diseases by reducing oxidative stress and neuroinflammation. We examined the neuroprotective effect of formulated CHM Shaoyao Gancao Tang (SG-Tang, made of Paeonia lactiflora and Glycyrrhiza uralensis at 1:1 ratio) in AD cell and mouse models. In Aß-GFP SH-SY5Y cells, SG-Tang reduced Aß aggregation and reactive oxygen species (ROS) production, as well as improved neurite outgrowth. When the Aß-GFP-expressing cells were stimulated with conditioned medium from interferon (IFN)-γ-activated HMC3 microglia, SG-Tang suppressed expressions of inducible nitric oxide synthase (iNOS), NLR family pyrin domain containing 1 (NLRP1) and 3 (NLRP3), tumor necrosis factor (TNF)-α, interleukin (IL)-1ß and IL-6, attenuated caspase-1 activity and ROS production, and promoted neurite outgrowth. In streptozocin-induced hyperglycemic APP/PS1/Tau triple transgenic (3×Tg-AD) mice, SG-Tang also reduced expressions of NLRP1, NLRP3, Aß and Tau in hippocampus and cortex, as well as improved working and spatial memories in Y maze and Morris water maze. Collectively, our results demonstrate the potential of SG-Tang in treating AD by moderating neuroinflammation.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Cognição , Medicamentos de Ervas Chinesas/uso terapêutico , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Neuroproteção , Doença de Alzheimer/fisiopatologia , Peptídeos beta-Amiloides/metabolismo , Animais , Linhagem Celular , Cognição/efeitos dos fármacos , Meios de Cultivo Condicionados/farmacologia , Modelos Animais de Doenças , Medicamentos de Ervas Chinesas/farmacologia , Proteínas de Fluorescência Verde/metabolismo , Humanos , Interferon gama/metabolismo , Memória/efeitos dos fármacos , Transtornos da Memória/complicações , Transtornos da Memória/fisiopatologia , Camundongos Transgênicos , Microglia/efeitos dos fármacos , Microglia/metabolismo , Modelos Biológicos , Crescimento Neuronal/efeitos dos fármacos , Neuroproteção/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Agregados Proteicos/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Aprendizagem Espacial/efeitos dos fármacos , Proteínas tau/metabolismo
10.
Clin Invest Med ; 44(2): E5-18, 2021 06 14.
Artigo em Inglês | MEDLINE | ID: mdl-34152702

RESUMO

PURPOSE: This literature review summarizes the main immunological characteristics of type III interferons (IFN) and highlights the clinically relevant aspects and future therapeutic perspectives for these inflammatory molecules. SOURCE: Relevant articles in PubMed MEDLINE from the first publication (2003) until 2020. N=101 articles were included in this review. PRINCIPAL FINDINGS: Type III IFNs represent a relatively newly described inflammatory cytokine family. Although they induce substantially similar signalling to the well-known type I IFNs, significant functional differences make these molecules remarkable. Type III IFNs have extensive biological effects, contributing to the pathogenesis of several diseases and also offering new diagnostic and therapeutic approaches: 1) their potent anti-viral properties make them promising therapeutics against viral hepatitis and even against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is causing the current coronavirus disease 2019 (COVID-19) pandemic; 2) imbalances in the IFN-λs contribute to several forms of chronic inflammation (e.g., systemic and organ-specific autoimmune diseases) and potentially predict disease progression and therapeutic response to biologic therapies; and 3) the antitumor properties of the type III IFNs open up new therapeutic perspectives against malignant diseases. CONCLUSION: Over the last 18 years, researchers have gathered extensive information about the presence and role of these versatile inflammatory cytokines in human diseases, but further research is needed to clarify the mechanistic background of those observations. Better understanding of their biological activities will permit us to use type III IFNs more efficiently in new diagnostic approaches and individualized therapies, consequently improving patient care.


Assuntos
COVID-19/metabolismo , Citocinas/metabolismo , Inflamação/metabolismo , Interferons/fisiologia , Animais , Antivirais/farmacologia , Doenças Autoimunes/metabolismo , Infecções Bacterianas/metabolismo , Progressão da Doença , Humanos , Interferon gama/metabolismo , Micoses/metabolismo , SARS-CoV-2 , Transdução de Sinais
11.
Nat Commun ; 12(1): 3754, 2021 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-34145241

RESUMO

Atherosclerosis is a major underlying cause of cardiovascular disease. Previous studies showed that inhibition of the co-stimulatory CD40 ligand (CD40L)-CD40 signaling axis profoundly attenuates atherosclerosis. As CD40L exerts multiple functions depending on the cell-cell interactions involved, we sought to investigate the function of the most relevant CD40L-expressing cell types in atherosclerosis: T cells and platelets. Atherosclerosis-prone mice with a CD40L-deficiency in CD4+ T cells display impaired Th1 polarization, as reflected by reduced interferon-γ production, and smaller atherosclerotic plaques containing fewer T-cells, smaller necrotic cores, an increased number of smooth muscle cells and thicker fibrous caps. Mice with a corresponding CD40-deficiency in CD11c+ dendritic cells phenocopy these findings, suggesting that the T cell-dendritic cell CD40L-CD40 axis is crucial in atherogenesis. Accordingly, sCD40L/sCD40 and interferon-γ concentrations in carotid plaques and plasma are positively correlated in patients with cerebrovascular disease. Platelet-specific deficiency of CD40L does not affect atherogenesis but ameliorates atherothrombosis. Our results establish divergent and cell-specific roles of CD40L-CD40 in atherosclerosis, which has implications for therapeutic strategies targeting this pathway.


Assuntos
Aterosclerose/patologia , Linfócitos T CD4-Positivos/metabolismo , Antígenos CD40/metabolismo , Ligante de CD40/metabolismo , Interferon gama/metabolismo , Placa Aterosclerótica/patologia , Animais , Plaquetas/metabolismo , Linfócitos T CD4-Positivos/citologia , Doenças Cardiovasculares/patologia , Células Dendríticas/imunologia , Camundongos , Camundongos Knockout , Miócitos de Músculo Liso/citologia , Transdução de Sinais/fisiologia , Trombose/patologia
12.
Int J Med Sci ; 18(12): 2561-2569, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34104087

RESUMO

SARS-CoV-2 infection poses a global challenge to human health. Upon viral infection, host cells initiate the innate antiviral response, which primarily involves type I interferons (I-IFNs), to enable rapid elimination of the invading virus. Previous studies revealed that SARS-CoV-2 infection limits the expression of I-IFNs in vitro and in vivo, but the underlying mechanism remains incompletely elucidated. In the present study, we performed data mining and longitudinal data analysis using SARS-CoV-2-infected normal human bronchial epithelial (NHBE) cells and ferrets, and the results confirmed the strong inhibitory effect of SARS-CoV-2 on the induction of I-IFNs. Moreover, we identified genes that are negatively correlated with IFNB1 expression in vitro and in vivo based on Pearson correlation analysis. We found that SARS-CoV-2 activates numerous intrinsic pathways, such as the circadian rhythm, phosphatidylinositol signaling system, peroxisome, and TNF signaling pathways, to inhibit I-IFNs. These intrinsic inhibitory pathways jointly facilitate the successful immune evasion of SARS-CoV-2. Our study elucidates the underlying mechanism by which SARS-CoV-2 evades the host innate antiviral response in vitro and in vivo, providing theoretical evidence for targeting these immune evasion-associated pathways to combat SARS-CoV-2 infection.


Assuntos
COVID-19/imunologia , Interações Hospedeiro-Patógeno/imunologia , Interferon gama/metabolismo , SARS-CoV-2/imunologia , Animais , Brônquios/citologia , COVID-19/virologia , Linhagem Celular , Conjuntos de Dados como Assunto , Modelos Animais de Doenças , Células Epiteliais , Furões , Regulação da Expressão Gênica/imunologia , Interações Hospedeiro-Patógeno/genética , Humanos , Imunidade Inata , Interferon gama/imunologia , RNA-Seq , Mucosa Respiratória/citologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia
13.
Int J Mol Sci ; 22(11)2021 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-34070449

RESUMO

Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells that are responsible for immunosuppression in tumor microenvironment. Here we report the impact of mucin 1 (MUC1), a transmembrane glycoprotein, on proliferation and functional activity of MDSCs. To determine the role of MUC1 in MDSC phenotype, we analyzed MDSCs derived from wild type (WT) and MUC1-knockout (MUC1KO) mice bearing syngeneic pancreatic (KCKO) or breast (C57MG) tumors. We observed enhanced tumor growth of pancreatic and breast tumors in the MUC1KO mice compared to the WT mice. Enhanced tumor growth in the MUC1KO mice was associated with increased numbers of suppressive MDSCs and T regulatory (Tregs) cells in the tumor microenvironment. Compared to the WT host, MUC1KO host showed higher levels of iNOS, ARG1, and TGF-ß, thus promoting proliferation of MDSCs with an immature and immune suppressive phenotype. When co-cultured with effector T cells, MDSCs from MUC1KO mice led to higher repression of IL-2 and IFN-γ production by T cells as compared to MDSCs from WT mice. Lastly, MDSCs from MUC1KO mice showed higher levels of c-Myc and activated pSTAT3 as compared to MDSCs from WT mice, suggesting increased survival, proliferation, and prevention of maturation of MDSCs in the MUC1KO host. We report diminished T cell function in the KO versus WT mice. In summary, the data suggest that MUC1 may regulate signaling pathways that are critical to maintain the immunosuppressive properties of MDSCs.


Assuntos
Neoplasias da Mama/metabolismo , Mucina-1/metabolismo , Células Supressoras Mieloides/imunologia , Neoplasias Pancreáticas/metabolismo , Linfócitos T Reguladores/imunologia , Microambiente Tumoral/imunologia , Animais , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Técnicas de Cocultura , Modelos Animais de Doenças , Feminino , Interferon gama/metabolismo , Interleucina-2/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mucina-1/genética , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Baço/citologia , Baço/metabolismo , Fator de Crescimento Transformador beta/sangue , Microambiente Tumoral/genética
14.
Int Immunopharmacol ; 96: 107797, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34162159

RESUMO

Specific antibodies against SARS-CoV-2 structural protein have a wide range of effects in the diagnose, prevention and treatment of the COVID-19 epidemic. Among them, egg yolk immunoglobulin Y (IgY), which has high safety, high yield, and without inducing antibody-dependent enhancement, is an important biological candidate. In this study, specific IgY against the conservative nucleocapsid protein (NP) of SARS-CoV-2 was obtained by immunizing hens. Through a series of optimized precipitation and ultrafiltration extraction schemes, its purity was increased to 98%. The hyperimmune IgY against NP (N-IgY) at a titer of 1:50,000 showed strong NP binding ability, which laid the foundation of N-IgY's application targeting NP. In an in vitro immunoregulatory study, N-IgY (1 mg/mL) modulated NP-induced immune response by alleviating type II interferon secretion stimulated by NP (20 µg/mL). In summary, N-IgY can be mass produced by achievable method, which endows it with potential value against the current COVID-19 pandemic.


Assuntos
Anticorpos/imunologia , Antivirais/imunologia , COVID-19/imunologia , Imunoglobulinas/imunologia , Fatores Imunológicos/imunologia , Interferon gama/metabolismo , SARS-CoV-2/imunologia , Animais , Anticorpos/farmacologia , Antivirais/farmacologia , COVID-19/terapia , Galinhas , Desenvolvimento de Medicamentos , Gema de Ovo/química , Gema de Ovo/metabolismo , Humanos , Imunidade , Imunoglobulinas/farmacologia , Fatores Imunológicos/farmacologia , Imunomodulação , Técnicas In Vitro , Proteínas do Nucleocapsídeo/imunologia , Proteínas do Nucleocapsídeo/metabolismo , SARS-CoV-2/metabolismo
15.
Front Immunol ; 12: 679482, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34177923

RESUMO

Infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes COVID-19, a disease that involves significant lung tissue damage. How SARS-CoV-2 infection leads to lung injury remains elusive. The open reading frame 8 (ORF8) protein of SARS-CoV-2 (ORF8SARS-CoV-2) is a unique accessory protein, yet little is known about its cellular function. We examined the cellular distribution of ORF8SARS-CoV-2 and its role in the regulation of human lung epithelial cell proliferation and antiviral immunity. Using live imaging and immunofluorescent staining analyses, we found that ectopically expressed ORF8SARS-CoV-2 forms aggregates in the cytosol and nuclear compartments of lung epithelial cells. Using in silico bioinformatic analysis, we found that ORF8SARS-CoV-2 possesses an intrinsic aggregation characteristic at its N-terminal residues 1-18. Cell culture did not reveal any effects of ORF8SARS-CoV-2 expression on lung epithelial cell proliferation and cell cycle progression, suggesting that ORF8SARS-CoV-2 aggregates do not affect these cellular processes. Interestingly, ectopic expression of ORF8SARS-CoV-2 in lung epithelial cells suppressed basal expression of several antiviral molecules, including DHX58, ZBP1, MX1, and MX2. In addition, expression of ORF8SARS-CoV-2 attenuated the induction of antiviral molecules by IFNγ but not by IFNß in lung epithelial cells. Taken together, ORF8SARS-CoV-2 is a unique viral accessory protein that forms aggregates when expressing in lung epithelial cells. It potently inhibits the expression of lung cellular anti-viral proteins at baseline and in response to IFNγ in lung epithelial cells, which may facilitate SARS-CoV-2 escape from the host antiviral innate immune response during early viral infection. In addition, it seems that formation of ORF8SARS-CoV-2 aggregate is independent from the viral infection. Thus, it would be interesting to examine whether any COVID-19 patients exhibit persistent ORF8 SARS-CoV-2 expression after recovering from SARS-CoV-2 infection. If so, the pathogenic effect of prolonged ORF8SARS-CoV-2 expression and its association with post-COVID symptoms warrant investigation in the future.


Assuntos
COVID-19/imunologia , Pulmão/patologia , Mucosa Respiratória/fisiologia , SARS-CoV-2/fisiologia , Proteínas Virais/metabolismo , COVID-19/virologia , Regulação da Expressão Gênica , Células HEK293 , Humanos , Imunidade , Interferon gama/metabolismo , Espaço Intracelular , Agregação Patológica de Proteínas , Mucosa Respiratória/virologia
16.
Nat Commun ; 12(1): 3299, 2021 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-34083537

RESUMO

Bioenergetic perturbations driving neoplastic growth increase the production of reactive oxygen species (ROS), requiring a compensatory increase in ROS scavengers to limit oxidative stress. Intervention strategies that simultaneously induce energetic and oxidative stress therefore have therapeutic potential. Phenformin is a mitochondrial complex I inhibitor that induces bioenergetic stress. We now demonstrate that inflammatory mediators, including IFNγ and polyIC, potentiate the cytotoxicity of phenformin by inducing a parallel increase in oxidative stress through STAT1-dependent mechanisms. Indeed, STAT1 signaling downregulates NQO1, a key ROS scavenger, in many breast cancer models. Moreover, genetic ablation or pharmacological inhibition of NQO1 using ß-lapachone (an NQO1 bioactivatable drug) increases oxidative stress to selectively sensitize breast cancer models, including patient derived xenografts of HER2+ and triple negative disease, to the tumoricidal effects of phenformin. We provide evidence that therapies targeting ROS scavengers increase the anti-neoplastic efficacy of mitochondrial complex I inhibitors in breast cancer.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Fenformin/farmacologia , Fator de Transcrição STAT1/metabolismo , Animais , Antineoplásicos/administração & dosagem , Linhagem Celular Tumoral , Sinergismo Farmacológico , Complexo I de Transporte de Elétrons/antagonistas & inibidores , Metabolismo Energético/efeitos dos fármacos , Feminino , Glutationa/antagonistas & inibidores , Glutationa/biossíntese , Humanos , Interferon gama/administração & dosagem , Interferon gama/deficiência , Interferon gama/metabolismo , Células MCF-7 , Neoplasias Mamárias Experimentais/tratamento farmacológico , Neoplasias Mamárias Experimentais/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Camundongos SCID , NAD(P)H Desidrogenase (Quinona)/antagonistas & inibidores , NAD(P)H Desidrogenase (Quinona)/metabolismo , Naftoquinonas/administração & dosagem , Estresse Oxidativo/efeitos dos fármacos , Fenformin/administração & dosagem , Poli I-C/administração & dosagem , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição STAT1/agonistas , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Nat Commun ; 12(1): 3010, 2021 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-34021148

RESUMO

Resident memory T cells (TRM) positioned within the respiratory tract are probably required to limit SARS-CoV-2 spread and COVID-19. Importantly, TRM are mostly non-recirculating, which reduces the window of opportunity to examine these cells in the blood as they move to the lung parenchyma. Here, we identify circulating virus-specific T cell responses during acute infection with functional, migratory and apoptotic patterns modulated by viral proteins and associated with clinical outcome. Disease severity is associated predominantly with IFNγ and IL-4 responses, increased responses against S peptides and apoptosis, whereas non-hospitalized patients have increased IL-12p70 levels, degranulation in response to N peptides and SARS-CoV-2-specific CCR7+ T cells secreting IL-10. In convalescent patients, lung-TRM are frequently detected even 10 months after initial infection, in which contemporaneous blood does not reflect tissue-resident profiles. Our study highlights a balanced anti-inflammatory antiviral response associated with a better outcome and persisting TRM cells as important for future protection against SARS-CoV-2 infection.


Assuntos
COVID-19/imunologia , Memória Imunológica/imunologia , Pulmão/imunologia , SARS-CoV-2/imunologia , Linfócitos T/imunologia , Apoptose/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , COVID-19/virologia , Movimento Celular/imunologia , Humanos , Interferon gama/imunologia , Interferon gama/metabolismo , Interleucina-4/imunologia , Interleucina-4/metabolismo , Pulmão/virologia , SARS-CoV-2/fisiologia , Linfócitos T/metabolismo
18.
Nat Commun ; 12(1): 2620, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33976173

RESUMO

Tumor associated macrophage responses are regulated by distinct metabolic states that affect their function. However, the ability of specific signals in the local tumor microenvironment to program macrophage metabolism remains under investigation. Here, we identify NAMPT, the rate limiting enzyme in NAD salvage synthesis, as a target of STAT1 during cellular activation by interferon gamma, an important driver of macrophage polarization and antitumor responses. We demonstrate that STAT1 occupies a conserved element within the first intron of Nampt, termed Nampt-Regulatory Element-1 (NRE1). Through disruption of NRE1 or pharmacological inhibition, a subset of M1 genes is sensitive to NAMPT activity through its impact on glycolytic processes. scRNAseq is used to profile in vivo responses by NRE1-deficient, tumor-associated leukocytes in melanoma tumors through the creation of a unique mouse strain. Reduced Nampt and inflammatory gene expression are present in specific myeloid and APC populations; moreover, targeted ablation of NRE1 in macrophage lineages results in greater tumor burden. Finally, elevated NAMPT expression correlates with IFNγ responses and melanoma patient survival. This study identifies IFN and STAT1-inducible Nampt as an important factor that shapes the metabolic program and function of tumor associated macrophages.


Assuntos
Citocinas/genética , Melanoma/genética , Nicotinamida Fosforribosiltransferase/genética , Fator de Transcrição STAT1/metabolismo , Neoplasias Cutâneas/genética , Macrófagos Associados a Tumor/imunologia , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Elementos Facilitadores Genéticos , Feminino , Regulação Neoplásica da Expressão Gênica/imunologia , Células HEK293 , Humanos , Interferon gama/metabolismo , Estimativa de Kaplan-Meier , Masculino , Melanoma/imunologia , Melanoma/mortalidade , Melanoma/patologia , Camundongos , Camundongos Knockout , Nicotinamida Fosforribosiltransferase/metabolismo , Células RAW 264.7 , RNA-Seq , Receptores de Interferon/genética , Receptores de Interferon/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/patologia , Células THP-1 , Macrófagos Associados a Tumor/metabolismo , Regulação para Cima , Efeito Warburg em Oncologia
19.
Nat Commun ; 12(1): 2537, 2021 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-33953170

RESUMO

Metastasis accounts for 90% of cancer-related deaths and, currently, there are no effective clinical therapies to block the metastatic cascade. A need to develop novel therapies specifically targeting fundamental metastasis processes remains urgent. Here, we demonstrate that Salmonella YB1, an engineered oxygen-sensitive strain, potently inhibits metastasis of a broad range of cancers. This process requires both IFN-γ and NK cells, as the absence of IFN-γ greatly reduces, whilst depletion of NK cells in vivo completely abolishes, the anti-metastatic ability of Salmonella. Mechanistically, we find that IFN-γ is mainly produced by NK cells during early Salmonella infection, and in turn, IFN-γ promotes the accumulation, activation, and cytotoxicity of NK cells, which kill the metastatic cancer cells thus achieving an anti-metastatic effect. Our findings highlight the significance of a self-regulatory feedback loop of NK cells in inhibiting metastasis, pointing a possible approach to develop anti-metastatic therapies by harnessing the power of NK cells.


Assuntos
Interferon gama/metabolismo , Interferon gama/farmacologia , Células Matadoras Naturais/imunologia , Ativação Linfocitária/efeitos dos fármacos , Metástase Neoplásica/imunologia , Infecções por Salmonella/metabolismo , Salmonella/genética , Animais , Citocinas/metabolismo , Feminino , Imunidade Inata , Imunoterapia/métodos , Interferon gama/genética , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Infecções por Salmonella/tratamento farmacológico
20.
J Med Chem ; 64(11): 7596-7616, 2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-34019405

RESUMO

Cyclic dinucleotides (CDNs) are second messengers that bind to the stimulator of interferon genes (STING) and trigger the expression of type I interferons and proinflammatory cytokines. Here we evaluate the activity of 3',3'-c-di(2'F,2'dAMP) and its phosphorothioate analogues against five STING allelic forms in reporter-cell-based assays and rationalize our findings with X-ray crystallography and quantum mechanics/molecular mechanics calculations. We show that the presence of fluorine in the 2' position of 3',3'-c-di(2'F,2'dAMP) improves its activity not only against the wild type (WT) but also against REF and Q STING. Additionally, we describe the synthesis of the acyloxymethyl and isopropyloxycarbonyl phosphoester prodrugs of CDNs. Masking the negative charges of the CDNs results in an up to a 1000-fold improvement of the activities of the prodrugs relative to those of their parent CDNs. Finally, the uptake and intracellular cleavage of pivaloyloxymethyl prodrugs to the parent CDN is rapid, reaching a peak intracellular concentration within 2 h.


Assuntos
Ésteres/química , Proteínas de Membrana/agonistas , Fosfatos/química , Pró-Fármacos/síntese química , Cristalografia por Raios X , Teoria da Densidade Funcional , Ésteres/farmacologia , Ésteres/uso terapêutico , Células HEK293 , Humanos , Interferon gama/metabolismo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Espectroscopia de Ressonância Magnética , Proteínas de Membrana/metabolismo , Fosfatos/metabolismo , Fosfatos/farmacologia , Fosfatos/uso terapêutico , Pró-Fármacos/química , Pró-Fármacos/metabolismo , Pró-Fármacos/farmacologia , Fator de Necrose Tumoral alfa/metabolismo
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