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1.
Mol Syst Biol ; 17(9): e10426, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34486798

RESUMO

Although 15-20% of COVID-19 patients experience hyper-inflammation induced by massive cytokine production, cellular triggers of this process and strategies to target them remain poorly understood. Here, we show that the N-terminal domain (NTD) of the SARS-CoV-2 spike protein substantially induces multiple inflammatory molecules in myeloid cells and human PBMCs. Using a combination of phenotypic screening with machine learning-based modeling, we identified and experimentally validated several protein kinases, including JAK1, EPHA7, IRAK1, MAPK12, and MAP3K8, as essential downstream mediators of NTD-induced cytokine production, implicating the role of multiple signaling pathways in cytokine release. Further, we found several FDA-approved drugs, including ponatinib, and cobimetinib as potent inhibitors of the NTD-mediated cytokine release. Treatment with ponatinib outperforms other drugs, including dexamethasone and baricitinib, inhibiting all cytokines in response to the NTD from SARS-CoV-2 and emerging variants. Finally, ponatinib treatment inhibits lipopolysaccharide-mediated cytokine release in myeloid cells in vitro and lung inflammation mouse model. Together, we propose that agents targeting multiple kinases required for SARS-CoV-2-mediated cytokine release, such as ponatinib, may represent an attractive therapeutic option for treating moderate to severe COVID-19.


Assuntos
Antivirais/farmacologia , Citocinas/metabolismo , Interações Hospedeiro-Patógeno/fisiologia , Animais , Azetidinas/farmacologia , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Humanos , Imidazóis/farmacologia , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Janus Quinase 1/metabolismo , Lipopolissacarídeos/toxicidade , Aprendizado de Máquina , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/virologia , Inibidores de Proteínas Quinases/farmacologia , Purinas/farmacologia , Pirazóis/farmacologia , Piridazinas/farmacologia , Células RAW 264.7 , SARS-CoV-2/patogenicidade , Glicoproteína da Espícula de Coronavírus/metabolismo , Sulfonamidas/farmacologia
2.
Immunol Lett ; 237: 33-41, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34228987

RESUMO

OBJECTIVE: In this study, we focused on the interaction between SARS-CoV-2 and host Type I Interferon (IFN) response, so as to identify whether IFN effects could be influenced by the products of SARS-CoV-2. METHODS: All the structural and non-structural proteins of SARS-CoV-2 were transfected and overexpressed in the bronchial epithelial cell line BEAS-2B respectively, and typical antiviral IFN-stimulated gene (ISG) ISG15 expression was detected by qRT-PCR. RNA-seq based transcriptome analysis was performed between control and Spike (S) protein-overexpressed BEAS-2B cells. The expression of ACE2 and IFN effector JAK-STAT signaling activation were detected in control and S protein-overexpressed BEAS-2B cells by qRT-PCR or/and Western blot respectively. The interaction between S protein with STAT1 and STAT2, and the association between JAK1 with downstream STAT1 and STAT2 were measured in BEAS-2B cells by co-immunoprecipitation (co-IP). RESULTS: S protein could activate IFN effects and downstream ISGs expression. By transcriptome analysis, overexpression of S protein induced a set of genes expression, including series of ISGs and the SARS-CoV-2 receptor ACE2. Mechanistically, S protein enhanced the association between the upstream JAK1 and downstream STAT1 and STAT2, so as to promote STAT1 and STAT2 phosphorylation and ACE2 expression. CONCLUSION: SARS-CoV-2 S protein enhances ACE2 expression via facilitating IFN effects, which may help its infection.


Assuntos
Enzima de Conversão de Angiotensina 2/metabolismo , Brônquios/efeitos dos fármacos , COVID-19/virologia , Células Epiteliais/efeitos dos fármacos , Interferon alfa-2/farmacologia , SARS-CoV-2/patogenicidade , Glicoproteína da Espícula de Coronavírus/metabolismo , Enzima de Conversão de Angiotensina 2/genética , Brônquios/enzimologia , Brônquios/virologia , COVID-19/enzimologia , Citocinas/genética , Citocinas/metabolismo , Células Epiteliais/enzimologia , Células Epiteliais/virologia , Células HEK293 , Interações Hospedeiro-Patógeno , Humanos , Janus Quinase 1/metabolismo , Fosforilação , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT2/metabolismo , Transdução de Sinais , Glicoproteína da Espícula de Coronavírus/genética , Ubiquitinas/genética , Ubiquitinas/metabolismo , Regulação para Cima
3.
J Virol ; 95(19): e0086221, 2021 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-34260266

RESUMO

SARS-CoV-2 can infect multiple organs, including lung, intestine, kidney, heart, liver, and brain. The molecular details of how the virus navigates through diverse cellular environments and establishes replication are poorly defined. Here, we generated a panel of phenotypically diverse, SARS-CoV-2-infectible human cell lines representing different body organs and performed longitudinal survey of cellular proteins and pathways broadly affected by the virus. This revealed universal inhibition of interferon signaling across cell types following SARS-CoV-2 infection. We performed systematic analyses of the JAK-STAT pathway in a broad range of cellular systems, including immortalized cells and primary-like cardiomyocytes, and found that SARS-CoV-2 targeted the proximal pathway components, including Janus kinase 1 (JAK1), tyrosine kinase 2 (Tyk2), and the interferon receptor subunit 1 (IFNAR1), resulting in cellular desensitization to type I IFN. Detailed mechanistic investigation of IFNAR1 showed that the protein underwent ubiquitination upon SARS-CoV-2 infection. Furthermore, chemical inhibition of JAK kinases enhanced infection of stem cell-derived cultures, indicating that the virus benefits from inhibiting the JAK-STAT pathway. These findings suggest that the suppression of interferon signaling is a mechanism widely used by the virus to evade antiviral innate immunity, and that targeting the viral mediators of immune evasion may help block virus replication in patients with COVID-19. IMPORTANCE SARS-CoV-2 can infect various organs in the human body, but the molecular interface between the virus and these organs remains unexplored. In this study, we generated a panel of highly infectible human cell lines originating from various body organs and employed these cells to identify cellular processes commonly or distinctly disrupted by SARS-CoV-2 in different cell types. One among the universally impaired processes was interferon signaling. Systematic analysis of this pathway in diverse culture systems showed that SARS-CoV-2 targets the proximal JAK-STAT pathway components, destabilizes the type I interferon receptor though ubiquitination, and consequently renders the infected cells resistant to type I interferon. These findings illuminate how SARS-CoV-2 can continue to propagate in different tissues even in the presence of a disseminated innate immune response.


Assuntos
COVID-19/metabolismo , Interações entre Hospedeiro e Microrganismos/fisiologia , Janus Quinases/metabolismo , SARS-CoV-2/metabolismo , Linhagem Celular , Regulação da Expressão Gênica , Humanos , Evasão da Resposta Imune , Imunidade Inata , Interferon Tipo I/metabolismo , Janus Quinase 1/metabolismo , Miócitos Cardíacos , Receptor de Interferon alfa e beta/metabolismo , Fator de Transcrição STAT1/metabolismo , Transdução de Sinais , TYK2 Quinase/metabolismo , Replicação Viral
4.
FASEB J ; 35(7): e21721, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34118090

RESUMO

Mineral homeostasis is regulated by a complex network involving endocrine actions by calcitriol, parathyroid hormone (PTH), and FGF23 on several organs including kidney, intestine, and bone. Alterations of mineral homeostasis are found in chronic kidney disease and other systemic disorders. The interplay between the immune system and the skeletal system is not fully understood, but cytokines play a major role in modulating calcitriol production and function. One of the main cellular signaling pathways mediating cytokine function is the Janus kinase (JAK)--signal transducer and activator of transcription (STAT) pathway. Here, we used a mouse model (Jak1S645P+/- ) that resembles a constitutive activating mutation of the Jak1/Stat3 signaling pathway in humans, and shows altered mineral metabolism, with higher fibroblast growth factor 23 (FGF23) levels, lower PTH levels, and higher calcitriol levels. The higher calcitriol levels are probably due to extrarenal calcitriol production. Furthermore, systemic Jak1/Stat3 activation led to growth impairment and skeletal alterations. The growth plate in long bones showed decreased chondrocyte proliferation rates and reduced height of terminal chondrocytes. Furthermore, we demonstrate that Jak1 is also involved in bone remodeling early in life. Jak1S645P+/- animals have decreased bone and cortical volume, imbalanced bone remodeling, reduced MAP kinase signaling, and local inflammation. In conclusion, Jak1 plays a major role in bone health probably both, directly and systemically by regulating mineral homeostasis. Understanding the role of this signaling pathway will contribute to a better knowledge in bone growth and in mineral physiology, and to the development of selective Jak inhibitors as osteoprotective agents.


Assuntos
Osso e Ossos/metabolismo , Osso e Ossos/fisiologia , Calcitriol/metabolismo , Transtornos do Crescimento/metabolismo , Janus Quinase 1/metabolismo , Transdução de Sinais/fisiologia , Animais , Remodelação Óssea/fisiologia , Proliferação de Células/fisiologia , Condrócitos/metabolismo , Condrócitos/fisiologia , Citocinas/metabolismo , Feminino , Fatores de Crescimento de Fibroblastos , Lâmina de Crescimento/metabolismo , Lâmina de Crescimento/fisiologia , Homeostase/fisiologia , Humanos , Inflamação/metabolismo , Rim/metabolismo , Rim/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C3H , Mutação/genética , Hormônio Paratireóideo/metabolismo , Fator de Transcrição STAT3/metabolismo
5.
Life Sci ; 281: 119754, 2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34174323

RESUMO

AIM: The present study was undertaken to elucidate the potential protective mechanism of berberine (BBR) and/or zinc (Zn) against methotrexate (MTX)-induced intestinal injury. METHODS: Five groups of rats were assigned; normal group (received vehicle), MTX group (20 mg/kg; i.p. single dose), and the other three groups received a single daily oral dose of BBR (50 mg/kg), Zn (5 mg/kg), and BBR plus Zn respectively, for 5 days before MTX and 5 days after. RESULTS: Our results emphasized the toxic effect of MTX on rat's intestine as shown by disturbance of oxidant/antioxidant status, down-regulation of NRF2, SIRT1, FOXO-3, Akt, and mTOR expressions, along with up-regulation of GSK-3ß, JAK1, and STAT-3 expressions. Besides, severe intestinal histopathological changes were also observed. On the contrary, BBR and/or Zn produced marked protection against MTX-induced intestinal toxicity via amelioration of oxidative stress, improving NRF2, SIRT1, FOXO-3, GSK-3ß, Akt, mTOR, JAK1, and STAT-3 alterations. Moreover, our treatments significantly restored histopathological abnormalities. Interestingly, combination therapy of BBR plus Zn exhibited higher effectiveness than mono-therapy. SIGNIFICANCE: BBR plus Zn could be used as a novel therapy for the treatment of MTX-induced intestinal damage through modulation of GSK-3ß/NRF2, Akt/mTOR, JAK1/STAT-3, and SIRT1/FOXO-3 signaling pathways.


Assuntos
Berberina/farmacologia , Proteína Forkhead Box O3/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Intestinos/efeitos dos fármacos , Janus Quinase 1/metabolismo , Metotrexato/toxicidade , Fator 2 Relacionado a NF-E2/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Zinco/farmacologia , Animais , Inflamação/prevenção & controle , Intestinos/enzimologia , Intestinos/patologia , Masculino , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Wistar , Sirtuína 1/metabolismo
6.
J Hepatol ; 75(3): 647-658, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-33991637

RESUMO

BACKGROUND AND AIMS: COVID-19 is associated with liver injury and elevated interleukin-6 (IL-6). We hypothesized that IL-6 trans-signaling in liver sinusoidal endothelial cells (LSECs) leads to endotheliopathy (a proinflammatory and procoagulant state) and liver injury in COVID-19. METHODS: Coagulopathy, endotheliopathy, and alanine aminotransferase (ALT) were retrospectively analyzed in a subset (n = 68), followed by a larger cohort (n = 3,780) of patients with COVID-19. Liver histology from 43 patients with COVID-19 was analyzed for endotheliopathy and its relationship to liver injury. Primary human LSECs were used to establish the IL-6 trans-signaling mechanism. RESULTS: Factor VIII, fibrinogen, D-dimer, von Willebrand factor (vWF) activity/antigen (biomarkers of coagulopathy/endotheliopathy) were significantly elevated in patients with COVID-19 and liver injury (elevated ALT). IL-6 positively correlated with vWF antigen (p = 0.02), factor VIII activity (p = 0.02), and D-dimer (p <0.0001). On liver histology, patients with COVID-19 and elevated ALT had significantly increased vWF and platelet staining, supporting a link between liver injury, coagulopathy, and endotheliopathy. Intralobular neutrophils positively correlated with platelet (p <0.0001) and vWF (p <0.01) staining, and IL-6 levels positively correlated with vWF staining (p <0.01). IL-6 trans-signaling leads to increased expression of procoagulant (factor VIII, vWF) and proinflammatory factors, increased cell surface vWF (p <0.01), and increased platelet attachment in LSECs. These effects were blocked by soluble glycoprotein 130 (IL-6 trans-signaling inhibitor), the JAK inhibitor ruxolitinib, and STAT1/3 small-interfering RNA knockdown. Hepatocyte fibrinogen expression was increased by the supernatant of LSECs subjected to IL-6 trans-signaling. CONCLUSION: IL-6 trans-signaling drives the coagulopathy and hepatic endotheliopathy associated with COVID-19 and could be a possible mechanism behind liver injury in these patients. LAY SUMMARY: Patients with SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) infection often have liver injury, but why this occurs remains unknown. High levels of interleukin-6 (IL-6) and its circulating receptor, which form a complex to induce inflammatory signals, have been observed in patients with COVID-19. This paper demonstrates that the IL-6 signaling complex causes harmful changes to liver sinusoidal endothelial cells and may promote blood clotting and contribute to liver injury.


Assuntos
COVID-19/complicações , Células Endoteliais/patologia , Interleucina-6/fisiologia , Hepatopatias/etiologia , SARS-CoV-2 , Adulto , Transtornos da Coagulação Sanguínea/etiologia , Fibrinogênio/análise , Humanos , Interleucina-6/sangue , Janus Quinase 1/metabolismo , Pirazóis/farmacologia , Estudos Retrospectivos , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/fisiologia , Fator de von Willebrand/análise
7.
Phytomedicine ; 87: 153574, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34006448

RESUMO

BACKGROUND: Leelamine (LEE) is a lipophilic diterpene amine phytochemical, which can be naturally extracted from pine's bark trees. It has been extensively studied recently for its promising chemopreventive and anti-cancer effects against various cancers such as that of prostate and breast. HYPOTHESIS: We examined the potential impact of LEE in affecting the activation of signal transducer and activator of transcription 3 (STAT3) and promoting apoptosis in human multiple myeloma (MM) cells. METHODS: We evaluated the effect of LEE on STAT3 signaling pathway in MM cells by using Western blot analysis and reverse transcription polymerase chain reaction (RT-PCR). Thereafter, apoptosis was evaluated using cell cycle analysis and Annexin V assay. RESULTS: We noted that LEE could attenuate the phosphorylation of STAT3 and other up-stream signaling molecules such as JAK1, JAK2, and Src activation in U266 and MM.1S cells. It also diminished STAT3 translocation into the nucleus and enhanced the expression of protein-tyrosine phosphatase epsilon (PTPε). Additionally, LEE caused cell cycle arrest and synergistically augmented the apoptotic actions of bortezomib against MM cells. CONCLUSIONS: Our data indicates that LEE could block STAT3 signaling cascade linked to tumorigenesis and can be used in combination with approved anti-cancer agents in attenuating MM growth and survival.


Assuntos
Abietanos/farmacologia , Janus Quinase 1/metabolismo , Janus Quinase 2/metabolismo , Mieloma Múltiplo/metabolismo , Fator de Transcrição STAT3/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Janus Quinase 1/genética , Janus Quinase 2/genética , Mieloma Múltiplo/tratamento farmacológico , Fosforilação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
8.
Biochem Pharmacol ; 188: 114575, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33887260

RESUMO

Hyperactivation of signal transducer and activator of transcription 3 (STAT3) is strongly associated with cancer initiation, progression, metastasis, chemoresistance, and immune evasion; thus, STAT3 has been intensely studied as a therapeutic target for cancer treatment. Berberine (BBR), an active component extracted from Coptis chinensis, has shown anti-tumor effects in multiple tumors. However, its underlying mechanisms have not yet been fully elucidated. In this study, we investigated the effects and the underlying mechanisms of BBR on bladder cancer (BCa) cells. We found that BBR showed significant cytotoxic effects against BCa cell lines both in vivo and in vitro, with much lower cytotoxic effects on the human normal urothelial cell line SV-HUC-1. BBR treatment induced DNA replication defects and cell cycle arrest, resulting in apoptosis or cell senescence, depending on p53 status, in BCa cells. Mechanistically, BBR exerted anti-tumor effects on BCa cells by inhibiting Janus kinase 1 (JAK1)-STAT3 signaling through the upregulation of miR-17-5p, which directly binds to the 3'UTR of JAK1 and STAT3, downregulating their expressions. Collectively, our results demonstrate that BBR exerts anti-tumor effects by perturbing JAK1-STAT3 signaling through the upregulation of miR-17-5p in BCa cells, and that BBR may serve as a potential therapeutic option for BCa treatment.


Assuntos
Berberina/farmacologia , Proliferação de Células/fisiologia , Janus Quinase 1/metabolismo , MicroRNAs/biossíntese , Fator de Transcrição STAT3/metabolismo , Neoplasias da Bexiga Urinária/metabolismo , Animais , Berberina/uso terapêutico , Linhagem Celular Transformada , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Janus Quinase 1/antagonistas & inibidores , Janus Quinase 1/genética , Camundongos , Camundongos Endogâmicos BALB C , MicroRNAs/genética , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologia , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/genética , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
9.
J Cell Mol Med ; 25(9): 4455-4465, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33797200

RESUMO

Myocardial infarction requires urgent reperfusion to salvage viable heart tissue. However, reperfusion increases infarct size further by promoting mitochondrial damage in cardiomyocytes. Exosomes from a wide range of different cell sources have been shown to activate cardioprotective pathways in cardiomyocytes, thereby reducing infarct size. Yet, it is currently challenging to obtain highly pure exosomes in quantities enough for clinical studies. To overcome this problem, we used exosomes isolated from CTX0E03 neuronal stem cells, which are genetically stable, conditionally inducible and can be produced on an industrial scale. However, it is unknown whether exosomes from neuronal stem cells may reduce cardiac ischaemia/reperfusion injury. In this study, we demonstrate that exosomes from differentiating CTX0E03 cells can reduce infarct size in mice. In an in vitro assay, these exosomes delayed cardiomyocyte mitochondrial permeability transition pore opening, which is responsible for cardiomyocyte death after reperfusion. The mechanism of MPTP inhibition was via gp130 signalling and the downstream JAK/STAT pathway. Our results support previous findings that exosomes from non-cardiomyocyte-related cells produce exosomes capable of protecting cardiomyocytes from myocardial infarction. We anticipate our findings may encourage scientists to use exosomes obtained from reproducible clinical-grade stocks of cells for their ischaemia/reperfusion studies.


Assuntos
Receptor gp130 de Citocina/metabolismo , Exossomos/fisiologia , Janus Quinase 1/metabolismo , Janus Quinase 2/metabolismo , Infarto do Miocárdio/prevenção & controle , Traumatismo por Reperfusão Miocárdica/complicações , Células-Tronco Neurais/fisiologia , Substâncias Protetoras/administração & dosagem , Animais , Receptor gp130 de Citocina/genética , Regulação da Expressão Gênica , Janus Quinase 1/genética , Janus Quinase 2/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/etiologia , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Células-Tronco Neurais/citologia
10.
Sci Immunol ; 6(58)2021 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-33827897

RESUMO

Patients with coronavirus disease 2019 (COVID-19) present a wide range of acute clinical manifestations affecting the lungs, liver, kidneys and gut. Angiotensin converting enzyme (ACE) 2, the best-characterized entry receptor for the disease-causing virus SARS-CoV-2, is highly expressed in the aforementioned tissues. However, the pathways that underlie the disease are still poorly understood. Here, we unexpectedly found that the complement system was one of the intracellular pathways most highly induced by SARS-CoV-2 infection in lung epithelial cells. Infection of respiratory epithelial cells with SARS-CoV-2 generated activated complement component C3a and could be blocked by a cell-permeable inhibitor of complement factor B (CFBi), indicating the presence of an inducible cell-intrinsic C3 convertase in respiratory epithelial cells. Within cells of the bronchoalveolar lavage of patients, distinct signatures of complement activation in myeloid, lymphoid and epithelial cells tracked with disease severity. Genes induced by SARS-CoV-2 and the drugs that could normalize these genes both implicated the interferon-JAK1/2-STAT1 signaling system and NF-κB as the main drivers of their expression. Ruxolitinib, a JAK1/2 inhibitor, normalized interferon signature genes and all complement gene transcripts induced by SARS-CoV-2 in lung epithelial cell lines, but did not affect NF-κB-regulated genes. Ruxolitinib, alone or in combination with the antiviral remdesivir, inhibited C3a protein produced by infected cells. Together, we postulate that combination therapy with JAK inhibitors and drugs that normalize NF-κB-signaling could potentially have clinical application for severe COVID-19.


Assuntos
COVID-19/metabolismo , Ativação do Complemento , Células Epiteliais/metabolismo , Janus Quinase 1/metabolismo , Janus Quinase 2/metabolismo , Pulmão/metabolismo , Sistema de Sinalização das MAP Quinases , SARS-CoV-2/metabolismo , COVID-19/patologia , Linhagem Celular Tumoral , Complemento C3a/metabolismo , Fator B do Complemento/metabolismo , Células Epiteliais/patologia , Humanos , Pulmão/patologia
11.
Exp Hematol ; 98: 36-46.e2, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33811972

RESUMO

The objective of this study was to examine the therapeutic effect of ruxolitinib, an orally administered selective Janus kinase (JAK) 1/2 inhibitor, on chronic graft-versus-host disease (cGVHD) using a murine model of sclerodermatous GVHD (scl-GVHD). Compared with scl-GVHD controls, ruxolitinib-treated recipients had scl-GVHD of significantly attenuated clinical and pathological severity in the skin and decreased frequencies of effector cells, CD4+ T cells, and CD11b+ macrophage/monocytes. Regulatory CD4+ Foxp3+ T cells were expanded whereas interferon-γ (IFN-γ)-producing CD4+ T cells were significantly decreased in ruxolitinib-treated recipients. Ruxolitinib suppressed not only the production of IFN-γ from CD4+ T cells and monocyte chemoattractant protein 1 (MCP-1) from CD11b+ macrophage/monocytes, but also the proliferation of these cells in vitro. Levels of both cytokines (IFN-γ and MCP-1) were also reduced in the spleen and skin of ruxolitinib-treated recipients in vivo. IFN-γ-induced MCP-1 production and migration of RAW 264.7 cells, a macrophage cell line, were inhibited by ruxolitinib. However, supplementation with MCP-1 restored this effect of ruxolitinib. In addition, blocking JAK-STAT signaling using ruxolitinib reduced the activation of STAT1 in stimulated immune effector cells. Taken together, these results suggest that ruxolitinib can prevent scl-GVHD by suppressing IFN-γ produced by T cells and MCP-1 expression in macrophage/monocytes via inhibition of JAK-STAT signaling.


Assuntos
Doença Enxerto-Hospedeiro/tratamento farmacológico , Janus Quinase 1/antagonistas & inibidores , Janus Quinase 2/antagonistas & inibidores , Pirazóis/farmacologia , Animais , Doença Crônica , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Feminino , Doença Enxerto-Hospedeiro/enzimologia , Janus Quinase 1/metabolismo , Janus Quinase 2/genética , Camundongos , Camundongos Endogâmicos BALB C
12.
Cell Biochem Funct ; 39(4): 555-561, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33724512

RESUMO

Alzheimer's disease (AD), the most prevalent dementia, is identified as a neurodegenerative disease arising from a degenerative disturbance in the central nervous system. A previous study reported that TTP488 could ameliorate symptoms in patients with mild AD, but the underlying mechanisms need to be studied further. Therefore, the objective of this study was to explore the role of TTP488 in the development of an AD cell model. Administration of TTP448 in an AD cell model reduced the expression of pro-inflammatory cytokines [interleukin (IL)-1ß, IL-6, and TNF-α], reversed the inhibitory role of Aß on cell proliferation and viability, and decreased Aß-triggered cell apoptosis and reactive oxygen species (ROS) production. Furthermore, Aß treatment induced activation of JAK1/STAT3/NFκB/IRF3 pathway as well as NLRP3 expression, and TTP488 administration partially reversed the activation of this pathway and NLRP3 expression. Use of WP1160, a STAT3 agonist, re-activated the downstream STAT3/NFκB/IRF3 pathway and NLRP3 expression. Moreover, we found that WP1160 counteracted the role of TTP488 in Aß-induced SH-SY5Y cells' viability, inflammation, apoptosis, and ROS production. SIGNIFICANCE OF THE STUDY: This study explores the role of TTP488 in the development of an Alzheimer's disease (AD) cell model and confirms that TTP488 administration notably promotes cell proliferation and reduces apoptosis, inflammatory factor expression, and reactive oxygen species generation. Further, this study suggests that the NLRP3-relevant JAK1/STAT3/P65/IRF3 signalling pathway is related to AD pathogenesis.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Apoptose/efeitos dos fármacos , Imidazóis/farmacologia , Inflamação/tratamento farmacológico , Fator Regulador 3 de Interferon/antagonistas & inibidores , Modelos Biológicos , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/antagonistas & inibidores , Peptídeos beta-Amiloides/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Humanos , Imidazóis/administração & dosagem , Inflamação/metabolismo , Inflamação/patologia , Fator Regulador 3 de Interferon/metabolismo , Janus Quinase 1/antagonistas & inibidores , Janus Quinase 1/metabolismo , NF-kappa B/antagonistas & inibidores , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/metabolismo
13.
Mol Immunol ; 133: 163-172, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33667986

RESUMO

Forkhead box P3 (Foxp3) expressing CD4+CD25+ regulatory T cells (Tregs), an essential subset of immune T cells for maintaining immune homeostasis is implicated as a negative regulator in an anti-tumor immune response. Current researches suggest that reducing tumor-infiltrating Tregs contribute to enhanced anti-cancer effect. However, the mechanism of infiltration of a large number of Tregs into tumor tissues is still unclear. In this study, human induced Tregs (iTregs) were co-cultured with human hepatocytes and various types of cancer cells (HepG2, NSCLC, and AsPC-1) supernatants. Foxp3, multiple cytokines, levels of apoptosis and suppressive ability of iTregs were detected by FACS. Western blot was employed to test of proteins. Impact of HepG2 supernatants on T cell subpopulations differentiation, cytokines in supernatants were examed by FACS and ELISA respectively. Anti-IL-10R antibody and JAK1 inhibitor were used to reconfirm the role of tumor-derived IL-10 play in the regulation on iTregs. Hepatocarcinoma cells (HCC) supernatants treatment increases Foxp3 stability and reduces apoptosis level in human iTregs without influencing its differentiation trend. Furthermore, IL-10 was found to be extremely higher in HCC supernatants than other groups, IL-10R blockade neutralize the effect of HCC supernatants on iTregs in vitro obviously. HCC supernatants also reversed IL-1ß/6 triggered decline on Foxp3 which may be related to higher expression of JAK1 and elevated phosphorylation level of STAT5 induced by IL-10. Our results suggest that improved stability and abnormal accumulation of Tregs in tumor microenvironment is IL-10/JAK1/STAT5 signal pathway-dependent and provide a novel approach for improving the efficiency of anti-tumor immunotherapy.


Assuntos
Interleucina-10/imunologia , Janus Quinase 1/metabolismo , Neoplasias Hepáticas/patologia , Fator de Transcrição STAT5/metabolismo , Linfócitos T Reguladores/imunologia , Microambiente Tumoral/imunologia , Proteínas Supressoras de Tumor/metabolismo , Adenocarcinoma/patologia , Adenoma/patologia , Apoptose/imunologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Técnicas de Cocultura , Meios de Cultivo Condicionados/química , Fatores de Transcrição Forkhead/análise , Células Hep G2 , Hepatócitos/metabolismo , Humanos , Imunoterapia/métodos , Interleucina-10/análise , Janus Quinase 1/antagonistas & inibidores , Neoplasias Pulmonares/patologia , Neoplasias Pancreáticas/patologia , Receptores de Interleucina-10/antagonistas & inibidores
14.
Drug Des Devel Ther ; 15: 1135-1147, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33731985

RESUMO

Atopic dermatitis (AD) is a chronic, pruritic, inflammatory skin disease that predominantly affects children. However, it can persist in adulthood and/or start at older ages. Both dysfunction of the epidermal barrier and immune dysregulation are known to play a role in the pathogenesis of AD. In the last years, numerous studies showed that Janus kinase (JAK) enzymes have a key role in AD pathogenesis. Therefore, oral and topical JAK inhibitors are new emerging treatments for AD. We report the data relating to abrocitinib, an oral JAK1 inhibitor. For this purpose, we examined articles already published concerning, in particular, concluded clinical trials. Furthermore, we also report the design of current ongoing clinical trials. The search was carried out considering the main search engines relating to medical literature and clinical trials. From all the data we collected, abrocitinib proved to be an effective drug in significantly reducing the severity of moderate-to-severe AD when compared to placebo. Furthermore, the efficacy was similar to other well-established treatment for AD, such as dupilumab. Adverse events were generally mild; indeed, the drug was definitively suspended only in few patients.


Assuntos
Dermatite Atópica/tratamento farmacológico , Janus Quinase 1/antagonistas & inibidores , Inibidores de Janus Quinases/farmacologia , Pirimidinas/farmacologia , Sulfonamidas/farmacologia , Ensaios Clínicos como Assunto , Dermatite Atópica/metabolismo , Humanos , Janus Quinase 1/metabolismo , Inibidores de Janus Quinases/química , Pirimidinas/química , Sulfonamidas/química
15.
Cancer Res ; 81(8): 2171-2183, 2021 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-33558334

RESUMO

Ewing sarcoma is the second most common pediatric bone cancer, with a 5-year survival rate for metastatic disease of only 20%. Recent work indicates that survival is strongly correlated with high levels of tumor-infiltrating lymphocytes (TIL), whose abundance is associated with IFN-inducible chemokines CXCL10 and CCL5. However, the tumor-intrinsic factors that drive chemokine production and TIL recruitment have not been fully elucidated. We previously showed that ubiquitin-specific protease 6 (USP6) directly deubiquitinates and stabilizes Jak1, thereby inducing an IFN signature in Ewing sarcoma cells. Here, we show that this gene set comprises chemokines associated with immunostimulatory, antitumorigenic functions, including CXCL10 and CCL5. USP6 synergistically enhanced chemokine production in response to exogenous IFN by inducing surface upregulation of IFNAR1 and IFNGR1. USP6-expressing Ewing sarcoma cells stimulated migration of primary human monocytes and T lymphocytes and triggered activation of natural killer (NK) cells in vitro. USP6 inhibited Ewing sarcoma xenograft growth in nude but not NSG mice and was accompanied by increased intratumoral chemokine production and infiltration and activation of NK cells, dendritic cells, and macrophages, consistent with a requirement for innate immune cells in mediating the antitumorigenic effects of USP6. High USP6 expression in patients with Ewing sarcoma was associated with chemokine production, immune infiltration, and improved survival. This work reveals a previously unrecognized tumor-suppressive function for USP6, which engenders an immunostimulatory microenvironment through pleiotropic effects on multiple immune lineages. This further raises the possibility that USP6 activity may be harnessed to create a "hot" tumor microenvironment in immunotherapy. SIGNIFICANCE: This study reveals a novel tumor-suppressive function for USP6 by inducing an immunostimulatory microenvironment, suggesting that USP6 activity may be exploited to enhance immunotherapy regimens.


Assuntos
Neoplasias Ósseas/genética , Linfócitos do Interstício Tumoral , Sarcoma de Ewing/genética , Proteínas Supressoras de Tumor/fisiologia , Ubiquitina Tiolesterase/fisiologia , Animais , Neoplasias Ósseas/imunologia , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/mortalidade , Movimento Celular/efeitos dos fármacos , Quimiocina CCL5/biossíntese , Quimiocina CXCL10/biossíntese , Células Dendríticas/efeitos dos fármacos , Humanos , Imunoterapia , Interferons/farmacologia , Janus Quinase 1/metabolismo , Células Matadoras Naturais/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Nus , Transplante de Neoplasias , Receptor de Interferon alfa e beta/metabolismo , Receptores de Interferon/metabolismo , Sarcoma de Ewing/imunologia , Sarcoma de Ewing/metabolismo , Sarcoma de Ewing/mortalidade , Microambiente Tumoral/imunologia , Ubiquitina Tiolesterase/imunologia , Ubiquitina Tiolesterase/metabolismo , Regulação para Cima/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Exp Cell Res ; 400(1): 112517, 2021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33582094

RESUMO

To investigate mechanisms that TMEM2 activation inhibits hepatitis B virus (HBV) infection in hepatocarcinoma (HCC) cells, co-immunoprecipitation (Co-IP) and mass spectrometry were used in screening interacting proteins for TMEM2. Levels of casein kinase 2 subunit α3 (CSNK2A3) in HCC cells were found to be inhibited or overexpressed using siRNAs and pcDNA3.1-CSNK2A3, respectively. Effect of CSNK2A3 expression on cell proliferation was analyzed using MTS, while its effect on HBV infection was measured using ddPCR and IHC. Western blotting and JAK inhibitor ruxolitinib were also used to determine whether TMEM2-regulated CSNK2A3 expression and HBV infection were affected by JAK-STAT signaling. Co-IP and mass spectrometry results showed that CSNK2A3 interacts with TMEM2. Moreover, overexpression of CSNK2A3 significantly inhibited cell proliferation, while inhibition of CSNK2A3 promoted proliferation of HCC cells. In addition, overexpression of CSNK2A3 was observed to significantly enhance HBV infection, while siRNA knockdown of CSNK2A3 inhibited HBV infection. Notably, effect of CSNK2A3 overexpression on HBV infection was suppressed by TMEM2 overexpression. Further mechanistic analyses have revealed that TMEM2 could antagonize the effects of CSNK2A3 on cell proliferation and HBV infection via JAK-STAT pathway activation. In conclusion, TMEM2 has been determined to bind to CSNK2A3 to inhibit HBV infection via activation of the JAK-STAT pathway.


Assuntos
Caseína Quinase II/metabolismo , Epilepsia do Lobo Temporal/prevenção & controle , Hepatite B/prevenção & controle , Janus Quinase 1/metabolismo , Proteínas de Membrana/metabolismo , Fatores de Transcrição STAT/metabolismo , Encéfalo/metabolismo , Encéfalo/patologia , Carcinoma Hepatocelular/complicações , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/virologia , Estudos de Casos e Controles , Caseína Quinase II/genética , Epilepsia do Lobo Temporal/genética , Epilepsia do Lobo Temporal/metabolismo , Epilepsia do Lobo Temporal/patologia , Hepatite B/metabolismo , Hepatite B/patologia , Hepatite B/virologia , Vírus da Hepatite B/fisiologia , Humanos , Janus Quinase 1/genética , Neoplasias Hepáticas/complicações , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/virologia , Proteínas de Membrana/genética , PPAR gama/genética , PPAR gama/metabolismo , Fatores de Transcrição STAT/genética , Células Tumorais Cultivadas
17.
J Cell Mol Med ; 25(5): 2645-2654, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33523587

RESUMO

Anwulignan is a monomer compound derived from Schisandra sphenanthera lignans. It has been reported to possess a spectrum of pharmacological activities, including anti-bacterial, anti-inflammatory, anticancer and hepatoprotective properties. However, its anticancer capacity and molecular mechanism(s) against non-small cell lung cancer (NSCLC) have not been fully elucidated. Anwulignan significantly inhibited cell growth and increased G1-phase cell cycle arrest in NSCLC cells. Anwulignan strongly attenuates the JAK1/STAT3 signalling pathway by directly targeting JAK1 protein kinase activity in vitro. The anticancer activity by Anwulignan is dependent upon the JAK1 protein expression. Remarkably, Anwulignan strongly inhibited tumour growth in vivo. In conclusion, Anwulignan is a novel JAK1 inhibitor that may have therapeutic implications for NSCLC management.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Janus Quinase 1/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Schisandra/química , Animais , Antineoplásicos Fitogênicos/química , Carcinoma Pulmonar de Células não Pequenas , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Humanos , Janus Quinase 1/genética , Janus Quinase 1/metabolismo , Neoplasias Pulmonares , Camundongos , Inibidores de Proteínas Quinases/química , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Mol Med Rep ; 23(3)2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33495807

RESUMO

In recent decades, the role of microRNAs (miRs) in the development of pneumonia has been reported by a number of researchers. The present study aimed to investigate the role of miR­409­3p in lipopolysaccharide (LPS)­induced human bronchial epithelial cells and the implication for bronchopneumonia. An in vitro inflammation model was established using LPS­induced BEAS­2B cells. Cell apoptosis was determined by flow cytometry. Inflammatory factors were detected by ELISA and reverse transcription­quantitative PCR. Protein levels of Janus kinase 1 (JAK1)/STAT3 and suppressor of cytokine signaling (SOCS)3 were determined by western blotting. Dual­luciferase reporter assay was performed to confirm the interaction between miR­409­3p and SOCS3. LPS treatment significantly increased miR­409­3p expression and decreased the expression levels of SOCS3 in BEAS­2B cells. Dual­luciferase reporter assay demonstrated that miR­409­3p directly targeted and negatively regulated SOCS3. Inhibition of miR­409­3p markedly decreased the levels of TNF­α, IL­6 and IL­1ß, and suppressed apoptosis induced by LPS, which was reversed by SOCS3­knockdown. The inhibition of SOCS3 significantly activated JAK1/STAT3 signaling, as well as enhancing the levels of TNF­α, IL­6 and IL­1ß, and promoting apoptosis, which was reversed by the JAK1 inhibitor Tofacitinib. Suppression of miR­409­3p improved LPS­induced inflammation through SOCS3 in LPS­treated BEAS­2B cells, and this may be caused by regulating JAK1/STAT3 signaling.


Assuntos
Brônquios/metabolismo , Regulação para Baixo/efeitos dos fármacos , Células Epiteliais/metabolismo , Janus Quinase 1/metabolismo , Lipopolissacarídeos/toxicidade , MicroRNAs/biossíntese , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína 3 Supressora da Sinalização de Citocinas/metabolismo , Brônquios/patologia , Linhagem Celular Transformada , Células Epiteliais/patologia , Humanos , Inflamação/induzido quimicamente , Inflamação/metabolismo , Inflamação/patologia
19.
FASEB J ; 35(2): e21302, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33475190

RESUMO

Fibroblast growth factor 23 (FGF23) is a main regulator of mineral homeostasis. Low and high circulating FGF23 levels are associated with bone, renal, cardiovascular diseases, and increased mortality. Understanding the factors and signaling pathways affecting FGF23 levels is crucial for the management of these diseases and their complications. Here, we show that activation of the Jak1/Stat3 signaling pathway leads to inflammation in liver and to an increase in hepatic FGF23 synthesis, a key hormone in mineral metabolism. This increased synthesis leads to massive C-terminal FGF23 circulating levels, the inactive C-terminal fragment, and increased intact FGF23 levels, the active form, resulting in imbalanced production and cleavage. Liver inflammation does not lead to activation of the calcineurin-NFAT pathway, and no signs of systemic inflammation could be observed. Despite the increase of active intact FGF23, excessive C-terminal FGF23 levels block the phosphaturic activity of FGF23. Therefore, kidney function and renal αKlotho expression are normal and no activation of the MAPK pathway was detected. In addition, activation of the Jak1/Stat3 signaling pathway leads to high calcitriol levels and low parathyroid hormone production. Thus, JAK1 is a central regulator of mineral homeostasis. Moreover, this study also shows that in order to assess the impact of high FGF23 levels on disease and kidney function, the source and the balance in FGF23 production and cleavage are critical.


Assuntos
Fatores de Crescimento de Fibroblastos/metabolismo , Inflamação/metabolismo , Janus Quinase 1/metabolismo , Fígado/imunologia , Fígado/metabolismo , Animais , Osso e Ossos/metabolismo , Linhagem Celular , Fatores de Crescimento de Fibroblastos/genética , Células HEK293 , Humanos , Imunoprecipitação , Inflamação/genética , Janus Quinase 1/genética , Rim/metabolismo , Camundongos , Fator de Transcrição STAT3/metabolismo
20.
Biomed Res Int ; 2021: 1419631, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33506010

RESUMO

Lung injury was the common and serious complication of sepsis, a systemic inflammatory response syndrome caused by severe infections. Chinese medicine had unique advantages in attenuating inflammatory response, such as Zuojinfang (ZJF). ZJF was a classical compound herb formula composed of Coptidis Rhizoma and Euodiae Fructus in a ratio of 6 : 1. In this paper, 15 ingredients in ZJF were identified and 8 of them absorbed into rat's serum were quantified by HPLC-MS/MS. Subsequently, sepsis-induced lung injury model was replicated in rats by cecal ligation and puncture. 60 SD rats were randomly divided into 6 groups (n = 10): control group (CON), sham group (Sham), model group (MOD), ZJF low-dose group (ZJF-L), ZJF high-dose group (ZJF-H), and prednisolone group (PNSL). Within the next 24 h, the levels of inflammatory factors, correlation between active ingredients and inflammatory cytokines, the pathological changes of lung tissue, and protein expression of the JAK1/STAT3 signaling pathways were analyzed one by one. Finally, the concentration order of components absorbed in rat serum was berberine > palmatine > jatrorrhizine > coptisine > evodin > chlorogenic acid > evodiamine. Compared with the MOD group, the TNF-α, IL-6, and IFN-γ in the ZJF-H group were significantly reduced (p < 0.05). Moreover, the TNF-α decreased significantly accompanied by the increase of berberine, chlorogenic acid, jatrorrhizine, palmatine, evodin, and evodiamine in serum (negative correlation, p < 0.05). Compared with the MOD, the area of lung injury, the expressions of JAK1, p-JAK1, STAT3, and p-STAT3 were significantly decreased under the treatment of ZJF (p < 0.05). Therefore, downregulating the JAK1/STAT3 signaling pathways was a potential avenue of ZJF in reversing lung injury induced by sepsis.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Medicamentos de Ervas Chinesas/farmacologia , Janus Quinase 1/metabolismo , Fator de Transcrição STAT3/metabolismo , Sepse/patologia , Lesão Pulmonar Aguda/metabolismo , Lesão Pulmonar Aguda/microbiologia , Lesão Pulmonar Aguda/patologia , Animais , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Janus Quinase 1/genética , Masculino , Substâncias Protetoras/farmacologia , Ratos , Ratos Sprague-Dawley , Fator de Transcrição STAT3/genética , Sepse/tratamento farmacológico , Sepse/metabolismo , Transdução de Sinais
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