RESUMO
Janus tyrosine kinase 3 (JAK3) is primarily expressed in immune cells and is needed for signaling by the common gamma chain (γc) family of cytokines. Abnormal JAK3 signal transduction can manifest as hematological disorders, e.g., leukemia, severe combined immunodeficiency (SCID) and autoimmune disease states. While regulatory JAK3 phosphosites have been well studied, here a functional proteomics approach coupling a JAK3 autokinase assay to mass spectrometry revealed ten previously unreported autophosphorylation sites (Y105, Y190, Y238, Y399, Y633, Y637, Y738, Y762, Y824, and Y841). Of interest, Y841 was determined to be evolutionarily conserved across multiple species and JAK family members, suggesting a broader role for this residue. Phospho-substitution mutants confirmed that Y841 is also required for STAT5 tyrosine phosphorylation. The homologous JAK1 residue Y894 elicited a similar response to mutagenesis, indicating the shared importance for this site in JAK family members. Phospho-specific Y841-JAK3 antibodies recognized activated kinase from various T-cell lines and transforming JAK3 mutants. Computational biophysics analysis linked Y841 phosphorylation to enhanced JAK3 JH1 domain stability across pH environments, as well as to facilitated complementary electrostatic JH1 dimer formation. Interestingly, Y841 is not limited to tyrosine kinases, suggesting it represents a conserved ubiquitous enzymatic function that may hold therapeutic potential across multiple kinase families.
Assuntos
Fator de Transcrição STAT5 , Transdução de Sinais , Fosforilação , Fator de Transcrição STAT5/genética , Janus Quinase 1/genética , Processamento de Proteína Pós-Traducional , Tirosina/metabolismoRESUMO
Based on Janus kinase 1/2-signal transducer and activator of transcription 1(JAK1/2-STAT1) signaling pathway, this study explored the immune mechanism of Maxing Shigan Decoction in alleviating the lung tissue and colon tissue damage in mice infected with influenza virus. The influenza virus infection was induced in mice by nasal drip of influenza virus. The normal group, model group, oseltamivir group, antiviral granule group, and Maxing Shigan Decoction group were designed. After intragastric administration of corresponding drugs or normal saline for 3 or 7 days, the body mass was measured, and lung index, spleen index, and thymus index were calculated. Based on hematoxylin-eosin(HE) staining, the pathological changes of lung tissue and colon tissue were observed. Enzyme-linked immunosorbent assay(ELISA) was used to detect serum levels of inflammatory factors interleukin-8(IL-8) and interferon-γ(IFN-γ), Western blot and real-time quantitative polymerase chain reaction(RT-qPCR) to determine the protein and mRNA levels of JAK1, JAK2, STAT1, interferon regulatory factor 9(IRF9), and IFN-γ in lung tissue and colon tissue. The results showed that after 3 and 7 days of administration, the body mass, spleen index, and thymus index were lower(P<0.05 or P<0.01), and the lung index was higher(P<0.01) in the model group than in the normal group. Moreover, the model group showed congestion, edema, and infiltration of a large number of lymphocytes and macrophages in the lung tissue, irregular structure of colon mucosa, ulceration and shedding of epithelial cells, and infiltration of a large number of inflammatory cells. The model group had higher levels of serum IFN-γ(P<0.01), higher protein and mRNA expression of JAK1, JAK2, STAT1, IRF9, IFN-γ in lung tissue(P<0.05 or P<0.01), higher level of JAK2 protein in colon tissue(P<0.01), and higher protein and mRNA levels of STAT1 and IRF9(P<0.05 or P<0.01) than the normal group. Compared with the model group, Maxing Shigan Decoction group had high body mass, spleen index, and thymus index(P<0.05 or P<0.01), low lung index(P<0.05 or P<0.01), and significant alleviation of pathological injury in lung and colon. Moreover, lower serum level of IFN-γ(P<0.05 or P<0.01), protein and mRNA levels of JAK1, JAK2, STAT1, IRF9, and IFN-γ in lung tissue(P<0.05 or P<0.01), JAK2 protein level in colon tissue(P<0.01), and protein and mRNA levels of STAT1 and IRF9(P<0.05 or P<0.01) were observed in the Maxing Shigan Decoction group than in the model group. After 3 days of administration, the level of serum IL-8 in the model group was significantly higher than that in the normal group(P<0.01), and the level in the Maxing Shigan Decoction group was significantly reduced(P<0.01). In conclusion, Maxing Shigan Decoction can significantly up-regulate body mass, spleen index, and thymus index, down-regulate lung index, reduce the levels of IL-8 and IFN-γ, and down-regulate protein and mRNA levels of JAK1, JAK2, STAT1, IRF9, and IFN-γ in lung tissue and protein and mRNA levels of JAK2, STAT1, and IRF9 in colon tissue, and alleviate pathological damage of lung tissue and colon tissue. The mechanism is the likelihood that it inhibits the activation of JAK1/2-STAT1 signaling pathway to alleviate the damage to lung and colon tissue damage.
Assuntos
Influenza Humana , Infecções por Orthomyxoviridae , Orthomyxoviridae , Camundongos , Animais , Humanos , Janus Quinase 1/genética , Fator de Transcrição STAT1/genética , Interleucina-8 , Transdução de Sinais , Interferon gama , Pulmão , RNA Mensageiro , ColoRESUMO
Cervical cancer (CC) is recognized as the most common neoplasm in the female reproductive system worldwide. The lack of chemotherapeutic agents with outstanding effectiveness and safety severely compromises the anti-cipated prognosis of patients. Aloperine (ALO) is a natural quinolizidine alkaloid with marked anti-cancer effects on multiple malignancies as well as favorable activity in relieving inflammation, allergies and infection. However, its therapeutic efficacy and underlying mechanism in CC are still unclear. In the current study, MTT assay was employed to evaluate the viability of HeLa cells exposed to ALO to preliminarily estimate the effectiveness of ALO in CC. Then, the effects of ALO on the proliferation and apoptosis of HeLa cells were further investigated by plate colony formation and flow cytometry, respectively, while the migration and invasion of ALO-treated HeLa cells were evaluated using Transwell assay. Moreover, nude mice were subcutaneously inoculated with HeLa cells to demonstrate the anti-CC properties of ALO in vivo. The molecular mechanisms underlying these effects of ALO were evaluated by Western blot and immunohistochemical analysis. This study experimentally demonstrated that ALO inhibited the proliferation of HeLa cells via G2 phase cell cycle arrest. Simultaneously, ALO promoted an increase in the percentage of apoptotic HeLa cells by increasing the Bax/Bcl-2 ratio. Additionally, the migration and invasion of HeLa cells were attenuated by ALO treatment, which was considered to result from inhibition of epithelial-to-mesenchymal transition. For molecular mechanisms, the expression and activation of the IL-6-JAK1-STAT3 feedback loop were markedly suppressed by ALO treatment. This study indicated that ALO markedly suppresses the proliferation, migration and invasion and enhances the apoptosis of HeLa cells. In addition, these prominent anti-CC properties of ALO are associated with repression of the IL-6-JAK1-STAT3 feedback loop.
Assuntos
Quinolizidinas/farmacologia , Neoplasias do Colo do Útero , Animais , Apoptose , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Retroalimentação , Feminino , Células HeLa , Humanos , Interleucina-6/genética , Janus Quinase 1/genética , Camundongos , Camundongos Nus , Fator de Transcrição STAT3/genética , Transdução de Sinais , Neoplasias do Colo do Útero/tratamento farmacológicoRESUMO
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 fármacosRESUMO
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 SinaisRESUMO
Benign prostatic hyperplasia (BPH) is a widespread disorder in elderly men. Cinnamaldehyde, which is a major constituent in the essential oil of cinnamon, has been previously reported to reduce xanthine oxidase activity, in addition to its anti-inflammatory, anti-oxidant, and anti-proliferative activities. Our study was designed to investigate the potential modulatory effects of cinnamaldehyde on testosterone model of BPH in rats through reduction of uric acid level, and suppression of IL-6/JAK1/STAT3 signaling pathway. Cinnamaldehyde (40 and 75 mg/kg) was orally administered to male Wistar rats for 3 weeks, and concurrently with testosterone (3 mg/kg, s.c.) from the second week. Cinnamaldehyde ameliorated the elevation in prostatic weight and index compared to rats treated with testosterone only, that was also confirmed by alleviation of histopathological changes in prostate architecture. The protective mechanisms of cinnamaldehyde were elucidated through inhibition of xanthine oxidase activity and reduced uric acid level. That was accompanied by reduction of the pro-inflammatory cytokines; interleukin-6 (IL-6), IL-1ß, tumor necrosis factor-alpha (TNF-α), and the nuclear translocation of the transcription factor NF-κB p65, that could be attributed also to the enhanced anti-oxidant defense by cinnamaldehyde. The protein expression of JAK1, which is IL-6 receptor linked protein, was reduced with subsequently reduced activation of STAT3 protein. That eventually suppressed the formation of the proliferation protein cyclin D1, while elevated Bax/Bcl2 ratio. It can be concluded that reducing uric acid level through xanthine oxidase inhibition and suppression of the inflammatory signaling cascade; IL-6/JAK1/STAT3; by cinnamaldehyde could be a novel and promising therapeutic approach against BPH.
Assuntos
Acroleína/análogos & derivados , Interleucina-6/metabolismo , Janus Quinase 1/metabolismo , Hiperplasia Prostática/prevenção & controle , Fator de Transcrição STAT3/metabolismo , Ácido Úrico/sangue , Acroleína/farmacologia , Animais , Biomarcadores/sangue , Proliferação de Células/fisiologia , Ciclina D1/genética , Ciclina D1/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Imuno-Histoquímica , Interleucina-6/genética , Janus Quinase 1/genética , Masculino , Próstata/efeitos dos fármacos , Próstata/enzimologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Fator de Transcrição STAT3/genética , Xantina Oxidase/genética , Xantina Oxidase/metabolismoRESUMO
Background: Cutaneous lupus erythematosus (CLE) is an interferon (IFN) -driven autoimmune skin disease characterized by an extensive cytotoxic lesional inflammation with activation of different innate immune pathways. Aim of our study was to investigate the specific role of Janus kinase 1 (JAK1) activation in this disease and the potential benefit of selective JAK1 inhibitors as targeted therapy in a preclinical CLE model. Methods: Lesional skin of patients with different CLE subtypes and healthy controls (N = 31) were investigated on JAK1 activation and expression of IFN-associated mediators via immunohistochemistry and gene expression analyses. The functional role of JAK1 and efficacy of inhibition was evaluated in vitro using cultured keratinocytes stimulated with endogenous nucleic acids. Results were confirmed in vivo using an established lupus-prone mouse model. Results: Proinflammatory immune pathways, including JAK/STAT signaling, are significantly upregulated within inflamed CLE skin. Here, lesional keratinocytes and dermal immune cells strongly express activated phospho-JAK1. Selective pharmacological JAK1 inhibition significantly reduces the expression of typical proinflammatory mediators such as CXCL chemokines, BLyS, TRAIL, and AIM2 in CLE in vitro models and also improves skin lesions in lupus-prone TREX1-/- -mice markedly. Conclusion: IFN-associated JAK/STAT activation plays a crucial role in the pathophysiology of CLE. Selective inhibition of JAK1 leads to a decrease of cytokine expression, reduced immune activation, and decline of keratinocyte cell death. Topical treatment with a JAK1-specific inhibitor significantly improves CLE-like skin lesions in a lupus-prone TREX1-/- -mouse model and appears to be a promising therapeutic approach for CLE patients.
Assuntos
Azetidinas/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Ácidos Isonicotínicos/uso terapêutico , Janus Quinase 1/antagonistas & inibidores , Lúpus Eritematoso Cutâneo/tratamento farmacológico , Animais , Azetidinas/farmacologia , Linhagem Celular , Citocinas/biossíntese , Citocinas/genética , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Indução Enzimática , Inibidores Enzimáticos/farmacologia , Exodesoxirribonucleases/deficiência , Regulação da Expressão Gênica , Humanos , Ácidos Isonicotínicos/farmacologia , Janus Quinase 1/biossíntese , Janus Quinase 1/genética , Queratinócitos/efeitos dos fármacos , Queratinócitos/enzimologia , Líquen Plano/enzimologia , Lúpus Eritematoso Cutâneo/enzimologia , Lúpus Eritematoso Discoide/enzimologia , Camundongos , Camundongos Endogâmicos C57BL , Modelos Imunológicos , Fosfoproteínas/deficiência , Organismos Livres de Patógenos EspecíficosRESUMO
Icariin (ICA) is obtained from Epimedium brevicornu maxim and exploited to remedy miscellaneous cancers. But the role of ICA in medulloblastoma remains hazy. The research delved into the antitumor activity of ICA in medulloblastoma DAOY cells. ICA with diverse concentrations was utilized to stimulate DAOY cells, and the biological functions of ICA in medulloblastoma DAOY cells were examined. Then, the relative SPARC expression was determined in ICA-managed DAOY cells, and the pc-SPARC vector was transfected into DAOY cells to further probe the influence of SPARC and JAK1/STAT3 and PI3K/AKT pathways in ICA-managed DAOY cells. A xenograft model was established to investigate the function of ICA in vivo. ICA restrained cell viability, expedited apoptosis, prohibited cell migration and invasion, and meanwhile affected the associative factors expression in DAOY cells. Additionally, SPARC expression was declined in ICA-stimulated DAOY cells. Overexpressed SPARC reversed the functions of ICA in above-involved cell behaviors of DAYO cells and the correlative protein levels. Besides, ICA notably frustrated JAK1/STAT3 and PI3K/AKT activations in DAOY cells. Beyond that, ICA prohibited tumor formation in vivo. The results concluded that ICA exhibited the antitumor activity in DAOY cells via decreasing SPARC and inactivating JAK1/STAT3 and PI3K/AKT pathways.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Neoplasias Cerebelares/tratamento farmacológico , Flavonoides/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Meduloblastoma/tratamento farmacológico , Osteonectina/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Humanos , Janus Quinase 1/genética , Janus Quinase 1/metabolismo , Invasividade Neoplásica/prevenção & controle , Proteína Oncogênica v-akt/genética , Proteína Oncogênica v-akt/metabolismo , Osteonectina/genética , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismoRESUMO
The current treatment options for inflammatory bowel disease (IBD) are unsatisfactory. Therefore, novel and safer therapies are needed. We previously reported that koreanaside A (KA) showed high radical scavenging activity and suppressed vascular cell adhesion molecule 1 (VCAM-1) expression in vascular smooth muscle cells. However, the molecular mechanisms involved in its anti-inflammatory effect have not been reported. KA inhibited pro-inflammatory mediators such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), nitric oxide (NO), and prostaglandin E2 (PGE2). KA inhibited the production and mRNA expression of interleukin (IL)-6 and tumor necrosis factor-α (TNF-α) induced by LPS. KA downregulated the myeloid differentiation primary response 88 (MyD88)-dependent inflammatory gene expressions in the MyD88-overexpressed cells. KA suppressed the LPS-induced transcriptional and DNA-binding activities of activator protein-1 (AP-1) and nuclear factor-kappa B (NF-κB). KA was found to inhibit the phosphorylation of Janus kinase 1/2 (JAK1/2) and signal transducers and activators of transcription 1/3 (STAT1/3). In DSS-induced colitis mice, KA relieved the symptoms of colitis by suppressing inflammatory cell infiltration, restoring tight junction (TJ)- and epithelial-mesenchymal transition (EMT)-related protein expression, and inactivating AP-1, NF-κB, and STAT1/3. Therefore, KA reduced inflammatory responses by downregulating AP-1, NF-κB, and JAK/STAT signaling in LPS-induced macrophages and DSS-induced colitis mice.
Assuntos
Anti-Inflamatórios/farmacologia , Colite/tratamento farmacológico , Regulação da Expressão Gênica/efeitos dos fármacos , Glicosídeos/farmacologia , Lignanas/química , Lignanas/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Anti-Inflamatórios/isolamento & purificação , Colite/induzido quimicamente , Colite/metabolismo , Colite/patologia , Sulfato de Dextrana/toxicidade , Flores/química , Forsythia/química , Glicosídeos/isolamento & purificação , Janus Quinase 1/genética , Janus Quinase 1/metabolismo , Lignanas/isolamento & purificação , Lipopolissacarídeos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/genética , NF-kappa B/metabolismo , Células RAW 264.7 , Fatores de Transcrição STAT/genética , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais , Fator de Transcrição AP-1/genética , Fator de Transcrição AP-1/metabolismoRESUMO
The flower of Trollius chinensis Bunge was widely used for the treatment of inflammation-related diseases in traditional Chinese medicine (TCM). In order to clarify the anti-inflammatory mechanism of this Chinese herbs, a comprehensive network pharmacology strategy that consists of three sequential modules (pharmacophore matching, enrichment analysis and molecular docking.) was carried out. As a result, Apoptosis signal-regulating kinase 1 (ASK1), Janus kinase 1 (JAK1), c-Jun N-terminal kinases (JNKs), transforming protein p21 (HRas) and mitogen-activated protein kinase 14 (p38α) that related to the anti-inflammatory effect were filtered out. In further molecular dynamics (MD) simulation, the conformation of CID21578038 and CID20055288 were found stable in the protein ASK1 and JNKs respectively. The current investigation revealed that two effective compounds in the flower of Trollius chinensis Bunge played a crucial role in the process of inflammation by targeting ASK1 and JNKs, the comprehensive strategy can serve as a universal method to guide in illuminating the mechanism of the prescription of traditional Chinese medicine by identifying the pathways or targets.
Assuntos
Flores/química , Regulação da Expressão Gênica/genética , Inflamação/tratamento farmacológico , Ranunculaceae/química , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Biologia Computacional , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/farmacologia , Flavonoides/química , Flavonoides/farmacologia , Flores/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Inflamação/genética , Inflamação/patologia , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Janus Quinase 1/genética , MAP Quinase Quinase Quinase 5/genética , Proteína Quinase 14 Ativada por Mitógeno/genética , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Ranunculaceae/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genéticaRESUMO
Pancreatic cancer is a leading cause of mortality and morbidity worldwide. Due to drug resistance, and the high toxicity and adverse side effects of existing chemotherapeutic drugs, the current treatment of highly aggressive pancreatic cancer is considered inadequate. Allergenremoved Rhus verniciflua Stokes (aRVS) has a strong antiproliferative effect in various cancer cells, and due to its low toxicity, it has emerged as an attractive candidate for cancer treatment. However, the potential use of aRVS as a treatment for pancreatic cancer is relatively unexplored. The present study examined the effects of aRVS on the invasion and migration of pancreatic cancer cells, and identified the molecular mechanisms underlying its anticancer effects. aRVS inhibited the Janus kinase/signal transducer and activator of transcription pathway in pancreatic cancer cells, and decreased the protein expression of mucin 4. In addition, it inhibited the activation of focal adhesion kinase and Src signaling, and decreased the expression of matrix metalloproteinase 9, which may reduce the migration and invasion of pancreatic cancer cells. In conclusion, the present study suggested that aRVS may be a potential treatment for aggressive pancreatic cancer.
Assuntos
Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias Pancreáticas/tratamento farmacológico , Extratos Vegetais/farmacologia , Rhus/química , Alérgenos/efeitos adversos , Alérgenos/química , Alérgenos/genética , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Proteína-Tirosina Quinases de Adesão Focal/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Janus Quinase 1/genética , Metaloproteinase 9 da Matriz/genética , Mucina-4/genética , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Extratos Vegetais/química , Extratos Vegetais/genética , Rhus/efeitos adversos , Rhus/genética , Transdução de Sinais/efeitos dos fármacos , Quinases da Família src/genéticaRESUMO
Macrophage polarization is flexible, and involves in different signaling pathways and various transcription factors. Suppressor of cytokine signaling (SOCS) is an important inhibitor of cytokine signaling pathways and also a key physiological regulator for natural and acquired immunity systems. Following transfection of SOCS1 short hairpin (sh)RNA into mouse macrophage cells, reverse transcriptionquantitative polymerase chain reaction demonstrated that the mRNA levels of Janus kinase (JAK)1 and signal transducer and activator of transcription (STAT)1 increased significantly. In addition, western blotting indicated that JAK1, STAT1 and pSTAT1 expression was significantly enhanced. Fludarabine can inhibit phosphorylation of STAT1 and SOCS1 expression. When fludarabine was added and SOCS1 shRNA was transfected, the inhibition of fludarabine was weakened, and pSTAT1 expression was upregulated. Flow cytometry detection indicated that, following the downregulation of SOCS1 expression, M1type cells significantly increased, but the proportion of M2type cells did not change significantly. Fludarabine can reduce the effect of SOCS1 shRNA on promoting M1type cell polarization, and macrophages can polarize into both M1 and M2 phenotypes. Further ELISA results presented that, when downregulating SOCS1 expression, interleukin (IL)4 and IL10 expression was both downregulated, and tumor necrosis factor (TNF)α and interferon (IFN)γ expression was significantly upregulated. When adding fludarabine or injecting with the traditional Chinese medicine Xuebijing, IL4 and IL10 expression was both significantly upregulated, and TNFα and IFNγ expression was significantly downregulated. When adding fludarabine and downregulating SOCS1, IL4, IL10, TNFα and IFNγ expression presented no significant changes. The above results indicated that, when SOCS1 expression is downregulated, it will activate the JAK1/STAT1 pathway, and thereby promote the polarization of macrophages into M1 type. The findings are of great importance for understanding occurrence, development and treatment of various immunerelated diseases.
Assuntos
Janus Quinase 1/imunologia , Macrófagos Peritoneais/imunologia , Fator de Transcrição STAT1/imunologia , Transdução de Sinais/imunologia , Proteína 1 Supressora da Sinalização de Citocina/imunologia , Animais , Anti-Inflamatórios/farmacologia , Diferenciação Celular , Medicamentos de Ervas Chinesas/farmacologia , Inibidores Enzimáticos/farmacologia , Regulação da Expressão Gênica , Interferon gama/genética , Interferon gama/imunologia , Interleucina-10/genética , Interleucina-10/imunologia , Interleucina-4/genética , Interleucina-4/imunologia , Janus Quinase 1/genética , Macrófagos Peritoneais/citologia , Macrófagos Peritoneais/efeitos dos fármacos , Camundongos , Fosforilação , Cultura Primária de Células , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/imunologia , Fator de Transcrição STAT1/agonistas , Fator de Transcrição STAT1/genética , Proteína 1 Supressora da Sinalização de Citocina/antagonistas & inibidores , Proteína 1 Supressora da Sinalização de Citocina/genética , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Vidarabina/análogos & derivados , Vidarabina/antagonistas & inibidores , Vidarabina/farmacologiaRESUMO
Importance: Castleman disease (CD) is an ultrarare, interleukin-6 (IL-6)-driven lymphoproliferative disorder whose underlying molecular alterations are unknown. Siltuximab (anti-IL-6 antibody) is approved for treatment of this disease. To our knowledge, genomic sequencing of CD has not been reported. Objective: To investigate and identify molecular aberration(s) that help explain the exceptional response to siltuximab in a patient with cutaneous CD. Design, Setting, and Participants: This case study examines data from comprehensive genomic profiling (using targeted next-generation sequencing) of tissue from a patient with cutaneous CD who demonstrated an exceptional response to siltuximab treated at a National Cancer Institute-designated Comprehensive Cancer Center. Interventions: Intravenous siltuximab 12 mg/kg every 3 weeks. Tissue from the patient was interrogated by next-generation sequencing (405 genes). Serum was evaluated for IL-6 levels by enzyme-linked immunoassay. Main Outcomes and Measures: Identification of pretreatment serum IL-6 levels and somatic variants that may explain the exceptional response to siltuximab in this patient with cutaneous CD. Results: Patient pretreatment serum IL-6 levels were normal. Treatment with siltuximab resulted in a complete response lasting 7 years. Next-generation sequencing demonstrated a JAK1V310I missense mutation. Janus Kinase 1 (JAK1) is a crucial signaling component of the IL-6/IL-6 receptor/gp130 machinery. JAK1V310I may induce a conformation change with functional activation effect leading to enhanced sensitivity to the IL-6 ligand. Conclusions and Relevance: Our observations suggest that a JAK1 alteration may explain the underlying biology of a patient's cutaneous CD, as well as the patient's exceptional response to siltuximab.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Hiperplasia do Linfonodo Gigante/tratamento farmacológico , Interleucina-6/imunologia , Janus Quinase 1/genética , Antineoplásicos/uso terapêutico , Hiperplasia do Linfonodo Gigante/genética , Hiperplasia do Linfonodo Gigante/patologia , Ensaio de Imunoadsorção Enzimática , Feminino , Genômica , Humanos , Interleucina-6/sangue , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Fatores de Tempo , Resultado do TratamentoRESUMO
BACKGROUND: Triptolide is a therapeutic diterpenoid derived from the Chinese herb Tripterygium wilfordii Hook f. Triptolide has been shown to induce apoptosis by activation of pro-apoptotic proteins, inhibiting NFkB and c-KIT pathways, suppressing the Jak2 transcription, activating MAPK8/JNK signaling and modulating the heat shock responses. RESULTS: In the present study, we used lymphoblast cell lines (LCLs) derived from 55 unrelated Caucasian subjects to identify genetic markers predictive of cellular sensitivity to triptolide using genome wide association study. Our results identified SNPs on chromosome 2 associated with triptolide IC50 (p < 0.0001). This region included biologically interesting genes as CFLAR, PPIl3, Caspase 8/10, NFkB and STAT6. Identification of a splicing-SNP rs10190751, which regulates CFLAR alternatively spliced isoforms predictive of the triptolide cytotoxicity suggests its role in triptolides action. Our results from functional studies in Panc-1 cell lines further demonstrate potential role of CFLAR in triptolide toxicity. Analysis of gene-expression with cytotoxicity identified JAK1 expression to be a significant predictor of triptolide sensitivity. CONCLUSIONS: Overall out results identified genetic factors associated with triptolide chemo-sensitivity thereby opening up opportunities to better understand its mechanism of action as well as utilize these biomarkers to predict therapeutic response in patients.
Assuntos
Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/genética , Genoma Humano , Estudo de Associação Genômica Ampla , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/antagonistas & inibidores , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Diterpenos/química , Diterpenos/toxicidade , Medicamentos de Ervas Chinesas , Compostos de Epóxi/química , Compostos de Epóxi/toxicidade , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Janus Quinase 1/genética , Janus Quinase 1/metabolismo , Fenantrenos/química , Fenantrenos/toxicidade , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas , Interferência de RNA , Splicing de RNA , RNA Interferente Pequeno/metabolismo , Tripterygium/química , Tripterygium/metabolismoRESUMO
OBJECTIVE: To observe effect of Liuweibuqi Capsule, a Traditional Chinese Medicine (TCM), on the janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway and matrix metalloproteinases (MMPs) in a chronic obstructive pulmonary disease (COPD) rat model with lung deficiency in terms of TCM's pattern differentiation. METHODS: Rats were randomly divided into a normal group, model group, Liuweibuqi group, Jinshuibao group, and spleen aminopeptidase group (n=10). Aside from the normal group, all rats were exposed to smoke plus lipopolysaccharide tracheal instillation to establish the COPD model with lung deficiency. Models were established after 28 days and then the normal and model groups were given normal saline (0.09 g/kg), Liuweibuqi group was given Liuweibuqi capsule (0.35 g/kg), Jinshuibao group was given Jinshuibao capsules (0.495 g/kg), and the spleen group was given spleen aminopeptidase (0.33 mg/kg), once a day for 30 days. Changes in symptoms, signs, and lung histology were observed. Lung function was measured with a spirometer. Serum cytokines were detected using enzyme-linked immunosorbent assay, and changes in the JAK/STAT pathway, MMP-9, and MMPs inhibitor 1 (TIMP1) were detected by immunohistochemistry, RT-PCR, and western blotting, respectively. RESULTS: Compared with the normal group, lung tissue was damaged, and lung function was reduced in the model control group. Additionally, the levels of interleukin (IL)-1beta, gamma interferon (IFN-gamma), and IL-6 were higher, while IL-4 and IL-10 were lower in the model control group than those in the normal group. The expressions of JAK1, STAT3, p-STAT3, and MMP-9 mRNA and protein in lung tissue were higher, and TIMP1 mRNA and protein was lower in the model group compared with the normal group. After treatment, compared with the model group, the expression of inflammatory cytokines was lower in each treatment group, and expressions of JAK/STAT pathway, MMPs were lower. Compared with the positive control groups, the Jinshuibao and spleen aminopeptidase groups, lung function was better, and JAK1, STAT3, and p-STAT3 protein were lower and TIMP1 was higher in the Liuweibuqi group. CONCLUSION: Liuweibuqi capsules can improve the symptoms of COPD possibly by regulating the expression of the JAK1/STAT3 pathway and MMP9/TIMP1.
Assuntos
Medicamentos de Ervas Chinesas/administração & dosagem , Janus Quinase 1/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Fator de Transcrição STAT3/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Animais , Cápsulas/administração & dosagem , Modelos Animais de Doenças , Humanos , Janus Quinase 1/genética , Masculino , Metaloproteinase 9 da Matriz/genética , Doença Pulmonar Obstrutiva Crônica/enzimologia , Doença Pulmonar Obstrutiva Crônica/genética , Doença Pulmonar Obstrutiva Crônica/metabolismo , Ratos , Ratos Sprague-Dawley , Fator de Transcrição STAT3/genética , Transdução de Sinais , Inibidor Tecidual de Metaloproteinase-1/genéticaRESUMO
Host factors required for viral replication are ideal drug targets because they are less likely than viral proteins to mutate under drug-mediated selective pressure. Although genome-wide screens have identified host proteins involved in influenza virus replication, limited mechanistic understanding of how these factors affect influenza has hindered potential drug development. We conducted a systematic analysis to identify and validate host factors that associate with influenza virus proteins and affect viral replication. After identifying over 1,000 host factors that coimmunoprecipitate with specific viral proteins, we generated a network of virus-host protein interactions based on the stage of the viral life cycle affected upon host factor downregulation. Using compounds that inhibit these host factors, we validated several proteins, notably Golgi-specific brefeldin A-resistant guanine nucleotide exchange factor 1 (GBF1) and JAK1, as potential antiviral drug targets. Thus, virus-host interactome screens are powerful strategies to identify targetable host factors and guide antiviral drug development.
Assuntos
Antivirais/farmacologia , Influenza Humana/metabolismo , Orthomyxoviridae/efeitos dos fármacos , Orthomyxoviridae/metabolismo , Mapeamento de Interação de Proteínas/métodos , Mapas de Interação de Proteínas/efeitos dos fármacos , Proteínas Virais/metabolismo , Avaliação Pré-Clínica de Medicamentos , Fatores de Troca do Nucleotídeo Guanina/antagonistas & inibidores , Fatores de Troca do Nucleotídeo Guanina/genética , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Interações Hospedeiro-Patógeno/efeitos dos fármacos , Humanos , Influenza Humana/tratamento farmacológico , Influenza Humana/genética , Influenza Humana/virologia , Janus Quinase 1/antagonistas & inibidores , Janus Quinase 1/genética , Janus Quinase 1/metabolismo , Orthomyxoviridae/genética , Ligação Proteica/efeitos dos fármacos , Proteínas Virais/genéticaRESUMO
Janus activated kinase/signal transducers and activators of transcription (JAK/STATs) pathway are associated with various neuronal functions including cell survival and inflammation. In the present study, it is hypothesized that protective action of aqueous extract of Hippophae rhamnoides in hippocampal neurons against hypoxia is mediated via JAK/STATs. Neuronal cells exposed to hypoxia (0.5% O2) display higher reactive oxygen species with compromised antioxidant status compared to unexposed control cells. Further, these cells had elevated levels of pro-inflammatory cytokines; tumor necrosis factor α and interleukin 6 and nuclear factor κappa B. Moreover, the expression of JAK1 was found to be highly expressed with phosphorylation of STAT3 and STAT5. Cells treated with JAK1, STAT3 and STAT5 specific inhibitors resulted in more cell death compared to hypoxic cells. Treatment of cells with extract prevented oxidative stress and inflammatory response associated with hypoxia. The extract treated cells had more cell survival than hypoxic cells with induction of JAK1 and STAT5b. Cells treated with extract having suppressed JAK1 or STAT3 or STAT5 expression showed reduced cell viability than the cell treated with extract alone. Overall, the findings from these studies indicate that the aqueous extract of Hippophae rhamnoides treatment inhibited hypoxia induced oxidative stress by altering cellular JAK1, STAT3 and STAT5 levels thereby enhancing cellular survival response to hypoxia and provide a basis for possible use of aqueous extract of Hippophae rhamnoides in facilitating tolerance to hypoxia.
Assuntos
Hipocampo/patologia , Hippophae/química , Janus Quinase 1/metabolismo , Neurônios/metabolismo , Oxigênio/farmacologia , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição STAT5/metabolismo , Animais , Antioxidantes/análise , Caspase 3/metabolismo , Hipóxia Celular/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Citocinas/metabolismo , Flavonoides/análise , Mediadores da Inflamação/metabolismo , Janus Quinase 1/genética , L-Lactato Desidrogenase/metabolismo , Camundongos , NF-kappa B/metabolismo , Neurônios/efeitos dos fármacos , Fenóis/análise , Fosforilação , Extratos Vegetais/farmacologia , Substâncias Protetoras/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT5/genética , Transdução de Sinais/efeitos dos fármacosRESUMO
The JAKs receive continued interest as therapeutic targets for autoimmune, inflammatory, and oncological diseases. JAKs play critical roles in the development and biology of the hematopoietic system, as evidenced by mouse and human genetics. JAK1 is critical for the signal transduction of many type I and type II inflammatory cytokine receptors. In a search for JAK small molecule inhibitors, GLPG0634 was identified as a lead compound belonging to a novel class of JAK inhibitors. It displayed a JAK1/JAK2 inhibitor profile in biochemical assays, but subsequent studies in cellular and whole blood assays revealed a selectivity of â¼30-fold for JAK1- over JAK2-dependent signaling. GLPG0634 dose-dependently inhibited Th1 and Th2 differentiation and to a lesser extent the differentiation of Th17 cells in vitro. GLPG0634 was well exposed in rodents upon oral dosing, and exposure levels correlated with repression of Mx2 expression in leukocytes. Oral dosing of GLPG0634 in a therapeutic set-up in a collagen-induced arthritis model in rodents resulted in a significant dose-dependent reduction of the disease progression. Paw swelling, bone and cartilage degradation, and levels of inflammatory cytokines were reduced by GLPG0634 treatment. Efficacy of GLPG0634 in the collagen-induced arthritis models was comparable to the results obtained with etanercept. In conclusion, the JAK1 selective inhibitor GLPG0634 is a promising novel therapeutic with potential for oral treatment of rheumatoid arthritis and possibly other immune-inflammatory diseases.
Assuntos
Inflamação/metabolismo , Janus Quinase 1/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Triazóis/farmacologia , Animais , Diferenciação Celular/efeitos dos fármacos , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Feminino , Inativação Gênica , Humanos , Inflamação/tratamento farmacológico , Concentração Inibidora 50 , Interleucina-6/farmacologia , Janus Quinase 1/genética , Janus Quinase 1/metabolismo , Masculino , Camundongos , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/administração & dosagem , Piridinas/administração & dosagem , Ratos , Fator de Transcrição STAT1/metabolismo , Linfócitos T Auxiliares-Indutores/citologia , Linfócitos T Auxiliares-Indutores/efeitos dos fármacos , Linfócitos T Auxiliares-Indutores/imunologia , Triazóis/administração & dosagemRESUMO
OBJECTIVE: To observe the influence of Sijunzi decoction and Yupingfeng powder on the expression of the relevant DNAs of janus kinase (JAK)-signal transducer and activator of transcription (STAT) signal pathway of the brain in spleen-deficiency model rats. METHODS: Eighty male Wistar rats of sanitary degree were divided randomly into four groups: normal group, model group, treatment group 1, treatment group 2. Besides the rats in the normal group, all the rats in other 3 groups were prepared as spleen deficiency model. The treatment group 1 were treated with Sijunzi decoction and the treatment group 2 were treated with Yupingfeng powder. After treatment for 6 weeks, perfusion was given and the brain was taken for detection of the expression of the relevant DNAs of JAK-STAT signal pathway of the brain in SD rats bygene chip method. RESULTS: Spleen deficiency could lead to increase expression of JAK1, STAT1 and Interleukin 4 (IL-4) in the brain, but the decrease expression of Suppressor of cytokine signaling 1 (SOCS1), prolactin receptor (PRLR) and binding protein 3 (GATA 3). Sijunzi decoction could increase expression of STAT3, Prolactin (PRL) and GATA3, but decrease expression of JAK1, STAT, STAT4, Interleukin 10 receptor, alpha (IL10RA), Coagulation factor II (F2), PRLR, MAD homolog 3 (SMAD3) and IL-4. Yupingfeng powder could decrease expression of JAK1, STAT1, STAT4, SOCS4_ predicted, Epidermal growth factor receptor (EGFR), PRLR, High mobility group AT-hook 1 (HMGA10), IL-4. CONCLUSION: Sijunzi decoction and Yupingfeng powder can improve immune function of the rat through influencing the genetic expression of JAK-STAT signal pathway.
Assuntos
Encéfalo/metabolismo , Medicamentos de Ervas Chinesas/administração & dosagem , Janus Quinase 1/metabolismo , Transdução de Sinais/efeitos dos fármacos , Deficiência da Energia Yin/tratamento farmacológico , Deficiência da Energia Yin/metabolismo , Animais , Encéfalo/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Humanos , Janus Quinase 1/genética , Masculino , Ratos , Ratos Wistar , Fatores de Transcrição STAT/genética , Fatores de Transcrição STAT/metabolismo , Baço/efeitos dos fármacos , Baço/fisiopatologia , Deficiência da Energia Yin/genéticaRESUMO
CRLF2 rearrangements, JAK1/2 point mutations, and JAK2 fusion genes have been identified in Philadelphia chromosome (Ph)-like acute lymphoblastic leukemia (ALL), a recently described subtype of pediatric high-risk B-precursor ALL (B-ALL) which exhibits a gene expression profile similar to Ph-positive ALL and has a poor prognosis. Hyperactive JAK/STAT and PI3K/mammalian target of rapamycin (mTOR) signaling is common in this high-risk subset. We, therefore, investigated the efficacy of the JAK inhibitor ruxolitinib and the mTOR inhibitor rapamycin in xenograft models of 8 pediatric B-ALL cases with and without CRLF2 and JAK genomic lesions. Ruxolitinib treatment yielded significantly lower peripheral blast counts compared with vehicle (P < .05) in 6 of 8 human leukemia xenografts and lower splenic blast counts (P < .05) in 8 of 8 samples. Enhanced responses to ruxolitinib were observed in samples harboring JAK-activating lesions and higher levels of STAT5 phosphorylation. Rapamycin controlled leukemia burden in all 8 B-ALL samples. Survival analysis of 2 representative B-ALL xenografts demonstrated prolonged survival with rapamycin treatment compared with vehicle (P < .01). These data demonstrate preclinical in vivo efficacy of ruxolitinib and rapamycin in this high-risk B-ALL subtype, for which novel treatments are urgently needed, and highlight the therapeutic potential of targeted kinase inhibition in Ph-like ALL.