Alpinetin inhibits neuroinflammation and neuronal apoptosis via targeting the JAK2/STAT3 signaling pathway in spinal cord injury.

BACKGROUND: A growing body of research shows that drug monomers from traditional Chinese herbal medicines have antineuroinflammatory and neuroprotective effects that can significantly improve the recovery of motor function after spinal cord injury (SCI). Here, we explore the role and molecular mechanisms of Alpinetin on activating microglia-mediated neuroinflammation and neuronal apoptosis after SCI. METHODS: Stimulation of microglia with lipopolysaccharide (LPS) to simulate neuroinflammation models in vitro, the effect of Alpinetin on the release of pro-inflammatory mediators in LPS-induced microglia and its mechanism were detected. In addition, a co-culture system of microglia and neuronal cells was constructed to assess the effect of Alpinetin on activating microglia-mediated neuronal apoptosis. Finally, rat spinal cord injury models were used to study the effects on inflammation, neuronal apoptosis, axonal regeneration, and motor function recovery in Alpinetin. RESULTS: Alpinetin inhibits microglia-mediated neuroinflammation and activity of the JAK2/STAT3 pathway. Alpinetin can also reverse activated microglia-mediated reactive oxygen species (ROS) production and decrease of mitochondrial membrane potential (MMP) in PC12 neuronal cells. In addition, in vivo Alpinetin significantly inhibits the inflammatory response and neuronal apoptosis, improves axonal regeneration, and recovery of motor function. CONCLUSION: Alpinetin can be used to treat neurodegenerative diseases and is a novel drug candidate for the treatment of microglia-mediated neuroinflammation.

Curcumin inhibits proliferation and invasion of papillary thyroid carcinoma cells by inhibiting the JAK2 / STAT3 pathway.

PURPOSE: The purpose of this study was to analyze the function of curcumin to suppress the proliferative and invasive abilities of papillary thyroid carcinoma (PTC) through inhibiting the JAK2/STAT3 pathway. METHODS: After treatment of different doses of curcumin in TPC-1 and SW1736 cells, changes in viability, clonality, cell cycle, apoptosis, wound healing and invasion were determined. Western blot analyses were performed to detect protein levels of apoptosis-associated genes, JAK2 and STAT3 in TPC-1 and SW1736 cells treated with different doses of curcumin. RESULTS: Curcumin treatment dose-dependently reduced viability, clonality and metastatic ability in TPC-1 and SW1736 cells. After treatment of 10 µM or 20 µM curcumin, PTC cells were blocked in G2/M phase, and their apoptotic rate increased. Curcumin treatment downregulated Bcl-2 and upregulated Bax in PTC cells. In addition, curcumin treatment downregulated p-JAK2 and p-STAT3 in TPC-1 and SW1736 cells. CONCLUSIONS: Curcumin treatment blocks PTC cells to proliferate and invade via inhibiting the JAK2/STAT3 pathway.

Notoginsenoside R1 counteracts mesenchymal stem cell-evoked oncogenesis and doxorubicin resistance in osteosarcoma cells by blocking IL-6 secretion-induced JAK2/STAT3 signaling.

Tumor microenvironment is a critical participant in the initiation, progression and drug resistance of carcinomas, including osteosarcoma. Notoginsenoside R1 (NGR1) is a proverbial active ingredient of the traditional Chinese medicine Panax notoginseng (PN) and possess undeniable roles in several cancers. Nevertheless, its function in osteosarcoma and tumor microenvironment remains elusive. In the current study, exposure to NGR1 dose-dependently inhibited osteosarcoma cell viability and migration, and induced apoptosis. Furthermore, osteosarcoma cells that were incubated with conditioned medium (CM) from bone marrow mesenchymal stem cells (BMSCs) exhibited greater proliferation, migration capacity and MMP-2 and MMP-9 expression relative to control cells, which was reversed when BMSCs were treated with NGR1. Notably, administration with NGR1 antagonized CM-evoked doxorubicin resistance in osteosarcoma cells by decreasing cell viability and increasing cell apoptosis and caspase-3/9 activity. Mechanically, NGR1 suppressed IL-6 secretion from BMSCs, as well as the subsequent activation of the JAK2/STAT3 signaling in osteosarcoma cells. In addition, blocking the JAK2 pathway by its antagonist AG490 reversed CM-induced osteosarcoma cell proliferation, migration and doxorubicin resistance. Moreover, exogenous supplementation with IL-6 engendered not only the reactivation of the JAK2/STAT3 signaling but also muted NGR1-mediated efficacy against osteosarcoma cell malignancy and doxorubicin resistance. Collectively, NGR1 may directly restrain osteosarcoma cell growth and migration, or indirectly antagonize MSC-evoked malignancy and drug resistance by interdicting IL-6 secretion-evoked activation of the JAK2/STAT3 pathway. Consequently, the current study may highlight a promising therapeutic strategy against osteosarcoma by regulating tumor cells and the tumor microenvironment.

Zeylenone inhibits proliferation and promotes apoptosis in ovarian carcinoma cells via Janus kinase 2 / signal transducers and activators of transcription 3 pathways.

AIM: Ovarian cancer is the fifth common cancer in females. The aim of our study was to determine function of Zeylenone on cell viability and apoptosis of ovarian carcinoma SKOV3 cells. METHODS: Cell viability was measured by Cell counting kit-8 (CCK8) assay; Mitochondrial membrane potential (MMP) and apoptosis were detected by flow cytometry. The mRNA and protein levels of related factors were determined by Real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot, respectively. RESULTS: Cell viability was decreased by Zeylenone in a dose-dependent manner. Zeylenone with concentrations of 2.5, 5 and 10 µmol/L was used to treat ovarian carcinoma SKOV3 cells for 24 h in the following study. The loss of MMP and apoptosis were both significantly increased by Zeylenone. The mRNA and protein levels of cytochrome c (cyto c) and apoptosis inducing factor (AIF) in cytosol were increased by Zeylenone. The mRNA and protein levels of Caspase-3, Fas, Fasl and Bax were increased; while the expression of Bcl-2 was decreased by Zeylenone. The expression of (Janus family of tyrosine kinase) p-JAK and signal transducer and activator of transcription (p-STAT) was decreased significantly by Zeylenone. CONCLUSION: Zeylenone inhibited cell proliferation and promoted apoptosis in ovarian carcinoma cells. The JAK-STAT pathway was involved in this progress.

Cryptotanshinone induces cell cycle arrest and apoptosis through the JAK2/STAT3 and PI3K/Akt/NFκB pathways in cholangiocarcinoma cells.

BACKGROUND: Cholangiocarcinoma (CCA) is the most common biliary tract malignancy in the world with high resistance to current chemotherapies and extremely poor prognosis. The main objective of this study was to investigate the inhibitory effects of cryptotanshinone (CTS), a natural compound isolated from Salvia miltiorrhiza Bunge, on CCA both in vitro and in vivo and to explore the underlying mechanisms of CTS-induced apoptosis and cell cycle arrest. METHODS: The anti-tumor activity of CTS on HCCC-9810 and RBE cells was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay and colony forming assays. Cell cycle changes were detected by flow cytometric analysis. Apoptosis was detected by annexin V/propidium iodide double staining and Hoechst 33342 staining assays. The efficacy of CTS in vivo was evaluated using a HCCC-9810 xenograft model in athymic nude mice. The expression of key proteins involved in cell apoptosis and signaling pathway in vitro was analyzed by Western blot analysis. RESULTS: CTS induced potent growth inhibition, S-phase arrest, apoptosis, and colony-forming inhibition in HCCC-9810 and RBE cells in a dose-dependent manner. Intraperitoneal injection of CTS (0, 10, or 25 mg/kg) for 4 weeks significantly inhibited the growth of HCCC-9810 xenografts in athymic nude mice. CTS treatment induced S-phase arrest with a decrease of cyclin A1 and an increase of cyclin D1 protein level. Bcl-2 expression was downregulated remarkably, while Bax expression was increased after apoptosis occurred. Additionally, the activation of JAK2/STAT3 and PI3K/Akt/NFκB was significantly inhibited in CTS-treated CCA cells. CONCLUSION: CTS induced CCA cell apoptosis by suppressing both the JAK2/STAT3 and PI3K/Akt/NFκB signaling pathways and altering the expression of Bcl-2/Bax family, which was regulated by these two signaling pathways. CTS may serve as a potential therapeutic agent for CCA.

Luhong formula inhibits myocardial fibrosis in a paracrine manner by activating the gp130/JAK2/STAT3 pathway in cardiomyocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Luhong formula (LHF)-a traditional Chinese medicine containing Cervus nippon Temminck, Carthamus tinctorius L., Cinnamomum cassia Presl, Codonopisis pilosula( Franch.) Nannf., Astragalus membranaceus ( Fisch.) Bge. var. mongholicus ( Bge.) Hsiao, Lepidium apetalum Willd-is used in the treatment of heart failure. AIM OF THE STUDY: To investigate the antifibrotic efficacy of LHF in a myocardial infarction-induced rat model of heart failure and to determine its mechanism of action. MATERIAL AND METHODS: Myocardial infarction was induced in rats by coronary artery ligation, and cardiac fibroblasts were isolated. Neonatal rat cardiomyocytes (NRCMs) were isolated from 2 to 3-day-old Sprague-Dawley male rats, and cardiomyocyte hypertrophy was induced by isoprenaline. Histological examination was carried out to estimate the degree of myocardial fibrosis. Expression of gp130/JAK2/STAT3 pathway proteins was measured by western blot. The mRNA levels of downstream genes of gp130/JAK2/STAT3 pathway (i.e., CTGF, TSP-1, and TIMP1) were determined by RT-PCR; while CTGF, TSP-1, and TIMP1 protein levels were measured by ELISA. To investigate paracrine effects, cell proliferation and collagen synthesis was measured after treating cardiac fibroblasts with the conditioned media from isoprenaline-treated NRCMs. RESULTS: Histopathological changes showed that LHF inhibited myocardial fibrosis in heart failure rats. Treatment with LHF up-regulated gp130, JAK2, and STAT3 protein expression in heart tissue, and down-regulated CTGF, TSP-1, and TIMP1 gene expression. Isoprenaline-treated NRCMs displayed lower expression of the gp130, JAK2, and STAT3 pathway proteins and higher secretion of its downstream signaling molecules (CTGF, TSP-1, TIMP1). LHF inhibited cardiac fibroblast proliferation and collagen synthesis after treatment with the conditioned media from isoprenaline-treated NRCMs. CONCLUSION: LHF treatment attenuates myocardial fibrosis in vivo. LHF inhibits cardiac fibroblasts proliferation and collagen synthesis in a paracrine manner by activating the gp130/JAK2/STAT3 pathway in cardiomyocytes, thereby inhibiting the secretion of downstream profibrogenic cytokines.

Farrerol inhibited angiogenesis through Akt/mTOR, Erk and Jak2/Stat3 signal pathway.

BACKGROUND: Farrerol is one of traditional Chinese medicines, isolated from Rhododendron dauricum L. It has been reported that Farrerol exerts multiple biological activities. Angiogenesis is an important drug target for cancer and inflammation therapy, the effect of Farrerol on angiogenesis is unknown. HYPOTHESIS/PURPOSE: We aimed to investigate whether Farrerol may have inhibitory effects against angiogenesis. STUDY DESIGN/METHODS: Two kinds of endothelial cells, named human umbilical vein endothelia cell and human micro vessel endothelial cells, were used to examine the effect and mechanism of Farrerol on angiogenesis. MTT assay was used to detect cell proliferation, wound healing assay and boyden's chamber assay were used to examine cell migration, Matrigel was used as basement membrane substratum in tube formation assay, Annexin V-FITC/PI dual staining assay and trypan blue staining were used to detect cell apoptosis, mouse aortic rings assay was performed as ex vivo assay, the expression of proteins involved in angiogenesis was tested using western blot, the binding of Farrerol to Stat3 was monitored by docking assay, molecular dynamics simulations and MM-GBSA method. RESULTS: Farrerol showed an inhibitory effect on proliferation, migration and tube formation of human umbilical vein endothelia cell and human micro vessel endothelial cells in a concentration-dependent manner. Farrerol induced cell cycle arrest and increased the apoptotic percentage of endothelial cells. Farrerol also suppressed the formation of new micro vessels from mouse aortic rings. Moreover, Farrerol reduced the phosphorylation levels of Erk, Akt, mTOR, Jak2 and Stat3 as well as protein expression of Bcl-2 and Bcl-xl. Docking assay, molecular dynamics simulations and MM-GBSA method showed that Farrerol bound to domain of Stat3, Ser613,Gln635, Glu638 and Thr714 are the main residues in Farrerol binding sites with the binding free energy -7.3 ∼ -9.0kcal/mol. CONCLUSIONS: In this study, we demonstrated that Farrerol inhibited angiogenesis through down regulation of Akt/mTOR, Erk and Jak2/Stat3 signal pathway. The inhibitory effect of Farrerol on angiogenesis suggested that this compound may be helpful to the angiogenesis-related diseases treatment, such as cancer and inflammations.

Anti-inflammatory effects and the underlying mechanisms of action of daidzein in murine macrophages stimulated with Prevotella intermedia lipopolysaccharide.

BACKGROUND AND OBJECTIVE: Host modulatory agents directed at inhibiting specific proinflammatory mediators could be beneficial in terms of attenuating periodontal disease progression and potentially enhancing therapeutic responses. The aim of this study was to investigate whether daidzein could modulate the production inflammatory mediators in macrophages stimulated with lipopolysaccharide (LPS) from Prevotella intermedia, a pathogen implicated in periodontal disease, and to delineate underlying mechanisms of action. MATERIAL AND METHODS: LPS was extracted from P. intermedia ATCC 25611 cells by the standard hot phenol-water method. The amounts of nitric oxide (NO) and interleukin-6 (IL-6) secreted into the culture medium were assayed. A real-time PCR was performed to quantify inducible nitric oxide synthase (iNOS) and IL-6 mRNA expression. We used immunoblot analysis to characterize iNOS protein expression, phosphrylation of c-Jun N-terminal kinase (JNK) and p38, degradation of inhibitory κB-α (IκB-α), nuclear translocation of nuclear factor-κB (NF-κB) subunits and phosphorylation of signal transducer and activator of transcription 1 (STAT1). The DNA-binding activity of NF-κB was assessed by using ELISA-based kits. RESULTS: Daidzein significantly inhibited the production of NO and IL-6, as well as their mRNA expression, in P. intermedia LPS-treated RAW264.7 cells. The JNK and p38 pathways were not involved in the regulation of LPS-induced NO and IL-6 release by daidzein. Daidzein inhibited the degradation of IκB-α induced by P. intermedia LPS. In addition, daidzein suppressed NF-κB transcriptional activity via regulation of the nuclear translocation and DNA-binding activity of NF-κB p50 subunit and blocked STAT1 phosphorylation. CONCLUSION: Although additional studies are required to dissect the molecular mechanism of action, our results suggest that daidzein could be a promising agent for treating inflammatory periodontal disease. Further research in animal models of periodontitis is necessary to better evaluate the potential of daidzein as a novel therapeutic agent to treat periodontal disease.