Expression Of p-JAK2 Research Articles

Inhibition of the SOCS1-JAK2-STAT3 Signaling Pathway Confers Neuroprotection in Rats with Ischemic Stroke

The present study aims to investigate the protective effects of the SOCS1-JAK2-STAT3 signaling pathway on neurons in a rat model of ischemic stroke. Our study was conducted using an ischemic stroke rat model. After the microglia were extracted, 40 neonatal Sprague-Dawley (SD) rats were assigned into the blank, AG490, model and negative control (NC) groups. The neurological function of all the rats was evaluated. Histopathological changes were observed. qRT-PCR and western blotting were applied to measure the expression of genes and proteins in the SOCS1-JAK2-STAT3 signaling pathway and related to apoptosis. The TUNEL assay was conducted to calculate the cellular morphology and apoptosis of neuronal cells. Cell viability was detected using the MTT assay. In addition, immunoassays were used to measure the content of superoxide dismutase (SOD), glutathione (GSH) and malondialdehyde (MDA) as well as the levels of oxidative stress. Compared with the blank group, the model and NC groups showed higher neurological function scores-the cytoplasm of the neurons were cavitated, the organelles were reduced with unclear margins, some of the neurons were necrotic, and apoptosis was increased. In addition, the NC and model groups exhibited decreased cell viability, lower mRNA and protein expression of SOCS1 SOCS3 and bcl-2 and reduced SOD and GSH levels but higher mRNA and protein expression levels of AK2, STAT3,Bax and caspase-3 as well as increased protein expression of P-JAK2, P-STAT3 and activated caspase-3 (c-caspase-3). Moreover, the MDA levels were up-regulated in the NC and model groups. In contrast, opposing trends were found in the AG490 group compared with the NC and model groups. These data demonstrate that inhibiting the SOCS1-JAK2-STAT3 signaling pathway can reduce the loss of nerve function and apoptosis of neuronal cells, which provides a new target for the clinical treatment of ischemic stroke.

Knockdown of KLF11 attenuates hypoxia/reoxygenation injury via JAK2/STAT3 signaling in H9c2.

KLF11 is a Krüppel-like factor (KLF) family member, which plays a central role in cardiac hypertrophy and cerebrovascular protection during ischemic insults. However, the roles of KLF11 in hypoxia/reoxygenation (H/R) injury of rat cardiomyocytes H9c2 have not been elucidated. The aim of this study was to evaluate the effects of KLF11 on H/R injury and investigate the molecular mechanisms involved. Here, we found that KLF11 was increased following H/R and reached the highest level with 24h hypoxia followed by 12h reoxygenation. Moreover, we found that inhibition of KLF11 by small RNA suppressed cell apoptosis, the activity of caspase3, the expression of cleaved-caspase3 and cytochrome C in the cytoplasm and the damage of mitochondrial membrane induced by H/R in H9c2, suggesting that KLF11 silencing protects against H/R injury. In addition, we observed that knockdown of KLF11 elevated the expression of p-JAK2 and p-STAT3 in H9c2, and AG490, a selective inhibitor of JAK2/STAT3 abrogated the potential roles of KLF11 in cell apoptosis and mitochondrial damage. In aggregates, our results showed that depletion of KLF11 protected H9c2 against H/R injury through activating the JAK2/STAT3 signaling pathway, suggesting that KLF11 may be provide therapeutic targets for H/R or other heart diseases.

Protective effect of diosgenin on chondrocytes mediated by JAK2/STAT3 signaling pathway in mice with osteoarthritis

Objective: To investigate the effect of diosgenin (Dgn) on chondrocytes and its relation to JAK2/STAT3 signaling pathway in mice with osteoarthritis (OA).Methods: Fifteen male C57BL/6 mice were randomly divided into three groups:control group, OA group and OA+Dgn group. After 4 weeks of treatment, the histopathological changes of cartilage tissue were observed by toluidine blue staining under light microscopy and the ultrastructure of chondrocytes was observed under electron microscopy. The primarily cultured chondrocytes of OA mice were randomly divided into 4 groups:(1) OA group, (2) Dgn group, (3) Dgn+AG490 group, (4) AG490 group. The expression of p-JAK2, p-STAT3, Bax, succinate dehydrogenase (SDH) and cytochrome c oxidase (COX) were detected by Western blotting, and superoxide dismutase (SOD) was detected using colorimetric method. Results: The morphological observation showed that the chondrocytes of OA group presented considerable pathological changes, while the chondrocytes in OA+Dgn group maintained intact membrane. Electron microscopy observation found obvious injury in cartilage tissues of OA group, while that in OA+Dgn group remained smooth. Compared with OA group, the expressions of p-JAK2 and p-STAT3 in chondrocytes of Dgn group were increased (all P<0.05), and the expressions of Bax protein, SDH, COX and SOD were decreased (all P<0.05). While compared with Dgn group, the expressions of p-JAK2, p-STAT3, SDH, COX and SOD in chondrocytes of Dgn+AG490 group were decreased (all P<0.05), and the expression of Bax protein was increased (P<0.05). Conclusion: Diosgenin can inhibit apoptosis and increase mitochondrial oxidative stress capacity of chondrocytes in mice with osteoarthritis, which is closely related to the activation of JAK2/STAT3 signaling pathway.

Effect of frequent intake of sweeteners on the JAK2/STAT3 signaling pathway in the central nervous system of mice

Sweeteners are food additives that are commonly used worldwide. There are discrepancies in epidemiological studies evaluating their effect on body weight and appetite; this generates the need to further study the mechanism by which sweeteners could affect satiety and energy balance.This study's main objective is to evaluate whether frequent intake of the commercial sweeteners; sucralose, steviol glycosides and sucrose could cause alterations in the leptin‐mediated JAK2/STAT3 signaling pathway, in the central nervous system of mice.For this purpose, three study groups comprised of 14‐week‐old female BALB/c mice were established: sucrose, steviol glycosides and sucralose groups were supplemented with those sweeteners in their daily water for 6 weeks (from week 8 to week 14), additionally, a control group was established without sweetener supplementation. Mice were fed ad libitum and weight was measured weekly. The amount of food and water intake was measured daily. After 6 weeks, body composition of mice was determined using bioelectrical impedance and total brain proteins were obtained to determine the expression of total and phosphorylated JAK2 and STAT3 proteins by western blot.Our preliminary results show that there is a higher consumption of water, but lower food intake in the sucrose group compared with the control and other experimental groups; there are no significant differences in body weight and adiposity among groups. Regarding JAK2/STAT3, expression of pJAK2 and JAK2 was found to be higher in the group supplemented with sucralose, whereas we did not observe changes in the STAT3 proteins. We also found that mice supplemented with steviol glycosides and sucrose had a lower expression of pJAK2 and JAK2 compared to control mice.Our results suggest that frequent intake of commercial sweeteners may cause alterations in energy balance and appetite by altering the phosphorylation of JAK2, however, we need further study the effect of these sweeteners in the leptin‐receptor expression and serum leptin concentrations.

In vivo treatment of rat arterial adventitia with interleukin‑1β induces intimal proliferation via the JAK2/STAT3 signaling pathway.

Previous studies have indicated that adventitial inflammation is involved in the development of atherosclerosis. The aim of this study was to investigate the effect of arterial adventitia inflammation induced by interleukin (IL)-1β on intimal proliferation and the mechanisms involved in this process. The left common carotid artery adventitia of male rats in the experimental and control groups (25 rats/group) was wrapped with agar containing or without a sustained-release suspension of 2.5 µg IL-1β, respectively. Five animals in each group were randomly selected for sacrifice at 2 h, 8 h, 24 h, 48 h, and 1 week post-treatment. Hematoxylin and eosin staining was performed for to analyze the morphology of the adventitia. The expression of janus kinase (JAK)2, signal transducer and activator of transcription (STAT)3, phosphorylated (p-)JAK2 and p-STAT3 were detected by western blot analysis or immunohistochemistry staining. A model of adventitial inflammation was successfully created by wrapping IL-1β around the rat carotid artery. IL-1β treatment induced vascular smooth muscle cell proliferation and migration as well as intimal proliferation. In addition, the expression of p-JAK2 and p-STAT3 increased after IL-1β treatment. Furthermore, an inhibitor of JAK2/STAT3 pathway, AG490, suppressed IL-1β-induced intimal proliferation and phosphorylation of JAK2 and STAT3. Thus, the JAK2/STAT3 signaling pathway is involved in intimal proliferation caused by vascular adventitial inflammation. Inhibiting the JAK2/STAT3 signaling pathway may be a novel method for the clinical treatment of artery atherosclerosis.

JAK2/STAT3 Pathway Mediates Protection of Metallothionein Against Doxorubicin-Induced Cytotoxicity in Mouse Cardiomyocytes.

Doxorubicin (Dox) is one of the most important anticancer agents; however, its clinical application is limited by its severe cardiotoxicity. In our previous study, we found that the gene expression levels of the Janus-activated kinase/signal transducer and activator of transcription 3 (JAK2/STAT3) pathway were different between MT(-/-) cardiomyocytes and MT(+/+) cardiomyocytes when they were treated with Dox. Thus, this study was intended to investigate the role of JAK2/STAT3 pathway in metallothionein (MT) protection of Dox-induced cardiotoxicity. Tyrphostin AG490 (α-cyano-(3,4-dihydroxy)-N-benzylcinnamide) is a synthetic protein tyrosine kinase inhibitor which at first has been considered as a specific JAK2 inhibitor and can inhibit the JAK2/STAT3 signaling pathway. In the present study, AG490 was used to assess the role of JAK2/STAT3 in MT protection against Dox-induced cardiotoxicity. The AG490 can attenuate the MT protection by increasing lactate dehydrogenase and the number of apoptotic cells. Interestingly, pretreated with AG490, MT(-/-) cardiomyocytes were more sensitive than MT(+/+) to Dox-induced cytotoxicity as measured by reactive oxygen species generation, lipid peroxidation, and protein carbonylation. Metallothionein 1 and MT-2 messenger RNA were upregulated by Dox, and AG490 decreased the protein expression of MT-1 and MT-2. After Dox treatment, the protein expression of p-Jak2 and p-Stat3 levels was significantly increased in MT(+/+) cardiomyocytes, suggesting that the JAK2/STAT3 pathway was partially involved in MT protection against Dox-induced cardiotoxicity.

The significance of microRNA-184 on JAK2/STAT3 signaling pathway in the formation mechanism of glioblastoma.

Glioblastoma is a type of glioma with a relatively higher degree of malignancy that may result in severe intracranial hypertension and focal symptoms. Surgery is the preferred treatment modality. Combination therapy including radiotherapy, chemotherapy, gene therapy, immunotherapy and targeted therapy have also been employed. However, due to the invasiveness and pathogenesis of the disease, such treatments do not yield satisfactory outcomes. The aim of the present study was to examine the expression of microRNA (miR)-184 in Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway in the mechanism of glioblastoma formation, thus providing a new basis for the mechanism of glioblastoma induction. The LN18 cell line was employed in the present study. After undergoing thawing, culturing and passaging processes, the cells were divided into the set control group, miR-184 mimic group (transfer miR-184 simulator) and miR-184 group. The expression of miR-184 was detected using quantitative polymerase chain reaction. An MTT assay was used to detect the proliferation ability of glioma cells, and clone formation ability was also detected. The cell scratch and invasion assays were used to identify the cell invasion ability. Western blotting was performed to detect the expression level of p-JAK2 and p-STAT3 proteins. The results showed that compared to the control group, the expression of miR-184 in the miR-184 mimic group increased. Cell proliferation, as well as clone formation and invasion ability were enhanced. The number of cells penetrating septum, as well as the expression of p-JAK2 and p-STAT3 proteins were increased. Differences were statistically significant (P<0.05). By contrast, compared to the control group, the expression of miR-184 in the miR-184 inhibitory group decreased. Cell proliferation, as well as clone formation and invasion ability were reduced. The number of cells penetrating septum, as well as the expression of p-JAK2 and p-STAT3 proteins were reduced. Differences were statistically significant (P<0.05). In conclusion, the results of the present study have shown that miR-184 may be involved in the formation of glioblastoma and influence the expression of JAK2/STAT3 signaling pathway.

Protective effect of Fenofibrate in renal ischemia reperfusion injury: Involved in suppressing kinase 2 (JAK2)/transcription 3 (STAT3)/p53 signaling activation

ObjectiveRenal ischemia reperfusion (I/R) injury is a common reason of acute kidney injury. Apoptosis play an important role in the IRI. Fenofibrate, one of agonist Peroxisome proliferator-activated receptor-alpha (PPARα) has the effect of anti-apoptosis. This study was to explore the effect of Fenofibrate on renal ischemia reperfusion injury and its mechanism. Materials and methodsIRI was induced by bilateral renal ischemia for 45min followed by reperfusion for 24h. Eighteen male C57BL/6 mice were randomly divided into three groups: Sham group (Sham), IRI group (IRI), I/R-Fenofibrate group (FEN). Fenofibrate was injected at 45min before renal ischemia. Renal histology, function, and the expression of Bax, Bcl-2, Bcl-xl p21, p53, Caspse3, CytC, p-JAK2, p-STAT3 and p-PPAR-α were assessed. ResultsFenofibrate precondition can significantly alleviate the renal dysfunction, the pathological change, up-regulate the expression of p-PPAR-α, Bcl-2, Bcl-xl, Caspase3 and down-regulate the expression of p-JAK2, p-STAT3, p53, p21, CytC and Bax induced by renal IR injury. ConclusionFenofibrate precondition can protect mice against IRI by suppressing p53-mediating apoptosis which was associated with inhibiting JAK2/STAT3 signaling activation though further activating PPAR-α. Our findings suggest that Fenofibrate could be useful for preventing IR-induced renal injury.

Nucleophosmin overexpression is associated with poor survival in astrocytoma.

The multiple functions of the protein nucleophosmin (NPM) include the regulation and balance of cell growth, proliferation, and apoptosis. Many cancers have suspected associations with overexpression of NPM or with mutation of the NPM gene. Although NPM and anaplastic lymphoma kinase fusion proteins are known to be related to the Janus Kinase/Signal Transduction and Activator of Transcription (JAK/STAT) signaling pathway, the relationships of NPM, JAK2, and STAT5 to astrocytoma remain unclear. Therefore, this study performed histochemical analyses of expressions of NPM, p-JAK2, and STAT5B proteins in patients with astrocytoma. The results showed that high NPM expression was significantly associated with high tumor grade (p = 0.000), old age (p = 0.000), low Karnofsky Performance Scale (KPS) score (p = 0.000), and tumor recurrence (p = 0.045). High p-JAK2 expression was significantly associated with old age (p = 0.000), high tumor grade (p = 0.000), low KPS score (p = 0.000), and tumor recurrence (p = 0.036). Expression of STAT5B was significantly correlated with tumor grade (p = 0.018) and KPS score (p = 0.002). High expressions of NPM, p-JAK2, and STAT5B were associated with a short survival time (p = 0.035, 0.003 and 0.002, respectively). In multivariable analysis, STAT5B expression was a significant predictor of survival time (p = 0.003). In conclusion, NPM and p-JAK2/STAT5B may have important roles in tumor progression, and STAT5B is an independent prognostic marker of astrocytoma.

Nimesulide inhibits the growth of human esophageal carcinoma cells by inactivating the JAK2/STAT3 pathway

Although selective COX-2 inhibitors have cancer-preventive effects and induce apoptosis, the mechanisms underlying these effects are not fully understood. This study investigated the effects of nimesulide, a selective COX-2 inhibitor, on apoptosis and on the JAK/STAT signaling pathway in Eca-109 human esophageal squamous carcinoma cells. The effects and mechanisms of nimesulide on Eca-109 cell growth were studied in culture and in nude mice with Eca-109 xenografts. Cells were cultured with or without nimesulide and/or the JAK2 inhibitor AG490. Cell proliferation was evaluated using the MTT assay, and apoptosis was investigated. COX-2 mRNA expression was measured using reverse transcription polymerase chain reaction, and protein expression was detected by Western blot analysis, immunohistochemistry, and flow cytometry. Nimesulide significantly inhibited Eca-109 cell viability in vitro in a dose- and time-dependent manner (P<0.05). Nimesulide also induced apoptosis, which was accompanied by a significant decrease in the expression of COX-2 and survivin and an increase in caspase-3 expression. Nimesulide downregulated the phosphorylation levels of JAK2 and STAT3, and JAK2 inhibition by AG490 significantly augmented both nimesulide-induced apoptosis and the downregulation of COX-2 and survivin (P<0.05). In vivo, nimesulide inhibited the growth of Eca-109 tumors and the expression of p-JAK2 and p-STAT3. Thus, nimesulide downregulates COX-2 and survivin expression and upregulates caspase-3 expression in Eca-109 cells, by inactivating the JAK2/STAT3 pathway. These effects may mediate nimesulide-induced apoptosis and growth inhibition in Eca-109 cells in vitro and in vivo.

Prolactin protects cardiomyocytes against intermittent hypoxia-induced cell damage by the modulation of signaling pathways related to cardiac hypertrophy and proliferation

ObjectivesProlactin (PRL) is a multifunctional hormone that influences multiple physiological processes. It has been shown to have a protective effect on the cardiovascular system; however, the mechanisms of this effect are poorly understood. The purpose of the study was to elucidate the role of PRL in intermittent hypoxia (IH)-induced apoptosis in the cardiovascular system. Method and resultsWe established a hyperprolactinemic rat model by implanting two anterior pituitary (AP) glands into the renal capsule of male Sprague–Dawley rats. The rats were kept under normoxic conditions for 4weeks after implantation in order to reach the expression plateau of PRL in the plasma, and then treated with IH for 7 or 14days. Their hearts were then removed for histological and protein expression analyses. Cerebral cortex (CX)-grafted control rats challenged with IH displayed unique phenotypes such as a thicker heart wall, an abnormal myocardial architecture and an increased interstitial space of the left ventricle. They exhibited reduced expressions of p-JAK2, p-STAT5, cell cycle-dependent proteins (cyclin D1, cyclin E and cyclin A), IGF-IRα, PI3Kα, p-AKT and p-ERK1/2 in cardiomyocytes at 7days. ConclusionsOur comprehensive analysis suggested that high plasma PRL can protect rat cardiomyocytes against IH through (1) the p-JAK2 and p-STAT5 pathways for transient cell proliferation, (2) the PI3Kα/AKT and MAPK survival pathways through IGF-I, and (3) the downregulation of IGF-II and ERK5, which inhibit cell hypertrophy.

Evaluation of cardio-protective effect of soybean oligosaccharides

The aim of the present study was to investigate whether soybean oligosaccharides (SO) protects heart function against myocardium ischemia reperfusion (MIR) injury. Hearts were 20min global ischemia and 50min reperfusion. Rats were fed for 30days with saline (sham and MIR groups) or the SO (200 or 400mg/kg body weight, daily). At the end of 30days, the left main coronary artery was occluded for 30min, followed by 24h reperfusion, in anesthetized rats. Sham operated animals were subjected to the same surgical procedures, except that the suture under the left anterior descending coronary artery was not tied. Results showed that SO decreased malondialdehyde (MDA) level and increased antioxidant enzymes activities in the SO-treatment group. Pre-treated with SO it showed a significant recovery in cardiac contractile function, reduction in infarct size, and decrease in creatine kinase (CK), aspartate transaminase (AST) and lactate dehydrogenase (LDH) activities. Moreover, SO also significantly increased the expression of p-JAK2 and p-STAT3 proteins in rat heart. However, no significant change in JAK2 and STAT3 levels was observed. Activation of JAK2/STAT3 pathway showed a significant protective role in the SO-treatment group. Perhaps, the altered activation of the JAK2/STAT3 pathway in ischemic myocardium is one mechanism by which SO is cardioprotective.

Acetylated derivative of glaucine inhibits joint inflammation in collagenase-induced arthritis

Context: Osteoarthritis (OA) has become by far the most common joint disorder. A number of studies using OA animal models have explored the effects of agents that can modulate bone metabolism.Objective: In the present study, we investigated the effect of acetylated derivative of plant alkaloid glaucine (ADG) on experimental OA in mice.Materials and methods: Arthritis was induced by two intraarticular (i.a.) injections of collaganase. Histopathological changes were observed through hematoxylin and eosine (H&E), safranin O and toluidine blue staining. Differentiation of bone marrow (BM) cells was evaluated by tartarate-resistant acid phosphatase (TRAP) assay. The expression of phospho-Janus kinase 2 (pJAK2) and phospho signal transducer and activator of transcription3 (pSTAT3) expression in the joints was determined by immunohistochemistry.Results: We established that ADG significantly decreased cell infiltration (2.32 ± 0.14 versus 1.62 ± 0.13), cartilage loss (2.42 ± 0.12 versus 1.12 ± 0.10) and bone erosion (1.76 ± 0.13 versus 1.04 ± 0.14) in arthritic mice. It appeared that the substance inhibited in a dose-dependent manner osteoclast differentiation in vitro. ADG suppressed the expression of pJAK2 in the joint and partially affected the expression of pSTAT3.Conclusion: Present results suggest that ADG is a suitable candidate for further development as an anti-arthritic agent.

Diosmin protects against cerebral ischemia/reperfusion injury through activating JAK2/STAT3 signal pathway in mice

Background and objectApoptosis is a major form of cell death in cerebral ischemia/reperfusion (I/R) pathogenesis and may represent a target for treatment. Diosmin (DM), a micronized purified flavonoid drug, possesses an anti-apoptotic effect in the treatment of varicose veins and renal injury. However, the effect of DM in the acute phase of cerebral I/R is not clear. This study investigated DM’s role in cerebral I/R and its potential mechanism. MethodsMale CD-1 mice were subjected to transient middle cerebral artery occlusion (tMCAO). Experiment 1 was used to evaluate the time course expression of Janus tyrosine kinase-2 (JAK2), signal transducer and activator of transcription-3 (STAT3), phosphorylated JAK2 (pJAK2) and phosphorylated STAT3 (pSTAT3) after cerebral I/R, and six time points were included. In experiment 2, DM was given orally at doses of 50mg/kg or 100mg/kg for 6 consecutive days before receiving tMCAO. At 24h after reperfusion, neurological deficit, Nissl staining, brain water content and infarct volume were examined. Bcl-2, Bax, pJAK2, and pSTAT3 were detected by immunohistochemistry, qRT-PCR and Western blot. Confocal microscope was used to observe the location of pSTAT3 in the cerebral cortex. ResultsCompared with Vehicle group, the high dose of DM significantly alleviated neurological deficit, brain water content, infarct volume, increased the Nissl-positive cells, upregulated the expression of pJAK2, pSTAT3 and Bcl-2 and downregulated Bax (P<0.05). ConclusionThese results showed that DM protected against cerebral I/R injury through activating JAK2/STAT3 signal pathway.

Plumbagin Inhibits Leptin-Induced Proliferation of Hepatic Stellate Cells via JAK2-STAT3 Pathway to Protect against Hepatic Fibrosis

Purpose: To investigate the protective effects of plumbagin against liver fibrosis and explore the influence of plumbagin on the proliferation of hepatic stellate cells (HSCs).Methods: HSC-LX2 cells were divided into blank/control group, 100 ng/ml leptin group, 100 ng/ml leptin + 2 μmol/L plumbagin group, 100 ng/ml leptin + 8 μmol/L plumbagin group and 100 ng/ml leptin + 6.25 μg/ml colchicines group. The expressions of leptin receptor protein (OB-Rb), p-JAK2, p-STAT3, JAK2, STAT3, p-ERK1/2, ERK1/2 and MMP-1 were detected by Western blot assay. The content of type I collagen in the supernatant of HSCs was also measured after stimulation by leptin.Results: Leptin induced OB-Rb expression in HSCs, which reached a peak level at 24 h (p &lt; 0.01). Leptininduced OB-Rb expression was significantly inhibited by plumbagin at concentrations of 2 μmol/L and 8 μmol/L, respectively. Western blot assay revealed that plumbagin significantly decreased pJAK2 expression in leptin-treated HSCs (p &lt; 0.01). Leptin induced expression of pSTAT3 significantly decreased after plumbagin treatment in HSC-LX2 (p &lt; 0.01). p-ERK1/2 expression markedly decreased in plumbagin-treated HSCs (p &lt; 0.01). Plumbagin significantly increased MMP-1 expression in leptin-treated HSCs (p &lt; 0.01).Conclusion: Plumbagin has an anti-fibrotic effect and may decrease the protein expressions of components in JAK2-STAT3 pathway to inhibit HSC proliferation. Thus, plumbagin may be useful in the clinical prevention and treatment of liver fibrosis.Keywords：Plumbagin, Leptin, Hepatic stellate cell, Proliferation, Signal transduction pathway, Antiinflammatory

Da-Huang-Fu-Zi-Tang attenuates liver injury in rats with severe acute pancreatitis

Ethnopharmacological relevanceDa-Huang-Fu-Zi-Tang (DHFZT) is a famous traditional Chinese prescription with strong anti-inflammatory effects. Our previous work found that DHFZT could act against pancreatic injury in rats with severe acute pancreatitis (SAP) via inhibiting the Janus kinase 2/signal transducers and activators of transcription 3 (JAK2/STAT3) signaling pathway in pancreatic tissues. Aim of the studyTo investigate the therapeutic effects of DHFZT on liver injury in SAP rats, and the effects on JAK2/STAT3 signaling in liver tissue and Kupffer cells (KCs). Materials and methodsFifty SD male rats were randomly divided into five groups: sham operation group (SO), SAP model group, DHFZT treatment groups (12, 24, and 48mg/kg body weight). The model of SAP was constructed by injecting sodium taurocholate (3.5%) into pancreatic and biliary ducts. One hour before constructing the model, DHFZT was perfused into the stomach. All rats were sacrificed after 24h following the operation; livers were examined with hematoxylin and eosin staining. The protein expression of pJAK2 and pSTAT3 in liver tissue was detected by immunohistochemical staining. The activity of ALT, IL-6 and TNF-α in serum was detected. KCs of each group were isolated. After culture for 4h, the protein expression of JAK2, pJAK2, STAT3 and pSTAT3, the mRNA expression of IL-6 and TNF-α in KCs were examined. ResultsSodium taurocholate induced liver injury concomitant with increased expression of pJAK2 and pSTAT3 in liver tissue and KCs. Pretreatment with DHFZT significantly attenuated liver injury induced by SAP, and concurrently, effectively lowered the serum ALT level. Furthermore, our studies showed that DHFZT obviously decreased the expression of pJAK2 and pSTAT3 in liver tissue and KCs. ConclusionsDHFZT could ameliorate liver injury in rats with SAP.

Germacrone induces apoptosis in human hepatoma HepG2 cells through inhibition of the JAK2/STAT3 signalling pathway

Previous studies have shown that STAT3 plays a vital role in the genesis and progression of cancer. In this study, we investigated the relationship between the JAK2/STAT3 signalling pathway and germacrone-induced apoptosis in HepG2 cells. HepG2 cells were incubated with germacrone for 24 h, the protein expression of p-STAT3, STAT3, p-JAK2 and JAK2 was detected by Western Blotting, and RT-PCR was used to determine the expression of STAT3, p53, Bcl-2 and Bax at transcriptional levels. Besides that, HepG2 cells were pre-treated with AG490 or IL-6 for 2 h, and then incubated with germacrone for 24 h. The expression of p-JAK2, JAK2, p-STAT3, STAT3, p53, Bax and Bcl-2 was detected by Western blotting. The activity of HepG2 cells was tested by MTT assay. The apoptosis of HepG2 cells and levels of reactive oxygen species (ROS) were flow cytometrically measured. The results showed that germacrone exposure decreased p-STAT3 and p-JAK2 and regulated expression of p53 and Bcl-2 family members at the same time. Moreover, IL-6 enhanced the activation of the JAK2/STAT3 signalling pathway and therefore attenuated the germacrone-induced apoptosis. Suppression of JAK2/STAT3 signalling pathway by AG490, an inhibitor of JAK2, resulted in apoptosis and an increase in ROS in response to germacrone exposure. We therefore conclude that germacrone induces apoptosis through the JAK2/STAT3 signalling pathway.

JAK2‐related pathway induces acquired erlotinib resistance in lung cancer cells harboring an epidermal growth factor receptor‐activating mutation

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors, such as gefitinib and erlotinib, are effective for non-small cell lung cancer with activating EGFR mutations. However, even in patients with an initial dramatic response to such a drug, acquired resistance develops after 6-12 months. A secondary mutation of T790M in EGFR and amplification of the MET gene account for this resistance; however, the mechanism(s) of approximately 30% of acquired resistance cases remain unknown. We established an erlotinib-resistant lung cancer cell line named PC-9/ER3 that harbors an EGFR mutation after continuously exposing PC-9 cells to erlotinib. PC-9/ER3 cells were 136-fold more resistant to erlotinib than the parental cells. Although the PC-9/ER3 cells did not carry the T790M mutation or MET amplification and had similar levels of phosphorylated (p) STAT3, pJAK2 increased in the resistant cells. It was found in the present study that 3-12 h of exposure to erlotinib in both cell lines did not affect pJAK2 expression, but did result in increased pSTAT3 expression. pAkt in PC-9/ER3 cells was less suppressed than in PC-9 cells, although pEGFR and pMAPK were markedly suppressed in both cell lines. The combined treatment of erlotinib plus a JAK2 inhibitor (JSI-124) suppressed pAkt in PC-9/ER3 cells. Similarly, the combination of erlotinib plus JSI-124 or siRNA against JAK2 restored sensitivity to erlotinib in PC-9/ER3 cells. The combination of erlotinib plus JSI-124 was also effective for reducing PC-9/ER3 tumors in a murine xenograft model. Our results suggest that the activation of JAK2 partially accounts for acquired erlotinib resistance.

Therapeutic effect of 7, 3′-dimethoxy hesperetin on adjuvant arthritis in rats through inhibiting JAK2-STAT3 signal pathway

In our previous study, we have demonstrated that 7, 3′-dimethoxy hesperetin (DMHP), an active derivative of hesperidin, showed pro-apoptotic effect on synoviocytes in vitro. The present study was to investigate the potential therapeutic effect of DMHP on adjuvant arthritis (AA) in rat and its possible mechanisms. Freund's complete adjuvant was used to induce AA in rats. DMHP were administered intragastrically once a day from days 12 to 21 after AA induction. Secondary paw swelling, arthritis index, and pathological assessments were observed. IL-6 production in serum and IL-6 mRNA expression in synovium was detected by ELISA and real-time RT-PCR respectively. The expression of mRNA (JAK2, STAT3) and protein (JAK2, p-JAK2, STAT3, p-STAT3) in synovium were determined. We found that DMHP significantly inhibited hind paw swelling and arthritis index, and ameliorated pathological changes of ankle joint in AA rats. DMHP suppressed the level of IL-6 in serum and the expression of IL-6 mRNA in synovium of AA rats in a dose-dependent manner. DMHP apparently decreased mRNA expression of JAK2 and STAT3 as well as protein expression of p-JAK2 and p-STAT3 in the synovium of the AA rats. Correlation analysis indicated that p-JAK2 or p-STAT3 protein expression was highly correlated with joint damage severity. In conclusion, DMHP has a powerful therapeutic effect on AA in rats and its mechanisms might be partly related to inhibiting excessive activation of JAK2-STAT3 pathway.

Prolactin Promotes Hepatocellular Carcinoma through Janus Kinase 2

Hepatocellular carcinoma (HCC) is one human cancer with obvious gender disparity. This study investigated the association of aberrant prolactin levels with HCC risk and the potential impacts on HCC of the prolactin receptor (PRLR)/Janus kinase 2 (JAK2) signaling. Serum prolactin of 63 HCC patients and 162 subjects without HCC was measured by radioimmunoassay. The expressions of PRLR and phosphorylated JAK2 (p-JAK2) in 82 retrospectively collected HCC specimens were evaluated by immunohistochemistry and further incorporated into the survival analysis. The immunoblotting and proliferation assays were used to analyze the effects of PRLR/JAK2 signaling on liver cancer cells with prolactin treatment. Serum prolactin level was significantly higher in HCC patients than in controls. Hepatocellular carcinoma patients with high p-JAK2 expression had a significantly higher postoperative risk than those with low p-JAK2 expression. Moreover, results from the multivariate analysis indicated the prognostic role of p-JAK2 expression with respect to overall survival in HCC patients. In addition, the Kaplan-Meier survival curve showed that high p-JAK2 expression was associated with poor survival in HCC patients with high PRLR expression. The immunoblotting assay showed that prolactin induced the expression of both p-JAK2 and cyclin D1 in Hep-G2 cells. Importantly, the proliferative effects induced by prolactin could be effectively attenuated by adding AG490, a JAK2 inhibitor. Increased circulating prolactin was found in HCC patients and high p-JAK2 expression could predict poor overall survival in those patients expressing high PRLR. In addition, prolactin contributed to the proliferation of liver cancer cells through PRLR/JAK2 signaling.