TWEAK/Fn14 mediates atrial‐derived HL‐1 myocytes hypertrophy via JAK2/STAT3 signalling pathway

Atrial myocyte hypertrophy is one of the most important substrates in the development of atrial fibrillation (AF). The TWEAK/Fn14 axis is a positive regulator of cardiac hypertrophy in cardiomyopathy. This study therefore investigated the effects of Fn14 on atrial hypertrophy and underlying cellular mechanisms using HL‐1 atrial myocytes. In patients with AF, Fn14 protein levels were higher in atrial myocytes from atrial appendages, and expression of TWEAK was increased in peripheral blood mononuclear cells, while TWEAK serum levels were decreased. In vitro, Fn14 expression was up‐regulated in response to TWEAK treatment in HL‐1 atrial myocytes. TWEAK increased the expression of ANP and Troponin T, and Fn14 knockdown counteracted the effect. Inhibition of JAK2, STAT3 by specific siRNA attenuated TWEAK‐induced HL‐1 atrial myocytes hypertrophy. In conclusion, TWEAK/Fn14 axis mediates HL‐1 atrial myocytes hypertrophy partly through activation of the JAK2/STAT3 pathway.     

Inhibition of JAK2/STAT3 signalling induces colorectal cancer cell apoptosis via mitochondrial pathway

Abnormalities in the JAK2/STAT3 pathway are involved in the pathogenesis of colorectal cancer (CRC), including apoptosis. However, the exact mechanism by which dysregulated JAK2/STAT3 signalling contributes to the apoptosis has not been clarified. To investigate the role of both JAK2 and STAT3 in the mechanism underlying CRC apoptosis, we inhibited JAK2 with AG490 and depleted STAT3 with a small interfering RNA. Our data showed that inhibition of JAK2/STAT3 signalling induced CRC cellular apoptosis via modulating the Bcl-2 gene family, promoting the loss of mitochondrial transmembrane potential (Δψm) and the increase of reactive oxygen species. In addition, our results demonstrated that the translocation of cytochrome c (Cyt c), caspase activation and cleavage of poly (ADP-ribose) polymerase (PARP) were present in apoptotic CRC cells after down-regulation of JAK2/STAT3 signalling. Moreover, inhibition of JAK2/STAT3 signalling suppressed CRC xenograft tumour growth. We found that JAK2/STAT3 target genes were decreased; meanwhile caspase cascade was activated in xenograft tumours. Our findings illustrated the biological significance of JAK2/STAT3 signalling in CRC apoptosis, and provided novel evidence that inhibition of JAK2/STAT3 induced apoptosis via the mitochondrial apoptotic pathway. Therefore, JAK2/STAT3 signalling may be a potential target for therapy of CRC.     

IGF2BP3 facilitates cell proliferation and tumorigenesis via modulation of JAK/STAT signalling pathway in human bladder cancer

Insulin‐like growth factor‐2 messenger RNA‐binding protein 3 (IGF2BP3) has been reported to contribute to tumorigenesis in several human cancers. However, the biological functions of IGF2BP3 in bladder cancer are poorly understood. We investigated the relation between IGF2BP3 expression and prognosis of bladder cancer patients. Cell proliferation, cell cycle and cell apoptosis assays were performed to assess IGF2BP3 functions. The results showed that IGF2BP3 was overexpressed in bladder cancer tissues compared with that in normal bladder tissues, and its higher expression was closely correlated with poor prognosis in bladder cancer patients. Overexpression of IGF2BP3 markedly promoted cell proliferation and cell cycle progression and inhibited cell apoptosis, while knockdown of IGF2BP3 notably suppressed the proliferation, promoted cell apoptosis and induced cell cycle arrest at the G0/G1 phase. Mechanistically, we revealed that IGF2BP3 promotes the activation of the JAK/STAT pathway in bladder cancer cells. Moreover, the JAK/STAT inhibitor dramatically blocked the tumour‐promoting activity of IGF2BP3. Tumour growth in vivo was also suppressed by knocking down of IGF2BP3. Hence, IGF2BP3 facilitated bladder cancer cell proliferation by activating the JAK/STAT signalling pathway. These findings suggest that IGF2BP3 exhibits an oncogenic effect in human bladder cancer progression.     

Functional 3‐D cardiac co‐culture model using bioactive chitosan nanofiber scaffolds

The in vitro generation of a three‐dimensional (3‐D) myocardial tissue‐like construct employing cells, biomaterials, and biomolecules is a promising strategy in cardiac tissue regeneration, drug testing, and tissue engineering applications. Despite significant progress in this field, current cardiac tissue models are not yet able to stably maintain functional characteristics of cardiomyocytes for long‐term culture and therapeutic purposes. The objective of this study was to fabricate bioactive 3‐D chitosan nanofiber scaffolds using an electrospinning technique and exploring its potential for long‐term cardiac function in the 3‐D co‐culture model. Chitosan is a natural polysaccharide biomaterial that is biocompatible, biodegradable, non‐toxic, and cost effective. Electrospun chitosan was utilized to provide structural scaffolding characterized by scale and architectural resemblance to the extracellular matrix (ECM) in vivo. The chitosan fibers were coated with fibronectin via adsorption in order to enhance cellular adhesion to the fibers and migration into the interfibrous milieu. Ventricular cardiomyocytes were harvested from neonatal rats and studied in various culture conditions (i.e., mono‐ and co‐cultures) for their viability and function. Cellular morphology and functionality were examined using immunofluorescent staining for alpha‐sarcomeric actin (SM‐actin) and gap junction protein, Connexin‐43 (Cx43). Scanning electron microscopy (SEM) and light microscopy were used to investigate cellular morphology, spatial organization, and contractions. Calcium indicator was used to monitor calcium ion flux of beating cardiomyocytes. The results demonstrate that the chitosan nanofibers retained their cylindrical morphology in long‐term cell cultures and exhibited good cellular attachment and spreading in the presence of adhesion molecule, fibronectin. Cardiomyocyte mono‐cultures resulted in loss of cardiomyocyte polarity and islands of non‐coherent contractions. However, the cardiomyocyte‐fibroblast co‐cultures resulted in polarized cardiomyocyte morphology and retained their morphology and function for long‐term culture. The Cx43 expression in the fibroblast co‐culture was higher than the cardiomyocytes mono‐culture and endothelial cells co‐culture. In addition, fibroblast co‐cultures demonstrated synchronized contractions involving large tissue‐like cellular networks. To our knowledge, this is the first attempt to test chitosan nanofiber scaffolds as a 3‐D cardiac co‐culture model. Our results demonstrate that chitosan nanofibers can serve as a potential scaffold that can retain cardiac structure and function. These studies will provide useful information to develop a strategy that allows us to generate engineered 3‐D cardiac tissue constructs using biocompatible and biodegradable chitosan nanofiber scaffolds for many tissue engineering applications. Biotechnol. Bioeng. 2013; 110: 637–647. © 2012 Wiley Periodicals, Inc.     

The JAK/STAT pathway is essential for opioid-induced cardioprotection: JAK2 as a mediator of STAT3, Akt, and GSK-3 beta

We examined the role for the JAK/STAT signaling pathway in acute opioid-induced cardioprotection (OIC) and whether opioid-induced glycogen synthase kinase-3beta (GSK-3 beta) inhibition is mediated by the JAK/STAT pathway. Rats underwent 30 min of ischemia and either 5 min or 2 h of reperfusion, followed by tissue isolation for molecular analysis or infarct size assessment, respectively. Rats were treated with vehicle, morphine (300 microg/kg), the delta-opioid agonist fentanyl isothiocynate (FIT, 10 microg/kg), or the GSK inhibitor SB-216763 (SB21, 600 microg/kg) 10 min before ischemia. Five minutes before opioid or SB21 treatment, some rats received the putative JAK2 inhibitor AG-490 (3 mg/kg) or the putative JAK3 inhibitor ZM-449829 (3 mg/kg). H9C2 cardiomyoblast cells were also used to investigate FIT-induced signaling (1 microM) in vitro via molecular analysis. Morphine induced the phosphorylation of JAK2, yet not JAK1, in the area at risk. Morphine, FIT, and SB21 also reduced infarct size compared with vehicle (water) when administered before ischemia [43.0 +/- 2.8, 39.1 +/- 3.1, and 42.1 +/- 2.5 (*P < 0.001, respectively) vs. 58.1 +/- 1.3%, respectively]. AG-490 abrogated OIC, whereas ZM-449829 had no effect on OIC. Cardioprotection was afforded by SB21 even in the presence of AG-490. Morphine phosphorylated STAT3, Akt, and GSK-3beta, and phosphorylation was abrogated by AG-490. FIT stimulation of H9C2 cells also caused a time-dependent phosphorylation of STAT3, Akt, and GSK-3beta, and this effect was abrogated by AG-490. STAT3 phosphorylation was also dependent on phosphatidylinositol 3-kinase (PI3K) activation in both tissue and H9C2 cells. These data suggest that OIC occurs via the JAK2 regulation of PI3K pathway-dependent STAT3, Akt, and GSK-3 beta, with GSK-3 beta contributing a central role in acute OIC.