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The extracts from Psoralea corylifolia Linn. (P. corylifolia) seeds have been shown to display antitumor activity. To date, the prospects of this plant and its active compounds in the treatment of non-small-cell lung cancer (NSCLC) have not been thoroughly studied. In this study, we identified a novel psorachromene compound that displays selective cytotoxic effects on all NSCLC cells tested, including NSCLC cells harboring epidermal growth factor receptor (EGFR) activation mutants (H1975L858R/T790M and H1975-MS35L858R/T790M/C797S ). Psorachromene induces G1 arrest in NSCLC cells harboring wild-type EGFR but induces apoptosis in NSCLC cells harboring activating EGFR mutations. Psorachromene inhibits activated EGFR signaling and kinase activity and suppresses tumor growth of implanted H1975-MS35L858R/T790M/C797S cells in nude mice. Molecular docking analysis revealed that psorachromene could form stronger bonds with mutant EGFR than wild-type EGFR, which might account for the greater cytotoxic effects observed in NSCLC cells harboring activating EGFR mutations (H1975 and H1975-MS35) than wild-type EGFR (A549). In conclusion, it is suggested that psorachromene is an attractive agent to be further explored for its use in the treatment of NSCLC patients harboring EGFR L858R/T790M/C797S. [ABSTRACT FROM AUTHOR]

Phosphatidylinositide 3-kinases (PI3Ks) are intracellular signal transducer enzymes that recruit protein kinase B (aka Akt) to the cell membrane, the subsequent activation of which regulates many cellular functions. PI3K/Akt activity is up-regulated within mesocorticolimbic structures in animal models of alcoholism, but less is known regarding PI3K/Akt activity in animal models of cocaine addiction. Given that prefrontal cortex (PFC) is grossly dysregulated in addiction, we studied how cocaine affects protein indices of PFC PI3K/Akt activity in rat and mouse models and examined the relevance of PI3K activity for cocaine-related learning. Immunoblotting of mouse medial PFC at 3 weeks withdrawal from a cocaine-sensitization regimen (seven injections of 30 mg/kg, intraperitoneal [IP]) revealed increased kinase activity, as did immunoblotting of tissue from the ventral PFC of rats with a history of long-access intravenous cocaine self-administration (0.25 mg/0.1 mL infusion; 10 days of 6 h/d cocaine access). Interestingly, increased Akt phosphorylation was observed in rat ventromedial PFC at both 3- and 30-day withdrawal only in animals re-exposed to cocaine-associated cues. A conditioned place-preference paradigm in mice and a cue-elicited drug-seeking test in rats were conducted to determine the functional relevance for elevated PI3K activity for addiction-related behavior. In both cases, an intra-PFC infusion of the PI3K inhibitor wortmannin (50μM) reduced drug-seeking behavior. Taken together, this cross-species, interdisciplinary, study provides convincing evidence that cocaine history produces an enduring increase in PI3K/Akt-dependent signaling within the more ventral aspect of the PFC that is relevant to behavioral reactivity to drug-associated cues/contexts. As such, PI3K inhibitors may well serve as an effective strategy for reducing drug cue reactivity and craving in cocaine addiction. [ABSTRACT FROM AUTHOR]

Background: Transient receptor potential vanilloid-1 (TRPV1), activated by heat, acidic pH, endogenous vanilloids and capsaicin, is essential for thermal hyperalgesia. Under inflammatory conditions, phosphorylation of TRPV1 by protein kinase C (PKC) can sensitize the channel and decrease the activation threshold. Src kinase also phosphorylates TRPV1, promoting channel trafficking to the plasma membrane. These post-translational modifications are important for several chronic pain conditions. This study presents a previously undescribed relationship between Src and PKC phosphorylation of TRPV1, influencing the thermal hypersensitivity associated with TRPV1 activation.Methods: We assessed TRPV1 channel activity using intracellular calcium imaging and patch-clamp electrophysiology in mouse dorsal root ganglion cultures. Additionally, we used behavioural experiments to evaluate plantar thermal sensitivity following intraplantar injections of activators of known modulators of TRPV1 with and without an Src antagonist.Results: Using calcium imaging and patch-clamp techniques, we demonstrated that pharmacological inhibition of Src kinase or mutation of the Src phosphorylation site on TRPV1 prevented PKC but not PKA-mediated sensitization of TRPV1 in vitro. We found that intraplantar injection of the PKC activator phorbol 12-myristate 13-acetate (PMA) or bradykinin produces thermal hypersensitivity that can be attenuated by pharmacological inhibition of Src. Additionally, complete Freund's Adjuvant (CFA)-induced inflammatory hypersensitivity could also be attenuated by local Src kinase inhibition.Conclusions: Our data demonstrate that Src phosphorylation is critical for PKC-mediated sensitization of TRPV1. Further, in a model of inflammatory pain, CFA, Src kinase inhibition could reduce thermal hypersensitivity. Targeting of Src kinase may have analgesic benefits in inflammatory pain conditions.Significance: Src kinase-mediated phosphorylation of TRPV1 is a critical regulator of the PKC-induced sensitization induced by multiple inflammatory mediators. This suggest a new regulatory mechanism governing TRPV1 function and a potential therapeutic target for inflammatory type pain, including cancer pain where Src antagonists are currently utilized. [ABSTRACT FROM AUTHOR]

Objective: Maternal high-fat diet (HFD) may alter the offspring intestinal immune system, thereby enhancing susceptibility toward inflammatory bowel disease. The objective of the current study was to investigate the impact of maternal HFD on offspring intestinal health using a mouse model of dextran sulfate sodium (DSS)-induced colitis.Methods: Dams were provided with either HFD (60%) or control diet. After weaning, female offspring from both groups were kept on 45% HFD. At 14 weeks of age, offspring were subjected to 2.5% DSS in drinking water for 5 days, followed by 5 days of recovery.Results: Offspring from maternal HFD had higher body weight gain before DSS induction and had higher liver and fat weights with increased adipocyte size at necropsy. When subjected to DSS treatment, HFD offspring had accelerated body weight loss and exaggerated disease activity index. HFD offspring had an elevated histopathological score and interleukin (IL)-1β, IL-6, and IL-17 expression with upregulated NF-κB signaling. Maternal HFD resulted in enhanced neutrophil infiltration associated with elevated expression of monocyte chemoattractant protein-1. Furthermore, maternal HFD suppressed AMP-activated protein kinase activity and decreased sirtuin 1 and p53 protein contents in offspring gut.Conclusions: Maternal HFD consumption predisposes offspring to a higher susceptibility to develop inflammatory bowel disease. [ABSTRACT FROM AUTHOR]

Background and Purpose: The pathogenesis of osteoarthritis implicates a low-grade inflammation associated to the innate immune system activation. Toll like receptor (TLR) stimulation triggers the release of inflammatory mediators, which aggravate osteoarthritis. We studied the preventive effect of 6-shogaol, a potential TLR4 inhibitor, on the treatment of experimental knee osteoarthritis.Experimental Approach: Osteoarthritis was induced in C57BL6 mice by surgical section of the medial meniscotibial ligament, which received 6-shogaol for eight weeks. Cartilage damage, inflammatory mediator presence and disease markers were assessed in joint tissues by immunohistochemistry. Computational modelling was used to predict binding modes of 6-shogaol into the TLR4/MD2 receptor and its permeability across cellular membranes. Employing LPS-stimulated chondrocytes and MAPK assay, we elucidated 6-shogaol action mechanisms.Key Results: 6-Shogaol treatment prevented articular cartilage lesions, synovitis and the presence of pro-inflammatory mediators, and disease markers in osteoarthritis animals. Molecular modelling studies predicted 6-shogaol interaction with the TLR4/MD-2 heterodimer in an antagonist conformation through its binding into the MD-2 pocket. In cell culture, we confirmed that 6-shogaol reduced LPS-induced TLR4 inflammatory signalling pathways. Besides, MAPK assay demonstrated that 6-shogaol directly inhibits the ERK1/2 phosphorylation activity.Conclusion and Implications: 6-Shogaol evoked a preventive action on cartilage and synovial inflammation in osteoarthritis mice. 6-shogaol effect may take place not only by hindering the interaction between TLR4 ligands and the TLR4/MD-2 complex in chondrocytes, but also through inhibition of ERK phosphorylation, implying a pleiotropic effect on different mediators activated during osteoarthritis, which proposes it as an attractive drug for osteoarthritis treatments. [ABSTRACT FROM AUTHOR]

Background and Purpose: Recent evidence links brain insulin resistance with neurodegenerative diseases, where hyperphosphorylated tau protein contributes to neuronal cell death. In the present study, we aimed to evaluate if d-pinitol inositol, which acts as an insulin sensitizer, affects the phosphorylation status of tau protein.Experimental Approach: We studied the pharmacological effect of d-pinitol on insulin signalling and tau phosphorylation in the hippocampus of Wistar and Zucker rats. To this end, we evaluated by western blotting the Akt pathway and its downstream proteins as being one of the main insulin-mediator pathways. Also, we explored the functional status of additional kinases phosphorylating tau, including PKA, ERK1/2, AMPK and CDK5. We utilized the 3xTg mouse model as a control for tauopathy, since it carries tau mutations that promote phosphorylation and aggregation.Key Results: Surprisingly, we discovered that oral d-pinitol treatment lowered tau phosphorylation significantly, but not through the expected kinase GSK-3 regulation. An extensive search for additional kinases phosphorylating tau revealed that this effect was mediated through a mechanism dependent on the reduction of the activity of the CDK5, affecting both its p35 and p25 subunits. This effect disappeared in leptin-deficient Zucker rats, uncovering that the association of leptin deficiency, obesity, dyslipidaemia and hyperinsulinaemia abrogates d-pinitol actions on tau phosphorylation. The 3xTg mice confirmed d-pinitol effectiveness in a genetic AD-tauopathy.Conclusion and Implications: The present findings suggest that d-pinitol, by regulating CDK5 activity through a decrease of CDK5R1, is a potential drug for developing treatments for neurological disorders such as tauopathies. [ABSTRACT FROM AUTHOR]

Insulin resistance, a smaller than expected response to a given dose of insulin, is associated with many common diseases including, ageing, polycystic ovarian disease, syndrome X, cancer, infections, trauma and, most significantly, obesity and type 2 diabetes mellitus. The biochemical basis of insulin resistance in type 2 diabetes has been the subject of many studies. Earlier studies have indicated that quantitative regulation of the insulin sensitive glucose transporters (Glut-4) and insulin receptors themselves may contribute to this disorder, however, these two factors are probably inadequate to explain the extent of insulin resistance. This point also became apparent by the development of only mild hyperinsulinaemia in mice with a targeted mutation in the Glut-4 gene. Studies on postreceptor defects in type 2 diabetes has recently focused on the intrinsic catalytic activity of the insulin receptor and downstream signalling events. A reduction in tyrosine phosphorylation of both the insulin receptor (IR) and the insulin receptor substrate-1 (IRS-1) has been noted in both animal and human type 2 diabetes. Importantly, this appears to occur in all of the major insulin-sensitive tissues, namely the muscle, fat and liver. It is now clear that decreased signalling capacity of the insulin receptor is an important component of this disease. I will review some of the potential mechanisms underlying this deficiency. [ABSTRACT FROM AUTHOR]

Background and Purpose: Kaempferol is a natural flavonoid widely investigated in various fields due to its antioxidant, anti-cancer, and anti-inflammatory activities, but few studies have shown its inhibitory effect on T cell activation. This study examined the therapeutic potential of kaempferol in atopic dermatitis by modulating T cell activation.Experimental Approach: Effects of kaempferol on T cell activation and the underlying mechanisms were investigated in Jurkat cells and mouse CD4+ T cells. A model of atopic dermatitis in mice was used to determine its therapeutic potential on T cell-mediated conditions in vivo. Western blots, RT-PCR, pulldown assays and ELISA were used, along with histological analysis of skin.Key Results: Pretreatment with kaempferol reduced CD69 expression and production of inflammatory cytokines including IL-2 from activated Jurkat cells and murine CD4+ T cells without cytotoxicity. Pulldown assays revealed that kaempferol physically binds to MRP-1 in T cells, inhibiting the action of MRP-1. In activated T cells, kaempferol suppressed JNK phosphorylation and the TAK1-IKKα mediated NF-κB pathway. Oral administration of kaempferol to mice showed improved manifestation of atopic dermatitis, a T cell-mediated condition. Western blot results showed that, as in the in vitro studies, decreased phosphorylation of JNK was associated with down-regulated MRP-1 activity in vivo, in the kaempferol-treated mice in the atopic dermatitis model.Conclusion and Implications: Kaempferol regulates T cell activation by inhibiting MRP-1 activity in activated T cells, thus showing protective effects against T cell mediated disease in vivo. [ABSTRACT FROM AUTHOR]

Berberine, a natural isoquinoline alkaloid isolated from the berberis species, has a wide array of biological properties such as anti-inflammatory, antibacterial, antifungal, and antihelminthic effects. We evaluated the antiviral effect of berberine against influenza A/FM1/1/47 (H1N1) in vivo and in vitro. The results showed that berberine strongly suppressed viral replication in A549 cells and in mouse lungs. Meanwhile, berberine relieved pulmonary inflammation and reduced necrosis, inflammatory cell infiltration, and pulmonary edema induced by viral infection in mice when compared with vehicle-treated mice. Berberine suppressed the viral infection-induced up-regulation of TLR7 signaling pathway, such as TLR7, MyD88, and NF-κB (p65), at both the mRNA and protein levels. Furthermore, berberine significantly inhibited the viral infection-induced increase in Th1/Th2 and Th17/Treg ratios as well as the production of inflammatory cytokines. Our data provide new insight into the potential of berberine as a therapeutic agent for viral infection via its antiviral activity. [ABSTRACT FROM AUTHOR]

Introduction: N-methyl-d-aspartate (NMDA) is expressed in sensory neurons and plays important roles in peripheral pain mechanisms. The aim of this study was to examine the effects and molecular mechanisms of NMDA on C2C12 myoblast proliferation and differentiation.Methods: Cytotoxicity and differentiation were examined by the MTT assay, reverse transcription-polymerase chain reaction, and immunofluorescence.Results: NMDA had no cytotoxicity (10-500 μM) and inhibited myoblastic differentiation of C2C12 cells, as assessed by F-actin immunofluorescence and levels of mRNAs encoding myogenic markers such as myogenin and myosin heavy-chain 2. It inhibited phosphorylation of mammalian target of rapamycin (mTOR) by inactivating mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38. It induced reactive oxygen species production. Furthermore, NMDA-suppressed expression of F-actin was reversed by adding the antioxidant N-acetylcysteine.Conclusions: Collectively, these results indicate that NMDA impairs myogenesis or myogenic differentiation in C2C12 cells through the mTOR/MAPK signaling pathways and may lead to skeletal muscle degeneration. Muscle Nerve 56: 510-518, 2017. [ABSTRACT FROM AUTHOR]

Duchenne muscular dystrophy is a lethal muscle-wasting disease that affects boys. Mutations in the dystrophin gene result in the absence of the dystrophin glycoprotein complex (DGC) from muscle plasma membranes. In healthy muscle fibers, the DGC forms a link between the extracellular matrix and the cytoskeleton to protect against contraction-induced membrane lesions and to regulate cell signaling. The absence of the DGC results in aberrant regulation of inflammatory signaling cascades. Inflammation is a key pathological characteristic of dystrophic muscle lesion formation. However, the role and regulation of this process in the disease time-course has not been sufficiently examined. The transcription factor nuclear factor-κB has been shown to contribute to the disease process and is likely involved with increased inflammatory gene expression, including cytokines and chemokines, found in dystrophic muscle. These aberrant signaling processes may regulate the early time-course of inflammatory events that contribute to the onset of disease. This review critically evaluates the possibility that dystrophic muscle lesions in both patients with Duchenne muscular dystrophy and mdx mice are the result of immune-mediated mechanisms that are regulated by inflammatory signaling and also highlights new therapeutic directions. [Copyright &y& Elsevier]

Background and Purpose: While the 5-HT and Rho-kinase (ROCK) pathways have been implicated in the development of pulmonary arterial hypertension (PAH), the nature of any interactions between them remain unclear. This study investigated a role for ROCK in 5-HT-regulated proliferative responses in lung fibroblasts in vivo and in vitro.Experimental Approach: PAH was examined in mice over-expressing human 5-HT transporters (SERT+), from which pulmonary artery fibroblasts (PFs) were isolated to assess ROCK expression. In vitro analysis of 5-HT signalling employed CCL39 hamster lung fibroblasts.Key Results: ROCK inhibition ablated increased pulmonary remodelling and hypertension observed in SERT+ mice, and ROCK1/2 protein levels were elevated in SERT+ PFs. ROCK inhibition also reduced 5-HT-stimulated proliferation by suppressing MEK-stimulated ERK phosphorylation. While optimal 5-HT-stimulated proliferation required 5-HT(1B) and 5-HT(2A) receptors and SERT, receptor sensitivity to Y27632 was restricted to the 5-HT(1B) receptor. Also, while hypoxia-induced pulmonary vascular remodelling and hypertension were sensitive to Y27632 in WT and SERT+ animals, the proportions sensitive to ROCK inhibition were increased by SERT over-expression.Conclusions and Implications: SERT over-expression increased ROCK-dependent pulmonary remodelling in normoxia and hypoxia and SERT over-expression was associated with elevated ROCK1/2 levels. ROCK also potentiated 5-HT(1B) receptor-stimulated ERK activation and proliferation in vitro by facilitating MEK-ERK interaction. [ABSTRACT FROM AUTHOR]

The proliferation of mouse submandibular gland carcinoma YT-12 cells was stimulated by endothelial cell growth factor (ECGF)/bovine brain-derived acidic fibroblast growth factor (aFGF) and recombinant human aFGF. To determine whether aFGF was capable of modifying salivary gland carcinogenesis, the effect of brain-derived aFGF was examined in vivo. Mice in Groups 1 and 2 were injected with 9,10-dimethyl-1,2-benzanthracene (DMBA) into the left submandibular gland, and then Group 1 mice received bovine brain-derived aFGF and Group 2 mice received vehicle subcutaneously for 10 weeks. Group 3 and 4 mice received either bovine brain-derived aFGF or vehicle only. Sixteen weeks after the start of the experiment, the incidence of submandibular gland carcinomas in Group 1 was significantly greater than that in Group 2. Immunohistochemical study indicated that ducts in the normal submandibular glands and carcinomas showed positive staining with anti-aFGF antibody. Immunoblot and reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed the expression of aFGF in these tissues. FGF receptor (FGFR)-1 and FGFR-4 were detectable in the mouse submandibular glands and carcinomas. These findings suggest that bovine brain-derived aFGF stimulates the proliferation of submandibular gland carcinoma cells and promotes mouse submandibular gland carcinogenesis. [ABSTRACT FROM AUTHOR]