Activated Protein Kinase Signaling Research Articles

Roles of cell signaling pathways in thyroid carcinoma

The intracellular signal transduction pathways play an important role in the occurrence and development of thyroid cancer. Mitogen activated protein kinase (MAPK) signal transduction pathway, phosphatidylinositol 3-kinase (PI3K)-AKT pathway, NF-κB pathway and RASSF1-MST1-FOXO3 pathway affect the occurrence and development of thyroid cancer by modulating the activity and expression of specifical proteins, and determine the growth and apoptosis of thyroid cancer cells. Key words: Thyroid neoplasms; Signal transduction pathways

Evaluating the Oxidative Stress in Inflammation: Role of Melatonin.

Oxygen is used by eukaryotic cells for metabolic transformations and energy production in mitochondria. Under physiological conditions, there is a constant endogenous production of intermediates of reactive oxygen (ROI) and nitrogen species (RNI) that interact as signaling molecules in physiological mechanisms. When these species are not eliminated by antioxidants or are produced in excess, oxidative stress arises. Oxidative stress can damage proteins, lipids, DNA, and organelles. It is a process directly linked to inflammation; in fact, inflammatory cells secrete a large number of cytokines and chemokines responsible for the production of ROI and RNI in phagocytic and nonphagocytic cells through the activation of protein kinases signaling. Currently, there is a wide variety of diseases capable of producing inflammatory manifestations. While, in the short term, most of these diseases are not fatal they have a major impact on life quality. Since there is a direct relationship between chronic inflammation and many emerging disorders like cancer, oral diseases, kidney diseases, fibromyalgia, gastrointestinal chronic diseases or rheumatics diseases, the aim of this review is to describe the use and role of melatonin, a hormone secreted by the pineal gland, that works directly and indirectly as a free radical scavenger, like a potent antioxidant.

Monocyte Tumor Necrosis Factor-α-Converting Enzyme Catalytic Activity and Substrate Shedding in Sepsis and Noninfectious Systemic Inflammation.

Objectives:To determine the effect of severe sepsis on monocyte tumor necrosis factor-α–converting enzyme baseline and inducible activity profiles.Design:Observational clinical study.Setting:Mixed surgical/medical teaching hospital ICU.Patients:Sixteen patients with severe sepsis, 15 healthy volunteers, and eight critically ill patients with noninfectious systemic inflammatory response syndrome.Interventions:None.Measurements and Main Results:Monocyte expression of human leukocyte antigen-D-related peptide, sol-tumor necrosis factor production, tumor necrosis factor-α–converting enzyme expression and catalytic activity, tumor necrosis factor receptor 1 and 2 expression, and shedding at 48-hour intervals from day 0 to day 4, as well as p38-mitogen activated protein kinase expression. Compared with healthy volunteers, both sepsis and systemic inflammatory response syndrome patients’ monocytes expressed reduced levels of human leukocyte antigen-D-related peptide and released less sol-tumor necrosis factor on in vitro lipopolysaccharide stimulation, consistent with the term monocyte deactivation. However, patients with sepsis had substantially elevated levels of basal tumor necrosis factor-α–converting enzyme activity that were refractory to lipopolysaccharide stimulation and this was accompanied by similar changes in p38-mitogen activated protein kinase signaling. In patients with systemic inflammatory response syndrome, monocyte basal tumor necrosis factor-α–converting enzyme, and its induction by lipopolysaccharide, appeared similar to healthy controls. Changes in basal tumor necrosis factor-α–converting enzyme activity at day 0 for sepsis patients correlated with Acute Physiology and Chronic Health Evaluation II score and the attenuated tumor necrosis factor-α–converting enzyme response to lipopolysaccharide was associated with increased mortality. Similar changes in monocyte tumor necrosis factor-α–converting enzyme activity could be induced in healthy volunteer monocytes using an in vitro two-hit inflammation model. Patients with sepsis also displayed reduced shedding of monocyte tumor necrosis factor receptors upon stimulation with lipopolysaccharide.Conclusions:Monocyte tumor necrosis factor-α–converting enzyme catalytic activity appeared altered by sepsis and may result in reduced shedding of tumor necrosis factor receptors. Changes seemed specific to sepsis and correlated with illness severity. A better understanding of how tumor necrosis factor-α–converting enzyme function is altered during sepsis will enhance our understanding of sepsis pathophysiology, which will help in the assessment of patient inflammatory status and ultimately may provide new strategies to treat sepsis.

Signal pathway defects of Toll-like receptors 2-p38 mitogen activated protein kinase signal pathway defects and immune suppression in children with measles

Objective To investigate the expression of Toll-like receptors 2(TLR2)mRNA, p38 mitogen activated protein kinase(p38 MAPK)mRNA and cytokine in peripheral blood of children with measles and to study the effect and possible mechanism for TLR2-p38 MAPK signal pathway defects on immune suppression in the children with measles during acute phase. Methods Thirty children with measles hospitalized in the department of infectious diseases from June 2012 to July 2013 were enrolled into the measles group, and 30 healthy children were chosen as the healthy control group.The mRNA expressions of TLR and p38 MAPK in peripheral blood mononuclear cells (PBMC)were detected by reverse transcription-polymerase chain reaction (RT-PCR). The protein levels of interferon-γ(IFN-γ), tumor necrosis factor-β(TNF-β), interleukin(IL)-12, IL-6 and IL-10 in plasma were measured by using enzyme linked immunosorbent assay (ELISA), and flow cytometry (FCM)was applied to detect the percentage of lymphocyte subpopulation.The serum IgG, IgA and IgM levels were detected by velocity scatter turbidimetry. Results (1)The expressions of TLR2 mRNA and p38 MAPK mRNA in the measles group were both significantly lower than those in the healthy control group (all P 0.05). Conclusions The mRNA expressions of TLR2 and p38 MAPK are low in PBMC in the measles children during acute phase.There are different degrees of suppression of cell immunity, humoral immune and cytokines disorder in children with measles.Defects of TLR2-p38 MAPK signal pathway may cause the formation of measles immune suppression. Key words: Measles; Toll-like receptors 2; p38 mitogen activated protein kinase; Immune suppression; Child

Treatment with hydrogen molecule attenuates cardiac dysfunction in streptozotocin-induced diabetic mice

Diabetic cardiomyopathy, a disorder of the heart muscle in diabetic patients, is one of the major causes of heart failure. The aim of present study was to investigate the therapeutic effect of hydrogen molecule on streptozotocin-induced diabetic cardiomyopathy in mice. Diabetes was induced in adult male mice by consecutive peritoneal injection of streptozotocin (50 mg/kg/day) for 5 days. Then, they were treated with hydrogen water (1.3±0.2 mg/l) for 8 weeks (four groups, n=83-88 in each group). Although treatment of diabetic mice with hydrogen water did not significantly affect blood glucose level, it significantly attenuated cardiac hypertrophy and reduced expression of atrial natriuretic factor and β-myosin heavy chain; it alleviated cardiac fibrosis and reduced expression of collagen I and III, transforming growth factor beta, alpha-smooth muscle actin, and osteopontin; it reduced cardiac caspase-3 activity and ratio of bax/bcl-2. Importantly, hydrogen water treatment improved cardiac function in streptozotocin-diabetic mice. Furthermore, it was found that hydrogen water treatment abated oxidative stress, suppressed inflammation, and attenuated endoplasmic reticulum stress in the hearts of streptozotocin-diabetic mice. In addition, hydrogen water treatment suppressed activation of Jun NH2-terminal kinase and p38 mitogen activated protein kinase signaling and nuclear factor κB signaling in the hearts of streptozotocin-diabetic mice. Treatment with hydrogen molecule attenuated cardiac dysfunction in streptozotocin-induced diabetic mice, which was independent of glycemic control. Treatment with hydrogen molecule attenuated cardiac dysfunction in streptozotocin-induced type 1 diabetic mice. Molecular hydrogen could thus be envisaged as a nutritional countermeasure for diabetic cardiomyopathy.

The reduction of the mitochondrial cytochrome B makes development of gastric cancerthrough mitogen-activated protein kinase pathway

Objective Collect clinical specimens of gastric cancer patients with metastasis and without metastasis, application of molecular biology method confirm cytochrome B in its role in the development of cancer of the stomach, provide experimental basis for exploring the mechanism of gastric cancer. Methods Collect 12 cases confirmed by pathology of gastric cancer surgery in patients with gastric cancer tissue, including specimens with and without lymph node metastasis specimens each six cases. Detect the expression of two groups of gastric cancer tissue apoptosis related factors B cell lymphoma/leukemia- 2 associated X protein(bax), B cell lymphoma/leukemia- 2(bcl- 2), Caspase and mitogen- activated protein kinase(MAPK)signaling pathway proteins level by Western blotting. Results Compared with the group without metastasis, the expression of promoting apoptosis factor bax was reduced in gastric cancer tissue(0.55:1.00, P< 0.05), and the expression of inhibiting apoptosis factor bcl-2 increased(1.32:1.00, P<0.05), and the level of cell apoptosis decreased(2.86:3.35, P< 0.05). MAPK pathway activation level in gastric cancer tissue with metastasis was significantly reduced by comparing with the gastric cancer tissue without metastasis(P< 0.05). Conclusion Cytochrone B expression level decrease, thus inhibit MAPK pathway phosphorylation and reduce the cell apoptosis. Gastric cancer metastasis is related to the above mechanism. Key words: Gastric cancer; Mitochondrial cytochrome B; Apoptosis; Mitogen-activated protein kinase pathway

Proteome mapping of epidermal growth factor induced hepatocellular carcinomas identifies novel cell metabolism targets and mitogen activated protein kinase signalling events.

BackgroundHepatocellular carcinoma (HCC) is on the rise and the sixth most common cancer worldwide. To combat HCC effectively research is directed towards its early detection and the development of targeted therapies. Given the fact that epidermal growth factor (EGF) is an important mitogen for hepatocytes we searched for disease regulated proteins to improve an understanding of the molecular pathogenesis of EGF induced HCC. Disease regulated proteins were studied by 2DE MALDI-TOF/TOF and a transcriptomic approach, by immunohistochemistry and advanced bioinformatics.ResultsMapping of EGF induced liver cancer in a transgenic mouse model identified n = 96 (p < 0.05) significantly regulated proteins of which n = 54 were tumour-specific. To unravel molecular circuits linked to aberrant EGFR signalling diverse computational approaches were employed and this defined n = 7 key nodes using n = 82 disease regulated proteins for network construction. STRING analysis revealed protein-protein interactions of > 70% disease regulated proteins with individual proteins being validated by immunohistochemistry. The disease regulated network proteins were mapped to distinct pathways and bioinformatics provided novel insight into molecular circuits associated with significant changes in either glycolysis and gluconeogenesis, argine and proline metabolism, protein processing in endoplasmic reticulum, Hif- and MAPK signalling, lipoprotein metabolism, platelet activation and hemostatic control as a result of aberrant EGF signalling. The biological significance of the findings was corroborated with gene expression data derived from tumour tissues to evntually define a rationale by which tumours embark on intriguing changes in metabolism that is of utility for an understanding of tumour growth. Moreover, among the EGF tumour specific proteins n = 11 were likewise uniquely expressed in human HCC and for n = 49 proteins regulation in human HCC was confirmed using the publically available Human Protein Atlas depository, therefore demonstrating clinical significance.ConclusionNovel insight into the molecular pathogenesis of EGF induced liver cancer was obtained and among the 37 newly identified proteins several are likely candidates for the development of molecularly targeted therapies and include the nucleoside diphosphate kinase A, bifunctional ATP-dependent dihydroyacetone kinase and phosphatidylethanolamine-binding protein1, the latter being an inhibitor of the Raf-1 kinase.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1312-z) contains supplementary material, which is available to authorized users.

Suppression of oxidative stress and improvement of liver functions in mice by ursolic acid via LKB1-AMP-activated protein kinase signaling.

Hepatic cirrhosis is the final stage of liver dysfunction, characterized by diffuse fibrosis, which is the main response to the liver injury. This study is to investigate the effects of ursolic acid (UA) on liver functions and fibrosis in bile duct ligation (BDL) mice and to determine the underlying mechanisms. Cultured hepatocytes were treated with lipopolysaccharide (LPS) in the presence or absence of UA. The reactive oxygen species (ROS) level, protein levels of IκBα, iNOS and Cox-2, and NF-κB activation were detected, respectively. C57/BL6 and AMP-activated protein kinase (AMPK)α2(-/-) mice were subjected to BDL for 14 days. UA was administered by gavage. The markers of liver function and oxidative stress, and liver histopathology were analyzed after treatment. Treatment of hepatocytes with UA dose-dependently activates AMPK, which is abolished by silence of liver kinase B1 (LKB1). LPS significantly increased ROS productions, apoptosis, NF-κB activation, and expressions of iNOS and Cox-2 in cultured hepatocytes. All these effects were blocked by co-incubation with UA. Importantly, silence of LKB1, AMPK, or iNOS/Cox-2 by small interference RNA transfection reversed UA-induced effects in cultured cells. In an animal study, 14-day BDL induced liver fibrosis and liver injury, accompanied with increased oxidative stress and protein expressions of iNOS and Cox-2 in liver. Treatment of UA significantly attenuated the BDL-induced detrimental effects in wild-type mice but not in AMPKα2(-/-) mice. UA via LKB1-AMPK signaling offers protective effects on BDL-induced liver injury in mice, which may be related to inhibition of oxidative stress.

V600E BRAF mutation in pilocytic astrocytoma is associated with a more diffuse growth pattern but does not confer a more aggressive clinical behavior.

Activation in mitogen activated protein kinase signaling pathway has recently been described as a predominant event in pilocytic astrocytoma (PA) and is commonly caused by constitutively active mutation in BRAF protein. Whereas PA of posterior fossa in children have a high prevalence of BRAF duplication and fusion, primary molecularm abnormalities in supratentorial tumors of adults are more diverse and also include BRAF V600E point mutation. In our study we evaluated 51 PAs for BRAF duplication and BRAF V600E point mutation. We found a relatively high frequency of V600E mutation in our cohort. Histologically, V600E-carrying PA appeared more infiltrative, yet our limited clinical follow-up failed to detect a deleterious prognostic significance.

MiR-155 induced transcriptome changes in the MCF-7 breast cancer cell line leads to enhanced mitogen activated protein kinase signaling.

A single microRNA (miRNA) has the potential to regulate thousands of genes and thus govern multiple signaling pathways at once. miR-155 is an oncogenic miRNA which regulates many cellular pathways, designating it as a multifaceted regulator of proliferation, chemo-resistance, and apoptosis. While many singular targeted effects of miR-155 have been defined and an oncogenic role has been attributed to miR-155 expression, the global effect of miR-155 on the cellular transcriptomes of an ER(+) breast cancer cell line has yet to be determined. Here we demonstrate that miR-155 expression increases tumorigenesis in vivo and we determine miR-155 mediated transcriptome changes through next generation sequencing analysis. miR-155 expression alters many signaling pathways, with the chief altered pathway being the MAPK signaling cascade and miR-155 induces shortening of target mRNA 3'UTRs and alternative isoform expression of MAPK related genes. In addition there is an observed increase in protein phosphorylation of components of MAPK signaling including ERK1/2 and AP-1 complex members (Fra-1 and c-Fos) as well as elevated gene expression of MAPK regulated genes Zeb1, Snail, Plaur, and SerpinE1.

Ethanol impairs intestinal barrier function in humans through mitogen activated protein kinase signaling: a combined in vivo and in vitro approach.

BackgroundEthanol-induced gut barrier disruption is associated with several gastrointestinal and liver disorders.AimSince human data on effects of moderate ethanol consumption on intestinal barrier integrity and involved mechanisms are limited, the objectives of this study were to investigate effects of a single moderate ethanol dose on small and large intestinal permeability and to explore the role of mitogen activated protein kinase (MAPK) pathway as a primary signaling mechanism.MethodsIntestinal permeability was assessed in 12 healthy volunteers after intraduodenal administration of either placebo or 20 g ethanol in a randomised cross-over trial. Localization of the tight junction (TJ) and gene expression, phosphorylation of the MAPK isoforms p38, ERK and JNK as indicative of activation were analyzed in duodenal biopsies. The role of MAPK was further examined in vitro using Caco-2 monolayers.ResultsEthanol increased small and large intestinal permeability, paralleled by redistribution of ZO-1 and occludin, down-regulation of ZO-1 and up-regulation of myosin light chain kinase (MLCK) mRNA expression, and increased MAPK isoforms phosphorylation. In Caco-2 monolayers, ethanol increased permeability, induced redistribution of the junctional proteins and F-actin, and MAPK and MLCK activation, as indicated by phosphorylation of MAPK isoforms and myosin light chain (MLC), respectively, which could be reversed by pretreatment with either MAPK inhibitors or the anti-oxidant L-cysteine.ConclusionsAdministration of moderate ethanol dosage can increase both small and colon permeability. Furthermore, the data indicate a pivotal role for MAPK and its crosstalk with MLCK in ethanol-induced intestinal barrier disruption.Trial RegistrationClinicalTrials.gov NCT00928733

The Ethanolic Extract of Bark from Salix Aegyptiaca L. Inhibits the Metastatic Potential and Epithelial to Mesenchymal Transition of Colon Cancer Cell Lines

Willow bark extracts have been used for centuries as a natural pain killer. Recently their potential as anticancer agents has been reported. We have shown the high antioxidant activity, phenolic and flavonoid content in the ethanolic extract of bark (EEB) from Salix aegyptiaca, a species endogenous to the Middle East. We have also reported that incubation with EEB resulted in a reduction in cell proliferation through the induction of apoptosis and cell cycle arrest via the inhibition of phosphatidyl inositol 3-kinase/Protein kinase B and mitogen activated protein kinases signaling pathways as strongly as commercial inhibitors. The current study demonstrates the robust inhibition of anchorage-independent growth, motility, migration, and adhesion of colon cancer cell lines HCT-116 and HT-29 by EEB. These in vitro functional changes were accompanied by a restoration of E-cadherin expression, a reduction in EGFR, SNAI1, SNAI2, and Twist1 and the matrix metalloproteases MMP9 and MMP2. Many of these proteins are involved in the process of epithelial to mesenchymal transition, which is considered as a critical step in the progression of noninvasive tumor cells into malignant, metastatic carcinomas. We therefore propose that EEB from Salix aegyptiaca is a potent nutraceutical causing cancer chemoprevention by inhibiting epithelial to mesenchymal transition and can be consumed for its health promoting effects.

Cholesterol selectively regulates IL-5 induced mitogen activated protein kinase signaling in human eosinophils.

Eosinophils function contributes to human allergic and autoimmune diseases, many of which currently lack curative treatment. Development of more effective treatments for eosinophil-related diseases requires expanded understanding of eosinophil signaling and biology. Cell signaling requires integration of extracellular signals with intracellular responses, and is organized in part by cholesterol rich membrane microdomains (CRMMs), commonly referred to as lipid rafts. Formation of these organizational membrane domains is in turn dependent upon the amount of available cholesterol, which can fluctuate widely with a variety of disease states. We tested the hypothesis that manipulating membrane cholesterol content in primary human peripheral blood eosinophils (PBEos) would selectively alter signaling pathways that depend upon membrane-anchored signaling proteins localized within CRMMs (e.g., mitogen activated protein kinase [MAPK] pathway), while not affecting pathways that signal through soluble proteins, like the Janus Kinase/Signal Transducer and Activator of Transcription [JAK/STAT] pathway. Cholesterol levels were increased or decreased utilizing cholesterol-chelating methyl-β-cyclodextrin (MβCD), which can either extract membrane cholesterol or add exogenous membrane cholesterol depending on whether MβCD is preloaded with cholesterol. Human PBEos were pretreated with MβCD (cholesterol removal) or MβCD+Cholesterol (MβCD+Chol; cholesterol delivery); subsequent IL-5-stimulated signaling and physiological endpoints were assessed. MβCD reduced membrane cholesterol in PBEos, and attenuated an IL-5-stimulated p38 and extracellular-regulated kinase 1/2 phosphorylation (p-p38, p-ERK1/2), and an IL-5-dependent increase in interleukin-1β (IL-1β) mRNA levels. In contrast, MβCD+Chol treatment elevated PBEos membrane cholesterol levels and basal p-p38, but did not alter IL-5-stimulated phosphorylation of ERK1/2, STAT5, or STAT3. Furthermore, MβCD+Chol pretreatment attenuated an IL-5-induced increase in cell survival at 48 hours, measured as total cellular metabolism. The reduction in cell survival following cholesterol addition despite unaltered STAT phosphorylation contradicts the current dogma in which JAK/STAT activation is sufficient to promote eosinophil survival, and suggests an additional, unidentified mechanism critically regulates IL-5-mediated human PBEos survival.

Thromboxane A2 stimulates neurite outgrowth in cerebral cortical neurons via mitogen activated protein kinase signaling

Thromboxane A2 (TXA2) is a central prostanoid in the cardiovascular system and is a crucial mediator of vascular homeostasis and platelet aggregation. In this study, we report a novel role for TXA2 in neurite outgrowth. TXA2 receptor is expressed in rat cortical neurons, and treatment with the TXA2 agonist U-46619 promotes neurite outgrowth in a concentration-dependent manner. We investigated the molecular mechanism underlying U-46619-induced neurite outgrowth in cortical neurons. Blockade of the phosphorylation of mitogen activated protein kinase (MAPK) prevents U-46619-mediated neurite outgrowth. These data indicates that TXA2 functions as a positive regulator of neurite outgrowth via a mechanism dependent on MAPKs in cortical neurons.

Maternal fructose intake during pregnancy modulates hepatic and hypothalamic AMP‐activated protein kinase signalling in a sex‐specific manner in offspring

Dietary fructose ingestion during gestation affects carbohydrate metabolism in the offspring. In the present study, we investigated the effects of excess fructose intake during pregnancy on hepatic and hypothalamic AMP-activated kinase (AMPK) expression and phosphorylation, as well as hepatic glucose-6-phosphatase (G6Pase) activity in offspring. Pregnant Wistar rats received normal chow and 100g/L fructose solution or normal water during gestation ad libitum. On gestational Day 21, some dams were killed and plasma samples and fetuses were collected. The remaining dams received normal water after spontaneous delivery during lactation. Pups were killed on postnatal Day 22 and the plasma, liver and hypothalamus were collected and analysed. Plasma glucose and insulin levels increased in female but not male offspring in the fructose group. Although the mRNA and total protein levels of AMPKα were unchanged, levels of phosphorylated AMPKα protein in the fructose group of female offspring were significantly lower in the liver and 4.6-fold higher in the hypothalamus. The hepatic protein level of sirtuin 1, which is involved in AMPK phosphorylation and activation, was significantly reduced in the fructose group of female offspring. The activity of G6Pase, which plays a role in gluconeogenesis, was significantly enhanced in the liver of female offspring from fructose-fed dams. These changes were not observed in male offspring. In conclusion, we found that excessively high fructose intake during pregnancy may modulate the hepatic and hypothalamic AMPK signalling pathways in female offspring after birth.

Identification and validation co-differentially expressed genes with NAFLD and insulin resistance

Insulin resistance is a major feature and pathogenic factor of nonalcoholic fatty liver disease (NAFLD). Theoretically, genetic variation in candidate genes related to insulin resistance may contribute to the pathogenesis of NAFLD. The purpose of this study was to identify potentially pathogenic genes involved in NAFLD and insulin resistance that have not yet been discovered. This study yielded five important discoveries. 1. A total of 21 co-differentially expressed genes in both the NAFLD and insulin-resistance groups were identified from the pool containing thousands of genes via the significance analysis of microarraysmethod. 2. MAP kinase-interacting serine/threonine kinase 2 (Mknk2) was the unique gene to be identified that is involved in the insulin signaling pathway and Mitogen Activated Protein Kinase signaling pathway according to the Kyoto Encyclopedia of Genes and Genomes database. 3. Mknk2 mRNA and protein expression were dose-dependently up-regulated by palmitic acid (PA) in mouse primary hepatocytes. 4. Western blotting analysis and quantitative real-time PCR confirmed that Mknk2 affected the expression of acetyl-CoA carboxylases-1 and fatty acid synthase. 5. The inhibition of Mknk2 alleviated PA-induced insulin resistance, whereas the overexpression of Mknk2 resulted in the aggravation of insulin resistance in PA-treated hepatocytes. Therefore, we predict that MKNK2 may be a key protein related to NAFLD and insulin resistance.

The effect of 5-aminoimidazole-4-carboxamide-ribonucleoside was mediated by p38 mitogen activated protein kinase signaling pathway in FRO thyroid cancer cells.

Background/Aims5'-Adenosine monophosphate (AMP)-activated protein kinase (AMPK) is a cellular energy sensor that monitors intracellular AMP/adenosine triphosphate (ATP) ratios and is a key regulator of the proliferation and survival of diverse malignant cell types. In the present study, we investigated the effect of activating AMPK by 5-aminoimidazole-4-carboxamide-ribonucleotide (AICAR) in thyroid cancer cells.MethodsWe used FRO thyroid cancer cells harboring the BRAFV600E mutation to examine the effect of AICAR on cell proliferation and cell survival. We also evaluated the involvement of mitogen-activated protein kinase (MAPK) pathways in this effect.ResultsWe found that AICAR treatment promoted AMPK activation and suppressed cell proliferation and survival by inducing p21 accumulation and activating caspase-3. AICAR significantly induced activation of p38 MAPK, and pretreatment with SB203580, a specific inhibitor of the p38 MAPK pathway, partially but significantly rescued cell survival. Furthermore, small interfering RNA targeting AMPK-α1 abolished AICAR-induced activation of p38 MAPK, p21 accumulation, and activation of caspase-3.ConclusionsOur findings demonstrate that AMPK activation using AICAR inhibited cell proliferation and survival by activating p38 MAPK and proapoptotic molecules in FRO thyroid cancer cells. These results suggest that the AMPK and p38 MAPK signaling pathways may be useful therapeutic targets to treat thyroid cancer.

Differential Host Cell Gene Expression and Regulation of Cell Cycle Progression by Nonstructural Protein 11 of Porcine Reproductive and Respiratory Syndrome Virus

Nonstructural protein 11 (nsp11) of porcine reproductive and respiratory syndrome virus (PRRSV) is a viral endoribonuclease with an unknown function. The regulation of cellular gene expression by nsp11 was examined by RNA microarrays using MARC-nsp11 cells constitutively expressing nsp11. In these cells, the interferon-β, interferon regulatory factor 3, and nuclear factor-κB activities were suppressed compared to those of parental cells, suggesting that nsp11 might serve as a viral interferon antagonist. Differential cellular transcriptome was examined using Affymetrix exon chips representing 28,536 transcripts, and after statistical analyses 66 cellular genes were shown to be upregulated and 104 genes were downregulated by nsp11. These genes were grouped into 5 major signaling pathways according to their functional relations: histone-related, cell cycle and DNA replication, mitogen activated protein kinase signaling, complement, and ubiquitin-proteasome pathways. Of these, the modulation of cell cycle by nsp11 was further investigated since many of the regulated genes fell in this particular pathway. Flow cytometry showed that nsp11 caused the delay of cell cycle progression at the S phase and the BrdU staining confirmed the cell cycle arrest in nsp11-expressing cells. The study provides insights into the understanding of specific cellular responses to nsp11 during PRRSV infection.

Plumbagin inhibits LPS-induced inflammation through the inactivation of the nuclear factor-kappa B and mitogen activated protein kinase signaling pathways in RAW 264.7 cells

Plumbagin (PL) has been reported to exhibit anti-carcinogenic, anti-inflammatory and analgesic activities, but little is known about its mechanism. In this study, we investigated the anti-inflammatory property of PL and its mechanism of action. Although no significant cytotoxicity of PL was observed over the concentration range tested, PL (2.5–7.5μM) significantly and dose-dependently suppressed the secretion of pro-inflammatory mediators and inhibited the expression of TNF-α, IL-1β, IL-6 and iNOS in LPS-stimulated RAW 264.7 cells. Furthermore, PL consistently suppressed the activity of iNOS in LPS-induced RAW 264.7 cells. To elucidate the mechanism underlying the anti-inflammatory activity of PL, we assessed the effects of PL on the MAPK pathway and the activity and expression of NF-κB. These experiments demonstrated that PL significantly reduced the luciferase activity of an NF-κB promoter reporter and p65 nuclear translocation. The LPS-induced phosphorylation of MAP kinases was also attenuated by PL; significant changes were observed in the levels of phosphorylated ERK1/2, JNK and p38 MAPK. Additionally, MAPK inhibitors confirmed the inhibitory effect of PL on the MAPK pathway. Taken together, these data suggest that PL exerts its anti-inflammatory effects by down-regulating the expression of pro-inflammatory mediators through inhibition of NF-κB and MAPK signaling in LPS-stimulated RAW 264.7 cells.

Protective cardiovascular effects of metformin in type 2 diabetic patients: evidence and underlying mechanisms

Metformin has been recommended as the first-line therapy for type 2 diabetes.Besides its ideal glucose-lowering effect,metformin has also been shown to exert beneficial effects on cardiovascular system,including preventing the occurrence and progression of atherosclerosis,reducing risk for cardiovascular events,attenuating myocardial ischemia-reperfusion injury,improving ventricular remodeling,and slowing the progress of heart failure.The activation of adenosine monophosphate activated protein kinase (AMPK) signaling pathway plays a pivotal role in the protective cardiovascular effects resulted from metformin. Key words: Metformin; Diabetes mellitus, type 2; Cardiovascular events; Heart failure; Adenosine monophosphate activated protein kinase