Tumor necrosis factor-α-induced nuclear factor-kappaB activation in human cardiomyocytes is mediated by NADPH oxidase

An elevated level of tumor necrosis factor (TNF)-α is implicated in several cardiovascular diseases including heart failure. Numerous reports have demonstrated that TNF-α activates nuclear factor (NF)-kappaB, resulting in the upregulation of several genes that regulate inflammation, proliferation, and apoptosis of cardiomyocytes. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, a major source of reactive oxygen species (ROS), is also activated by TNF-α and plays a crucial role in redox-sensitive signaling pathways. The present study investigated whether NADPH oxidase mediates TNF-α-induced NF-kappaB activation and NF-kappaB-mediated gene expression. Human cardiomyocytes were treated with recombinant TNF-α with or without pretreatment with diphenyleneiodonium (DPI) and apocynin, inhibitors of NADPH oxidase. TNF-α-induced ROS production was measured using 5-(and-6)-chloromethyl-2’, 7’-dichlorodihydrofluorescein diacetate assay. TNF-α-induced NF-kappaB activation was also examined using immunoblot; NF-kappaB binding to its binding motif was determined using a Cignal reporter luciferase assay and an electrophoretic mobility shift assay. TNF-α-induced upregulation of interleukin (IL)-1β and vascular cell adhesion molecule (VCAM)-1 was investigated using real-time PCR and immunoblot. TNF-α-induced ROS production in cardiomyocytes was mediated by NADPH oxidase. Phosphorylation of IKK-α/β and p65, degradation of IkappaBα, binding of NF-kappaB to its binding motif, and upregulation of IL-1β and VCAM-1 induced by TNF-α were significantly attenuated by treatment with DPI and apocynin. Collectively, these findings demonstrate that NADPH oxidase plays a role in regulation of TNF-α-induced NF-kappaB activation and upregulation of proinflammatory cytokines, IL-1β and VCAM-1, in human cardiomyocytes.     

Expression profile of MAGI2 gene as a novel biomarker in combination with major deregulated genes in prostate cancer

Complex molecular changes that occur during prostate cancer (PCa) progression have been described recently. Whole genome sequencing of primary PCa samples has identified recurrent gene deletions and rearrangements in PCa. Specifically, these molecular events disrupt the gene loci of phosphatase and tensin homolog (PTEN) and membrane-associated guanylate kinase inverted-2 (MAGI2). In the present study, we analyzed the expression profile of MAGI2 gene in a cohort of clinical PCa (n = 45) and benign prostatic hyperplasia (BPH) samples (n = 36) as well as three PCa cell lines. We also studied the expression of PCa-related genes, including PTEN, NKX3.1, SPINK1, DD3, AMACR, ERG, and TMPRSS2-ERG fusion in the same samples. The expression of MAGI2 mRNA was significantly down-regulated in PC3, LNCaP and DU-145 PCa cell lines (p = 0.000), and also in clinical tumor samples (Relative expression = 0.307, p = 0.002, [95 % CI 0.002–12.08]). The expression of PTEN, NKX3.1, SPINK1, DD3, and AMACR genes was significantly deregulated in prostate tumor samples (p range 0.000–0.044). A significant correlation was observed between MAGI2 and NKX3.1 expression in tumor samples (p = 0.006). Furthermore, the inclusion of MAGI2 in the gene panel improved the accuracy for discrimination between PCa and BPH samples with the sensitivity and specificity of 0.88 [CI 0.76–0.95] and 0.83 [CI 0.68–0.92], respectively. The data presented here suggest that MAGI2 gene can be considered as a novel component of gene signatures for the detection of PCa.     

Negative Purifying Selection Drives Prion and Doppel Protein Evolution

The prion protein (PrP) when misfolded into the pathogenic conformer PrP^Sc is the major causative agent of several lethal transmissible spongiform encephalopathies in mammals. Studies of evolutionary pressure on the corresponding gene using different datasets have yielded conflicting results. In addition, putative PrP or PrP interacting partners with strong similarity to PrP such as the doppel protein have not been examined to determine if the same evolutionary mechanisms apply to prion paralogs or if there are coselected sites that might indicate how and where the proteins interact. We examined several taxonomic groups that contain model organisms of prion diseases focusing on primates, bovids, and an expanded dataset of rodents for selection pressure on the prion gene (PRNP) and doppel gene (PRND) individually and for coevolving sites within. Overall, the results clearly indicate that both proteins are under strong selective constraints with relaxed selection on amino acid residues connecting α-helices 1 and 2.     

Neurotrophic Factors,Clinical Features and Gender Differences in Depression

Recent studies have evaluated the role of brain-derived neurotrophic factor (BDNF) in mood disorders; however, little is known about alterations in nerve growth factor (NGF) and glial cell line-derived neurotrophic factor (GDNF). The aim of this study was to evaluate differences among serum neurotrophic factors (BDNF, NGF and GDNF) in depressed patients and healthy controls and to verify the association between serum neurotrophic levels and clinical characteristics in a young, depressed population stratified by gender. This is a cross-sectional study with depressed patients and population controls 18–29 years of age. The concentrations of neurotrophic factors were determined by the ELISA method. The diagnosis of depression and the duration of the disease were assessed by the Structured Clinical Interview according to the diagnostic and statistical manual of mental disorders. Depression severity was measured with the 17-item Hamilton Rating Scale for Depression, and the severity of anxiety symptoms was measured using the Hamilton Anxiety Rating Scale. Serum BDNF and GDNF were lower in major depressive disorder (MDD) patients compared to controls (p ≤ 0.001). Serum NGF levels were higher in MDD patients versus controls (p ≤ 0.001). BDNF was associated with the duration of disease only in women (p = 0.005). GDNF was not associated with clinical characteristics in either gender. In women, NGF was associated with the severity of depressive symptoms (p = 0.009), anxiety (p = 0.011) and disease duration (p = 0.005). NGF was associated with disease duration in men (p = 0.026). Our results demonstrated that significant neurochemical differences in NGF and BDNF, but not in GDNF, were associated with the clinical features of MDD when patients were stratified by gender.     

Rutin Alleviates Prion Peptide-Induced Cell Death Through Inhibiting Apoptotic Pathway Activation in Dopaminergic Neuronal Cells

Prion disorders are progressive neurodegenerative diseases characterized by extensive neuronal loss and accumulation of the abnormal form of the scrapie prion protein (PrP). Rutin is a flavonoid that occurs naturally in plant-derived beverages and foods and is used in traditional and folkloric medicine worldwide. In the present study, we evaluated the protective effects of rutin against PrP fragment (106–126)-induced neuronal cell death. Rutin treatment blocked PrP(106–126)-mediated increases in reactive oxygen species production and nitric oxide release and helped slowing the decrease of neurotrophic factors that results from PrP accumulation. Rutin attenuated PrP(106–126)-associated mitochondrial apoptotic events by inhibiting mitochondrial permeability transition and caspase-3 activity and blocking expression of the apoptotic signals Bax and PARP. Additionally, rutin treatment significantly decreased the expression of the death receptor Fas and its ligand Fas-L. Overall, our results demonstrated that rutin protects against the neurodegenerative effects of prion accumulation by increasing production of neurotropic factors and inhibiting apoptotic pathway activation in neuronal cells. These results suggested that rutin may have clinical benefits for prion diseases and other neurodegenerative disorders.     

Bone Metabolism Status and Associated Risk Factors in Elderly Patients with Chronic Obstructive Pulmonary Disease (COPD)

The prevalence of osteoporosis in older patients with chronic obstructive pulmonary disease (COPD) is higher than in the age-matched elderly patients, but the exact cause in relation to COPD is not clear. We hypothesized that the underlying causes for this difference are related to bone metabolism with the possible risk factors that include the duration of COPD, GOLD grade, cor pulmonale, the frequencies of acute exacerbations within the past year, smoking and inhaled corticosteroid therapy. We conducted a matched-pair study of 100 patients aged older than 65 years at the Southwest Hospital from May to November 2012. The enrolled patients with COPD were matched to controls for age and gender. Clinical characteristics of cohorts were recorded. Bone mineral density (BMD) was measured using dual-energy X-ray absorptiometry and osteoporosis was diagnosed according to the definition of WHO. All cohorts accepted bone metabolism marker measurement, including Procollagen type 1 aminoterminal propeptide (P1NP), β-C-telopeptides of type I collagen (βCTX), and N-terminal midmolecule fragment osteocalcin (N-MID OC). Statistical analysis was calculated using the student’s t test, ANOVA and multiple regression analysis at a significance level set at a p < 0.05. Circulating biochemical markers of bone formation (P1NP), resorption (βCTX) and turnover (N-MID OC) were significantly lower in the COPD group than control group, while mean 25-OH Vitamin D was similar in two groups. The P1NP, βCTX, and N-MID OC were still lower in men with COPD, but only P1NP was lower in women with COPD compared to that of controls. Multiple regression analysis in COPD group suggests that age, the frequency of acute exacerbation, and BMD are independent risk factors for P1NP. The frequency of acute exacerbation within the past one year and 25-OH D level are independent risk factors for βCTX; the frequency of acute exacerbation is the only independent risk factor for N-MID OC. These were significant differences in bone metabolism in patients with or without COPD. These results should help us to further understand the cause of osteoporosis and fractures and conduce to prevent osteoporosis in patients with COPD.     

Variation in organisation and copy number of ribosomal RNA genes inPetunia hybrida somaclones

The copy number of genes encoding 5S ribosomal RNA has been found to be constant in Petunia hybrida plants regenerated from protoplast and leaf disc-derived callus cultures. However, in one somaclone a heritable change in the length of the major 5S rDNA repeat has arisen. Despite the constant copy number of 5S rRNA genes, that of the 18S–25S rRNA genes is found to very by at least ten-fold. The relevance of these findings to ribosomal RNA gene variability and to somaclonal variation is discussed.     

Enhanced biotransformation of dehydroepiandrosterone to 3β,7α,15α-trihydroxy-5-androsten-17-one with Gibberella intermedia CA3-1 by natural oils addition

Dihydroxylation of dehydroepiandrosterone (DHEA) is an essential step in the synthesis of many important pharmaceutical intermediates. However, the solution to the problem of low biohydroxylation conversion in the biotransformation of DHEA has yet to be found. The effects of natural oils on the course of dihydroxylation of DHEA to 3β,7α,15α-trihydroxy-5-androsten-17-one (7α,15α-diOH-DHEA) were studied. With rapeseed oil (2 %, v/v) addition, the bioconversion efficiency was improved, and the 7α,15α-diOH-DHEA yield was increased by 40.8 % compared with that of the control at DHEA concentration of 8.0 g/L. Meantime, the ratio of 7α,15α-diOH-DHEA to 7α-OH-DHEA was also increased by 4.5 times in the rapeseed oil-containing system. To explain the mechanism underlying the increase of 7α,15α-diOH-DHEA yield, the effects of rapeseed oil on the pH of the bioconversion system, the cell growth and integrity of Gibberella intermedia CA3-1, as well as the membrane composition were systematically studied. The addition of rapeseed oil enhanced the substrate dispersion and maintained the pH of the system during bioconversion. Cells grew better with favorable integrity. The fatty acid profile of G. intermedia cells revealed that rapeseed oil changed the cell membrane composition and improved cell membrane permeability for lipophilic substrates.     

Recent advances in biochemistry and biotechnological synthesis of avermectins and their derivatives

Avermectins (AVMs), produced by Streptomyces avermitilis MA-4680 (or ATCC 31267, NRRL 8165, NCBIM 12804), are 16-member macrocylic lactones that play very important functions as bactericidal and antiparasitic agents against nematodes and anthropods, as well as Mycobacterium tuberculosis H37Rv. Since its discovery in 1975, use of AVM has been widely spreading around the globe. To date, the whole genome sequence of S. avermitilis K139 has been acquired, in which the AVM biosynthetic gene cluster was the most highly investigated to mine the genes responsible for functional as well as regulatory roles. Therefore, significant progress has been achieved for understanding and manipulating the biosynthesis, improved production, regulation mechanism, side effects, as well as the resistance of AVMs and their derivatives. These findings will facilitate further strain improvement and biosynthesis of novel derivatives bearing stable and improved biological activities, as well as overcoming the resistance mechanism to open up a bright period for these compounds. In this review, we have summarized and analyzed the update in advanced progress in biochemistry and biotechnological approaches used for the production of AVMs and their derivatives.     

The checkpoint 1 kinase inhibitor LY2603618 induces cell cycle arrest,DNA damage response and autophagy in cancer cells

Chemotherapy- or radiotherapy-induced DNA damage activates the Chk1-dependent DNA damage response (DDR) and cell cycle checkpoints to facilitate cell survival. Numerous attempts have been made to identify specific Chk1 inhibitors to enhance the efficiency of chemotherapy or radiotherapy. In this study, we investigated the molecular mechanisms underlying the antitumor activity of LY2603618, a potent and selective small molecule inhibitor of Chk1 protein kinase, in human lung cancer cells. Treatment of cancer cells with LY2603618 caused cell cycle arrest in the G2/M phase. A marked induction of DDR, including the phosphorylation of ATM, Chk2, p53 and histone H2AX, was observed after LY2603618 treatment. LY2603618 inhibited Chk1 autophosphorylation (S296 Chk1) and increased DNA damage-mediated Chk1 phosphorylation (S345 Chk1). In addition, LY2603618-treated lung cancer cells transitioned from LC3-I to LC3-II, a hallmark of autophagy. Blocking autophagy with chloroquine (CQ) further enhanced LY2603618′s inhibitory effect on cell viability/proliferation. LY2603618 also significantly increased p38 and c-Jun N-terminal kinase (JNK) phosphorylation. Pretreatment with the JNK inhibitor reduced cleavage of caspase-3 and PARP levels in LY2603618-treated cells. These results suggest the following: (i) the biological consequences of LY2603618 in lung cancer cells is associated with both inhibition of Chk1 phosphorylation on S296 and activation of the DNA damage response network; and (ii) the anticancer property of LY2603618 might be increased by inhibiting autophagy.     

Batimastat Nanoparticles Associated with Transcatheter Arterial Chemoembolization Decrease Hepatocellular Carcinoma Recurrence

Hepatocellular carcinoma (HCC) is a malignant tumor characterized by easy metastasis and frequent recurrence. Transarterial chemoembolization (TACE) remains the routine treatment for patients with HCC who are not eligible for surgical resection or percutaneous tumor ablation; however, 5-year survival rates following interventional therapy are only 17–38.8 %, with liver recurrence due to incomplete embolization and tumor angiogenesis being a significant reason for treatment failure. Ischemia and hypoxia induced by TACE is correlated with an increased expression of angiogenic factor and stimulates an increase in angiogenesis, including endothelial cells (ECs) proliferation. Matrix metalloproteinases (MMPs) are zinc-dependent proteolytic endopeptidases involved in tumor angiogenesis. In addition, MMPs stimulate tumor cell growth, migration and invasion, and metastasis. Hypoxia enhanced EC migration in a MMP-2-dependent manner while MMP inhibitors (MMPIs) significantly decreased the number of migrating cells in hypoxic cultures. We hypothesize batimastat (synthetic MMPI) nanoparticles associated with TACE could decrease HCC recurrence and metastasis. At first, batimastat nanoparticles were made from batimastat and poly(lactic-co-glycolic acid). Then, nanoparticles were mixed with lipiodol and chemotherapeutic drugs solution. The mixture was infused super-selectively into supplied artery of HCC through catheter. The disseminated area of batimastat might be same with TACE-induced hypoxia area. In the hypoxia area, batimastat inhibited the activity of MMPs, weakened the angiogenesis of tumor vascular system and migration of HCC cells. HCC cells could not escape from hypoxia area and tumor angiogenesis inhibited could not supply sufficient nutrients and O₂ to residual HCC cells. With the help of batimastat, the killing effect of chemotherapeutic drugs might be enhanced. The rate of complete necrosis of HCC lesion might be increased and local recurrence and metastasis of HCC might be reduced. The hypothesis might increase the clinical efficacy of TACE and improve the prognosis of HCC patients.     

The mitochondrial complex I of trypanosomatids - an overview of current knowledge

The contribution of trypanosomatid mitochondrial complex I for energy transduction has long been debated. Herein, we summarize current knowledge on the composition and relevance of this enzyme. Bioinformatic and proteomic analyses allowed the identification of many conserved and trypanosomatid-specific subunits of NADH:ubiquinone oxidoreductase, revealing a multifunctional enzyme capable of performing bioenergetic activities and possibly, also of functioning in fatty acid metabolism. A multimeric structure organized in 5 domains of more than 2 MDa is predicted, in contrast to the 1 MDa described for mammalian complex I. The relevance of mitochondrial complex I within the Trypanosomatidae family is quite diverse with its NADH oxidation activity being dispensable for both procyclic and bloodstream Trypanosoma brucei, whereas in Phytomonas serpens the enzyme is the only respiratory complex able to sustain membrane potential. Aside from complex I, trypanosomatid mitochondria contain a type II NADH dehydrogenase and a NADH-dependent fumarate reductase as alternative electron entry points into the respiratory chain and thus, some trypanosomatids may have bypassed the need for complex I. The involvement of each of these enzymes in the maintenance of the mitochondrial redox balance in trypanosomatids is still an open question and requires further investigation.     

Differentiation between attention-deficit/hyperactivity disorder and autism spectrum disorder by the Social Communication Questionnaire

The differentiation of attention-deficit/hyperactivity disorder (ADHD) and autism spectrum disorder (ASD) poses a clinical challenge. In children, overlap of psychopathological and cognitive findings has been found for both disorders. In addition, some children suffer from both disorders. The Social Communication Questionnaire (SCQ) is a screening instrument for ASD symptoms which indicates the presence of ASD in a rapid and economic way. However, validity to differentiate ASD and ADHD as differential or comorbid diagnoses has not been studied. Here, the differential validity was compared in groups of children with ASD, ADHD, ASD + ADHD, and typically developing (TD) children and IQ > 70. ROC analyses indicated an excellent differentiation between ASD and TD with ROC–AUC = .941 and between ASD + ADHD with ROC–AUC = .993. The optimal cutoff was below the originally recommended one of 15. The differentiation between children with ASD with (ROC–AUC = .982) or without ADHD (ROC–AUC = .864) and ADHD alone also showed acceptable differential validity, and here, the optimal cutoff corresponded to the recommended. Taken together, the SCQ can be recommended as a screening instrument for a first differentiation between children with ASD and typically developing children as well as children with ADHD.     

Analysis of the Relationship Between Ultrasound of Breast Cancer DOT-SDI and the Expression of MVD,VEGF and HIF-1α

Ultrasonic light scattering tomography system is a new imaging technique for breast function, which associates with diffused optical tomography (DOT) with ultrasonic examination. A recent study found microvessel density (MVD), vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1 alpha (HIF-1α) play a major role in angiogenesis of breast cancer. This study analyses the relationship between breast cancer ultrasound synthesis diagnostic index (SDI) and the expression of MVD, VEGF and HIF-1α by testing the expression level of the breast cancer gene MVD, VEGF and HIF-1α. In this study, we disclose the relationship between ultrasound of breast cancer DOT-SDI and the expression of MVD, VEGF and HIF-1α.     

High pH (and not free ammonia) is responsible for Anammox inhibition in mildly alkaline solutions with excess of ammonium

Ammonium is a substrate of the anaerobic ammonium oxidation (Anammox) process but it has been suggested as a substrate-inhibitor because of the action of its unionized form, free ammonia. High pH of the medium is also an important limiting factor of the Anammox bacteria. Both effects are difficult to discriminate. In this work the inhibitory effects of high pH, total ammonia (TA) and NH₃ on the Anammox process were investigated simultaneously. Results confirmed that TA caused no inhibition and high pH is a much more important inhibiting factor than NH₃ in mildly alkaline conditions, based on a multi-factorial analysis. Values of pH higher than 7.6 caused Anammox inhibition >10 % and should be avoided during the application of the Anammox process in practice.     

Reaction time and replenishment time of SP and CGRP after incision in rat skin

Background. The skin neurogenic inflammation is mainly related to Substance P (SP) and Calcitonin Gene-related Peptide (CGRP). There is no data on their availability in the dynamics of skin nerve endings, concerning their release and replenishment after a nociceptive stimulus, so this was investigated. Materials and methods. 25 rats were randomly distributed in 5 groups. The animals of the control group (CG) determined the baseline levels of neuropeptides in the skin. The groups S0 and S30 did not receive any cutaneous stimulus at 30 and 60 minutes, respectively. In the group S1, an “incision stimulus” was made at 30 minutes. In the group S31, a nociceptive stimulus was performed by subdermal scratching at 30 minutes and, at 60 minutes, the “incision stimulus” was carried out in the same location (“nociceptive hyperstimulation”). The skin samples of the other animals were harvested from the back 1 minute after their death. SP, pro-CGRP and CGRP were quantified by Western Blotting. Results. The “incision stimulus” released SP, S1 compared to S0 (p <0.05) detected in the first minute, and the replenishment time was more than 30 minutes. Also, it cleaved pro-CGRP, S1 compared to S31 (p <0.05) in the first minute, and its replenishment time less than 30 minutes. Release of CGRP was not detected. Conclusion. The incision released SP already detected in the first minute; its replenishment time is more than 30 minutes. The incision decreased pro-CGRP, also detected in the first minute; and its replenishment time is less than 30 minutes.     

Secretion of adipocytes and macrophages under conditions of inflammation and/or insulin resistance and effect of adipocytes on preadipocytes under these conditions

The purpose of the present study was to examine changes in preadipocytes following the coculture of preadipocytes and adipocytes and the effects on the secretion of adipocytes and macrophages following induction of inflammation and insulin resistance. Mature adipocytes and RAW264.7 macrophages were treated with lipopolysaccharide and insulin to establish models of inflammation and insulin resistance, respectively. The mRNA expression levels of IL-6, MCP-1, and TNF-α in all adipocyte treatment groups were significantly greater compared with the control, and that of adiponectin was less (P < 0.05). In the RAW264.7 macrophages, the mRNA expression levels of IL-6 and TNF-α were greater than those in the control group (P < 0.05). Moreover, the results of this study confirmed that adipocytes and macrophages increased the secretion of inflammatory factors under conditions of induced inflammation and insulin resistance. In addition, 3T3-L1 adipocytes inhibited the proliferation and differentiation of preadipocytes when cocultured with adipocytes under conditions of inflammation and/or insulin resistance, and the phenotype of preadipocytes did not change.     

CD39/NTPDase-1 expression and activity in human umbilical vein endothelial cells are differentially regulated by leaf extracts from Rubus caesius and Rubus idaeus

Many experimental studies have demonstrated the favorable biological activities of plants belonging to the genus Rubus, but little is known of the role of Rubus leaf extracts in the modulation of the surface membrane expression and activity of endothelial apyrase. The aim of this study was to assess the influence of 1–15 μg/ml Rubus extracts on CD39 expression and enzymatic activity, and on the activation (ICAM-1 expression) and viability of human umbilical vein endothelial cells (HUVEC). The polyphenolic contents and antioxidative capacities of extracts from dewberry (R. caesius L.) and raspberry (R. idaeus L.) leaves were also investigated. The techniques applied were flow cytometry (endothelial surface membrane expression of ICAM-1 and CD39), malachite green assay (CD39 activity), HPLC-DAD (quantitative analysis of polyphenolic extract), ABTS, DPPH and FRAP spectrometric assays (antioxidant capacity), and the MTT test (cell viability). Significantly increased CD39 expressions and significantly decreased ATPDase activities were found in the cells treated with 15 μg/ml of either extract compared to the results for the controls. Neither of the extracts affected cell proliferation, but both significantly augmented endothelial cell ICAM-1 expression. The overall antioxidant capacities of the examined extracts remained relatively high and corresponded well to the determined total polyphenol contents. Overall, the results indicate that under in vitro conditions dewberry and raspberry leaf extracts have unfavorable impact on endothelial cells.