Knockdown of phosphotyrosyl phosphatase activator induces apoptosis via mitochondrial pathway and the attenuation by simultaneous tau hyperphosphorylation

Phosphotyrosyl phosphatase activator (PTPA) is decreased in the brains of Alzheimer's disease (AD) and the AD transgenic mouse models. Here, we investigated whether down‐regulation of PTPA affects cell viability and the underlying mechanisms. We found that PTPA was located in the integral membrane of mitochondria, and knockdown of PTPA induced cell apoptosis in HEK293 and N2a cell lines. PTPA knockdown decreased mitochondrial membrane potential and induced Bax translocation into the mitochondria with a simultaneous release of Cyt C, activation of caspase‐3, cleavage of poly (DNA ribose) polymerase (PARP), and decrease in Bcl‐xl and Bcl‐2 protein levels. Over‐expression of Protein phosphatase 2A (PP2A) catalytic subunit (PP2A_C) did not rescue the apoptosis induced by PTPA knockdown, and PTPA knockdown did not affect the level of and their phosphorylation of mitogen‐activated protein kinases (MAPKs), indicating that PP2A and MAPKs were not involved in the apoptosis induced by PTPA knockdown. In the cells with over‐expression of tau, PTPA knockdown induced PP2A inhibition and tau hyperphosphorylation but did not cause significant cell death. These data suggest that PTPA deficit causes apoptotic cell death through mitochondrial pathway and simultaneous tau hyperphosphorylation attenuates the PTPA‐induced cell death.

     

Characterization of the Grp94/OS-9 Chaperone–Lectin Complex

Grp94 is a macromolecular chaperone belonging to the hsp90 family and is the most abundant glycoprotein in the endoplasmic reticulum (ER) of mammals. In addition to its essential role in protein folding, Grp94 was proposed to participate in the ER-associated degradation quality control pathway by interacting with the lectin OS-9, a sensor for terminally misfolded proteins. To understand how OS-9 interacts with ER chaperone proteins, we mapped its interaction with Grp94. Glycosylation of the full-length Grp94 protein was essential for OS-9 binding, although deletion of the Grp94 N-terminal domain relieved this requirement suggesting that the effect was allosteric rather than direct. Although yeast OS-9 is composed of a well-established N-terminal mannose recognition homology lectin domain and a C-terminal dimerization domain, we find that the C-terminal domain of OS-9 in higher eukaryotes contains “mammalian-specific insets” that are specifically recognized by the middle and C-terminal domains of Grp94. Additionally, the Grp94 binding domain in OS-9 was found to be intrinsically disordered. The biochemical analysis of the interacting regions provides insight into the manner by which the two associate and it additionally hints at a plausible biological role for the Grp94/OS-9 complex.     

Integrating Multi-Omics for Uncovering the Architecture of Cross-Talking Pathways in Breast Cancer

Cross-talk among abnormal pathways widely occurs in human cancer and generally leads to insensitivity to cancer treatment. Moreover, alterations in the abnormal pathways are not limited to single molecular level. Therefore, we proposed a strategy that integrates a large number of biological sources at multiple levels for systematic identification of cross-talk among risk pathways in cancer by random walk on protein interaction network. We applied the method to multi-Omics breast cancer data from The Cancer Genome Atlas (TCGA), including somatic mutation, DNA copy number, DNA methylation and gene expression profiles. We identified close cross-talk among many known cancer-related pathways with complex change patterns. Furthermore, we identified key genes (linkers) bridging these cross-talks and showed that these genes carried out consistent biological functions with the linked cross-talking pathways. Through identification of leader genes in each pathway, the architecture of cross-talking pathways was built. Notably, we observed that linkers cooperated with leaders to form the fundamentation of cross-talk of pathways which play core roles in deterioration of breast cancer. As an example, we observed that KRAS showed a direct connection to numerous cancer-related pathways, such as MAPK signaling pathway, suggesting that it may be a central communication hub. In summary, we offer an effective way to characterize complex cross-talk among disease pathways, which can be applied to other diseases and provide useful information for the treatment of cancer.     

实时灰阶超声造影股浅静脉瓣膜返流时间与病变严重程度的相关性探讨

目的:探讨超声造影股浅静脉瓣膜返流与病变严重程度的关系。方法:回顾性分析55例63条诊断为原发性深静脉功能不全患者的临床和超声造影资料,记录股浅静脉瓣膜返流时间和CEAP临床表现分级,并进行两者的相关性分析。结果:统计学分析表明,CEAP临床表现分级与瓣膜返流时间无相关性;但溃疡肢体组与非溃疡肢体组的股浅静脉瓣膜返流时间比较,两组有显著性差异(P0.05)。结论:超声造影检查股静脉瓣膜返流时间有可能预测患肢静脉性溃疡的发生。     

综述: 自噬参与心脏疾病调控的研究进展

细胞自噬(autophagy)是将细胞内受损、变性或衰老的蛋白质以及细胞器运输到溶酶体内进行消化降解的过程．细胞自噬既是一种广泛存在的正常生理过程,又是细胞对不良环境的一种防御机制,参与多种疾病的病理过程．正常水平的自噬可以保护细胞免受环境刺激的影响,但自噬过度和自噬不足却可能导致疾病的发生．在心脏中,心肌细胞自噬对维持心肌功能具有重要的作用,自噬的异常可能导致各种心肌疾病如溶酶体储积症(Danon disease)等．各种心血管刺激如心肌缺血(ischemia)、再灌注(reperfusion)损伤、慢性缺氧(chronic hypoxia)等均可诱导心肌细胞自噬增强．而这些情况下心肌细胞自噬的作用还不清楚：它是否是一种潜在的细胞存活机制还是导致细胞死亡或疾病发生的病理性机制,或者是同时具有两种作用,目前还没有定论．心脏疾病是心肌功能出现异常时产生的各种病理状态的总称．在多种心脏疾病中,均伴随有心肌细胞自噬的改变,且影响着疾病的发生发展．在心肌肥厚(hypertrophic cardiomyopathy)中,细胞自噬程度降低而加剧心肌肥厚;在心力衰竭(heart failure,HF)中,细胞自噬增强可导致心肌细胞自噬性死亡;而在心肌梗死(myocardial infarction,MI)中,细胞自噬增强可减小梗死面积．但是细胞自噬在心脏疾病中到底扮演着怎样的角色,取决于细胞自噬发生的水平及病理状态．目前越来越多的人开始关注药物与细胞自噬调节之间的联系,且主要集中于抗肿瘤药物及心血管调节药物的研究．另外,有报道维生素类以及雌激素受体拮抗剂他莫西芬对细胞自噬也具有调节作用．研究心肌细胞自噬与心脏疾病的关系,以及药物对细胞自噬的调节,将有利于从自噬的角度探讨心脏疾病的发生发展过程及机制,开发出治疗心脏疾病的药物．     

Novel matrine derivative MD-1 attenuates hepatic fibrosis by inhibiting EGFR activation of hepatic stellate cells

Matrine (MT), the effective component of Sophora flavescens Ait, has been shown to have anti-inflammation, immune-suppressive, anti-tumor, and anti-hepatic fibrosis activities. However, the pharmacological effects of MT still need to be strengthened due to its relatively low efficacy and short half-life. In the present study, we report a more effective thio derivative of MT, MD-1, and its inhibitory effects on the activation of hepatic stellate cells (HSCs) in both cell culture and animal models. Cytological experiments showed that MD-1 can inhibit the proliferation of HSC-T6 cells with a half-maximal inhibitory concentration (IC50) of 62 μmol/L. In addition, MD-1 more strongly inhibits the migration of HSC-T6 cells compared to MT and can more effectively induce G0/G1 arrest and apoptosis. Investigating the biological mechanisms underlying anti-hepatic fibrosis in the presence of MD-1, we found that MD-1 can bind the epidermal growth factor receptor (EGFR) on the surface of HSC-T6 cells, which can further inhibit the phosphorylation of EGFR and its downstream protein kinase B (Akt), resulting in decreased expression of cyclin D1 and eventual inhibition of the activation of HSC-T6 cells. Furthermore, in rats with dimethylnitrosamine (DMN)-induced hepatic fibrosis, MD-1 slowed the development and progression of hepatic fibrosis, protecting hepatic parenchymal cells and improving hepatic functions. Therefore, MD-1 is a potential drug for anti-hepatic fibrosis.     

Pyrroloquinoline quinone inhibits oxygen/glucose deprivation-induced apoptosis by activating the PI3K/AKT pathway in cardiomyocytes

The purposes of this study were to examine the protective effect of pyrroloquinoline quinone (PQQ) on oxygen/glucose deprivation (OGD)-induced injury to H9C2 rat cardiomyocytes and to investigate the mechanism. Using H9C2 cells cultured in vitro, we examined changes in cell viability with an MTT assay at 12, 24, and 48 h after injury induced by OGD. Various concentrations of PQQ (1, 10, and 100 μM) were added, and the effect of PQQ on cell viability after OGD was assessed using the MTT assay. Thus, the optimal concentration of PQQ for the protection of cardiomyocytes against oxygen and glucose deprivation injury was determined. We also used flow cytometry analysis to examine the effect of PQQ on H9C2 cells with OGD-induced injury. The molecular probe 2′,7′-dichlorofluorescin diacetate was used to label the H9C2 cells, and flow cytometry was used to detect the effect of PQQ on reactive oxygen species (ROS) content. After labeling the H9C2 cells using a mitochondrial green fluorescent probe (Mito-Tracker Green), we measured the change in the mitochondrial content of PQQ-treated H9C2 cells. Western blotting was used to examine the effect of PQQ on the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in the H9C2 cells. The results of the MTT assay showed that 48 h of OGD significantly injured the H9C2 cells (p < 0.01) and that treatment with 100 μM PQQ effectively decreased the level of OGD-induced injury (p < 0.01). The results of the flow cytometry analysis showed that PQQ significantly reduced apoptosis in H9C2 cells subjected to OGD (p < 0.05). In addition, OGD significantly increased the ROS level in H9C2 cells (p < 0.01), and PQQ significantly inhibited this increase (p < 0.05). The results of the Mito-Tracker Green staining suggested that PQQ effectively inhibited the decrease in mitochondrial content caused by OGD (p < 0.05). Western blot analysis showed that PQQ partially reversed the decrease in Akt phosphorylation that was caused by OGD (p < 0.05). PQQ treatment dose-dependently protects H9C2 cells from OGD-induced injury by reducing apoptosis, decreasing intracellular ROS levels, and rescuing the OGD-induced decrease in mitochondrial content. The protective effect of PQQ may be related to its effects on the PI3K/Akt pathway.     

A Multi-Area Stochastic Model for a Covert Visual Search Task

Decisions typically comprise several elements. For example, attention must be directed towards specific objects, their identities recognized, and a choice made among alternatives. Pairs of competing accumulators and drift-diffusion processes provide good models of evidence integration in two-alternative perceptual choices, but more complex tasks requiring the coordination of attention and decision making involve multistage processing and multiple brain areas. Here we consider a task in which a target is located among distractors and its identity reported by lever release. The data comprise reaction times, accuracies, and single unit recordings from two monkeys’ lateral interparietal area (LIP) neurons. LIP firing rates distinguish between targets and distractors, exhibit stimulus set size effects, and show response-hemifield congruence effects. These data motivate our model, which uses coupled sets of leaky competing accumulators to represent processes hypothesized to occur in feature-selective areas and limb motor and pre-motor areas, together with the visual selection process occurring in LIP. Model simulations capture the electrophysiological and behavioral data, and fitted parameters suggest that different connection weights between LIP and the other cortical areas may account for the observed behavioral differences between the animals.     

A facile method for controlling the reaction equilibrium of sphingolipid ceramide N-deacylase for lyso-glycosphingolipid production

Lyso-glycosphingolipids (lyso-GSLs), the N-deacylated forms of glycosphingolipids (GSLs), are important synthetic intermediates for the preparation of GSL analogs. Although lyso-GSLs can be produced by hydrolyzing natural GSLs using sphingolipid ceramide N-deacylase (SCDase), the yield for this reaction is usually low because SCDase also catalyzes the reverse reaction, ultimately establishing an equilibrium between hydrolysis and synthesis. In the present study, we developed an efficient method for controlling the reaction equilibrium by introducing divalent metal cation and detergent in the enzymatic reaction system. In the presence of both Ca²⁺ and taurodeoxycholate hydrate, the generated fatty acids were precipitated by the formation of insoluble stearate salts and pushing the reaction equilibrium toward hydrolysis. The yield of GM1 hydrolysis can be achieved as high as 96%, with an improvement up to 45% compared with the nonoptimized condition. In preparative scale, 75 mg of lyso-GM1 was obtained from 100 mg of GM1 with a 90% yield, which is the highest reported yield to date. The method can also be used for the efficient hydrolysis of a variety of GSLs and sphingomyelin. Thus, this method should serve as a facile, easily scalable, and general tool for lyso-GSL production to facilitate further GSL research.