Salidroside Inhibits Reactive Astrogliosis and Glial Scar Formation in Late Cerebral Ischemia via the Akt/GSK-3β Pathway

Neurochemical Research - Cerebral ischemia leads to reactive astrogliosis and glial scar formation. Glial scarring can impede functional restoration during the recovery phase of stroke. Salidroside...     

BI1 alleviates cardiac microvascular ischemia-reperfusion injury via modifying mitochondrial fission and inhibiting XO/ROS/F-actin pathways

Pathogenesis of cardiac microvascular ischemia-reperfusion (IR) injury is associated with excessive mitochondrial fission. However, the upstream mediator of mitochondrial fission remains obscure. Bax inhibitor 1 (BI1) is linked to multiple mitochondrial functions, and there have been no studies investigating the contribution of BI1 on mitochondrial fission in the setting of cardiac microvascular IR injury. This study was undertaken to establish the action of BI1 on the cardiac microvascular reperfusion injury and figure out whether BI1 sustained endothelial viability via inhibiting mitochondrial fission. Our observation indicated that BI1 was downregulated in reperfused hearts and overexpression of BI1 attenuated microvascular IR injury. Mechanistically, reperfusion injury elevated the levels of xanthine oxidase (XO), an effect that was followed by increased reactive oxygen species (ROS) production. Subsequently, oxidative stress mediated F-actin depolymerization and the latter promoted mitochondrial fission. Aberrant fission caused mitochondrial dysfunction and ultimately activated mitochondrial apoptosis in cardiac microvascular endothelial cells. By comparison, BI1 overexpression repressed XO expression and thus neutralized ROS, interrupting F-actin-mediated mitochondrial fission. The inhibitory effect of BI1 on mitochondrial fission sustained endothelial viability, reversed endothelial barrier integrity, attenuated the microvascular inflammation response, and maintained microcirculation patency. Altogether, we conclude that BI1 is essential in maintaining mitochondrial homeostasis and alleviating cardiac microvascular IR injury. Deregulated BI1 via the XO/ROS/F-actin pathways plays a causative role in the development of cardiac microvascular reperfusion injury.     

Purification of a bioactive recombinant human Reg IV expressed in Escherichia coli

Regenerating gene (Reg) IV is a newly discovered member of the regenerating gene family belonging to the calcium (C-type) dependent lectin superfamily. Reg IV is highly expressed in the gastrointestinal tract and markedly up-regulated in colon adenocarcinoma, pancreatic cancer, gastric adenocarcinoma, and inflammatory bowel disease. However, the physiological and pathological functions of Reg IV are largely unknown, partly due to the limited access of the bioactive protein. We report here the first expression and purification of Reg IV proteins using a prokaryotic system. Human Reg IV was expressed in Escherichia coli as an insoluble protein which was identified in the fraction of inclusion body after ultrasonication of the bacteria. After the protein aggregate was solubilized by guanidine–HCl, it was refolded by sucrose and arginine-assisted procedures and purified using cation-exchange chromatography. The protein identity and purity of the final preparation were confirmed by analysis of the protein mass and immune specificity in SDS–PAGE, Western blotting, and HPLC assay. The biological activity of the protein was determined by the HCT116 and HT29 cell proliferation assays. The highly purified bioactive human Reg IV should aid in further characterization of its physiological and pathological functions.     

Expressions and purification of a mature form of recombinant human Chemerin in Escherichia coli

Chemerin is a novel chemokine that binds to the G protein-coupled receptor (GPCR) ChemR23, also known as chemokine-like receptor 1 (CMKLR1). It is secreted as a precursor and executes pro-inflammatory functions when the last six amino acids are removed from its C-terminus by serine proteases. After maturation, Chemerin attracts dendritic cells and macrophages through binding to ChemR23. We report a new method for expression and purification of mature recombinant human Chemerin (rhChemerin) using a prokaryotic system. After being expressed in bacteria, rhChemerin in inclusion bodies was denatured using 6 M guanidine chloride. Soluble rhChemerin was prepared by the protein-specific renaturation solution under defined conditions. It was subsequently purified using ion-exchange columns to more than 95% purity with endotoxin level <1.0 EU/μg. We further demonstrated its biological activities for attracting migration of human dendritic cells and murine macrophages in vitro using established chemotaxis assays.     

杨树基因组AMT转运蛋白的生物信息学特性

本研究中,通过隐马尔科夫模型(HMM)和杨树蛋白质库搜索,共找到17个杨树铵转运体蛋白(PtAMTs).利用生物信息学方法,我们对杨树家族17条AMT蛋白序列的系统发生和AMT基因组定位进行分析,然后对其氨基酸组成成分、理化性质以及二级结构进行预测和分析,同时还分析了杨树与拟南芥、水稻、番茄、百脉根和欧洲油菜的AMT基因家族之间的联系.二级结构预测结果发现不同成员间氨基酸数目、氨基酸序列间的疏水性存在一定的差异;α-螺旋和无规则卷曲为主要二级结构组成部分.同源性比对分析表明,PtAMT基因家族主要分为2个亚家族,AMT1 (11个成员)和AMT2 (6个成员),基因结果分析表明AMT2亚家族成员不含内含子.杨树AMT蛋白的亚细胞定位分析表明PtAMT主要定位于膜结构上.电子表达图谱分析结果表明:只有XP_002309151和 XP_002334025基因有对应的EST序列,并有相应的电子表达谱,并主要在花蕾表达.     

Fluvoxamine increased glutamate release by activating both 5-HT(3) and sigma-1 receptors in prelimbic cortex of chronic restraint stress C57BL/6 mice

Emerging evidence from therapeutic trials in humans and animal models suggests that in the treatment of depression, antidepressants play a role by targeting the glutamatergic system. Fluvoxamine is one of the widely used SSRIs which has been considered to target monoamine neurotransmitter reuptake mechanisms. However, whether fluvoxamine has an effect on the glutamate release is still unclear. The present experiment studied the effect of fluvoxamine on presynaptic glutamate release in prelimbic cortex, both in control C57BL/6 mice and chronic restraint stress C57BL/6 mice, and further investigated the mechanism underlying this effect by using patch clamp, on-line fluorimetry, pharmacological approaches combined with other techniques. The results showed that fluvoxamine increased the glutamate release in the depression model mice but it had no effect on the glutamate release in the control mice. The mechanism underlying these effects in depression model mice was that, fluvoxamine firstly activated presynaptic 5-HT(3) receptors, which transiently increased the Ca(2+) concentration. The increase of Ca(2+) concentration via 5-HT(3) receptors caused the activation of sigma-1 receptors, which were activated by fluvoxamine. The activation of sigma-1 receptors increased the intrasynaptosomal Ca(2+) concentration significantly through the outflow of endoplasmic reticulum calcium and finally activated PKC. These results suggested that fluvoxamine may have a selective effect and different mechanism based on the condition of animal.     

Expression and purification of recombinant human Mig in Escherichia coli and its comparison with murine Mig

Monokine induced by IFN-γ (Mig) is a member of CXC-chemokines and recruits T-lymphocytes to activate the immune response. In recent years, it has raised much interest in the areas of autoimmune disease and allograft rejection, as the production of recombinant human Mig (rHuMig) would be of considerable significance for both research and potential clinical use. Here we report the expression, preparation and characterization of non-tagged recombinant human Mig (rHuMig) using a prokaryotic expression system. Following expression in Escherichia coli (E. coli) BL21, the 103 amino acid residue of rHuMig was purified from bacteria inclusion bodies with a one-step S-Sepharose cation exchange chromatography. The product was immunologically characterized via Western blot and its purity was determined via SDS-PAGE and silver staining to be above 99%, with an endotoxin level <0.5EU/μg via a chemotaxis assay, rHuMig demonstrated chemotactic activity on mouse spleen lymphocytes with an ED50 of 15 ng/mL. Additionally, using a proliferation assay, rHuMig significantly inhibited proliferation of the human bladder cell line T24. In vivo experiments revealed that rHuMig could inhibit mouse bone marrow mononuclear cells cycling into the S-phase and reduced intestinal cell proliferation. Our results demonstrate that rHuMig is fully functional in the mouse model.     

Blood-based gene expression profiles models for classification of subsyndromal symptomatic depression and major depressive disorder

Subsyndromal symptomatic depression (SSD) is a subtype of subthreshold depressive and also lead to significant psychosocial functional impairment as same as major depressive disorder (MDD). Several studies have suggested that SSD is a transitory phenomena in the depression spectrum and is thus considered a subtype of depression. However, the pathophysioloy of depression remain largely obscure and studies on SSD are limited. The present study compared the expression profile and made the classification with the leukocytes by using whole-genome cRNA microarrays among drug-free first-episode subjects with SSD, MDD, and matched controls (8 subjects in each group). Support vector machines (SVMs) were utilized for training and testing on candidate signature expression profiles from signature selection step. Firstly, we identified 63 differentially expressed SSD signatures in contrast to control (P< = 5.0E-4) and 30 differentially expressed MDD signatures in contrast to control, respectively. Then, 123 gene signatures were identified with significantly differential expression level between SSD and MDD. Secondly, in order to conduct priority selection for biomarkers for SSD and MDD together, we selected top gene signatures from each group of pair-wise comparison results, and merged the signatures together to generate better profiles used for clearly classify SSD and MDD sets in the same time. In details, we tried different combination of signatures from the three pair-wise compartmental results and finally determined 48 gene expression signatures with 100% accuracy. Our finding suggested that SSD and MDD did not exhibit the same expressed genome signature with peripheral blood leukocyte, and blood cell-derived RNA of these 48 gene models may have significant value for performing diagnostic functions and classifying SSD, MDD, and healthy controls.     

Investigation of the cyt gene in Bacillus thuringiensis and the biological activities of Bt isolates from the soil of China

The presence of cyt genes was investigated in 80 type strains of Bacillus thuringiensis and 143 isolates obtained from soil samples of China by PCR amplification using two pairs of primers for the cyt1 and cyt2 genes. Three type strains of serotypes H11ac, H14 and H36, eight isolates belonging to H3, H14, H18 and H21, and one isolate of unknown serotype harbored cyt genes. We also tested the cytolytic activity for mammal cells, the hemolytic activity for sheep erythrocytes and insecticidal activity against mosquitoes of five isolates that contained cyt genes but did not belong to B. thuringiensis serovar israelensis. The protein profiles of the five isolates were different from those of the type strains of B. thuringiensis serovar israelensis, and among the five isolates, only Y-5 showed mosquitocidal activity against larvae of Culex quinquefasciatus. All five of the isolates exhibited hemolytic activity, but only three could cause the cell death of A549 cells. The cytopathological changes induced by NX-4 in some A549 cells were characterized with cell-ballooning.     

The 5-HTTLPR Confers Susceptibility to Anorexia Nervosa in Han Chinese: Evidence from a Case-Control and Family-Based Study

Accumulating evidence has implied that serotonin system dysfunction may be involved in the etiology of anorexia nervosa (AN). Serotonin-transporter-linked promoter region (5-HTTLPR) polymorphism is the genetic variant coding for the serotonin transporter and has a modulatory effect on its expression. This study aimed to investigate the possible association between the 5-HTTLPR and the susceptibility and severity of AN in Han Chinese using a case-control (255 patients and 351 controls) and family based study (198 trios). Eating disorder examination was used to measure the severity of AN behavioral symptoms. For the case-control study, the 5-HTTLPR showed significant association with AN in our sample (genotypic P = 0.03). The frequency of S allele was significantly higher in patients than that in controls (OR = 1.38, 95%CI: 1.06–1.79, P = 0.017). For the family-based study, the S allele of 5-HTTLPR was preferentially transmitted rather than non-transmitted from the parents to affected offspring (P = 0.013). The results of ANCOVA test revealed no significant association between the 5-HTTLPR polymorphism and severity of AN. Our findings suggested that 5-HTTLPR is able to confer susceptibility to AN in Han Chinese.