A proteomic approach in investigating the hepatoprotective mechanism of Schisandrin B: role of raf kinase inhibitor protein

To identify key proteins involved in the hepatoprotection afforded by schisandrin B (Sch B), we used a proteomic approach to screen proteins that were specifically regulated by Sch B in mouse livers and to investigate the role of the proteins in hepatoprotection. Thirteen proteins were specifically activated or suppressed by Sch B treatment. Among the 13 proteins, Raf kinase inhibitor protein (RKIP) was postulated to be the key regulator involved in the development of hepatotoxin-induced cellular damage. The results indicated that the downregulation of RKIP by antisense RKIP vector transfection led to the activation of the Raf-1/MEK/ERK signaling pathway, as evidenced by increases in the level of MEK/ERK phosphorylation and the level of nuclear factor erythroid 2-related factor 2 in the nucleus. The signaling effect produced by RKIP downregulation resembled that triggered by Sch B, wherein both treatments resulted in a decrease in the extent of carbon tetrachloride-induced apoptotic cell death in AML12 hepatocytes. Overexpression of RKIP by the sense RKIP transfection vector or the inhibition of MEK kinase by PD98059 was able to abrogate the cytoprotective effect of Sch B in the hepatocytes. The results indicate that Sch B triggers the Raf/MEK/ERK signaling pathway, presumably by downregulating RKIP, thereby protecting against carbon tetrachloride-induced cytotoxicity.     

Decoy engineering of the receptor-like cytoplasmic kinase StPBS1 to defend against virus infection in potato

Potato virus Y (PVY) is an important pathogen of potato (Solanum tuberosum). Although the PBS1–RPS5 immune system is well documented in Arabidopsis thaliana, it has not been reported in potato. In Arabidopsis, the bacterial effector AvrPphB cleaves AtPBS1 to trigger an immune response. Here, we show that the AvrPphB-triggered immune response is mediated by StPBS1, a close homologue of AtPBS1 in potato. However, downstream signalling of StPBS1 was mediated by unknown resistance (R) proteins other than potato orthologues of AtRPS5 and HvPBR1, which is important for HvPBS1 signalling in barley. Immune signalling of StPBS1 is mediated by the AvrPphB C-terminal cleavage domain and an STKPQ motif, in contrast to AtPBS1-mediated immunity in which both AvrPphB cleavage fragments and an SEMPH motif are essential. The cleavage sequence of AvrPphB in StPBS1 was replaced with that of the PVY NIa-Pro protease to obtain StPBS1^NIa. StPBS1^NIa overexpression potato displayed stronger immunity to PVY infection than did the StPBS1 transgenic lines. StPBS1^NIa was cleaved at the expected target site by NIa-Pro protease from PVY. Thus, we characterized the function of StPBS1 in potato immunity and provide a biotechnology control method for PVY via transformation of decoy-engineered StPBS1^NIa.     

检测乙肝表面抗原夹心法中使用不同抗体的对比配伍研究

在提高乙型肝炎表面抗原（ＨＢｓＡｇ）酶联免疫诊断盒灵敏度的研究中对包被抗体、酶标记用抗体作了大量的筛选、对比实验。结果表明,包被单克隆抗体配伍酶标记山羊或豚鼠多克隆抗体可使乙肝表面抗原检测灵敏度达到０．２ｎｇ,超过中国药品生物制品检定所要求的１ｎｇ批批检合格标准,同时,特异性及变异系数均合乎要求,从而提高了试剂盒的质量。这一研究结果对提高其它以夹心法为原理的检测灵敏度有重要意义。     

全雌系苦瓜ACC氧化酶基因cDNA克隆及序列分析

为研究1-氨基环丙烷-1-羧酸氧化酶(ACC氧化酶,ACO)基因在苦瓜性别分化中的作用,以全雌系苦瓜‘X-Hei-d-d’花蕾为试材,采用RT-PCR和RACE技术获得了ACC氧化酶基因(Mc-ACO1)的全长cDNA序列。该序列为1 137 bp(GenBank登录号:FJ459813),其完整开放阅读框长1 005 bp,编码334个氨基酸,预测分子量为37.30 kD,肽链N端缺失ACOs家族典型的1个保守区和2个保守二价铁离子/抗坏血酸依赖型双加氧酶氨基酸残基。序列分析表明,植物ACO基因的演化与其来源植物亲缘关系的一致性较高;与其他物种相比,苦瓜Mc-ACO1不同的氨基酸序列主要表现在肽链N端;苦瓜Mc-ACO1应属于黄瓜Cs-ACO2类成员,功能可能与性别分化相关。     

Diversity and dynamics of the bacterial community involved in pig manure biodegradation in a microbial fermentation bed system

Overproduction of livestock manures with unpleasant odors causes significant environmental problems. The microbial fermentation bed (MFB) system is considered an effective approach to recycling utilization of agricultural byproducts and pig manure (PM). To gain a better understanding of bacterial communities present during the degradation of PM in MFB, the PM bacterial community was evaluated at different fermentation stages using 16S rRNA high throughput sequencing technology. The heatmap plot clustered five samples into short-term fermentation stage of 0–10 days and long-term fermentation stage of 15–20 days. The most abundant OTUs at the phylum level were Firmicutes, Actinobacteria and Proteobacteria in the long-term fermentation stage of PM, whereas Firmicutes, Bacteroidetes, and Proteobacteria predominated in the short-term fermentation stage of PM. At the genus level, organic degradation strains, such as Corynebacterium, Bacillus, Virgibacillus, Pseudomonas, Actinobacteria, Lactobacillus, Pediococcus were the predominate genera at the long-term fermentation stage, but were found only rarely in the short-term fermentation stage. C/N ratios increased and the concentration of the unpleasant odor substance 3-hydroxy-5-methylisoxazole (3-MI) decreased with prolonged period of fermentation. Redundancy analysis (RDA) demonstrated that the relative abundance of Firmicutes, Actinobacteria, Acidobacteria and Proteobacteria had a close relationship with degradation of 3-MI and increasing C/N ratio. These results provide valuable additional information about bacterial community composition during PM biodegradation in animal husbandry.     

Diazoxide Pretreatment Prevents Aβ1–42 Induced Oxidative Stress in Cholinergic Neurons Via Alleviating NOX2 Expression

The aggregation and accumulation of amyloid-β (Aβ) plays a significant role in the pathogenesis of Alzheimer’s disease. Aβ is known to increase free radical production in neuronal cells, leading to oxidative stress and cell death. Diazoxide (DZ), a highly selective drug capable of opening mitochondrial ATP-sensitive potassium channels, has neuroprotective effects against neuronal cell death. However, the mechanism through which DZ protects cholinergic neurons against Aβ-induced oxidative injury is still unclear. The present study was designed to investigate the effects of DZ pretreatment against Aβ_1–42 induced oxidative damage and cytotoxicity. Through measures of DZ effects on Aβ_1–42 induced cellular damage, reactive oxygen species (ROS) and MDA generation and expressions of gp91phox and p47phox in cholinergic neurons, new insights into the neuroprotective mechanisms can be derived. Aβ_1–42 significantly decreased 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide levels and increased ROS and MDA production; all effects were attenuated by pretreatment with DZ or diphenyleneiodonium chloride (a NOX2 inhibitor). Pretreatment with DZ also attenuated the upregulation of NOX2 subunits (gp91phox and p47phox) induced by Aβ_1–42. Since NOX2 is one of the main sources of free radicals, these results suggest that DZ can counteract Aβ_1–42 induced oxidative stress and associated cell death by reducing the level of ROS and MDA, in part, by alleviating NOX2 expression.     

柱前衍生化HPLC法测定黄芪中黄芪甲苷的含量

目的：建立黄芪中黄芪甲苷的柱前衍化高效液相色谱测定法,并用该法对不同产地黄芪中黄芪甲苷的含量进行比较,方法：以吡啶－苯甲酰氯（２．５：１）为衍生化试剂,对黄芪甲苷分子中的羟基进行苯甲酰化,以甲醇－四氢呋喃－水（９０：４：６,０．２％三乙胺）为流动相,ＶＤ３为内标物,在２３０ｎｍ波长处检测。结果黄芪甲苷在０．００４－０．０８０ｍｇ．ｍＬ＾－１范围内呈线性,相关系数为０．９９９９,平均回收率为９４．７     

低氧对肺动脉内皮细胞PDGF—B链释放及平滑肌细胞生长的影响

应用蛋白ｄｏｔｂｌｏｔ技术检测了低氧内皮细胞条件培养液（ＨＥＣＣＭ）和常氧内皮细胞条件培养液（ＮＥＣＣＭ）内ＰＤＧＦ相对含量,并利用［３Ｈ］－ＴｄＲ掺入法和流式细胞术观察了ＨＥＣＣＭ和ＮＥＣＣＭ及加入特异ＰＤＧＦ抗体对肺动脉平滑肌细胞（ＰＡＳＭＣ）生长的影响。结果表明,ＨＥＣＣＭ中的ＰＤＧＦ含量明显高于ＮＥＣＣＭ;ＨＥＣＣＭ能明显增强ＰＡＳＭＣ内ＤＮＡ合成,促进ＰＡＳＭＣ从Ｇｏ／Ｇ１期进入Ｓ期;当预先加入ＰＤＧＦ－Ｂ链抗体时,则会明显地抑制ＨＥＣＣＭ对ＰＡＳＭＣ的ＤＮＡ合成,阻止ＰＡＳＭＣ从Ｇｏ／Ｇ１期进入Ｓ期。结果提示,低氧时ＰＡＳＭＣ增殖与肺动脉内皮细胞分泌释放ＰＤＧＦ增加有关