石蜡油对离子注入动物细胞真空环境的保护效应初探

利用石蜡油的高效护水功能进行动物细胞(HeLa细胞)的真空及注入保护,研究了不同的预处理时间对细胞活性的影响,并将石蜡油浸泡细胞2 m in~6 m in选为最佳保护时间。当细胞经过2 m in~6 m in的预处理后再进行真空、注入,结果显示成活率大幅度提高。     

Skp2-mediated degradation of p27 regulates progression into mitosis

Although Skp2 has been thought to mediate the degradation of p27 at the G(1)-S transition, Skp2(-/-) cells exhibit accumulation of p27 in S-G(2) phase with overreplication. We demonstrate that Skp2(-/-)p27(-/-) mice do not exhibit the overreplication phenotype, suggesting that p27 accumulation is required for its development. Hepatocytes of Skp2(-/-) mice entered the endoduplication cycle after mitogenic stimulation, whereas this phenotype was not apparent in Skp2(-/-)p27(-/-) mice. Cdc2-associated kinase activity was lower in Skp2(-/-) cells than in wild-type cells, and a reduction in Cdc2 activity was sufficient to induce overreplication. The lack of p27 degradation in G(2) phase in Skp2(-/-) cells may thus result in suppression of Cdc2 activity and consequent inhibition of entry into M phase. These data suggest that p27 proteolysis is necessary for the activation of not only Cdk2 but also Cdc2, and that Skp2 contributes to regulation of G(2)-M progression by mediating the degradation of p27.     

ASC is a Bax adaptor and regulates the p53-Bax mitochondrial apoptosis pathway

The apoptosis-associated speck-like protein (ASC) is an unusual adaptor protein that contains the Pyrin/PAAD death domain in addition to the CARD protein-protein interaction domain. Here, we present evidence that ASC can function as an adaptor molecule for Bax and regulate a p53-Bax mitochondrial pathway of apoptosis. When ectopically expressed, ASC interacted directly with Bax, colocalized with Bax to the mitochondria, induced cytochrome c release with a significant reduction of mitochondrial membrane potential and resulted in the activation of caspase-9, -2 and -3. The rapid induction of apoptosis by ASC was not observed in Bax-deficient cells. We also show that induction of ASC after exposure to genotoxic stress is dependent on p53. Blocking of endogenous ASC expression by small-interfering RNA (siRNA) reduced the apoptotic response and inhibited translocation of Bax to mitochondria in response to p53 or genotoxic insult, suggesting that ASC is required to translocate Bax to the mitochondria. Our findings demonstrate that ASC has an essential role in the intrinsic mitochondrial pathway of apoptosis through a p53-Bax network.     

免疫法筛选酵母HSA-IL-11融合蛋白转化子

报道了一种筛选高表达融合蛋白HSA-IL-11的毕赤酵母转化子的免疫双膜筛选法.将生长在醋酸纤维素滤膜上的转化子进行原位诱导,再用硝酸纤维素滤膜对表达的蛋白进行原位捕捉,并经封闭过夜后使用抗HSA抗体进行免疫杂交,再用标记二抗进行显色.根据显色强弱将转化子分为强阳性、中等和阴性三类,再用抗IL-11抗体进行复筛验证.结...     

创伤患者急性肾功能损伤诊断与治疗

目的:探讨患者创伤后发生急性肾损伤的患者发病率、临床特点以及发病危险因素,以便有效预防和及早治疗。方法:回顾性分析我院重症监护室2004年1月至2010年12月收治的创伤患者相关临床资料,分析创伤后急性肾损伤的发病率以及发病危险因素。结果:共有106例患者纳入我们的研究,其中47例患者创伤后并发急性肾损伤。在发生急性肾损伤患者中,平均年龄为31±19岁,84.6%为男性;其中25例为脓毒血症引起,18例是因为低血压导致急性肾功能损伤。所有患者中,24例患者出现了少尿的症状,19例患者进行了透析治疗。腹部外伤[(OR)=3.66,P=0.027]和应用呋塞米[(OR=4.10,P=0.026)]是发生急性肾损伤的危险因素。结论:急性肾损伤时创伤后的严重并发症之一,死亡率高。只有找到创伤后发生急性肾损伤的危险因素,才能有效预防和及早治疗。     

ABCA1基因多态性R219K与帕金森症和阿尔兹海默症发病率的相关性研究

目的:探讨与研究三磷酸腺苷结合盒转运体A1 (Adenosine triphosphate (ATP)-binding cassette transporter A1)基因多态性R219K与帕金森症(Parkinson disease,PD)和阿尔兹海默症(Alzheimer disease,AD)发病率的相关性。方法:选择2016年2月到2019年8月在本院门诊与住院的帕金森症患者42例作为PD组,同期选择本院门诊与住院的阿尔兹海默症患者42例作为AD组,同期选择本院门诊健康体检者84例作为对照组。调查入选者的一般资料,检测三组血液样本的ABCA1基因多态性R219K情况并进行相关性分析。结果:AD组低密度脂蛋白(low-density lipoprotein,LDL-C)、总胆固醇(total cholesterol,TC)、甘油三酯(triglyceride,TG)与尿酸(Uric acid,UA)均低于对照组,而高密度脂蛋白(high-density lipoprotein,HDL-C)、同型半胱氨酸(homocysteine,Hcy)值高于对照组(P0.05);AD组TC均低于PD组,而HDL高于PD组。PD患者HDL-C均低于对照组,而LDL、TC和TG与对照组无差异(P0.05),三组空腹血糖(Fasting blood glucose,FBG)值对比差异无统计学意义(P0.05)。PD组与AD组的ABCA1 R219K GA基因型、A等位基因频率都显著高于对照组(P0.05),PD组与AD组对比差异无统计学意义(P0.05)。在168例入选者中,直线相关分析显示ABCA1 R219K GA基因型与A等位基因与帕金森症或阿尔兹海默症发生有显著相关性(P0.05)。结论:ABCA1基因多态性R219K在帕金森症和阿尔兹海默症患者中比较常见,ABCA1 R219K GA基因型与A等位基因可诱发帕金森症和阿尔兹海默症的发生。     

Diversity of culturable bacterial communities in the intestinal tracts of goldfish (<Emphasis Type="Italic">Carassius auratus</Emphasis>) and their ability to produce <Emphasis Type="Italic">N</Emphasis>-acyl homoserine lactone

Intestinal bacteria isolated from goldfish (Carassius auratus) were identified based on 16 ribosomal RNA (rRNA) gene sequences and screened for their ability to produce N-acyl homoserine lactone (AHL), an autoinducer of the quorum sensing (QS) system. The 230 aerobes/facultative anaerobes that were isolated comprised members of the genera Aeromonas (184 isolates), Citrobacter (11), Enterobacter (2), Shewanella (28), Vagococcus (1), and Vibrio (4). Among these genera, the two most abundant species were Aeromonas veronii (163 isolates) and Shewanella xiamenensis (27). In addition, 142 obligate anaerobes consisting of Cetobacterium somerae (139 isolates), Clostridium frigidicarnis (2), and Cetobacterium sp. (1) were also isolated. One hundred seventy isolates (74.2%) belonging to the genera Aeromonas, Citrobacter, Enterobacter, Shewanella, and Vibrio produced AHL, while 155 (67.7%) and 91 (39.7%) isolates possessed the luxR and luxI gene homologs, respectively. None of the obligate anaerobes produced AHL or possessed luxRI homologs. Total viable counts ranged from 1.2 × 10⁷ to 2.2 × 10⁹ CFU/g, which were accounted for 0.8 to 15.2% of direct counts. Aeromonas veronii, S. xiamenensis, and C. somerae were detected from five goldfish at densities ranging from 4.0 × 10⁶ to 1.7 × 10⁹ CFU/g, indicating that these bacteria are dominant components of the culturable gut flora in goldfish. In addition, members of the genera Aeromonas and Shewanella appeared to communicate with each other by using the QS system to some extent when the concentration of AHL reaches a certain threshold. It is therefore suggested that bacteria with the ability to disrupt AHL secretion in intestinal environments are potential candidates for probionts for preventing opportunistic infections in freshwater fish such as goldfish.     

定向进化提高灰盖鬼伞过氧化物酶染织废水脱色效率

【目的】获得染织废水脱色能力增强的灰盖鬼伞过氧化物酶。【方法】使用基因合成及定点突变平台合成突变灰盖鬼伞过氧化物酶基因CIPmt4(I49S、V53A、M166F和M242I),并调整密码子至毕赤酵母偏好性。以CIPmt4为模板进行定向进化,经过三轮易错PCR和高通量筛选得到一个酶学性质显著改善的突变体(CIPmt5)。通过3D建模和分子动力学模拟分析蛋白的结构及热稳定性,并进一步研究CIPmt5和野生型CIP对刚果红、氨基黑、甲基橙、次甲基蓝、苯胺蓝、结晶紫、溴酚蓝共7种染料的脱色能力。【结果】序列分析显示该突变体积累了I49S、V53A、T121A、M166F和Y272F共5个氨基酸突变,与野生型灰盖鬼伞过氧化物酶相比,以ABTS为底物酶活性是野生型的2.01倍(24.44 U/mg),最适反应p H由5.0提高到6.5,最适反应温度由25°C提高到45°C。除次甲基蓝外对其它染料脱色的最适p H都往中、碱性方向偏移,脱色率普遍高于野生型。模型分析显示CIPmt5活性中心更开放,热稳定性增强。【结论】突变体酶CIPmt5能够更好地替代野生型灰盖鬼伞过氧化物酶应用于染织业染料脱色、化工废水和染织废水的生物修复。