Construction of recombinant plasmids containing rat thyroglobulin mRNA sequences

Two plasmids containing rat thyroglobulin cDNA sequences have been constructed and characterized. A plasmid with a 500-bp insert (pRT6) was isolated and identified as thyroglobulin-specific on the basis of the tissue specificity of the inserted sequence and of its ability to retain thyroglobulin mRNA on a nitrocellulose filter. The cDNA insert in pRT6 was subsequently used to screen a rat thyroid cDNA library constructed with large cDNA. A plasmid was found containing a 1700-bp insert. The polarity and the fidelity of the insert is demonstrated by S1 mapping.     

A critical role of protein-bound water in the catalytic cycle of cytochrome P-450 camphor.

The rates of NADH oxidation during the hydroxylation of camphor by cytochrome P-450cam were followed in the presence of co-solvents used to increase the osmotic pressure surrounding the protein-bound water. As a result, the measured Vmax decreases independently of the perturbant tested. Roughly 28 molecules of water, involved during the catalytic cycle, are deduced from the variation of Vmax as a function of osmotic pressure. These molecules, in part, could be those present in the cytochrome P-450cam-putidaredoxin interface.     

紫背金盘(Ajuga nipponensis)次生物质对桔全爪螨(Panonychus citri)的驱避作用

研究了紫背金盘Ajuga nipponensis Makino各溶剂提取物和部分化合物对桔全爪螨Panonychus citri McGregor雌成螨及其产卵的驱避作用.结果表明,石油醚萃取物、乙酸乙酯萃取物具有较强的生物活性.在0.1 g · L-1时, 石油醚和乙酸乙酯萃取物对该螨处理1d后的产卵忌避率分别为:84.86%、69.88%;2d后为89.49%、82.19%;对雌成螨驱避率分别为:85.08%、68.66%;2d后为50.96%、69.84%.乙酸乙酯萃取物经分离得到四类化合物,结果表明:馏分Ⅰ为长链脂肪酸混合物,具有较强生物活性,2000μg/ml和1000μg/ml处理1d后,产卵忌避率分别为:80.77%、74.77%;2d后为73.81%、72.59%.2000μg/ml处理1d后对雌成螨的驱避率为:69.88%;2d后为74.24%.刺槐素Ⅱ、新克罗烷化合物Ⅲ和β-蜕皮甾酮Ⅳ在2000μg/ml均不表现活性.对馏分Ⅰ中的4个主要化合物单体进行活性测定,结果表明:十六烷酸、十六烷酸甲酯、十六烷酸乙酯和十八烷酸甲酯在2000μg/ml处理时,1d后,产卵驱避率分别为:75.18%、61.76%、59.18%和66.49%;2d后产卵驱避率为:66.67%、31.15%、46.75%和44.84%;雌成螨驱避率分别为:1d后,67.53%、63.79%、59.26%和68.00;2d后,67.23%、43.96%、48.23%和64.19%.在1000μg/ml处理时,1d 后,产卵驱避率分别为:59.21%、59.16%、57.02%和61.40%;1d后,雌成螨驱避率分别为:69.64%、61.43%、55.76%和64.00%.     

RBM10 inhibits cell proliferation of lung adenocarcinoma via RAP1/AKT/CREB signalling pathway

Initial functional studies have demonstrated that RNA‐binding motif protein 10 (RBM10) can promote apoptosis and suppress cell proliferation; however, the results of several studies suggest a tumour‐promoting role for RBM10. Herein, we assessed the involvement of RBM10 in lung adenocarcinoma cell proliferation and explored the potential molecular mechanism. We found that, both in vitro and in vivo, RBM10 overexpression suppresses lung adenocarcinoma cell proliferation, while its knockdown enhances cell proliferation. Using complementary DNA microarray analysis, we previously found that RBM10 overexpression induces significant down‐regulation of RAP1A expression. In this study, we have confirmed that RBM10 decreases the activation of RAP1 and found that EPAC stimulation and inhibition can abolish the effects of RBM10 knockdown and overexpression, respectively, and regulate cell growth. This effect of RBM10 on proliferation was independent of the MAPK/ERK and P38/MAPK signalling pathways. We found that RBM10 reduces the phosphorylation of CREB via the AKT signalling pathway, suggesting that RBM10 exhibits its effect on lung adenocarcinoma cell proliferation via the RAP1/AKT/CREB signalling pathway.     

光动力疗法治疗感染性疾病的动物模型

光动力疗法(photodynamic therapy,PDT)是利用特定波长的激发光照射生物靶标上的光敏剂,从而产生活性氧并有效杀伤多种耐药病原体的新型治疗方式,具有作用广、安全可控、不易耐受等优点。大量体外实验已证实了PDT疗效,但目前动物实验数据较少,且治疗参数不一,一定程度上影响了PDT在临床治疗中的广泛应用。本文综述近年来PDT用于体内抗感染治疗的动物模型构建、治疗方案设计等方面的研究进展,为未来PDT抗感染研究及临床应用提供参考。     

卵巢上皮癌中ING4 基因启动子的甲基化状态及其临床意义

目的：探讨卵巢上皮癌中ING4 基因启动子的甲基化状态及其临床意义。方法：收集2005 年7 月至2012 年6 月哈尔滨医科 大学附属第一医院行全面分期手术并经病理检查确诊的150 例卵巢上皮癌组织标本,并以同期因子宫肌瘤或子宫腺肌症行子宫 全切除术或次全切除术并经病理检查确诊为正常卵巢组织的150 例标本作为对照组。采用甲基化特异性PCR（MSP）技术检测卵 巢上皮癌组织与正常卵巢组织中ING4 基因启动子的甲基化状态,蛋白印迹法检测ING4 蛋白的表达,并分析ING4 基因启动子 的甲基化状态与卵巢上皮癌临床病例特征的关系。结果：卵巢上皮癌组织中ING4 基因启动子的甲基化阳性率为42.7%（64/150）, 明显高于正常卵巢组织（4%,6/150）,差异有统计学意义（P<0.05）。ING4 基因启动子甲基化阳性的卵巢上皮癌组织中ING4蛋白 表达阴性或弱阳性;ING4 基因启动子甲基化阴性的卵巢上皮癌和正常卵巢组织中ING4 蛋白表达阳性;在64 例ING4 基因启动 子甲基化的卵巢上皮癌组织中,ING4 蛋白表达强度与ING4 基因启动子的甲基化程度呈负相关（r=-0.435,P<0.05）。卵巢上皮癌 组织中,ING4 基因甲基化的阳性率随着手术病理分期和组织学分级的增加而增加（P<0.05）;卵巢透明细胞癌（55.6%,10/18）和卵 巢子宫内膜样癌（59.3%,16/27）中ING4 基因甲基化的阳性率显著高于浆液性囊腺癌（33.9%,20/59）和粘液性囊腺癌（39.1%, 18/46）（P<0.05）;ING4基因启动子的甲基化状态与患者的年龄、有无腹水及淋巴结转移均无显著相关性（P>0.05）。结论：ING4 基 因启动子的甲基化可能促进了其在卵巢上皮癌组织中的表达失活,进而促进了卵巢上皮癌的生长和分化。     

Sex determination of buffalo embryos (Bubalus bubalis) by polymerase chain reaction

The aim of this study was to identify a simple, rapid method for sex determination of in vitro produced buffalo embryos, amplifying Y-chromosome-specific repeat sequences by polymerase chain reaction (PCR). Buffalo oocytes collected from slaughtered animals were matured, fertilised and cultured in vitro for 7 days. On day 7 embryos were evaluated and divided in to six groups according to developmental stage (2, 4, 8, 16 cells, morulae and blastocyst). Each embryo was stored singly in phosphate-buffered saline at -20 degrees C until PCR. Two different methods of extraction of DNA were compared: a standard procedure (ST), using a normal extraction by phenol-chloroform, isoamyl alcohol and final precipitation in absolute ethanol and a direct procedure (DT), using a commercial kit (Qiaquik-Qiagen mini blood). A pair of bovine satellite primers and two pairs of different bovine Y-chromosome-specific primers (BRY4.a and BRY.1) were used in the PCR assay on embryos and on whole blood samples collected from male and female adult buffaloes, used as control. The trial was carried out on 359 embryos (193 for ST and 166 for DT). When DNA samples from blood were amplified, the sex determined by PCR always corresponded to the anatomical sex. Embryo sexing was not possible in two embryos in ST and one embryo in DT. Both extraction protocols recovered sufficient quantities of target DNA at all developmental stages, but the time required for the ST (24 h) limits its use in embryo sexing and supports the use of commercial extraction kits (5 h).     

Development of a new class of potential antiatherosclerosis agents: NO-donor antioxidants

A new class of NO-donor phenol derivatives is described. The products were obtained by joining appropriate phenols with either nitrooxy or 3-phenylsulfonylfuroxan-4-yloxy moieties. All the compounds proved to inhibit the ferrous salt/ascorbate induced lipidic peroxidation of membrane lipids of rat hepatocytes. They were also capable of dilating rat aorta strips precontracted with phenylephrine.     

Therapeutics development for triplet repeat expansion diseases

The underlying genetic mutations for many inherited neurodegenerative disorders have been identified in recent years. One frequent type of mutation is trinucleotide repeat expansion. Depending on the location of the repeat expansion, the mutation might result in a loss of function of the disease gene, a toxic gain of function or both. Disease gene identification has led to the development of model systems for investigating disease mechanisms and evaluating treatments. Examination of experimental findings reveals similarities in disease mechanisms as well as possibilities for treatment.