小型猪冠脉支架模型中即刻OCT检查评价支架贴壁不良的安全性和有效性

目的评价中华小型猪冠状动脉支架植入术后即刻行光学相干断层成像扫描（OCT）的安全性和有效性。方法健康中华小型猪22只,经股动脉途径行右冠状动脉西罗莫司药物洗脱支架植入术,术后即刻行OCT检查评价支架贴壁不良情况,同时记录OCT检查时间和相关并发症。结果 OCT共检查25段支架,失败3例（1例指引导丝断裂于支架内,2例发生冠脉痉挛导致阻断球囊送入困难未能成像）,OCT检查并发症包括一过性ST段抬高（5例）和室颤（3例）,死亡1例,平均手术时间49.7±12.9 min,其中OCT检查耗时17.9±4.9 min,OCT共检测522 mm支架,定量测量4514个支架丝,其中66个存在贴壁不良,即刻支架贴壁不良率1.46%。结论利用冠脉内OCT技术可以有效评价支架术后即刻贴壁不良情况,安全性良好。     

大豆GmOLPa基因的蛋白序列分析及原核表达

以盐诱导后的大豆根为材料,提取总RNA,RT-PCR得到GmOLPa目的片段。对其蛋白序列分析表明,该蛋白属于GH64-TLP-SF同源超家族中的PR-5蛋白,第25-244位氨基酸是其保守结构域,与番茄中的PR-5亲缘关系最为接近。构建GmOLPa基因ORF的原核表达载体pET-28-GP,并对其在大肠杆菌BL21中的表达条件进行优化。SDS-PAGE证实重组质粒能够在大肠杆菌BL21中表达,目的蛋白分子量约为30 kD,最佳诱导时间为4 h,最佳IPTG诱导浓度为1.0 mmol/L。     

Genetic relationships among twelve Chinese indigenous goat populations based on microsatellite analysis

Twelve Chinese indigenous goat populations were genotyped for twenty-six microsatellite markers recommended by the EU Sheep and Goat Biodiversity Project. A total of 452 goats were tested. Seventeen of the 26 microsatellite markers used in this analysis had four or more alleles. The mean expected heterozygosity and the mean observed heterozygosity for the population varied from 0.611 to 0.784 and 0.602 to 0.783 respectively. The mean F_ST (0.105) demonstrated that about 89.5% of the total genetic variation was due to the genetic differentiation within each population. A phylogenetic tree based on the Nei (1978) standard genetic distance displayed a remarkable degree of consistency with their different geographical origins and their presumed migration throughout China. The correspondence analysis did not only distinguish population groups, but also confirmed the above results, classifying the important populations contributing to diversity. Additionally, some specific alleles were shown to be important in the construction of the population structure. The study analyzed the recent origins of these populations and contributed to the knowledge and genetic characterization of Chinese indigenous goat populations. In addition, the seventeen microsatellites recommended by the EU Sheep and Goat Biodiversity Project proved to be useful for the biodiversity studies in goat breeds.     

红藻氨酸致敏癫痫大鼠海马内AP-1 DNA结合活性和成分的改变

一次皮下注射惊厥剂量（７．５ｍｇ／ｋｇ）的红藻氨酸（ｋａｉｎｉｃ ａｃｉｄ,ＫＡ）诱发Ｆｉｓｈｅｒ３４４大鼠出现急性癫痫发作,７ｄ后即可形成癫痫敏感大鼠,继用Ｇｅｌ ｓｈｉｆｔ、Ｓｕｐｅｒ－ｓｈｉｆｔ和Ｗｅｓｔｅｍ ｂｌｏｔ方法测定大鼠海马内ＡＰ－１ ＤＮＡ结合活性及其组成成分。Ｇｅｌ ｓｈｉｆｔ结合显示,癫痫敏感大鼠海马内ＡＰ－１ ＤＮＡ结合活性的基础水平较对照组为高;Ｓｕｐｅｒ－ｓｈｉｆｔ实研     

Renal response to head-out water immersion in Korean women divers

Head-out water immersion (HOI) induces a profound diuresis and natriuresis, which may endanger the body fluid balance of breath-hold divers during prolonged diving work. To investigate if adaptation is acquired by professional breath-hold divers, we have evaluated renal responses to 3-h HOI in 5 Korean women divers (Amas) and 11 nondiving housewives (controls). In both control and diver groups, the average urine flow during 3-h immersion was four times greater and Na⁺ excretion was 70%–80% greater than the pre-immersion value [urine flow: 3.7 (SD 1.0) ml·min^–1 vs 0.9 (SD 0.4), P<0.001, in controls; 4.3 (SD 0.9) vs 1.1 (SD 0.4), P<0.001, in divers; Na⁺ excretion: 270 (SD 176) mol· min^–1 vs 161 (SD 84), P<0.025, in controls; 303 (SD 31) vs 164 (SD 62), P<0.005, in divers]. In all cases, the values for a given period were not significantly different between the two groups. The plasma concentrations of Na⁺ and osmolality, and renal clearance of creatinine did not change significantly. However, the osmolal clearance increased [from 2.0 (SD 0.8) ml·min^–1 to 2.8 (SD 0.7), P<0.05, in the controls; from 2.2 (SD 0.4) to 2.6 (SD 0.4), P<0.05, in the divers] and free water clearance changed from negative to positive values [from -1.1 (SD 0.5) ml·min^–1 to 1.2 (SD 0.3), P<0.005, in the controls; from -1.2 (SD 0.4) to 1.6 (SD 1.1), P<0.01, in the divers] during immersion, again the pattern of change being similar in the two groups. It was, therefore, concluded from our study that the renal response to HOI was unchanged in the Korean women professional breath-hold divers compared to the nondiving women.     

Characterization of S-Triazine Herbicide Metabolism by a Nocardioides sp. Isolated from Agricultural Soils

Atrazine, a herbicide widely used in corn production, is a frequently detected groundwater contaminant. Nine gram-positive bacterial strains able to use this herbicide as a sole source of nitrogen were isolated from four farms in central Canada. The strains were divided into two groups based on repetitive extragenic palindromic (rep)-PCR genomic fingerprinting with ERIC and BOXA1R primers. Based on 16S ribosomal DNA sequence analysis, both groups were identified as Nocardioides sp. strains. None of the isolates mineralized [ring-U-¹⁴C]atrazine. There was no hybridization to genomic DNA from these strains using atzABC cloned from Pseudomonas sp. strain ADP or trzA cloned from Rhodococcus corallinus. S-Triazine degradation was studied in detail in Nocardioides sp. strain C190. Oxygen was not required for atrazine degradation by whole cells or cell extracts. Based on high-pressure liquid chromatography and mass spectrometric analyses of products formed from atrazine in incubations of whole cells with H₂¹⁸O, sequential hydrolytic reactions converted atrazine to hydroxyatrazine and then to the end product N-ethylammelide. Isopropylamine, the putative product of the second hydrolytic reaction, supported growth as the sole carbon and nitrogen source. The triazine hydrolase from strain C190 was isolated and purified and found to have a K_m for atrazine of 25 μM and a V_max of 31 μmol/min/mg of protein. The subunit molecular mass of the protein was 52 kDa. Atrazine hydrolysis was not inhibited by 500 μM EDTA but was inhibited by 100 μM Mg, Cu, Co, or Zn. Whole cells and purified triazine hydrolase converted a range of chlorine or methylthio-substituted herbicides to the corresponding hydroxy derivatives. In summary, an atrazine-metabolizing Nocardioides sp. widely distributed in agricultural soils degrades a range of s-triazine herbicides by means of a novel s-triazine hydrolase.     

细胞生成素3'端增强子在肺动脉平滑肌细胞低氧性增殖调控中的作用

用噻唑蓝比色法(MTT法)、H3-胸腺嘧啶核苷(H3-TdR)掺入法和流式细胞术, 观察红细胞生成素(EPO)3'端增强子片段对培养的猪肺动脉平滑肌细胞(PASMCs)的内皮依赖性和非内皮依赖性低氧性增殖的影响.结果为: (1)低氧24 h后PASMCs明显增殖, 转入野生型EPO3'端增强子片段可被抑制, 而转入突变型片段无此作用;(2)肺动脉内皮细胞(PAECs)低氧24 h, 其条件培养液有明显的促PASMCs增殖作用, 将野生型EPO3'端增强子片段先转入PAECs, 此作用明显减弱, 而转入突变型片段则无明显影响.提示: (1)PAECs低氧条件培养液可促进PASMCs增殖, PASMCs也可直接感受低氧而增殖, PAECs和PASMCs的低氧反应均被外源性EPO3'增强子片段抑制;(2)由于EPO3'增强子上有低氧诱导因子-1(HIF-1)结合位点, PAEC和PASMC低氧反应可能是通过HIF-1低氧信号转导的共同通路.     

Human p53 is phosphorylated on serines 6 and 9 in response to DNA damage-inducing agents

To characterize the sites in human p53 that become phosphorylated in response to DNA damage, we have developed polyclonal antibodies that recognize p53 only when it is phosphorylated at specific sites. Several attempts to generate an antibody to p53 phosphorylated at Ser(6) using a phosphoserine-containing peptide as an immunogen were unsuccessful; however, phosphorylation-specific antibodies were produced by using the phosphoserine mimetic, l-2-amino-4-phosphono-4, 4-difluorobutanoic acid (F(2)Pab), in place of phosphoserine. Fmoc-F(2)Pab was prepared by an improved synthesis and chemically incorporated using solid phase peptide synthesis. Affinity-purified antibodies elicited by immunizing rabbits with an F(2)Pab peptide coupled to keyhole limpet hemocyanin recognized a p53(1-39) peptide phosphorylated only at Ser(6) but not the unphosphorylated peptide or the same peptide phosphorylated at Ser(9), Ser(15), Ser(20), Ser(33), or Ser(37). Untreated A549 cells exhibited a background of constitutive phosphorylation at Ser(6) that increased approximately 10-fold upon exposure to either ionizing radiation or UV light. Similar results were obtained for Ser(9) using antibodies raised against a conventional phosphopeptide. Ser(9) was phosphorylated by casein kinase 1 in vitro in a phosphoserine 6-dependent manner. Our data identify two additional DNA damage-induced phosphorylations in human p53 and show that F(2)Pab-derivatized peptides can be used to develop phosphorylation site-specific polyclonal antibodies.     

The binding of substrate analogs to phosphotriesterase

Phosphotriesterase (PTE) from Pseudomonas diminuta catalyzes the detoxification of organophosphates such as the widely utilized insecticide paraoxon and the chemical warfare agent sarin. The three-dimensional structure of the enzyme is known from high resolution x-ray crystallographic analyses. Each subunit of the homodimer folds into a so-called TIM barrel, with eight strands of parallel beta-sheet. The two zinc ions required for activity are positioned at the C-terminal portion of the beta-barrel. Here, we describe the three-dimensional structure of PTE complexed with the inhibitor diisopropyl methyl phosphonate, which serves as a mimic for sarin. Additionally, the structure of the enzyme complexed with triethyl phosphate is also presented. In the case of the PTE-diisopropyl methyl phosphonate complex, the phosphoryl oxygen of the inhibitor coordinates to the more solvent-exposed zinc ion (2.5 A), thereby lending support to the presumed catalytic mechanism involving metal coordination of the substrate. In the PTE-triethyl phosphate complex, the phosphoryl oxygen of the inhibitor is positioned at 3.4 A from the more solvent-exposed zinc ion. The two structures described in this report provide additional molecular understanding for the ability of this remarkable enzyme to hydrolyze such a wide range of organophosphorus substrates.     

Dual anticancer activity of 8-Cl-cAMP: inhibition of cell proliferation and induction of apoptotic cell death

8-Cl-cAMP induces apoptotic cell death in human cancer cells. To look at this more closely, we examined the changes in the levels of Bcl-2 family proteins during 8-Cl-cAMP-induced apoptosis of SH-SY5Y human neuroblastoma cells. Following the treatment with 8-Cl-cAMP, Bcl-2 was transiently down-regulated and Bad was increased continuously up to day 5. In addition, overexpression of Bcl-2 efficiently blocked the 8-Cl-cAMP-induced apoptosis, suggesting Bcl-2 family proteins may be involved in the 8-Cl-cAMP-induced apoptosis. The contribution of the apoptotic cell death and the inhibition of cell proliferation in the 8-Cl-cAMP-induced growth inhibition was closely monitored in the Bcl-2-overexpressing cells. Though the apoptosis was reduced significantly, no significant difference was observed in the inhibition of cell proliferation up to day 2 of 8-Cl-cAMP treatment. These results suggest that 8-Cl-cAMP exerts anticancer activity by two distinct mechanisms, i.e. , through the inhibition of cell proliferation as well as the induction of apoptosis. Supporting this notion was the observations that (1) suppression of apoptosis by zVAD did not abrogate 8-Cl-cAMP-induced inhibition of cell proliferation, and (2) 8-Cl-cAMP did not show additive inhibition of cell proliferation in RIIbeta-overexpressing cells.