人野生型parkin基因真核表达载体的构建及其在PC12细胞中的表达

目的 克隆人野生型parkin基因并构建真核表达载体pCDNA3．1—parkin，将重组质粒转染PC12细胞获得高表达人野生型parkin基因的PC12细胞克隆。方法 从胎脑组织中提取总RNA，用RT—PCR方法获得人野生型parkin基因的全长cDNA，插入pCR2．1—TA克隆载体中进行序列测定，测序正确后将其亚克隆至表达载体pCD—NA3．1，利用脂质体将重组质粒转染PC12细胞，经G418筛选获得抗性细胞克隆，采用RT—PCR和Western Blot方法鉴定人野生型parkin基因在PC12细胞中的过表达。结果 经限制性内切酶酶切图谱分析和DNA序列测定证实目的基因已插入重组质粒，RT—PCR和Western Blot证明经G418筛选得到的转基因PC12细胞克隆中存在人野生型parkin基因的表达。结论 成功构建了人野生型parkin基因的真核表达载体，获得了稳定表达人野生型parkin基因的PC12细胞克隆，为进一步研究parkin的生物学功能以及parkin在帕金森病发病机制中的作用奠定了良好的基础。     

甲基强的松龙治疗实验性变态反应性脑脊髓炎的作用机制

目的:研究细胞因子、T细胞凋亡和淋巴细胞增殖在实验性变态反应性脑脊髓炎(EAE)形成中的作用及甲基强的松龙(MP)治疗EAE的作用机制。方法:采用人脑纯化的髓鞘碱性蛋白(MBP)与完全福氏佐剂免疫Lewis大鼠,建立EAE动物模型。用双抗体夹心ELISA法检测各组大鼠血清中IL-10、TNF-α、IFN-γ的含量:流式细胞仪检测外周血T细胞凋亡;3H-TdR释放法检测外周血淋巴细胞转化率。结果:与对照组比较,EAE组的外周血IFN-γ、TNF-α水平明显增高,IL-10水平明显降低,MP治疗后IFN-γ和TNF-α水平下降,IL-10浓度上调。MP还诱导外周血T细胞凋亡和抑制MBP致敏淋巴细胞增殖并呈剂量依赖性。结论:应用人MBP成功建立EAE大鼠模型,MP可能通过调节Th细胞因子格局、促进Th2细胞因子分泌、抑制MBP致敏淋巴细胞增殖及外周血T细胞凋亡而发挥治疗多发性硬化的作用。     

中国男性原发不育的遗传高风险因子:Y染色体DAZ基因拷贝缺失

目的 探讨Y染色体无精症因子C区(azoospermia factor C，AZFe)无精症缺失基因(deleted-in-azoospermia，DAZ)家族基因拷贝缺失与中国男性原发不育之间的关系。方法 运用多重PCR与PCR-RFLP检测技术，对210例已生育男性、216例原发无精症与189例严重少精症患者Y染色体AzFc区域DAZ基因家族的基因拷贝数进行分析。结果 在所有已生育男性中未检出DAZ基因拷贝的完全或部分缺失，而在原发无精症与严重少精症患者中DAZ基因拷贝完全缺失率分别为8．8％和12．2％，DAZ1／DAZ2共缺失率分别为8．3％和5．3％。结论 在中国男性原发无精症与严重少精症患者中存在较高频率的DAZ基因拷贝缺失现象，提示Y染色体AZFc区域DAZ基因家族基因拷贝的完全与部分缺失是中国男性原发不育的遗传高风险因子。     

11β-羟类固醇脱氢酶与2型糖尿病

11β-羟类固醇脱氢酶(11β-HSDs)催化氢化可的松(皮质酮)和可的松(脱氢皮质酮)的相互转化，对调节局部糖皮质激素的活性起着重要作用，而糖皮质激素可诱导胰岛素抵抗。肝、脂肪等组织内11β-HSD活性的改变与肥胖、2型糖尿病等疾病密切相关。     

粗糙脉孢菌漆酶基因的克隆及在毕赤酵母中的初步表达

采用连续延伸PCR方法克隆到粗糙脉孢菌（Neurospora crassa）漆酶基因，并将其克隆到表达载体pPIC9k，重组质粒经线性化、电激转化Pichia pastoris KM71，部分阳性克隆的PCR结果表明：漆酶基因已整合到巴斯德毕赤酵母染色体上，重组菌经甲醇诱导后3～5d产漆酶量最高，为2-3U／mL。     

49Sibutramine accelerates gastric emptying, inhibits intestinal motility in conscious dogs

Fasting interdigestive myoelectric complex (IMC) and postprandial gastroduodenal myoelectric activities are regulated by motilin and leptin, respectively (L. Zhou 2005). This study is to observe whether electrical acupuncture points could increase fasting gastroduodenal IMC and postprandial activities, as well as increase the release of motilin and leptin. Methods: Bipolar platinum electrodes were implanted on the serosa of antrum and duodenum in 10 conscious Wistar rats to record IMC and postprandial digestive myoelectric activities. We acupunctured Points Zusanli (S‐36), Tianshu (S‐25), and Liangmen (S‐21) during phase I of IMC (Frequency 20~100 Hz, Strengthen 12~16 mA, period 90 min) and compared with non‐specific electric stimulation (sham acupuncture) group. Serum motilin and leptin level was measured by RIA. Results: (1) Acupuncture points could shorten markedly phase I of antroduodenal IMC and increase the period of phase III (P < 0.01). Acupuncture points also increased the number of spike burst of antral and duodenal slow wave by (121.24 ± 20.0)% and (97.34 ± 15.20)% (compared with control group, P < 0.01). In postprandial period, acupuncture points could increase the number of spike burst of antral and duodenal slow wave by (142.52 ± 23.50)% and (102.48 ± 13.25)% (compared with control group, P < 0.01). (2) When acupuncture points increased the number of spike burst of IMC in fasting state, serum motilin concentration also increased, which was (74.56 ± 8.20)% more than control group (P < 0.01). When acupuncture points with meal, serum leptin concentration increased with myoelectric activity, which was (139.84 ± 20.25)% more than control group (P < 0.01). There was no change of motilin and leptin concentration in non‐specific electric stimulation group. (3) Cutting off subphrenic vagus nerves could totally block the effect of acupuncture points on antroduodenal myoelectric activities and release of motilin and leptin. Conclusions: Acupuncture points could effectively increase fasting and postprandial antroduodenal myoelectric activities. This effect is mediated by motilin and leptin, respectively.     

Genome-wide scan identified QTLs underlying femoral neck cross-sectional geometry that are novel studied risk factors of osteoporosis.

A genome-wide screen was conducted using a large white sample to identify QTLs for FNCS geometry. We found significant linkage of FNCS parameters to 20q12 and Xq25, plus significant epistatic interactions and sex-specific QTLs influencing FNCS geometry variation. INTRODUCTION: Bone geometry, a highly heritable trait, is a critical component of bone strength that significantly determines osteoporotic fracture risk. Specifically, femoral neck cross-sectional (FNCS) geometry is significantly associated with hip fracture risk as well as genetic factors. However, genetic research in this respect is still in its infancy. MATERIALS AND METHODS: To identify the underlying genomic regions influencing FNCS variables, we performed a remarkably large-scale whole genome linkage scan involving 3998 individuals from 434 pedigrees for four FNCS geometry parameters, namely buckling ratio (BR), cross-sectional area (CSA), cortical thickness (CT), and section modulus (Z). The major statistical approach adopted is the variance component method implemented in SOLAR. RESULTS: Significant linkage evidence (threshold LOD = 3.72 after correction for tests of multiple phenotypes) was found in the regions of 20q12 and Xq25 for CT (LOD = 4.28 and 3.90, respectively). We also identified eight suggestive linkage signals (threshold LOD = 2.31 after correction for multiple tests) for the respective geometry traits. The above findings were supported by principal component linkage analysis. Of them, 20q12 was of particular interest because it was linked to multiple FNCS geometry traits and significantly interacted with five other genomic loci to influence CSA variation. The effects of 20q12 on FNCS geometry were present in both male and female subgroups. Subgroup analysis also revealed the presence of sex-specific quantitative trait loci (QTLs) for FNCS traits in the regions such as 2p14, 3q26, 7q21 and 15q21. CONCLUSIONS: Our findings laid a foundation for further replication and fine-mapping studies as well as for positional and functional candidate gene studies, aiming at eventually finding the causal genetic variants and hidden mechanisms concerning FNCS geometry variation and the associated hip fractures.     

Clinical report of cervical arthroplasty in management of spondylotic myelopathy in Chinese

Objectives  

To investigate clinical effects and manual operational point of Bryan cervical disc prosthesis in Chinese, to observe the stability and range of movement (ROM) post-operatively.     

Arterial effects of salt restriction in hypertensive patients. A 9-week, randomized, double-blind, crossover study.

OBJECTIVE: To investigate the hemodynamic effects of a moderately low-salt diet in a 9-week, randomized, double-blind, crossover study in 20 hypertensive, ambulatory patients. METHODS: All subjects followed a 9-week, low-salt diet. During this period, they received capsules containing either lactose or salt in 4-week treatment periods, separated by a 1-week washout period. Hemodynamic and biological parameters were evaluated on the day of randomization and at the end of weeks 4 and 9. We defined a low-sodium diet (LSD) as a salt-restriction period with lactose capsules, and a normal-sodium diet (NSD) as a salt-restriction period with capsular salt supplementation. RESULTS: Blood pressure was significantly lower during LSD compared with NSD. This fall in blood pressure was associated with a decrease in peripheral resistance in carotid and forearm circulation. Brachial artery diameter was larger during LSD whereas carotid artery diameter remained unchanged. The changes in brachial artery were: (1) not related to blood pressure changes; (2) positively related to age, and (3) negatively correlated with baseline intracellular sodium content. CONCLUSIONS: These results suggest that moderate low-salt restriction is capable of decreasing blood pressure and peripheral resistance in carotid and forearm circulation. The increase in brachial, but not carotid, artery diameter following salt restriction suggests a difference in salt dependence among different arteries.