P268S突变型NOD2/CARD15真核表达载体的构建及其体外表达

背景：NOD2/CARD15基因序列单核苷酸多态性（SNP）与欧美人群的克罗恩病（CD）明显相关．其中R702W、G908R和3020insC3个SNP位点与CD的相关性尤为显著。而13本、韩国以及我国香港和浙江地区的研究均未发现上述3个SNP的改变，但最近研究发现了可能与中国人CD相关的P268S突变。目的：构建P268S突变型NOD2／CARD15真核表达载体和体外转染体系，为研究突变型NOD2／CARD15的功能提供实验基础。方法：应用定点诱变技术构建P268S突变型NOD2／CARD15真核表达载体，以阳离子脂质体介导体外转染技术瞬时转染人胚肾细胞HEK293T．以蛋白质印迹法和逆转录聚合酶链反应（RT—PCR）检测HEK293T细胞NOD2／CARD15的表达。结果：经克隆、酶切、测序证实获得P268S突变型NOD2／CARD15基因，突变载体转入HEK293T细胞后，NOD2／CARD15有效表达。结论：成功构建了P268S突变型NOD2／CARD15真核表达载体，阳离子脂质体是人胚肾细胞有效的体外转染体系。     

Acetylene hydrochlorination over boron-doped Pd/HY zeolite catalysts

A novel boron-doped Pd/HY zeolite catalyst for acetylene hydrochlorination was prepared and exhibited an outstanding catalytic performance (the acetylene conversion was maintained at >95% for about 30 h). The boron species can stabilize catalytically active Pd²⁺ species and weaken carbon deposition and Pd²⁺ reduction during the reaction, thus improving the catalytic stability.

B doping partly weakens carbon deposition and Pd²⁺ reduction, thus enhancing catalytic stabilities of Pd/HY catalysts for acetylene hydrochlorination.

The coal-based acetylene hydrochlorination process is a major route for the production of vinyl chloride monomer (VCM) in China, which is essential for polyvinyl chloride (PVC) production. However, this process is restricted by the application of toxic and scarce mercury chloride, which is used as the active ingredient of catalysts in the industrial process.^1,2 Hence, it is imperative to explore environmentally friendly catalysts for acetylene hydrochlorination.Since Hutchings et al. suggested that the activities of metal catalysts were associated with the standard electrode potentials of the related metal ions,³ numerous studies have been carried out to develop non-mercury catalysts for efficient and sustainable hydrochlorination of acetylene in recent years, such as Au,^4–11 Pd,^12–19 Pt,²⁰ Ru,²¹ Bi,²² Cu,^23,24etc. Based on the detailed study by Hutchings'' team,³ metal chloride/activated carbon catalysts (including Pd²⁺, Hg²⁺, Cu²⁺, Cu⁺, Ag⁺) had superior catalytic activity and the order of their activities was Pd²⁺ > Hg²⁺ > Cu²⁺ > Cu⁺ > Ag⁺. Therefore, palladium-based catalysts were reported to be active for acetylene hydrochlorination.^12–19 Wang et al. prepared Pd-based/C catalysts via impregnation method and suggested that the Pd loss and the carbon deposition were main reasons for the catalyst deactivation, and the adding of K and La could improve the stability and regeneration of Pd/C catalysts.¹⁸ Mitchenko et al. reported that Pd(ii) chloro complexes were the active sites of catalysts¹⁹ and the (NH₄)₂PdCl₄ complex could stabilize the Pd species resulted in improving the catalytic performance over carbon-based materials.¹³ In our previous study, zeolites as new supports were prepared Pd/HY catalysts applied for acetylene hydrochlorination and the catalysts stabilities could be enhanced after modification with NH₄F or polyaniline.^14–17 While the carbon deposition and the active component reduction is the major obstacle to improve activity and stability of catalysts. It reported that heteroatom doping of support materials was effective for the improvement of the catalytic performance of metal catalysts²⁵ and few reports were studied on the B-doped zeolite-based catalysts for acetylene hydrochlorination at present. In this work, a novel boron-doped Pd/HY zeolite-based catalyst for acetylene hydrochlorination was prepared and its outstanding catalytic performance was further studied.The boron-doped HY zeolites supports were prepared using boron oxide (B2) as the boron source. The boron oxide aqueous solution was quantitatively (the weight percentage of B2 is 1.0%, 3.0%, 5.0%, 7.0%, 9.0%) added into HY zeolites (purchased by Nankai University, Si/Al = 9, 5.0 g) under stirring for 4 h, then washed with distiller water to pH about 7 and water was evaporated at 80 °C for 10 h, the B-doped HY zeolites supports were obtained. The Pd/B2-HY catalysts were prepared via ultrasonic-assisted impregnation using the B-doped HY or the original HY supports, as described in our previous work.^14–17 Based on the weight percentage of the boron oxide, the obtained B-doped Pd/HY catalysts were named as Pd/1B2-HY, Pd/3B2-HY, Pd/5B2-HY, Pd/7B2-HY and Pd/9B2-HY, respectively. The Pd loading in all catalysts was 0.9 wt%.X-ray diffraction (XRD) data were collected using a M18XHF22-SRA diffractometer with Cu–K irradiation at 50 kV and 100 mA in the scan range of 2 between 10°–80°. Scanning electron microscopy (SEM) and energy dispersive X-ray (EDX) analyses were performed using a LEO1450VP detector to determine the morphology. High-resolution transmission electron microscopy (HRTEM) experiment was performed using a JEM-2100F (JEOL, Japan) working at 200 kV. BET surface areas analysis was performed by JW-BK Brunauer–Emmett–Teller (BET) equipment. Thermogravimetric (TG) tests were detected by a NETZSCH SAT 449F3 multifunctional thermal analyzer in an air atmosphere at a flow rate of 50 mL min⁻¹. The nature of carbon deposition was determined by an Agilent 7890A/5975C gas chromatography-mass spectrometry (GC-MS). Palladium contents were detected using an inductively coupled plasma (ICP-6300) instrument. X-ray photoelectron spectroscopy (XPS) data were recorded by AXIS ULTRA spectrometer (Kratos Analytical Ltd) and binding energies were referred to the C 1s line at 284.8 eV. Fourier transform infrared spectrometer (FTIR) was used EQUINOX-55 (Bruker Company, Germany) in transmittance (%) mode in the range 4000–400 cm⁻¹ and temperature programmed decomposition (TPD) was determined using a TP-5080 adsorption instrument with ammonia over a temperature ramp of 0–900 °C, rate ramp of 10 °C min⁻¹ and flow of 100 mL min⁻¹.The catalytic performances of catalysts were evaluated in a fixed bed with 10 mm-diameter a quartz tube micro reactor. The reactor temperature was calibrated by a CKW-110 temperature controller (Chaoyang automation instrument factory, Beijing, China) and the gas flow during the reaction was regulated by a D08-1F mass flow controller (Sevenstar Huachuang electronics. co. Ltd, Beijing, China). After N₂ flow (20 mL min⁻¹) purged, hydrogen chloride (12.6 mL min⁻¹) and acetylene (10.1 mL min⁻¹) were fed through a mixing vessel containing catalyst (3.0 g), giving a temperature at 160 °C, feed volume ratio V_HCl : V_C2H2 = 1.25 and the C₂H₂ gas hourly space velocity (GHSV) of 110 h⁻¹. The exit gas mixture from the reactor was passed through an absorption bottle filled with sodium hydroxide solution and then set into a gas chromatography (GC 2010, Shimadzu) to analyse the acetylene conversion and the VCM selectivity immediately.^13–17 Fig. 1 and S1^† display the catalytic performance of undoped and B-doped catalysts. The undoped Pd/HY catalyst exhibited a poor catalytic stability and its C₂H₂ conversion dropped dramatically from 93% to 13% in 3 h. After B doping, the C₂H₂ conversion was significantly enhance from 93% of the undoped Pd/HY catalyst to 96% of the B-doped Pd/B2-HY catalyst under the same conditions. It was clear that the Pd/7B2-HY catalyst displayed the optimal catalytic performance with the C₂H₂ conversion of 96% and the VCM selectivity of 98% after 10 h and its activity kept over 95% within 30 h (Fig. S2^†). It is suggested that the appropriate amount of B doping can improve the activity and stability of Pd-based catalysts for acetylene hydrochlorination.Open in a separate windowFig. 1The catalytic performances of the B-doped Pd/B2-HY catalysts.It is obvious that no other diffraction peaks of Pd species were detected, except the strongest characteristic peak of HY zeolites, which could be attributed to the low concentration or high dispersion of Pd species^26,27 (Fig. 2a). For the fresh Pd/7B2-HY catalyst, numerous Pd-based particles with the size about 2.81 nm were distributed uniformly on the HY support (Fig. 2b and d), which was slight bigger than that of the average pore width (by BET results, 2.49 nm) of the 7B2-HY zeolites, deducing that most Pd species involved in reaction might be on the support surface. In addition, the Pd species on the HY zeolite were visualized with well-defined fringes, which was ascribed to the (111) plane of face-centered cubic Pd with a d spacing of 0.23 nm (Fig. 2c).²⁸ The characterization of HRTEM verifies that XRD pattern is reasonable and suggests that the Pd species are highly dispersed in the fresh Pd/7B2-HY catalyst (Open in a separate windowFig. 2The characterization on the fresh catalysts: (a) XRD patterns of Pd-based catalysts. (b and c) HRTEM images of Pd/7B2-HY catalyst. (d) Pd particle size distribution of Pd/7B2-HY catalyst.Porous structure parameters of samples

大肠癌组织中P16蛋白和血管内皮生长因子的表达及其临床意义

目的：探讨大肠癌组织中P16蛋白和血管内皮生长因子（VEGF）表达及其临床意认。方法：用S－P免疫组织化学方法测定66例大肠癌组织和20例正常大肠组织中P16蛋白和VEGF的表达。结果：大肠癌中P16蛋白阳性率为48．5％（32／66）明显低于对照组的70．0％（14／20）（P＜0．01），VEGF阳性率为72．7％（48／66）则明显高于对照组的15．0％（3／20）（P＜0．01）：P16蛋白和VEGF在大肠癌中表达具有明显负相关性；P16蛋白和VEGF表达与大肠癌组织学类型、肿瘤直径、肿瘤部位无关（P＞0．05），而与淋巴结转移、Duke's分期五年生存率有明显的关系（P＜0．01）。结论：大肠癌中存在P16蛋白下调和VEGF上调，P16蛋白和VEGF表达可作为反映大肠癌生物学行为的指标之一。     

Protective Role of Artemisia Afra Aqueous Extract on Tissue Antioxidant Defense Systems in Streptozotocin-Induced Diabetic Rats

Changes in antioxidant capacity in the body as a result of oxidative stress play an important role in the development of diabetic complications. The aim of this study was to evaluate the effect of aqueous extract of Artemisia afra Jacq. ex Willd. on antioxidant defense systems in the liver and kidney of streptozotocin-induced diabetic rats. Administration of the extract to diabetic rats for 21 days significantly reduced blood glucose levels and increased body weight. The diabetic animals exhibited decreased levels of glutathione reductase (GR), glutathione peroxidase (GPx), superoxide dismutase (SOD) and reduced glutathione (GSH) in the liver and kidney, which were restored to near normal levels following treatment with the herb. The increased levels of lipid peroxidation observed in the tissues of diabetic rats were also reverted back to near normalcy after administering the extract. These findings revealed the protective role of A. afra on tissues by reducing oxidative stress which could be attributed to its flavonoids content. The efficacy of the plant compared favourably well with glibenclamide, a standard hypoglycemic drug.     

Healthcare Workers Knowledge and Attitude Toward Palliative Care in an Emerging Tertiary Centre in South-West Nigeria

Background:

Palliative care is an emerging area of medicine with potential to affect positively many chronically ill patients. This study investigated the knowledge and attitude of healthcare workers in a tertiary level hospital in Nigeria where a palliative care unit is being established.

Material and Methods:

The study was a cross-sectional questionnaire-based study carried out among healthcare workers in Ekiti State University Teaching Hospital, Ado-Ekiti, south-west Nigeria. The questionnaire had sections about definition of palliative care, its philosophy, communication issues, medications, and contexts about its practice. The information obtained from the questionnaire was coded, entered, and analyzed using IBM SPSS version 19.

Results:

A total of 170 questionnaires were returned within the stipulated time frame with response rate of 66.7%. Majority, (135, 86%) respondents felt palliative care was about the active management of the dying while 70.5% of respondents equated palliative care to pain management. Regarding the philosophy of palliative care, 70 (57.9%) thought that it affirms life while 116 (78.4%) felt palliative care recognizes dying as a normal process. One hundred and twenty-two (78.7%) respondents were of the opinion that all dying patients would require palliative care. The patient should be told about the prognosis according to 122 (83%) respondents and not doing so could lead to lack of trust (85%). Regarding the area of opioid use in palliative care, 76% of respondents agreed that morphine improves the quality of life of patients.

Conclusion:

There are plausible gaps in the knowledge of the healthcare workers in the area of palliative care. Interventions are needed to improve their capacity.     

养阴清热降糖胶囊的提取工艺考察

目的：探讨养阴清热降糖胶囊的最佳提取工艺。方法：采用正交设计优化养阴清热降糖胶囊的水提取工艺,以多糖含量为考察指标,比较溶剂用量、提取时间及提取次数对提取效果的影响。结果：最佳提取条件为8倍水量,回流提取3次,每次1h。结论：该工艺所得多糖含量较高,稳定性、重复性较好,可为养阴清热降糖胶囊的制备提供参考资料。     

The BH3-only protein, PUMA, is involved in oxaliplatin-induced apoptosis in colon cancer cells

Oxaliplatin, the first line chemotherapeutic of colon cancer, induces damage to tumors via induction of apoptosis. PUMA (p53 up-regulate modulator of apoptosis) is an important pro-apoptotic member of Bcl-2 family and regulated mainly by p53. Here we investigated the role of PUMA in oxalipaltin-induced apoptosis and the potential mechanism. We showed that oxaliplatin-induced PUMA expression in a time- and dose-dependent manner and suppression of PUMA expression by stable transfecting anti-sense PUMA plasmid decreased oxaliplatin-induced apoptosis in colon cancer cells. By abrogating the function of p53, we further demonstrated that the induction was p53-independent. We also found that oxaliplatin could inactivate ERK and suppression of ERK activity by its specific inhibitor (PD98059), and dominant negative plasmid (DN-MEK1) enhanced the oxaliplatin-induced PUMA expression and apoptosis in a p53-independent manner. Taken together, our data suggest that PUMA plays an important role in oxaliplatin-induced apoptosis and the induction could be both p53-dependent and p53-independent. Moreover, PUMA expression and apoptosis in oxaliplatin-treated colon cancer cells could be regulated partly by ERK inactivation. Identification of the molecular components involved in regulating the cellular sensitivity to oxaliplatin may provide potential targets for development of novel compounds that may be useful in enhancement of oxaliplatin cytotoxicity in p53 deficient colon cancer.     

幽门螺杆菌过氧化氢酶基因的克隆、高效表达及活性评价

目的：构建高效表达幽门螺杆菌（Hp）过氧化氢酶重组蛋白的候选菌株并测定其活性。方法：用PCR方法从Hp染色体DNA上扩增过氧化氢酶基因，将其定向插入表达pET-22b(＋），并在BL21（DE3）大肠杆菌中表达，进一步利用贝尔斯－西策尔斯法测定其活性。结果：DNA序列分析表明，所克隆的过氧化氢酶基因序列与基因库公布的一致。在37℃诱导表达3h后，过氧化氢酶重组蛋白表达量占苗体总蛋白的24．4％，并显示了良好的活性。结论：本研究获得了表达高活性Hp过氧化氢酶的克隆，为深入研究其相关功能奠定了良好基因。     

����Һ��ʧѪ���ݿ˵Ľ�չ

补液是治疗各类休克的主要手段 ,是综合治疗休克的重要组成部分。因休克发生与发展的各环节均不同程度地伴有容量的丧失。　　输入等渗溶液作为抗休克的标准疗法 ,一直沿用至今[1] 。等渗溶液即为按正常人体渗透压而配制的等渗性溶液 (或生理性溶液 ) [2 ] ,而用生理性溶液 (包括血液 )来拮抗已发生明显病理生理改变的状态 ,这是值得探讨的。按正常人配制的生理性溶液 ,理应给予正常人似乎更为合理 ,但正常人又不需要这些液体。因而 ,在对发生轻度病理生理变化时作为补充替代或作为载体 (如溶入维生素及抗生素等 )是可行的。而对已发生明显…     

乳腺癌患者化疗前后血清sVCAM-1和CA15-3表达及其临床意义

目的观察可溶性血管细胞粘附分子-1(sVCAM-1)和癌相关糖蛋白抗原(CA15-3)在乳腺癌患者化疗前后浓度的变化,探讨二者水平与临床分期、转移部位的关系,并初步探讨晚期转移性乳腺癌患者治疗效果与二者水平变化的关系。方法采用酶联免疫吸附试验(ELISA)和化学发光免疫法(CLIA)测定15例健康人、30例早期乳癌患者(Ⅰ、Ⅱ)、40例晚期乳癌患者(Ⅲ、Ⅳ)化疗前后sVCAM-1、CA15-3血清水平。结果治疗前,Ⅰ、Ⅱ期乳腺癌患者血清sVCAM-1和CA15-3浓度与健康者比较差异无显著性(P>0.05);Ⅲ、Ⅳ期乳腺癌患者血清sVCAM-1和CA15-3浓度均显著高于健康者,并随着分期渐晚二者浓度逐渐升高,差异有显著性(P<0.05),其中,乳腺癌患者各组血清CA15-3阳性率均显著高于正常对照组(P<0.05)。检测了12例内脏转移和7例骨转移晚期乳腺癌患者血清sVCAM-1和CA15-3水平,发现内脏转移组二者水平高于骨转移组,但差异无显著性(P>0.05)。治疗后,各乳腺癌患者组血清sVCAM-1和CA15-3水平均较前降低,Ⅰ、Ⅱ组差异无显著性,Ⅲ、Ⅳ组差异显著(P<0.05),转移复发乳腺癌患者下降尤为明显。24例晚期转移患者采用以TA为主的联合方案化疗,治疗后19例患者疗效评价为PR CR,化疗后sVCAM-1和CA15-3水平均下降,差异显著(P<0.001)。结论乳腺癌患者血清sVCAM-1和CA15-3水平与病期早晚及治疗反应相关,与转移部位关系不明显,二者有可能成为评价乳腺癌临床分期和血清学疗效的指标。