HPLC-ELSD法测定舒血宁注射液配伍溶液中内酯含量

目的:建立高效液相色谱-蒸发光散射检测法(HPLC-ELSD)同时测定舒血宁注射液中银杏内酯A、B、C的含量。结果:舒血宁注射液中银杏内酯A进样量在2～32μL范围内线性关系良好,r=0.9998;银杏内酯B进样量在2～32μL范围内线性关系良好,r=1;银杏内酯C进样量在2～32μL范围内线性关系良好,r=0.9996。方法重复性良好。银杏内酯A、B、C平均回收率分别为97.08%、102.06%、101.6%;RSD分别为2.53%、2.91%、2.25%。结论:该法专属性强,灵敏、准确,可作为舒血宁注射液配伍溶液中内酯含量的检测方法。     

银杏达莫注射液对血管性痴呆大鼠的学习记忆及海马区神经元干预作用

目的:观察银杏达莫注射液对血管性痴呆大鼠的学习记忆及海马区神经元干预作用。方法:大鼠分为三组:假手术组、安慰剂组、银杏达莫组。采用改良双侧颈总动脉阻断方法制备血管性痴呆大鼠模型。利用Morris水迷宫评价大鼠的学习记忆能力,显微镜观察海马区CA1区神经元的组织形态结构。结果 :与假手术相比,安慰剂组大鼠平均逃避潜伏期明显延长(P<0.05),而银杏达莫组大鼠平均潜伏期较安慰剂组明显减少(P<0.05)。形态学观察显示安慰剂组大鼠海马CA1区神经细胞排列紊乱、数量较少,甚至神经细胞坏死、凋亡。银杏达莫组大鼠海马CA1区神经元数目、形态较安慰剂组均有明显改善。结论 :银杏达莫注射液对VD大鼠海马CA1区神经细胞具有保护作用,从而改善血管性痴呆大鼠的学习记忆能力。     

液相色谱-串联质谱法测定人血浆中银杏内酯B的含量

建立测定人血浆中银杏内酯B的含量的LC-MS/MS方法。取正常人血浆50μL,加入2.5mol/L盐酸溶液10μL酸化,再加入内标2μg/mL银杏内酯C20μL,以1mL乙醚萃取后,取提取液500μL挥干溶剂,然后用30%的乙腈500μL溶解,取10μL进行LC-MS/MS测定。LC条件:色谱柱为ZorbaxC18柱(4.5*150mm,5μm);流动相:乙腈-水梯度洗脱;流速:0.6mL/min,柱温:35℃。质谱条件:ESI电离源;负离子模式;用于定量分析的离子对分别为m/z423.1→m/z367.2(银杏内酯B)和m/z439.1→383.2(内标:银杏内酯C)。该方法血浆中银杏内酯B的最低定量限为5ng,线性范围为5ng/mL~0.5μg/mL。本方法操作简便,灵敏度高,结果准确可靠,可用于该药物的含量测定,同时也可为临床药代动力学研究提供参考。     

银杏内酯B注射液在生理盐水中的稳定性的研究

目的:采用高效液相法对银杏内酯B注射液在生理盐水中的稳定性进行研究,又采用HPLC-MS法对银杏内酯B注射液在生理盐水中经加热后所生成的降解产物进行研究。方法:高效液相条件为:色谱柱:Venusil XBP-C18(5μm,4.6×250mm),柱温:25℃,流动相∶甲醇∶水(40∶60),流速:1.0mL/min,检测波长:220nm。结果:在上述色谱条件下,本品在0.1~2mg/mL范围呈线性关系,回归方程为:Y=313.95X+0.8054,r=0.9999;精密度良好,RSD:0.86%;主峰与其它杂质峰分离良好。结论:银杏内酯B注射液在生理盐水中至少可放置3个月。     

应用流式细胞术检测大鼠心肌细胞缺氧/复氧后内质网应急引起细胞凋亡

利用流式细胞仪观察西洋参茎叶总皂甘通诱导大鼠心肌细胞后内质网应急引起细胞凋亡。选用原代培养心肌细胞,将心肌细胞分为正常细胞对照组,缺氧/复氧组和缺氧/复氧后加入西洋参茎叶总皂苷组。按照40mg/L,80mg/L,160mg/L浓度培养24h后,应用流式细胞仪采用Annexin-V/PI诱导凋亡。结果显示应用流式细胞仪检测最终浓度为160mg/L西洋参茎叶总皂苷凋亡率明显低于缺氧/复氧组。流式细胞仪检测由内质网应急诱导细胞凋亡对研究细胞凋亡的机理提供一种快速、准确检测方法。     

环十二酮肟及其羧酸脂的EIMS研究

对环十二酮肟（A）及其行生物（B1～B9）进行了EIMS裂解规律的研究，十二酮肟环开裂后主要形成CnH2n-1C＝NOH及CnH2nCNOH系列的离子，羟基转移后脱C7H14CN形成基峰．羟基氢原子被OC（O）R取代后，取代基的a位及等基裂解成为主要方式。     

关于GB13319—91《形状和位置公差位置度公差》的商榷

对于GB13319-91《形状和位置公差位置度公差》，笔者发现有几个不确切处，并拟建议几处图形变更，现提出与标准制订部门进行商榷。（1）标准的第20页图35：建议位置度公差带图改成图35b下方的公差带图，见图1。说明：在一般机械加工中，4－10H8均布孔的另一定心尺寸为450，该尺寸精度一般按GB／T18O4－92《一般公差线性尺寸的未注公差》的m级，即为450士0．3，虽然位置度为单方向标注，但四个孔的轴线应在图2所示的变动范围内。（2）标准的第20页图36：建议位置度公差带图改成图36b下方的公差带图，见图3。几）位置度公差带续图3说明：根…     

共沉淀法制备Ca0．6Mg0．4Zr4（P04）6纳米粉体

采用共沉淀法，以Ca（NO3）2·4H2O、Mg（NO3）2·6H2O、ZrO（NO3）2·8H2O和NH4H2PO4为原料合成Ca0．6Mg0．4Zr4（P04）6（C0．6M0．4ZP）纳米粉体，通过TGDSC、XRD、TEM和纳米粒度／Zeta电位分析手段研究了反应过程pH值、反应物浓度配比和沉淀反应方式等对合成粉体的相组成、平均颗粒尺寸及其分布等的影响。结果表明：将Ca^2＋、Mg^2＋、Zr4＋离子混合溶液加入到不同pH值的NH4H2PO4溶液中，并控制pH=3～11，沉淀物经900℃煅烧3h后，可合成单相的C0．6M0．4ZP纳米粉体；当Zr4十浓度为0．5mol／L、沉淀剂NH4H2PO4浓度为1．0mol／L、pH=9时，合成粉体平均颗粒尺寸和粒径分布范围最小，分别为40nm和20-70nm。     

畜禽肠道定位驱虫剂对猪肠道寄生虫驱杀效果及增重试验

通过粪检选择自然感染胃肠道寄生虫病60头杜长大三元杂交猪。分为A组（项目产品组）、B组（药物对照组）和C组（空白对照）三个组。结果表明：三个组在驱虫前寄生虫的感染率均为100％，在投药后的第6天粪检，A组和B组的虫卵减少率分别为98．8％和94．7％，A组和B组相比，均差异显著（P〈0．05）；第31天粪检A组和B组的虫卵减少率分别为100％和99．3％；在第55天时粪检时，A组和B组虫卵减少率均为100％，与C组相比差异极显著（P〈O．01）。A组和B组相比，差异显著（P〈0．05）；A、B两组与C组相比，差异极显著（P〈0．01）。     

关于“光学仪器现行国家标准和行业标准”清理整顿结果的通报

根据国家技术监督局关于清理整顿现行标准的要求，机械部对1992年底以前批准发布的机械工业现行国家标准、行业标准进行了清理整顿，现将光学仪器国家标准和行业标准清理整顿结果通报如下：1光学仪器强制性标准1．1国家标准：＊B7667──87电子显微镜x射线泄漏剂量规定1．2行业标准：JB5517──引光学仪器电气防护基本安全要求2光学仪器推荐性标准2．1国家标准：GB779──89光学计GB903—87无色光学玻璃GBI185—89光学零件表面疵病GB］240──76光学样板GB2831──阳光学零件的面形偏差检验方法（光圈识别）onzsas──sl生物显微镜GB…     

Effects of docosahexaenoic acid or arachidonic acid supplementation on gene expression and contractile force of rat cardiomyocytes in primary culture

While fatty acids play essential roles in the physiology of the myocardium, conventional culture media contain little lipid. We previously revealed that rat neonatal myocardium mainly contains docosahexaenoic (DHA), linoleic (LA), and arachidonic (AA) acids as polyunsaturated fatty acids (PUFAs), and these contents in cultured cardiomyocytes derived from fetal rats were markedly lower than those in the neonatal myocardium. In this study, we first assessed the effects of supplementation of DHA, LA, or AA on the fatty acid contents and the percentage change of contractile area in primarily cultured rat cardiomyocytes. Based on this assessment, we then evaluated the effects of DHA or AA supplementation on mRNA expression and further directly measured the contractile force of cardiomyocytes with the supplementations. This study revealed that percentage change of contractile area was maximized under 20 μM DHA or 50 μM AA supplementation while LA supplementation did not affect this contraction index, and that a widespread upregulation tendency of the mRNA expression related to differentiation, maturity, fatty acid metabolism, and cell adhesion was seen in the cultured cardiomyocytes with supplementation of DHA or AA. In particular, upregulation of the gene expression of cellular adhesion molecules connexin43 and N-cadherin were remarkable, whereas the effects on differentiation and maturation were less pronounced. Correspondingly, the increase of the percentage change of the contractile area of cardiomyocyte clusters in culture dishes with the supplementations was significant, whereas the enhancement of the contractile force was modest. These results suggest that supplementation of DHA or AA to the fetal cardiomyocyte culture may play effective roles in preventing the de-differentiation of the cardiomyocytes in culture and that the enhancement of the contractile performance may be mainly attributed to the improvement of intercellular connection.     

Hypoxia induced apoptosis of rat gastric mucosal cells by activating autophagy through HIF-1α/TERT/mTORC1 pathway

The pathogenesis of high altitude-related gastric mucosal injury remains poorly understood, this study aimed to investigate the role of autophagy in hypoxia-induced apoptosis of rat gastric mucosal cells. Rats were randomized into four groups which were maintained at an altitude of 400 m (P) or received no treatment (H), autophagy inducer rapamycin (H+AI) or autophagy inhibitor 3-MA (H+AB) at an altitude of 4,300 m for 1, 7, 14 and 21 days, respectively, and the morphology, ultrastructure, autophagy, and apoptosis of gastric mucosal tissues were examined. Gastric mucosal epithelial cells CC-R039 were cultured under conditions of normoxia, 2% O2 (hypoxia), or 2% O2+anti-mTORC1 for 0, 24, 48, and 72 h, respectively, and the autophagy and apoptosis were analyzed. CC-R039 cells were transfected with siHIF-1α, siTERT, or siRNA and the autophagy was examined. The results showed that the exposure to hypoxia increased the autophagy and apoptosis of gastric mucosal cells in rats, and apoptosis was aggravated by rapamycin treatment but alleviated by 3-MA treatment. Increased duration of hypoxia from 0 to 72 h could increase the autophagy and apoptosis but decrease the proliferation of gastric mucosal cells. Inhibition of mTORC1 with rapamycin led to further increase in apoptosis and even substantial cell death, and inhibition of HIF- 1α and TERT increased mTORC1 expression and reduced autophagy. Moreover, the inhibition of HIF-1α reduced TERT expression. In conclusion, hypoxia could induce apoptosis of rat gastric mucosal cells by activating autophagy through HIF-1α/TERT/mTORC1 pathway     

Cardioprotective potential of curcumin against norepinephrine‐induced cell death: a microscopic study

Cardiomyopathy and associated heart failure continues to be one of the most severe complications that threaten a large population. Curcumin, one of the three curcuminoids of the spice turmeric, is very well known for a multitude of health benefits and functions. Norepinephrine (NE), a catecholamine and also a stress hormone may cause the cardiomyocytes to develop increased sensitivity to death with its increasing concentrations. In this study, we investigated the cardioprotective effect of curcumin in NE‐induced cardiac apoptosis using several fluorescent and nonfluorescent microscopic techniques like DAPI, PI, Giemsa, PicroSirius and TUNEL. The aim of the study was to assess the effect of curcumin in preventing the occurrence of features underlying apoptosis such as nuclear disruption, chromatin condensation, DNA fragmentation and alterations in mitochondrial membrane permeability. Our results show that curcumin protects the cardiomyocytes against apoptosis significantly and also helps them to revert to their normal physiological state. Hence, we propose that curcumin has the potential to act as a therapeutic agent for the attenuation of NE‐induced cardiac cell death and modulation of apoptosis in H9c2 cardiomyocytes.     

心肌细胞传感器及其在生物医学中的应用

传统心肌细胞电生理的研究,因其对细胞的穿刺损害作用,难以实现长时程测量,同时也难以实现对细胞群体的多位点同时测量.该研究采用微机械加工技术开展了基于细胞传感器(cell-based biosensor)的微电极阵列(microelectrode array, MEA)和光寻址电位传感器(light addressable potentiometric sensor, LAPS)等生物传感器技术,在传感器芯片表面培养了心肌细胞,并对其电生理特性进行了传感测量.该技术可对多个细胞同时进行长期、无损检测,在药物筛选、环境检测等生物医学领域具有良好的应用前景.     

High passage numbers induce resistance to apoptosis in C2C12 muscle cells

Cell lines with high passage numbers exhibit alterations in cell morphology and functions. In the present work, C2C12 skeletal muscle cells with either low (<20) or high (>60) passage numbers (identified as lC2C12 or h-C2C12, respectively) were used to investigate the apoptotic response to H2 O2 as a function of culture age h-C2C12. We found that older cultures (h-C2C12 group) were depleted of mitochondrial DNA (mtDNA). When we analyzed the behavior of Bad, Bax, caspase-3 and mitochondrial transmembrane potential, we observed that cells in the h-C2C12 group were resistant to H2 O2 induction of apoptosis. We propose serially cultured C2C12 cells as a refractory model to H2 O2 -induced apoptosis. In addition, the data obtained in this work suggest that mtDNA is required for apoptotic cell death in skeletal muscle C2C12 cells.     

Natriuretic peptides in ectopic myocardial tissues originating from mouse embryonic stem cells

In a previous report we described the survival and contractile function of mouse embryonic stem cell-derived cardiomyocytes in the host retroperitoneum. To further understand the nature of embryonic stem cell-derived cardiomyocytes, the study assessed the synthesis of natriuretic peptides in ectopic myocardial tissues of embryonic stem cell origin. Cardiomyocytes formed in embryoid body outgrowths were transplanted into the retroperitoneum of adult nude mice, and the myocardial tissues that developed were characterized by RT-PCR and immunohistochemistry concerning atrial and brain natriuretic peptides (ANP, BNP). In the outgrowths of embryoid bodies in vitro, gene expression of ANP and BNP was detected by RT-PCR and granules positive for the peptides were identified in a few cardiomyocytes by light and electron microscopic immunocytochemistry. Seven days after transplantation the transplants exhibited multidifferentiated teratoma tissues. Developing chamber myocardial tissues positive for cardiac troponin I, cadherin, and connexin 43 were evident in the transplants, which contained ANP-positive cardiomyocytes. Transplants with beating bundles were observed 30 days after transplantation, in which gene expression of both natriuretic peptides was detected. Myocardial tissues with abundant ANP-immunoreactivity, as well as with BNP-immunoreactivity to a lesser extent, were evident in the transplants. Also, myocardial tissues without immunoreactivity for natriuretic peptides were observed. Immunoelectron microscopy showed discernible secretory granules containing ANP and/or BNP in the cardiomyocytes. These results showed that part of the cardiomyocytes in embryonic stem cell-derived ectopic myocardial tissues are capable of producing natriuretic peptides, which suggests that they may be used as an endocrine source for cardiac hormones.     

Ginsenoside Rg1 protects against ischemia-induced neuron damage by regulating the rno-miRNA-27a-3p/PPARγ axis

Background: A preliminary miRNA screening showed that expression levels of rno-miRNA-27a-3p were significantly increased in the serum and brain tissues of rats undergoing cerebral ischemia. In recent years, there is evidence of the protective capacity of the saponins extracted from panax ginseng and its primary active ingredient ginsenosideRg1oncerebral ischemic injury. Methods: Fetal rat neurons (FRNs) were cultured in glucose-and-serum-free medium and exposed to hypoxia to establish a cerebral ischemia model in vitro (oxygen and glucose deprivation model, OGD). Antioxidant indexes (CAT, SOD), inflammatory markers (MPO, TNF-α and IL-6), and the expression of apoptosis and proliferation associated proteins (NF kB-p65, Caspase 3-cleaved, BCL-2) were examined. Results: Pre-treatment of Rg1 (30–100 μg/mL) could effectively inhibit the decline of antioxidant indexes (CAT, SOD) and increase in inflammatory markers (MPO, TNF-α and IL-6), and effectively inhibited the apoptosis in FRNs induced by OGD in a gradient-dependent manner. The mechanism analysis showed that the role of Rg1 in protecting against ischemia-induced neuron damage depends on its indirect up-regulation of PPAR protein via suppression of rno-miRNA-27a-3p. Moreover, these effects of Rg1 could be reversed by exogenous rno-miRNA-27a-3p and PPAR gene silencing in FRNs exposed to OGD. Conclusion: To summarize, our study demonstrates that Rg1 could effectively attenuate neuronal damage caused by cerebral ischemia via the rno-miRNA-27a-3p/PPARγ pathway. Further, clarification of the novel mechanism will certainly improve our previous understanding of the role of Rg1 and enhancing its level in treatments for alleviating ischemic brain injury.     

Differential expression of CD95, Bcl-2, and Bax in rat gastric chief and parietal cells

Apoptotic cell death is common in the inflamed gastric mucosa, but its role in the regulation of cell homeostasis in normal gastric mucosa is unknown. We investigated the expression of CD95, Bcl-2, and Bax and their roles in the regulation of apoptosis in normal rat gastric mucosa and in cultures of highly enriched rat chief and parietal cells by immunostaining, Western blotting, and FACS. In intact tissue CD95, Bcl-2, and Bax were localized predominantly in the glandular base region in chief cells. In freshly isolated cells, expression of CD95, Bcl-2, and Bax was much more pronounced in chief cells than in parietal cells. A lower intracellular Bcl-2/Bax ratio suggesting a higher susceptibility to apoptosis was noticed in chief rather than in parietal cells. In extended cultures of parietal and chief cells, Bax expression was upregulated and Bcl-2 expression was downregulated. These regulatory changes, presumably caused by in vitro effects, were not associated with an increase in spontaneous apoptosis. Treatment of chief and parietal cells with Fas-ligand induced apoptosis of all CD95 expressing cells. Expression of CD95, Bcl-2, and Bax predominantly in chief cells suggests that in this cell type regulation of apoptosis may differ from that in parietal cells. Binding of FasL with functionally active CD95 receptors on chief and parietal cells may be relevant for induction of apoptosis in inflamed gastric mucosa.     

Application of regularized Richardson-Lucy algorithm for deconvolution of confocal microscopy images

Although confocal microscopes have considerably smaller contribution of out-of-focus light than widefield microscopes, the confocal images can still be enhanced mathematically if the optical and data acquisition effects are accounted for. For that, several deconvolution algorithms have been proposed. As a practical solution, maximum-likelihood algorithms with regularization have been used. However, the choice of regularization parameters is often unknown although it has considerable effect on the result of deconvolution process. The aims of this work were: to find good estimates of deconvolution parameters; and to develop an open source software package that would allow testing different deconvolution algorithms and that would be easy to use in practice. Here, Richardson-Lucy algorithm has been implemented together with the total variation regularization in an open source software package IOCBio Microscope. The influence of total variation regularization on deconvolution process is determined by one parameter. We derived a formula to estimate this regularization parameter automatically from the images as the algorithm progresses. To assess the effectiveness of this algorithm, synthetic images were composed on the basis of confocal images of rat cardiomyocytes. From the analysis of deconvolved results, we have determined under which conditions our estimation of total variation regularization parameter gives good results. The estimated total variation regularization parameter can be monitored during deconvolution process and used as a stopping criterion. An inverse relation between the optimal regularization parameter and the peak signal-to-noise ratio of an image is shown. Finally, we demonstrate the use of the developed software by deconvolving images of rat cardiomyocytes with stained mitochondria and sarcolemma obtained by confocal and widefield microscopes.