Antifungal Activity of Aspidin BB from Dryopteris fragrans against Trichophyton rubrum Involved Inhibition of Ergosterol Biosynthesis

Objectives Aspidin BB, a typical phloroglucinol derivative from Dryopteris fragrans, possesses significant antifungal property. This study aimed to investigate potential mechanism of antifungal activity of Aspidin BB against Trichophyton rubrum which is the most common pathogens responsible for chronic dermatophytosis. Methods The minimum inhibitory concentration(MIC) of Aspidin BB against strains was determined by broth microdilution. The effects of Aspidin BB on ergosterol biosynthesis were investigated by content determination based on UPLC method. Besides, the effects of drugs on squalene epoxidase(SE) in T. rubrum cell membrane were analyzed. Results MIC value of Aspidin BB against T. rubrum was 25.0 μg/m L. Aspidin BB reduced ergosterol content significantly, but no notable effect on squalene epoxidase activity. Conclusion The results suggested that Aspidin BB inhibited ergosterol biosynthesis.However, it was not squalene epoxidase but other components may sever as possible targets in ergosterol biosynthesis pathway.     

建泽泻鲨烯合酶基因克隆及其生物信息学分析

目的 对建泽泻原萜烷型三萜类成分生物合成关键酶鲨烯合酶(squalene synthase,SS)进行基因全长的克隆和生物信息学分析.方法 以建泽泻总RNA为模板,运用同源克隆法和RACE技术克隆建泽泻SS基因的cDNA全长,并通过DNAMAN软件及ExPASy在线分析等方法对其进行生物信息学研究.结果 获得建泽泻SS基因全长cDNA,GenBank注册号为JX866770,序列分析表明,所克隆的cDNA序列全长为1577bp,包含一个长1230bp的开放阅读框架,编码409个氨基酸的蛋白,与其他药用植物具有较高的同源性.预测该蛋白的相对分子质量为4.68×104,等电点为5.97,无信号肽,包含2个富含天冬氨酸(DXXDD)的保守功能域.结论 首次克隆获得建泽泻SS基因的全长cDNA,为泽泻原萜烷型三萜类成分生物合成途径阐明与生物工程应用提供科学依据.     

毛细管气相色谱法测定角鲨烯软胶囊中角鲨烯的含量

目的 建立毛细管气相色谱法测定角鲨烯软胶囊中角鲨烯的含量.方法 采用毛细管气相色谱法;色谱柱为RTX-5弹性石英毛细管柱(30 m×0.32 mm,0.25 μm);载气为氮气;检测器为FID;进样口和检测器温度均为280℃;柱温为250℃.结果 角鲨烯在1～50 mg·L-1与峰面积线性关系良好,r=0.997 6;加样回收率平均为100.8％(n=9);检测限为0.4 ng;定量限为2ng.结论 本法灵敏度高,简单易行,结果准确可靠,可为角鲨烯的质量控制提供依据.     

Simvastatin disrupts cytoskeleton and decreases cardiac fibroblast adhesion, migration and viability

Statins reduce the isoprenoids farnesyl and geranylgeranyl pyrophosphate, essential intermediates, which control a diversity of cellular events such as cytoskeleton integrity, adhesion, migration and viability. Cardiac fibroblasts are the major non-myocyte cell constituent in the normal heart, and play a key role in the maintenance of extracellular matrix. The effects of simvastatin on cardiac fibroblast processes previously mentioned remain unknown. Our aims were to investigate the effects of simvastatin on cytoskeleton structure and focal adhesion complex assembly and their relationships with cell adhesion, migration and viability in cultured cardiac fibroblasts. To this end, cells were treated with simvastatin for 24 h and changes in actin cytoskeleton, levels of vimentin and paxillin as well as their subcellular localization were analyzed by Western blot and immunocytochemistry, respectively. Cell adhesion to plastic or collagen coated dishes, migration in Transwell chambers, and cell viability were analyzed after simvastatin treatment. Our results show that simvastatin disrupts actin cytoskeleton and focal adhesion complex evaluated by phalloidin stain and immunocytochemistry for paxillin and vinculin. All these effects occurred by a cholesterol synthesis-independent mechanism. Simvastatin decreased cell adhesion, migration and viability in a concentration-dependent manner. Finally, simvastatin decreased angiotensin II-induced phospho-paxillin levels and cell adhesion. We concluded that simvastatin disrupts cytoskeleton integrity and focal adhesion complex assembly in cultured cardiac fibroblasts by a cholesterol-independent mechanism and consequently decreases cell migration, adhesion and viability.     

桑黄鲨烯环氧酶基因克隆与序列分析

目的对参与桑黄三萜合成途径的关键酶鲨烯环氧酶(squalene epoxidase,SE)进行基因全长克隆和生物信息学分析。方法以桑黄总RNA为模板,采用RT-PCR和RACE技术克隆桑黄SE基因的全长c DNA和DNA序列,并通过Ex PASy在线分析等方法对其进行生物信息学分析。结果序列分析表明,SE基因全长2 145 bp,包含6个外显子和5个内含子;所克隆的c DNA全长为1 856 bp,包含1个1 452 bp的开放阅读框,编码483个氨基酸的蛋白,命名为Ib SE1,预测该蛋白的相对分子质量为5.3×104,等电点(p I)为8.41,无信号肽。结论首次克隆并获得桑黄鲨烯环氧酶基因全长序列,为进一步阐明桑黄三萜代谢途径和改善中药材品质奠定基础。     

Influence of proton pump inhibitors and VKORC1 mutations on CYP2C9‐mediated dose requirements of vitamin K antagonist therapy: a pilot study

Interindividual variations in dose requirements of oral vitamin K antagonists (VKA) are attributed to several factors, including genetic variant alleles of vitamin K epoxide reductase complex subunit 1 (VKORC1) and cytochrome P450 2C9 (CYP2C9), but also interaction with co‐medications. In this context, proton pump inhibitor (PPI)‐related alterations of VKA maintenance dose requirements have been published. The present investigation aimed to test for an interaction profile of oral VKA‐therapy and PPIs in relation to the CYP2C9 genotype. Median weekly stable VKA dose requirements over 1 year were recorded in 69 patients. Patients were genotyped for CYP2C9*2, CYP2C9*3, VKORC1c.‐1639G>A and VKORC1c.174‐136C>T and assessed for an association with PPI use and total VKA maintenance dose requirements. PPI users with CYP2C9 genetic variations required significantly lower weekly VKA maintenance doses than those with the wild‐type genotype (t‐test: P = 0·02). In contrast, in subjects without PPI use, the CYP2C9 genotype had no significant influence on oral VKA dose requirements. Further, the combined CYP2C9/VKORC1 genotype was a significant predictor for VKA dose requirements [linear regression: estimate: ?1·47, standard error: 0·58 (P = 0·01)]. In conclusion, in carriers of CYP2C9 gene variations, the interference with the VKA metabolism is modified by PPI co‐medication and the VCKORC1 genotype. Preceding knowledge of the genetic profile and the awareness for potentially occurring severe over‐anticoagulation problems under PPI co‐medication could contribute to a safer and personalized VKA pharmacotherapy.     

Reduction of Squalene Epoxidase by Cholesterol Accumulation Accelerates Colorectal Cancer Progression and Metastasis

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Novel agents for managing dyslipidaemia

An elevated low-density lipoprotein (LDL) cholesterol level is a strong predictor of coronary heart disease (CHD) risk. Over the past seven years, equally strong evidence has accumulated that lowering LDL cholesterol with HMG-CoA reductase inhibitors or statins reduces CHD risk and there is now widespread use of these agents for the primary and secondary prevention of CHD. Treatment issues remain regarding the appropriate degree of LDL cholesterol reduction and whether, in people with very high levels, it would be preferable to achieve the LDL cholesterol goal with a powerful statin alone or combined with an agent that lowers LDL cholesterol by a different mechanism. The main focus in the development of novel agents is the patient with low high-density lipoprotein (HDL) cholesterol, usually associated with hypertriglyceridaemia. Already prevalent as a risk factor for CHD, this abnormality has been linked with insulin resistance, which is likely to increase greatly over the next decade, along with increasing obesity and diabetes. Agents that have potent HDL cholesterol raising capacity include cholesteryl ester transfer protein (CETP) inhibitors, retinoid X receptor (RXR) selective agonists, specific peroxisome proliferator-activated receptor (PPAR) agonists and oestrogen-like compounds. Another area of development involves agents that will lower both cholesterol and triglyceride levels, such as partial inhibitors of microsomal triglyceride transfer protein (MTP) and perhaps squalene synthase inhibitors and agonists of AMP kinase. Future emphasis will be on correcting all lipid abnormalities for the prevention of CHD, not just lowering LDL cholesterol.     

2个丹参鲨烯合酶基因的克隆和鉴定

目的 克隆与鉴定丹参Salvia miltiorrhiza鲨烯合酶基因。方法 基于丹参基因组的基因预测结果设计引物,通过RT-PCR方法,克隆丹参鲨烯合酶基因。利用生物信息软件对基因序列进行了结构分析,对基因编码多肽进行了序列同源性比较和保守结构域分析,利用实时定量RT-PCR方法对基因进行表达模式研究。结果 通过RT-PCR方法扩增得到2个丹参鲨烯合酶基因（SmSQS1和SmSQS2）,它们编码的多肽具有鲨烯合酶相关结构域和保守基序。这2个基因具有不同的外显子/内含子结构特征,具有不同的组织特异性表达模式和时间表达模式。结论 丹参基因组中存在2个鲨烯合酶基因,它们具有不同的基因结构特征和表达模式,可能在丹参甾体类和三萜类化合物生物合成中起不同作用。