樟芝多糖通过CD4+CD25+Foxp3+调节性T细胞对小鼠非酒精性脂肪性肝病的保护作用

目的 研究樟芝多糖通过CD4⁺CD25⁺Foxp3⁺调节性T细胞（Treg）的调控对小鼠非酒精性脂肪性肝病（NAFLD）的保护作用。方法 高脂饮食构建小鼠NAFLD模型,设置对照组、模型组、低剂量组、高剂量组。樟芝多糖干预1~4周中每周流式细胞术检测外周血Treg细胞的比例,谷丙转氨酶（ALT）、谷草转氨酶（AST）的表达,外周血中转化生长因子β（TGF-β）、白介素-6（IL-6）的表达。樟芝多糖干预4周后,小鼠处死,取肝脏进行油红染色,Western blot法检测肝组织中TGF-β和Foxp3、Smad3蛋白的表达,RT-qPCR检测IL-6、TGF-β、Foxp3、Smad3的mRNA表达。结果 高脂饮食喂养4周后成功构建NAFLD小鼠模型,且模型组中Terg比例显著低于对照组（P<0.05）,樟芝多糖干预后Treg比例相比模型组显著增高（P<0.05）,小鼠肝功能得到显著改善,外周血中TGF-β表达上调、IL-6的表达下调,肝组织中TGF-β和Foxp3、Smad3蛋白和mRNA均上调,而IL-6的mRNA表达下调。结论 樟芝多糖可以通过Treg和TGF-β-Smad3信号对NAFLD起到保护作用,作用机制和免疫改善有关。     

樟芝多糖保护小鼠急性肝损伤的机制研究

目的 研究樟芝多糖保护小鼠急性肝损伤的机制。方法 通过D-氨基半乳糖构建急性肝损伤小鼠,C57BL/6小鼠分为对照组、模型组、樟芝多糖高剂量组、樟芝多糖低剂量组,樟芝多糖高、低剂量组每日分别给予50,25 mg·kg^-1的樟芝多糖2次,对照组和模型组给予等体积的生理盐水。给药7 d后紫外比色法测试盒检测大鼠肝组织中谷丙转氨酶（ALT）、谷草转氨酶（AST）水平;TBA法试剂盒检测肝组织中丙二醛（MDA）的水平;ELISA试剂盒检测肝组织中白介素-6（IL-6）的表达;Western-Blot检测肝脏组织中NLRP-3、白介素-1β（IL-1β）、pro-caspase-1和caspase-1的表达;HE染色观察小鼠肝脏病理;RT-qPCR检测肝脏组织中Bcl-2、Bax、caspase-3、caspase-1的mRNA表达。结果 与模型组相比,樟芝多糖可以明显改善急性肝损小鼠的肝脏病理,降低ALT、AST的表达以及MDA和IL-6的表达（P<0.01）,肝脏组织中NLRP-3、IL-1β以及pro-caspase-1和caspase-1的表达相比模型组显著降低（P<0.05）,Bax、caspase-3、caspase-1的mRNA表达相比模型组显著降低（P<0.01）,Bcl-2显著升高（P<0.01）。结论 樟芝多糖对于小鼠急性肝损伤有着很好的保护作用,其作用机制与炎性小体NLRP-3及其相关炎症因子的抑制有关。     

樟芝多糖辅助bFGF对于大鼠机械性脊髓损伤后神经功能修复的作用研究

目的 研究樟芝多糖联合碱性成纤维细胞因子（basic fibroblast growth factor,bFGF）对于机械性脊髓损伤的大鼠神经功能修复的作用。方法 采用改良的Allens法构建大鼠脊髓机械性损伤模型,将大鼠分为对照组、模型组、bFGF组、樟芝多糖组、bFGF+樟芝多糖组,药物干预时间为30 d。在药物干预后第1,7,14,21,30天对大鼠进行脊髓损伤（Basso-Beattie-Bresnahan,BBB）评分、诱发电位（somatosensory evoked potential,SEP）试验、运动诱发电位（motor evokedpotential,MEP）试验,30 d后处死大鼠,提取脊髓组织后进行HE染色和Nissl染色,Elisa检测组织中炎症因子肿瘤坏死因子（TNF-α）、白介素1β（IL-1β）、趋化因子10（CXCL-10）、集落刺激因子（GM-CSF）和白介素10（IL-10）的表达。结果 bFGF、樟芝多糖单一使用对小鼠神经功能BBB评分和SEP、MEP试验结果均优于模型组（P<0.05）,而bFGF+樟芝多糖组的BBB评分和SEP、MEP试验优于单一用药组（P<0.05）。bFGF组、樟芝多糖组、bFGF+樟芝多糖组的脊髓组织中炎症因子TNF-α、IL-1β、CXCL-10的表达显著低于模型组（P<0.05）,而GM-CSF和IL-10的表达高于模型组（P<0.05）,且bFGF+樟芝多糖组显著优于bFGF组和樟芝多糖组（P<0.05）。结论 樟芝多糖辅助bFGF对于机械性脊髓损伤大鼠有着良好的神经功能修复作用,且优于使用单一的bFGF,其作用可能与抗炎症因子的表达,促进神经细胞的存活有关。     

排脓散对葡聚糖硫酸钠诱导结肠炎小鼠的治疗作用

目的 考察排脓散对结肠炎小鼠的治疗作用，为其临床使用提供依据。方法 采用♂BALB/c小鼠，设置正常组、模型组、柳氮磺吡啶组和排脓散高、中、低剂量组，在日常饮水中给予3%葡聚糖硫酸钠（dextran sodium sulfate，DSS）构建结肠炎模型，通过测定小鼠体质量、疾病活动指数（disease activity index，DAI）评分情况、结肠长度和结肠炎性因子表达水平，研究排脓散对小鼠结肠炎的治疗作用。结果 与模型组相比，排脓散各剂量组和柳氮磺吡啶组的小鼠体质量下降均有不同程度的缓解，DAI评分均显著降低（P<0.05），并部分恢复结肠长度。排脓散可以减少结肠部位IL-1β、TNF-α和IL-6表达，提高IL-10表达。结论 排脓散可以减轻DSS诱导的结肠炎症状，改善炎症相关因子表达。     

乙醇分级沉淀的樟芝多糖对于小鼠急性肝损伤的保护作用

目的 研究不同浓度乙醇沉淀获得的樟芝多糖对于急性肝损伤小鼠的保肝作用。方法 采用不同浓度的乙醇沉淀获得樟芝多糖,建立D-氨基半乳糖的急性肝损伤小鼠,Elisa法检测各组血清的谷丙转氨酶（ALT）,谷草转氨酶（AST）,肿瘤坏死因子（TNF-α）、白介素-6（IL-6）、白介素-10（IL-10）的表达以及肝组织中SOD、CAT、GSH-Px、GSH的表达,肝脏HE染色检查组织病理,qPCR检测肝组织中Bcl-2、Bax、Caspase-3的mRNA的表达。确定樟芝多糖中对于小鼠急性肝损伤有效的部分。结果 樟芝多糖的各浓度的乙醇沉淀物均有一定的保肝作用,其中90%乙醇沉淀的多糖对于小鼠模型ALT、AST的含量具有显著的降低作用且优于其他组多糖（P<0.05）。4种多糖可以显著降低血清中肿瘤坏死因子（TNF-α）、白介素-6（IL-6）、和肝组织中Bax、Caspase-3mRNA的表达,提高白介素-10（IL-10）和肝组织中SOD、CAT、GSH-Px、GSH蛋白表达以及Bcl-2mRNA的表达（P<0.05）。且PW90多糖效果较好。结论 不同浓度乙醇沉淀的樟芝多糖对于急性肝损伤有着一定的保护作用,90%乙醇沉淀获得的多糖保肝作用优于其他浓度乙醇沉淀获得的多糖。     

盐酸小檗碱对溃疡性结肠炎小鼠结肠组织TNF-α、IL-1β和IL-10表达的影响

目的 观察盐酸小檗碱（berberine hydrochloride,BBR）对溃疡性结肠炎模型小鼠结肠组织TNF-α、IL-1β和IL-10表达的影响,探讨其治疗UC的可能作用机制。方法 BALB/c小鼠随机分为模型对照组、BBR低、高剂量组、柳氮磺吡啶阳性对照组和空白对照组。采用右旋葡聚糖硫酸钠法复制UC小鼠模型后,每日灌胃给药1次,连续7 d。实验期间每天观察小鼠一般情况并评估疾病活动指数（DAI）;末次给药后解剖观察并评估结肠大体形态损伤指数（CMDI）;组织切片染色,光镜下观察结肠组织的病理学变化并评估组织损伤指数（TDI）;ELISA酶联法检测结肠组织中细胞因子TNF-α、IL-1β和IL-10的含量。结果 与模型对照组比较,BBR治疗组小鼠的结肠炎临床表现明显改善,其DAI、CMDI和TDI评分显著下降,小鼠结肠组织中TNF-α、IL-1β水平均明显降低,IL-10水平升高（P<0.05）。结论 BBR能有效治疗UC小鼠的结肠炎症,其机制可能与其抑制结肠组织中TNF-α和IL-1β、提高IL-10的表达有关。     

樟芝多糖抑制6-OHDA诱导的多巴胺能神经元细胞炎症反应的作用机制研究

目的 探究樟芝多糖通过抑制ROS-NLRP3-caspase-1途径调节6-羟基多巴胺（6-hydroxydopamine,6-OHDA）诱导的多巴胺能神经元（dopaminergic neurons,DAN）细胞炎症反应的作用。方法 分离小鼠中脑DAN细胞,采用6-OHDA体外构建帕金森病细胞模型,将细胞分为正常组、模型组、对照组、实验组。正常组为常规培养的DAN细胞,模型组为6-OHDA处理的DAN细胞,对照组为ROS抑制剂乙酰半胱氨酸（NAC）+6-OHDA处理的DAN细胞,实验组为6-OHDA+樟芝多糖处理的DAN细胞。采用CCK-8法检测细胞活力,流式细胞术和免疫荧光染色法检测ROS的水平,流式细胞术检测细胞凋亡水平,Hoechst 33342染色活细胞,蛋白免疫印迹（Western-bolt）法检测细胞中NLRP3、caspase-1、pro-caspase-1的表达水平,酶联免疫吸附（Elisa）法检测上清中IL-1β、IL-6和IL-18的分泌水平。结果 模型组中6-OHDA可以诱导DAN细胞炎症反应,ROS表达增高,NLRP3-caspase-1炎性小体水平增高,细胞凋亡率增高,相比正常组具有显著性差异（P<0.05）。樟芝多糖干预后,ROS的水平下调,NLRP3-caspase-1炎性小体水平降低,细胞凋亡率下调,相比模型组具有显著性差异（P<0.05）。结论 樟芝多糖可以通过抑制ROS-NLRP3-caspase-1途径调节6-OHDA诱导DAN炎症反应,这可能是樟芝多糖在帕金森病炎症反应中的作用机制之一。     

葡萄籽提取物与柳氮磺吡啶联合用药对实验性溃疡性结肠炎的作用

目的 研究葡萄籽提取物（GSPE）与柳氮磺吡啶（SASP）联合用药治疗葡聚糖硫酸钠（DSS）诱导的小鼠溃疡性结肠炎的效果及作用机制。方法 将SPF级C57小鼠随机分为对照组、模型组、GSPE （250 mg/kg）组、SASP （250 mg/kg）组、联合用药（SASP 250 mg/kg+SASP 250 mg/kg）组,对照组给予正常饮用水,其他组均自由饮用3% DSS水溶液,连续饮用7 d,诱发小鼠溃疡性结肠炎模型;造模第1天同步开始ig给药,每天记录小鼠体质量、便血、便型等症状变化;7 d后断头取血,收集结肠和脾脏,记录结肠长度、脾脏质量变化情况。HE染色评估小鼠结肠黏膜组织病理变化,ELISA法检测血清和结肠组织中炎症因子肿瘤坏死因子（TNF-α）、白细胞介素（IL）-1β、IL-6和NO的表达变化以及丙二醛（MDA）、超氧化物歧化酶（SOD）细胞因子的含量变化,免疫组化分析结肠组织上皮细胞中NF-κB-p65、Nrf2、HO-1和Keap-1含量的变化。应用金氏Q值法分析联合用药的作用效果。结果 与模型组比较,各给药组的小鼠体质量下降幅度显著降低（P<0.01）,小鼠腹泻、便血症状改善明显,其中联合用药组小鼠体质量较单独给药组下降更缓,小鼠腹泻、便血程度改善更明显（P<0.05）。各给药组小鼠血清及结肠组织中IL-1β、IL-6、TNF-α、NO、MDA含量较模型组显著降低,SOD含量则显著升高（P<0.01）;病理组织切片分析发现,各给药组小鼠结肠黏膜病理损伤较模型组显著减轻,其中联合用药组小鼠结肠黏膜病理损伤较单独给药组降低更为显著（P<0.05）。免疫组化分析结果也显示,各给药组小鼠的NF-κB-p65、Keap-1蛋白表达与模型组比较显著下调（P<0.01）,Nrf2、HO-1蛋白表达显著上调（P<0.01）。联合给药组与单独给药组比较,NF-κB-p65、Keap-1蛋白表达显著降低（P<0.05）,Nrf2、HO-1蛋白表达显著升高（P<0.05）。金氏Q值法评价两药合用后对IL-1β、IL-6、TNF-α、NO、MDA、NF-κB-p65、Keap-1的作用效果均大于1.15,说明GSPE与SASP具有协同作用。结论 GSPE与SASP联合用药治疗实验性溃疡性结肠炎效果优于单独给药,联合用药增强GSPE和SASP的抗氧化和抗炎作用,GSPE与SASP配伍治疗溃疡性结肠炎可进一步发挥协同增效的作用。     

氧化苦参碱介导细胞自噬减轻溃疡性结肠炎小鼠结肠黏膜氧化性损伤的作用机制研究

目的 探讨氧化苦参碱介导细胞自噬减轻溃疡性结肠炎（ulcerative colitis,UC）小鼠结肠黏膜氧化性损伤的作用机制。方法 采用2,4,6-三硝基苯磺酸灌肠复制小鼠UC模型,将造模成功小鼠按体质量随机分为模型组、氧化苦参碱组（50 mg·kg^-1·d^-1,ig）、羟基氯喹组（50 mg·kg^-1·d^-1,ig）、氧化苦参碱+羟基氯喹组,另设正常组,每组10只。治疗1周后测定各组小鼠疾病活动度（disease activity,DAI）、结肠质量系数及病理形态;MitoSOX Red法测定结肠ROS含量,ELISA法测定结肠组织SOD、MDA、MPO、GSH-PX含量;透射电镜结合免疫荧光观测结肠细胞自噬程度,Western blot测定Atg5和Beclin-1蛋白表达。结果 与模型组和羟基氯喹组比较,氧化苦参碱组小鼠DAI和结肠质量系数均有显著减小（P<0.01）,结肠病理损伤明显减轻;ROS、MDA和MPO含量极显著降低（P<0.01）,SOD和GSH-PX含量极显著增加（P<0.01）;结肠黏膜细胞自噬,程度显著增强,Atg5和Beclin-1蛋白表达极显著上调（P<0.01）。结论 氧化苦参碱可促进UC小鼠细胞自噬,减轻小鼠结肠黏膜氧化性损伤。     

全反式维甲酸对兔颈动脉粥样硬化斑块中TGF-β1、IL-10、MMP-2和MMP-9表达的影响

目的 采用实时荧光定量PCR（qRT-PCR）法探索全反式维甲酸（ATRA）对兔颈动脉粥样硬化斑块组织炎症因子表达的影响。方法 选取24只纯种雄性新西兰白兔随机分为对照组、模型组、ATRA组。对照组给予基础饲料、其余两组给予高脂饲料，共饲养12周。饲养4周后，ATRA组ig 5 mg/kg的ATRA，模型组和对照组ig等量溶剂，每天1次，至12周末处死各组动物。取颈动脉粥样硬化斑块组织，进行HE染色，显微镜下观察管壁结构；提取组织RNA，qRT-PCR法测定组织中转化生长因子-β1（TGF-β1）、白细胞介素-10（IL-10）、基质金属蛋白酶-2（MMP-2）和基质金属蛋白酶-9（MMP-9） mRNA的相对表达量。结果 模型组可见典型的AS斑块，可见薄层的纤维帽及大的脂质核心，以及大量炎症细胞浸润，内弹力纤维膜断裂；ATRA组内皮细胞形态基本完整，可见少量炎性细胞浸润。与对照组比较，模型组中TGF-β1和IL-10 mRNA的相对表达量明显降低（P<0.05），而MMP-2和MMP-9 mRNA的相对表达量明显升高（P<0.05）；与模型组比较，ATRA组的TGF-β1和IL-10 mRNA的相对表达量明显升高（P<0.05），MMP-2和MMP-9 mRNA的相对表达量明显降低（P<0.05）。结论 ATRA可通过调节炎症因子的表达抗动脉粥样硬化。     

Allele and genotype frequency of CYP2C9 in Tamilnadu population

Objectives To identify the frequency of CYP2C9*1, *2 and *3 alleles and the genotype of CYP2C9 gene in the Tamilian population.Methods The study was conducted on 135 unrelated healthy human volunteers. DNA was extracted from the peripheral leukocytes samples and was analyzed using the polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) protocol. The PCR products were digested with AvaII, KpnI or NsiI restriction enzymes. The digested products were separated using 8% polyacrylamide gel and stained by ethidium bromide. Genotyping of the subjects was done based on DNA fragment size.Results The frequencies of CYP2C9*1, *2 and *3 alleles in the Tamilian population were 0.907, 0.026 and 0.067, respectively. The distribution of CYP2C9*1/*1, *1/*2, *1/*3 and *2/*3 genotypes were 0.823, 0.044, 0.126 and 0.007, respectively.Conclusion CYP2C9*3 is the most frequent mutant allele found in the Tamilian population. The distribution of this mutant allele in the Tamilian population was found to be lesser than in Caucasians but higher than in Chinese.     

11-酮基-△9-炔诺酮的合成

We have introduced a carbonyl group at the 11-position and a double bond at the 9-position of norethisterone in order to increase its antifertility potency. The starting material for the preparation of 11-keto-Δ⁹-noret-histerone(9) was 19-hydroxy-Δ⁴-androstene-3,17-dione(1), which was converted to the title compound by partial synthesis through the sequence of reactions:(1)→(9).11-Keto-Δ⁹-norethisterone was found to possess very potent antifertility effect by preliminary screening tests.     

CYP2C9*3 allelic variant is associated with metabolism of irbesartan in Chinese population

Objective There is considerable variability in the individual pharmaceutical dosages required to achieve optimal therapeutic effects, which may be due to environmental or genetic factors. The objective of this study was to test the presence of the CYP2C9*3 allelic variant in the Chinese population and to investigate the association of this variant with both metabolism and therapeutic efficacy of irbesartan on essential hypertension. Methods In this study, we enrolled 711 subjects from Taihu County and 376 subjects from Dongzhi County in Anhui Province, China. All subjects received a single oral dose of 150 mg irbesartan daily for 28 days. The plasma concentration of irbesartan at 24 h after dosing on the 27th day and at 6 h after dosing on the 28th day was detected using fluorescence-high-performance liquid chromatography. CYP2C9 genotypes were determined using polymerase chain reaction—restriction fragment length polymorphism. Results No CYP2C9*2 allele was found in 235 Chinese samples and was removed from further study. The mean frequency of the CYP2C9*3 allele was 3.65%, while no CYP2C9*3/*3 genotype was detected. Multiple linear regression analyses revealed that the CYP2C9*3 allele carriers had significantly higher irbesartan concentrations in plasma at 6 h (Taihu: P<0.0001; Dongzhi: P=0.03) and 24 h (Taihu: P<0.0001; Dongzhi: P=0.00013) after dosing. No significant association was found between the CYP2C9*3 allelic variant and the therapeutic effect of irbesartan on essential hypertension. Conclusion Our study suggests that the CYP2C9*3 plays an important role in the metabolism of irbesartan and/or is in linkage disequilibrium with another potential CYP2C9 allele, both of which possibly modify the pharmacokinetics of irbesartan.     

CYP2C9 genotypes and the quality of anticoagulation control with warfarin therapy among Brazilian patients

Objective To evaluate the impact of the two most common CYP2C9 variant alleles (*2 and *3) on the maintenance dose of warfarin and on the quality of anticoagulation control in Brazilians. Methods Patients (n = 103) initiated warfarin therapy with 5 mg/day (or 2.5 mg/day when over 80 years old). The international normalized ratio (INR) was targeted between 2 and 3, monitored every week until four consecutive adequate measures had been obtained, and then monthly. Serious hemorrhagic events were defined by the need for inpatient hospitalization. CYP2C9 genotyping was obtained by PCR-RFLP. Results The frequencies of CYP2C9*2 and CYP2C9*3 were 0.097 and 0.073, respectively, with genotypic distribution fitting Hardy-Weinberg equilibrium. CYP2C9 genotype was the only clinical feature associated with the risk of severe bleeding (one-sided P = 0.019, Fisher exact method), with an odds ratio of 4.8 (95% confidence interval of 1.4–16.6) for any variant genotype as compared to CYP2C9*1*1. Patients with either CYP2C9*2 or CYP2C9*3 were equally difficult to maintain in the INR target range, showing significantly (one-sided P = 0.038, Mann-Whitney U-test) reduced ratio of adequate INR measures (0.54 ± 0.2), when compared to CYP2C9*1*1 patients (0.63 ± 0.2). Patients with CYP2C9*3, but not CYP2C9*2, required significantly (one-sided P = 0.001, Mann-Whitney U-test) lower warfarin maintenance doses (3.1 ± 1.8 mg) than CYP2C9*1*1 patients (5.3 ± 2.1 mg). Conclusion Patients with either CYP2C9*2 or CYP2C9*3 show higher risk of over-anticoagulation compared to CYP2C9*1*1 subjects and could benefit from a reduction in the initial warfarin standard dose (e.g., to 2.5 mg/day).     

662 A/G gene variation in human tumor necrosis factor receptor superfamily,member 9 (TNFRSF9)

The aim of this paper is to report a new coding variance of the TNFRSF9 gene, a candidate for auto-immune diseases. We found the variation in two families with type 2 diabetes mellitus by D-HPLC mutation screening method and confirmed our results by direct sequencing and PCR-RFLP. Although without changing the amino acid coding, the variance may have an effect on codon usage and play a role in disease development, such as type 2 diabetes mellitus. However, we cannot define the role of this variance because the frequency of the minor allele is low in the Chinese population and no homozygote of the variance was found. More research in multiple populations will be necessary to define the role of this variance.     

Cytochrome P 450 2C9 phenotyping using low-dose tolbutamide

Objectives The hypoglycaemic drug tolbutamide is used for assessment of CYP2C9 activity in vivo. However, therapeutically active doses of 500 mg bear the risk of hypoglycaemia, and a tolbutamide-derived parameter based on a single plasma or urine concentration reflecting CYP2C9 activity accurately is lacking.Methods We examined tolbutamide and its metabolites 4-hydroxy-tolbutamide and carboxytolbutamide in plasma and urine of 26 healthy, male volunteers up to 24 h after intake of 125 mg tolbutamide using liquid chromatography–tandem mass spectrometry. CYP2C9 genotypes were determined by sequencing of exons 3 and 7. Raw plasma and urine data were compared with pharmacokinetic parameters, CYP2C9 genotypes, and data from a study in 23 volunteers with all six CYP2C9*1–*3 combinations who received 500 mg tolbutamide.Results Plasma clearance and tolbutamide plasma concentrations 24 h after drug intake reflected the genotypes: 0.85 l/h and 1.70 µg/ml (95% confidence interval, CI, 0.80–0.89 l/h and 1.50–1.90 µg/ml) for CYP2C9*1 homozygotes (n=15), 0.77 l/h and 2.14 µg/ml (95%CI, 0.67–0.88 l/h and 1.64–2.63 µg/ml) for *1/*2 genotypes (n=7), 0.60 l/h and 3.13 µg/ml (95%CI, 0.58–0.62 l/h and 2.68–3.58 µg/ml) for *1/*3 genotypes (n=3), and 0.57 l/h and 3.27 µg/ml in the single *2/*2 carrier. Natural logarithms of tolbutamide plasma concentrations 24 h after intake correlated to plasma clearance (r²=0.84, P<0.0000001). This correlation was confirmed in the comparison data set (r²=0.97, P<0.0000001).Conclusions A low dose of 125 mg tolbutamide can safely and accurately be used for CYP2C9 phenotyping. As a simple metric for CYP2C9 activity, we propose to determine tolbutamide in plasma 24 h after drug intake.     

Induction of several cytochrome P450 genes by doxorubicin in H9c2 cells

Doxorubicin is a potent anti-neoplastic antibiotic used to treat a wide variety of malignancies; however, its use is limited by dose dependent cardiotoxicity. There is indirect evidence suggesting that doxorubicin cardiotoxicity is CYP-mediated. In the current study, we investigated the effect of doxorubicin on hypertrophic markers, and different CYP gene expression in cardiac derived H9c2 cells. H9c2 cells were incubated with increasing concentrations of doxorubicin and the expressions of different genes were determined by real-time PCR. Our results demonstrate that multiple CYP genes are expressed in H9c2 cells and the level of expression from the highest to the lowest were; CYP1B1, CYP2B1, CYP2J3, CYP1A1, CYP2C11, CYP2C23, CYP2E1, CYP1A2, and CYP2B2. Doxorubicin treatment caused an induction of the hypertrophic markers, ANP and BNP. In addition, doxorubicin caused a significant induction of CYP1A1, CYP1A2, CYP1B1, CYP2B2, CYP2E1, and CYP2J3 gene expression in a concentration-dependent manner. However, only the highest concentration tested, 10 μM, caused an induction of CYP2C11; whereas, CYP2B1 and CYP2C23 were not altered. Our findings demonstrate that doxorubicin induces the hypertrophic markers, ANP and BNP as well as several CYP genes in H9c2 cells. Doxorubicin-mediated CYP induction may represent a novel mechanism by which this drug induces cardiotoxicity.     

13-Hydroxy-9Z,11E,15E-octadecatrienoic acid from the leaves of Cucurbita moschata

A new unsaturated hydroxy fatty acid was isolated from the leaves of Cucurbita moschata through repeated silica gel column chromatography and chemical methods. The structure of the new fatty acid was determined as 13-hydroxy-9,11,15-octadecatrienoic acid on the basis of several spectral data including 2D-NMR. The stererostructures of double bonds were determined to be 9Z, 11E and 15E by coupling patterns of related proton signals in the 1H-NMR and NOESY experiments.     

Varying the Unsaturation in N 4,N 9-Dioctadecanoyl Spermines: Nonviral Lipopolyamine Vectors for More Efficient Plasmid DNA Formulation

Purpose The aim of the study is to analyze the effect of varying the degree of unsaturation in synthesized N⁴,N⁹-dioctadecanoyl spermines on DNA condensation and then to compare their transfection efficiency in cell culture. Methods The N⁴,N⁹-di-C18 lipopolyamines—saturated (stearoyl), C9-cis- (oleoyl), and C9,12-di-cis- (linoleoyl)—were synthesized from the naturally occurring polyamine spermine. The ability of these novel compounds to condense DNA and form nanoparticles was studied using ethidium bromide fluorescence quenching and nanoparticle characterization techniques. Transfection efficiency was studied in several primary skin cells (FEK4, FCP4, FCP5, FCP7, and FCP8) and in an immortalized cancer cell line (HtTA) and was compared with the commercially available nonliposomal transfection formulation Transfectam^? (dioctadecylamidoglycyl spermine), which also contains two saturated C18 lipid chains. Results N⁴,N⁹-Dilinoleoyl spermine (C18, di-cis-9,12) is efficient at circular plasmid DNA (pEGFP) condensation and gives the most effective transfection in a series of primary skin cells and cancer cell lines at low charge ratios of 5.5 (± ammonium/phosphate). Conclusions The dienoic fatty acyl spermine conjugate N⁴,N⁹-dilinoleoyl spermine efficiently condenses DNA and achieves the highest transfection levels among the studied lipopolyamines in cultured cells.     

Effects of fluvastatin and cigarette smoking on CYP2C9 activity measured using the probe S-warfarin

Objective The effect of cigarette smoking on CYP2C9 activity is unknown. We conducted a study to evaluate whether there is a difference in CYP2C9 activity in smokers versus non-smokers by examining S-warfarin AUC after CYP2C9 inhibition with fluvastatin. In addition, the effect of the CYP2C9 inhibitor fluvastatin was evaluated using S-warfarin as a probe.Methods A randomized, single dose, two-treatment crossover study of warfarin with a washout period of 21 days was performed. Eighteen healthy Caucasian smokers and non-smokers, genotyped as CYP2C9*1/*1 or CYP2C9*1/*2, received warfarin 10 mg plus vitamin K 10 mg to measure baseline CYP2C9 activity. Warfarin dosing was repeated after 18 days of fluvastatin 40 mg twice daily to evaluate CYP2C9 activity after inhibition.Results The S-warfarin between smokers and non-smokers did not differ by >25% after inhibition. There was no difference in S-warfarin during baseline (p=0.45) or inhibition (p=0.19) periods for smokers versus non-smokers. Fluvastatin increased the AUC of S-warfarin by 42±29% and 26±18% in smokers and nonsmokers, respectively. Linear regression analyses showed significant but weak correlations between peak concentrations (C_{at 1 h}) or (−) 3S,5R-fluvastatin AUC_{0–12 h} and extent of warfarin inhibition. For (+) 3R,5S-fluvastatin, a weak correlation was found between C_{at 1 h} and extent of warfarin inhibition.Conclusions Cigarette smoking does not affect CYP2C9 activity as evaluated using S-warfarin as a CYP2C9 probe. Fluvastatin is a weak inhibitor of CYP2C9 activity in both smokers and non-smokers.