Auto-inhibition of verapamil metabolism in rat hepatocytes of gel entrapment culture

Mechanism-based inactivation (MBI) of cytochrome P450 (CYP) 3A by verapamil and the resulting drug-drug interactions have been studied in vitro, but the inhibition of verapamil on its own metabolic clearance in clinic, namely auto-inhibition of verapamil metabolism, has never been reproduced in vitro. This paper aimed to evaluate the utility of gel entrapped rat hepatocytes in reflecting such metabolic auto-inhibition using hepatocyte monolayer as a control. Despite being a similar concentration- and time-dependent profile, auto-inhibition of verapamil metabolism showed apparent distinctions between the two culture models. Firstly, gel entrapped hepatocytes were more sensitive to such inhibition, which could be largely due to their higher CYP3A activity detected by the formation rates of 6β-hydroxy testosterone and 1’-hydroxy midazolam. Furthermore, the inhibitory effect of ketoconazole and verapamil on CYP 3A activity as well as the reduction of verapamil intrinsic clearance (CL_int) by ketoconazole was only observed in gel-entrapped hepatocytes. In this respect, the involvement of CYP3A in auto-inhibition of verapamil metabolism could be illustrated in gel-entrapped hepatocytes but not in hepatocyte monolayer. All of these results indicated that hepatocytes of gel entrapment reflected more of verapamil metabolic auto-inhibition than hepatocyte monolayer and could serve as a suitable system for investigating drug metabolism.     

New perspectives in bio-analytical techniques for preclinical characterization of a drug candidate: UPLC-MS/MS in in vitro metabolism and pharmacokinetic studies

Lead optimization requires rapid bio-analytical turnover for the generation of early absorption, distribution, metabolism, excretion (ADME) and pharmacokinetics (PK) data maintaining a high quality level. Therefore, one of the major challenges in the bio-analytical field is to achieve faster and more sensitive quantification protocols. In the present communication, a comparison between HPLC and ultra performance liquid chromatography (UPLC) performances in terms of sensitivity and resolution is shown using a pharmakokinetic study and a metabolism study as models. The studies highlight the features of the new technology and the resulting impact in the PK throughput and in the characterization of isomeric metabolites using UPLC/MS/MS technique.     

Shortening of the ventricular fibrillatory intervals after administration of verapamil in a patient with Brugada syndrome and vasospastic angina

A 43-year-old man presented with electrocardiographic findings consistent with Brugada syndrome. Though the baseline coronary angiogram was normal, intracoronary infusion of ergonovine maleate caused complete occlusion of the left anterior descending and a 99% occlusion of the proximal right coronary artery, each relieved by intracoronary isosorbide dinitrate. Double extrastimuli delivered at the right ventricular outflow tract induced ventricular fibrillation terminated by a 200-J shock. Verapamil, 10 mg IV, increased ST-segment elevation and programmed stimulation repeated after the drug induced ventricular fibrillation with shorter F-F intervals and lower amplitude signals, which was not terminated by 200 J and required an additional 360-J shock. Ca²⁺ antagonism may have been adverse in this patient with Brugada syndrome because the drug has the potential to increase the voltage gradient through the right ventricle and to slow intraventricular conduction at very fast heart rates.     

低氧时钙离子对冠状动脉内皮细胞释放ET-1、NO、PGI2的影响

目的:探讨低氧引起冠状动脉内皮细胞(CAEC)释放ET-1、NO和PGI₂改变的机制。方法:运用放射性[⁴⁵Ca²⁺]掺入、放免和比色等方法, 测定常氧组、低氧组冠状动脉内皮细胞钙摄取率, 以及常氧组、低氧组、低氧+维拉帕米组冠状动脉内皮细胞培养液中ET-1、NO和PGI₂含量。结果:低氧组CAEC[⁴⁵Ca²⁺]摄取率为(29.6±3.7)ng/gprotein/30min, 常氧对照组为(15.2±1.1)ng/gprotein(P<0.01).低氧+维拉帕米组ET-1分泌量显著多于低氧组(P<0.01).低氧+维拉帕米组分泌的NO显著低于低氧组(P<0.05), 与常氧组相差不显著。低氧+维拉帕米组分泌的PGI₂显著多于低氧组, 与常氧组相差不显著。结论:低氧时引起的NO、ET-1、PGI₂分泌改变与低氧引起的冠状动脉内皮细胞Ca²⁺内流增加有关。     

维拉帕米对耐苯妥英钠和卡马西平癫(癎)模型大鼠电生理和行为的影响

目的 观察P-糖蛋白(PGP)拮抗剂维拉帕米对耐苯妥英钠(PHT)和卡马西平(CBZ)癫(癎)大鼠电生理指标和行为学的影响.方法 建立慢性杏仁核点燃癫(癎)大鼠模型,筛选出耐药组和治疗有效组,给予维拉帕米,观察其对各组大鼠后放电阈值(after discharge threshold,ADT)、后放电时程(after discharge duration,ADD)等电生理指标和行为学的影响.结果 相对于耐药对照组,耐药组大鼠在预先给予维拉帕米后,使用抗癫(癎)药物后ADT[(238.0±32.2)μA]明显高于耐药对照组[(177.0±23.3)μA,P＜0.05];ADD时程明显缩短,Racine行为分级明显下降(P＜0.05).结论 维拉帕米可以协助抗癫(癎)药物(AEDs)改善耐药大鼠的电生理活动,降低点燃后发作分级,提示有效抑制PGP有助于改善多药转运体高表达导致的癫(癎)耐药.     

维拉帕米对家兔快速心房起搏所致心房结构和电重构的影响

目的:观察L-钙离子通道阻断剂维拉帕米对家兔快速心房起搏所致结构和电重构的作用. 方法:将24只家兔随机分为:对照组、快速起搏组和维拉帕米组.经颈内静脉将电极置入兔右心房.分别测定各组在0、2、4、6和8 h(记为P0、P2、P4、P6、P8)时的心房有效不应期(AERP200,和AERP150).左心房和肺静脉心肌袖组织行HE染色观察组织学改变. 结果:快速心房起搏组在不同基础刺激作用下AERP缩短,AERP200-AERP150的频率适应性不良,P8与起搏前P0比较差异显著(P<0.05).左心房和肺静脉心肌袖组织学改变明显.维拉帕米组AERP基本无改变(P>0.05),AERP200-150,频率适应性维持.左心房和肺静脉心肌袖与起搏组相比组织学改变较轻.结论:维拉帕米可以抑制快速心房起搏所致电重构,但不能逆转结构重构.     

Embryonic stem cell-derived cardiomyocytes as a model system to study cardioprotective effects of dexamethasone in doxorubicin cardiotoxicity

Embryonic stem cell (ESC)-derived beating cardiomyocytes may be considered as a suitable model for in vitro assessment of pharmacological and toxicological studies. In this model, laboratory animals are not required. In addition, physiological functions, such as heart beat, are assessed rather than single parameters such as cell viability. Here we report that doxorubicin (DOX) cardiotoxicity on mouse ESC-derived beating cardiomyocytes can be ameliorated by treatment with dexamethasone (DEX) when DEX is administrated only before DOX and not in combination with DOX. DEX effect appears to be mediated via glucocorticoid receptor and increases cardiomyocyte-specific gene expression. Cardiotoxicity of DOX can be augmented by calcium channel blocker, verapamil (VER) which also decreases the expression of cardiac gene markers. This model provides us with a clinical suggestion which proposes that the beneficial effect of DEX is obtained when DEX was added before DOX administration.     

冠状动脉内注射腺苷与维拉帕米治疗急性冠状动脉 综合征介入术中缓再流现象的对照研究

目的对照观察冠状动脉内注射腺苷与维拉帕米对急性冠状动脉综合征(ACS)介入术中缓再流现象的疗效、副反应及对患者预后的影响.方法选取33例接受经皮冠状动脉腔内成形术和支架术(PTCA/STENT)后发生缓再流现象的急性冠状动脉综合征患者,随机分为腺苷组(17例)和维拉帕米组(16例).腺苷组在发生缓再流现象的病变血管内注射腺苷3 mg,维拉帕米组在病变血管内注射维拉帕米2.5 mg.注射后1、5、10 min后重复冠状动脉造影,记录注药前后病变血管内血流TIMI分级,并应用TIMI血流计帧法(TIMI frame count, TFC)定量测定血流速度,进行比较.详细记录用药前后患者血压、心率和心电图变化.结果注药前腺苷组和维拉帕米组TIMI血流分级分别为1.65±0.38和1.56±0.33,血流帧数分别为87.9±15.3和89.5±17.4,两组血流速度无明显差异(P>0.05).腺苷组在冠脉内注药后有9例(53%)患者获得TIMI3级血流,注药后1、5、10 min时血流帧数分别为58.5±13.4、45.8±9.7、50.2±11.6;维拉帕米组注药后有7例(44%)患者获得TIMI3级血流,注药后1、5、10 min时血流帧数分别为68.9±15.6、59.7±14.5、62.3±16.2.两组冠状动脉内注药后血流均有明显改善(血流帧数与注药前比较P<0.05),腺苷组血流改善较维拉帕米组更为显著(P<0.05).冠脉内注射2.5 mg维拉帕米较注射3 mg腺苷对患者血压和心率的影响更明显(P<0.05).住院期间两组均无患者死亡,出院前心脏超声测LVEF在腺苷组为(46.5±10.2)%,维拉帕米组为(43.1±9.8)%,随访6个月,两组心血管事件(心绞痛、心肌梗死、心衰、心因性死亡)发生率分别为29.4%和43.8%,无统计学差异.结论腺苷与维拉帕米均能有效改善急性冠状动脉综合征介入术中缓再流现象.与维拉帕米相比较,腺苷能更安全、有效地改善冠状动脉血流.     

1,6-二磷酸果糖对心肌细胞、淋巴细胞和血清钙含量的影响

1,6-二磷酸果糖(FDP)为对多种代谢径路有调节作用的高能底物,已应用于休克、缺氧等的治疗。肺心病呼吸衰竭应用常规治疗加FDP(5g/日)9例,2周后外围血淋巴细胞(PBL)内Ca含量显著高于应用前和常规治疗组。用10～15kg全麻开胸健康雄犬10条,隔2h先后静注FDP(250mg/kg)和FDP 异搏停(V;0.1mg/kg)。FDP注射后15、30、45分,左室心肌和PBL平均细胞内Ca含量分别较药前增高144—293％和231—259％(P<0.01),血清Ca含量均低于药前(P<0.01)。注射FDP V后则细胞内及血清Ca含量均未见与药前有显著差异。所见提示FDP促使细胞内钙含量增高和细胞外含量降低的效应,可能在于药物作用于慢通道而增强Ca内流所致。