Renal targeting of captopril selectively enhances the intrarenal over the systemic effects of ACE inhibition in rats

1 In previous studies on the renal targeting of the ACE inhibitor captopril, we demonstrated that a 6 fold increased concentration of this drug could be obtained in the kidney after conjugation to the low-molecular-weight protein lysozyme. In this study, we investigated in unrestrained rats whether systemic administration of captopril-lysozyme also results in an enhanced effect on renal parameters, relative to the systemic effects. 2 Renal effects: intravenous infusion of captopril-lysozyme for 6 h resulted in a more pronounced increment of renal blood flow (31+/-2% vs 17+/-4% at 0.5 mg kg(-1) 6h(-1), P<0.01) and an approximately 5 fold enhanced natriuresis (167+/-17% vs 36+/-7% at 1 mg kg(-1) 6 h(-1), P<0.001) in comparison with equimolar amounts of captopril as a free drug. In correspondence with these findings, renal ACE inhibition was potentiated approximately 5 fold (-50+/-4% vs -22+/-3% at 1 mg kg(-1) 6 h(-1), P<0.001). 3 Systemic effects: conjugated captopril did not affect blood pressure in dosages up to 5 mg kg(-1) 6 h(-1). This effect coincided with a less pronounced inhibition of the pressor response to intravenously administered angiotensin I (-12+/-3% vs -66+/-5% at 1 mg kg(-1) 6 h(-1), P<0.001), and a markedly attenuated plasma ACE inhibition (-19+/-2% vs -37+/-3% at 1 mg kg(-1) 6 h(-1), P<0.001) compared to an equivalent dose of free captopril. 4 An experiment of continued intravenous administration of captopril-lysozyme for 7 days in nephrotic syndrome demonstrated that the conjugate is also active in renal disease: the antiproteinuric response was substantially augmented (-67+/-5% vs -15+/-7% at 4 mg kg(-1) 24 h(-1), P<0.001) compared to the free drug, in the absence of blood pressure reduction. 5 These data demonstrate that intravenous administration of a captopril-lysozyme conjugate leads to more selective renal ACE inhibition and enhanced renal effects as well as less systemic effects compared to captopril itself.     

The progression of adriamycin-induced nephrotic syndrome in rats and the effect of captopril

The progression of adriamycin-induced nephrotic syndrome in rats was studied over a 3-month period. The effect of an angiotensin-converting enzyme inhibitor, captopril, on this model of renal disease, was also studied. Two weeks following a single iv injection of adriamycin, rats were divided into two treatment groups: one received a daily po dose of captopril and the other received a placebo. Measurements of renal function were performed at 4, 8, and 11 weeks following the initiation of therapy. Necropsies and light microscopic evaluation of the kidneys were performed at the end of the treatment period. Functional and morphologic alterations in both groups of rats were compared to each other and to normal age/weight-matched control rats studied over the same time period. At 13 weeks following the administration of adriamycin, both treatment groups had significant renal dysfunction when compared to normal controls. In addition to severe proteinuria, rats receiving adriamycin exhibited polyuria, polydipsia, increased plasma urea nitrogen and plasma creatinine, and decreased endogenous creatinine clearance. They had severe generalized kidney lesions characterized by tubular dilation and atrophy, cast formation, interstitial fibrosis and lymphocytic infiltration, and focal, global glomerulosclerosis. The histopathologic ranking of the kidneys was correlated with some antemortem laboratory parameters but not with the degree of proteinuria. Captopril had no ameliorating effects on the progression of renal disease. Certain findings indicate that captopril may actually have promoted the deterioration of renal function. We conclude that adriamycin-induced nephrotic syndrome in the rat is a progressive disease resulting in generalized renal dysfunction, and that captopril, at the dose given in this experiment, is unable to slow the progression of the disease.     

贝那普利、螺内酯联用对阿霉素肾病大鼠肾小管保护作用的研究

目的:观察贝那普利、螺内酯联合应用于阿霉素肾病大鼠降低蛋白尿及增加的肾小管保护作用。方法:42只SD大鼠随机抽取7只为正常对照组,余下经尾静脉注射阿霉素制备肾病模型。6周后29只造模成功的大鼠(24h尿蛋白>100mg)随机分为:模型组(n=7),贝那普利组(n=8),螺内酯组(n=7),贝那普利和螺内酯联合治疗组(联合用药组,n=7)。分别于6、12、18周末收集24h尿液,检测24h尿蛋白,并于18周末测尿视黄醛结合蛋白(RBP)后,处死大鼠经腹主动脉取血液标本检测生化指标。取出肾组织HE染色,并采用免疫组织化学方法检测肾小管间质转化生长因子-β1(TGF-β1)的表达。结果:各治疗组均能显著降低阿霉素肾病大鼠血压、蛋白尿,升高血清白蛋白(P<0.05),其中贝那普利组和联合治疗组疗效优于螺内酯组,以联合治疗组疗效最为显著;与模型组比较各治疗组尿RBP水平均下降(P<0.01),其中联合治疗组疗效优于单独治疗组(P<0.01)。联合治疗组抑制TGF-β1的表达优于治疗组和模型组。治疗组尿RBP和TGF-β1均表现为强相关(r=0.735、r=0.845、r=0.585)。结论:在阿霉素大鼠模型中,贝那普利和螺内酯联合应用能降低蛋白尿,抑制TGF-β1合成,减轻肾小管损伤。     

槲皮素对阿霉素致小鼠心肌损伤的保护作用及其机制

观察槲皮素对阿霉素致小鼠心肌损伤的保护作用并初步探讨其机制。腹腔注射阿霉素(20 mg·kg^-1)复制小鼠心肌损伤模型，检测心电图、心肌超微结构，血清NO含量和iNOS活性，心肌组织LDH、SOD、MDA的水平和p53蛋白表达。并观察槲皮素(50，100及200 mg·kg^-1)对上述指标的影响。阿霉素可导致小鼠心律失常和心肌超微结构损伤；使NO、iNOS、MDA和LDH的水平升高，SOD的水平降低；p53蛋白表达增强。槲皮素(50，100及200 mg·kg^-1)可拮抗阿霉素所致的上述变化。槲皮素对阿霉素性小鼠心肌损伤具有保护作用，其机制与增强SOD活力、降低iNOS活性、抑制p53蛋白表达等有关。     

The antiproteinuric effect of ACE inhibition in renal disease

Proteinuria due to non-steroid-responsive renal disease may be harmful. Firstly, because it may cause a nephrotic syndrome, and, secondly, because it is, like hypertension, associated with an increased risk of progressive renal damage and loss of renal function over the years. Until recently, only non-steroidal antiinflammatory drugs (NSAIDs) were available to reduce proteinuria in such patients. Due to their potential side-effects, however, NSAIDs have never been widely used as antiproteinuric agents. In 1985 some studies were published showing that inhibition of angiotensin-converting enzyme (ACE) not only reduced the elevated blood pressure in rats with chronic renal failure (experimentally induced by renal ablation or by induced diabetic nephropathy), but also prevented the development of glomerular damage with proteinuria and loss of renal function in these animals. This beneficial effect of ACE inhibition was attributed to the prevention of glomerular hypertension. At the same time it was reported that ACE inhibition could reduce proteinuria in patients with advanced diabetic nephropathy. ACE inhibitors might thus be an attractive alternative for NSAIDs as antiproteinuric treatment, possibly being renoprotective, and being generally well-tolerated.     

生脉注射液对大鼠阿霉素心肌损伤的保护作用及其抗细胞凋亡机制研究

目的探讨生脉注射液对阿霉素心肌损伤的作用是否为通过抗细胞凋亡机制。方法腹腔注射阿霉素建立大鼠阿霉素心肌损伤模型。观察大鼠心率变化,计算死亡率;测定各组大鼠肌酸激酶同工酶（CK-MB）;TUNEL法检测细胞凋亡;免疫组化检测Bax、Bcl-2蛋白水平表达;RT-PCR检测Bax、Bcl-2mRNA水平表达。结果生脉注射液在蛋白、mRNA水平可明显使Bcl-2表达增加,Bax表达减少;减少CK-MB释放,减少死亡率。与模型组比较均有显著性意义（P〈0.01或0.05）,其疗效成一定剂量依赖性。结论生脉注射液对大鼠阿霉素心肌损伤有明显疗效,其机制可能与抑制细胞凋亡等作用有关。     

生物技术药物的药代动力学研究近况

近年来,生物大分子药物发展迅猛,受到的关注也越来越多。与传统小分子药物相比,生物大分子药物具有相对分子质量大、不易透过生物膜、给药剂量低、易在体内降解等特点,这导致其具有与小分子药物不同的药代动力学特征。以蛋白多肽药物、单克隆抗体药物、抗体药物偶联物和核酸药物4类生物大分子药物为例,综述近年来生物大分子药物的药代动力学研究进展,旨在为生物大分子药物及生物类似药的研发提供参考。

     

The protective effect of glutathione administration on adriamycin-induced acute cardiac toxicity in rats.

Adriamycin (ADR) is a potent antitumor antibiotic drug known to cause severe cardiac toxicity. Although ADR generates free radicals, the role of these radicals in the development of cardiac toxicity is still not well understood. In the present study, we evaluated the effect of glutathione (GSH) supplementation or depletion on ADR-induced cardiotoxicity in male Wistar rats. Cardiac toxicity was induced by a single intraperitoneal injection of ADR (20 mg kg(-1)) and manifested by an increase in heart rate, blood pressure elevation, and increased serum creatine kinase (CK) and lactate dehydrogenase (LDH). The extent of lipoprotein oxidation, lipid peroxide measured as malondialdhye (MDA), total homocysteine (tHcy), lipid profile, and atherogenic index were markedly elevated, whereas cardiac GSH content was dramatically decreased in ADR rats. Pre- and co-treatment of ADR rats with GSH (5 mm kg(-1)) (ADR +GSH) markedly reduced the levels of CK, LDH, lipoprotein oxidation susceptibility, cardiac MDA, tHcy and atherogenic index, and elevated GSH levels in cardiac tissues. In contrast, GSH depletion through administration of l-buthionine-(S,R)-sulfoximine (BSO) (15 mm kg(-1)) before and after ADR injection (ADR +BSO) greatly exacerbated ADR cardiotoxicity compared to the control and ADR groups. Finally, there were also severe cardiac histopathological changes in ADR and ADR +BSO groups, which were nearly restored by GSH treatment. These results suggest that GSH inhibits ADR cardiotoxicity and might serve as a novel combination with ADR to limit free radical-mediated organ injury.     

Kinins are involved in the antiproteinuric effect of angiotensin-converting enzyme inhibition in experimental diabetic nephropathy

The present study examined non-insulin-treated streptozotocin (STZ)-induced diabetic rats to determine the role of kinins in diabetic nephropathy. Their involvement in the renoprotective effect of the angiotensin-converting enzyme inhibitor (ACEI) ramipril was investigated using the bradykinin (BK) B(2)-receptor antagonist, icatibant (HOE 140), or a combination of the two drugs.Although, none of the treatments prevented the decline of the glomerular filtration rate (GFR) in diabetic rats, ramipril (3 mg/kg/day), but not icatibant (HOE 140; 500 microg/kg/day), prevented proteinuria in these animals. However, the antiproteinuric effect of ramipril was reduced by 45% when combined with icatibant. To explore whether the renal kallikrein-kinin system (KKS) belongs to the underlying mechanisms of these findings, we also determined urinary BK levels, renal kallikrein (KLK) and angiotensin-converting enzyme (ACE) activity as well as renal cortical mRNA levels of neutral endopeptidase 24.11 (NEP) and low-molecular weight (LMW) kininogen. STZ led to a reduction of renal KLK and ACE activity and NEP expression and to a three-fold increase of urinary BK excretion and renal kininogen expression. Icatibant given alone had no effect on these parameters. In contrast, ramipril treatment normalized urinary protein and BK excretion as well as kininogen mRNA expression without affecting NEP mRNA expression or KLK and ACE activity.Our data demonstrate that renal BK is increased in severe STZ-induced diabetes mellitus, but may affect glomerular regulation only to a minor degree under this condition. However, kinins are partly involved in the antiproteinuric action of ACEI at this stage of diabetic nephropathy.     

大蒜多糖对阿霉素所致小鼠心脏毒性的拮抗作用

目的　研究大蒜多糖 (GP)对中毒性心肌炎的拮抗作用并探讨其机制。方法　建立小鼠阿霉素 (ADR)中毒性心肌炎模型,测定血清、心肌多项生化指标,并观察心肌结构变化。结果　ADR(3mg·kg-1ip, qod×7)可致小鼠血清肌酸激酶(CK)、乳酸脱氢酶(LDH)、谷草转氨酶(GOT)和诱导型一氧化氮合酶(iNOS)活力升高(P<0 01),同时心肌超氧化物歧化酶(SOD)活力下降而丙二醛 (MDA)含量升高 (P<0 01),线粒体水肿明显。GP( 0 75 ~3 0g·kg-1 ig, qd×15)能逆转ADR所致的上述改变,表现为剂量相关性降低血清CK、LDH、GOT和iNOS活力,增加心肌SOD活力和降低MDA含量,尤其以GP大剂量组作用明显 (P<0 05或P<0 01)。光镜和电镜结果也证实了GP的保护作用。结论　GP能拮抗阿霉素所致的小鼠中毒性心肌炎,其作用机制与增强心肌SOD活力和抗心肌脂质过氧化有关。     

TLC determination of iodochlorhydroxyquin and its conjugate in plasma.

A simple, specific, reliable, and sensitive method for the determination of iodochlorhydroxyquin and/or its conjugate in biological fluids is described. The method is based on a quantitative ether-acetone (1:1) extraction of plasma samples followed by TLC separation, visualization, elution, and determination at 267 nm. Iodochlorhydroxyquin released by hydrolysis of its conjugate was analyzed. Both compounds are detectable in amounts as low as 0.04 microgram/ml. Application of the one-compartment open model to the data (assuming the biotransformation of the drug in conjugated form) provides a pharmacokinetic profile for the 50-mg/kg dose of iodochlorhydroxyquin in Wistar male rats.     

Dose-response effect of sublingual captopril in hypertensive crises

Forty-one patients, presenting in the Emergency Service of the Hospital General y Clinico, Tenerife, with symptoms of hypertensive crisis and supine diastolic blood pressure (DBP) greater than 120 mm Hg, were studied. They received 12.5 mg of sublingual captopril and 30 minutes later, if diastolic blood pressure (DBP) was not 100 mm Hg or less, the same dose was repeated by the same route. Supine systolic blood pressure (SBP), DBP and heart rate (HR) were monitored at 0, 5, 10, 15, 30, 45, 60, and 120 minutes after each administration of captopril. In 27 patients (66%) had a satisfactory response (DBP less than or equal to 100 mm Hg), after a single dose, less than 30 minutes after administration, which persisted at 120 minutes. In 14, a second administration was necessary after 30 minutes, and a satisfactory response to the second dose, defined again as DBP reaching values of 100 mm Hg or less, was achieved in 12 of them (29% of the total group). In two patients (5% of total) no full response was obtained. The observed pattern of response suggests that a sublingual dose of 25 mg of captopril is the minimum effective dose, but it is also possible that administration of 12.5 mg of sublingual captopril at 10 to 15 minute intervals, perhaps up to a maximum dose of 37.5 mg, might be considered as an alternative treatment in hypertensive crises.     

白藜芦醇对小鼠阿霉素性心肌损伤的保护作用及机制

目的研究白藜芦醇(Res)对阿霉素(ADM)诱导的心肌损伤的保护作用及机制。方法一次性腹腔注射ADM(15mg·kg-1),建立小鼠阿霉素急性心肌损伤模型,并观察白藜芦醇预防性给药的保护作用。结果与正常对照组相比,ADM可使小鼠心电图QRS波电压幅度下降(P<0.01),心律失常率发生达60%;心肌超微结构损伤明显;血清中MDA、NO含量及LDH活性升高,SOD活性降低;p53蛋白表达升高(P<0.01)。与ADM损伤组相比,5、10、15mg·kg-1白藜芦醇呈剂量依赖性降低血清LDH活性和MDA、NO含量,增加SOD活性;减少QRS波电压下降幅度和心律失常发生率;下调p53蛋白表达(P<0.01或P<0.05);减轻电镜下心肌超微结构损伤。白藜芦醇对正常小鼠仅升高SOD活性,对其余指标无明显影响。结论Res对阿霉素性心脏损伤具有保护作用,其机制可能与其增强心肌SOD活力、抗脂质过氧化和抑制心肌细胞凋亡有关。     

卡托普利对对氧磷所致血管内皮损伤的保护作用及机制探讨

目的探讨卡托普利对对氧磷(paraoxon)所致血管内皮功能损伤的保护作用及其机制。方法用大鼠离体血管环和培养的人脐静脉内皮细胞(HUVEC)为实验模型,以血管内皮依赖性舒张(EDR)反应、HUVEC的通透性、内皮细胞活力、内皮细胞的形态学改变及生化参数为指标,用对氧磷作为损伤因子,用血管紧张素转化酶抑制剂(ACEI)卡托普利(captopril)作为保护药,观察了对氧磷对内皮的损伤作用及卡托普利的保护作用。用非巯基类ACEI(依那普利拉,enalaprilat)和抗氧化酶为对照,探讨了卡托普利对对氧磷所致血管内皮损伤的保护作用的机制。结果对氧磷(3.63μmol.L-1)与血管环或内皮细胞共孵30 min,显著性地抑制了血管EDR反应和增加了单层内皮细胞的通透性。卡托普利与血管环共孵30 min,剂量依赖性(0.1、1.0、10μmol.L-1)地显减轻了对氧磷(3.63μmol.L-1)对血管EDR的抑制作用、保护了培养的HUVEC中NO的释放。对氧磷和卡托普利对硝普钠(SNP)诱导的非内皮依赖性舒张反应没有影响。卡托普利(10μmol.L-1)显著性地阻滞了对氧磷所致的单层HU-VEC通透性的增加以及内皮细胞活力的降低、保护了超氧化物歧化酶(SOD)活性、阻滞了丙二醛(MDA)浓度的升高。超氧化物歧化酶、过氧化物酶(catalase)有与卡托普利相似的抗对氧磷损伤作用,但依那普利拉对对氧磷所致损伤无明显保护作用,巯基抑制剂(4-羟基汞苯甲酸普罗比妥钠,cystain)能显著性拮抗卡托普利的保护作用。结论对氧磷能直接损伤血管内皮细胞,卡托普利对氧磷所致的血管内皮细胞损伤有显著性保护作用,其机制可能主要依赖于其所含的巯基的抗氧化作用,而非依赖于抑制血管紧张素转化酶。     

影响血管紧张素转化酶抑制剂抗尿蛋白作用的因素

目的；分析、探讨影响血管紧张素转化酶抑制剂（ACEI）抗尿蛋白作用的因素。方法：原发性肾小球疾病患者136例，分别实施卡托普利，西拉普利和依那普利各两个剂量组治疗4个月。治疗前及治疗后每月检测平均动脉压（MAP），尿蛋白、尿白蛋白排泄率（UAER），尿钠和肾功能，并检测患者ACE基因表型，依据尿蛋白减少率（UPDR）分为：显效组（UPDR＞50％）33例，良效组（UPDP：30％－50％）49例，有效组（UPDP；10％－30％）23例，无效组（UPDR：－10－10％）21例，恶化组（UPDP＜－10％）10例，分析各组间ACEI种类，剂量以及上述各检测指标的变化。结果：1．ACEI的抗尿蛋白作用，与ACEI种类，剂量和患者ACE基因多态性无关；2．与其他组相比，显效组患者治疗前肾功能良好，MAP较低；3．无效组和恶化组治疗过程中食盐摄入控制不良。结论：ACEI种类，剂量和患者ACE基因多态性不影响ACEI的抗尿蛋白作用，肾脏疾病早期应用以及严格限制食盐摄入可提高ACEI抗尿蛋白疗效。     

Antithrombotic effect of captopril and enalapril in young rats

Angiotensin converting enzyme inhibitors (ACE-Is) are the main drugs used in the treatment of essential hypertension and congestive heart failure in adults. Recently, we have demonstrated the antithrombotic effect of captopril (CAP) and enalapril (ENA) in venous thrombosis model in adult rats. One might also suggest the beneficial effect of those drugs on hemostasis in young individuals. Two months old male Wistar rats were used in the study. Acute administration of CAP at a dose of 50 and 100 mg kg(-1) significantly reduced the venous thrombus weight. Dose-dependent reduction in the thrombus weight was also observed in ENA (3, 10, 30 mg kg(-1))-treated rats. Strong reduction in the thrombus weight was also seen after chronic administration of CAP (2 x 25 mg kg(-1)) and ENA (1 x 15 mg kg(-1)). Both drugs given chronically reduced the frequency of thrombi. Systolic blood pressure was reduced to similar extent after acute and chronic application of the drugs. CAP shortened euglobulin clot lysis time (ECLT) when given acutely (100 mg kg(-1)) and chronically (2 x 25 mg kg(-1)). ENA decreased ECLT only when given at multiple doses (1 x 15 mg kg(-1)). None of the drugs changed prothrombin time or activated partial tromboplastin time. We conclude that CAP and ENA possess antithrombotic effect in young individuals. Activation of the fibrinolytic pathway seems to play an important role in the mechanism of their antithrombotic action.     

Quantitative determination of captopril in blood and captopril and its disulfide metabolites in plasma by gas chromatography

A sensitive, quantitative gas chromatographic-electron capture (GC-EC) method for the determination of captopril in blood and captopril and its disulfide metabolites (collectively) in plasma was developed. After addition of an internal standard and N-ethylmaleimide to the biological samples, excess N-ethylmaleimide and naturally occurring interfering substances were removed by extraction with benzene followed by acidification and extraction with hexane. The N-ethylmaleimide adducts of captopril and of the internal standard were then extracted with benzene and converted to their hexafluoroisopropyl esters. For the assay of captopril and its disulfide metabolites, tributylphosphine was used to reduce the disulfide metabolites to captopril prior to derivatization. The hexafluoroisopropyl esters of the N-ethylmaleimide adducts of captopril and of the internal standard, the 4-ethoxyproline analogue of captopril, were separated by GC on a column packed with 3% OV-101 on Chromosorb W-HP. The lower limits of sensitivity were 20 ng/mL for captopril in blood and 50 ng/mL for captopril and its disulfide metabolites in plasma. Linearity, precision, and accuracy were excellent. The method was validated by comparison of results obtained for total captopril in dog plasma by the GC-EC assay with results obtained by a published GC-MS method. The assay was applied to dog and human samples to explore its general utility.     

Antithrombotic effect of captopril and enalapril in old rats

In the present study, we have shown considerably accelerated thrombosis in old rats in comparison with adult rats, which may be related to the impaired hemostatic balance in these animals. In old rats, captopril and enalapril caused a marked reduction of venous thrombus weight. The mechanism of antithrombotic action of these drugs seems to be dependent on the suppression of coagulation cascade and the enhancement of the fibrinolytic processes.