Flunixin对猪肝微粒体细胞色素P450酶系的影响

采用cocktail探针药物法研究了氟尼辛葡甲铵对猪肝微粒体细胞色素（CY）P450酶系的作用.将12头猪随机分为2组,试验组每日肌肉注射氟尼辛葡甲铵1次,对照组给予等体积的生理盐水,连续给药10 d.通过高效液相色谱法检测探针药物的代谢率,评价各组CYP450酶的活性水平.结果显示,试验组的氨苯砜代谢减慢,消除半衰期t1/2延长;而氯唑沙宗代谢加快,t1/2缩短.说明氟尼辛葡甲铵对猪的CYP3A4具有抑制作用,而对CYP2E1存在诱导效应.     

几种临床常用药物对猪肝细胞色素P450酶系的影响

研究了几种临床常用药物对猪肝微粒体细胞色素P450酶系的影响。红霉素、环丙沙星、地塞米松和乙醇,按5倍治疗剂量进行肌肉注射每天1次,连续给药10 d。差速离心法制备猪肝微粒体,B radford法测定蛋白浓度,分光光度法检测6种肝细胞色素P450酶含量及活性。结果表明,4种药物除环丙沙星外,均对P450酶的总活性产生显著影响(P<0.05),同时地塞米松组的微粒体蛋白含量和红霉素-N-脱甲基酶活性,乙醇组的苯胺-4-羟化酶活性和红霉素组的红霉素-N-脱甲基酶活性,与对照组相比均差异显著(P<0.05)。说明环丙沙星和红霉素对猪肝细胞色素P450酶系有选择性抑制作用,而地塞米松和乙醇对其起选择性诱导作用。     

Effects of dapagliflozin in combination with insulin on cytochrome P450 activities in a diabetes type 1 rat model

Previous studies reported that diabetes alters the activities of hepatic cytochrome P450 (CYP) enzymes, which, in turn, affects the disposition of some drugs. We herein examined and compared the effects of the combination of dapagliflozin with a low insulin dose, a full dose of insulin alone, and dapagliflozin alone for 3 and 8 weeks on CYP activities in a diabetes type 1 rat model. We induced type 1 diabetes in rats using a single intraperitoneal injection of 60 mg/kg streptozotocin (STZ). Daily treatment with the full dose of insulin alone, dapagliflozin alone, or dapagliflozin in combination with a low dose of insulin was then initiated. STZ-induced rats developed marked hyperglycemia and altered CYP2E activities. Dapagliflozin in combination with a low dose of insulin stabilized hyperglycemia and CYP1A, 2D, 2E and 3A activities. However, dapagliflozin alone did not improve blood glucose levels or CYP activities. These results suggest that the effects of dapagliflozin in combination with a low dose of insulin are similar to those of a full dose of insulin, and stabilize CYP activities in type 1 diabetes.     

乐果亚急性暴露对大鼠胆碱酯酶浓度及细胞色素P450亚型表达量的影响

对6周龄雄性Wistar大鼠5次灌胃给予20％LD50的乐果，观察给药后大鼠的毒性反应，每日测定实验鼠采食量、饮水量和体重。最后一次给药后第24h，采用丁酰硫代胆碱法测定大鼠红细胞、血清和脑组织中的胆碱酯酶浓度，并采用定量RT-PCR法检测大鼠肝中细胞色素P450基因8个亚型的表达量。结果表明，大鼠用药后不同时间内出现了有机磷药物中毒的典型症状，其采食量、饮水量和体重明显下降，红细胞、血清和脑组织中胆碱酯酶浓度均明显低于对照组，肝细胞色素P450的8个亚型中CYP2B1／2的表达量明显高于对照组。     

Characteristics of cytochrome P450-dependent metabolism in the liver of the wild raccoon,Procyon lotor

Wildlife is exposed to a wide range of xenobiotics in the natural environment. In order to appropriately assess xenobiotic-induced toxicity in wildlife, it is necessary to understand metabolic capacities. Carnivores, in general, have low metabolic abilities, making them vulnerable to a variety of chemicals. Raccoons (Procyon lotor) in the wild have been found to have high levels of xenobiotics. However, little is known about the metabolic capacity of the cytochrome P450 (CYP) enzymes in this species. Thus, this study used liver samples to investigate the characteristics of CYP enzymes in wild raccoons. In 22 wild raccoons, CYP concentrations in hepatic microsomes were examined. To better understand the properties of CYP-dependent metabolism, in vitro metabolic activity studies were performed using ethoxyresorufin, pentoxyresorufin and testosterone as substrates. In addition, three raccoons were fed commercial dog food in the laboratory for one week, and the effects on CYP-dependent metabolism were investigated. In comparison to other mammalian species, raccoons had very low concentrations of CYP in their livers. In an in vitro enzymatic analysis, raccoons’ ethoxyresorufin O-deethylase (EROD) and pentoxyresorufin O-depentylase (PROD) metabolic capacities were less than one-fifth and one-tenth of rats’, respectively. These results indicate the possible high risk in raccoons if exposed to high levels of environmental xenobiotics because of their poor CYP activity. In this study, the features of CYP-dependent metabolism in wild raccoons are described for the first time.     

Effects of bovine cytochrome P450 single-nucleotide polymorphism, forage type and body condition on production traits in cattle

Relating single-nucleotide polymorphisms (SNP) to cows with acceptable productivity could benefit cattle breeders in areas where tall fescue is the predominant forage. This study aimed to (i) identify SNPs in bovine cytochrome P450 3A28 (CYP3A28) and (ii) determine the associations between SNP genotype, forage and cow body condition (BC). Genotype (CC, CG or GG) and forage [Kentucky-31 wild-type endophyte-infected tall fescue (KY+) vs. bermudagrass] effects on milk volume and quality were determined in Herd 1 cows (123 cows); in Herd 2 (99 cows), genotype and BC (low vs. moderate) effects on ovarian follicle size, calving date and calving per cent were determined; and in Herd 3 (114 cows), effects of genotype and fescue cultivar [KY+ vs. non-toxic endophyte-infected tall fescue (HiMag4)] were related to calving per cent, calving date and weaning weights of both cow and her calf. A cytosine (C) to guanine (G) transversion at base 994 (C994G) in CYP3A28 was identified. There was a genotype × forage type interaction (p < 0.05) on milk protein in Herd 1 cows; CC cows grazing bermudagrass had greater milk protein percentage in relation to other cows in the herd. In Herd 2, BC and genotype × BC tended (p < 0.10) to influence follicle size and Julian calving date respectively. Diameter of the largest follicle tended to be larger in moderate BC than in low-BC cows; whereas, CC and CG cows in moderate BC and homozygous (CC and GG) cows in low BC tended to calve 14 days earlier in relation to CG cows in low BC. In Herd 3, there was a genotype × forage type interaction (p < 0.05) on calving per cent, Julian calving date and calf weaning weight. In this study, genetic alterations (G allele at C994G) coupled with nutritional factors (low BC and toxic tall fescue) resulted in overall lower productivity in cows.     

Research Progress on Cytochrome P450 2E1

Cytochrome P450 2E1 (CYP2E1) is one of the body's metabolite of drugs and low molecular weight compounds mainly enzymes, and the increasing of activity or expression of CYP2E1 is closely related with the clinical variety of diseases. This paper reviews the research progress of protein structure, function, positioning mechanism and other aspects of genetic polymorphisms of CYP2E1 at home and abroad. Thereby the paper could lay the foundation for further exploration of the biological effects of CYP2E1 and its relationship with clinical disease.     

Effects of forage type,body condition and single‐nucleotide polymorphisms in the bovine cytochrome P450 regulatory region on cow productivity*

Single‐nucleotide polymorphisms (SNP) in the coding sequence of cytochrome p450 (CYP3A28) have been associated with milk yield and composition, and calving traits in cows. In this study, we aimed to determine whether (i) the CYP3A28 regulatory region was polymorphic and (ii) SNP genotype, forage type, body condition and their interactions affect cow productivity. Primers for CYP3A28 promoter were designed to amplify a 483‐bp segment by PCR. Amplicon sequences revealed seven SNP (T‐318C, T‐113A, C‐189T, T‐78G, A6G, G17A and T21C) in Brahman (38 cows), Brahman x Angus reciprocal crosses (47 cows) and crossbreds (98 cows). Angus cows (n = 41) appeared to be fixed at those SNP locations. Genotype and forage {endophyte‐infected tall fescue [KY+; Lolium arundinaceum (Schreb.) S. J. Darbyshire] vs. bermudagrass [Cynodon dactylon (L.) Pers.]} effects on lifetime (8‐years) calving rate, and calf weaning weights and heights were determined in Herd 1 (126 cows); genotype and BC (low vs. moderate) effects on calving date and calving percent were determined in Herd 2 (98 cows). Four SNP (T‐318C, T‐113A, A06G and T21C) appeared to be related to cattle productivity, CC cows at T‐318C having a lower (p < 0.05) lifetime calving rate than TC or TT cows (65%, 85% and 81% respectively). Cows that grazed KY+ and were TT at T‐318C produced calves that tended (p < 0.07) to weigh less than their contemporaries. Moreover, calves of TT cows were shorter (p < 0.05) at weaning than calves of CC or TC cows. In Herd 2, moderate‐BC cows that were TT or AA at T‐318C, T‐113A, T‐78G, A6G and T21C had greater (p < 0.05) calving rates (74–80%) than heterozygous cows (46–60%), and low‐BC cows that were AA at G17A calved at least 6 days earlier (p < 0.05) than heterozygous cows. Our findings suggest that SNP in the CYP3A28 regulatory region of Brahman‐influenced cows are associated with cattle productivity.     

实验性雏鸡铜中毒的病理学研究

选用1日龄艾维菌肉鸡180只随机分为3组，分别以对照日粮（Cu 11．97mg／kg）、铜中毒Ⅰ组日粮（Cu 650mg／kg）和铜中毒Ⅱ组日粮（Cu 850mg／kg）饲喂6周，以实验病理学方法系统研究了铜中毒对雏鸡组织器官和某些血液指标的影响。2个铜中毒组在试验的第2、3周先后出现临床症状；铜中毒Ⅱ组雏鸡的发病率和死亡率高于铜中毒Ⅰ组。病理形态学观察，2个铜中毒组的病变基本一致，表现为肌胃角质层增厚、龟裂，肠绒毛裸露断裂，伴有坏死；肝细胞脂肪变性；肾小管上皮细胞变性，坏死脱落；淋巴免疫器官体积缩小、重量减轻、淋巴细胞数量减少。2个铜中毒组雏鸡血清谷草转氨酶（AST）活性显著升高，红细胞数量、血红蛋白含量显著降低（P〈0．05或P〈0．01）。表明，雏鸡在生长发育的2～6周对高剂量铜较为敏感，毒性损坏的靶器官是肝、肾、胃肠道和淋巴免疫器官，铜中毒导致雏鸡组织器官受损、功能障碍及免疫功能低下，最终导致发病、死亡。     

Novobiocin Inhibits Both UDP-Glucuronosyltransferase and Cytochrome P450-Mediated Enzyme Activities in Pig Liver Microsomes

The effects of novobiocin (range 0.0125–2 mmol/L) on the hydroxylation of testosterone, the N-demethylation of erythromycin, and the glucuronidation of -naphthol and paracetamol were studied using pig hepatic microsomes, pooled from five animals. The final concentrations of these substrates in the incubation mixtures were selected to meet V_max conditions. Novobiocin caused a concentration-dependent inhibition of the glucuronidation of paracetamol; the formation of -naphthol-glucuronide was reduced to a lesser degree. These results confirm and extend earlier findings in laboratory animal species that novobiocin inhibits UDP-glucuronosyltransferases (UDPGTs). Moreover, novobiocin strongly inhibited 6-hydroxylation of testosterone. The microsomal N-demethylation of erythromycin and hydroxylation of testosterone at the 15 position were less affected by novobiocin. These results suggest that novobiocin inhibits not only UDPGTs, but also cytochrome P450 (CYP) enzyme activities, probably those belonging to the CYP3A subfamily. More research is needed to reveal which CYPs and UDPGTs are affected by novobiocin in vivo, in order to improve the understanding and probably the predictability of potential drug interactions with this antibiotic.     

家蚕氟中毒后中肠NADPH-细胞色素P450还原酶和NADPH-细胞色素C还原酶活性的变化

为了探究家蚕对NaF的代谢机制,以家蚕耐氟品种T6和氟化物敏感品种734为材料,从5龄起蚕开始分别添食50、100、200、400 mg/kg NaF溶液处理后的桑叶,检测蚕体中肠微粒体酶液中的黄素蛋白NADPH-细胞色素P450还原酶(CPR)和NADPH-细胞色素C还原酶(CR)的活性变化。氟物化敏感品种734的4个NaF处理组第3天的中肠CPR活性低于对照组,其余时间几乎都高于对照组,400 mg/kg NaF处理组在第4天的CPR活性最高,且各NaF处理组的CPR活性差异显著(P<0.05);耐氟品种T6 NaF处理组和对照组的中肠CPR活性整体上呈先升后降的趋势,几乎都在第2天达到最高值,各组之间的差异不显著(P>0.05)。氟化物敏感品种734的4个NaF处理组的中肠CR活性呈现先升后降的趋势,而对照组呈下降趋势;耐氟品种T6的50、100、400 mg/kg NaF处理组在第1～2天的中肠CR活性呈明显下降趋势,之后的变化相对较小,对照组的CR活性仅在第3～4天略高于NaF处理组,而其余时间NaF处理组的CR活性较高。2个家蚕品种添食不同浓度NaF后的中肠CR活性差异均不显著(P>0.05)。试验结果显示,耐氟品种T6在NaF作用下中肠的CPR和CR活性变化范围(分别为对照组的0.4～2.0倍和0.3～2.9倍)远小于氟化物敏感品种734这2种酶的活性变化范围(分别为对照组的0.6～9.3倍和0.4～4.6倍)。初步推测CPR和CR与家蚕对氟化物代谢具有一定的关联。     

Development of genotyping method for functionally relevant variants of cytochromes P450 in cynomolgus macaques

In cynomolgus macaques (Macaca fascicularis), widely used in drug metabolism studies, CYP2C9, CYP2C76, CYP2D6, CYP3A4, and CYP3A5, important drug‐metabolizing enzymes, are abundantly expressed in liver and metabolize cytochrome P450 substrates. CYP2C9 (c.334A>C), CYP2C76 (c.449TG>A), CYP2D6 (c.891A>G), CYP3A4 (IVS3 + 1G>del), and CYP3A5 (c.625A>T) substantially influence metabolic activity of enzymes, and thus are important variants in drug metabolism studies. In this study, a real‐time PCR method was developed for genotyping these variants. The validity of the methods was verified by genotyping two wild type, two heterozygous, and two homozygous DNAs and was used to genotype 41 cynomolgus macaques (from Cambodia, Indonesia, the Philippines, or Vietnam) for the five variants, along with another important variant CYP2C19 (c.308C>T). The CYP2C9 and CYP2C19 variants were found only in Cambodian and Vietnamese animals, while the CYP2C76 and CYP2D6 variants were found only in Indonesian and Philippine animals. The CYP3A4 and CYP3A5 variants were not found in any of the animals analyzed. Mauritian animals, genotyped using next‐generation sequencing data for comparison, possessed the CYP2C19 and CYP2D6 variants, but not the other variants. These results indicated differences in prevalence of these important variants among animal groups. Therefore, the genotyping tool developed is useful for drug metabolism studies using cynomolgus macaques.     

铜中毒对雏鸡淋巴细胞凋亡的影响

选用1日龄艾维菌肉鸡健雏180只，随机分为3组，分别喂以对照日（Cu 11．97mg／kg）、中毒Ⅰ日（Cu 650mg／kg）和中毒Ⅱ日粮（Cu 850mg／kg），为期6周，以流式细胞术的5r法观察研究了铜中毒对雏鸡免疫器官淋巴细胞凋亡的影响。研究结果表明，铜中毒可诱导淋巴细胞凋亡，2个中毒组免疫器官淋巴细胞凋亡百分率显著高于对照组（P〈0．05或P〈0．01）。     

大豆黄酮对伊莎鸡卵泡发育及其P450arom基因表达的影响

实验探讨了大豆黄酮(DAI)对伊莎鸡卵泡发育及其芳香化酶(P450arom)mRNA表达的影响。实验选取16只产蛋后期伊莎鸡,等分为对照组和DAI处理组。对照组饲喂基础日粮,实验组在基础日粮中添加10 mg/kgDAI。实验持续7周后,分离排卵前卵泡(F1、F2、F3……)的颗粒层及小黄卵泡和大白卵泡,通过RT-PCR法检测P450arom mRNA表达的相对丰度。结果表明:DAI明显提高了伊莎鸡小黄卵泡和大白卵泡的数量,P450arommRNA在伊莎鸡不同发育阶段卵泡中的表达存在差异,部分卵泡P450arom mRNA表达的相对丰度显著增加。因此,在产蛋后期伊莎鸡基础日粮中添加DAI可增加不同发育阶段卵泡的数目,上调部分卵泡中与发育相关的基因表达以促进卵泡发育。     

Analysis of polymorphisms of canine Cytochrome P 450-CYP2D15

Cytochrome P450 (CYP) proteins constitute a large ancient family of oxidative enzymes essential for the efficient elimination of a wide variety of clinically used drugs. Polymorphic variants of human CYP2D6 are associated with the conversion rate and efficacy of several drugs such as antidepressants. Polymorphisms of the canine orthologue CYP2D15 are of interest because these antidepressants are also used in dogs with behavioral problems and the outcome of the treatment is variable. However, the annotated CYP2D15 gene is incomplete and inaccurately assembled in CanFam3.1, hampering DNA sequence analysis of the gene in individual dogs. We elucidated the complete exon–intron structure of CYP2D15 to enable comprehensive genotyping of the gene using genomic DNA. We surveyed variations of the gene in four diverse dog breeds and identified novel polymorphisms in exon 2 in border collies. Further investigation to establish the impact of these canine CYP2D15 polymorphisms on interindividual variability in expression and function of this metabolizing enzyme is now feasible. Further knowledge of CYP pharmacogenetics will help individualize therapy and thereby increase therapeutic efficacy, especially in the use of antidepressants in veterinary behavioral medicine.