The role of polymorphic cytochrome P450 gene(CYP2B6)in B-chronic lymphocytic leukemia(B-CLL)incidence and outcome among Egyptian patients

Cytochromes P450(CYPs)play a prominent role in catalyzing phase I xenobiotic biotransformation and account for about 75%of the total metabolism of commercially available drugs,including chemotherapeutics.The gene expression and enzyme activity of CYPs are variable between individuals,which subsequently leads to different patterns of susceptibility to carcinogenesis by genotoxic xenobiotics,as well as differences in the efficacy and toxicity of clinically used drugs.This research aimed to examine the presence of the CYP2B6*9 polymorphism and its possible association with the incidence of B-CLL in Egyptian patients,as well as the clinical outcome after receiving cyclophosphamide chemotherapy.DNA was isolated from whole blood samples of 100 de novo B-CLL cases and also from 100 sex-and age-matched healthy individuals.The presence of the CYP2B6*9(G516T)polymorphism was examined by PCR-based allele specific amplification(ASA).Patients were further indicated for receiving chemotherapy,and then they were followed up.The CYP2B6*9 variant indicated a statistically significant higher risk of B-CLL under different genetic models,comprising allelic(T-allele vs.G-allele,OR=4.8,p<0.001)and dominant(GT+TT vs.GG,OR=5.4,p<0.001)models.Following cyclophosphamide chemotherapy,we found that the patients with variant genotypes(GT+TT)were less likely to achieve remission compared to those with the wild-type genotype(GG),with a response percentage of(37.5%vs.83%,respectively).In conclusion,our findings showed that the CYP2B6*9(G516T)polymorphism is associated with B-CLL susceptibility among Egyptian patients.This variant greatly affected the clinical outcome and can serve as a good therapeutic marker in predicting response to cyclophosphamide treatment.

Cardioprotection of Shenfu preparata on cardiac myocytes through cytochrome P450 2J3

To evaluate whether Shenfu injection （SFI） protects against cardiac myocyte injury induced by Fupian injection （FPI） in vitro. METHODS： H9c2 cells were separately treated with FPI, Renshen injection （RSI） and SFI. Cell viability, lactate dehydrogenase （LDH） release, spontaneous beating rate of primative cardical cells, caspase-3/7 activity, cell apoptosis, and cytochrome P450 2J3 （CYP2J3） mRNA expression were analyzed. RESULTS： The viability of H9c2 cells treated with SFI （37 and 75 mg/mL） was significantly higher than that of H9c2 cells treated with FPI （25 and 50 mg/mL） （P〈0.05, P〈0.01, respectively）. LDH activity of H9c2 cells treated with SFI （75 mg/mL） was significantly decreased （P〈0.01） compared with that of H9c2 cells treated with FPI （50 mg/mL）. SFI （150 mg/mL） significantly attenuated FPI （100 mg/mL）-induced spontaneous beating rate decrease in primary myocardial cells after 4-hour treatment. Compared with FPI （12 and 25 mg/mL）, SFI （18 and 37 mg/mL） treatment could effectively reverse the change of caspase-3/7 activity （P〈0.01 and P〈0.01, respectively）. Compared with FPI （6 and 25 mg/mL）, apoptotic cells decreased significantly （P〈0.05, P〈0.01, respectively） when H9c2 cells were incubated with SFI （9 and 37 mg/mL）. The expression of CYP2J3 mRNA was down-regulated by FPI, while RSI and SFI could up-regulate the expression of CYP2J3 （P〈0.01）, which suggested the potential mechanism of protection of RSI against cardiac myocyte damage induced by FPI treatment. CONCLUSION： These observations indicate that SFI has the potential to exert cardioprotective effects against FPI toxicity. The effect was possibly correlated with the activation of CYP2J3.

Inhibitive Effect of Cremophor RH40 or Tween 80-based Self-microemulsiflying Drug Delivery System on Cytochrome P450 3A Enzymes in Murine Hepatocytes

This study examined the effect of self-microemulsiflying drug delivery system (SMEDDS) containing Cremophor RH40 or Tween 80 at various dilutions on cytochrome P450 3A (CYP3A) enzymes in rat hepatocytes, with midazolam serving as a CYP3A substrate.The particle size and zeta potential of microemulsions were evaluated upon dilution with aqueous medium.In vitro release was detected by a dialysis method in reverse.The effects of SMEDDS at different dilutions and surfactants at different concentrations on the metabolism of MDZ were investigated in murine hepatocytes.The cytotoxicity of SMEDDS at different dilutions was measured by LDH release and MTT technique.The effects of SMEDDS on the CYP3A enzymes activity were determined by Western blotting.Our results showed that dilution had less effect on the particle size and zeta potential in the range from 1:25 to 1:500.The MDZ was completely released in 10 h.A significant decrease in the formation of 1’-OH-MDZ in rat hepatocytes was observed after treatment with both SMEDDS at dilutions ranging from 1:50 to 1:250 and Cremophor RH 40 or Tween 80 at concentrations ranging from 0.1% to 1% (w/v), with no cytotoxicity observed.A significant decrease in CYP3A protein expression was observed in cells by Western blotting in the presence of either Cremophor RH40 or Tween 80-based SMEDDS at the dilutions ranging from 1:50 to 1:250.This study suggested that the excipient inhibitor-based formulation is a potential protective platform for decreasing metabolism of sensitive drugs that are CYP3A substrates.

In vitro and in vivo cytochrome P450 3A enzyme inhibition by Aframomum melengueta and Dennettia tripetala extracts

Objective: To evaluate the in vitro and in vivo inhibitory effects of two commonly used herbs, Aframomum melengueta(A. melengueta) and Dennettia tripetala(D. tripetala) on CYP 3A enzymes. Methods: In vitro inhibition of the enzymes were assessed with microsomes extracted from female albino rats using erythromycin-N-demethylation assay(EMND) method while their in vivo effects were measured by estimating simvastatin plasma concentrations in rats. Pharmacokinetic parameters were determined using non-compartmental anaysis as implemented in Win Nonlin pharmacokinetic program. Results: EMND assay with intestinal microsomes indicated that aqueous extracts of D. tripetala and A. melengueta significantly(P < 0.05) inhibited intestinal CYP 3A activity at both 50 μg and 100 μg concentrations. Petroleum ether extract of D. tripetala and ethanol extracts of A. melengueta inhibited intestinal CYP3 A activity at 100 μg but not at 50 μg concentrations. All the extracts showed an in vitrodose dependent CYP 3A inhibition with liver microsomes. In vivo analysis showed that pretreatment with the extracts enhanced systemic absorption of simvastatin with reductions in metabolizing enzymes activity as indicated in significant increases in maximal concentration, area under curve, area under moment curve and mean resident time of simvastatin(P < 0.05). Conclusions: Herbal preparations containing these plants' extracts should be used with caution especially in patients on CYP450 3A substrate medications.

酒精性肝损伤大鼠细胞色素P450 CYP2E1和细胞色素P450 CYP3A的代谢活性

目的观察酒精性肝损伤对大鼠细胞色素P450CYP3A(CYP3A)和细胞色素P450CYP2E1(CYP2E1)代谢活性的影响。方法采用ig给予白酒制备大鼠酒精性肝损伤模型,检测血清中谷丙转氨酶(GPT)和谷草转氨酶(GOT)活性,采用HE染色法光镜下观测酒精对肝脏损伤程度。大鼠ip给予CYP3A探针药物咪达唑仑10mg·kg-1或ig给予CYP2E1探针药物氯唑沙宗50mg·kg-1后,采用高效液相色谱法测定不同时间点大鼠血浆中咪达唑仑和氯唑沙宗的血药浓度,并应用3P87软件计算其药代动力学参数,以考察CYP2E1和CYP3A的代谢活性的变化。大鼠ig给予氯唑沙宗80mg·kg-1后,热板方法测定大鼠添足次数和添足反射潜伏期。结果酒精性肝损伤可致大鼠肝小叶结构不清,肝索排列紊乱,肝细胞体积增大,呈弥漫性中度水变性,肝窦受压,大部分肝细胞胞浆内见大小不等的脂肪空泡;与正常对照组相比,酒精性肝损伤组大鼠GPT和GOT活性分别增加了16.0%和20.0%(P<0.05,P<0.01)。酒精性肝损伤致大鼠CYP2E1对探针药物氯唑沙宗的代谢活性增强,AUC,t1/2和cmax分别降低了38.0%,30.5%和35.0%(P<0.05);酒精肝损伤组大鼠氯唑沙宗镇痛效果明显降低;酒精性肝损伤致大鼠CYP3A对探针药物咪达唑仑的代谢活性增强,AUC,t1/2和cmax分别降低了122.6%,54.9%和56.9%(P<0.01,P<0.05)。结论酒精性肝损伤可使大鼠CYP2E1和CYP3A代谢活性增强。

Identification of Cytochrome P450 (CYP) Genes in Zhikong Scallop (Chlamys farreri)

Cytochrome P450 （CYP） superfamily is one of the membership largest and function most diverse protein superfamily recogniozed among living beings. Members of this superfamily were further assigned to different families and subfamilies based on their amino acid similarities. According to their phylogenetic relationships, the CYP genes which likely diverged from common ancestor gene and may share common functions were grouped into one clan. Widely distributing scallops are a group of the most conspicuous bivalve; however the studies on their CYP is acarce. In this study, we searched the genome and expressed sequence tags of Zhikong scallop （Chlamysfarreri） for CYP genes. In total, 88 non-redundant CIfP were identified, which were homed in 13 CYPs gene families. Phylogenetic analysis divided these genes into 4 CYP clans. As in deuterostomes, Clan 2 was the largest, which contained 33 genes belonging to CYP1, CYP2, CYP17 and CYP356 families. Clan 3 contgained 19 genes belonging to CYP3, CYP5 and CYP30 families. Clan 4 contained 23 genes, all belonging to CYP4 family. The mitochondrial CYP clan contained 9 genes belonging to CYPIO and CYP24 families. In comparison, protostomes （C. farreri, D. pluex, D. melanogaster） contained more CYP genes than deuterostomes （S. purpuratus and vertebrates） in Clan 2 but less genes in Clan 3 and Clan 4. Our findings will aid to deciphering CYP function and evolution in scallops and bivalves.

Substrate-dependent inhibition of human cytochrome P450 3A catalytic activity by specific isomers of vitamin E

OBJECTIVE Lithocholic acid,which is a secondary bile acid,has been reported to be hepatotoxic and carcinogenic.It is metabolized by human cytochrome P450 3A(CYP3A)to form 3-ketocholanoic acid.A previous study suggests that vitamin E isomers(tocotrienols and tocopherols)are metabolized by CYP3 A.Given that substrates of an enzyme may competitively inhibit the enzyme,we determined whether alpha-tocotrienol,gamma-tocotrienol,delta-tocotrienol,tocotrienol-rich mixture(a mixture consisting of 25.7% d-α-tocotrienol,2.6% d-β-tocotrienol,28.6% d-γ-tocotrienol,8.4% d-δ-tocotrienol,25.6% d-α-tocopherol,and 4.3% d-α-tocomonoenol),and alpha-tocopherol inhibit human liver microsomal CYP3Aactivity,as assessed by the enzymatic conversion of lithocholic acid to 3-ketocholanoic acid and of testosterone to6β-hydroxytestosterone.METHODS Enzymatic formation of 3-ketocholanoic acid via lithocholic acid 3-oxidation was determined in pooled human liver microsomes and recombinant CYP3A4 and CYP3A5.Enzyme inhibition assay was conducted in a mixture containing potassium phosphate buffer(pH 7.4),human liver microsomes,NADPH,lithocholic acid,and various concentrations of a test chemical.The amount of 3-ketocholanoic acid formed was quantified by a novel,validated ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS-MS)method.RESULTS Lithocholic acid was metabolized to 3-ketocholanoic acid by human recombinant CYP3A4 and CYP3A5enzymes and human liver microsomes.Alpha-tocotrienol,gamma-tocotrienol,delta-tocotrienol,and tocotriernol-rich mixture,but not alpha-tocopherol,inhibited 3-ketocholanoic acid formation in human liver microsomes.Concentration-response experiments indicated that tocotrienol-rich mixture and delta-tocotrienol inhibited 3-ketocholanoic acid formation with IC50 values of 6.6±2.1μg·mL-1 and 19.0±1.0μmol·L-1,respectively.CONCLUSION Tocotrienols inhibited CYP3A-catalyzed lithocholic acid 3-oxidation but not CYP3A-catalyzed testosterone 6-beta-hydroxylation.This suggests that lithocholic acid and testosterone bind to different CYP3 Abinding sites and that tocotrienols preferentially inhibit the lithocholic acid binding site on CYP3 Aenzymes.

注射用丹参总酚酸(冻干)对人CYP450酶和P-糖蛋白体外抑制作用及对大鼠CYP1A2和CYP3A体内诱导作用(英文)

目的探讨注射用丹参总酚酸(冻干)(SLI)对人CYP450酶和P-糖蛋白体外抑制作用以及对大鼠CYP1A2和CYP3A体内诱导作用。方法①应用P450-GloTMCYP450检测试剂盒,通过化学发光法测定SLI和经典抑制剂对细胞色素P4501A2(CYP1A2),CYP2D6,CYP3A4,CYP2C19和CYP2C9的IC50值,通过比较SLI和经典抑制剂对相应细胞色素P450亚型的IC50值来判断SLI对人CYP450酶的体外抑制作用。②Wistar大鼠分别iv给予SLI 3,10和30 mg·kg-1和诱导剂苯巴比妥钠20 mg·kg-1,采用探针底物法,通过比较代谢产物的生成速率来评价SLI对大鼠CYP1A2和CYP3A的诱导作用。③应用ATP酶检测试剂盒,通过化学发光法测定ATP酶活性来评价SLI是否为P-gp的底物或抑制剂。结果①CYP1A2,CYP2C9,CYP2C19,CYP2D6和CYP3A4抑制剂的IC50与SLI对其的IC50进行比较(CYP1A2:0.12μmol·L-1vs 840μmol·L-1;CYP2C9:3.362μmol·L-1vs 704μmol·L-1;CYP2C19:3.236μmol·L-1vs 306μmol·L-1;CYP2D6:0.117μmol·L-1vs 2660μmol·L-1;CYP3A4:0.078μmol·L-1vs 1780μmol·L-1)。②与空白对照组(86.4±6.3)nmol·g-1.min-1相比,SLI 3,10和30 mg·kg-1组CYP1A2活性分别为83.4±6.6,82.5±4.0和(83.4±6.6)nmol·g-1.min-1。与空白对照组(16.1±0.9)nmol·g-1.min-1比较,SLI 3,10和30 mg·kg-1组CYP3A活性分别为15.7±0.6,15.9±0.7和(15.9±1.0)nmol·g-1.min-1,无显著性差异。③以临床血药浓度为依据设计的一系列浓度的SLI 0.0002,0.0006,0.002,0.006,0.017,0.052,0.156和0.468 g.L-1的ATP酶活性分别与空白对照组进行比较(5.8,5.3,5.8,5.5,5.8,5.2,,5.8,5.3,vs 5.75μmol·g-1.min-1),无显著性差异。结论SLI临床给药剂量既不能体外抑制人CYP1A2,CYP2D6,CYP3A4,CYP2C19和CYP2C9酶活性,也不能诱导大鼠CYP1A2和CYP3A,同时也不是P-gp的体外抑制剂或底物。

细胞色素P450CYP3A65斑马鱼模型建立及对环境污染物的生物响应

目的：建立细胞色素P450CYP3A65荧光标记转基因斑马鱼用于快速、直观检测重金属（铜、镉、锌）及类二噁英化合物五氯联苯（ PCB126）等环境污染物。方法 PCR方法钓取斑马鱼CYP3A65基因调控序列，构建于pT2AL200R150G转座载体克隆位点，将pTol2-CYP3A65-EGFP 质粒与pCS-TP 转座酶mRNA共同注射入斑马鱼胚胎单细胞，通过荧光筛选，遗传选育和建立荧光标记转基因斑马鱼品系 Tg （ CYP3A65-EGFP）；利用该转基因斑马鱼胚胎及幼鱼检测上述3种重金属及 PCB126的生物学响应。结果成功建立了CYP3A65荧光标记转基因斑马鱼品系，绿色荧光在肝和消化道表达；该转基因斑马鱼对3种重金属及PCB126产生生物学响应。染毒96 h后，铜0.1和0.2μmol·L^－1、镉0.35和0.7μmol·L^－1组、锌1.5和3μmol·L^－1组、PCB126所有浓度组荧光相对表达量显著增强（ P＜0.01）；铜0.9μmol·L^－1组、镉2.7和5.4μmol·L^－1组及锌24μmol·L^－1组荧光相对表达量显著减弱（ P＜0.01）；染毒168 h 后，铜0.1和0.2μmol·L^－1组、镉0.35和0.7μmol·L^－1组、锌1.5和3μmol·L^－1组及PCB1262～32μmol·L^－1浓度组荧光相对表达量显著增强（ P＜0.01），铜0.9μmol·L^－1组、镉2.7和5.4μmol·L^－1组、锌12和24μmol·L^－1组荧光相对表达量减弱（ P＜0.05），随着浓度增加荧光减弱，呈现出量效关系。结论 CYP3A65荧光标记转基因斑马鱼模型可以用来检测铜、镉、锌及PCB126等环境污染物。

孕烷X受体对细胞色素P450 CYP3A基因表达的调控在中药配伍禁忌及中药毒性早期预测中的应用

中药配伍禁忌是药物与机体相互作用本质的具体体现,是中药配伍理论的重要组成内容。毒性早期预测是药物安全性评价的重要组成部分。快速、早期获得药物可能的毒性反应数据,建立一种简单、可靠地中药配伍禁忌和中药毒性早期预测方法,是当今毒理学领域研究的重点之一。孕烷X受体(PXR)是一种配体依赖性转录因子,大量临床药物作为其配体或激活剂通过激活PXR而诱导细胞色素P450 CYP3A(CYP3A)基因表达。联合用药过程中,PXR的激活可能会增加药物发生相互作用和不良反应的风险,导致药效降低甚至产生毒性。本文从PXR-CYP3A途径为药物相互作用研究提供分子学机制,以及为中药配伍禁忌的研究和药物毒性早期预测提供了新思路。

Effect of Intestinal Cytochrome P450 3A on Phytochemical Presystemic Metabolism

Phytochemicals, orally administered substances, are found to undergo presystemic metabolism mainly in the intestine. Although early researches confirmed the role of intestinal bacteria in phytochemical presystemic metabolism, along with the development of molecular biology in investigating intestinal metabolism, a breakthrough has been won in research into metabolizing enzymes and transporters in intestine, which demands more attention and further studies. Recently, Cytochrome P450 3A has been found to be the most effective enzyme in mediating both oxidative （Phase Ⅰ ） and conjugative （Phase Ⅱ ） metabolism in the intestine. The present review summarizes the current findings correlated with the effect of intestinal cytochrome P450 3A on phytochemical presystemic metabolism, which provides a good basis for further research on phytochemical pharmacokinetics.

Expression of cytochrome P450 2A13 in human non-small cell lung cancer and its clinical significance

Lung cancer is one of the most important causes of cancer-related mortality worldwide. Human cytochrome P450 2A13 enzyme （CYP2A13） is predominantly expressed in the respiratory tract and could catalyze various carcinogens. In this study, we quantified CYP2A13 expression in non-small cell lung cancer （NSCLC） tissues and examined the relation between CYP2A13 and clinicopathologic factors. Thirty-five paired lung cancer and normal tissues were studied for the expression of the CYP2A13 gene by using real-time PCR and Western blot- ting assays. We also investigated the relationship between CYP2A13 expression and clinicopathologic factors such as age, gender, histology and lymph node status in tumor tissues. SPSS （17.0） statistical software was applied for data analysis. The real-time PCR results showed that there was no significant difference in the CYP2A13 mRNA transcript levels between tumor and paired normal tissues in the 35 samples and in 12 paired squamous cell car- cinomas. In adenocarcinoma, the expression of CYP2A13 mRNA in tumor tissues was 12.5% of that in adjacent tissues （P 〈 0.05） and it was not associated with age, gender, histology and lymph node status of the patients. The amounts of CYP2A13 proteins detected by Western blotting assays correlated well with those of the correspond- ing mRNAs. In conclusion, the expression of CYP2A13 was downregulated in lung adenocarcinoma. CYP2A13 may be involved in the development and progression of lung adenocarcinoma.

细胞色素P450异构体CYP3A4^(*)1G基因多态性对老年直肠癌手术患者芬太尼用药量及术后镇痛效果的影响

目的探讨细胞色素P450异构体CYP3A4~*1G基因多态性对老年直肠癌手术患者芬太尼用药量及术后镇痛效果的影响。方法选取2016年2月至2018年5月于我院行直肠癌根治术的老年患者共173例,通过术前外周血PCR和测序评估CYP3A4~*1G的基因分型,对比CYP3A4~*1G突变(突变组)和野生型(野生组)患者其术中芬太尼用药量,采用视觉模拟评分(VAS)用于在术后不同时间点评估患者的疼痛程度,对比患者自控镇痛(PCA)的初次镇痛时间和术后24 h芬太尼使用量。结果在所有173例患者中,CYP3A4~*1G变异等位基因的频率为0.191,其中突变组共61例,野生组共112例。其中突变组术中使用的平均芬太尼药物总量显著低于野生组,差异具有统计学意义(t=7.662,P <0.001),突变组患者术后初次追加镇痛时间显著高于野生组(t=7.279,P <0.001),且突变组术后24 h芬太尼用量显著低于野生组(t=13.847,P <0.001),突变组在术后2、4和8 h的疼痛VAS评分均显著低于野生组(P <0.05)。结论 CYP3A4~*1G基因多态性在老年直肠癌手术麻醉中与芬太尼的药代动力学密切相关,CYP3A4~*1G遗传多态性可能是导致患者对芬太尼镇痛反应的个体差异的重要因素之一。

小型猪肝脏细胞色素P450基因CYP3A29克隆与序列分析

设计引物通过RT-PCR扩增巴马香猪和贵州小型猪香猪药物代谢关键P450基因CYP3A29基因的全长编码区,纯化并克隆入pMD 18-T载体,通过序列测定和比对分析其序列变异情况。17头小型猪共发现CYP3A29基因的5个单核苷酸变异位点,均表现为同义替换,对编码的氨基酸序列无影响;猪CYP3A29的单核苷酸变异和品系相关,在不同品系中可表现出品系特有的单核苷酸变异;贵州小型香猪G3 a2902发现1个可变剪接,缺失89 bp并产生移码,改变编码蛋白。本研究克隆的巴马香猪和贵州小型香猪CYP3A29基因及其序列变异情况,可为其作为药物代谢模型的应用提供技术资料。

人细胞色素P450 3A4突变体CYP3A4.3,CYP3A4.4,CYP3A4.5和CYP3A4.18重组酶的异源表达与活性

目的重组表达人细胞色素P450(CYP)3A4突变体CYP3A4.3,CYP3A4.4,CYP3A4.5和CYP3A4.18蛋白,为CYP3A4代谢活性的体外研究提供单一酶源。方法应用杆状病毒表达系统构建含有上述各CYP3A4突变体基因序列的重组病毒,将其连同含人源还原型烟酰胺腺嘌呤二核苷磷酸-P450氧化还原酶(POR)和细胞色素b5基因的重组病毒共同感染昆虫草地夜蛾细胞Sf9得到CYP3A4突变体与POR和细胞色素b5共表达的重组蛋白,分别以高效液相色谱法和荧光分析法测定各重组酶对睾酮和7-甲氧基-4-三氟甲基香豆素的代谢活性。结果在mRNA分子水平上验证了CYP3A4突变体CYP3A4*3,CYP3A4*4,CYP3A4*5和CYP3A4*18基因在Sf9细胞中的转录。感染了各重组病毒的Sf9细胞裂解液对睾酮和7-苄氧基-4-三氟甲基香豆素有明显代谢。结论应用杆状病毒-昆虫细胞表达系统在体外成功表达了具有催化活性的人CYP3A4突变体CYP3A4.3,CYP3A4.4,CYP3A4.5和CYP3A4.18蛋白,其中CYP3A4.5活性显著低于野生型蛋白,CYP3A4.18活性显著高于野生型蛋白,而CYP3A4.3和CYP3A4.4与野生型蛋白活性近似。

白纹伊蚊细胞色素P450 CYP6N3基因的原核表达及鉴定

目的 对白纹伊蚊细胞色素P4 5 0CYP6N3基因进行原核表达 ,以获得高效表达蛋白。方法 根据CYP6N3基因的全长cRNA为模板进行RT -PCR。产物经T -A克隆测序鉴定后 ,亚克隆入原核融合表达载体 pGEX - 4T - 1(含有编码 2 6KDaGST的基因序列 )中 ,在大肠杆菌BL2 1(DE3)中进行原核表达。将细菌总蛋白进行SDS聚丙烯酰胺凝胶电泳分析 ,通过Westernblot分析鉴定目的蛋白的位置 ,并运用核酸蛋白分析仪扫描凝胶以确定表达产物的含量。结果 获得了高效表达的融合蛋白GST -CYP6N3,表达量占菌体总蛋白的 37 4 9%。结论 本实验成功地异源表达了白纹伊蚊CYP6N3基因 ,为体外重建细胞色素P4 5 0CYP6N3单加氧酶系 。

黑龙江省寒地高血压患者细胞色素P450 CYP3A5基因多态特征性分析

目的:分析黑龙江省寒地原发性高血压患者CYP3A5基因分布频率,研究该地区CYP3A5基因特征性,为个体化精准治疗高血压病提供临床依据。方法:应用原位杂交荧光染色技术检测入组的高血压患者及健康者CYP3A5基因型,根据基因型分组对比基因型频率及等位基因分布频率,与已报道的我国其他非高寒地区、不同民族进行比较,探讨黑龙江省寒地高血压患者CYP3A5基因特征性。结果:入组的原发性高血压患者共512例,其中GG型284例(55.47%),AG型181例(35.35%),AA型47例(9.18%),G、A等位基因频率分别为73.14%,26.86%。与我国其他学者已报导的湖南长沙、四川南充高血压患者对比CYP3A5基因型频率及等位基因分布频率均具有显著统计学差异(P<0.05)。与新疆哈萨克族高血压患者CYP3A5等位基因频率和基因型分布频率对比均存在统计学差异(P=0.000,P=0.000)。结论:(1)CYP3A5基因分布频率存在南北地域性差异;(2)CYP3A5基因分布频率存在民族性差异。

2种烷基多环芳烃对仿刺参CYP450和p53基因表达的影响研究

海洋环境中多环芳烃类(PAHs)主要来源于海洋溢油事故以及沿海石油化工企业的废水排放,国内外大量研究发现海洋中的多环芳烃对海洋生物造成了潜在的生态风险。为了揭示不同浓度多环芳烃类污染物对海参的生态毒理效应,将仿刺参(Apostichopusjaponicus)分别暴露于不同浓度的2种烷基多环芳烃3-甲基菲(5、10、100μg·L^-1)和2-甲基蒽(5、10、50μg·L^-1)中,检测暴露3d、7d和14d后,3-甲基菲和2-甲基蒽胁迫下仿刺参CYP450和p53基因的相对表达量。结果表明,3-甲基菲和2-甲基蒽胁迫下,仿刺参CYP450和p53基因的表达均对毒物产生了不同程度的响应。与对照组相比,3-甲基菲各处理组对仿刺参CYP450和p53基因的表达均产生显著的抑制作用(P<0.05);2-甲基蒽各处理组对仿刺参CYP450和p53基因的表达影响作用不同,暴露7d后,2-甲基蒽各处理组对仿刺参CYP450基因的表达表现出抑制作用,对p53基因的表达表现出诱导作用。相同浓度与时间胁迫下,2-甲基蒽对仿刺参CYP450和p53基因表达的影响比3-甲基菲的影响大。上述研究结果表明,3-甲基菲和2-甲基蒽均可不同程度影响仿刺参CYP450和p53基因的表达,且与3-甲基菲相比,2-甲基蒽对仿刺参CYP450和p53基因表达的影响较明显。上述结果为多环芳烃类污染物对仿刺参的生物毒性评价提供了基础数据。

CYP3A5在胰腺癌中调节葡萄糖代谢的新机制

胰腺导管腺癌是一种高度侵袭性的恶性肿瘤,其致死性主要源于早期诊断困难和治疗手段的局限。在胰腺癌中,糖酵解过程的改变被视为其主要的代谢特征,而葡萄糖转运则是该过程中的一个关键限速步骤。CYP3A5作为一种细胞色素P450酶,参与了药物代谢和外源性致癌物的解毒过程,但其在胰腺导管腺癌发生和发展中的直接生理作用尚不明确。

柴胡水提液对大鼠肝CYP450酶含量及其亚型CYP3A、CYP2E1和CYP1A2活性的影响

目的:研究柴胡水提物对大鼠细胞色素CYP450酶含量及对亚型CYP3A、CYP2E1和CYP1A2活性的影响。方法:大鼠灌胃给予柴胡水提液7d后取肝脏称重并制备肝微粒体,紫外分光光度法测定肝微粒体细胞色素b5(Cytb5)、P450的含量及CYP3A的活性,高效液相法测定CYP2E1和CYP1A2的活性。结果:与空白对照组比较,柴胡水提液组大鼠肝指数无显著变化,CYP450含量减少但统计学意义(P>0.05),CYP1A2活性极显著降低(P<0.01),CYP2E1活性明显降低(P<0.05)。结论:柴胡水提液降低大鼠肝脏CYP450酶含量并抑制CYP2E1和CYP1A2的活性,由于抑制CYP450酶活性,减少对其他药代谢,提示若柴胡水提液与经CYP2E1和CYP1A2代谢的药物同用,有可能会影响这些药物的临床疗效,临床用药时应慎重考虑。