Roe A L, Howard G, Blouin R, Snawder J E
Graduate Center for Toxicology, Division of Pharmacology and Experimental Therapeutics, University of Kentucky, Lexington 40536-0082, USA.
Int J Obes Relat Metab Disord. 1999 Jan;23(1):48-53. doi: 10.1038/sj.ijo.0800756.
To characterize the effect(s) of gender, age (glycemic status) and obese state, on hepatic biotransformation activities, expression of cytochrome P450 (CYP450) mRNAs and glutathione transferase activity in the ob/ob mouse.
Male and female, ob/ob or ob/+ mice were killed at 3-4 months or 7-8 months of age. Hepatic microsomes, cytosol and RNA were prepared from each animal.
Male and female ob/ob and ob/+ mice, 3-4 or 7-8 months of age.
CYP450 form-specific activities of CYP1A1/1A2, CYP3A and CYP2B were estimated by determining the 0-dealkylation of alkoxyresorufin substrates (ethoxy-EROD, benzoxy-BROD and pentoxy-resorufin, PROD, respectively). CYP2E1-dependent, 4-nitrophenol hydroxylase (PNP-OH) and CYP3A-dependent erythromycin N-demethylase (ERY-DM) were also measured in hepatic microsomes. CYP1A2, CYP2E1 and CYP3A protein in microsomal fractions was determined by ELISA. Glutathione transferase activity (GST) was determined in hepatic cytosol and CYP1A2 and CYP2E1 mRNA was estimated by Northern blot analysis.
Female mice, regardless of glycemic status, showed an obesity enhanced level of CYP2E1-dependent PNP-OH activity and CYP2E1 protein as shown by ELISA. These increases were observed to be independent of the diabetic state, since 7-8 month-old mice had blood glucose levels identical to lean mice. The mRNA level of CYP2E1 in female mice also exhibited age-and obesity-influenced decreases in expression. No significant differences in CYP2E1 activity or expression were observed in male mice. CYP3A-dependent ERY-DM activity was significantly higher in young males, regardless of phenotype. CYP3A and CYP2B activities did not differ among any animals; however, CYP1A activity, while depressed in obese animals of both genders, was significantly different in old animals. Glutathione S-transferase activity was lower in obese male mice, whereas no difference was observed between lean and obese females
This study supports earlier observations in man and rats that the obese state produces alterations in the expression of important oxidation and conjugation pathways. In addition, this report more thoroughly examines the role of gender and glycemic status on biotransformation activities in the ob/ob mouse as demonstrated by increased CYP2E1 protein and CYP2E1-dependent activity in obese females, decreased CYP1A2 protein and CYP1A2-dependent activity in obese animals, and obesity had no effect of glutathione transferase in female mice, in contrast with the previously reported obesity-dependent decrease of this activity in male mice.
研究性别、年龄（血糖状态）和肥胖状态对ob/ob小鼠肝脏生物转化活性、细胞色素P450（CYP450）mRNA表达及谷胱甘肽转移酶活性的影响。
将3 - 4月龄或7 - 8月龄的雄性和雌性ob/ob或ob/+小鼠处死。从每只动物中制备肝脏微粒体、胞质溶胶和RNA。
3 - 4月龄或7 - 8月龄的雄性和雌性ob/ob及ob/+小鼠。
通过测定烷氧基试卤灵底物（分别为乙氧基 - EROD、苯氧基 - BROD和戊氧基试卤灵，PROD）的O - 脱烷基化反应来估计CYP1A1/1A2、CYP3A和CYP2B的CYP450形式特异性活性。还测定了肝脏微粒体中CYP2E1依赖性的4 - 硝基苯酚羟化酶（PNP - OH）和CYP3A依赖性的红霉素N - 脱甲基酶（ERY - DM）。通过ELISA测定微粒体组分中的CYP1A2、CYP2E1和CYP3A蛋白。在肝脏胞质溶胶中测定谷胱甘肽转移酶活性（GST），并通过Northern印迹分析估计CYP1A2和CYP2E1 mRNA。
无论血糖状态如何，雌性小鼠均表现出肥胖导致的CYP2E1依赖性PNP - OH活性和CYP2E1蛋白水平升高，ELISA结果显示如此。观察到这些升高与糖尿病状态无关，因为7 - 8月龄小鼠的血糖水平与瘦小鼠相同。雌性小鼠中CYP2E1的mRNA水平也呈现出受年龄和肥胖影响的表达下降。在雄性小鼠中未观察到CYP2E1活性或表达的显著差异。无论表型如何，年轻雄性小鼠中CYP3A依赖性ERY - DM活性显著更高。CYP3A和CYP2B活性在任何动物之间均无差异；然而，CYP1A活性在肥胖动物中降低，在老年动物中存在显著差异，且在两种性别中均如此。肥胖雄性小鼠的谷胱甘肽S - 转移酶活性较低，而瘦小鼠和肥胖雌性小鼠之间未观察到差异。
本研究支持了早期在人类和大鼠中的观察结果，即肥胖状态会导致重要氧化和结合途径的表达发生改变。此外，本报告更全面地研究了性别和血糖状态对ob/ob小鼠生物转化活性的作用，表现为肥胖雌性小鼠中CYP2E1蛋白和CYP2E1依赖性活性增加、肥胖动物中CYP1A2蛋白和CYP1A2依赖性活性降低，并且肥胖对雌性小鼠的谷胱甘肽转移酶无影响，这与先前报道的肥胖导致雄性小鼠该活性降低形成对比。
