转基因作物标记蛋白潮霉素磷酸转移酶活性的测定方法

潮霉素磷酸转移酶 (HPT)是转基因作物中重要的标记蛋白 ,其活性的测定对含有该标记基因的转基因作物的植株构建、基因沉默、标记基因去除及安全性分析等具有重要的作用。本文综述了潮霉素磷酸转移酶活性测定的几种方法 ,包括放射化学法、抑菌生长法及酶耦联法 ,其中放射化学法用于检测组织粗提蛋白中的HPT活性 ,而其他两种方法适用于纯化过的HPT蛋白活性的检测     

转Xa21基因大米亚慢性毒性实验

目的　观察Xa2 1转基因大米对大鼠的亚慢性毒性损害。方法　将Wistar大鼠按性别体重随机分为转基因大米组、非转基因大米组和AIN93G正常对照组。分别饲相应饲料 90天 ,自由进食。观察大鼠体重、身长、血常规、血生化、脏器重量、骨密度和脏器的病理学检查。结果　与非转基因大米组相比 ,实验中期转基因大米组血糖降低 ,胆固醇和高密度脂蛋白升高 ,实验结束时上述差异消失 ,但雌性组谷草转氨酶活性显著升高。与AIN93G正常对照组相比 ,实验中期转基因大米组体重、甘油三酯、高密度脂蛋白显著增高 ,血糖降低 ;实验结束时上述差异消失。脑、心、脾、肺、肾、胃、十二指肠、肾上腺、睾丸、卵巢病理检查未见异常。结论　现有实验结果不能证实转基因大米对大鼠有亚慢性毒性作用     

Xa21转基因大米对大鼠致畸作用的实验研究

目的　观察Xa2 1转基因大米对大鼠胚胎生长、发育的影响。方法　将初断乳Wistar大鼠按雌雄分别随机分为 4组 :转基因大米组、非转基因大米组、AIN93G对照组和敌枯双阳性对照组。单笼喂养 ,饲相应鼠料 ,喂满 90天 ,雌雄合笼。观察母鼠和胎鼠的生长发育情况。结果　转基因大米组孕鼠增重、活胎体重、身长、尾长均显著高于阳性对照组 ,而死胎数、吸收胎数、畸形率 (外观、内脏、骨骼 )均显著低于阳性对照组。转基因大米组与非转基因大米组、AIN93G对照组相比 ,所有观察指标均无统计学差异。结论　转Xa2 1基因大米与非转基因大米相比 ,对大鼠受孕率、胚胎生长发育无显著性差异。     

转豇豆胰蛋白酶抑制剂大米的致畸作用研究

目的　用表达杀虫蛋白CpTI(豇豆胰蛋白酶抑制剂 )的大米喂养Wistar大鼠 ,研究其是否具有致畸作用。方法　将断乳的Wistar大鼠随机分为 4组 ,转基因大米组、非转基因大米组、阴性对照组和阳性对照组 ,敌枯双为阳性对照物。转基因大米组含 78 3%转基因大米 ,非转基因大米组含 74 7%与转基因大米同品系的非转基因大米 ,两个对照组的饲料为AIN93G ,三种饲料的宏量及微量营养素的含量相同。大鼠性成熟后 ,进行传统的致畸实验 ,观察母鼠及胎鼠的情况。结果　转基因大米组的孕鼠增重、胎鼠体重、身长和尾长显著高阳性对照组 ,畸形率 (包括外观畸形、骨骼畸形和内脏畸形 )显著低于阳性对照组 (P <0 0 1) ,转基因大米组、非转基因大米组及阴性对照组间的所有指标均无显著性差异 (P >0 0 5 )。结论　此种转CpTI基因的大米对大鼠无母体毒性、胚胎毒性和致畸作用。     

肝部分切除术对大鼠门静脉途径肝细胞转基因效率的影响

目的 :探讨肝部分切除术对大鼠肝细胞门静脉途径转基因效率的影响。方法 :Wistar大鼠分 4组 ,每组 5只 :A1组 :30 %切肝并于门静脉注射脂质体—质粒 DNA混合物 (lipofectamine- p EGFP- N1DNA complexes,lipoplexes) ;A2组 :30 %切肝并于门静脉注射生理盐水 (NS) ;B1组 :70 %切肝并于门静脉注射 lipoplexes;B2组 :70 %切肝并于门静脉注射 NS。转染后 15 d均以胶原酶消化、Percoll液梯度离心法获取纯化肝细胞悬液 ;以 NS组为对照并以 GFP为荧光标记物 ,用流式细胞仪 (flow cytometry,FCM)分析各实验组肝细胞的转染率。结果 :A1组和 B1组的平均转染率分别为 :(1.12± 0 .5 0 ) %、(3.91± 1.6 8) % ;B1组明显高于 A1组 (P<0 .0 5 )。结论 :与 30 %肝部分切除术相比 ,70%切肝可显著提高大鼠肝细胞门静脉途径转基因的效率     

乙肝病毒X基因转基因树鼩模型的建立和病理学研究

目的:探索以精原细胞作为载体建立带乙肝病毒X基因的树鼩转基因动物模型的可行性.方法:首先构建带乙肝病毒pcDNA3.1- HBx基因的表达载体;然后以脂质体2000包被之.将此复合物以微量注射器注入8只雄性树鼩之睾丸组织中,1周后重复注射1次.注射后6周,使处理过的雄树鼩与雌性树鼩交配.在仔树鼩出生后1个月,取其肝组织作活检,并以多聚酶链式反应(PCR)对其DNA表达进行鉴定,观察其内是否存在乙肝病毒X基因.结果:处理的8只雄性树鼩中,6只具有生育能力,产仔树鼩11只.其中,1只仔树鼩乙肝病毒X基因阳性(阳性率9.09%).基因测序结果证实为乙肝病毒X基因.病理检查结果显示仔树鼩肝组织的改变包括:肝细胞坏死、慢性炎细胞在汇管区的浸润.结论:以精原干细胞作为载体,建立带乙肝病毒X基因的树鼩转基因动物模型是可行的.     

砷致癌动物模型研究现状

人类流行病学资料显示,长期暴露于砷化合物可以导致人类皮肤、肺、膀胱、肝和肾脏的癌症。近来报道指出,砷在小鼠和大鼠的相应器官也可诱发促癌或完全致癌作用,给予p53+/-和K6/ODC转基因小鼠无机砷的甲基化代谢产物二甲基胂酸(DMA)或亚砷酸盐,对转基因小鼠的皮肤或膀胱有一定程度的致癌、联合致癌或促癌作用。由于到目前为止尚无无机砷化合物致癌的动物模型,致使其致癌机制的研究一直滞后,所以大量研究人员致力于野生型和转基因动物的砷致癌动物模型研究,本文就这一方面的研究进展做一综述。     

The use of genetically modified mice in cancer risk assessment: Challenges and limitations

Abstract

The use of genetically modified (GM) mice to assess carcinogenicity is playing an increasingly important role in the safety evaluation of chemicals. While progress has been made in developing and evaluating mouse models such as the Trp53^+/?, Tg.AC and the rasH2, the suitability of these models as replacements for the conventional rodent cancer bioassay and for assessing human health risks remains uncertain. The objective of this research was to evaluate the use of accelerated cancer bioassays with GM mice for assessing the potential health risks associated with exposure to carcinogenic agents. We compared the published results from the GM bioassays to those obtained in the National Toxicology Program’s conventional chronic mouse bioassay for their potential use in risk assessment. Our analysis indicates that the GM models are less efficient in detecting carcinogenic agents but more consistent in identifying non-carcinogenic agents. We identified several issues of concern related to the design of the accelerated bioassays (e.g., sample size, study duration, genetic stability and reproducibility) as well as pathway-dependency of effects, and different carcinogenic mechanisms operable in GM and non-GM mice. The use of the GM models for dose-response assessment is particularly problematic as these models are, at times, much more or less sensitive than the conventional rodent cancer bioassays. Thus, the existing GM mouse models may be useful for hazard identification, but will be of limited use for dose-response assessment. Hence, caution should be exercised when using GM mouse models to assess the carcinogenic risks of chemicals.     

Lactate detection in inducible and orthotopic Her2/neu mammary gland tumours in mouse models

This study compared the steady state concentration of lactate in an inducible Her2/nue transgenic breast cancer mouse model and in tumours from the same Her2/neu model grown orthotopically. In vivo lactate was detected by MRS using the Hadamard encoded selective multiple quantum coherence pulse sequence (HadSelMQC) recently developed by our laboratory. A lower lactate signal was observed in the inducible tumours compared to orthotopic tumours in vivo, while ex vivo analysis of perchloric acid extracts revealed similar amounts of this metabolite in both models. Histological staining of mammary tumour specimens showed a much higher level of fat tissue in inducible tumours compared to the orthotopic model. Phantom studies with [3‐¹³C] lactate indicated that a lipid environment could significantly reduce the T2 of lactate and impede its detection. The transgenic inducible model for breast cancer not only better recapitulated the biological aspects of the human disease but also provided additional characteristics related to in vivo detection of lactate that are not available in orthotopic or xenograft models. This study suggests that the level of lactate measured by the HadSelMQC pulse sequence may be underestimated in human patients in the presence of high lipid levels that are typically encountered in the breast. Copyright © 2012 John Wiley & Sons, Ltd.     

The value of animal models to study immunopathology of primary human Sjögren's syndrome symptoms

Sjögren's syndrome (SjS) is a complex chronic autoimmune disease of multifactorial etiology that results in eventual loss of secretory function in the exocrine glands. The challenges towards finding a therapeutic prevention or treatment for SjS are due primarily to a lack of understanding in the pathophysiological and clinical progression of the disease. In order to circumnavigate this problem, there is a need for appropriate animal models that resemble the major phenotypes of human SjS and deliver a clear underlying biological or molecular mechanism capable of defining various aspects for the disease. Here, we present an overview of SjS mouse models that are providing insight into the autoimmune process of SjS and advance our focus on potential diagnostic and therapeutic targets.