Analysis of p53 inactivation in a human T-cell leukemia virus type 1 Tax transgenic mouse model.
Analysis of p53 inactivation in a human T-cell leukemia virus type 1 Tax transgenic mouse model.
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人类 T 细胞白血病病毒 1 型 Tax 转基因小鼠模型中 p53 失活的分析。
DOI：
10.1128/jvi.75.5.2185-2193.2001
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发表时间：
2001
影响因子：
5.4
通讯作者：
Ratner,L
中科院分区：
文献类型：
--
作者：
Portis,T;Grossman,WJ;Harding,JC;Hess,JL;Ratner,L
材料和方法如前所述产生粒酶B-Tax转基因小鼠（Tax）（14）。从杰克逊实验室购买了p53纯合缺失的小鼠（p53/）（18）。将Tax小鼠与p53/小鼠交配，并将所得p53/Tax后代交配以产生F2后代。每周监测F2小鼠的肿瘤形成率、发病率和死亡率。比较p53/Tax、p53/Tax、p53/Tax、p53/、p53/和p53/转基因同窝小鼠肿瘤的病理学特征。所有基因分型均按先前所述进行（14;杰克逊实验室方案）。将组织固定在10%中性缓冲福尔马林中，包埋在石蜡中进行切片，并如前所述用苏木精和伊红染色（14）。根据华盛顿大学动物护理指南，在无病原体的条件下饲养和维持所有小鼠。使用SPSS统计分析程序（SPSS，Inc.）进行Kaplan-Meier分析和统计计算。组织和细胞系。将从小鼠移除的新鲜肿瘤和组织释放到补充有10%胎牛血清、1%L-谷氨酰胺、1%丙酮酸钠和1%青霉素-链霉素的培养基（RPMI培养基; Life Technologies）中。用155 mM氯化铵裂解脾细胞制备物中的红细胞，并在培养前洗涤。肿瘤衍生的F8和SC大颗粒淋巴细胞（LGL）细胞系已在其他地方描述，并维持在RPMI培养基中（14，15）。含有野生型（204-3-1）和突变型（143- 2 M）p53的Abelson鼠白血病病毒（MLV）转化的前B细胞系由Naomi Rosenberg（Tufts University，Boston，MA）提供。(43)并维持在RPMI培养基中。人H1299非小细胞肺癌细胞系由Rainer Brachmann（华盛顿大学，圣路易斯，密苏里州）提供。并维持在补充有10%胎牛血清、1%小牛血清和1%小牛血清的Dulbecco改良Eagle培养基（DMEM）（Life Technologies）中。
MATERIALS AND METHODSMice. Granzyme B-Tax transgenic mice (Tax) were generated as previously described (14). Mice containing a homozygous deletion in p53 (p53/) were purchased from Jackson Laboratories (18). Tax mice were mated with p53/mice, and the resulting p53/Tax progeny were mated for production of F2 progeny. F2 mice were monitored weekly for rates of tumor formation, morbidity, and mortality. Pathological characteristics of tumors were compared among p53/Tax, p53/Tax, p53/Tax, p53/, p53/, and p53/transgenic littermates. All genotyping was performed as described previously (14; Jackson Laboratories protocol). Tissues were fixed in 10% neutral-buffered formalin, embedded in paraffin for sectioning, and stained with hematoxylin and eosin as described previously (14). All mice were bred and maintained under pathogen-free conditions in accordance with Washington University animal care guidelines. Kaplan-Meier analysis and statistical calculations were carried out using the SPSS statistical analysis program (SPSS, Inc.). Tissues and cell lines. Fresh tumors and tissues removed from mice were released into medium supplemented with 10% fetal bovine serum, 1% L-glutamine, 1% sodium pyruvate, and 1% penicillin-streptomycin (RPMI medium; Life Technologies). Erythrocytes in splenocyte preparations were lysed with 155 mM ammonium chloride and washed prior to culture. The tumor-derived F8 and SC large granular lymphocytic (LGL) cell lines have been described elsewhere and were maintained in RPMI medium (14, 15). Abelson murine leukemia virus (MLV)-transformed pre-B-cell lines containing wild-type (204-3-1) and mutant (143-2M) p53 were provided by Naomi Rosenberg (Tufts University, Boston, Mass.)(43) and maintained in RPMI medium. The human H1299 non-small-cell lung carcinoma cell line was provided by Rainer Brachmann (Washington University, St. Louis, Mo.) and maintained in Dulbecco modified Eagle medium (DMEM)(Life Technologies) supplemented with 10% fetal bovine serum, 1%