Mechanism of cell death induced by aflatoxin B1

描述：（根据申请人的要求改编）s摘要）：本研究的目的是 以确定黄曲霉毒素B1（AFB 1）如何杀死肝细胞。首先，我将探索 导致肝细胞死亡的一系列生化事件。Seond，自从 黄曲霉毒素被认为通过造成DNA损伤来发挥其生物学作用， 我将确定黄曲霉毒素引起的几种DNA损伤中的哪一种 诱导死亡前的基因表达模式。第三，我将 筛选在AFB 1诱导的细胞死亡过程中mRNA水平发生变化的新基因， 使用DNA微阵列方法。细胞凋亡是一种机制， 肝脏消除癌前细胞，并且已知是由AFB 1诱导的， 转化的肝细胞最初的实验将检验这一假设， 细胞凋亡是AFB 1细胞毒性的原因。基因的表达知道 参与细胞凋亡是黄曲霉毒素B1细胞毒性的主要原因。 已知参与细胞凋亡的基因的表达将通过 定量RT-PCR，以确定参与的特定途径， 诱导细胞凋亡，从而确定哪些基因容易受到影响 致癌突变将采用原代大鼠肝细胞进行建模 体内黄曲霉毒素B1的暴露。为了精确定位 从DNA损伤到细胞死亡，含有AFB 1的DNA诱导的病变 将AFB 1-N7-Gua、AFB 1-FAPY和AP位点引入肝细胞。的 每种损伤对细胞死亡和凋亡基因表达的影响， 测量以确定哪种特异性DNA加合物诱导细胞死亡。最后， 将构建大鼠基因的微阵列，并用于寻找新的 在AFB 1诱导的细胞死亡过程中被诱导或抑制的基因。的基因 表达被AFB 1暴露改变，特别是那些似乎 与已知的凋亡基因共同调节，可能参与了 细胞对毒素的反应

DESCRIPTION: (Adapted from the Applicant?s Abstract): The goal of this study is to determine how aflatoxin B1 (AFB1) kills hepatocytes. First, I shall probe the sequence of biochemical events that leads to liver cell death. Seond, since aflatoxin is believed to exert its biological effects by inflicting DNA damage, I shall determine which of the several types of DNA damage induced by aflatoxin induces the pattern of gene expression that precedes death. Third, I shall screen for novel genes whose mRNA levels change during AFB1-induced cell death, employing a DNA microarray approach. Apoptosis is the mechanism by which the liver eliminates pre-neoplastic cells, and is known to be induced by AFB1 in transformed liver cells. Initial experiments will test the hypothesis that apoptosis is responsible for the cytotoxicity of AFB1. Expression of genes know to participate in apoptosis is responsible for the cytotoxicity of AFB1. Expression of genes known to participate in apoptosis will be examined by quantitative RT-PCR in order to identify the specific pathways involved in the induction of apoptosis, thereby establishing which genes are vulnerable to oncogenic mutations. Primary rat hepatocytes will be employed in order to model in vivo AFB1 exposure realistically. In order to pinpoint the molecular pathway from DNA damage to cell death, DNA containing the AFB1-induced lesions AFB1-N7-Gua, AFB1-FAPY, and AP sites will be introduced into hepatocytes. The impact of each lesion on cell death, and apoptosis gene expression, will be measured to determine which specific DNA adduct(s) induce cell death. Finally, a microarray of rat genes will be constructed and used to search for novel genes that are induced or repressed during AFB1-induced cell death. Genes whose expression is altered by AFB1 exposure, and especially those that appear to be co-regulated with known apoptosis genes, are likely to participate in the cellular response to the toxin.