Analysis Of Mechanisms Of Testicular Toxicity Using DNA

该项目的一组目标是确定已知生殖毒物诱导的各种睾丸病变中基因表达的变化，确定暴露于毒物后改变的细胞类型和机制，并定义基因表达变化的顺序，以区分睾丸病变中的早期效应和晚期效应。我们假设基因表达模式的改变可用于识别原发性和继发性反应，并识别睾丸中毒物的初始靶细胞，并且基因表达变化的时间模式可用于定义毒性机制。这些研究正在确定睾丸中基因表达模式的变化，以响应镉，一种已知的男性生殖毒物。更长期的目标是确定不同毒物对睾丸单个细胞类型中基因表达的具体影响，毒物在睾丸不同细胞类型中诱导的基因表达变化的时间过程，不同剂量毒物的基因表达变化模式，以及其他已知或疑似导致睾丸损伤的化学物质诱导的基因表达变化模式的相似性和差异。 另一组目标是制备小鼠睾丸微阵列，这将为男性生殖毒物的研究以及基因敲除、内分泌操纵和其他治疗的影响的研究提供高度信息。用从小鼠睾丸和不同发育阶段分离的生殖细胞的mRNA构建的8个cDNA文库从5'端进行单程测序，以鉴定用于此目的的克隆。对28，000多个克隆进行了测序，并将22，000多个EST存入公共数据库。利用该序列信息和公共数据库中的其他信息构建了小鼠睾丸22K基因芯片。研究正在进行中，使用这种微阵列来检查雄性生殖毒物对小鼠睾丸中基因表达的影响。

One set of goals of this project is to identify the changes in gene expression in various testis lesions induced by known reproductive toxicants, determine which cell types and mechanisms are altered after exposure to a toxicant, and define the sequence of changes in gene expression to separate earlier from later effects in testis lesions. We hypothesize that altered patterns of gene expression can be used to identify primary and secondary responses and to identify the initial target cell of a toxicant in the testis, and that the temporal pattern of changes in gene expression can be used to define the mechanisms of toxicity. These studies are identifying changes in patterns of gene expression in the testis in response to cadmium, a known male reproductive toxicant. Longer term goals are to determine the specific effects of different toxicants on gene expression in individual cell types of the testis, the time course of changes in gene expression induced by toxicants in different cell types of the testis, the patterns of changes in gene expression with different doses of toxicants, and the similarities and differences in the patterns of change in gene expression induced by other chemicals known or suspected to cause injury to the testis. Another set of goals is to prepare a mouse testis microarray that will be highly informative for studies of male reproductive toxicants, as well as for studies on effects of gene knockouts, endocrine manipulations, and other treatments. Eight cDNA libraries constructed with mRNA from mouse testis and germ cells isolated at different stages of development were subjected to single-pass sequencing from the 5' end to identify clones to be used for this purpose. Over 28,000 clones were sequenced and over 22,000 ESTs were deposited in public databases. A mouse testis 22K microarray has been constructed using this sequence information and other information public databases. Studies are underway to use this microarray to examine the effects of male reproductive toxicants on gene expression in the mouse testis.