CHRARACTERIZATION OF FOLLICULAR STEM CELLS IN TG.AC MICE

v- ha = Tg。研究表明，AC小鼠在特定化学物质、全层损伤和紫外线照射下会发生皮肤肿瘤，肿瘤的发生依赖于转基因表达的激活。先前的研究表明，转基因表达可以在增生毛囊的一个区域检测到，一些人认为这是一个假定的干细胞库。这些研究的目的是利用ras基因在Tg中的诱导表达。交流小鼠皮肤作为鉴定和表征居住在毛囊中的潜在干细胞群的手段。我们已经证明，通过毛囊间表皮的去除，转基因表达的发展，这是直接的证据，鳞状病变起源于毛囊起源的小鼠。此外，我们通过原位杂交在磨损后6天的再上皮化皮肤中发现了转基因表达，加强了潜在肿瘤细胞起源于毛囊并表达ras转基因的假设。进一步表征Tg的乳头状瘤前体细胞。在AC小鼠皮肤中，我们从tpa处理的非肿瘤皮肤中分离出角质形成细胞，并结合流式细胞术和β - 1整合素（一种假定的滤泡干细胞标记物）特异性抗体的孵育，表明转基因表达与β - 1阳性细胞同时发生。这进一步证明了滤泡干细胞表达ras基因。最后，为了解决毛囊干细胞具有哪些特征为造血启动子驱动的转基因转录激活提供了一个允许的微环境的问题，我们已经证明了CD34，一种造血干细胞标记物，在毛囊的一个非常特定的区域表达。由于有证据表明β - 1整合素和CD34在造血干细胞中共表达，我们建议结合流式细胞术、免疫组织化学和激光捕获显微解剖来更紧密地定义和表征乳头状瘤前体细胞。——转基因;表皮干细胞；角化细胞;激光捕获显微解剖

The v-Ha-ras Tg.AC mouse has been shown to develop cutaneous neoplasms in response to specific chemicals, full-thickness wounding, and UV radiation exposure, and tumor development is dependent upon activation of expression of the transgene. Previous work has demonstrated that transgene expression can be detected in a region of hyperplastic hair follicles identified by some as a putative stem cell resevoir. The goal of these studies is to utilize inducible expression of the ras transgene in Tg.AC mouse skin as a means to identify and characterize potential stem cell populations residing in the hair follicle. We have shown through the use of removal of the interfollicular epidermis through felt-wheel abrasion that transgene-expressing develop, which is direct evidence that squamous lesions arise from hair follicle origin in these mice. In addition, we have found transgene expression by in situ hybridization in re-epithelialising skin as early as 6 days following abrasion, strengthening the hypothesis that latent neoplastic cells originate from the hair follicle and express the ras transgene. To further characterize papilloma precursor cells in Tg.AC mouse skin, we have isolated keratinocytes from TPA-treated, non-tumor bearing skin and using a combination of flow cytometry and incubation with an antibody specific to Beta-one integrin, a putative follicular stem cell marker, have shown that transgene expression is co-incident with beta- one positive cells. This is further evidence that follicular stem cells express the ras transgene. Finally, in order to address the issue of what characteristics follicular stem cells possess that provides a permissive microenvironment for transcriptional activation of a transgene driven by a hematopoietic promoter, we have demonstrated that CD34, a hematopoietic stem cell marker, is expressed in a very specific region of the hair follicle. Since there is evidence that Beta-one integrin and CD34 are co-expressed in hematopoietic stem cells, we propose to use a combination of flow cytometry, immunohistochemistry, and laser capture microdissection to more closely define and characterize papilloma precursor cells. - Transgenic; epidermal stem cells; keratinocyte; laser capture microdissection