Kobayashi Tetsuro, Shimizu Atsushi, Nishifuji Koji, Amagai Masayuki, Iwasaki Toshiroh, Ohyama Manabu
Department of Veterinary Medicine, Faculty of Agriculture, Tokyo University of Agriculture and Technology, Fuchu, Tokyo 183-8509, Japan.
Vet Dermatol. 2009 Oct;20(5-6):338-46. doi: 10.1111/j.1365-3164.2009.00815.x.
Homeostasis of the epidermis and skin appendages is maintained by tissue-specific stem cells. In mice and humans, two populations of epithelial stem cells have been identified: one in the basal layer of the interfollicular epidermis and another in the bulge area of hair follicles. However, our understanding of canine epithelial stem cells is extremely limited. In this study, in vitro colony-forming assays were performed to locate highly proliferative keratinocytes in canine skin. Their phenotypic resemblance to epithelial stem cells in other species was also assessed. When equal numbers of epidermal or hair-follicle keratinocytes were cultured, colonies derived from follicular keratinocytes were significantly larger both in total numbers and size, than those derived from epidermal keratinocytes. In addition, immunoreactivity for CD34, a putative bulge stem-cell marker in the mouse, was predominantly detected in follicular keratinocytes. Thus in dogs, follicular keratinocytes were distinct from epidermal keratinocytes in proliferative capacity and CD34 expression. Using microdissection, highly proliferative keratinocytes were located within the middle portion of hair follicles, including the bulge area. Immunohistochemical study revealed that keratin 15, an established marker of bulge stem cells in mice and humans, was also predominantly expressed in the canine bulge area. Flow cytometry analysis revealed high numbers of keratin-15-positive cells in the highly proliferative keratinocyte compartment. Of note, keratin 15(high) cells possessed the phenotypic characteristics of putative stem cells. This study represents the first in vitro identification and isolation of highly proliferative canine keratinocytes, which represent candidate epithelial stem cells.
表皮和皮肤附属器的稳态由组织特异性干细胞维持。在小鼠和人类中，已鉴定出两种上皮干细胞群体：一种位于毛囊间表皮的基底层，另一种位于毛囊的隆突区。然而，我们对犬上皮干细胞的了解极为有限。在本研究中，进行了体外集落形成试验以定位犬皮肤中高度增殖的角质形成细胞。还评估了它们与其他物种上皮干细胞的表型相似性。当培养等量的表皮或毛囊角质形成细胞时，来自毛囊角质形成细胞的集落在总数和大小上均明显大于来自表皮角质形成细胞的集落。此外，在小鼠中作为假定隆突干细胞标志物的CD34的免疫反应性主要在毛囊角质形成细胞中检测到。因此，在犬中，毛囊角质形成细胞在增殖能力和CD34表达方面与表皮角质形成细胞不同。通过显微切割，高度增殖的角质形成细胞位于毛囊的中部，包括隆突区。免疫组织化学研究表明，角蛋白15，一种在小鼠和人类中已确定的隆突干细胞标志物，也主要在犬的隆突区表达。流式细胞术分析显示在高度增殖的角质形成细胞区室中有大量角蛋白15阳性细胞。值得注意的是，角蛋白15高表达细胞具有假定干细胞的表型特征。本研究首次在体外鉴定和分离了高度增殖的犬角质形成细胞，它们代表了候选上皮干细胞。
