Kawakami Tamihiro, Soma Yoshinao, Kawa Yoko, Ito Masaru, Yamasaki Emiko, Watabe Hidenori, Hosaka Eri, Yajima Kenji, Ohsumi Kayoko, Mizoguchi Masako
Department of Dermatology, St Marianna University School of Medicine, Kanagawa, Japan.
J Invest Dermatol. 2002 Mar;118(3):471-8. doi: 10.1046/j.0022-202x.2001.01696.x.
Stem cell factor is essential to the migration and differentiation of melanocytes during embryogenesis based on the observation that mutations in either the stem cell factor gene, or its ligand, KIT, result in defects in coat pigmentation in mice. Stem cell factor is also required for the survival of melanocyte precursors while they are migrating towards the skin. Transforming growth factor beta1 has been implicated in the regulation of both cellular proliferation and differentiation. NCC-melb4, an immortal cloned cell line, was cloned from a mouse neural crest cell. NCC-melb4 cells provide a model to study the specific stage of differentiation and proliferation of melanocytes. They also express KIT as a melanoblast marker. Using the NCC-melb4 cell line, we investigated the effect of transforming growth factor beta1 on the differentiation and proliferation of immature melanocyte precursors. Immunohistochemically, NCC-melb4 cells showed transforming growth factor beta1 expression. The anti-transforming growth factor beta1 antibody inhibited the cell growth, and downregulated the KIT protein and mRNA expression. To investigate further the activation of autocrine transforming growth factor beta1, NCC-melb4 cells were incubated in nonexogenous transforming growth factor beta1 culture medium. KIT protein decreased with anti-transforming growth factor beta1 antibody concentration in a concentration-dependent manner. We concluded that in NCC-melb4 cells, transforming growth factor beta1 promotes melanocyte precursor proliferation in autocrine and/or paracrine regulation. We further investigated the influence of transforming growth factor beta1 in vitro using a neural crest cell primary culture system from wild-type mice. Anti-transforming growth factor beta1 antibody decreased the number of KIT positive neural crest cell. In addition, the anti-transforming growth factor beta1 antibody supplied within the wild-type neural crest explants abolished the growth of the neural crest cell. These results indicate that transforming growth factor beta1 affect melanocyte precursor proliferation and differentiation in the presence of stem cell factor/KIT in an autocrine/paracrine manner.
基于以下观察结果，干细胞因子对于胚胎发育过程中黑素细胞的迁移和分化至关重要：干细胞因子基因或其配体KIT发生突变会导致小鼠皮毛色素沉着缺陷。在黑素细胞前体向皮肤迁移的过程中，干细胞因子也是其存活所必需的。转化生长因子β1与细胞增殖和分化的调节均有关联。NCC-melb4是一种永生化克隆细胞系，从一只小鼠神经嵴细胞中克隆而来。NCC-melb4细胞为研究黑素细胞分化和增殖的特定阶段提供了一个模型。它们还表达KIT作为成黑素细胞标志物。利用NCC-melb4细胞系，我们研究了转化生长因子β1对未成熟黑素细胞前体分化和增殖的影响。免疫组织化学检测显示，NCC-melb4细胞表达转化生长因子β1。抗转化生长因子β1抗体抑制细胞生长，并下调KIT蛋白和mRNA表达。为了进一步研究自分泌转化生长因子β1的激活情况，将NCC-melb4细胞置于无外源性转化生长因子β1的培养基中培养。KIT蛋白随着抗转化生长因子β1抗体浓度的增加呈浓度依赖性降低。我们得出结论，在NCC-melb4细胞中，转化生长因子β1以自分泌和/或旁分泌调节方式促进黑素细胞前体增殖。我们进一步利用野生型小鼠的神经嵴细胞原代培养系统在体外研究了转化生长因子β1的影响。抗转化生长因子β1抗体减少了KIT阳性神经嵴细胞的数量。此外，在野生型神经嵴外植体中加入抗转化生长因子β1抗体可抑制神经嵴细胞的生长。这些结果表明，在干细胞因子/KIT存在的情况下，转化生长因子β1以自分泌/旁分泌方式影响黑素细胞前体的增殖和分化。
