The role of Mitf and Tyrp1 as survival factor on melanocytes

它被证明是因为Mitf不仅仅是酪氨酸酶、Tyrp 1和Tyrp 2的转录因子，但也是黑色素酶的存活因子。我们以前曾报道过，细胞对每个试剂盒、酪氨酸酶、Tyrp 1或Tyrp 2并不出现在神经细胞（NCC）中的主要文化中，Mitfmi甚至在Kitl和/或Edn 3中添加了良好的文化，如先前的报道，但一些Tyrp 1阳性细胞和少量的Kit-positive细胞出现时，他们用TPA和Cholera毒素进行了文化。这些结果建议Tyrp 1可能在黑色素细胞生存中扮演一个角色。To clarify the mechanisms of Mitf and Tyrp1 in maintaining melanocytes， we knocked-down the expression of Mitf or Tyrp1 in a melanoblast cell line （NCCmelb4M5： Kawa， et）。Al.，2005） and a melanocyte cell line （NCCmelan5： Ooka， et。Al.，（2001年）使用RNA干扰方法。NCCmelb 4 M5是一个工具包和DOPA负细胞线。NCCmelan 5是一个工具包和DOPA阳性细胞线。在Mitf-M或Tyrp 1 siRNA的转染中，这两个细胞线在96小时内没有存活，完全被禁用。免疫抑制显示，caspase 3-阳性细胞是在时间依赖的情况下增加的。实时PCR分析也在48小时内显示了干扰素刺激转录因子3和FasmRNA的显著增强。在对比、试剂盒和bcl-2分解在NCCCmelan 5细胞中，但不在NCmelb 4 M5细胞中。我们没有发现细胞被转染的GL 3作为负控制的任何变化。基于这些发现，我们建议Tyrp 1也是一个很好的生存因素，作为黑色素瘤及其在两个细胞中的生存机制都可以与Fathway主要相关。

It has been demonstrated that Mitf is not only a transcription factor for tyrosinase, Tyrp1 and Tyrp2 but also a survival factor for melanocytes. We previously reported that cells positive for either Kit, tyrosinase, Tyrp1 or Tyrp2 did not appear in neural crest cell (NCC) primary cultures in Mitfmi・ew mice even when Kitl and/or Edn3 were added to the cultures as well as previous reports but that some Tyrp1-positive cells and fewer numbers of Kit-positive cells appeared when they were cultured with TPA and cholera toxin. These results suggest that Tyrp1 may play a role in melanocytes survival. To clarify the mechanisms of Mitf and Tyrp1 in maintaining melanocytes, we knocked-down the expression of Mitf or Tyrp1 in a melanoblast cell line (NCCmelb4M5: Kawa, et. Al., 2005) and a melanocyte cell line (NCCmelan5: Ooka, et. Al., 2001) using the RNA interference method. NCCmelb4M5 is a Kit- and DOPA-negative cell line. NCCmelan5 is a Kit- and DOPA-positive cell line. In transfection of Mitf-M or Tyrp1 siRNA, these two cell lines did not survive and were completely abolished by 96 hrs. Immunostaining showed that caspase 3-positive cells increased in time-dependently. Real-time PCR analysis also revealed that interferon-stimulated transcription factor 3 and Fas mRNA greatly enhanced at 48 hours in the both cells. In contrast, Kit and bcl-2 decreased in NCCmelan5 cells but not in NCCmelb4M5 cells. We did not detect any change in the cells transfected GL3 as negative control. Based on these findings, we suggest that Tyrp1 also survival factor as well as Mtf for melanocyte and their survival mechanisms in both cells could mainly correlate with Fas pathway.