Identification of genes and proteins related to hair growth and analysis of alopecia inducing and preventing factor

首先，我们成功地从发根和外根鞘中直接提纯了mRNA。以正常男性头发和男型脱发患者头发组织中的mRNA为模板，通过DNA末端分析，分析了不同物种间基因表达的差异。我们的重点是角蛋白表达的差异，这被认为是影响头发质量的因素。我们还重点研究了与黑色素合成相关的基因，如酪氨酸酶(TYR)和酪氨酸酶相关蛋白1(TYRP1)，因为男性型秃发经常会观察到头发颜色的变化。TYR是这一级联反应的限速酶，而TYRP1是维持TYR功能所必需的。在DNA末端分析中，发角蛋白KA36和角蛋白25在正常头发中大量表达。头发角蛋白酸性5(KRTA5)、碱性5(KRTB5)和角蛋白15(KRT15)在脱发中大量表达，而TYR和TYRP I不表达。下一步，我们尝试使用RT-PCR来定量它们的表达。KRTA5和B5的表达大致相同。P-肌动蛋白在正常头发中的表达是正常头发的3.5倍，而在脱发头发中的表达约为正常头发的3.5倍。5.5倍。KRT15在正常头发和脱发头发中的表达分别约为1/2和1/2。分别是β-肌动蛋白的0.5倍和6.5倍。脱发组织中KRT的表达是正常头发的13倍以上。Tyr和TYRP1的表达大致相同。分别是(-肌动蛋白)的0.1倍和0.3倍。并不是所有的脱发患者都能观察到，但有些脱发患者的头发几乎不表达。即使在脱发患者中，从非秃发区域获得的头发也表现出与正常头发相似的表达模式。因此，我们认为，对这些基因的分析可以为我们提供区分男型脱发影响区域和非影响区域的方法，并适用于专利。

At first, we succeed to purify mRNA directly from hair root and outer root sheath. Using mRNA from the normal male scalp hair and the scalp hair from male-type alopecia, expression difference among mRNA species were analyzed by DNA tip analysis. We focused on difference of expression of keratin that is supposed to affect the quality of hair. We also focused on genes related to melanin synthesis like tyrosinase (TYR) and tyrosinase related protein 1 (TYRP1) because hair color changes were frequently observed in male-type alopecia. TYR is a rate-limiting enzyme of this cascade and TYRP1 is essential to maintain TYR function. On DNA tip analysis, hair keratin KA36 and keratin 25 showed abundant expression in normal hair. In alopecia pair, hair keratin acidic 5 (KRTA5) and basic 5 (KRTB5) as well as keratin 15 (KRT15) were abundantly expressed but TYR and TYRP I were not. Next, we tried to quantitate their expression using RT-PCR. Expression of KRTA5 and B5 was approx. 3.5 times as much and p-actin expression in normal hair, whereas that in alopecia hair was approx. 5.5 times. KRT15 expression in normal hair and alopecia hair was respectively approx. 0.5 times and 6.5 times as that of β-actin, viz. KRT expression in alopecia was more than 13 times higher than that in normal hair. TYR and TYRP1 expression was approx. 0.1 and 0.3 times as that of (-actin, respectively. It is not always observed in all alopecia hair but some alopecia hair showed almost null expression of them. Even in alopecia patient, hair obtained from the non-bald area displayed the expression pattern similar with normal hair. Thus, we consider that the analysis of these genes can show us the way to distinguish the area affected and non-affected by male-type alopecia and apply to the patent.