六味地黄方体外对人正常黑素细胞中褪黑素合成限速酶ANNAT、HIOMT活性的影响

Effects of Liu Wei Di Huang Decoction on the rate limiting enzyme ANNAT and HIOMT activity of melatonin in human normal melanocytes in vitro

目的观察六味地黄方对体外培养的人正常黑素细胞中ANNAT、HIOMT活性的影响。方法体外培养人正常黑素细胞,CCK-8法测药物法测定不同浓度的六味地黄方含药血清对黑素细胞增殖的影响,实时荧光PCR定量法测定不同浓度的六味地黄方含药血清对黑素细胞中ANNAT、HIOMT活性的影响。结果六味地黄方对于黑素细胞的增殖药物剂量的增加先促进了黑素细胞的增殖,在中剂量达到最高(P<0.05),继而高剂量组药物抑制其生长(P<0.05)。AANAT活性在低剂量组及中剂量组表达下调,与低中剂量促进黑素细胞增殖情况一致。分析结果表明,六味地黄方体外对黑素细胞增殖与AANAT活性存在相关性,且差异有统计学意义(P<0.05)。但六味地黄方含药血清对黑素细胞各用药剂量组HIOMT的mRNA含量无明显变化。结论 AANAT的活性在六味地黄方体外对黑素细胞影响的过程中可能起到限制性调节作用。

Objective To investigate the effects of Liu Wet Di Huang Decoction on the rate limiting enzyme ANNAT and HIOMT activity of melatonin in human normal melanoeytes in vitro. Methods Culture human melanocytes in vitro, CCK - 8 assay was used to assess the proliferation of melanocytes treated with Liu Wet Di Huang Decoction, real time fluorescent PCR quantitative method was employed to determine the rate limiting enzyme ANNAT and HIOMT activity of melatonin in human normal melano- cytes. Results The proliferation of melanocytes was promoted by the increase of different concentrations Liu Wet Di Huang Decoc- tion, which reached the highest in the middle dose（ P 〈 0.05 ） , and then the high dose group was inhibited（ P 〈 0.05 ）. AANAT activity in low dose group and middle dose group was down regulated, which was consistent with the low concentration of middle concentration to promote the proliferation of melanocytes. The results showed that there was a correlation between the proliferation of Liu Wet Di Huang Decoction and the activity of AANAT, and the difference was statistically significant（ P 〈 0.05 ）. Conclusion The activity of AANAT in the process of in vitro effects of the Liu Wet Di Huang Decoction on the melanocytes may play a restric- tive regulatory role.