PARP1抑制PTX3表达在黑素瘤发病中的作用及机制研究

Melanoma, a malignancy deriving from melanocytes, has the highest mortality among skin cancers. Thus exploring its pathogenesis is of great significance to our country. Other research and our previous study have found that PARP1 plays a promoting role in melanoma cells clone formation and tumorigenesis. However, the downstream molecular and regulatory mechanism of PARP1 is still unclear. Through preliminary mRNA expression profile selection and verification, we found that PARP1 interference could promote the expression of tumor suppressor factor PTX3, which hints that PARP1 could promote melanoma pathogenesis via inhibiting the expression of PTX3. To explore the mechanism, we found PARP1 enzyme activity is not involved in the regulation of PTX3. Further analysis found suspicious PARP1 sites in promoter region of PTX3, which hint that PARP1 may combine to specific sequence of PTX3 promoter region and inhibits the transcription of PTX3 thus to mediating melanoma pathogenesis. Based on this hypothesis, we intend to prove that PTX3 as downstream molecules involved in regulation of PARP1 mediated melanomatosis in vivo and in vitro. Further, we will clarify the specific mechanism by which PARP1 mediates the PTX3 expression. The study may be essentially to clarify the new pathogenesis of melanoma and provide new ideas for the treatment of melanoma, have important theoretical and practical significance.

黑素瘤致死率高、治疗困难，探索其发病机制对我国黑素瘤的防治意义重大。结合文献及我们前期研究发现PARP1在促进黑素瘤体内成瘤及克隆形成中发挥重要作用，但其下游分子及调控机制尚不明确。通过前期mRNA表达谱芯片筛选及验证我们发现敲低PARP1可促进抑癌因子PTX3表达，提示PARP1可能通过抑制PTX3表达促进黑素瘤发病。对其机制进行探索我们发现PARP1酶活性并不参与调控PTX3表达，进一步分析发现PTX3启动子区存在可疑PARP1结合序列。提示PARP1可能通过与PTX3启动子区特定序列结合抑制其转录因子介导的PTX3表达进而促进黑素瘤发病。本项目拟通过临床及基础研究在体内外水平证实PARP1通过抑制PTX3表达促进黑素瘤发病这一现象，并进一步阐明PARP1抑制PTX3表达的具体机制。通过本项目研究有望丰富并完善黑素瘤的发病机制，并为黑素瘤的靶向治疗提供新的思路及靶点。

黑色素瘤是致死率最高的皮肤肿瘤。 PARP1在肿瘤中起致癌作用，但其在黑色素瘤中的作用机制尚不清楚。 阐明PARP1的作用机制可能为提高黑色素瘤患者生存率提供新的治疗靶点。 首先，利用TCGA数据库、GEO12391数据集以及western blot检测PARP1的表达水平及其与黑色素瘤预后的相关性。 然后，用芯片载玻片筛选PARP1干预后的差异表达基因(DEGs)。 并对DEGs进行GSEA、GO和KEGG通路富集分析。 同时，将这些DEGs与GEO59455中与PARP1相关的DEGs进行进一步比较，并对交叉基因进行相关分析，筛选PARP1的靶基因。 最后，通过Western blot和RT-qPCR检测PARP1与靶基因的关系。 CCK8法检测PARP1及其靶基因在黑色素瘤中的生物学作用。 我们发现PARP1在黑色素瘤和黑色素瘤细胞系中过表达，并且PARP1的表达上调与黑色素瘤的更高病理分期和不良预后相关。 筛选出NFATc2、ITGAX、CYP26B1、SYT17和VGF 5个基因为PARP1靶基因，证实NFATc2受PARP1调控。 CCK8进一步检测显示，下调PARP1或NFATc2的表达可抑制黑色素瘤的增殖。 综上所述，本研究表明PARP1是一种预后标志物，并通过调控NFATc2促进黑色素瘤增殖。