ZNF652/Sonic Hedgehog轴介导慢性N-二甲基亚硝胺诱导肝癌干细胞及莱菔硫烷干预作用的机制研究

肝癌干细胞在肝癌的发生、发展过程中起决定性作用。亚硝基化合物是肝癌的关键膳食致癌因素。迄今从肝癌干细胞角度探讨亚硝基化合物诱发肝癌的分子机制及其干预研究尚未见报道。我们前期研究发现， N-二甲基亚硝胺慢性暴露诱导正常人肝细胞恶性转化且具有肝癌干细胞特性，此表型与重要转录因子ZNF652下调及关键调控信号Sonic Hedgehog通路异常激活有关；莱菔硫烷明显抑制恶转的肝癌干细胞活性。然ZNF652 /Sonic Hedgehog是否真正调控慢性N-二甲基亚硝胺诱导肝癌干细胞的活性及莱菔硫烷的干预效应尚属未知。本课题采用体外细胞模型和动物模型相结合的方法，创新性地探讨ZNF652 /Sonic Hedgehog轴调控N-二甲基亚硝胺诱导肝癌干细胞的分子机制，并在此基础上研究莱菔硫烷的干预作用。旨在从肝癌干细胞角度探索亚硝基化合物诱发肝癌的作用机制及其靶向干预提供新的研究策略和科学依据。

Cancer stem cells (CSCs) play central role in the formation and development of cancer. Liver cancer stem cells (LCSCs) are considered to be responsible for the development of liver cancer. N-Nitrosodimethylamine (NDMA) has drawn the most attention due to its high lifetime cancer risk level and its frequent detection in foods，which is the definite risk factor of liver cancer. It is not clear that the function of ZNF652, a key transcription factor, in regulating Sonic Hedgehog signal pathway and the LCSCs induced by NDMA. The interventional effects of sulforaphane (SFN) on NDMA-induced LCSCs also remain to be explored. In our preliminary studies, human liver cell (LO2) underwent malignant transformation with chronic exposure to NDMA, from which we successfully isolated and enriched NDMA-induced LCSCs. We found that NDMA-induced LCSCs downregulated ZNF652 expression, increased Sonic Hedgehog expression level as well as its target LCSCs-related genes. ZNF652 transcriptionally repressed Gli1 and Gli2 expression, the key regulators of Sonic Hedgehog pathway. We also showed that SFN reduced the sphere formation of lung cancer cells cultured in serum-free-medium, decreased LCSCs markers’ expression, increased ZNF652 expression, and suppressed Sonic Hedgehog expression. However, the role and regulation of ZNF652/Sonic Hedgehog axis in NDMA-induced LCSCs activity and SFN intervention is totally unknown. Using serum-free-medium cultured lung CSCs and nude mouse xenograft models, the proposed research will for the first time in the field to investigate the role of ZNF652 in Sonic Hedgehog inactivation, target gene expression and NDMA-induced LCSCs activity. Regulation of Sonic Hedgehog signaling activity in LCSCs activity will be examined. The transcriptional regulation of ZNF652 on Sonic Hedgehog will be further explored. Moreover, this research will illustrate the role of ZNF652/Sonic Hedgehog in the interventional effects of SFN on NDMA-induced LCSCs. Findings from this research will provide important new insights into the molecular mechanisms of ZNF652/Sonic Hedgehog axis in regulating NDMA-induced LCSCs activity as well as its target intervention.