OBSL1/IGFBP2上调肝星状细胞VEGF表达促进胰腺癌肝转移及纳米示踪与靶向阻断

Liver metastasis usually occurs in pancreatic cancer, which is related to the richness of hepatic stellate cells in the microenvironment of liver matrix，and liver metastases are rich in matrix tissue. We found that insulin-like growth factor binding protein 2 (IGFBP2) is likely to be key molecular in communication of pancreatic cancer and liver matrix by the transcriptome sequencing and bioinformatics analysis of the primary focal pancreatic cancer and liver metastases. Further experiments showed that the expression of IGFBP2 in pancreatic cancer cells was regulated by OBSL1 (Obscurin like 1), and IGFBP2 could increase VEGF expression of hepatic stellate cells to remodel microvasculars of liver metastases to promote pancreatic cancer growth and the formation of liver metastasis. This research intends to further explore the specific effect of OBSL1/IGFBP2 on increasing VEGF expression and the liver matrix microenvironment related with liver metastasis, so that we can elucidate the critical pathway of liver metastasis of pancreatic cancer. We established simulation of transgenic mouse models of pancreatic cell malignant transformation and tumor metastasis, and used our nano technology platform to realize the body tracer of liver metastasis in pancreatic cancer. By using nanoparticles conjugating the single chain antibodies of IGFBP2 and VEGF to cut off the pancreatic cancer - hepatic stellate cell contact, we explore the new strategy in the treatment for liver metastasis in pancreatic cancer.

胰腺癌易发生肝转移，与肝脏基质微环境富含星状细胞相关。我们通过胰腺癌原发灶和肝转移灶转录组测序和生物信息学分析，发现胰岛素样生长因子结合蛋白2（IGFBP2）很可能是沟通胰腺癌-肝脏基质关键分子。进一步实验显示胰腺癌细胞IGFBP2表达受OBSL1(Obscurin like 1)调控，IGFBP2可上调肝星状细胞VEGF表达，重塑肝转移微血管，促进胰腺癌生长与肝转移灶形成。本课题拟进一步研究胰腺癌OBSL1/IGFBP2上调肝星状细胞VEGF表达的具体机制、IGFBP2/VEGF在肝脏转移微环境中的作用，阐明胰腺癌肝转移关键通路。建立模拟胰腺细胞恶变进程的转基因小鼠模型，利用我们已建立的纳米技术平台实现体内示踪胰腺癌肝转移，通过联合负载抗IGFBP2和VEGF单链抗体纳米微粒切断胰腺癌-肝星状细胞联系，为寻找胰腺癌肝转移治疗新靶点提供新思路。

胰腺导管腺癌早期即可发生肝脏转移，预后极差。但是胰腺癌肝转移的相关分子生物学机制仍未明了。本课题旨在进一步阐明胰腺癌肝转移过程的潜在机制，为胰腺癌治疗提供新方向。本研究构建胰腺癌肝转移模型，通过转录组测序及生物信息学分析技术检测肝转移灶与正常肝脏组织差异基因表达情况，富集分析差异基因功能；胰腺癌细胞株与肝星状细胞共培养，检测肝星状细胞促胰腺癌细胞转移能力；转录组测序和生物信息学分析胰腺原发肿瘤组织与肝转移组织差异表达基因，并行免疫组化验证。OBSL1与IGFBP2基因表达相关性分析，实验验证OBSL1调控IGFBP2表达。细胞共培养验证胰腺癌细胞分泌IGFBP2调控肝星状细胞VEGF表达。免疫荧光技术证实肝星状细胞胞吞IGFBP2并转运入细胞核，上调VEGF表达。胰腺癌原位移植瘤技术构建胰原位腺癌模型，活体荧光及MR显影观察胰腺癌肝转移形成情况。免疫组化染色技术证实VEGF募集骨髓源性炎症细胞机制。研究结果显示：肝星状细胞与胰腺癌细胞共培养后可增强胰腺癌细胞迁移、侵袭能力，且癌细胞发生EMT相关改变；相比胰腺癌原位肿瘤组织，IGFBP2在肝转移灶高表达，且主要分布在胞浆；胰腺癌细胞存在OBSL/IGFBP2调控，但IGFBP2无法调控胰腺癌细胞自身VEGF合成和分泌；胰腺癌细胞外分泌IGFBP2作用于肝星状细胞，上调肝星状细胞VEGF表达；胰腺癌细胞分泌IGFBP2被星状细胞胞吞，并进入细胞核，调控星状细胞VEGF表达，促进胰腺癌肝转移灶形成。本项目运用转录组测序和生物信息学技术筛选胰腺癌肝转移关键细胞、基因以及因子，从体内、外实验说明胰腺癌OBSL1/IGFBP2上调肝星状细胞VEGF表达，从而改变肝脏转移微环境，促胰腺癌肝转移发生。