MON-314 Insulin-like Growth Factor 2 Drives Hepatocellular Carcinoma In Hepatocyte Srsf3 Ko Mice

Abstract

Background: Hepatocellular Carcinoma (HCC) is the most common liver cancer and is the second highest cause of cancer-related death worldwide. Previously we demonstrated that hepatocyte-specific deletion of serine/arginine rich splicing factor 3 (SRSF3-HKO) caused spontaneous hepatocellular carcinoma in aging with very high tumor expression of insulin-like growth factor 2 (IGF2). Elevated IGF2 has been reported in human HCC and we have also shown loss of SRSF3 occurs in human HCC, so the SRSF3HKO mouse represents a useful model for human HCC. Whether the elevated IGF2 is involved in HCC development or growth remains to be determined however. Here, we aimed to test whether IGF2 drives tumor development in the SRSF3HKO mouse by creating a double hepatocyte deletion of SRSF3 and IGF2. Methods: We generated hepatocyte specific SRSF3-IGF2-DKO mice to assess the role of IGF2 in tumor growth in the SRSF3-HKO mice over 12 months. Proliferation, apoptosis, steatosis, fibrosis and tumor phenotype were assessed by immunohistochemistry and immunoblot. We measured IGF1, Insulin, IGF2 and growth hormone levels in plasma. In addition we measured gene expression of lipid storage, oxidation, transportation, and inflammation. Results: None of the SRSF3-IGF2-DKO mice developed tumors over 12 months while all SRSF3-HKO mice developed HCC. Histology of livers showed (1) nuclear morphology in the SRSF3-IGF2-DKO mice resembling that of the SRSF3-HKO mice, (2) reduced fibrosis as indicated by decreased collagen staining in SRSF3-IGF2-DKO mice livers at 6 and 12 month of age, (3) SRSF3-IGF2-DKO mice livers have greater steatosis than SRSF3-HKO, and (4) SRSF3-IGF2-DKO mice livers have lower inflammation than SRSF3-HKO. Gene expression by qRT-PCR suggested (1) higher expression of lipid synthesis and accumulation in SRSF3-IGF2-DKO livers, (2) lower levels of inflammation in SRSF3-IGF2-DKO livers. Immunohistochemistry showed more Ki67-positive staining for proliferation and also for apoptosis (TUNEL and caspase3) in SRSF3-HKO mice compared to WT and SRSF3-IGF2-DKO mice. ELISA data showed the elevated levels of IGF1 and insulin in plasma of SRSF3-IGF2-DKO.We also confirmed over expression of IGF2 in human early liver diseases as has been shown in HCC. We furthermore showed that H19 expression was high in both SRSF3 HKO and SRSF3-IGF2-DKO mice livers suggesting altered epigenetic imprinting of the IGF2-H19 locus. Conclusion: IGF2 over expression is essential for HCC formation in SRSF3HKO mice.