Implicating the cholecystokinin B receptor in liver stem cell oncogenesis

Introduction: Hepatocellular carcinoma (HCC) remains one of the leading causes of cancer-related deaths worldwide. Although the cellular origin of HCC remains uncertain, the self-renewal capacity of the liver after injury suggests that it may arise from liver stem cells. The cholecystokinin-B receptor (CCK-BR) is not found in normal liver tissue but becomes expressed in HCC, where it mediates tumor growth. Given the involvement of CCK-BR in proliferation, we believe that CCK-BR expression in hepatic stem cells represents a key step in hepatic carcinogenesis. Furthermore, if the CCK-BR is implicated in HCC development, treatment with the CCK receptor antagonist, proglumide, may be a viable approach for HCC prevention. Hypothesis: We hypothesize that (1) liver injury induces proliferation of CCK-BR+ hepatic stem cells; (2) this proliferation, if not repressed, has the propensity to become pro-oncogenic; and (3) proglumide treatment reduces liver injury and hepatic stem cell proliferation. Methods: C57BL/6 male mice (N=40) were randomized to four groups (N=10 each) for four weeks: control, normal chow-fed; a liver injury diet of 3,5-diethoxycarbonyl-1,4-dihydrocollidine (0.1%, DDC); normal chow and proglumide-supplemented drinking water (0.1 mg/mL); or the combination of the 0.1% DDC diet and proglumide. At necropsy, blood was collected for evaluation of transaminases. Livers were excised to analyze differentially expressed genes (DEGs) by RNAseq and RT-qPCR. Histology was used to confirm liver injury, and immunohistochemistry and flow cytometry were used to assess CD 133+ stem cells. The ability of liver cells to form spheroids and proliferate was examined in vitro with MTT assays. Results: Mice fed the DDC diet had elevated serum liver enzymes, and the effect was reduced by proglumide. Flow cytometry confirmed liver injury increases CCK-BR expression in liver stem cells, and proglumide treatment decreases this expression. Spheroids from the DDC mice exhibited increased proliferation while liver stem cells from proglumide-treated mice had a reduced proliferative capacity. DEGs of livers from the DDC-fed/proglumide-treated mice showed decreased expression of inflammatory cytokines and downregulation of tumor microenvironment genes compared to DDC-fed mice without proglumide. Conclusions: These results show that CD 133+ liver stem cells express CCK-BRs, and this expression is elevated during liver injury. The increased proliferative capacity of CCK-BR+ stem cells from liver injury mice suggests that the CCK-BR may play a role in HCC development. CCK-BR blockade with proglumide may provide a novel approach to prevent stem cell over-activation and risk for HCC. Department of Defense CA200380; American Physiological Society Summer Undergraduate Research Fellowship This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.