Abstract 3746: Influence of diethylnitrosamine on Mucin 13 expression in hepatocellular carcinoma
Authors: Malik Shabnam , Sikander Mohammed , Khan Parvez , Yallapu Murali M. , Jain Swatantra , Katare Deepshikha P. , Chauhan Subhash C. , Jaggi Meena 
Published: June 2022

Background: Hepatocellular carcinoma (HCC) is regarded as environmental-related cancer due to involvement of chemical carcinogen and viral components in multistage process. Accumulating studies have shown that obesity and non-alcoholic fatty liver disease, which can lead to HCC, have been linked to modern diets that are high in fat and charcoal-grilled processed meat, however, the exact processes through which diet contributes to hepato-carcinogenesis remain unclear. In the present study, we demonstrated that diethylnitrosamine (DEN), a subgroup of the N-nitroso compounds presents in food additive/preservative, tobacco smoke, and pharmaceutical products, upregulates the Mucin 13, transmembrane mucin, via miR-145 suppression accompanied by hepatocarcinogenesis events.

Methodology: Western blotting and qPCR were performed to examine the treatment effects of DEN on Mucin 13 and associated effector proteins in hepatocellular carcinoma cell (HepG2, Hep3B, C3A, and SK-HEP-1) lines. Chemically induced (DEN; 200 mg/kg bw; 2-AAF (150 mg/kg bw) animal model was used to further investigate the Mucin 13 expression. The serum AST, ALT and ALP activity were measured using kits provided by Span diagnostics. Immunohistochemistry was used to stain tumor tissue sections for Mucin 13 expression, and in situ hybridization was performed to detect miR-145 levels. Molecular docking studies were performed using Auto dock 4 package.

Results: Our study demonstrated that DEN treatment facilitates the DNA adducts formation in liver cancer cell lines as compared to vehicle controls. Moreover, DEN treatment upregulates Mucin 13 and associated effector proteins as analyzed by qPCR and Western blotting. Liver section of DEN+2-AAF-administered group showed vacuolization of hepatocytes in the centrizonal area with variation of nuclear size. Clear nuclear atypia in the adenoma was seen. The serum activities of AST, ALP and ALT in diseased animals correlated with the liver cancer progression. Additionally, Mucin 13 was also upregulated by DEN treatment in liver tissues. Intriguingly, nuclear Mucin 13 staining was very less in normal liver tissues, whereas its levels were significantly increased DEN+2-AAF induced HCC. These observations indicate that nuclear localization of Mucin 13 may contribute to the malignant transformation of hepatocytes during carcinogenesis. This revealed significant downregulation of tumor suppressors miR-145 in tumor tissues as compared to normal control. Molecular modelling analysis revealed that DEN interacts with Mucin 13 and forms a stable complex by offering numerous interactions with the residues of Mucin 13.

Conclusions: We propose that DEN modulates Mucin 13 and associated effector proteins in hepatocarcinogenesis.

Citation Format: Shabnam Malik, Mohammed Sikander, Parvez Khan, Murali M. Yallapu, Swatantra Jain, Deepshikha P. Katare, Subhash C. Chauhan, Meena Jaggi. Influence of diethylnitrosamine on Mucin 13 expression in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3746.