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Diethylnitrosamine-treated male mice were assigned to 4 groups: a casein-based 35% high fat ethanol liquid diet (EtOH), an EtOH diet made with soy protein isolate protein (EtOH/SOY), an EtOH liquid diet supplemented with genistein (EtOH/GEN) and a chow group. EtOH feeding, final concentration 5% (v/v), continued for 16 wks. EtOH increased incidence and multiplicity of basophilic lesions and adenomas compared to the chow group, (p < 0.05). The EtOH/SOY group had reduced adenoma progression when compared to the EtOH and EtOH/GEN group, (p < 0.05). Genistein supplementation had no protective effect. Soy feeding significantly reduced serum ALT concentrations (p < 0.05), decreased hepatic TNFα and CD-14 expression and decreased nuclear accumulation of NFκB protein in EtOH/SOY-treated mice compared to the EtOH group (p < 0.05). With respect to ceramides, high resolution MALDI-FTICR Imaging mass spectrometry revealed changes in the accumulation of long acyl chain ceramide species, in particular C18, in the EtOH group when compared to the EtOH/SOY group. Additionally, expression of acid ceramidase and sphingosine kinase 1 which degrade ceramide into sphingosine and convert sphingosine to sphingosine-1-phosphate (S1P) respectively and expression of S1P receptors S1PR2 and S1PR3 were all upregulated by EtOH and suppressed in the EtOH/SOY group, p < 0.05. EtOH feeding also increased hepatocyte proliferation and mRNA expression of β-catenin targets, including cyclin D1, MMP7 and glutamine synthase, which were reduced in the EtOH/SOY group, p < 0.05. These findings suggest that soy prevents tumorigenesis by reducing inflammation and by reducing hepatocyte proliferation through inhibition of EtOH-mediated β-catenin signaling. These mechanisms may involve blockade of sphingolipid signaling.

Alcohol is a well-established risk factor for hepatocellular carcinoma, and the mechanisms by which alcohol liver cancer is complex. It has been suggested that ethanol (EtOH) metabolism may enhance tumor progression by increasing hepatocyte proliferation. To test this hypothesis, ethanol (EtOH) feeding of male mice began 7 weeks post-injection of the chemical carcinogen diethylnitrosamine (DEN), and continued for 16 weeks, with a final EtOH concentration of 28% of total calories. As expected, EtOH increased the total number of cancerous foci and liver tumors identified in situ fixed livers from the EtOH+DEN group compared to corresponding pair-fed (PF)+DEN and chow+DEN control groups. In the EtOH+DEN group, tumor multiplicity corresponded to a 3- to 4-fold increase in proliferation and immunohistochemical staining of β-catenin in non-tumorigenic hepatocytes when compared to the PF+DEN and chow+DEN groups, p<0.05. Analysis of EtOH-treated livers from a previously published rat model of chronic liver disease revealed increases in hepatocyte proliferation accompanied by a hepatic depletion of retinol and retinoic acid stores (p<0.05), nuclear accumulation of β-catenin (p<0.05), increased cytosolic expression p-GSK3β (p<0.05), significant upregulation of soluble Wnts, Wnt2, and Wnt7a, and increased expression of several β-catenin targets involved in tumor promotion and progression, cyclin D1, c-myc, WISP1, and MMP7 (p<0.05). These data suggest that chronic EtOH consumption activates the Wnt/β-catenin signaling pathway, which increases hepatocyte proliferation thus promoting tumorigenesis following an initiating insult in the liver.