Genotoxic and Anti-Genotoxic Assessments of Fermented Houttuynia cordata Thunb. Leaf Ethanolic Extract and Its Anti-Cancer Effect in a Dual-Organ Carcinogenesis Model of Colon and Liver in Rats

The incidence of multiple-organ cancers has recently increased due to simultaneous exposure to various environmental carcinogens. Houttuynia cordata Thunb. (H. cordata) is recognized for its many health benefits, including its anti-cancer properties. The fermentation of its leaves has been shown to significantly enhance the bioflavonoid content and its bioactivities. This study aimed to evaluate the effectiveness of fermented H.cordata leaf (FHCL) extracts against combined carcinogens and investigate the underlying mechanisms. The crude ethanolic extract of FHCL was partitioned to obtain hexane- (HEX), dichloromethane- (DCM), ethyl acetate- (ETAC), butanol- (nBA), and residue fractions. The crude ethanolic extract (200-250 μg/mL) and the DCM fraction (50 μg/mL) significantly reduced NO production in RAW264.7 macrophages. In addition, the crude extract and the DCM and ETAC fractions showed anti-genotoxicity against aflatoxin B₁ and 2-amino-3,4-dimethylimidazo [4,5-f]quinoline (MeIQ) in Salmonella typhimurium assays (S9+). Despite demonstrating genotoxicity in the Salmonella mutation assay (with and without S9 activation), oral administration of the crude extract at 500 mg/kg of body weight (bw) for 40 days in rats did not induce micronucleated hepatocytes, indicating that the extract is non-genotoxic in vivo. Moreover, the crude extract significantly decreased Phase I but increased Phase II xenobiotic-metabolizing enzyme activities in the rats. Next, the anti-cancer effects of FHCL were evaluated in a dual-organ carcinogenesis model of the colon and liver in rats induced by 1,2-dimethylhydrazine (DMH) and diethylnitrosamine (DEN), respectively. The crude extract significantly reduced not only the number and size of glutathione S-transferase placental form positive foci in the liver (at doses of 100 and 500 mg/kg bw) but also the number of aberrant crypt foci in rat colons (at 500 mg/kg bw). Furthermore, FHCL significantly reduced the expression of proliferating cell nuclear antigen (PCNA) in the colon (at 100 and 500 mg/kg bw) and liver (at 500 mg/kg bw) of the treated rats. In conclusion, FHCL exhibits significant preventive properties against colon and liver cancers in this dual-organ carcinogenesis model. Its mechanisms of action may involve anti-inflammatory effects, the prevention of genotoxicity, the modulation of xenobiotic-metabolizing enzymes, and the inhibition of cancer cell proliferation. These findings support the use of FHCL as a natural supplement for preventing cancer.