Red Mold Rice Mitigates Oral Carcinogenesis in 7,12-Dimethyl-1,2-Benz[a]anthracene-Induced Oral Carcinogenesis in Hamster

Abstract

The prevalence of oral tumor has exponentially increased in recent years; however, the effective therapies or prevention strategies are not sufficient. Red mold rice is a traditional Chinese food, and several reports have demonstrated that red mold rice had an anti-tumor effect. However, the possible anti-tumor mechanisms of the red mold rice are unclear. In this study, we examined the anti-tumor effect of red mold rice on 7,12-dimethyl-1,2-benz[a]anthracene (DMBA)-induced oral tumor in hamster. The ethanol extract of red mold rice (RMRE) treatment significantly decreases the levels of DMBA-induced reactive oxygen species, nitro oxide and prostaglandin E(2) than those of the lovastatin-treated group (P < .001). Moreover, RMRE decreases the formation of oral tumor induced by DMBA. Monacolin K, monascin, ankaflavin or other red mold rice metabolites had been reported to decrease inflammation and oxidative stress and exerted anti-tumor effects. Therefore, we evaluated the anti-inflammation and anti-oxidative stress effects of monacolin K, monascin, ankaflavin and citrinin in lipopolysaccharide-treated RAW264.7 cells. We found that RMRE reduced the LPS-induced nitrite levels in RAW264.7 cells better than monacolin K, monascin, ankaflavin or citrinin (P < .05).

Figure 1

The flowchart of oral tumor induction and drug treatment. To induce oral tumor formation (DMBA-induced phase), 0.5% DMBA solution was prepared in mineral oil and applied to the entire mucosal surface of the right buccal pouch for 3 times/week. After 6 weeks, 6% celecoxib, RMRE (22.7 mg kg⁻¹) or lovastatin (0.15 mg kg⁻¹) was administrated with mineral oil to DMBA-induced oral tumor hamsters (experiment phase).

Figure 2

RMRE decreased ROS, NO₂ ⁻NO₃ ⁻ and PGE₂ levels in experimental hamsters. The homogenates of buccal pouch was measured levels of ROS (a), NO₂ ⁻/NO₃ ⁻ (b) and PGE₂ (c). Results are expressed as the mean ± SD, n = 5. *P < .05, **P < .01, ***P < .001 versus the DMBA treatment group.

Figure 3

The effects of RMRE on LPS-induced nitrite and PGE₂ generation in RAW264.7 cells. Various concentrations of RMRE affected the LPS-induced levels of nitrite (a) and PGE₂ (b), determined at 48 h. Results are expressed as the mean ± SD for at least three independent experiments. **P < .01, ***P < .001 versus the LPS treatment group.

Figure 4

The effects of RMRE on iNOS/COX-2 expression in RAW264.7 cells. Equal amounts of total proteins (80 μg per lane) were subjected to 10% SDS-PAGE and expression of iNOS, COX-2 and β-actin was detected by western blotting analysis. β-actin was used as an internal control. Results are expressed as the mean ± SD for at least three independent experiments. Treatment of RMRE for 24 h decreased LPS-induced iNOS expression. **P < .01, ***P < .001 versus the LPS treatment group.

Figure 5

Inhibitory effects of bioactive compounds on LPS-induced nitrite and PGE₂ generation in RAW264.7 cells. Treatment of various concentrations of different compounds (lovastatin: MK, monascin: MS, ankaflavin: AK and citrinin: CT) for 48 h decreased LPS-induced nitrite (a) and PGE₂ (b) generation. Results are expressed as the mean ± SD for at least three independent experiments. *P < .05, **P < .01, ***P < .001 versus the LPS treatment group.

Figure 6

The correlations of RMRE inhibiting ROS, NO and PGE₂ levels in DMBA-induced oral tumor model. iNOS expression induced NO generation and high NO levels might lead to oral squamous cell carcinoma development. On the other hand, COX-2 expression promoted PGE₂ synthesis, and high PGE₂ expression was increased in oral SCC tissue as compared with normal tissue. In this study, RMRE treatment had pharmacological inhibition of ROS, NO and PGE₂   in vivo. It might be a possible strategy for oral cancer prevention.