Cancer Letters

Cancer Letters

Volume 128, Issue 2, 19 June 1998, Pages 155-160
Cancer Letters

Inhibition by an extract of Ocimum sanctum of DNA-binding activity of 7,12-dimethylbenz[a]anthracene in rat hepatocytes in vitro

https://doi.org/10.1016/S0304-3835(98)00068-8Get rights and content

Abstract

Ocimum sanctum is a traditional medicinal plant. Previous studies have shown that extracts of O. sanctum inhibit the induction of skin papillomas in mice by 7,12-dimethylbenz[a]anthracene (DMBA). In the present study, primary cultures of rat hepatocytes were treated with 0–500 μg of O. sanctum extract for 24 h and then with DMBA (10 or 50 μg) for 18 h. Cells were then harvested and their DNA was isolated and analyzed by 32P-postlabelling. A significant reduction in the levels of DMBA–DNA adducts was observed in all cultures pretreated with O. sanctum extract. This effect was more pronounced at the lower dose of DMBA (10 μg). Hepatocytes which were treated with the highest dose of extract (500 μg) showed a maximum reduction of 93% in the mean values of DMBA–DNA adducts. The viability of the cells was not adversely affected by pretreatment with extract. Our findings suggest that O. sanctum leaf extract blocks or suppresses the events associated with chemical carcinogenesis by inhibiting metabolic activation of the carcinogen.

Introduction

Earlier studies have shown that induction of drug metabolizing and conjugative enzymes is a critical determinant of the carcinogenicity of chemicals 1, 2, 3, 4. Therefore, it is postulated that the risk of tumour development in humans could be reduced by compounds that can modulate the activities of these enzymes. Several plant components including those present in our diets have been found to decrease the incidence and mortality of certain cancers in high-risk human populations 5, 6. We have reported the modulatory influence of alcoholic extract of the leaves of Ocimum sanctum on the activities of cytochrome P450, cytochrome b5 and aryl hydrocarbon hydroxylase in liver and glutathione-S-transferase (GST) and a reduced glutathione level in the liver, lung and stomach of mice; all of these enzymes and cofactors play an important role in the detoxification of carcinogens and mutagens [7]. The chemopreventive property of O. sanctum leaf extract on 7,12-dimethylbenz[a]anthracene (DMBA)-induced skin papillomagenesis in male Swiss albino mice has also been reported by us 8, 9. A significant reduction in tumour formation (average number of tumours per mouse and the cumulative number of papillomas) was observed in mice treated topically or through gastric intubation with the leaf extract 8, 9.

Studies of over a dozen polycyclic aromatic hydrocarbons have either identified or implicated bay-region diol epoxides as the ultimate carcinogenic metabolites 10, 11, 12, 13. These electrophiles react with nucleophilic sites in cells to form adducts 14, 15. It is the formation of carcinogen–DNA adducts that is thought to initiate carcinogenesis [15].

In this study, we have examined the effect of pretreating the cells with O. sanctum leaf extract on the formation of DMBA–DNA adducts in rat hepatocytes.

Section snippets

Chemicals

DMBA was purchased from Sigma (Poole, Dorset, UK). Reagents and materials for 32P-postlabelling were from sources mentioned previously [16]. An ethanolic leaf extract of O. sanctum was prepared by distilling the dried leaf powder of O. sanctum using absolute alcohol for 12×3 h at 60°C. The left-over residue was filtered and the remaining alcohol was allowed to evaporate. The thick paste obtained was used to treat the rat hepatocytes.

Isolation and treatment of rat hepatocytes

Female Fischer F-344 rats (8–10 weeks old) were used. Livers

Results

When DNA isolated from rat hepatocytes treated with DMBA was analyzed by 32P-postlabelling, a pattern of four adduct spots was observed on thin-layer chromatograms (Fig. 1A). The same pattern was observed in hepatocytes that had been treated with the O. sanctum extract prior to treatment with DMBA, although the intensities of the adduct spots were diminished, indicating that the level of adduct formation was reduced (Fig. 1B–E). However, the viability and morphology of the hepatocytes were not

Discussion

Ocimum sanctum L. (Sacred basil, green tulsi) is a traditional medicinal plant. On steam distillation its leaves yield a bright yellow oil possessing a pleasant odour characteristic of the plant. The oil is reported to possess antibacterial and antifungal properties [20]. The juice of the leaves possesses diaphoretic, antiperiodic, stimulating and expectorant properties. It is used to treat bronchitis and is applied to the skin in the treatment of ringworm and other cutaneous diseases [21]. Its

Acknowledgements

R.P. gratefully acknowledges the receipt of an ICRETT fellowship (No. 367 1996) from UICC, Geneva.

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