Acanthus ilicifolius plant extract prevents DNA alterations in a transplantable Ehrlich ascites carcinoma-bearing murine model

AIM： To investigate the chemopreventive efficacy of the Indian medicinal plant Acanthus ilicifolius L Acanthaceae in a transplantable Ehrlich ascites carcinoma （EAC）- bearing murine model.METHODS： Male Swiss albino mice were divided into four groups： Group A was the untreated normal control; Group B was the EAC control mice group that received serial, intraperitoneal （ip） inoculations of rapidly proliferating 2 × 10^5 viable EAC cells in 0.2 mL of sterile phosphate buffered saline; Group C was the plant extract-treated group that received the aqueous leaf extract （ALE） of the plant at a dose of 2.5 mg/kg body weight by single ip injections, once daily for 10, 20 and 30 consecutive days following tumour inoculation （ALE control）; and Group D was the EAC ＋ ALE- treatment group. The chemopreventive potential of the ALE was evaluated in a murine model by studying various biological parameters and genotoxic markers, such as tumour cell count, mean survival of the animals, haematological indices, hepatocellular histology, immunohistochemical expression of liver metallothionein （MT） protein, sister-chromatid exchanges （SCEs）, and DNA alterations.RESULTS： Treatment of the EAC-bearing mice with the ALE significantly （P 〈 0.001） reduced viable tumour cell count by 68.34% （228.7 × 10^6 ± 0.53） when compared to EAC control mice （72.4 × 10^6 ± 0.49）, and restored body and organ weights almost to the normal values. ALE administration also increased （P 〈 0.001） mean survival of the hosts from 35 ± 3.46 d in EAC control mice to 83 ± 2.69 d in EAC ＋ ALE-treated mice. Haematological indices also showed marked improvement with administration of ALE in EAC-bearing animals. There was a significant increase in RBC count （P 〈 0.001）, hemoglobin percent （P 〈 0.001）, and haematocrit value （P 〈 0.001） from 4.3 ± 0.12, 6.4 ± 0.93, and 17.63 ± 0.72 respectively in EAC control mice to 7.1 ± 0.13, 12.1 ± 0.77, and 30.23 ± 0.57 respectively in EAC ＋ ALE-treated group, along with concurrent decrement （P 〈 0.001） in WBC count from 18.8 ± 0.54 in EAC control to 8.4 ± 0.71 in EAC ＋ ALE. Furthermore, treatment with ALE substantially improved hepatocellular architecture and no noticeable neoplastic lesions or foci of cellular alteration were observed. Daily administration of the ALE was found to limit liver MT expression, an important marker of cell proliferation with concomitant reduction in MT immunoreactivity （62.25 ± 2.58 vs 86.24 ± 5.69, P 〈 0.01）. ALE was also potentially effective in reducing （P 〈 0.001） the frequency of SCEs from 14.94 ± 2.14 in EAC control to 5.12 ± 1.16 in EAC ＋ ALE-treated group. Finally, in comparison to the EAC control, ALE was able to suppress in vivo DNA damage by abating the generations of＇tailed＇ DNA by 53.59% （98.65 ± 2.31 vs 45.06 ± 1.14, P 〈 0.001）, and DNA single-strand breaks （SSBs） by 38.53% （3.14 ± 0.31 vs 1.93 ± 0.23, P 〈 0.01） in EAC-bearing murine liver.CONCLUSION： Our data indicate that, ALE is beneficial in restoring haematological and hepatic histological profiles and in lengthening the survival of the animals against the proliferation of ascites tumour in vivo. Finally, the chemopreventive efficacy of the ALE is manifested in limiting MT expression and in preventing DNA alterations in murine liver. The promising results of this study suggest further investigation into the chemopreventive mechanisms of the medicinal plant A. ilicifolius in vivo and in vitro.

1α,25-dihydroxyvitamin D_(3) prevents DNA damage and restores antioxidant enzymes in rat hepatocarcinogenesis induced by diethylnitrosamine and promoted by phenobarbital

AIM:To investigate the chemopreventive iffects of 1α,25-dihydroxyvitamin D3 in diethylnitrosamine induced,phenobarbital promoted rat hepatocarcinogenesis.METHODS:The rats were randomly divided into 6 different groups(A-F).Group A,C and E rats received a single intraperitoneal (i.p) injection of diethylnitrosamine (DEN) at a dose of 200 mg/kg body mass in 9 g/L NaCl solution at 4 wk of age ,while group B served as normal vehicle control received normal saline once.After a brief recovery of 2 WK, all the DEN treated rats were given phenobarbital (PB) at 0.5g/L daily in the basal diet till WK 20.Group C was started 4 wk prior to DEN injection and continued thereafter till wk 20 at a dose (os) twice a week.Group E received the treatment of 1α,25-(OH)2D3 at the same dose mentioned as above for 15 wk.The rats in group D and F received 1α,25-(OH)2D3 alone as in group C without DEN injection.RESULTS:The comet assay showed statistically higher mean values for length to width ratios (L:W) of DNA mass and tailed cells (p<0.01;p<0.01 respectively) in DEN treated rats as compared to their normal controls.Continuous supplementation of 1α,25-dihydroxyvitaminD3 showed a significant (p<0.01) decrease in L:W ratio of DNA mass tailed cells.Furthermore,1α,25-(OH)2D3 supplementations elevated the super oxide dismutase (SOD) activety,hepatic malondialdehyde(MDA) levle,reduced glutathione (GSH) and glutathione S-transferase (GST) activety (p<0.01,p<0.01,p<0.05 and p<0.05 respectively).As an endpoint marker histological changes were observed to establish the chemopreventive effects of 1α,25-dihydroxyvitaminD3.CONCLUSION:Supplementations of 1α,25-(OH)2D3 has a marked protection against hepatic nodelogenesis,antioxidant enzymes and DNA damages in DEN induced rat hepatocarcinogenesis promoted by phenobarbital.