Hepatic Dysfunction Induced by 7, 12-Dimethylbenz(α)anthracene and Its Obviation with Erucin Using Enzymatic and Histological Changes as Indicators

<div><p>The toxicity induced by 7, 12-dimethylbenz(α)anthracene (DMBA) has been widely delineated by a number of researchers. This potent chemical damages many internal organs including liver, by inducing the production of reactive oxygen species, DNA-adduct formation and affecting the activities of phase I, II, antioxidant and serum enzymes. Glucosinolate hydrolytic products like isothiocyanates (ITCs) are well known for inhibiting the DNA-adduct formation and modulating phase I, II enzymes. Sulforaphane is ITC, currently under phase trials, is readily metabolized and inter-converted into erucin upon ingestion. We isolated erucin from <i>Eruca sativa</i> (Mill.) Thell. evaluated its hepatoprotective role in DMBA induced toxicity in male wistar rats. The rats were subjected to hepatic damage by five day regular intraperitoneal doses of DMBA. At the end of the protocol, the rats were euthanized, their blood was collected and livers were processed. The liver homogenate was analyzed for phase I (NADPH-cytochrome P450 reductase, NADH-cytochrome b5 reductase, cytochrome P450, cytochrome P420 and cytochrome b5), phase II (DT diaphorase, glutathione-S-transferase and γ-glutamyl transpeptidase) and antioxidant enzymes (superoxide dismutase, catalase, guaiacol peroxidise, ascorbate peroxidise, glutathione reductase and lactate dehydrogenase). The level of thiobarbituric acid reactive substances, lipid hydroperoxides, conjugated dienes and reduced glutathione in the liver homogenate was also analyzed. The serum was also analyzed for markers indicating hepatic damage (alkaline phosphatase, serum glutamic oxaloacetic transaminase, serum glutamic pyruvic transaminase, direct bilirubin and total bilirubin). Erucin provided significant protection against DMBA induced damage by modulating the phase I, II and antioxidant enzymes. The histological evaluation of liver tissue was also conducted, which showed the hepatoprotective role of erucin.</p></div

Comparative changes noted on histology- Normal Vs Abnormal at H&E 400 X.

<p>(<b>A</b>) Normal portal triad comprising of hepatic duct, artery and portal vein without any inflammatory infiltrate.; (<b>B</b>) Portal tracts showing mild to moderate periportal inflammation by mononuclear inflammatory cells; (<b>C</b>) Normal central vein which is surrounded by unremarkable looking hepatocytes maintaining their normal trabecular architecture and orientation; (<b>D</b>) Central vein is dilated and congested and shows loss of normal trabecular architecture along with focal necrosis.</p

Structural representation of (A) DMBA (7, 12-dimethylbenz(a)anthracene) and (B) erucin.

<p>Structural representation of (A) DMBA (7, 12-dimethylbenz(a)anthracene) and (B) erucin.</p

Treatment schedule of DMBA (7, 12-dimethylbenz(a)anthracene), erucin alone and in combination of both DMBA and erucin.

<p>Treatment schedule of DMBA (7, 12-dimethylbenz(a)anthracene), erucin alone and in combination of both DMBA and erucin.</p

Effect of DMBA and erucin alone and in combination on lipid and reduced glutathione content.

<p>Effect of DMBA and erucin alone and in combination on lipid and reduced glutathione content.</p

Effect of erucin on DMBA induced alterations in serum enzymes (A) Alkaline phosphatase, (B) Serum glutamic oxaloacetic transaminase, (C) Serum glutamic pyruvic transaminase, (D) Direct bilirubin, (E) Total bilirubin.

<p>Treatment groups with same alphabet show no significant difference, while the groups with different alphabets show significant difference at p≤0.05. Error bars indicate SEM (n = 6).</p

Depicts the graphical representation of modulation of different parameters in response to the interaction of DMBA (7, 12-dimethylbenz(a)anthracene) and erucin in the <i>in vivo</i> system.

<p>↑ -Shows increase in the activity: ↓ -Shows decrease in the activity. Yellow arrow represents general effect of reactive oxygen species. Blue arrows represents effect of DMBA and green arrows represents the effect of erucin.</p

Effect of erucin on DMBA induced alterations in hepatic phase I enzymes (A) NADH-cytochrome b5 reductase, (B) NADPH-cytochrome P450 reductase, (C) Cytochrome b5, (D) Cytochrome P450, (E) Cytochrome P420.

<p>Treatment groups with same alphabet show no significant difference, while the groups with different alphabets show significant difference at p≤0.05. Error bars indicate SEM (n = 6).</p

Effect of erucin on DMBA induced alterations in hepatic phase II enzymes (A) Glutathione-S-transferase, (B) DT-diaphorase, (C) γ-glutamyl transpeptidase.

<p>Treatment groups with same alphabet show no significant difference, while the groups with different alphabets show significant difference at p≤0.05. Error bars indicate SEM (n = 6).</p

Modified hepatic activity index grading: Necroinflammatory scores.

<p>Modified hepatic activity index grading: Necroinflammatory scores.</p