PASS-predicted hepatoprotective activity of Caesalpinia sappan in thioacetamide-induced liver fibrosis in rats

Abstract

The antifibrotic effects of traditional medicinal herb Caesalpinia sappan (CS) extract on liver fibrosis induced by thioacetamide (TAA) and the expression of transforming growth factor β1 (TGF-β1), α-smooth muscle actin (αSMA), and proliferating cell nuclear antigen (PCNA) in rats were studied. A computer-aided prediction of antioxidant and hepatoprotective activities was primarily performed with the Prediction Activity Spectra of the Substance (PASS) Program. Liver fibrosis was induced in male Sprague Dawley rats by TAA administration (0.03% w/v) in drinking water for a period of 12 weeks. Rats were divided into seven groups: control, TAA, Silymarin (SY), and CS 300 mg/kg body weight and 100 mg/kg groups. The effect of CS on liver fibrogenesis was determined by Masson's trichrome staining, immunohistochemical analysis, and western blotting. In vivo determination of hepatic antioxidant activities, cytochrome P450 2E1 (CYP2E1), and matrix metalloproteinases (MPPS) was employed. CS treatment had significantly increased hepatic antioxidant enzymes activity in the TAA-treated rats. Liver fibrosis was greatly alleviated in rats when treated with CS extract. CS treatment was noted to normalize the expression of TGF-β1, αSMA, PCNA, MMPs, and TIMP1 proteins. PASS-predicted plant activity could efficiently guide in selecting a promising pharmaceutical lead with high accuracy and required antioxidant and hepatoprotective properties.

Figure 1

Photomicrographs of livers from the different experimental groups: (a) control group: showing normal liver architecture, (b) TAA group (hepatotoxic group): showing proliferation of bile duct and thick fibrous septa, (c) SY + TAA group: showing mild fibrous septa, (d) CS100 group: showing normal liver architecture, (e) CS100 + TAA group: showing multiple nodules with moderate fibrous septa and fewer cirrhotic nodules, (f) CS300 group: showing normal liver architecture, and (g) CS300 + TAA: showing mild fibrous septa. Masson's trichrome stain (Bar scale: 100 μm).

Figure 2

Photomicrographs showing immunohistochemistry staining of TGF-β1 of livers from the different experimental groups. Control group: showing normal liver architecture, (b) TAA group (hepatotoxic group): showing more TGF-β1 expression, (c) SY + TAA group: showing mild TGF-β1-positive hepatocytes in the liver, (d) CS100 + TAA group: showing moderate TGF-β1 expression, and (e) CS300 + TAA group: showing mild TGF-β1 expression. (Bar scale: 50 μm).

Figure 3

Photomicrographs showing immunohistochemical staining of PCNA of spleens from the different experimental groups. Control group: showing normal spleen architecture, (b) TAA group (hepatotoxic group): showing more PCNA expression (white arrows), (c) SY + TAA group: showing mild PCNA positive in the spleen, (d) CS100 + TAA group: showing moderate PCNA expression, and (e) CS300 + TAA group: showing mild PCNA expression. (Bar scale: 10 μm).

Figure 4

Western blot analysis of TGF-β1 and αSMA levels in liver tissue for all experimental groups. (1) Normal control group, (2) TAA group (hepatotoxic group), (3) SY + TAA group, (4) CS100 + TAA group, and (5) CS300 + TAA group.

Figure 5

Protein expressions from western blots in all experimental groups are quantitated using Image J program. (a) TGF-β1 density. (b) αSMA density. The data were stated as mean ± S.E.M. (n = 3). Means with different superscripts are significantly different. ^a P < 0.05 versus normal control group and ^b P < 0.05 versus TAA control group. SY stands for Silymarin (standard hepatoprotective drug).

Figure 6

Effect of CS crude extract on hepatic levels of CYP2E1 in rats at the end of 12 weeks. The data were stated as mean ± S.E.M. Means with different superscripts are significantly different. ^a P < 0.05 versus normal control group and ^b P < 0.05 versus TAA control group. SY stands for Silymarin (standard hepatoprotective drug).