Resveratrol regulates antioxidant status, inhibits cytokine expression and restricts apoptosis in carbon tetrachloride induced rat hepatic injury

Abstract

Recent studies indicate the chemopreventive role of resveratrol in many animal models like ischemia, rheumatoid arthritis, human cancer, and diabetes. The present study was designed to investigate the chemopreventive potential of resveratrol in rat hepatic injury model by carbon tetrachloride. Male Wistar rats were treated with carbon tetrachloride (0.4 g/kg body weight) intraperitoneally daily for 8 weeks. Resveratrol (100 mg/kg, 200 mg/kg body weight) was given orally from first day until the last day of experiment. The investigation assesses the effect of resveratrol on morphological, oxidative status, histopathological, immunohistochemical, and apoptotic analysis in carbon tetrachloride-challenged liver tissue. The study indicated that the inflammatory cytokines TNF-α and IL-6 were profoundly expressed in experimental rats, whereas resveratrol decreases the immunopositivity of TNF-α and IL-6 and restored the altered architectural structure of challenged hepatic tissue. Resveratrol also protects liver cells by suppressing oxidative stress and apoptosis.

Figure 1

Structure of trans resveratrol.

Figure 2

Schematic representation of experimental regimen.

Figure 3

Effect of resveratrol on body weight gain during hepatic damage in rats. No significant difference in body weight was detected among the four groups.

Figure 4

Morphological examination of rat liver tissue at the end of the study. Macroscopically visible hepatic nodules were shown by arrows. Representative livers are taken from several groups: (a) Normal control (group A); (b) CCl₄ control (group B); (c) Resveratrol 100 mg/kg + CCl₄ (0.4 g/kg) (group C); (d) Resveratrol 200 mg/kg + CCl₄ (0.4 g/kg) (group D).

Figure 5

Histological profile of representative liver tissue in experimental animals. (a) Normal control rat liver (group A). Arrow 1 indicates the normal structure of central vein with radially arranged hepatocytes around it. Arrow 2 indicates normal hepatocytes (H & E X 200); (b) CCl₄ control (group B). Arrow 3 showed dilation of the central vein with few regenerative areas of hepatocytes around the central vein and distal part of liver. 4, 5 arrow showed extensive centrilobular necrosis around the central vein of liver (H & E X 200); (c) section from resveratrol (100 mg/kg) + CCl₄ (group C) arrow 6 represents the magnitude of dilation of the central vein (H & E X 200); (d) section from resveratrol (200 mg/kg) + CCl₄ ( group D) arrow 7 showed good protection of hepatocellular necrosis with a poorly dilated central vein and regular arrangement of hepatocytes around the central vein and also the distal part of the liver from the central vein along with a scattered regenerative zone of hepatocytes; very few necrotic zones were prominent showing hepatocytes maintaining nearnormal architecture (H & E X 200).

Figure 6

Immunohistochemical localisation of TNF-α in the rat liver tissue (H & E X 100). The figure shows representative staining for TNF-α (DAB) against counterstain (hematoxylin). (a) Represents normal liver (group A); (b) represents CCl₄ control liver (group B); (c) represents resveratrol (100 mg/kg) + CCl₄ (group C); (d) represents section from resveratrol (200 mg/kg) + CCl₄ (group D). The liver sections of control animals are mainly negative. Liver sections of CCl₄-exposed rats show more intensive staining.

Figure 7

Immunohistochemical localisation of IL-6 in the rat liver tissue (H & E X 100). The figure shows representative staining for TNF-α (DAB) against counterstain (hematoxylin). (a) Represents normal liver (group A); (b) represents CCl₄ control liver (group B); (c) represents resveratrol (100 mg/kg) + CCl₄ (group C); (d) represents section from resveratrol (200 mg/kg) + CCl₄ (group D). The liver sections of control animals are mainly negative. Liver sections of CCl₄ exposed rats show more intensive staining.

Figure 8

Levels of inflammatory cytokines TNF-α (a), and IL-6 (b) by ELISA in liver homogenate. Group A—Normal control; Group B—CCl₄ control; Group C—resveratrol (100 mg/kg) + CCl₄; Group D—resveratrol (200 mg/kg) + CCl₄. The values are expressed as mean ± SEM (n = 10); *P < 0.01 as compared with Group B; ^# P < 0.01 as compared to group C.

Figure 9

Apoptotic assay was performed by using TUNEL assay (H.E. X 400); brown stained cells (indicated by arrows) are undergoing apoptosis. (a) Immunostaining of rat liver from normal control (group A) showed absent of brown staining, (b) CCl₄ control (group B) liver showing more staining indicative cells undergoing apoptosis; (c) resveratrol (100 mg/kg) + CCl₄ (group C) exhibited less cells undergoing apoptosis; (d) resveratrol (200 mg/kg) + CCl₄ (group D) exhibited very few number of brown-stained nuclei.