Hesperidin ameliorates cisplatin induced hepatotoxicity and attenuates oxidative damage, cell apoptosis, and inflammation in rats

Abstract

Cisplatin is one of the most widely used chemotherapeutic anti-cancer drugs that is associated with multiple systemic toxicities limiting its use. The present study aimed to evaluate the hepato-protective effect of hesperidin against cisplatin-induced toxicity. Thirty-two adult male albino rats were equally split into four groups, the first group served as control received normal saline, the second group (CIS) received a single intraperitoneal dose of cisplatin (7.5 mg/kg bw) on the 22nd day of the experiment, the third group (HES) treated once daily with hesperidin (200 mg/kg bw, orally) for 21 days, and the last group (HES + CIS) pretreated once daily with hesperidin followed by a single intraperitoneal dose of cisplatin. Twenty-four hours later, samples were collected for further investigations. CIS-intoxication resulted in a significant decrease in the erythrogram along with thrombocytopenia leukopenia, and lymphopenia. Furthermore, CIS administration significantly elevated serum activity of liver enzymes, total, and indirect bilirubin as well serum glucose, total cholesterol, and triglycerides levels, meanwhile serum total protein, and globulin levels were significantly reduced. The hepatic MDA was markedly elevated with a concomitant decline in the hepatic antioxidant enzymes and severe alterations in the hepatic tissue architecture in CIS-intoxicated rats. Additionally, CIS-induced overexpression of hepatic Bax, caspase-3, and TNF-α, with no effect on hepatic expression of IL-10. Interestingly, HES pretreatment improved the CIS-induced hemato-biochemical, molecular and histopathological alterations. In conclusion, hesperidin hepato-protective effects against CIS might be mediated by its antioxidant, anti-inflammatory, and anti-apoptotic properties.

Keywords: Apoptosis; Cisplatin; Hepatic oxidative stress; Hesperidin; Inflammation.

Fig. 1

Effect of hesperidin on the hepatic oxidative stress and antioxidant markers in cisplatin-intoxicated rats (A) MDA content, (B) CAT activity (C) GSH level (D) SOD activity. Control; normal, CIS; Cisplatin, HIS; Hesperidin, MDA; malondialdehyde, CAT; catalase, GSH; reduced glutathione, SOD; superoxide dismutase. Data are presented as mean ± standard error of the mean. Values with different letters within the same figure are significantly different (p < 0.05).

Fig. 2

Effect of hesperidin on relative gene expression of hepatic IL-10 and TNF-α in cisplatin-intoxicated rats (A) Relative IL-10 expression normalized to β-actin (B) Relative TNF- α expression normalized to β-actin in all experimental groups. Each bar represents mean of fold change ± SEM (n = 8). Bars with different superscript letters within the same figure are significantly different (p < 0.05). Control; normal, CIS; Cisplatin, HIS; Hesperidin, IL-10; inerleukin-10, TNF- α; tumor necrosis factor-alpha.

Fig. 3

Effect of hesperidin on the hepatic histopathological alterations in CIS-intoxicated rats. micrograph of hepatic tissues displays normal hepatocytes (arrow) and normal hepatic architecture (H&E staining) in (A) Normal control rats and (C) Hesperidin group. (B) The micrograph is taken from cisplatin group, displays local extensive necrosis of hepatocytes (arrow), (D) The micrograph shows focal necrosis of hepatocytes (arrow) in Hesperidin + Cisplatin group. (E) Hepatic necrosis scoring in the different experimental groups. Each bar represents mean ± SEM (n = 8). Bars with different superscript letters are significantly different (p < 0.05). (0, absent; 1, spotty necrosis; one or few necrotic hepatocytes; 2, confluent necrosis; 3, bridging necrosis). Control; normal, CIS; Cisplatin, HIS; Hesperidin.

Fig. 4

Effect of hesperidin on the immunohistochemical expression of Bax in rat livers. Representative images for the immunohistochemical expression of Bax (A-D) (IHC, DAB immunostaining, hematoxylin as counter stain, 100x). (A): liver of control rats displays negative immunostaining against Bax, (B): liver of cisplatin group shows strong immunostaining in the cytoplasm of hepatocytes against Bax, (C): liver of HES displays mild immunostaining against Bax, (D): liver of CIS + HES displays moderate immunostaining in the hepatocytes against Bax. Graph (E) counting (number of immunopositive cells per 1000 cells) for Bax in liver tissue in all groups that was done by Image J analysis. Each bar represents mean ± SEM (n = 8). Bars with different superscript letters are significantly different (p < 0.05). Control; normal, CIS; Cisplatin, HIS; Hesperidin.

Fig. 5

Effect of hesperidin on the immunohistochemical expression of Caspase-3 in rat livers. Representative images for the immunohistochemical expression of Caspase-3 (IHC, DAB immunostaining, hematoxylin as counter stain, 100x). (A): liver of Control group displays negative immunostaining against caspase-3 (B) liver of CIS group displays strong immunostaining in the cytoplasm and nucleus of hepatocytes against caspase-3, (C) liver of HES group displays mild immunostaining against caspase-3, (D) liver of HES + CIS group displays moderate immunostaining in the cytoplasm and nucleus of hepatocytes against caspase-3. (E) counting (number of immunopositive cells per 1000 cells) for Caspase-3 in liver tissue in all experimental groups that was done by Image J analysis. Each bar represents mean ± SEM (n = 8). Bars with different superscript letters are significantly different (p < 0.05). Control; normal, CIS; Cisplatin, HIS; Hesperidin.