Protective effect of flavonoid-rich-fraction of Polygonum minus against cisplatin-induced hepatotoxicity and neurotoxicity in Sprague Dawley rats

Abstract

Cisplatin (CP) is a chemotherapeutic drug widely used for various type of cancers. Despite its effectiveness, CP produces undesirable adverse effects such as hepatotoxicity and neurotoxicity. The mechanism of CP-induced toxicities remains unclear, however few mechanisms are proposed including oxidative stress, inflammation, DNA damage and apoptosis. This study was aimed to minimize the adverse effects of CP on liver and brain by supplementing Polygonum minus (Pm) in normal rats. Pm is a local herb with potent antioxidant activity. The research project was divided into 3 phases: (I) a pilot study to determine the optimum dose and duration of CP to induce hepatotoxicity and neurotoxicity, (II) identification of phytochemicals and antioxidant activity of Pm flavonoid-rich-fraction (Pm-FRF) and (III) a protective study of Pm-FRF against CP-induced hepatotoxicity and neurotoxicity. In phase I, 72 male Sprague Dawley rats were randomly divided into four groups with 18 rats in each group i.e. control and CP groups (10, 20 and 25 mg/kg BW). All rats received single CP injection intraperitoneally (IP) while the control group received normal saline (NS). Rats were sacrificed at day 1, 3 and 5 post-CP injection. Physiological parameters, liver enzymes and histology analyses were performed. In phase II, Pm-FRF extraction, antioxidant analyses and identification of flavonoids by Liquid Chromatography Time of-Flight Mass Spectrometry (LC-TOF-MS) were performed. In phase III, 56 male Sprague Dawley rats were randomly divided into: (1) Control (distilled water, dH2O+NS) (2) CP 10 mg/kg BW (dH2O+CP) (3) Gallic acid 20 mg/kg (GA+CP) (4) Pm-FRF 400 mg/kg (Pm-FRF400+NS) (5) Pm-FRF 100 mg/kg (Pm-FRF100+CP) (6) Pm-FRF 200 mg/kg (Pm-FRF 200+CP) and (7) Pm-FRF 400 mg/kg (Pm-FRF400+CP). The groups were administered with dH2O, GA and Pm-FRF 100, 200 and 400 mg/kg via oral gavage for 14 days. At day 15, a single IP administration of NS was given to groups 1 and 4, while groups 2, 3, 5, 6 and 7 received a single CP injection IP. Physiological parameters were recorded throughout the experiment. At day 18, all rats were sacrificed. Blood was collected for liver enzymes test. Liver and brain were harvested for measurement of organ weight ratio, histological and ultrastructural changes, oxidative stress, inflammatory markers, DNA damage and apoptotic proteins and genes expression levels. In Phase I, the mortality rate, histological and liver enzymes tests showed that CP 10 mg/kg was a suitable dose to induce hepatotoxicity and neurotoxicity which were based on histology analysis and liver enzymes test. The optimum duration required to observe the toxicities was 3 days. In phase II, Pm-FRF possessed 98% total phenolic content and 96% radical scavenging activity. Twenty-two flavonoids were identified from LC-TOF-MS such as quercetin and flavone. In phase III, supplementation of Pm-FRF100 and Pm-FRF200 mg/kg showed protective effects against CP-induced macroscopic and microscopic changes, oxidative stress, inflammation, DNA damage and apoptosis. These findings could be due to the antioxidant property of Pm-FRF that able to protect liver and brain from CP. Supplementation with Pm-FRF 400 mg/kg showed detrimental effect against CP induced toxicities. These finding could be due to the prooxidant property of flavonoid that might contribute to the damaging effect in Pm-FRF400 group. In conclusion, Pm FRF supplementation should be used between 100 to 200 mg/kg to explicit its hepatoprotective and neuroprotective effects.