METHANOL EXTRACT OF Parquetina nigrescens (AFZEL.) BULLOCK LEAF AND SQUALENE AMELIORATE ARSENIC TRIOXIDE-INDUCED REPRODUCTIVE TOXICITY IN MALE WISTAR RATS

Abstract

In Nigeria, the concentration of arsenic in ground water, which is the major source of drinking water, is high. Arsenic causes male reproductive dysfunction by inducing oxidative stress. Parquetina nigrescens leaf, reported to have antioxidant property, is used in African traditional medicine to improve sexual performance, which is strongly related to gonadal functions. This study was designed to investigate the effects of Methanol Extract of Parquetina nigrescens Leaf (MEPL) on gonadal functions in arsenic trioxide-treated (As2O3) male Wistar rats. Parquetina nigrescens plant was collected within the premises of the University of Ibadan and authenticated at Forestry Research Institute of Nigeria (FHI: 109785). The MEPL was obtained by soxhlet extraction and concentrated. An aliquot of MEPL was subjected to gas chromatography mass spectrometry for identification of its constituents. Analytical grade squalene was used for the study. Fifty male Wistar rats (150-180 g), divided into ten groups (n=5), were treated orally for 54 days as follows: group 1 (distilled water, 3 mL/kg), group 2 (3 mg/kg As2O3), group 3 (250 mg/kg MEPL), group 4 (500 mg/kg MEPL), group 5 (1000 mg/kg MEPL), group 6 (100 mg/kg squalene), group 7 (As2O3+250 mg/kg MEPL), group 8 (As2O3+500 mg/kg MEPL), group 9 (As2O3+1000 mg/kg MEPL) and group 10 (As2O3+100 mg/kg squalene). The animals were anaesthetised using sodium thiopental, prior to collection of blood, testes and epididymes. Epididymal sperm analysis was done using computer-aided sperm analyser; testicular malondialdehyde, glutathione peroxidase, superoxide dismutase, catalase and 17-beta hydroxysteroid dehydrogenase (17β-HSD) were assayed spectrophotometrically; serum testosterone and 8-oxo-2-deoxyguanosine (8-OHdG) were assayed using ELISA. Testicular Bax and Bcl-2 expression were assessed by immunohistochemistry. Testicular and epididymal tissues were examined microscopically. Data were analysed using ANOVA at α0.05. Twenty-four compounds including squalene (4.0%) were identified in MEPL. Sperm motility increased in As2O3+250 mg/kg MEPL, As2O3+500 mg/kg MEPL and As2O3+squalene treated groups when compared with As2O3 (91.2±3.3, 86.3±2.8, 88.1±2.1 against 70.9±4.5%). Testicular malondialdehyde (2.1±0.6; 1.6±9.5; 0.5±0.3 vs. 6.5±1.0 x10-3 nM/mg tissue) and 8– OHdG (2.1±0.1, 1.1±0.2, 2.1±0.1 against 4.6±0.7 ng/mL) decreased, while activities of glutathione peroxidase (1.4±0.0, 1.3±0.0, 1.4±0.0 vs.1.2±0.0 x10-3U/mg protein) and 17β-HSD (268.5±39.0, 268.4±40.1, 261.0±18.4 against 123.9±16.1%) increased in As2O3+250 mg/kg MEPL, As2O3+500 mg/kg MEPL and As2O3+squalene compared with As2O3. Superoxide dismutase activity decreased significantly in As2O3+500 mg/kg MEPL but increased in As2O3+squalene when compared with As2O3 (50.4±1.1, 71.1±8.60 against 50.7±9.0 U/mg) while catalase level increased significantly in As2O3+500 mg/kg MEPL compared with As2O3 (9.6±1.3, 7.1±0.2 µ/mg tissue). Testosterone level increased in As2O3+250 mg/kg MEPL and As2O3+500 mg/kg MEPL compared with As2O3 (1.0±0.1, 1.0±0.1 vs. 0.5±0.1nmol/L). Expression of Bcl-2 increased, while Bax expression reduced in As2O3+250 mg/kg MEPL, As2O3+500 mg/kg MEPL and As2O3+squalene compared with As2O3. Atrophy of seminiferous tubules, depletion of germ cell layers and absence of spermatozoa were observed in As2O3 group, but not in groups co-treated with MEPL or squalene. Methanol Extract of Parquetina nigrescens leaf and squalene ameliorated arsenic trioxide induced gonadal toxicity via the prevention of cell death and oxidative stress in male rats.