EFFECTS OF Nᴳ NITRO-LARGINE METHYL ESTER ON NICOTINE-INDUCED INFERTILITY IN MALE WISTAR RATS

Abstract

Despite worldwide anti-smoking campaigns, cigarette smoking is still common. Nicotine, a major chemical constituent of cigarette has been shown to adversely affect male reproductive activities but there is paucity of information on the mode of action. In this study, the probable mechanisms underlying the harmful reproductive effects of nicotine and intervention was investigated in male rats. Sixty male Wistar rats (180-200g) were randomly assigned to six groups and treated orally for 30 days with saline (control), nicotine (0.5mg/kg, 1.0mg/kg) with or without Nᴳ Nitro-L-Arginine Methyl Ester (L-NAME, 50mg/Kg). Treated male rats were cohabited with untreated females in ratio 1:2 for fertility studies. Sperm analysis was done by microscopy. Serum Follicle Stimulating Hormone (FSH), Luteinizing Hormone (LH), testosterone and prolactin levels were measured by radioimmunoassay. Serum and testicular glutathione peroxidase (GPₓ), glutathione reductase (GSH-R), superoxide dismutase (SOD), malondialdehyde, serum concentrations of Total Cholesterol (TC), High Density Lipoprotein Cholesterol (HDL-C), triglyceride and calcium levels were estimated by spectrophotometry while Nitric Oxide (NO) was analysed by Griess reaction. Histology of the testis was done using haematoxylin and eosin stain. Sodium and potassium concentrations were determined by flame photometry while Low Density Lipoprotein Cholesterol (LDL-C) was calculated using Friedwald's formula. Data were analysed using descriptive statistics and ANOVA at p=0.05. Epididymal sperm motility and count in the control (93.0±3.6% and 108.8±4.5 x10⁶mL respectively) were significantly higher than those of the 0.5 and 1.0 mg/kg nicotine-treated groups (57.0±4.2%, 65.8±4.1 x10⁶mL and 45.0±3.6%. 50.8±4.5 x10⁶mL) respectively. The serum testosterone, FSH, LH and prolactin levels in the control group were 4.9±0.2 ng/mL, 2.0±0.1 mIU/mL, 1.7±0.1 mIU/mL and 4.2±0.3 mIU/mL respectively. Testosterone (2.2±0.2 ng/mL) and FSH (0.9±0.1 mIU/mL) levels were significantly decreased while LH (3.5±0.2 miU/mL) and prolactin (8.1±0.3 IU/mL) levels were significantly increased when nicotine (1.0 mg/kg) treated rats were compared with control. Administration of L- NAME with nicotine (1.0 mg/kg) significantly reversed nicotine-induced decrease of testosterone (4.5±0.2 ng/mL) when compared with nicotine treated group (1.0 mg/kg). There was a significant decrease in testicular GPₓ (1910.4±53.8 lU/mL), GSH-R (1071.0±57.1 IU/mL) and SOD (1.4±0.1 IU/mL) in nicotine treated group (1.0mg/kg) when compared with the controls (2489.8±60.1, 1487.6±55.0, 2.9±0.1 IU/mL) respectively. Testicular NO and malondialdehyde were significantly increased in nicotine-treated groups (37.7±2.1, 25.4±1.8 umol/L) and (45.9±3.6, 30.8±1.7 umol/L) respectively when compared with the controls (15.7±1.4, 17.9±1.5 umol/L). Total cholesterol, triglyceride, LDL-C, TC/HDL-C, LDL-C/HDL-C where significantly increased while sodium (113.2±2.6 mmol), potassium (4.4±0.6 mmol/L) and calcium (1.2± 0.2 mmol/L) levels were significantly decreased in nicotine treated groups when compared with the controls (137.2±2.0, 6.2±01, 1.9±0.5 mmol/L) respectively. Treatment with L-NAME revealed the testicular degeneration and disorganization in the cytoarchitecture that was observed in nicotine treated groups. Mating mates from nicotine treated group (1.0mg/kg) with untreated females yielded 0% fertility while L-NAME restored fertility to 70%. Nicotine-induced infertility was associated with altered hormonal profile and decreased testicular antioxidant enzymes. These effects were ameliorated by inhibiting systemic nitric oxide synthesis. Decreased serum levels of sodium, potassium and calcium and hyperlipidaemia are underlying risk factors associated with infertility during nicotine administration.