BIOCHEMICAL AND PHARMACOLOGICAL EFFECTS OF Glyphaea brevis (SPRENG) IN 1,4-DINITROBENZENE-INDUCED TESTICULAR TOXICITY AND DEGENERATION IN RATS

Abstract

Environmental toxicants have been reported to induce male-related infertility. 1,4-Dinitrobenzene (1,4-DNB), widely used in dye, plastic and explosives manufacturing industries has been identified as an environmental toxicant and industrial pollutant. This research investigated the effect of Glyphaea brevis, a plant traditionally used as an aphrodisiac and in the treatment of impotence, on 1,4-DNB-induced testicular toxicity and degeneration in Wistar rats. Aqueous-methanol leaf extract of Glyphaea brevis (GB) was prepared and phytochemically screened, high performance liquid chromatography with diode-array detector (HPLC-DAD) phenolic profiling, in silico analyses as well as in vitro antioxidant activity tests. Toxicological evaluations of GB (300, 750 and 1500 mg/kg) and 1,4-DNB (25, 50, 75, 1000 and 2000 mg/kg mg/kg) were also performed and biochemical estimations and histopathological examination were carried out on testes, epididymis, liver and kidneys of rats after exposure to extract or toxicant. The effect of GB on 1,4-DNB-induced testicular toxicity and degeneration was assessed by oral administration of 300, 750 and 1500 mg/kg GB for 21 days to rats after 48 h of oral administration of 50 mg/kg 1,4-DNB with vitamin E (300 mg/kg, p.o.) serving as the reference standard. Biochemical estimations of oxidative stress markers and serum reproductive hormones in testes and epididymis, analyses of sperm characteristics, and determination of hematological indices were carried out on serum and tissues of animals after sacrifice. Histopathological examination of testes and epididymis was also performed. The mechanism of action of 1,4-DNB and GB was evaluated in TM4 Sertoli cells and TM3 Leydig cells by assaying for cell viability, apoptosis, transepithelial electrical resistance, gamma glutamyl transferase, and inhibin B hormone. Results indicated that GB contained bioactive phytochemicals and displayed good antioxidant activity in vitro. HPLC-DAD profiling revealed three major phenolic compounds in GB: ferulic acid (605.97 μg/g extract), coumaric acid (860.49 μg/g extract) and catechuic acid (216.30 μg/g extract). In silico bioactivity predictions of major compounds revealed that GB possessed antioxidant, antimutagenic and free radical scavenging effects. In addition, the major compounds in GB showed no detected risks of genotoxicity, reprotoxicity, hepatotoxicity and dermal toxicity. In the in vivo toxicological evaluation, GB significantly improved (P < 0.05) body weight gain, levels of serum reproductive hormones, antioxidant activity, sperm quality and haematological parameters in treated rats when compared to the control. However, 1,4-DNB significantly increased (P < 0.05) spermic and testicular oxidative stress and decreased antioxidant activity and sperm quality. Post-treatment of rats with GB ameliorated 1,4-DNB-induced biochemical, hormonal, hematological and morphological aberrations in plasma/reproductive tissues of rats. GB significantly increased (p< 0.05) cell viability, inhibin B hormone level and gamma glutamyl transferase activity in TM3 and TM4 cells exposed to 1,4-DNB. In conclusion, this study demonstrated that exposure of rats to 1,4-DNB induced testicular toxicity and degeneration in rats and that GB significantly improved sperm production and testicular function via its androgenic property in rats exposed to 1,4-DNB. The reproductive toxicity of 1,4-DNB and the potential therapeutic efficacy of GB in the management of male infertility were highlighted.