Effets oestrogénique et anti-ostéoporotique de l’extrait éthanolique des écorces du tronc de Lannea acida A. Rich. (Anacardiaceae) : études in vitro et in vivo

Abstract

Estrogen decline is recognized as the main cause of physiological complaints associated with menopause. Due to the limits of conventional medicine, phytoestrogens, plant-derived compounds with estrogenic activity, are the subject of intensive research. L. acida A. Rich (Anacardiaceae) is a medicinal plant in the Cameroon pharmacopoeia, used to treat many pathologies such as dysmenorrhea, infertility, rheumatism and musculo-squeletic troubles. The aim of this study was to evaluate the estrogenic and antiosteoporotic effects of the ethanol extract of L. acida A. The animals used were female albinos Wistar rats (8 - 12 weeks; 200 ±10 g), who were tread by esophagi gavage. The estrogenicity of the ethanol extract of L. acida was evaluated in vivo following the 3-days uterotrophic test and in vitro following the "EScreen" method on the mammary adenocarcinoma cells MCF-7. The determination of the physical parameters (relative mass of femur, tibia and their mineral density) and biochemical parameters (calcium and inorganic phosphorus content and serum alkaline phosphatase activity) of bone remodeling. The hematological test and the histopathological analysis of the bone (tibia, femur, third lumber vertebrae (VL-3) and mandible) have permitted to evaluate the protective effects of the ethanol extract of L. acida on the model of osteoporosis induced by ovariectomy for 84 days. The effects of the ethanol extract of L acida were equally evaluated on the femur healing fracture model induced on normal and ovariectomized rat, by measuring bone mass, serum and bone alkaline phosphatase activity, calcium and inorganic phosphorus levels. The histological and radiographic analysis of femur, the determination of some oxidative stress parameters such as catalase (CAT), glutathione peroxidase activities (GPx) and total antioxidant capacity (FRAP) were equally assessed. The ability of the extracts to activate estrogen receptor α and β were evaluated in vitro on the cells lines HEK 293T. In vivo rats were treated with L. acida extract in the present of antagonist (faslodex). A phytochemical screening and study of acute oral toxicity were realized. The results obtained from the estrogenic test showed that the ethanolic extract of L. acida has estrogenic activities in vivo by the increase (p ˂ 0,05; 50 and 200 mg/kg BW) of relative uterine mass and the height of uterine (p ˂ 0,05; 200 mg/kg) and vaginal epithelia (p ˂ 0,001). In vitro, the ethanolic extract of L. acida induced significant increase of MCF-7 cells number at the concentration of 10, 100 (p ˂ 0,05) and 200 μg/mL (p ˂ 0,01). L. acida have induced the protective effects on a model of osteoporosis by preventing the decrease (p ˂ 0,05; 200 mg/kg) of the relative mass of femur, tibia as well as their mineral density. L. acida has equally prevent the augmentation of serum alkaline phosphatase activity (50 mg/kg) and reduction of calcium and inorganic phosphorus rate (p <0,01; 200 mg/kg). The hematological analysis showed the significant decrease (p ˂ 0,01) of hemoglobin rate (50 mg/kg), the percentage of lymphocyte and granulocyte (200 mg/kg) and hematocrit percentage (50, 200 mg/kg). The histological analysis of the bones (tibia, femur, VL3, Mandible) has showed that L. acida has prevented disorganization of their microarchitecture. At the end of 60-days of treatment, the L. acida extract prevented (p ˂ 0,001) on the OVX-fractured animals the decrease of GPx, FRAP and catalase activities (p ˂ 0,001;100 mg/kg BW). The extract has induced augmentation of trabecular bone thickness at the doses of 50 (p ˂ 0,05),100 and of 200 mg/kg (p ˂ 0,001), protected against femur microarchitecture disorganization, and induced total femur healing at the doses of 50, 100 and 200mg/kg. On the normal-fractured animals, the extract has equally prevented decrease of GPx (100 and 200mg/kg), FRAP (50, 100 and 200 mg/kg) and catalase (100 mg/kg) activities. These beneficial effects were due to phytoconstituents (totals polyphenols (786,75 ± 82,33) and flavonoids (250,61 ± 48,17) contains in the extract. L. acida has significantly decreased (p ˂ 0,001) the luciferase activity for the two subtypes of estrogens receptors α and β at the concentrations ranging from 0,00001 to 0,1μg/mL. No anti-estrogenic effect was observed in vivo on uterine, vaginal and mammary gland after treatment with L. acida at the dose of 50 mg/kg in the present of antagonist (faslodex). The acute oral toxicity evaluation showed that the ethanol extract of L. acida would be slightly toxic. Its DL50 was estimated higher than 2000 mg/kg BW. These results obtained showed that this extract has estrogenic and antiosteoporotic effects. These results provide the scientific basis for the use of the ethanol extract of L. acida to improve women health during menopause.