Study on the Effect of Alcoholic Extract of Flaxseed and Aerobic Exercise on Osteoporosis Parameters on Ovariectomized Rats

Background & Aims: Osteoporosis is a systemic skeletal disease associated with decreased bone mass and loss of bone tissue structure, resulting in increased susceptibility to fractures. Depending on the etiological factors, the disease may occur in primary (menopausal and aging) and secondary forms. One of the important etiologies of this disease is estrogen deficiency, in which estrogen deficiency is considered the main determinant of bone destruction in postmenopausal women. One of the major causes of this disease is estrogen deficiency, in which estrogen deficiency is considered the main cause of bone destruction in postmenopausal women (1, 2). The flax plant, with the scientific name of Linum usitatissimum, is an annual plant of the genus Linaceae that grows like a plant in the climatic conditions of Iran (10). Flaxseed is rich in phytoestrogenic compounds. Flaxseed phytoestrogens reduce post-menopausal complications in women by increasing sex hormone-binding globulin (SHBG) (11). Flaxseed is also a rich source of various phenols such as lignans, phenolic acids, flavonoids, phenylpropanoids, and tannins, known as a rich source of natural antioxidants (12). Regular exercise and physical activity can prevent diseases such as sarcopenia, obesity, and mitochondrial dysfunction, which are the main drivers of oxidative stress and inflammation in aging (13). It has also been suggested that exercise is a safe and effective way to prevent bone destruction in postmenopausal women (14). Given that ovariectomized rats have many similarities in the pathophysiological mechanisms of this complication with the human model, it is a suitable model for studying human osteoporosis (2, 16). On the other hand, since flaxseed has antioxidant and phytoestrogenic properties and exercise also leads to the prevention and treatment of osteoporosis complications, this study aimed to investigate the effect of aerobic exercise and flaxseed extract on osteoporosis index parameters in ovariectomized rats. Methods: The present study is an experimental intervention. For the study, 36 adult female Wistar rats with an average weight of 250 ±20 prepared from the Laboratory Animal Breeding Center of the Faculty of Veterinary Medicine, Islamic Azad University, Tabriz Branch, were used. Rats were randomly divided into 6 equal groups, including control, sham (only opening and closing the abdomen), ovariectomy (Ovx), ovariectomy + Flaxseed extract (Ovx + Flaxseed), ovariectomy + aerobic exercise (Ovx + Exercise), and ovariectomy + aerobic exercise + flaxseed extract groups (Ovx + Flaxseed + Exercise) were divided. The treatment protocol was started after ovariectomy surgery on the study groups and ten days postoperative recovery (17). The ovariectomy + Flaxseed extract and ovariectomy + aerobic exercise + flaxseed extract groups received 400 mg/kg of flaxseed extract daily by gavage. Rats in the control group and the ovariectomized group received the same volume of normal saline by gavage. In the exercise groups, aerobic exercise was performed on a treadmill for five days each week for eight weeks. The training process was incremental. Groups that did not practice were placed on the treadmill for the same amount of time as the exercise groups while the machine was off (18). After sacrificing and taking blood samples from rats, the serum was immediately separated by centrifugation (4°C) at 1500 rpm for 15 minutes. The serum was transferred to a freezer at 20 ° C and stored. Serum alkaline phosphatase (ALP) levels were measured using an automated analyzer. Serum calcium concentration was measured using the modified atomic absorption method of 180-80 Ziman using a spectrophotometric device. Serum osteocalcin was measured using a Radioimmunoassay kit, and parathormone (PTH) was measured using an ELISA kit. Data were analyzed by one-way ANOVA followed by Tukey test at the level of p<0.05 by GraphPad software. Results: Serum calcium and PTH concentrations in ovariectomy mice were significantly lower than controls (p<0.05). Also, serum concentrations of osteocalcin and ALP in mice in the ovariectomy group were significantly higher than in the control group (p<0.05). Serum concentrations of parathyroid hormone and osteocalcin decreased significantly, and alkaline phosphatase and calcium in the treated groups increased significantly compared to the ovariectomized group (p<0.05). The best results were obtained in the Ovx + Flaxseed + Exercise group. One of the sites of the effect of PTH is bone tissue, which increases the uptake of calcium and phosphorus from bone tissue (22). A study has shown that ovariectomized rats have a high aerobic capacity and are also more resistant to disease, and have a reduced response to adverse environmental conditions (22). Xiaoli et al. showed that ovariectomy significantly increased osteocalcin compared to the sham group (23). These results are consistent with the results of the present study. In another study, Qi and Zheng stated that ovariectomy in rats resulted in a significant increase in serum osteocalcin levels, which was consistent with the results of our study (24). The researchers also said that treating osteoporotic rats after ovariectomy with phytoestrogens such as genistein and silicon would improve osteocalcin factor, which is consistent with the present study (24). In the present study, decreased osteocalcin levels after ovariectomy improved after treatment with flaxseed extract due to its phytoestrogenic properties. A study by jian feng et al. showed that the serum concentration of ALP enzyme in the ovariectomy group increased significantly. ALP is produced by osteoblasts and is essential for bone mineralization, which is consistent with our study (21). In another study, Qi and Zheng stated that ovariectomy significantly increased serum ALP levels, which was in line with the results of our study. Also, treatment with phytoestrogens will improve the ALP factor, which is in line with the present study (24). In the present study, elevated ALP improved after ovariectomy after treatment with flaxseed extract. Lihui et al. found that exercise in ovariectomized rats increased bone density by reducing bone resorption and increased bone formation (26). Decreased serum concentrations of PTH and osteocalcin and increased serum concentrations of calcium and ALP in the treated groups compared to the ovariectomy group showed an improvement in the condition in the treated groups compared to the ovariectomy group. Conclusion: According to the results of the present study, it can be concluded that the combination of aerobic exercise and flaxseed extract has improved the complications of ovariectomy and estrogen deficiency in the studied rats. However, more research is needed to generalize the results in humans and to find the effective composition in the extract.