Mechanism Of Melatonin In The Treatment Of Postmenopausal Osteoporosis Through Intestinal Microbiota And Metabolites

Objective: Postmenopausal osteoporosis(PMOP)is a bone metabolism disease caused by the decline of ovarian function,mainly manifested as bone loss and bone microstructure disorders.Recent studies have found a significant correlation between intestinal flora and bone metabolism.However,the bacterial species that affect bone metabolism and their regulation mechanism are still unknown.Studies have found that the human intestine is the main place where melatonin is produced in addition to the pineal gland.The expression of melatonin and its receptors in the gastrointestinal tract is400 times that of the pineal gland.There is an intestinal bacterial ecosystem in the intestine,and it also has its own biological clock rhythm,which interacts with the host rhythm.The balance of the intestinal flora can promote the health of the host,and the imbalance of the intestinal flora may cause obesity,diabetes,tumors and other metabolic diseases.Melatonin is the main factor affecting the rhythm of the biological clock.Our previous studies have confirmed that melatonin can restore bone loss in postmenopausal osteoporosis.However,whether menopause will cause changes in the intestinal flora,and whether melatonin can treat osteoporosis through the intestinal flora still needs further research.Methods: Collect blood and stool samples from ovariectomized mice and mice in the sham operation group.16 Sr RNA high-throughput sequencing technology was used to detect the differences in the composition and abundance of the intestinal flora of the two groups of mice.UHPLC-MS technology was used to detect the serum and fecal metabolite levels of the two groups of mice,and the correlation analysis of the above results was performed.Using LC-MS to quantitatively detect 33 bile acids in the two groups of mice.The CCK-8 experiment detects the effects of differential bile acids on the proliferation of osteoblasts.Ovariectomized mice were modeled,melatonin gavage and intraperitoneal injection of melatonin receptor antagonist were performed micro-CT detection.16 Sr RNA high-throughput sequencing technology was used to detect the effect of melatonin on the composition and abundance of intestinal flora in ovariectomized mice.UHPLC-MS technology was used to detect the levels of serum and fecal metabolites in the three groups of mice,and the correlation analysis of the above results was carried out.Westernbolt and immunofluorescence staining were used to analyze the expression level of Bmal1 /Rev-erb? in the bone tissue of ovariectomized mice treated with melatonin.Results: The experimental results showed that: 1.The composition and abundance of the intestinal flora of ovariectomized mice were significantly different from those in the sham operation group,and the proportion of posterior wall bacteria of Bacteroides was significantly increased.2.Intestinal metabolites and serum metabolites of ovariectomized mice are significantly different from those of sham-operated mice,including a variety of bile acids.3.Correlation analysis of the above results found that postmenopausal osteoporosis may be significantly related to the imbalance of bile acid metabolism,and the difference in bile acids is significantly related to the same family of bacteria Eggerthellaceae.4.After quantitative detection of 33 bile acids in the two groups of mice,8 different bile acids in serum and 9 different fecal bile acids were found.All of them increased in Ovx group.5.Hydrodeoxycholic acid have a significant effect on the proliferation of osteoblasts.6.Oral melatonin has a reversal effect on bone loss in ovariectomized mice and there is a receptor-independent pathway.7.Oral melatonin can significantly reverse the imbalance of intestinal flora after ovariectomy.8.Oral melatonin significantly reversed the level of serum and intestinal bile acid metabolism.9.Oral melatonin can reverse the decrease of Bmal-1/Rev-erb? expression caused by ovariectomy.Conclusion: This study shows that the occurrence and development of osteoporosis is significantly related to the imbalance of intestinal flora and the disorder of bile acid metabolism.Oral melatonin can affect the proliferation of osteoblasts by reversing the imbalance of intestinal flora and restoring bile acid expression levels.This process is achieved through Bmal-1/Rev-erb? regulation.