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Eldecalcitol (ED-71) is a new type of vitamin D analog, and vitamin D has been reported to have therapeutic effects in infectious disease, autoimmune disease, and cancer. However, the anti-cancer effect of ED-71 remains unclear. The objective of this study was to explore the anti-cancer effect of ED-71 in human osteosarcoma cells and to identify the related mechanism. The CCK8 assay results showed that ED-71 inhibited MG-63 cell viability in dose and time dependent manners. Cloning and Transwell invasion assays showed that ED-71 inhibited clonal and invasion ability of MG-63 cells. Flow cytometry results showed ED-71 the G2/M cycle arrest rate, apoptosis, and intracellular ROS. Western blot was used to detect cleaved-caspase-3, Bax, Bcl-2, LC3-II/LC3-I, and P62 levels and the mTOR pathway. The increase of LC3-II and P62 indicated that ED-71 induced the formation of autophagosomes and inhibited autophagy flux. Furthermore, ED-71-induced apoptosis was weakened after adding 3-methyladenine and ED-71-induced early autophagy was weakened by caspase-3 inhibitor (Z-VAD-FMK), which indicated the two processes active each other in the presence of ED-71. Furthermore, N-acetylcysteine (NAC) pretreatment reversed the ED-71-treatment outcomes, including increased apoptosis and autophagy and inhibition of the PI3K/Akt/mTOR pathway. In conclusion, our results reveal that ED-71 induced G2/M arrest, apoptosis and autophagy in MG-63 cells by accumulating ROS to suppress the PI3K/Akt/mTOR signaling pathway., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Purpose: Periodontitis is a major chronic oral disease that is accelerated by activation of the NLRP3 inflammasome and the resulting pyroptosis. According to recent studies, active vitamin D and its analogs have been reported to have great anti-inflammatory effects. However, the anti-inflammatory mechanism of a newly found vitamin D analog, eldecalcitol (ED-71), is still unclear. This study investigates whether ED-71 could protect human gingival fibroblasts (HGFs) from LPS-induced pyroptosis and, if so, determine its underlying mechanism., Methods: After HGFs were treated with LPS alone or with LPS and ED-71, their viability was measured by CCK8 assay. The degrees of inflammation and pyroptosis were measured via LDH assay, H 2 O 2 assay, fluorescent staining, flow cytometry, and Western blots. Intracellular ROS, Hoechst 33,342, and PI stains were assessed with a fluorescence microscope. ROS inhibitor NAC, NLRP3 inhibitor MCC950, and Nrf2 inhibitor ML385 were added to further clarify the mechanism., Results: LPS induced cytotoxicity in HGFs, as shown by CCK8 assay. LPS also increased intracellular ROS, H 2 O 2 levels, release of LDH, and expression of the pyroptosis-related proteins NLRP3, caspase-1, and IL-1β. NAC and MCC950 reduced LPS-induced NLRP3, caspase-1, and IL-1β. Pretreatment with ED-71 effectively inhibited the LPS-induced pyroptosis and was associated with activation of the Nrf2/HO-1 signaling pathway. This beneficial effect of ED-71 was suppressed by ML385., Conclusion: This study demonstrates the therapeutic effect of ED-71 on LPS-induced NLRP3 inflammasome-dependent pyroptosis in HGFs and further reveals that ED-71 can inhibit pyroptosis by activating the Nrf2/HO-1 pathway. Our results thus suggest that ED-71 is a potential candidate for the treatment of periodontitis., Competing Interests: The authors report no conflicts of interest in this work., (© 2020 Huang et al.)

Background: The incidence of diabetic osteoporosis is increasing. This article evaluates the effect of combination treatment with the hypoglycemic drug exendin-4 (Ex-4) and the vitamin D analog eldecalcitol (ED-71) on improving diabetic osteoporosis and explores the relevant mechanism of action. Method: Micro-CT, HE staining, immunohistochemistry, qPCR and ELISA were used to evaluate the impact of Ex-4 and ED-71 on bone formation and macrophage polarization in a mouse model of diabetic osteoporosis in vivo. Immunofluorescence, flow cytometry and qPCR were used to characterize the polarization type of macrophages treated with Ex-4 and ED-71 in vitro. A co-culture system of BMSCs and macrophages was established. Subsequently, crystal violet staining, alkaline phosphatase staining and alizarin red staining were used to evaluate the migration and osteogenesis differentiation of BMSCs. Results: Ex-4 combined with ED-71 significantly reduced blood glucose levels and enhanced bone formation in mice with diabetic osteoporosis. In addition, Ex-4 synergized with ED-71 to induce the polarization of macrophages into M2 through the PI3K/AKT pathway. Macrophages treated with the combination of Ex-4 and ED-71 can significantly induce the osteogenic differentiation of BMSCs. Conclusion: Ex-4 synergized with ED-71 to reduce blood glucose levels significantly. And this combination therapy can synergistically induce osteogenic differentiation of BMSCs by promoting M2 macrophages polarization, thereby improving diabetic osteoporosis. Therefore, the combination of Ex-4 and ED-71 may be a new strategy for the treatment of diabetic osteoporosis. [ABSTRACT FROM AUTHOR]