PMID- 32995951 OWN - NLM STAT- MEDLINE DCOM- 20210714 LR - 20210714 IS - 1432-0827 (Electronic) IS - 0171-967X (Linking) VI - 107 IP - 5 DP - 2020 Nov TI - Iron Overload-Induced Osteocyte Apoptosis Stimulates Osteoclast Differentiation Through Increasing Osteocytic RANKL Production In Vitro. PG - 499-509 LID - 10.1007/s00223-020-00735-x [doi] AB - Iron overload is closely associated with osteoporosis, the potential cellular mechanism involved in decreased osteoblast differentiation and increased osteoclast formation. However, the effect of iron overload on the biological behavior in osteocytes has not been reported. This study aims to investigate the changes of osteocytic activity, apoptosis, and its regulation on osteoclastogenesis in response to iron overload. MLO-Y4 osteocyte-like cells and primary osteocytes from mice were processed with ferric ammonium citrate (FAC) and deferoxamine (DFO), the conditioned medium (CM) was harvested and co-cultured with Raw264.7 cells and bone marrow-derived macrophages (BMDMs) to induce them to differentiate into osteoclasts. Osteocyte apoptosis, osteoclast differentiation, osteocytic gene expression and protein secretion of receptor activator of nuclear factor kappaB ligand (RANKL) and osteoprotegerin (OPG) was examined. Excessive iron has a toxic effect on MLO-Y4 osteocyte-like cells. Increased cell apoptosis in MLO-Y4 cells and primary osteocytes was induced by iron overload. The osteoclastic formation, differentiation-related gene expression, and osteoclastic bone-resorption capability were significantly increased after treated with the CM from iron overload-exposed osteocytes. Excessive iron exposure significantly promoted the gene expression and protein secretion of the RANKL in MLO-Y4 cells. Addition of RANKL-blocking antibody completely abolished the increase of osteoclast formation and bone resorption capacity induced by the CM from osteocytes exposed to excessive iron. Moreover, the pan-caspase apoptosis inhibitor, QVD (quinolyl-valyl-O-methylaspartyl-[-2,6-difluorophenoxy]-methylketone) was used to inhibit osteocyte apoptosis. The results showed osteocyte apoptosis induced by iron overload was reduced by QVD and accompanied by the decrease of soluble RANKL (sRANKL) in supernatant. The increase of osteoclast formation and bone resorption capacity induced by the CM from osteocytes exposed to excessive iron was significantly decreased by QVD. These results indicated that iron overload-induced osteocyte apoptosis is required to regulate osteoclast differentiation by increasing osteocytic RANKL production. This study, for the first time, reveals the indirect effect of iron overload on osteoclast differentiation through regulating osteocytes. FAU - Yang, Jiancheng AU - Yang J AD - Department of Spine Surgery, People's Hospital of Longhua, Affiliated Hospital of Southern Medical University, No. 38, Jinglong Construction Road, Shenzhen, 518109, Guangdong, China. AD - Key Laboratory for Space Bioscience and Biotechnology, School of Life Sciences, Northwestern Polytechnical University, 127 West Youyi Road, Beilin District, Xi'an, 710072, Shaanxi, China. FAU - Dong, Dandan AU - Dong D AD - Key Laboratory for Space Bioscience and Biotechnology, School of Life Sciences, Northwestern Polytechnical University, 127 West Youyi Road, Beilin District, Xi'an, 710072, Shaanxi, China. FAU - Luo, Xinle AU - Luo X AD - Department of Spine Surgery, People's Hospital of Longhua, Affiliated Hospital of Southern Medical University, No. 38, Jinglong Construction Road, Shenzhen, 518109, Guangdong, China. AD - The Third School of Clinical Medicine, Southern Medical University, Shenzhen, China. FAU - Zhou, Jianhua AU - Zhou J AD - Department of Spine Surgery, People's Hospital of Longhua, Affiliated Hospital of Southern Medical University, No. 38, Jinglong Construction Road, Shenzhen, 518109, Guangdong, China. FAU - Shang, Peng AU - Shang P AUID- ORCID: 0000-0001-5418-6240 AD - Key Laboratory for Space Bioscience and Biotechnology, School of Life Sciences, Northwestern Polytechnical University, 127 West Youyi Road, Beilin District, Xi'an, 710072, Shaanxi, China. shangpeng@nwpu.edu.cn. AD - Research & Development Institute of Northwestern Polytechnical University in Shenzhen, Shenzhen, China. shangpeng@nwpu.edu.cn. FAU - Zhang, Hao AU - Zhang H AD - Department of Spine Surgery, People's Hospital of Longhua, Affiliated Hospital of Southern Medical University, No. 38, Jinglong Construction Road, Shenzhen, 518109, Guangdong, China. zhanghaodoctor@hotmail.com. AD - The Third School of Clinical Medicine, Southern Medical University, Shenzhen, China. zhanghaodoctor@hotmail.com. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20200929 PL - United States TA - Calcif Tissue Int JT - Calcified tissue international JID - 7905481 RN - 0 (Osteoprotegerin) RN - 0 (RANK Ligand) RN - 0 (Tnfrsf11b protein, mouse) RN - 0 (Tnfsf11 protein, mouse) SB - IM MH - Animals MH - *Apoptosis MH - Cell Differentiation MH - *Iron Overload MH - Mice MH - Osteoclasts/*cytology MH - Osteocytes/*cytology MH - Osteoprotegerin MH - *RANK Ligand MH - RAW 264.7 Cells OTO - NOTNLM OT - Bone resorption OT - Cell apoptosis OT - Iron overload OT - Osteoclastogenesis OT - Osteocytes OT - Osteoporosis EDAT- 2020/10/01 06:00 MHDA- 2021/07/15 06:00 CRDT- 2020/09/30 06:17 PHST- 2020/05/13 00:00 [received] PHST- 2020/07/21 00:00 [accepted] PHST- 2020/10/01 06:00 [pubmed] PHST- 2021/07/15 06:00 [medline] PHST- 2020/09/30 06:17 [entrez] AID - 10.1007/s00223-020-00735-x [pii] AID - 10.1007/s00223-020-00735-x [doi] PST - ppublish SO - Calcif Tissue Int. 2020 Nov;107(5):499-509. doi: 10.1007/s00223-020-00735-x. Epub 2020 Sep 29.