PMID- 37312126 OWN - NLM STAT- MEDLINE DCOM- 20230615 LR - 20230616 IS - 1749-799X (Electronic) IS - 1749-799X (Linking) VI - 18 IP - 1 DP - 2023 Jun 13 TI - TNF-alpha activates RELA expression via TNFRSF1B to upregulate OPA1 expression and inhibit chondrogenic differentiation of human adipose stem cells. PG - 430 LID - 10.1186/s13018-023-03846-x [doi] LID - 430 AB - BACKGROUND: Tumor necrosis factor-alpha (TNF-alpha), one of the pro-inflammatory cytokines mediating the local inflammatory process in joints, inhibits cartilage formation and has a detrimental effect on stem cell-based cartilage regeneration for the treatment of osteoarthritis (OA). However, the mechanisms behind this inhibitory effect are still poorly understood. Mitochondrial morphological changes mediated by mitochondrial fusion and fission are highly plastic, are quite sensitive to environmental stimuli and play a crucial role in maintaining cell structure and function. In our study, chondrogenic differentiated human adipose stem cells (hADSCs) were exposed to TNF-alpha and the effect of TNF-alpha on the ability of hADSCs to chondrogenic differentiate and on mitochondrial fusion and fission was observed and analyzed. The aim was to investigate the role and mechanisms of mitochondrial fusion and fission regulation in the chondrogenic differentiation of hADSCs under normal conditions and under exposure to TNF-alpha. METHODS: We used flow cytometry to identify hADSCs immunophenotypes CD29, CD44, CD34, CD45, and HLA-DR. Alcian blue staining and Sirius red staining were used to observe the formation of proteoglycans and collagen during the chondrogenic differentiation of hADSCs, respectively. The mRNA and protein expression levels of the cartilage formation marker SOX9, type II collagen (COL2A1), and Aggrecan were measured by real-time fluorescent quantitative PCR (RT-qPCR) and western blot, respectively. The fluorescent probes MitoTracker(R) Red CMXRos and JC-1 were used to visualize mitochondria morphology and detect mitochondrial membrane electricity (MMP). Affymetrix PrimeView chips were used for gene expression profiling. RESULTS: The results showed that the chondrogenic differentiation of hADSCs was inhibited in the presence of TNF-alpha that optic atrophy 1 (OPA1) expression was significantly upregulated and mitochondria were prolonged and interconnected during this process. Gene microarray and RT-qPCR data showed that the presence of TNF-alpha led to increased expression of TNFalpha receptor 2 (TNFRSF1B) and RELA during chondrogenic differentiation of hADSCs. CONCLUSIONS: TNF-alpha inhibits chondrogenic differentiation of human adipose stem cells by activating RELA expression through TNFRSF1B upregulating OPA1 expression thereby increasing mitochondrial fusion. CI - (c) 2023. The Author(s). FAU - Guo, Jiajia AU - Guo J AD - Medical College of Guizhou University, Guiyang, 550025, Guizhou, China. FAU - Ye, Wang AU - Ye W AD - Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China. FAU - Wu, Xinglin AU - Wu X AD - Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China. FAU - Huang, Haifeng AU - Huang H AD - Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China. FAU - Li, Bo AU - Li B AD - Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China. FAU - Sun, Zeyu AU - Sun Z AD - Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China. FAU - Ren, Zhijing AU - Ren Z AD - Department of Clinical Laboratory, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China. rzj123276719@163.com. FAU - Yang, Zhen AU - Yang Z AD - Department of Orthopedics, Guizhou Provincial People's Hospital, Guiyang, 550002, Guizhou, China. Y1234560603@163.com. LA - eng GR - 31660265/National Natural Science Foundation of China/ GR - 31960208/National Natural Science Foundation of China/ GR - GZSYQN(2015)06/Youth Fund of Guizhou Provincial People's Hospital/ GR - Guizhou Science and Technology Platform (2017)5724/Subsidy Foundation of National Natural Science Foundation of Guizhou Provincial People's Hospital/ GR - Guizhou Science and Technology J Word (2015)2096/Science and Technology Foundation of Guizhou Province/ PT - Journal Article DEP - 20230613 PL - England TA - J Orthop Surg Res JT - Journal of orthopaedic surgery and research JID - 101265112 RN - 0 (Tumor Necrosis Factor-alpha) RN - 0 (Cytokines) RN - EC 3.6.1.- (OPA1 protein, human) RN - EC 3.6.1.- (GTP Phosphohydrolases) RN - 0 (TNFRSF1B protein, human) RN - 0 (Receptors, Tumor Necrosis Factor, Type II) RN - 0 (RELA protein, human) RN - 0 (Transcription Factor RelA) SB - IM MH - Humans MH - *Tumor Necrosis Factor-alpha/pharmacology MH - *Adipocytes MH - Cell Differentiation/genetics MH - Cytokines MH - Mitochondria MH - GTP Phosphohydrolases MH - Receptors, Tumor Necrosis Factor, Type II MH - Transcription Factor RelA PMC - PMC10262578 OTO - NOTNLM OT - Chondrogenic differentiation OT - Human adipose-derived stem cells OT - Mitochondrial fusion OT - OPA1 OT - RELA OT - TNFRSF1B COIS- We declared that there were no conflicts of interest in this paper. EDAT- 2023/06/14 01:10 MHDA- 2023/06/15 06:41 PMCR- 2023/06/13 CRDT- 2023/06/13 23:40 PHST- 2023/03/08 00:00 [received] PHST- 2023/05/09 00:00 [accepted] PHST- 2023/06/15 06:41 [medline] PHST- 2023/06/14 01:10 [pubmed] PHST- 2023/06/13 23:40 [entrez] PHST- 2023/06/13 00:00 [pmc-release] AID - 10.1186/s13018-023-03846-x [pii] AID - 3846 [pii] AID - 10.1186/s13018-023-03846-x [doi] PST - epublish SO - J Orthop Surg Res. 2023 Jun 13;18(1):430. doi: 10.1186/s13018-023-03846-x.