Both JNK1 and JNK2 Are Indispensable for Sensitized Extracellular Matrix Mineralization in IKKβ-Deficient Osteoblasts

Abstract

Extracellular matrix mineralization is critical for osteogenesis, and its dysregulation could result in osteoporosis and vascular calcification. IKK/NF-κB activation inhibits differentiation of osteoblasts, and reduces extracellular matrix mineralization, however the underlying mechanisms are poorly understood. In this study, we used CRISPR/Cas9 system to permanently inactivate IKKβ in preosteoblast cells and confirmed that such cells displayed dramatic increase in extracellular matrix mineralization associated with JNK phosphorylation. Such observation was also found in our study using IKKβ-deficient primary murine osteoblasts. Interestingly, we found that in Ikbkb −/− Mapk8 −/− or Ikbkb −/− Mapk9 −/− double knockout cells, the enhanced mineralization caused by IKKβ deficiency was completely abolished, and deletion of either Mapk8 or Mapk9 was sufficient to dampen c-Jun phosphorylation. In further experiments, we discovered that absence of JNK1 or JNK2 on IKKβ-deficient background resulted in highly conserved transcriptomic alteration in response to osteogenic induction. Therefore, identification of the indispensable roles of JNK1 and JNK2 in activating c-Jun and promoting osteoblast differentiation on IKKβ-deficient background provided novel insights into restoring homeostasis in extracellular matrix mineralization.

TGF-Beta Induced Key Genes of Osteogenic and Adipogenic Differentiation in Human Mesenchymal Stem Cells and MiRNA–mRNA Regulatory Networks

“…In a previous study, optimized ECM could induce stronger osteogenic effects in mesenchymal stem cells ( Freeman et al, 2019 ). In another recent study, it was reported that ECM mineralization was critical for osteogenesis, and its dysregulation could result in osteoporosis ( Hao et al, 2020 ). The results of the current study are concordant with those previous results.…”

Exploring the Potential Mechanism of Artemisinin and Its Derivatives in the Treatment of Osteoporosis Based on Network Pharmacology and Molecular Docking

“…DHA can increase the expression of CASP3 during LPS-induced osteoclastogenesis [13]. MAPK8 can regulate osteoblast autophagy and mitophagy, promoting extracellular matrix mineralization [27][28][29]. Therefore, EGFR, CASP3, and MAPK8 were focused on the next experiments exploring the anti-OP mechanism of ARS and its derivatives.…”

Roxadustat promotes osteoblast differentiation and prevents estrogen deficiency-induced bone loss by stabilizing HIF-1α and activating the Wnt/β-catenin signaling pathway

“…Then, the cells were washed with distilled water three times to eliminate nonspecific staining. To quantify extracellular matrix mineralization, the bound stain was eluted with 10% (w/v) cetylpyridinium chloride (Sigma-Aldrich, St. Louis, MO, USA) in water at room temperature for 40 min followed by dilution with the same volume of 10% (w/v) cetylpyridinium chloride [ 18 ] and then quantified by measuring absorbance at 570 nm.…”