No abstract

Activation of nuclear-factor-E2-related factor 2 (Nrf2) cascade can alleviate dexamethasone (DEX)-induced oxidative injury and death of human osteoblasts. A recent study has shown that phosphoglycerate kinase 1 (PGK1) inhibition/depletion will lead to Kelch-like ECH-associated protein 1 (Keap1) methylglyoxal modification, thereby activating Nrf2 signaling cascade. Here, in OB-6 osteoblastic cells and primary human osteoblasts, PGK1 silencing, by targeted shRNA, induced Nrf2 signaling cascade activation, causing Nrf2 protein stabilization and nuclear translocation, as well as increased expression of ARE-dependent genes (HO1, NQO1, and GCLC). Functional studies demonstrated that PGK1 shRNA largely attenuated DEX-induced oxidative injury and following death of OB-6 cells and primary osteoblasts. Furthermore, PGK1 knockout, by the CRISPR/Cas9 method, similarly induced Nrf2 signaling activation and protected osteoblasts from DEX. Importantly, PGK1 depletion-induced osteoblast cytoprotection against DEX was almost abolished by Nrf2 shRNA. In addition, Keap1 shRNA mimicked and nullified PGK1 shRNA-induced anti-DEX osteoblast cytoprotection. At last we show that PGK1 expression is downregulated in human necrotic femoral head tissues of DEX-taking patients, correlating with HO1 depletion. Collectively, these results show that PGK1 depletion protects human osteoblasts from DEX via activation of Keap1-Nrf2 signaling cascade.

Background/Aims: Congenital scoliosis (CS) is a result of anomalous development of vertebrae and is frequently associated with somitogenesis malformation. Although noncoding RNAs (ncRNAs) have been recently determined to be involved in the pathogenesis of CS, the competing endogenous RNA (ceRNA) regulatory networks in CS remain largely unknown. Methods: Sequencing was conducted to explore the ncRNA expression profiles in rat embryos (gestation day 9) following vitamin A deficiency (VAD) (n = 9 for the vitamin A deficiency-induced congenital scoliosis (VAD-CS) group and n = 4 for the control group). Real-time reverse transcriptase polymerase chain reaction (RT-PCR) was conducted to verify the expression levels of selected mRNAs, long noncoding RNAs (lncRNAs), circular RNAs (circRNAs), and microRNAs (miRNAs). Bioinformatics analysis was used to discover the possible relationships and functions of the ceRNAs. Results: A total of 749 mRNAs, 56 miRNAs, 685 lncRNAs, and 70 circRNAs were identified to have significantly different expression levels in the two groups. Wnt, PI3K-ATK, FoxO, EGFR, and mTOR were found to be the most significant pathways involved in VAD-CS pathogenesis. The circRNA/miRNA/mRNA and lncRNA/miRNA/mRNA networks of CS were built, and the gene expression mechanisms regulated by ncRNAs were unveiled via the ceRNA regulatory networks. Conclusion: We comprehensively identified ceRNA regulatory networks of embryonic somite development in VAD-CS as well as revealed the contribution of different ncRNA expression profiles. Our data demonstrate the association between mRNAs and ncRNAs in the pathogenic mechanism of CS.