Chaperonin 60 sustains osteoblast autophagy and counteracts glucocorticoid aggravation of osteoporosis by chaperoning RPTOR

Abstract

Glucocorticoid excess medication interrupts osteoblast homeostasis and exacerbates bone mass and microstructure loss ramping up the pathogenesis of osteoporotic disorders. Heat shock protein 60 (HSP60) is found to maintain protein function within cellular microenvironment upon encountering detrimental stress. In this study, we revealed that supraphysiological dexamethasone decreased HSP60 expression along with deregulated autophagy in osteoblasts cultures. This chaperonin is required to sustain autophagic markers Atg4, and Atg12 expression, LC3-II conversion, and autophagic puncta formation, and alleviated the glucocorticoid-induced loss of osteogenic gene expression and mineralized matrix accumulation. Regulator-associated protein of mTOR complex 1 (RPTOR) existed in HSP60 immunoprecipitate contributing to the HSP60-promoted autophagy and osteogenesis because knocking down RPTOR impaired autophagic influx and osteogenic activity. HSP60 shielded from RPTOR dysfunction by reducing the glucocorticoid-induced RPTOR de-phosphorylation, aggregation, and ubiquitination. In vivo, forced RPTOR expression attenuated the methylprednisolone-induced loss of osteoblast autophagy, bone mass, and trabecular microstructure in mice. HSP60 transgenic mice displayed increased cortical bone, mineral acquisition, and osteoblast proliferation along with higher osteogenesis of bone marrow mesenchymal cells than those of wild-type mice. HSP60 overexpression retained RPTOR signaling, sustained osteoblast autophagy, and compromised the severity of glucocorticoid-induced bone loss and sparse trabecular histopathology. Taken together, HSP60 is essential to maintain osteoblast autophagy, which facilitates mineralized matrix production. It fends off glucocorticoid-induced osteoblast apoptosis and bone loss by stabilizing RPTOR action to autophagy. This study offers a new insight into the mechanistic by which chaperonin protects against the glucocorticoid-induced osteoblast dysfunction and bone loss.

Vitamin K2 Can Rescue the Dexamethasone-Induced Downregulation of Osteoblast Autophagy and Mitophagy Thereby Restoring Osteoblast Function In Vitro and In Vivo

“…Autophagy is a highly conserved homeostatic process that plays an essential part in the regulation of osteoblast differentiation and mineralization function (Liu et al, 2013;Nollet et al, 2014;Han et al, 2018;Lian et al, 2018). It has consistently been shown as a protective mechanism in maintaining osteoblast viability by allowing cells to survive various stresses (Todde et al, 2009;White and Lowe, 2009;Gu et al, 2016;Yang et al, 2016;Zhu et al, 2019;Zhao et al, 2020).…”

“…It has consistently been shown as a protective mechanism in maintaining osteoblast viability by allowing cells to survive various stresses (Todde et al, 2009;White and Lowe, 2009;Gu et al, 2016;Yang et al, 2016;Zhu et al, 2019;Zhao et al, 2020). Furthermore, GC usage have been reported to inhibit osteoblast autophagy and this effect plays a pivotal role in the pathogenesis of GIOP (Han et al, 2018;Lian et al, 2018;Gremminger et al, 2019;Wang et al, 2019). Recently a specialized form of autophagy that specifically degrades dysfunctional or redundant mitochondria, termed mitophagy, was found to contribute to the mineralization function of osteoblasts (Pei et al, 2018a).…”

Punicalagin Regulates Apoptosis-Autophagy Switch via Modulation of Annexin A1 in Colorectal Cancer

“…The four proteins that were significantly different between the treated (PU and FPR inhibitors) and untreated control were HSP 27, HSP 60, catalase and TNF RI/ TNFRSF1A. Our results showed that both HSP 27 and HSP 60 were significantly decreased in the treated group compared to the untreated control, and it is worth taking note that the modulation of HSP 27 [22] and HSP 60 [23] are crucial for the induction of autophagy. On the other hand, catalase, which is often degraded in activation of autophagy, seemed to be upregulated in the PU and FPR inhibitors-treated groups.…”

Bromodomain Protein BRD4 Accelerates Glucocorticoid Dysregulation of Bone Mass and Marrow Adiposis by Modulating H3K9 and Foxp1

“…Cells in each well were harvested and lysed to extract total RNA using TRI reagent ® (Sigma-Aldrich), as previously described [ 24 ]. An amount of 1 μg of total RNA was reversely transcribed using ReadScript ® Two-Step cDNA Synthesis Kits (Sigma-Aldrich, St. Louis, MO, USA).…”

“…The ABI 7900 Detection System (Applied Biosystems, Waltham, MA, USA) and 2× TaqMan ® Universal PCR Master Mix, together with customized primers for PCR analysis of BRD4 (forward 5′-ACACCTGCA CCTACCAGACTC-3′; reverse, 5′-CATCGGCTACAGTCTAGGCC-3′), Runx2 (forward, 5′-CCAG CAGCACTCCATAT CTC-3′; reverse, 5′-CAGCGTCAACACCATCATTC-3′), OCN (forward, 5′-TAC AACTCACCAATC AT AAC-3′; reverse, 5′-ACCTTCATACTTCAACTACT-3′), PPARγ2 (forward: 5′-ACCATGGTTGACACAGAGATGCCA-3′; reverse: 5′-AGGAATGCGAGTGGTCTTCCATCA-3′), Foxp1 (forward, 5′-ATAGCTAACCTCGCTCGCCTA-3′; reverse, 5′-CTCTTCATACTACGATGC ATA-3′), and β-actin (forward, 5′-GACGGCCAGGTCATCACTAT-3′; reverse: 5′-CTTCTGCATCCT GTCAGCAA-3′) were used for computing threshold values (C t ) of PCR amplification, according to the manufacturer’s manuals. Fold changes of mRNA expression were calculated using 2 −ΔΔC t (ΔC t , C t of gene—C t of housekeeping gene; ΔΔC t , C t of glucocorticoid group—C t of vehicle group), as previously described [ 24 ].…”