Jul 2026· International Immunopharmacology· Vol 187, pp.
117174
· 0 citations· 52 references
Medicine
TL;DR
By inciting mitochondrial damage, provoking ROS buildup, and decoupling the protective autophagy-Nrf2/xCT axis, PNPT1 promotes osteoclastogenesis, thereby introducing a promising immunopharmacological target for restraining pathological bone resorption.
Abstract
Background
Polynucleotide phosphorylase 1 (PNPT1) functions as a crucial mitochondrial enzyme; nevertheless, its potential genetic correlation with osteoporosis and its specific regulatory impact on osteoclastogenesis remain to be elucidated.
Methods
We executed a two-sample Mendelian randomization (MR) strategy to interrogate the causal link connecting PNPT1 expression to osteoporosis risk. For in vivo substantiation, we utilized both an ovariectomized (OVX) murine model and an adeno-associated virus (AAV)-driven overexpression system. Extensive in vitro assays employing RANKL-stimulated RAW264.7 macrophages were conducted to evaluate osteoclast differentiation, mitochondrial dynamics, autophagic flux, and intracellular oxidative stress through molecular and morphological analyses.
Results
MR evaluations pinpointed genetically predicted elevated PNPT1 expression as a potential genetic risk factor for osteoporosis. In vivo observations revealed a significant surge of PNPT1 within the osteoclast precursors of OVX subjects. In vitro, the ectopic overexpression of PNPT1 significantly enhanced osteoclastogenesis and bone degradation while simultaneously triggering severe mitochondrial depolarization alongside the accumulation of reactive oxygen species (ROS). On the contrary, targeted Pnpt1 silencing markedly suppressed osteoclast maturation. Mechanistic probes demonstrated that PNPT1 disrupted autophagic flux, marked by p62 accumulation. Notably, even with a compensatory transcriptional rise in Nrf2 mRNA, PNPT1 overexpression provoked a marked downregulation of Nrf2 and xCT proteins, suggesting a potent post-transcriptional suppression of the cellular antioxidant shield. This uncoupling invariably precipitated sub-lethal lipid peroxidation that amplifies osteoclastogenic signaling. Concordantly, AAV-mediated systemic PNPT1 amplification aggravated trabecular bone deterioration in vivo.
Conclusion
Guided by our MR findings and validated through our functional models, PNPT1 emerges as a potential genetic risk factor for osteoporosis. By inciting mitochondrial damage, provoking ROS buildup, and decoupling the protective autophagy-Nrf2/xCT axis, PNPT1 promotes osteoclastogenesis, thereby introducing a promising immunopharmacological target for restraining pathological bone resorption.
OBJECTIVES
Ponatinib is an effective tyrosine kinase inhibitor for chronic myeloid leukemia with the T315I mutation, but its clinical use is often limited by serious cardiovascular toxicity. Although mitochondrial dysfunction has been implicated in this process, the upstream stress-sensing mechanism that converts ponatinib exposure into collapse of mitochondrial quality control (MQC) remains poorly defined. We therefore investigated whether the PGAM5/VDAC1 axis mediates ponatinib-induced cardiac injury by coordinately disrupting mitophagy and the mitochondrial unfolded protein response (UPRmt).
METHODS
Cardiomyocyte-specific PGAM5 knockout mice (Pgam5_cko) and littermate controls (Pgam5_f/f) were fed a high-fat diet and then exposed to ponatinib. Cardiac function and adult cardiomyocyte contractility were assessed by echocardiography and IonOptix analysis. Single-cell RNA sequencing, in vivo genetic loss-of-function models, and HL-1 cells with stable Pgam5 knockdown were used to define the underlying mechanism. MQC status, including mitophagy and UPRmt, was evaluated by fluorescence imaging, RT-qPCR, western blotting, and biochemical assays.
RESULTS
Ponatinib markedly increased PGAM5 expression in the heart and induced contractile dysfunction, inflammatory activation, and cardiomyocyte apoptosis. These changes were substantially attenuated in Pgam5_cko mice. Mechanistically, ponatinib promoted pathological oligomerization of the outer mitochondrial membrane protein VDAC1 in a PGAM5-dependent manner. This event was accompanied by simultaneous suppression of PINK1/Parkin-related mitophagy and the UPRmt program, resulting in mitochondrial fragmentation, oxidative stress, and impaired bioenergetic function. At the functional level, loss of PGAM5 restored MQC and preserved cardiac performance under ponatinib stress. Importantly, forced VDAC1 oligomerization with arsenic trioxide largely abolished the protective effects of PGAM5 deficiency, supporting VDAC1 oligomerization as a critical downstream event in this pathway.
CONCLUSION
These findings identify the PGAM5/VDAC1 axis as a key mechanism linking ponatinib stress to coordinated failure of MQC in the heart. By simultaneously disabling mitophagy and UPRmt, this pathway drives mitochondrial dysfunction and cardiac injury. Targeting PGAM5-dependent VDAC1 oligomerization may therefore represent a potential strategy for limiting ponatinib-associated cardiotoxicity.
BACKGROUND/AIM
Osteoporosis is characterized by disrupted bone homeostasis while dysregulated autophagy contributes to impaired bone remodeling and bone loss. This study investigates the role of the RNA-binding protein polypyrimidine tract binding protein 1 (PTBP1) in osteoporosis progression through regulation of protein kinase C zeta (PRKCZ), and elucidates whether PTBP1 modulates the osteogenic-adipogenic fate commitment of bone marrow mesenchymal stem cells (BMSCs) via autophagy.
METHODS
In vitro, BMSCs were subjected to lentiviral infection and/or autophagy inhibitor 3-methyladenine (3-MA) treatment to systematically evaluate autophagic activity and osteogenic and adipogenic differentiation. RNA immunoprecipitation coupled with quantitative polymerase chain reaction (RIP-qPCR) and RNA stability assays were performed to clarify the molecular regulatory relationship between PTBP1 and PRKCZ. The molecular regulatory relationship between PTBP1 and PRKCZ was verified. In vivo, a bilateral ovariectomy (OVX)-induced osteoporotic mouse model was established. Adeno-associated virus-mediated modulation of PTBP1 and PRKCZ expression was conducted. Osteogenic-adipogenic balance and bone mass were comprehensively assessed.
RESULTS
Knockdown of PRKCZ markedly enhanced autophagic activity, promoted osteogenic differentiation, and suppressed adipogenic differentiation of BMSCs. These effects were reversed by 3-MA treatment. In OVX mice, PRKCZ expression was upregulated and was accompanied by impaired autophagy and osteogenic-adipogenic imbalance, whereas PRKCZ silencing substantially attenuated bone loss and reduced bone marrow adiposity. Mechanistically, PTBP1 increased PRKCZ expression by enhancing the stability of PRKCZ messenger RNA. Silencing PTBP1 activated autophagy via downregulation of PRKCZ, correcting the imbalance in BMSC fate commitment.
CONCLUSION
Inhibition of the PTBP1-PRKCZ axis activates autophagy to reprogram the osteogenic-adipogenic differentiation balance of BMSCs, thereby mitigating osteoporosis.
Inflammatory osteoporosis, also known as "immunoporosis," is a condition characterized by chronic inflammation and mitochondrial dysfunction, leading to impaired bone regeneration. Mitochondrial DNA (mtDNA) leakage activates the cGAS-STING pathway, triggering excessive inflammatory responses and hindering osteogenesis. Concurrently, defective mitophagy exacerbates mitochondrial damage, perpetuating a cycle of bone loss. This study investigated the role of the mtDNA-cGAS-STING axis in lipopolysaccharide (LPS)-induced bone marrow mesenchymal stem cells (BMSCs) dysfunction and inflammatory bone loss. We exposed BMSCs to LPS and assessed mitochondrial function, mtDNA release, and cGAS-STING activation using RT-qPCR, Western blotting, and immunofluorescence. We then engineered exosomes to co-deliver siRNA targeting STING and PINK1 mRNA, testing their effects on osteogenesis and mitochondrial homeostasis in vitro and in a mouse model of LPS-induced osteoporosis. LPS exposure resulted in mitochondrial damage, mtDNA leakage, and cGAS-STING activation, impairing osteogenic differentiation and increasing inflammatory cytokine expression. While STING inhibition reduced inflammatory signaling, it did not restore mitochondrial function, whereas PINK1 overexpression improved mitophagy and partially suppressed cGAS-STING activation. Dual regulation through siSTING/PINK1@Exo synergistically reduced mitochondrial ROS, restored membrane potential, promoted osteogenic marker expression, and enhanced mineralization in vitro. Dual-regulated exosomes significantly improved trabecular bone microarchitecture, reduced STING expression, and enhanced RUNX2 and OCN expression compared to single treatments through in vivo model. These results demonstrate that combining mitophagy activation with STING inhibition via engineered exosomes offers a promising therapeutic strategy for inflammatory osteoporosis by addressing both mitochondrial dysfunction and chronic inflammation.
Kaiwen Zheng, Benchi Che, Yongzhi Cui et al.· Free Radical Biology & Medic...· 0 citations
BACKGROUND
This study aimed to investigate the effects of retinoic acid receptor responder 2 (RARRES2) on osteoclast differentiation in postmenopausal osteoporosis (PMOP) and to elucidate the underlying mechanisms.
METHODS
A rat model of PMOP was induced by bilateral ovariectomy (OVX) method, and an in vitro osteoclast differentiation model was induced by treating RAW264.7 cells with nuclear factor kappa-B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF). Femoral bone structure, biomechanical properties, histology, osteoclast activity, co‑localization of RARRES2/CMKLR1 with TRAP+ osteoclasts and Perilipin‑1+ bone marrow adipocytes, gene expression, Fe2+ levels, reactive oxygen species (ROS) level, and lipid peroxidation level were detected.
RESULTS
OVX model rats exhibited significant trabecular bone loss, compromised bone biomechanical properties, increased osteoclast numbers, elevated osteoclast differentiation markers, and upregulated RARRES2 expression compared with Sham controls. Co‑localization immunofluorescence revealed that RARRES2 was predominantly co‑localized with TRAP+ osteoclasts and Perilipin‑1+ bone marrow adipocytes. In vitro, RARRES2 knockdown markedly suppressed osteoclast differentiation, as evidenced by decreased osteoclast-related markers (NFATc1, CTSK, c-Fos, and F-actin) and reduced TRAP activity. Mechanistically, RARRES2 knockdown increased ferroptosis in osteoclasts, manifesting by decreased Nrf2, GPX4, HO-1, NQO1, and FTH1, along with increased ACSL4, Fe2+ accumulation , ROS production, and lipid peroxidation. These findings were also confirmed in vivo. Activation of Nrf2 with DMF partially reversed the ferroptotic alterations and osteoclast‑inhibiting effects induced by RARRES2 knockdown both in vitro and in vivo.
CONCLUSION
RARRES2 knockdown inhibits osteoclast differentiation and alleviates PMOP progression by suppressing the activation of Nrf2/GPX4 and promoting osteoclast ferroptosis.
Yifan Wang, Jiabin Lin, Xinyao Cui et al.· Connective Tissue Research· 0 citations
Neddylation regulates diverse cellular processes, yet its role in osteoclast-mediated bone resorption is poorly understood. Here, we identify NAE1 (NEDD8 activating enzyme E1 subunit 1)-mediated neddylation as a critical regulator of postmenopausal osteoporosis and osteoclast differentiation through two distinct regulatory mechanisms. Pharmacological inhibition of Nae1 or myeloid-specific genetic ablation of Nae1 attenuated osteoclastogenesis in vitro and ameliorated ovariectomy (OVX)-induced osteoporosis in vivo without impairing osteoblast function. Mechanistically, Nae1 depletion disrupted intracellular iron metabolism, thereby suppressing ferritinophagy initiation in osteoclast precursors. Concurrently, integrated transcriptomics and affinity purification-mass spectrometry revealed ACSL3 as a direct neddylation substrate. Nae1-mediated neddylation modulates monounsaturated fatty acid (MUFA) biosynthesis, regulating the sensitivity of bone marrow-derived macrophages (BMDMs) to ferroptosis. This dual regulatory mechanism coordinately governs ferritinophagy initiation in iron metabolism and the sensitivity to ferroptosis mediated by ACSL3 neddylation, thereby critically influencing osteoclastogenesis. Clinically, serum MUFA levels positively correlated with bone mineral density (r = 0.329, p < 0.05). These findings support MLN4924, a clinical-stage NAE inhibitor, as a potential therapeutic strategy for osteoporosis and define an Nae1-ACSL3-MUFA-ferroptosis axis regulating osteoclast metabolism.Abbreviations: 4-HNE: 4-hydroxynonenal; ACP5/TRAP: acid phosphatase, tartrate resistant; ACSL3: acyl-CoA synthetase long chain family member 3; ACSL4: acyl-CoA synthetase long chain family member 4; BGLAP/OCN: bone gamma-carboxyglutamate protein; BMD: bone mineral density; BMDMs: bone marrow-derived macrophages; BV/TV: bone volume per total volume; CHX: cycloheximide; cKO: conditional knockout; co-IP: co-immunoprecipitation; CTSK: cathepsin K; DFO: deferoxamine; MDS: myelodysplastic syndrome; MUFA: monounsaturated fatty acid; NAE1: NEDD8 activating enzyme E1 subunit 1; NCOA4: nuclear receptor coactivator 4; NEDD8: NEDD8 ubiquitin like modifier; NFE2L2: NFE2 like bZIP transcription factor 2; NFATC1: nuclear factor of activated T cells 1; OC: osteoclast; OVX: ovariectomy; PUFA: polyunsaturated fatty acid; ROS: reactive oxygen species; RUNX2: RUNX family transcription factor 2; SLC40A1: solute carrier family 40 member 1; SLC7A11: solute carrier family 7 member 11; Tb.N: trabecular number; Tb.Sp: trabecular separation; Tb.Th: trabecular thickness; TFRC: transferrin receptor; TNFSF11/RANKL: TNF superfamily member 11; UBE2M: ubiquitin conjugating enzyme E2 M.
Hongwei Xie, Jiacheng Xie, Zihan Dai et al.· Autophagy· 0 citations
Osteoarthritis (OA) is driven in part by chondrocyte senescence, mitochondrial dysfunction and chronic inflammation, yet disease-modifying therapies that directly target these ageing mechanisms are lacking. Here, we identify the grape-seed-derived polyphenol procyanidin C1 (PCC1) as a senotherapeutic candidate that preserves chondrocyte mitochondrial homoeostasis and attenuates senescence via activation of the Nrf2/HO-1 axis. In tert-butyl hydroperoxide-induced senescent chondrocytes, PCC1 reduced senescence-associated β-galactosidase activity, dampened secretion of senescence-associated secretory phenotype factors, restored mitochondrial membrane potential and dynamics, limited extracellular matrix degradation and suppressed NF-κB activation. Network pharmacology and transcriptomic analyses converged on Nrf2 as a potential molecular target of PCC1, which we validated using cellular thermal shift and drug affinity responsive target stability assays. Genetic silencing of Nrf2 abrogated PCC1-mediated protection in vitro, confirming that Nrf2 is required for the anti-senescent and mitochondrial effects of PCC1. In a surgically induced anterior cruciate ligament transection model of OA, PCC1 administration reduced cartilage erosion, preserved matrix organisation and subchondral bone structure, and mitigated synovial inflammation in Nrf2-sufficient mice, whereas Nrf2 deficiency abolished these benefits. Together, these findings establish a mechanistic link between PCC1, Nrf2/HO-1 activation and chondrocyte mitochondrial homoeostasis in osteoarthritis, and support Nrf2-directed senotherapies as a regenerative strategy to slow OA progression.
Yu-Biao Zhang, Mao Chen, Wenxiang Cai et al.· Cell Death & Disease· 0 citations