HIF-2α/TFR1 mediated iron homeostasis disruption aggravates cartilage endplate degeneration through ferroptotic damage and mtDNA release: A new mechanism of intervertebral disc degeneration.

Backgroud: Iron overload is a prevalent condition in the elderly, often associated with various degenerative diseases, including intervertebral disc degeneration (IDD). Nevertheless, the mechanisms responsible for iron ion accumulation in tissues and the mechanism that regulate iron homeostasis remain unclear. Transferrin receptor-1 (TFR1) serves as the primary cellular iron gate, playing a pivotal role in controlling intracellular iron levels, however its involvement in IDD pathogenesis and the underlying mechanism remains obscure. Methods: Firstly, IDD mice model was established to determine the iron metabolism associated proteins changes during IDD progression. Then CEP chondrocytes were isolated and treated with TBHP or pro-inflammatory cytokines to mimic pathological environment, western blotting, immunofluorescence assay and tissue staining were employed to explore the underlying mechanisms. Lastly, TfR1 siRNA and Feristatin II were employed and the degeneration of IDD was examined using micro-CT and immunohistochemical analysis. Results: We found that the IDD pathological environment, characterized by oxidative stress and pro-inflammatory cytokines, could enhance iron influx by upregulating TFR1 expression in a HIF-2α dependent manner. Excessive iron accumulation not only induces chondrocytes ferroptosis and exacerbates oxidative stress, but also triggers the innate immune response mediated by c-GAS/STING, by promoting mitochondrial damage and the release of mtDNA. The inhibition of STING through siRNA or the reduction of mtDNA replication using ethidium bromide alleviated the degeneration of CEP chondrocytes induced by iron overload. Conclusion: Our study systemically explored the role of TFR1 mediated iron homeostasis in IDD and its underlying mechanisms, implying that targeting TFR1 to maintain balanced iron homeostasis could offer a promising therapeutic approach for IDD management. The translational potential of this article: Our study demonstrated the close link between iron metabolism dysfunction and IDD, indicated that targeting TfR1 may be a novel therapeutic strategy for IDD.

TfR1 mediated iron metabolism dysfunction as a potential therapeutic target for osteoarthritis.

OBJECTIVE: Transferrin receptor-1 (TfR1) plays important roles in controlling cellular iron levels, but its role in OA pathology is unknown. Herein we aim to investigate the role of TfR1 in OA progression and its underlying mechanisms. METHODS: TfR1 expression in cartilage during OA development were examined both in vivo and in vitro. Then IL-1ß was used to induce chondrocytes degeneration in vitro and TfR1 siRNA was used for observing the effect of TfR1 in modulating iron homeostasis, mitochondrial function and degrading enzymes expression. Also the inhibitor of TfR1 was exploited to analyze the protective effect of TfR1 inhibition in vivo. RESULTS: TfR1 is elevated in OA cartilage and contributes to OA inflammation condition. Excess iron not only results in oxidative stress damage and sensitizes chondrocytes to ferroptosis, but also triggers c-GAS/STING-mediated inflammation by promoting mitochondrial destruction and the release of mtDNA. Silencing TfR1 using TfR1 siRNA not only reduced iron content in chondrocytes and inhibited oxidative stress, but also facilitated the mitophagy process and suppressed mtDNA/cGAS/STING-mediated inflammation. Importantly, we also found that Ferstatin II, a novel and selective TfR1 inhibitor, could substantially suppress TfR1 activity both in vivo and in vitro and ameliorated cartilage degeneration. CONCLUSION: Our work demonstrates that TfR1 mediated iron influx plays important roles in chondrocytes degeneration and OA pathogenesis, suggesting that maintaining iron homeostasis through the targeting of TfR1 may represent a novel therapeutic strategy for the treatment of OA.

Nrf2 protects against cartilage endplate degeneration through inhibiting NCOA4­mediated ferritinophagy.

Iron overload and ferroptosis are associated with intervertebral disc degeneration (IDD); however, the mechanism underlying the regulation of iron homeostasis remains to be elucidated. Nuclear factor erythroid 2­related factor 2 (Nrf2) has been reported to regulate cellular iron homeostasis; however, its impact on IDD pathology and the underlying mechanism of action requires further investigation. In the present study, immunohistochemistry analysis of Nrf2 expression in the cartilage endplate (CEP) was conducted and it was demonstrated that Nrf2 expression was increased in the CEP at the early stages of the development of IDD, whereas it was decreased at the late stages of the development of IDD. The results of western blot analysis indicated that the inadequate activation of Nrf2 may aggravate mitochondrial dysfunction and oxidative stress, thus promoting CEP chondrocyte degeneration and calcification. It was also revealed that Nrf2 was involved in TNF­α­induced CEP chondrocyte iron metabolism dysfunction and ferroptosis. Inhibition of Nrf2 expression using Nrf2 small interfering RNA could enhance the process of nuclear receptor coactivator 4 (NCOA4)­mediated ferritinophagy and increase ferrous ion content, which may promote CEP chondrocyte ferroptotic cell death and extracellular matrix degradation. Furthermore, a decrease in cellular iron concentration may inhibit CEP chondrocyte ferroptosis, and CEP degeneration and calcification. The present study highlights the role of the Nrf2/NCOA4 axis in chondrocyte ferroptosis and IDD pathogenesis, thus suggesting that activation of Nrf2 may be a promising strategy for IDD treatment.

Activation of the Nrf-2 pathway by pinocembrin safeguards vertebral endplate chondrocytes against apoptosis and degeneration caused by oxidative stress.

AIM: The occurrence and progression of intervertebral disc degeneration (IDD) are significantly influenced by the cartilaginous endplate (CEP). Pinocembrin (PIN), a type of flavonoid present in propolis and botanicals, demonstrates both antioxidant and anti-inflammatory characteristics, which could potentially be utilized in management. Therefore, it is crucial to investigate how PIN protects against CEP degeneration and its mechanisms, offering valuable insights for IDD therapy. MATERIALS AND METHODS: To investigate the protective impact of PIN in vivo, we created the IDD mouse model through bilateral facet joint transection. In vitro, an IDD pathological environment was mimicked by applying TBHP to treat endplate chondrocytes. KEY FINDINGS: In vivo, compared with the IDD group, the mouse in the PIN group effectively mitigates IDD progression and CEP calcification. In vitro, the activation of the Nrf-2 pathway improves the process of Parkin-mediated autophagy in mitochondria and decreases ferroptosis in chondrocytes. This enhancement promotes cell survival by addressing the imbalance of redox during pathological conditions related to IDD. Knocking down Nrf-2 with siRNA fails to provide protection to endplate chondrocytes against apoptosis and degeneration. SIGNIFICANCE: The Nrf-2-mediated activation of mitochondrial autophagy and suppression of ferroptosis play a crucial role in safeguarding against oxidative stress-induced degeneration and calcification of CEP through the protective function of PIN. To sum up, this research offers detailed explanations about how PIN can protect against apoptosis and calcification in CEP, providing valuable information about the development of IDD and suggesting possible treatment approaches.

Astaxanthin suppresses oxidative stress and calcification in vertebral cartilage endplate via activating Nrf-2/HO-1 signaling pathway.

BACKGROUND: Cartilage endplate (CEP) degeneration is an important initiating factor leading to intervertebral disc degeneration (IVDD). Astaxanthin (Ast) is a natural lipid-soluble and red-orange carotenoid which possesses various biological activities, including antioxidant, anti-inflammatory, and anti-aging effects in multiple organisms. However, the effects and mechanism of Ast on endplate chondrocytes remain largely unknown. The objective of the current study was to investigate the effects and of Ast on CEP degeneration and its underlying molecular mechanisms. METHODS: Tert-butyl hydroperoxide (TBHP) was used to mimic the IVDD pathological environment. We investigated the effects of Ast on the Nrf2 signaling pathway and damage-associated events. The IVDD model was constructed by surgical resection of L4 posterior elements to explore the role of Ast in vivo. RESULTS: We found that the activation of the Nrf-2/HO-1 signaling pathway was enhanced by Ast, thus promoted mitophagy process, inhibited oxidative stress and CEP chondrocytes ferroptosis, eventually ameliorated extracellular matrix (ECM) degradation, CEP calcification and endplate chondrocytes apoptosis. Knockdown of Nrf-2 using siRNA inhibited Ast induced mitophagy process and its protective effect. Moreover, Ast inhibited oxidative stimulation-induced NF-κB activity and could ameliorate the inflammation response. The results also were confirmed by experiments in vivo, Ast alleviated IVDD development and CEP calcification. CONCLUSIONS: Ast could protect vertebral cartilage endplate against oxidative stress and degeneration via activating Nrf-2/HO-1 pathway. Our results imply that Ast may serve as a potential therapeutic agent for IVDD progression and treatment.

Clinical Characteristics of Minimal Lumbar Disc Herniation and Efficacy of Percutaneous Endoscopic Lumbar Discectomy via Transforaminal Approach: A Retrospective Study.

OBJECTIVE: To define the characteristics of Mini LDH, develop new diagnostic references and examine the clinical efficacy of percutaneous endoscopic lumbar discectomy via a transforaminal approach (TF-PELD) for it. METHODS: A total of 72 patients who underwent TF-PELD with Mini LDH from September 2019 to October 2022 were enrolled in this retrospective study. The patients' basic information, symptoms, number of outpatient visits, duration of conservative treatment, physical examination findings and so on were obtained from the medical records. Clinical effects of TF-PELD for Mini LDH were assessed by means of the following: the Visual Analog Scale (VAS) for low back pain (LBP) and leg pain, Oswestry Disability Index (ODI) for functional status assessment and Modified Mac Nab criteria for patient satisfaction. RESULTS: Mini LDH have specific clinical characteristics and imaging features. All included patients achieved obvious pain relief after TF-PELD surgery. Pain scores were repeated at postoperative day 1 and 1, 3, 6, 12 and 24 months later. Results were statistically analyzed. The average VAS-Back, VAS-Leg and ODI scores were all significantly reduced at the first postoperative day and gradually decreased with the follow-up time continuing. In total, 66 out of 72 patients received an excellent or good recovery and no poor result was reported according to the Modified Mac Nab criteria. CONCLUSIONS: Mini LDH is a type of LDH with special characteristics and in need of correct diagnosis and active treatment in clinical work. TF-PELD was also found to be an effective procedure for the treatment of Mini LDH.

PTEN inhibitor VO-OHpic protects endplate chondrocytes against apoptosis and calcification via activating Nrf-2 signaling pathway.

Cartilage endplate (CEP) degeneration and calcification is an important contributor to the onset and pathogenesis of intervertebral disc degeneration (IDD). However, the underlying mechanisms of CEP degeneration remain elusive, let alone according treatment strategies to prevent CEP degeneration. Phosphatase and tensin homolog (PTEN) is a tumor suppressor gene that promotes cell apoptosis, and recent studies indicated that PTEN is overexpressed in degenerated intervertebral disc. However, whether direct inhibition of PTEN attenuates CEP degeneration and IDD development remains largely unknown. In the present study, our in vivo experiments demonstrated that VO-OHpic could attenuate IDD progression and CEP calcification. We also found that VO-OHpic inhibited oxidative stress induced chondrocytes apoptosis and degeneration by activating Nrf-2/HO-1 pathway, thus promoted parkin mediated mitophagy process and inhibited chondrocytes ferroptosis, alleviated redox imbalance and eventually improved cell survival. Nrf-2 siRNA transfection significantly reversed the protective effect of VO-OHpic on endplate chondrocytes. In conclusion, our study demonstrated that inhibition of PTEN with VO-OHpic attenuates CEP calcification and IDD progression. Moreover, VO-OHpic protects endplate chondrocytes against apoptosis and degeneration via activating Nrf-2/HO-1 mediated mitophagy process and ferroptosis inhibition. Our results suggest that VO-OHpic may be a potential effective medicine for IDD prevention and treatment.

Compressive stress induces spinal vertebral growth plate chondrocytes apoptosis via Piezo1.

Many clinical studies have indicated an association between biomechanical factors and the incidence and pathological progression of adolescent idiopathic scoliosis (AIS). However, at present, the research on AIS is mainly focused on the etiology, and there are few studies reporting the causes of progressive aggravation of AIS. In the present study, we aim to investigate the role of Piezo1 in compressive stress-induced mouse spinal vertebral growth plate chondrocytes apoptosis. First, a scoliosis mouse model was established, and the expression of Piezo1 as well as the degree of apoptosis were investigated. We found that the expression of Piezo1 and the degree of apoptosis were significantly higher on the concave sides than that on the convex sides of the vertebral growth plate in mice with scoliosis. Spinal vertebral growth plate chondrocytes were further isolated and treated with Yoda1 to mimic Piezo1 overload. Excess Piezo1 significantly promoted apoptosis of spinal vertebral growth plate chondrocytes. Moreover, static gas compressive stress was used to simulate the increased concave compressive stress in the process of scoliosis with or without GsMTx4, a Piezo inhibitor. It was observed that with the increase of static compressive stress, the expression of Piezo1 increased, and the chondrocytes of vertebral growth plate treated with Piezo1 inhibitor GsMTx4 weakened the above phenomena. In conclusion, our results indicated that compressive stress is strongly associated with the different degrees of apoptosis on both sides on the convex and concave sides of the vertebral growth plate in scoliosis via inducing different expressions of Piezo1. Reducing the expression of Piezo1 in the concave side of the vertebral growth plate and inhibiting the apoptosis of chondrocytes in the bilateral vertebral growth plate caused by asymmetric stress on both sides of the concave vertebral body may be a promising treatment strategy for AIS.

Desferoxamine protects against hemophilic arthropathy through the upregulation of HIF-1α-BNIP3 mediated mitophagy.

AIMS: Hemophilic arthropathy (HA) is a typically iron overload induced joint disease secondary to continuous joint bleeding, however, the exact role of iron chelators in HA has not been fully elucidated. In the present study, we investigated whether desferoxamine (DFO), an iron chelator, could limit the development of HA and the underlying mechanisms. MATERIALS AND METHODS: A HA mice model was established by needle puncture in the left knees of FVIII-deficient hemophilic mice. HA progression was evaluated at 8 weeks after DFO administration. Moreover, chondrocytes were treated with ferric ammonium citrate (FAC) to mimic iron overload in vitro. Modulating effect of DFO on iron overload induced oxidative stress, chondrocytes apoptosis and extracellular matrix (ECM) degradation and the role of HIF-1α-BNIP3 mediated mitophagy were examined. KEY FINDINGS: We found that DFO limited the development of HA and protected iron overload induced ECM degradation, chondrocytes apoptosis and oxidative stress. Besides chelating Fe2+, we found that HIF-1α-BNIP3 mediated mitophagy played important roles in the protective effect of DFO. HIF-1α inhibition suppressed chondrocytes mitophagy process and partly diminished the protective effect of DFO on chondrocytes iron overload. SIGNIFICANCE: In conclusion, DFO could protect against HA development via HIF-1α-BNIP3 mediated mitophagy, suggesting DFO might be a potential therapeutic supplement for HA treatment.

Icariin protects vertebral endplate chondrocytes against apoptosis and degeneration via activating Nrf-2/HO-1 pathway.

Cartilage endplate (CEP) plays important roles in the onset and progression of intervertebral disc degeneration (IVDD). Icariin (ICA) is the major active ingredient of Herba Epimedii and has various biological activities such as anti-inflammatory and antioxidant, which is used to treat many degenerative diseases. However, the effects and mechanism of ICA on endplate chondrocytes are still unclear. Herein, we studied the effects of ICA on CEP degeneration and elucidated the underlying mechanisms. Endplate chondrocytes were isolated, and TNF-α and TBHP were applied to mimic an IVDD pathological environment. Also, an IVDD mice model was established by transection of bilateral facet joints to investigate the protective effect of ICA in vivo. We found that ICA treatment inhibited the chondrocytes apoptosis and the decrease of extracellular matrix production in a dose-dependent manner. Our in vivo experiments demonstrated that ICA could ameliorate IVDD development and CEP calcification. We also found that the ICA-activated Nrf-2/HO-1 pathway thus promoted the Parkin-mediated mitophagy process and inhibited chondrocytes ferroptosis, thus alleviated redox imbalance and mitochondrial dysfunction and eventually improved cell survival. Knockdown of Nrf-2 using siRNA reversed the protective effect of ICA on endplate chondrocytes apoptosis and degeneration. In conclusion, our study demonstrated that ICA could protect against CEP degeneration and calcification under IVDD pathological conditions, the associated mechanism may be related to Nrf-2/HO-1-mediated mitophagy activation and ferroptosis inhibition. Our results suggest that ICA may be a potential effective medicine for IVDD prevention and treatment.

Iron overload promotes intervertebral disc degeneration via inducing oxidative stress and ferroptosis in endplate chondrocytes.

Iron overload is a common phenomenon in the elderly population. Many clinical studies have indicated an association between iron overload and the incidence and pathological progression of intervertebral disc degeneration (IVDD). However, the role and underlying mechanism by which iron participates in the progression of IVDD has not yet been reported. In the present study, we aimed to elucidate the connection between iron overload and IVDD, and explore the underlying mechanisms of disease. Firstly, a clinical epidemiology study was conducted and revealed that iron overload is an independent risk factor for human IVDD. To elucidate the role of iron overload in IVDD, an iron overload mouse model was established, and we observed that iron overload promoted IVDD and cartilage endplate degeneration in a dose dependent manner. Endplate chondrocytes were further isolated and treated with FAC to mimic iron overload in vitro. Excess iron significantly promoted mineralization of endplate chondrocytes in addition to their degeneration via oxidative stress. Moreover, a high dose of excess iron promoted chondrocytes ferroptosis. An iron chelator (DFO), an antioxidant (NAC) and a ferroptosis inhibitor (Fer-1) demonstrated effective inhibition of endplate chondrocyte degeneration induced by iron overload, and our in vivo studies further demonstrated that DFO, NAC and Fer-1 could rescue high dose iron-induced IVDD and cartilage endplate calcification. In conclusion, our results indicate that iron overload is strongly associated with the onset and development of IVDD via oxidative stress and ferroptosis. Inhibiting oxidative stress or ferroptosis could therefore be promising therapeutic strategies for IVDD induced by iron overload.

Multifactor Analysis of Costal Pain in Osteoporotic Fracture of Thoracic Vertebra.

BACKGROUND: The costal pain is common in thoracic osteoporotic fracture patients. It is unclear why vertebral fracture patients without any specific nerve impingement on magnetic resonance imaging (MRI) present with costal pain. OBJECTIVES: The aim of this study was to investigate the potential causes of costal pain in patients with osteoporotic fracture of thoracic vertebra. STUDY DESIGN: A retrospective study. SETTING: Shandong province, China. METHODS: In this retrospective study, 100 patients with thoracic osteoporotic fractures were collected and assigned into 2 groups on the basis of pain patterns noted during medical history and physical examination. Group A was comprised of 50 patients with costal pain. Group B was comprised of 50 patients without costal pain. The Visual Analog Scale and Oswestry Disability Index scores were recorded to assess the pattern and severity of pain. The gender, age, presence or absence of trauma, time of fracture, fracture segments, and analgesic application were recorded. Computed tomography data including changes in fracture vertebral body shape (height, width, and length), intervertebral foramen shape (height and width), wedge shape of fractured vertebral body, and local kyphosis angle were recorded. The fracture edema signal was determined by MRI. Multivariate analysis was performed for all the above parameters. RESULTS: There was a statistically significant difference in the vertebral body width between the 2 groups. LIMITATIONS: The number of patients enrolled is not large enough. We also have limitations in interpreting all pains resulting from osteoporotic vertebral compression fractures, because all pain mechanisms are not fully understood. Further work is needed to improve the accuracy of locating pain sources and distinguishing pain patterns which may result from other spinal structures. CONCLUSION: The incidence of costal pain is significantly and positively associated with the width of the fractured vertebra in patients with osteoporotic thoracic vertebrae fracture.

Calcium chelator BAPTA­AM protects against iron overload­induced chondrocyte mitochondrial dysfunction and cartilage degeneration.

Osteoarthritis (OA) is a common joint disease that is characterized by cartilage degradation. Iron deposition in the joints is common during the pathogenic progression of OA and recent studies have indicated that iron overload is an important contributor to OA progression. Calcium chelators have been reported to inhibit iron influx via modulating transferrin receptor protein 1 internalization, and they have been identified as a potential approach to the treatment of iron overload­induced diseases. The aim of the present study was to investigate the effect of calcium chelators on the progression of iron overload­induced OA. Primary chondrocytes were treated with various concentrations of ferric ammonium citrate (FAC) to mimic iron overload in vitro, followed by co­treatment with the calcium chelator BAPTA acetoxymethyl ester (BAPTA­AM). Subsequently, intracellular iron levels, cell viability, reactive oxygen species (ROS) levels, mitochondrial function and morphological changes, as well as MMP levels, were detected using commercial kits. It was demonstrated that FAC treatment significantly promoted chondrocyte apoptosis and the expression of MMPs, and these effects were reversed by co­treatment with BAPTA­AM. Moreover, BAPTA­AM suppressed iron influx into chondrocytes and inhibited iron overload­induced ROS production and mitochondrial dysfunction. These results indicated that calcium chelators may be of value in the treatment of iron metabolism­related diseases and iron overload­induced OA progression.

Association Between Vertebral Rotatory Subluxation and the Apical Vertebra in Degenerative Lumbar Scoliosis.

OBJECTIVE: To identify the apical vertebra is relate to the occurrence of vertebral rotatory subluxation (VRS) in degenerative lumbar scoliosis. METHODS: In total, 86 patients with degenerative lumbar scoliosis included in this retrospective study were divided into 2 groups: with VRS and without VRS. Coronal and sagittal parameters were measured on whole-spine anteroposterior and lateral radiographs. The impact of VRS on the spine and pelvis parameters was statistically analyzed, and the correlation between the occurrence of VRS and the apical vertebra was analyzed by using logistic regression. RESULTS: VRS was present in 49 patients with degenerative lumbar scoliosis and absent in 37 patients. Compared with the patients without VRS, the patients with VRS had a smaller lumbar lordosis and thoracic kyphosis (P < 0.05) and a larger apical vertebra translation, apical vertebra tilt angle, Cobb angle, and pelvic incidence-lumbar lordosis (P < 0.05). Logistic regression analysis showed that apical vertebra translation and apical vertebra tilt angle were risk factors for the occurrence of VRS. The receiver operating characteristic curve analysis showed that apical vertebra translation greater than 23.2 mm and apical vertebra tilt angle greater than 11.8° were related to a greater probability of VRS in patients with degenerative lumbar scoliosis. CONCLUSIONS: VRS is an important characteristic to consider in degenerative lumbar scoliosis that can affect the coronal and sagittal alignment. The apical vertebra is correlated with the occurrence of VRS; an apical vertebra translation greater than 23.2 mm and apical vertebra tilt angle greater than 11.8° are highly correlate with the occurrence of VRS.

The detrimental effect of iron on OA chondrocytes: Importance of pro-inflammatory cytokines induced iron influx and oxidative stress.

Iron overload is common in elderly people which is implicated in the disease progression of osteoarthritis (OA), however, how iron homeostasis is regulated during the onset and progression of OA and how it contributes to the pathological transition of articular chondrocytes remain unknown. In the present study, we developed an in vitro approach to investigate the roles of iron homeostasis and iron overload mediated oxidative stress in chondrocytes under an inflammatory environment. We found that pro-inflammatory cytokines could disrupt chondrocytes iron homeostasis via upregulating iron influx transporter TfR1 and downregulating iron efflux transporter FPN, thus leading to chondrocytes iron overload. Iron overload would promote the expression of chondrocytes catabolic markers, MMP3 and MMP13 expression. In addition, we found that oxidative stress and mitochondrial dysfunction played important roles in iron overload-induced cartilage degeneration, reducing iron concentration using iron chelator or antioxidant drugs could inhibit iron overload-induced OA-related catabolic markers and mitochondrial dysfunction. Our results suggest that pro-inflammatory cytokines could disrupt chondrocytes iron homeostasis and promote iron influx, iron overload-induced oxidative stress and mitochondrial dysfunction play important roles in iron overload-induced cartilage degeneration.

Mitophagy in degenerative joint diseases.

Mitochondrial dysfunction is involved in aging and multiple degenerative diseases, including intervertebral disc degeneration (IVDD) and osteoarthritis (OA). Thus, the maintenance of mitochondria homeostasis and function is important. Mitophagy, a process that selectively clears damaged or dysfunctional mitochondria through autophagic machinery, functions to maintain mitochondrial quality control and homeostasis. IVDD and OA are similar joint diseases involving the degradation of cartilaginous tissues that are mainly caused by oxidative stress, cell apoptosis and extracellular matrix (ECM) degradation. Over the past decade, accumulating evidence indicates the essential role of mitophagy in the pathogenesis of IVDD and OA. Importantly, strategies by the regulation of mitophagy exert beneficial effects in the pre-clinical experiments. Given the importance and novelty of mitophagy, we provide an overview of mitophagy pathways and discuss the roles of mitophagy in IVDD and OA. We also highlight the potential of targeting mitophagy for the treatment of degenerative joint diseases.Abbreviations: AD: Alzheimer disease; AF: annulus fibrosus; ADORA2A/A2AR: adenosine A2a receptor; AMBRA1: autophagy and beclin 1 regulator 1; BMSCs: bone marrow mesenchymal stem cells; BNIP3: BCL2 interacting protein 3; BNIP3L/NIX: BCL2/adenovirus E1B interacting protein 3-like; CDH6: cadherin 6; CEP: cartilaginous endplates; circRNA: circular RNA; DNM1L/DRP1: dynamin 1-like; ECM: extracellular matrix; HIF1A: hypoxia inducible factor 1: alpha subunit; IL1B: interleukin 1 beta; IMM: inner mitochondrial membranes; IVDD: intervertebral disc degeneration; MAPK8/JNK: mitogen-activated protein kinase 8; MFN1: mitofusin 1; MFN2: mitofusin 2; MIA: monosodium iodoacetate; RHOT/MIRO: ras homolog family member T; MMP: mitochondrial transmembrane potential; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; NFE2L2: nuclear factor: erythroid 2 like 2; NP: nucleus pulposus; OA: osteoarthritis; OPA1: OPA1: mitochondrial dynamin like GTPase; OPTN: optineurin; PRKN: parkin RBR E3 ubiquitin protein ligase; PD: Parkinson disease; PGAM5: PGAM family member 5; PPARGC1A/PGC-1A: peroxisome proliferator activated receptor: gamma: coactivator 1 alpha; PHF23: PHD finger protein 23; PINK1: PTEN induced putative kinase 1; ROS: reactive oxygen species; SfMSCs: synovial fluid MSCs; SIRT1: sirtuin 1; SIRT2: sirtuin 2; SIRT3: sirtuin 3; SQSTM1/p62: sequestosome 1; TNF: tumor necrosis factor; Ub: ubiquitin; UBL: ubiquitin-like; VDAC: voltage-dependent anion channel.

Hyperoside ameliorates the progression of osteoarthritis: An in vitro and in vivo study.

BACKGROUND: Osteoarthritis (OA) is a common degenerative joint disease. The pathogenesis of OA is closely related to inflammatory responses and apoptosis of chondrocytes. Hyperoside (Hyp), a natural flavonoid compound, exerts multiple bioactivities in various diseases. PURPOSE: Our study aims to investigate the anti-arthritic effects of Hyp and delineate the potential mechanism at the cellular level. METHODS: Murine chondrocytes were stimulated with interleukin-1ß (IL-1ß) with or without Hyp treatment. CCK-8 assay was used to evaluate the cytotoxic effect of Hyp. DCFH-DA was used to detect intracellular ROS. Annexin V-FITC/PI method was applied to examine apoptosis of chondrocytes. The anti-arthritic effects of Hyp and related mechanisms were investigated by examining and analyzing relative markers through quantitative PCR, western blot analysis and immunofluorescent staining. C57BL/6 mice were performed the destabilized medial meniscus (DMM) surgery to establish OA model and then injected intraperitoneally with Hyp (20 mg/kg)) for 4 or 8 weeks. Finally, mice were sacrificed and knee joints were collected for histological observation and analysis. RESULTS: Hyp inhibited IL-1ß-induced expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Additionally, Hyp attenuated IL-1ß-induced destruction of the extracellular matrix (ECM) by downregulating the expression of MMPs and ADAMTS5, and meanwhile upregulating the expression of collagen II, aggrecan, and SOX9. Also, Hyp pretreatment reduced IL-1ß-induced overproduction of ROS and apoptosis of chondrocytes. Mechanistically, Hypexerted anti-inflammatory effects by partly suppressing the PI3K/AKT/NF-κB and the MAPK signaling pathways, enhancing the Nrf2/HO-1 to limit the activation of NF-κB. Moreover, Hyp played an anti-apoptotic effect via the Nrf2/ROS/BAX/Bcl-xlaxis. In vivo, cartilage degradation was attenuated with a lower OARSI score in Hyp-treated group compared to the DMM group. CONCLUSION: Our study demonstrated that anti-arthritic effects of Hyp in vitro and in vivo, indicating Hyp might serve as a potential agent for the treatment of OA.

Desferoxamine protects against glucocorticoid-induced osteonecrosis of the femoral head via activating HIF-1α expression.

Glucocorticoid-induced osteonecrosis of the femoral head (GIOFH) is one of the most common complications of glucocorticoid administration. By chelating Fe2+ , desferoxamine (DFO) was reported to be able to activate the HIF-1α/VEGF pathway and promote angiogenesis. In the present study, we examined whether DFO administration could promote angiogenesis and bone repair in GIOFH. GIOFH was induced in rats by methylprednisolone in combination with lipopolysaccharide. Bone repair was assessed by histologic analysis and microcomputed tomography (micro-CT). Vascularization was assessed by Microfil perfusion and micro-CT analysis. Immunohistochemical staining was performed to analyze the expression of HIF-1α, VEGF, and CD31. Our in vivo study revealed that DFO increased HIF-1α/VEGF expression and promoted angiogenesis and osteogenesis in GIOFH. Moreover, our in vitro study revealed that DFO restored dexamethone-induced HIF-1α downregulation and angiogenesis inhibition. Besides, our in vitro study also demonstrated that DFO could protect bone marrow-derived stem cells from dexamethone-induced apoptosis and mitochondrial dysfunction by promoting mitophagy and mitochondrial fission. In summary, our data provided useful information for the development of novel therapeutics for management of GIOFH.

Mechanical loading induces HIF-1α expression in chondrocytes via YAP.

OBJECTIVES: To investigate the role of YAP in cyclic mechanical stress induced up-regulation of HIF-1α in rat cartilage chondrocytes. RESULTS: Our in vitro and in vivo experiments demonstrated that cyclic mechanical stress promoted HIF-1α and YAP proteins expression in a magnitude dependent manner. Cyclic mechanical stress at 4000µ strain exhibited most significant effect in promoting HIF-1α and YAP up-regulation. Activation of YAP using LPA significantly promoted HIF-1α stabilization and expression, while YAP siRNA treatment suppressed the up-regulation of HIF-1α induced by cyclic mechanical stress. CONCLUSION: Our results indicated that cyclic mechanical stress promoted HIF-1α stabilization and YAP is involved in mechanical stress induced HIF-1α up-regulation.

PTEN inhibitor VO-OHpic attenuates GC-associated endothelial progenitor cell dysfunction and osteonecrosis of the femoral head via activating Nrf2 signaling and inhibiting mitochondrial apoptosis pathway.

BACKGROUND: Glucocorticoid (GC)-associated osteonecrosis of the femoral head (ONFH) is the most common in non-traumatic ONFH. Despite a strong relationship between GC and ONFH, the detailed mechanisms have remained elusive. Recent studies have shown that GC could directly injure the blood vessels and reduce blood supply in the femoral head. Endothelial progenitor cells (EPCs), which were inhibited quantitatively and functionally during ONFH, play an important role in maintaining the normal structure and function of vascular endothelium. Phosphatase and tensin homolog (PTEN) is a tumor suppressor gene that promotes cell apoptosis, and its expression was found to be elevated in GC-associated ONFH patients. However, whether direct inhibition of PTEN attenuates GC-associated apoptosis and dysfunction of the EPCs remains largely unknown. METHODS: We investigated the effect of, VO-OHpic, a potent inhibitor of PTEN, in attenuating GC-associated apoptosis and dysfunction of EPCs and the molecular mechanism. SD rats were used to study the effect of VO-OHpic on angiogenesis and osteonecrosis in vivo. RESULTS: The results revealed that methylprednisolone (MPS) obviously inhibit angiogenesis of EPCs by inducing apoptosis, destroying the normal mitochondrial structure, and disrupting function of mitochondria. VO-OHpic treatment is able to reverse the harmful effects by inhibiting the mitochondrial apoptosis pathway and activating the NF-E2-related factor 2 (Nrf2) signaling. Si-Nrf2 transfection significantly reduced the protective effects of VO-OHpic on EPCs. Our in vivo studies also showed that intraperitoneal injection of VO-OHpic obviously attenuates the osteonecrosis of the femoral head induced by MPS and potently increases the blood supply in the femoral head. CONCLUSION: Taken together, the data suggests that inhibition of PTEN with VO-OHpic attenuates apoptosis and promotes angiogenesis of EPCs in vitro via activating Nrf2 signaling pathway and inhibiting the mitochondrial apoptosis pathway. Moreover, VO-OHpic also mitigates GC-associated ONFH and potentiates angiogenesis in the femoral head.