The mitochondrial UPR induced by ATF5 attenuates intervertebral disc degeneration via cooperating with mitophagy.

Intervertebral disc degeneration (IVDD) is an aging disease that results in a low quality of life and heavy socioeconomic burden. The mitochondrial unfolded protein response (UPRmt) take part in various aging-related diseases. Our research intents to explore the role and underlying mechanism of UPRmt in IVDD. Nucleus pulposus (NP) cells were exposed to IL-1ß and nicotinamide riboside (NR) served as UPRmt inducer to treat NP cells. Detection of ATP, NAD + and NADH were used to determine the function of mitochondria. MRI, Safranin O-fast green and Immunohistochemical examination were used to determine the degree of IVDD in vivo. In this study, we discovered that UPRmt was increased markedly in the NP cells of human IVDD tissues than in healthy controls. In vitro, UPRmt and mitophagy levels were promoted in NP cells treated with IL-1ß. Upregulation of UPRmt by NR and Atf5 overexpression inhibited NP cell apoptosis and further improved mitophagy. Silencing of Pink1 reversed the protective effects of NR and inhibited mitophagy induced by the UPRmt. In vivo, NR might attenuate the degree of IDD by activating the UPRmt in rats. In summary, the UPRmt was involved in IVDD by regulating Pink1-induced mitophagy. Mitophagy induced by the UPRmt might be a latent treated target for IVDD.

Effectiveness of Physiotherapeutic Scoliosis-Specific Exercises on 3-Dimensional Spinal Deformities in Patients With Adolescent Idiopathic Scoliosis: A Systematic Review and Meta-analysis.

OBJECTIVE: To investigate the effects of physiotherapeutic scoliosis-specific exercises (PSSE) on coronal, horizontal, and sagittal deformities of the spine in adolescent idiopathic scoliosis (AIS) as well as how curve severity, intervention duration, and intervention type could modify these effects. DATA SOURCES: Data sources included PubMed, Web of Science, Embase, Cochrane Library, and Scopus databases, which were searched from their inception to September 5, 2023. STUDY SELECTION: Clinical controlled trials reporting the effects of PSSE on the Cobb angle, angle of trunk rotation (ATR), thoracic kyphosis (TK), or lumbar lordosis in patients with AIS aged 10-18 years. The experimental groups received PSSE; the control groups received standard care (observation or bracing) or conventional exercise such as core stabilization exercise, Pilates, proprioceptive neuromuscular facilitation, and other nonspecific exercises. DATA EXTRACTION: Two researchers independently extracted key information from eligible studies. The quality of the studies was assessed using the Cochrane Handbook version 5.1.0 risk of bias assessment and the JBI Center for Evidence-Based Health Care (2016) of quasi-experimental research authenticity assessment tool. The level and certainty of evidence were rated according to the Grading of Recommendations, Assessment, Development, and Evaluation framework. We followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement. The protocol for this study was registered in PROSPERO (CRD42023404996). DATA SYNTHESIS: Twelve randomized controlled trials (RCTs) and 5 non-RCTs were meta-analyzed separately. The results indicated that compared with other nonsurgical management, PSSE significantly improved the Cobb angle, ATR, and TK, whereas the lumbar lordosis improvement was not statistically significant. Additionally, the efficacy of PSSE on Cobb angle was not significant in patients with curve severity ≥30° compared with controls. Nevertheless, the pooled effect of PSSE on Cobb angle was not significantly modified by intervention duration and intervention type and that on ATR was not significantly modified by intervention duration. The overall quality of evidence according to Grading of Recommendations, Assessment, Development, and Evaluation was moderate to low for RCT and very low for non-RCT. CONCLUSIONS: PSSE exhibited positive benefits on the Cobb angle, ATR, and TK in patients with AIS compared with other nonsurgical therapies. In addition, the effectiveness of PSSE may be independent of intervention duration and intervention type but may be influenced by the initial Cobb angle. However, more RCTs are needed in the future to validate the efficacy of PSSE in moderate AIS with a mean Cobb angle ≥30°. Current evidence is limited by inconsistent control group interventions and small sample size of the studies.

Safety and Efficacy of Polyetheretherketone (PEEK) Cages and Cadaveric Allografts in Transforaminal Lumbar Interbody Fusion (TLIF) for Treating Lumbar Pyogenic Spondylodiscitis.

Purpose: There have been many studies in the operative management of pyogenic spondylodiscitis with foreign materials. However, it still remains an issue of debate on whether the allografts may be used in pyogenic spondylodiscitis. This study sought to evaluate the safety and effectiveness of PEEK cages and the cadaveric allograft in transforaminal lumbar interbody fusion (TLIF) for treating lumbar pyogenic spondylodiscitis. Methods: From January 2012 to December 2019, 56 patients underwent surgery for lumbar pyogenic spondylodiscitis. The posterior debridement of all patients and their fusion with allografts, local bone grafts, and bone chip cages were performed before posterior pedicle screw fusion. An assessment of the residual pain, the grade of neurological injury, and the resolution of infection was conducted on 39 patients. The clinical outcome was evaluated using a visual analog scale (VAS) and the Oswestry Disability Index (ODI), and neurological outcomes were appraised based on Frankel grades. The radiological outcomes were evaluated via focal lordosis, lumbar lordosis, and the state of the fusion. Results: Staphylococcus aureus and Staphylococcus epidermidis were the most common causative organisms. The mean preoperative focal lordosis was -1.2° (-11.4° to 5.7°), and the mean postoperative focal lordosis increased to 10.3° (4.3°-17.2°). At the final follow-up, there were five cases with subsidence of the cage, no case of recurrence, and no case with cage and screw loosening or migration. The mean preoperative VAS and ODI scores were 8.9 and 74.6%, respectively, and improvements in VAS and ODI were 6.6 ± 2.2 and 50.4 ± 21.3%, respectively. The Frankel grade D was found in 10 patients and grade C in 7. Following the final follow-up, only one patient improved from Frankel grade C to grade D while the others recovered completely. Conclusion: The PEEK cage and cadaveric allograft combined with local bone grafts is a safe and effective choice for intervertebral fusion and restoring sagittal alignment without increased incidence of relapse for treating lumbar pyogenic spondylodiscitis.

The Effects of Bone Cement Volume in Percutaneous Vertebroplasty for Thoracolumbar Junction Vertebral Compression Fractures: A Clinical Comparative Study.

We compared the outcomes of patients treated with different volumes of polymethyl methacrylate bone cement during percutaneous vertebroplasty (PVP) for thoracolumbar vertebral compression fractures. We performed a comparative, retrospective study of 316 patients who underwent PVP for a single-level thoracolumbar vertebral compression fracture. Patients were divided into two groups: group A (≤5 mL; n = 146) and group B (>5 mL; n = 170). The visual analogue scale (VAS) for pain and the Roland-Morris Disability Questionnaire (RDQ) scores were compared between the two groups at 1 week and at 1, 6, 12, and 24 months after PVP. The incidence of cement leakage into the intervertebral discs was evaluated by a postoperative lateral radiograph assessment. Patients were evaluated for new fractures 1 and 2 years after PVP or when new fractures were suspected. Among the 316 patients enrolled, 245 completed the clinical research. No difference between groups A and B in terms of the VAS, RDQ, and rate of complications at all time points after surgery was observed. The presence of intervertebral disc leakage was a relative risk (RR) for subsequent total vertebral fracture (RR, 6.42; 95% confidence interval (CI), 2.72-14.19; P < 0.0001) and adjacent vertebral fracture (RR, 8.03; 95% CI, 2.74-23.54; P = 0.0001). A high volume of bone cement may increase the rate of subsequent total and adjacent vertebral fractures. However, the occurrence of intervertebral disc leakage is the principal risk factor for these negative outcomes of PVP.

Involvement of oxidative stress-induced annulus fibrosus cell and nucleus pulposus cell ferroptosis in intervertebral disc degeneration pathogenesis.

Ferroptosis is a necrotic form of regulated cell death that was associated with lipid peroxidation and free iron-mediated Fenton reactions. It has been reported that iron deficiency had been implicated in the pathogenesis of intervertebral disc degeneration (IVDD) by activating apoptosis. However, the role of ferroptosis in the process of IVDD has not been illuminated. Here, we demonstrate the involvement of ferroptosis in IVDD pathogenesis. Our in vitro models show the changes in protein levels of ferroptosis marker and enhanced lipid peroxidation level during oxidative stress. Safranin O staining, hematoxylin-eosin staining, and immunohistochemical were used to assess the IVDD after 8 weeks of surgical procedure in vivo. Treatment with ferrostatin-1, deferoxamine, and RSL3 demonstrate the role of ferroptosis in tert-butyl hydroperoxide (TBHP)-treated annulus fibrosus cells (AFCs) and nucleus pulposus cells (NPCs). Ferritinophagy, nuclear receptor coactivator 4 (NCOA4)-mediated ferritin selective autophagy, is originated during the process of ferroptosis in response to TBHP treatment. Knockdown and overexpression NCOA4 further prove TBHP may induce ferroptosis of AFCs and NPCs in an autophagy-dependent way. These findings support a role for oxidative stress-induced ferroptosis in the pathogenesis of IVDD.

Exosomes derived from vascular endothelial cells antagonize glucocorticoid-induced osteoporosis by inhibiting ferritinophagy with resultant limited ferroptosis of osteoblasts.

High dose and long-term steroid treatment can alter antioxidative ability and decrease the viability and function of osteoblasts, leading to osteoporosis and osteonecrosis. Ferroptosis, a new type of cell death characterized by excessive lipid peroxidation due to the downregulation of GPX4 and system Xc- , is involved in glucocorticoid-induced osteoporosis. Endothelial cell-secreted exosomes (EC-Exos) are important mediators of cell-to-cell communication and are involved in many physiological and pathological processes. However, the effect of EC-Exos on osteoblasts exposed to glucocorticoids has not been reported. Here, we explored the role of EC-Exos in glucocorticoid-induced osteoporosis. In vivo and in vitro experiments indicated that EC-Exos reversed the glucocorticoid-induced osteogenic inhibition of osteoblasts by inhibiting ferritinophagy-dependent ferroptosis.

Autophagy activation facilitates mechanical stimulation-promoted osteoblast differentiation and ameliorates hindlimb unloading-induced bone loss.

Autophagy has been indicated to be involved in regulating bone metabolism. However, little is known about the role of autophagy in mechanical stimulation-influenced osteoblast differentiation and bone formation. In the present study, we first demonstrated that autophagy activation was essential for cyclic mechanical stretching-promoted osteoblast differentiation of bone marrow mesenchymal stem cells. To explore the in vivo role of autophagy in osteoblast differentiation, the hindlimb unloading-induced disuse osteoporosis model was used. Compared to the normal controls, hindlimb unloading led to abundant bone loss as well as lessened autophagy activation of osteoblasts. However, the activation of autophagy by ULK1 overexpression or in the presence of rapamycin significantly increased osteoblast differentiation activity and restored the bone volume. The findings implicate autophagy as a novel mechanosensitive pathway that regulates osteoblast differentiation. The pharmacological activation of autophagy may be an interesting approach for the prevention and treatment of disuse osteoporosis.

Evidence of the Role of R-Spondin 1 and Its Receptor Lgr4 in the Transmission of Mechanical Stimuli to Biological Signals for Bone Formation.

The bone can adjust its mass and architecture to mechanical stimuli via a series of molecular cascades, which have been not yet fully elucidated. Emerging evidence indicated that R-spondins (Rspos), a family of secreted agonists of the Wnt/ß-catenin signaling pathway, had important roles in osteoblastic differentiation and bone formation. However, the role of Rspo proteins in mechanical loading-influenced bone metabolism has never been investigated. In this study, we found that Rspo1 was a mechanosensitive protein for bone formation. Continuous cyclic mechanical stretch (CMS) upregulated the expression of Rspo1 in mouse bone marrow mesenchymal stem cells (BMSCs), while the expression of Rspo1 in BMSCs in vivo was downregulated in the bones of a mechanical unloading mouse model (tail suspension (TS)). On the other hand, Rspo1 could promote osteogenesis of BMSCs under CMS through activating the Wnt/ß-catenin signaling pathway and could rescue the bone loss induced by mechanical unloading in the TS mice. Specifically, our results suggested that Rspo1 and its receptor of leucine-rich repeat containing G-protein-coupled receptor 4 (Lgr4) should be a novel molecular signal in the transmission of mechanical stimuli to biological signal in the bone, and this signal should be in the upstream of Wnt/ß-catenin signaling for bone formation. Rspo1/Lgr4 could be a new potential target for the prevention and treatment of disuse osteoporosis in the future.

Neuropeptide Y1 Receptor Regulates Glucocorticoid-Induced Inhibition of Osteoblast Differentiation in Murine MC3T3-E1 Cells via ERK Signaling.

High dose glucocorticoid (GC) administration impairs the viability and function of osteoblasts, thus causing osteoporosis and osteonecrosis. Neuropeptide Y1 receptor (Y1 receptor) is expressed in bone tissues and cells, and regulates bone remodeling. However, the role of Y1 receptor in glucocorticoid-induced inhibition of osteoblast differentiation remains unknown. In the present study, osteoblastic cell line MC3T3-E1 cultured in osteogenic differentiation medium was treated with or without of 10-7 M dexamethasone (Dex), Y1 receptor shRNA interference, Y1 receptor agonist [Leu31, Pro34]-NPY, and antagonist BIBP3226. Cell proliferation and apoptosis were assessed by cell counting kit-8 (CCK-8) assay and cleaved caspase expression, respectively. Osteoblast differentiation was evaluated by Alizarin Red S staining and osteogenic marker gene expressions. Protein expression was detected by Western blot analysis. Dex upregulated the expression of Y1 receptor in MC3T3-E1 cells associated with reduced osteogenic gene expressions and mineralization. Blockade of Y1 receptor by shRNA transfection and BIBP3226 significantly attenuated the inhibitory effects of Dex on osteoblastic activity. Y1 receptor signaling modulated the activation of extracellular signal-regulated kinases (ERK) as well as the expressions of osteogenic genes. Y1 receptor agonist inhibited ERK phosphorylation and osteoblast differentiation, while Y1 receptor blockade exhibited the opposite effects. Activation of ERK signaling by constitutive active mutant of MEK1 (caMEK) abolished Y1 receptor-mediated Dex inhibition of osteoblast differentiation in MC3T3-E1 cells. Taken together, Y1 receptor regulates Dex-induced inhibition of osteoblast differentiation in murine MC3T3-E1 cells via ERK signaling. This study provides a novel role of Y1 receptor in the process of GC-induced suppression in osteoblast survival and differentiation.

Significance of hypoxia in the physiological function of intervertebral disc cells.

The intervertebral disc (IVD) is the largest avascular structure in the body, and IVD cells reside in vivo in an environment that is considered to be hypoxic. However, the role of oxygen in IVD cell biology remains an issue of debate. By reviewing the available literature about the effect of oxygen tension on regulating the phenotype, energy metabolism, matrix production, and survival of IVD cells, as well as on the expression and function of hypoxia-inducible factor in IVD cells, we conclude that hypoxia is essential in maintaining the physiological function of IVD cells. Modulating the oxygen tension of the IVD or the activity of hypoxia-inducible factor in IVD cells may be a promising strategy for the prevention and treatment of IVD degeneration.

The responses of autophagy and apoptosis to oxidative stress in nucleus pulposus cells: implications for disc degeneration.

BACKGROUND/AIMS: Apoptosis and autophagy are two patterns of programmed cell death which play important roles in the intervertebral disc degeneration. Oxidative stress is an important factor for the induction of programmed cell death. However, the cellular reactions linking autophagy to apoptosis of disc cells under oxidative stress have never been described. This study investigated the responses of autophagy and apoptosis and their interactions in the nucleus pulposus cells (NP cells) under oxidative stress, with the aim to better understand the mechanism of disc degeneration. METHODS: NP cells isolated from rat lumbar discs were subjected to different concentrations of H2O2 for various time periods. Cell viability was determined by CCK-8 assay, and their apoptosis and autophagy responses were evaluated by fluorescent analysis, flow cytometry and western blotting, et al. The interactions of autophagy and apoptosis and the possible signaling pathways were also investigated by using autophagy modulators. RESULTS: H2O2 increased the lysosomal membrane permeability in the NP cells and induced apoptosis through the mitochondrial pathway subsequently. Meanwhile, H2O2 stimulated an early autophagy response through the ERK/m-TOR signaling pathway. Autophagy inhibition significantly decreased the apoptosis incidence in the cells insulted by H2O2. CONCLUSION: These results suggested that controlling the autophagy response in the NP cells under oxidative stress should be beneficial for the survival of the cells and probably delay the process of disc degeneration.

Increased activity of osteocyte autophagy in ovariectomized rats and its correlation with oxidative stress status and bone loss.

OBJECTIVES: The objectives of the present study were to investigate ovariectomy on autophagy level in the bone and to examine whether autophagy level is associated with bone loss and oxidative stress status. METHODS: 36 female Sprague-Dawley rats were randomly divided into sham-operated (Sham), and ovariectomized (OVX) rats treated either with vehicle or 17-ß-estradiol. At the end of the six-week treatment, bone mineral density (BMD) and bone micro-architecture in proximal tibias were assessed by micro-CT. Serum 17ß-estradiol (E2) level were measured. Total antioxidant capacity (T-AOC), superoxide dismutase (SOD) activity, catalase (CAT) activity in proximal tibia was also determined. The osteocyte autophagy in proximal tibias was detected respectively by Transmission Electron Microscopy (TEM), immunofluorescent histochemistry (IH), realtime-PCR and Western blot. In addition, the spearman correlation between bone mass, oxidative stress status, serum E2 and autophagy were analyzed. RESULTS: Ovariectomy increased Atg5, LC3, and Beclin1 mRNA and proteins expressions while decreased p62 expression. Ovariectomy also declined the activities of T-AOC, CAT, and SOD. Treatment with E2 prevented the reduction in bone mass as well as restored the autophagy level. Furthermore, LC3-II expression was inversely correlated with T-AOC, CAT, and SOD activities. A significant inverse correlation between LC3-II expression and BV/TV, Tb.N, BMD in proximal tibias was found. CONCLUSIONS: Ovariectomy induced oxidative stress, autophagy and bone loss. Autophagy of osteocyte was inversely correlated with oxidative stress status and bone loss.

Decreased activity of osteocyte autophagy with aging may contribute to the bone loss in senile population.

Age-related bone loss is a major cause of osteoporosis and osteoporotic fractures in the elderly. However, the underlying molecular mechanism of age-related bone loss is still poorly understood. The aim of this study was to clarify whether autophagy in osteocytes was involved in age-related bone loss. Male Sprague-Dawley (SD) rats in 3, 9, and 24 month old were used to mimic the age-related bone loss in men. Micro-CT evaluation, histomorphometric analysis, and measurement of bone turnover rate verified age-related bone loss in the male SD rats. Immunofluorescent histochemistry, RT-PCR, and Western blot assessment demonstrated that the expression of LC3-II, LC3-II/I, Beclin-1, and Ulk-1 in the osteocytes decreased with age, while SQSTM1/p62 and apoptosis in the osteocytes increased. A significant correlation between the markers of osteocyte autophagy and bone mineral density in the proximal tibia was revealed. However, osteocyte autophagy was not correlated with osteocyte apoptosis in the process of aging. These results suggested that osteocyte autophagy was possibly involved in the age-related bone loss. Decreased activity of osteocyte autophagy independent of apoptosis might contribute to the age-related bone loss in senile osteoporosis.

The effect of transforming growth factor ß1 on the crosstalk between autophagy and apoptosis in the annulus fibrosus cells under serum deprivation.

Autophagy and apoptosis are important in maintaining the metabolic homeostasis of intervertebral disc cells, and transforming growth factor-ß1 (TGF-ß1) is able to delay intervertebral disc degeneration. This study determined the effect of TGF-ß1 on the crosstalk between autophagy and apoptosis in the disc cells, with the aim to provide molecular mechanism support for the prevention and treatment of disc degeneration. Annulus fibrosus (AF) cells were isolated and cultured under serum starvation. 10 ng/mL TGF-ß1 reduced the apoptosis incidence in the cells under serum starvation for 48 h, down-regulated the autophagy incidence in the cells pretreated with 3-methyladenine (3-MA) or Bafilomycin A (Baf A), partly rescued the increased apoptosis incidence in the cells pretreated with 3-MA, while further reduced the decreased apoptosis incidence in the cells pretreated with Baf A. Meanwhile, TGF-ß1 down-regulated the expressions of autophagic and apoptotic markers in the cells under starvation, partly down-regulated the expressions of Beclin-1, LC3 II/I and cleaved caspase-3 in the cells pretreated with 3-MA or Baf A, while significantly decreased the expression of Bax/Bcl-2 in the cells pretreated with Baf A. 3-MA blocked the phosphorylation of both AKT and mTOR and partly reduced the inhibitory effect of TGF-ß1 on the expression of LC3 II/I and cleaved caspase-3. TGF-ß1 enhanced the expression of p-ERK1/2 and down-regulated the expressions of LC3 II/I and cleaved caspase-3. U0126 partly reversed this inhibitory effect of TGF-ß1. In conclusion, TGF-ß1 protected against apoptosis of AF cells under starvation through down-regulating excessive autophagy. PI3K-AKT-mTOR and MAPK-ERK1/2 were the possible signaling pathways involved in this process.

Intraobserver and interobserver reliability of measures of cervical sagittal rotation.

BACKGROUND: Diagnosis and treatment decisions of cervical instability are made, in part, based on the clinician's assessment of sagittal rotation on flexion and extension radiographs. The objective of this study is to evaluate the intraobserver and interobserver reliability of three measurement techniques in assessing cervical sagittal rotation. METHODS: Fifty lateral radiographs of patients with single-level cervical degenerative disc were selected and measured on two separate occasions by three spine surgeons using three different measurement techniques. Cervical sagittal rotation was measured using three different techniques. RESULTS: Intraclass correlation coefficients were most consistent for Method 2 (ICC 0.93-0.96) followed by Method 1 (ICC 0.88-0.91) and Method 3 (ICC 0.81-0.87). Intraobserver agreement (% of repeated measures within 0.5° of the original measurement) ranged between 76% and 96% for all techniques, with Method 2 showing the best agreement (92%-96%). Paired comparisons between observers varied considerably with interobserver reliability correlation coefficients ranging from 0.54 to 0.89. Method 2 showed the highest interobserver reliability coefficient (0.82, range 0.73-0.88). Method 2 was also more reliable for the classification of "instability". Intraobserver percent agreements ranged from 94 to 98% for Method 2 versus 84% to 90% for Method 1 and 78% to 86% for Method 3, while interobserver percent agreements ranged from 90% to 98% for Method 2 versus 86% to 94% for Method 1 and 74% to 84% for Method 3. CONCLUSIONS: Method 2 (measuring the angle from the inferior endplate of the vertebra above the degenerative disc and the inferior endplate of the vertebra below the degenerative disc) showed the best intraobserver and interobserver reliability overall in assessing cervical sagittal rotation.

Estradiol inhibits osteoblast apoptosis via promotion of autophagy through the ER-ERK-mTOR pathway.

Estradiol could protect osteoblast against apoptosis, and apoptosis and autophagy were extensively and intimately connected. The aim of the present study was to test the hypothesis that autophagy was present in osteoblasts under serum deprivation and estrogen protected against osteoblast apoptosis via promotion of autophagy. MC3T3-E1 osteoblastic cells were cultured in a serum-free and phenol red-free minimal essential medium (α-MEM). Ultrastructural analysis, lysosomal activity assessment and monodansycadaverine (MDC) staining were employed to determine the presence of autophagy, and real time PCR was used to evaluate the expression of autophagic markers. Meanwhile, the osteoblasts were transferred in a serum-free and phenol red-free α-MEM containing either vehicle or estradiol. Apoptosis and autophagy was assessed by using the techniques of real-time PCR, Western blot, immunofluorescence assay, and flow cytometry. The possible pathway through which estrogen promoted autophagy in the serum-deprived osteoblasts was also investigated. Real-time PCR demonstrated the expression of LC3, beclin1 and ULK1 genes in osteoblasts under serum deprivation, and immunofluorescence assay verified high expression of proteins of these three autophagic bio-markers. Lysosomes and autolysosomes accumulated in the cytoplasm of osteoblasts were also detected under transmission electron microscopy, MDC staining and lysosomal activity assessment. Meanwhile, estradiol significantly decreased the expression of proteins of the bio-markers of apoptosis, and at the same time increased the expression of proteins of the bio-markers of autophagy in the serum-deprived osteoblasts. Furthermore, the estradiol-promoted autophagy in serum-deprived osteoblasts could be blocked by estrogen receptor (ER) antagonist (ICI 182780), and estradiol failed to rescue the cells pretreated with an inhibitor of vacuolar ATPase (bafilomycin A) from apoptosis. Serum deprivation resulted in apoptosis through activation of Caspase-3 and induced autophagy through inhibition of phospho-mammalian target of rapamycin (p-mTOR). Both 3-methyladenine (3MA) and U0126 led to increase of apoptosis in osteoblasts with serum deprivation. Estradiol failed to over-ride the inhibitory effect of 3MA on phosphorylation of AKT but directly led to dephosphorylation of mTOR and upregulation of LC3 protein expression. However, the estradiol-enhanced LC3 protein expression was significantly suppressed by U0126 through inhibition of phosphorylation of extracellular signal-regulated kinase (ERK). Estradiol rescued osteoblast apoptosis via promotion of autophagy through the ER-ERK-mTOR pathway.

Isolated osteoblasts from spinal cord-injured rats respond less to mechanical loading as compared with those from hindlimb-immobilized rats.

BACKGROUND: The pathogenesis of osteoporosis after spinal cord injury (SCI) may be different from disuse osteoporosis. OBJECTIVE: To investigate whether there is the differential anabolic response to mechanical loading between osteoblasts from SCI rats and those from hindlimb-immobilized rats. METHODS: Femoral bone-marrow was harvested for osteoblast culture from SCI rats, hindlimb-immobilized rats, and control rats 3 weeks after animal model creation. At the stage of differentiation, rat osteoblasts were plated in six-well plates for stretching. Cyclic strains were applied for 48 hours, and then alkaline phosphatase (ALPase) activity, procollagen, and osteocalcin production, and gene expression of osteocalcin, runt-related transcription factor 2 (Runx2), and osterix were measured in osteoblasts from SCI rats, hindlimb-immobilized rats, and control rats. RESULTS: ALPase activity, procollagen, and osteocalcin production, and gene expression of osteocalcin, Runx2, and osterix were significantly lower in osteoblasts after stretching from SCI rats compared with those from hindlimb-immobilized rats. However, there was no significant difference of these parameters between isolated osteoblasts from hindlimb-immobilized rats and those from control rats. CONCLUSION: The activity of isolated osteoblasts from SCI rats was lower than control rats, and this suggested that osteoblasts from SCI rats responded less to mechanical loading as compared with those from control rats.

Acceptable Fusion Rate of Single-Level OLIF Using Pure Allograft Combined with Posterior Instrumentation through the Wiltse Approach: A 2-Year Follow-Up Study.

OBJECTIVE: Autogenic bone grafts have shown successful fusion rates in the treatment of degenerative lumbar disorders, but taking too many autogenic bones may result in donor site ischemia or infection. This study aimed to evaluate the outcomes of single-level oblique lumbar interbody fusion (OLIF) using pure allograft combined with posterior pedicle screw instrumentation through the Wiltse approach. METHODS: A retrospective case analysis was performed on a series of consecutive patients who received a single-level OLIF procedure combined with posterior pedicle screw instrumentation through the Wiltse approach between July 1, 2017, and December 31, 2019, in which pure allogenic bone graft was used and filled in the large window of the cage. The patients were followed up as scheduled at 1 day and 3, 6, 12, 24 months after operation. Clinical outcome was assessed by multiple questionnaires, including Oswestry disability index (ODI), Japanese Orthopaedic Association (JOA) score rating system, short form-36 health survey (SF-36), and visual analog scale (VAS) for low back pain. Radiographic outcome was evaluated by measuring the parameters such as disc height, lumbar lordosis, and segmental angle on the standard standing lateral radiographs, and the space angle of the fusion level on the dynamic views of the lateral radiographs. Subsidence of the cage and intervertebral fusion status were evaluated on both the radiographic and CT scan images. RESULTS: A total of 34 patients were finally included in this study. At 2-year follow-up, the VAS for low back pain, ODI, JOA, and SF-36 scores all had significant improvement (p < 0.001). Substantial increase of anterior and posterior disc heights was observed (p < 0.001). Both lumbar lordosis and segmental angle became larger (p < 0.05). No visible change of the space angle of the fusion level was found on the dynamic views. The 1-year fusion rate of 73.5% on CT scans proceeded to 82.4% at 2-year follow-up. The fusion rate was as high as 91.2% according to Bridwell interbody fusion grading system on radiographic images. The clinical outcomes in patients with incomplete fusion were just as good as those with complete fusion. The six patients with cage subsidence had higher ODI (p < 0.001) and lower JOA (p < 0.001) and SF-36 PCS (p = 0.011) scores than those without cage subsidence. CONCLUSION: The use of pure allograft in single-level OLIF resulted in an acceptable fusion rate and satisfactory clinical effect at 2-year follow-up. Supplementation of posterior pedicle screw through the minimally invasive Wiltse approach ensured the favorable outcomes both clinically and radiographically.

High Endplate Hounsfield Units Value Indicate Intervertebral Disc Degeneration Following Transforaminal Lumbar Interbody Fusion Surgery.

OBJECTIVE: Lumbar disc degeneration (LDD) is a common cause of low back pain and disability, and its prevalence increases with age. The aim of this study is to investigate whether endplate Hounsfield unit (HU) values have an effect on lumbar disc degeneration (LDD) after transforaminal lumbar interbody fusion (TLIF) surgery in patients with degenerative lumbar stenosis. METHODS: This study was a retrospective analysis of patients who underwent TLIF surgery in January 2016 to October 2019. One hundred and fifty-seven patients who underwent TLIF surgery for degenerative lumbar stenosis were enrolled in this study. Demographic data was recorded. VAS and ODI values were compared to assess the surgical outcomes in patients with or without process of LDD after TLIF surgery. Correlation analysis was performed to investigate associations between LDD and endplate HU value. Binary logistic regression analysis was carried out to study relationships between the DDD and the multiple risk factors. RESULTS: There was a statistically significant correlation between LDD, body mass index (BMI), age, paraspinal muscle atrophy, and total endplate scores (TEPS). Also, a strong and independent association between endplate HU value and LDD was found at every lumbar disc level (p < 0.01). After conditioning on matching factors, multivariate logistic regression analysis showed that higher endplate HU (odds ratio [OR]: 1.003, p = 0.003), higher TEPS (OR: 1.264, p = 0.002), higher BMI (odds ratio [OR]: 1.202, p = 0.002), a smaller cross-sectional area (CSA) of the paraspinal muscle preoperatively (OR: 0.096, p < 0.001) were significant predictors of LDD development after TLIF surgery. CONCLUSIONS: There is a significant association between LDD and endplate HU value after TLIF surgery in patients with degenerative lumbar stenosis. Beyond that, results from this study provide a mechanism by which high endplate HU value predisposes to LDD after TLIF surgery.

Sesn2 Serves as a Regulator between Mitochondrial Unfolded Protein Response and Mitophagy in Intervertebral Disc Degeneration.

Mitochondrial unfold protein response (UPRmt) can induce mitophagy to protect cell from unfold protein. However, how UPRmt induces mitophagy to protect cell is not yet clear. Herein, Sesn2 was considered to be a key molecule that communicated UPRmt and mitophagy in the intervertebral disc. Silencing of Sesn2 was able to reverse the protective effects of Nicotinamide riboside (NR) on nucleus pulposus (NP) cells and inhibit mitophagy induced by UPRmt. UPRmt upregulated Sesn2 through Eif2ak4/eIF2α/Atf4, and further induced mitophagy. Sesn2 promoted the translocation of cytosolic Parkin and Sqstm1 to the defective mitochondria respectively, thereby enhancing mitophagy. The translocation of cytosolic Sqstm1 to the defective mitochondria was dependent on Parkin. The two functional domains of Sesn2 were necessary for the interaction of Sesn2 with Parkin and Sqstm1. The cytosolic interaction of Sesn2 between Parkin and Sqstm1 was independent on Pink1 (named as PINK1 in human) but the mitochondrial translocation was dependent on Pink1. Sesn2-/- mice showed a more severe degeneration and NR did not completely alleviate the intervertebral disc degeneration (IVDD) of Sesn2-/- mice. In conclusion, UPRmt could attenuate IVDD by upregulation of Sesn2-induced mitophagy. This study will help to further reveal the mechanism of Sesn2 regulating mitophagy, and open up new ideas for the prevention and treatment of IVDD.