Quercetin ameliorates oxidative stress-induced senescence in rat nucleus pulposus-derived mesenchymal stem cells via the miR-34a-5p/SIRT1 axis.

BACKGROUND: Intervertebral disc degeneration (IDD) is a main contributor to low back pain. Oxidative stress, which is highly associated with the progression of IDD, increases senescence of nucleus pulposus-derived mesenchymal stem cells (NPMSCs) and weakens the differentiation ability of NPMSCs in degenerated intervertebral discs (IVDs). Quercetin (Que) has been demonstrated to reduce oxidative stress in diverse degenerative diseases. AIM: To investigate the role of Que in oxidative stress-induced NPMSC damage and to elucidate the underlying mechanism. METHODS: In vitro, NPMSCs were isolated from rat tails. Senescence-associated ß-galactosidase (SA-ß-Gal) staining, cell cycle, reactive oxygen species (ROS), real-time quantitative polymerase chain reaction (RT-qPCR), immunofluorescence, and western blot analyses were used to evaluated the protective effects of Que. Meanwhile the relationship between miR-34a-5p and Sirtuins 1 (SIRT1) was evaluated by dual-luciferase reporter assay. To explore whether Que modulates tert-butyl hydroperoxide (TBHP)-induced senescence of NPMSCs via the miR-34a-5p/SIRT1 pathway, we used adenovirus vectors to overexpress and downregulate the expression of miR-34a-5p and used SIRT1 siRNA to knockdown SIRT1 expression. In vivo, a puncture-induced rat IDD model was constructed, and X rays and histological analysis were used to assess whether Que could alleviate IDD in vivo. RESULTS: We found that TBHP can cause NPMSCs senescence changes, such as reduced cell proliferation ability, increased SA-ß-Gal activity, cell cycle arrest, the accumulation of ROS, and increased expression of senescence-related proteins. While abovementioned senescence indicators were significantly alleviated by Que treatment. Que decreased the expression levels of senescence-related proteins (p16, p21, and p53) and senescence-associated secreted phenotype (SASP), including IL-1ß, IL-6, and MMP-13, and it increased the expression of SIRT1. In addition, the protective effects of Que on cell senescence were partially reversed by miR-34a-5p overexpression and SIRT1 knockdown. In vivo, X-ray, and histological analyses indicated that Que alleviated IDD in a puncture-induced rat model. CONCLUSION: In summary, the present study provides evidence that Que reduces oxidative stress-induced senescence of NPMSCs via the miR-34a/SIRT1 signaling pathway, suggesting that Que may be a potential agent for the treatment of IDD.

Safety and Efficacy of Local Steroid Application on Dysphagia Following Anterior Cervical Discectomy and Fusion: A Systematic Review and Meta-analysis.

STUDY DESIGN: A systematic review and meta-analysis. OBJECTIVE: To evaluate the safety and efficacy of local steroid application (LSA) on dysphagia after anterior cervical discectomy and fusion (ACDF). SUMMARY OF BACKGROUND DATA: Dysphagia is one of the most common adverse events in the early postoperative period of ACDF. LSA is reported as an effective method to reduce the swelling of soft tissues, thereby decreasing the incidence of dysphagia. However, the safety and efficacy of LSA on dysphagia after ACDF need to be systematically reviewed and analyzed. METHODS: A comprehensive literature search was carried out in the database PubMed, Web of Science, EMBASE, Clinical key, Cochrane library, and Wiley Online Library to screen papers that report LSA in ACDF surgery. The Cochrane Collaboration tool and a methodological index for nonrandomized studies were used for the assessment of study quality. Data were analyzed with the Review Manager 5.3 software. RESULTS: A total of 10 studies were included. The results revealed no significant differences between the steroid group and the control group in ACDF regarding postoperative drainage, estimated blood loss, and neck disability index score ( P > 0.05). LSA significantly alleviates visual analog scale score for neck pain (or odynophagia) ( P < 0.05), reduces the length of hospital stay (weighted mean difference, -1.00 (-1.05 to -0.95); P < 0.001), and mitigates dysphagia rate and prevertebral soft-tissue swelling in the early postoperative period ( P < 0.05). There seemed to be no significant increase in the complication rate and steroid-related adverse events in the steroid group compared with the control group ( P < 0.05). CONCLUSIONS: LSA shows advantages in reducing the length of hospital stay, decreasing dysphagia rate, and mitigating prevertebral soft-tissue swelling in the early postoperative period of ACDF. Further large-scale studies are urgently required for the development of a standard protocol for LSA and further analysis of potential delay complications.

Influence of the facet joint angle on facet joint degeneration following pedicle screw fixation without fusion in thoracolumbar fractures.

BACKGROUND: Posterior approach pedicle screw fixation without fusion is widely used in the treatment of neurologically intact type A3 thoracolumbar fractures. OBJECTIVE: To analyze the influence of the facet joint (FJ) angle on FJ degeneration following posterior approach pedicle screw fixation without fusion in neurologically intact type A3 thoracolumbar fractures. METHODS: Fifty-eight patients who underwent pedicle screw fixation via the traditional posterior approach (n= 28) or the Wiltse approach (n= 30) were enrolled. A CT scan was performed before fixation and before fixation removal (Within 1.5 to 2 years after fixation) to evaluate the FJs parameters, including FJ inclination (FJI), FJ tropism (FJT), FJ violation, and FJ degeneration grade (FJDG), of three fixed segments and the adjacent segment below the fixed segments. RESULTS: There was no significant difference in FJ violation rate, FJDG deterioration, or FJ angle between the two groups (P> 0.05). FJDG deterioration showed a weak positive correlation with FJI and FJT before fixation, and the angular change in FJI (P< 0.05); and FJT before fixation and the angular change in FJI were risk factors for FJDG deterioration (P< 0.01). CONCLUSION: The Wiltse approach did not increase the rate of FJDG deterioration and FJs angle changes. However, the FJT before fixation and the angular change in FJI were risk factors for FJDG deterioration.

Reactive Oxygen Species Contributes to Type 2 Diabetic Neuropathic Pain via the Thioredoxin-Interacting Protein-NOD-Like Receptor Protein 3- N -Methyl-D-Aspartic Acid Receptor 2B Pathway.

BACKGROUND: The number of patients with diabetic neuropathic pain (DNP) continues to increase, but available treatments are limited. This study aimed to examine the influence of reactive oxygen species (ROS)-thioredoxin-interacting protein (TXNIP)-NOD-like receptor protein 3 (NLRP3)- N -methyl-D-aspartic acid receptor 2B (NR2B) pathway on type 2 DNP. METHODS: Male Sprague-Dawley rats were fed with a high-fat and high-sugar diet for 8 weeks. Then, rats were intraperitoneally injected with streptozotocin (STZ, 35 mg/kg) to induce type 2 diabetes mellitus in rats. Diabetic rats with <85% of their basic levels in mechanical withdrawal threshold and thermal withdrawal latency were classified as DNP rats on day 14 after STZ injection. DNP rats were treated with ROS scavenger N-tert-Butyl-α-phenylnitrone (PBN, 100 mg·kg -1 ·d -1 ) or TXNIP small interfering ribonucleic acid (10 µg/d) once daily for 14 days. The level of ROS, protein levels of NLRP3, TXNIP, cysteinyl aspartate-specific proteinase-1 (caspase-1), interleukin-1ß (IL-1ß), NR2B phosphorylation at Tyr1472 (p-NR2B), total NR2B (t-NR2B), and distribution of NLRP3 in the spinal cord were examined. In vitro experiments, BV2 cells and PC12 cells were individually cultured and cocultured in a high-glucose environment (35 mmol/L D-glucose). The level of ROS and protein levels of NLRP3, TXNIP, caspase-1, and IL-1ß in BV2 cells, and p-NR2B, t-NR2B in PC12 cells were detected. The level of ROS was detected by the flow cytometry approach. The protein levels were detected by the Western blot technique. The location of NLRP3 was observed by immunofluorescent staining. The interaction between TXNIP and NLRP3 was detected by coimmunoprecipitation assay. RESULTS: The level of spinal ROS increased in DNP rats. The mechanical allodynia and thermal hyperalgesia of DNP rats were alleviated after systemic administration of PBN. This administration decreased protein levels of NLRP3, TXNIP, caspase-1, IL-1ß, and p-NR2B and the coupling of TXNIP to NLRP3 in spinal cords of DNP rats. Furthermore, knockdown of spinal TXNIP alleviated nociceptive hypersensitivity and decreased protein levels of NLRP3, TXNIP, caspase-1, IL-1ß, and p-NR2B in DNP rats. The level of ROS and protein levels of NLRP3, TXNIP, caspase-1, IL-1ß, the coupling of TXNIP to NLRP3, and the IL-1ß secretion increased in BV2 cells, and the protein expression of p-NR2B increased in cocultured PC12 cells in a high-glucose environment. All of these in vitro effects were significantly blocked after treatment of PBN. CONCLUSIONS: Our findings suggest that spinal ROS can contribute to type 2 DNP through TXNIP-NLRP3-NR2B pathway.

1,25(OH)2D3 Mitigates Oxidative Stress-Induced Damage to Nucleus Pulposus-Derived Mesenchymal Stem Cells through PI3K/Akt Pathway.

Intervertebral disc degeneration (IVDD) is one of the main causes of low back pain. The local environment of the degenerated intervertebral disc (IVD) increases oxidative stress and apoptosis of endogenous nucleus pulposus-derived mesenchymal stem cells (NPMSCs) and weakens its ability of endogenous repair ability in degenerated IVDs. A suitable concentration of 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3) has been certified to reduce oxidative stress and cell apoptosis. The current study investigated the protective effect and potential mechanism of 1,25(OH)2D3 against oxidative stress-induced damage to NPMSCs. The present results showed that 1,25(OH)2D3 showed a significant protective effect on NPMSCs at a concentration of 10-10 M for 24 h. Protective effects of 1,25(OH)2D3 were also exhibited against H2O2-induced NPMSC senescence, mitochondrial dysfunction, and reduced mitochondrial membrane potential. The Annexin V/PI apoptosis detection assay, TUNEL assay, immunofluorescence, western blot, and real-time quantitative polymerase chain reaction assay showed that pretreatment with 1,25(OH)2D3 could alleviate H2O2-induced NPMSC apoptosis, including the apoptosis rate and the expression of proapoptotic-related (Caspase-3 and Bax) and antiapoptotic-related (Bcl-2) proteins. The intracellular expression of p-Akt increased after pretreatment with 1,25(OH)2D3. However, these protective effects of 1,25(OH)2D3 were significantly decreased after the PI3K/Akt pathway was inhibited by the LY294002 treatment. In vivo, X-ray, MRI, and histological analyses showed that 1,25(OH)2D3 treatment relieved the degree of IVDD in Sprague-Dawley rat disc puncture models. In summary, 1,25(OH)2D3 efficiently attenuated oxidative stress-induced NPMSC apoptosis and mitochondrial dysfunction via PI3K/Akt pathway and is a promising candidate treatment for the repair of IVDD.

Outcomes of Oblique Lateral Interbody Fusion for Adult Spinal Deformity: A Systematic Review and Meta-Analysis.

STUDY DESIGN: A systematic review and meta-analysis. OBJECTIVES: To evaluate clinical and radiographic outcomes, and perioperative complications of oblique lateral interbody fusion (OLIF) for adult spinal deformity (ASD). METHODS: We performed a systematic review and meta-analysis of related studies reporting outcomes of OLIF for ASD. The clinical outcomes were assessed by visual analogue scale (VAS) and Oswestry Disability Index (ODI). The radiographic parameters were evaluated by sagittal vertical axis (SVA), pelvic tilt (PT), sacral slope (SS), thoracic kyphosis (TK), lumbar lordosis (LL), pelvic incidence-lumbar lordosis (PI-LL), Cobb angle and fusion rate. A random effects model and 95% confidence intervals (CI) were performed to investigate the results. RESULTS: A total of 16 studies involving 519 patients were included in the present study. The mean difference of VAS-back score, VAS-leg score and ODI score before and after surgery was 5.1, 5.0 and 32.3 respectively. The mean correction of LL was 20.6°, with an average of 6.9° per level and the mean correction of Cobb was 16.4°, with an average of 4.7° per level. The mean correction of SVA, PT, SS, TK and PI-LL was 59.3 mm, 11.7°, 6.9°, 9.4° and 20.6° respectively. The mean fusion rate was 94.1%. The incidence of intraoperative and postoperative complications was 4.9% and 29.6% respectively. CONCLUSIONS: OLIF is an effective and safe surgery method in the treatment of mild or moderate ASD and it has advantages in less intraoperative blood loss and lower perioperative complications.

Perioperative Hidden Blood Loss in Lumbar Disk Herniation Patients With Percutaneous Endoscopic Transforaminal Discectomy and Influencing Factors.

STUDY DESIGN: This was a retrospective study. OBJECTIVES: This study aimed to evaluate hidden blood loss (HBL) and its influencing factors in lumbar disk herniation (LDH) patients treated with percutaneous endoscopic transforaminal discectomy (PETD). SUMMARY OF BACKGROUND DATA: PETD is a minimally invasive spine surgery and is widely used to treat LDH. It is generally believed that there is less bleeding during PETD. However, HBL during the perioperative period is always ignored. MATERIALS AND METHODS: From January 2018 to March 2021, 74 LDH patients treated with PETD was selected. The patient's sex, age, height, weight, previous medical history (hypertension and diabetes) and other basic information were recorded. The preoperative fibrinogen (FIB) level, activated partial thromboplastin time and prothrombin time were recoded. The hemoglobin, hematocrit, and platelet immediately after admission and the next day postoperative were recorded. The surgical time, intraoperative blood loss, intervertebral disk degeneration grade and soft tissue thickness of the PETD approach were recorded. The total blood loss was calculated according to the Gross formula, and then HBL was calculated based on total blood loss and visible blood loss (VBL). The influencing factors were analyzed by single factor correlation analysis and multivariate linear regression analysis. RESULTS: Among the 74 patients, there were 34 males (20-68 y old) and 40 females (26-75 y old). The mean amount of VBL was (85.04±26.53) mL and HBL was (341.04±191.15) mL. There were statistically significant differences between HBL and VBL (P=0.000). Multiple linear regression analysis showed that sex (P=0.000), disk degeneration grade (P=0.000), preoperative FIB level (P=0.022) and preoperative platelet (P=0.026) were independent risk factors that contributed to HBL, but age (P=0.870), BMI (P=0.480), hypertension (P=0.867), diabetes (P=0.284), soft tissue thickness (P=0.701), preoperative prothrombin time (P=0.248) and preoperative activated partial thromboplastin time (P=0.521) were not. CONCLUSIONS: There was a large amount of HBL during the perioperative period of PETD in patients with LDH. Sex, disk degeneration grade, preoperative FIB level and preoperative platelet are the independent risk factors of HBL in the perioperative period of PETD. More attention should be paid to the patients with risk factors to ensure perioperative safety.

Urolithin a alleviates oxidative stress-induced senescence in nucleus pulposus-derived mesenchymal stem cells through SIRT1/PGC-1α pathway.

BACKGROUND: In degenerative intervertebral disc (IVD), an unfavorable IVD environment leads to increased senescence of nucleus pulposus (NP)-derived mesenchymal stem cells (NPMSCs) and the inability to complete the differentiation from NPMSCs to NP cells, leading to further aggravation of IVD degeneration (IDD). Urolithin A (UA) has been proven to have obvious effects in delaying cell senescence and resisting oxidative stress. AIM: To explore whether UA can alleviate NPMSCs senescence and to elucidate the underlying mechanism. METHODS: In vitro, we harvested NPMSCs from rat tails, and divided NPMSCs into four groups: the control group, H2O2 group, H2O2 + UA group, and H2O2 + UA + SR-18292 group. Senescence-associated ß-Galactosidase (SA-ß-Gal) activity, cell cycle, cell proliferation ability, and the expression of senescence-related and silent information regulator of transcription 1/PPAR gamma coactivator-1α (SIRT1/ PGC-1α) pathway-related proteins and mRNA were used to evaluate the protective effects of UA. In vivo, an animal model of IDD was constructed, and X-rays, magnetic resonance imaging, and histological analysis were used to assess whether UA could alleviate IDD in vivo. RESULTS: We found that H2O2 can cause NPMSCs senescence changes, such as cell cycle arrest, reduced cell proliferation ability, increased SA-ß-Gal activity, and increased expression of senescence-related proteins and mRNA. After UA pretreatment, the abovementioned senescence indicators were significantly alleviated. To further demonstrate the mechanism of UA, we evaluated the mitochondrial membrane potential and the SIRT1/PGC-1α pathway that regulates mitochondrial function. UA protected mitochondrial function and delayed NPMSCs senescence by activating the SIRT1/PGC-1α pathway. In vivo, we found that UA treatment alleviated an animal model of IDD by assessing the disc height index, Pfirrmann grade and the histological score. CONCLUSION: In summary, UA could activate the SIRT1/PGC-1α signaling pathway to protect mitochondrial function and alleviate cell senescence and IDD in vivo and vitro.

Research on decision-making of emergency plan for waterlogging disaster in subway station project based on linguistic intuitionistic fuzzy set and TOPSIS.

Targeted at emergency plans for rainstorm and waterlogging disasters in subway station projects, this work proposes a group decision-making method that uses linguistic intuitionistic fuzzy sets, structural entropy weights, and TOPSIS. An evaluation index system of emergency plans was constructed based on four aspects, namely a scientific basis, completeness, operability, and flexibility. A linguistic interval intuitionistic fuzzy set approach was then used to qualitatively present the decision-makers' understanding of, attitudes about, and preferences for emergency plans. The uncertainty was comprehensively and intuitively represented by the dimensions of the degrees of membership and non-membership. The structural entropy weight method was applied and improved to fully reflect the influences of experts with different characteristics on the index weights. Finally, the TOPSIS method, with a background context of linguistic interval intuitionistic fuzzy sets, was applied. The calculation results of benchmark and verification case highlight the rationality and scientificity of the method proposed in this paper. The emergency decisions regarding waterlogging in 2018 for the Huilong Road West Station Project of Chengdu Metro Line 11 in China were selected as a case study. The case study demonstrates that operability is the most critical of the four primary indicators, and that flexible response to changes in the emergency response level is the most important of the secondary indicators. The uncertainty analysis of data revealed that with the increase of uncertainty, the difference between each scheme and the ideal solution decreased. Compared with the classical TOPSIS method, the new model proposed in this paper is robust and effective, and can be used for similar projects in the future.

Kalirin-7 contributes to type 2 diabetic neuropathic pain via the postsynaptic density-95/N-methyl-D-aspartate receptor 2B-dependent N-methyl-D-aspartate receptor 2B phosphorylation in the spinal cord in rats.

OBJECTIVE: Diabetic neuropathic pain (DNP) is one of the common complications in type 2 Diabetes Mellitus (DM) patients. However, molecular mechanisms in underlying diabetic neuropathic pain are still poorly understood. Kalirin-7, a multifunctional Rho GDP/GTP exchange factor, located at the excitatory synapses, was reported to modulate the neuronal cytoskeleton. Therefore, in this study, we explored the effects of Kalirin-7 on type 2 diabetic neuropathic pain and the mechanisms in spinal cord in rats. METHODS: The type 2 diabetic neuropathic pain model was established in rats by feeding them with a high-sugar and high-fat diet for 8 weeks, and then fasting them for 12 hours, followed by a single intraperitoneal injection of STZ. Kalirin-7 was knocked down in the spinal cord by an intrathecal administration of Kalirin-7 siRNA. RESULTS: The levels of Kalirin-7, p-NR2B and PSD-95 as well as the PSD-95-NR2B coupling were significantly increased in the spinal cord of type 2 DM rats. The knockdown of Kalirin-7 expression in the spinal cord by the intrathecal administration of Kalirin-7 siRNA not only reduced the levels of p-NR2B and the PSD-95-NR2B coupling in the spinal cord, but also relieved mechanical allodynia and thermal hyperalgesia in type 2 DM rats. CONCLUSIONS: Our findings suggest that spinally expressed Kalirin-7 likely contributes to type 2 diabetic neuropathic pain through regulating the PSD-95/NR2B interaction-dependent NR2B phosphorylation in the spinal cord.

Caveolin-1 in spinal cord modulates type-2 diabetic neuropathic pain through the Rac1/NOX2/NR2B signaling pathway.

OBJECTIVE: The present study determines whether Cav-1 modulates the initiation, development and maintenance of type-2 DNP via the Rac1/NOX2-NR2B signaling pathway. METHODS: After regular feeding for three days, these rats were randomly divided into two groups: control group with normal-diet (maintenance feed) (n=8); type-2 DM group (n=8). In the type-2 DM group, the rats were fed with a high-fat and high-sugar diet, and received a single intraperitoneal streptozotocin (STZ) injection (35 mg/kg). At two weeks after STZ injection, these diabetic neuropathic pain (DNP) rats were treated with daidzein (0.4 mg/kg/day) and N-tert-Butyl-α-phenylnitrone (PBN, 100 mg/kg/day) for 14 days. After the type-2 DNP model was successfully established, the rats were assigned into four groups: DNP group, DNP+Da group (DNP rats with Cav-1 specific inhibitor daidzein), DNP+PBN group (DNP rats treated with ROS scavenger PBN), and SC group (solvent control group). Then, the mechanical and thermal hyperalgesia were assayed to evaluate the function of the caveolin 1-Recombinant Human Ras-Related C1/nicotinamide adenosine diphosphate oxidase 2-NR2B gene (Cav-1-Rac1/NOX2-NR2B) signaling pathway. In the mechanism study, the protein expression levels of p-Caveolin-1, Rac1, NOX2, p-NR2B and t-NR2B, the production of ROS, and the distribution of Cav-1 and NOX2 in the spinal cord were observed. RESULTS: The present study revealed that p-Cav-1 was persistently upregulated and activated in the spinal cord microglia in type-2 DNP rats. The use of the pharmacological inhibitor of Cav-1 and a ROS scavenger resulted to a significantly relieved mechanical allodynia and thermal hyperalgesia. In addition, it was demonstrated that Cav-1 promoted ROS generation via the activation of Rac1-dependent NADPH oxidase (NOX). CONCLUSION: The present data suggests that Cav-1 in the spinal cord modulates type-2 DNP via regulating the Rac1/NOX2-NR2B pathway.

Activation of CaMKII and GluR1 by the PSD-95-GluN2B Coupling-Dependent Phosphorylation of GluN2B in the Spinal Cord in a Rat Model of Type-2 Diabetic Neuropathic Pain.

The mechanisms underlying type-2 diabetic neuropathic pain (DNP) are unclear. This study investigates the coupling of postsynaptic density-95 (PSD-95) to N-methyl-D-aspartate receptor subunit 2B (GluN2B), and the subsequent phosphorylation of GluN2B (Tyr1472-GluN2B) in the spinal cord in a rat model of type-2 DNP. Expression levels of PSD-95, Tyr1472-GluN2B, Ca2+/calmodulin-dependent protein kinase II (CaMKII) and its phosphorylated counterpart (Thr286-CaMKII), and α-amino-3-hydroxy-5-methyl-4-soxazole propionic acid receptor subtype 1 (GluR1) and its phosphorylated counterpart (Ser831-GluR1) were significantly increased versus controls in the spinal cord of type-2 DNP rats whereas the expression of total spinal GluN2B did not change. The intrathecal injection of Ro25-6981 (a specific antagonist of GluN2B) or Tat-NR2B9c (a mimetic peptide disrupting the interaction between PSD-95 and GluN2B) induced an antihyperalgesic effect and blocked the increased expression of Tyr1472-GluN2B, CaMKII, GluR1, Thr286-CaMKII, and Ser831-GluR1 in the spinal cords; the increase in spinal cord PSD-95 was not affected. These findings indicate that the PSD-95-GluN2B interaction may increase phosphorylation of GluN2B, and subsequently induce the expression of phosphorylation of CaMKII and GluR1 in the spinal cord of type-2 DNP rats. Targeting the interaction of PSD-95 with GluN2B may provide a new therapeutic strategy for type-2 DNP.