Serglycin secreted by late-stage nucleus pulposus cells is a biomarker of intervertebral disc degeneration.

Intervertebral disc degeneration is a natural process during aging and a leading cause of lower back pain. Here, we generate a comprehensive atlas of nucleus pulposus cells using single-cell RNA-seq analysis of human nucleus pulposus tissues (three males and four females, age 41.14 ± 18.01 years). We identify fibrotic late-stage nucleus pulposus cells characterized by upregulation of serglycin expression which facilitate the local inflammatory response by promoting the infiltration of inflammatory cytokines and macrophages. Finally, we discover that daphnetin, a potential serglycin ligand, substantially mitigates the local inflammatory response by downregulating serglycin expression in an in vivo mouse model, thus alleviating intervertebral disc degeneration. Taken together, we identify late-stage nucleus pulposus cells and confirm the potential mechanism by which serglycin regulates intervertebral disc degeneration. Our findings indicate that serglycin is a latent biomarker of intervertebral disc degeneration and may contribute to development of diagnostic and therapeutic strategies.

Acetylshikonin promoting PI3K/Akt pathway and inhibiting SOX4 expression to delay intervertebral disc degeneration and low back pain.

This study investigated the molecular mechanism by which acetylshikonin inhibits SOX4 expression via the PI3K/Akt pathway to delay intervertebral disc degeneration (IVDD) and low back pain (LBP). Bulk RNA-seq, RT-qPCR, Western blot analysis, immunohistochemical staining, small interfering RNA (siSOX4), lentivirus (lentiv-SOX4hi ), and imaging techniques were used to assess SOX4 expression and validate its upstream regulatory pathway. Acetylshikonin and siSOX4 were injected into the IVD to measure IVDD. SOX4 expression significantly increased in degenerated IVD tissues. TNF-α increased SOX4 expression and apoptosis-related proteins in nucleus pulposus cells (NPCs). siSOX4 reduced TNF-α-induced NPCs apoptosis, while Lentiv-SOX4hi increased it. The PI3K/Akt pathway was significantly correlated with SOX4, and acetylshikonin upregulated PI3K/Akt pathway while inhibiting SOX4 expression. In the anterior puncture IVDD mouse model, SOX4 expression was upregulated, and acetylshikonin and siSOX4 delayed IVDD-induced LBP. Acetylshikonin delays IVDD-induced LBP by inhibiting SOX4 expression through the PI3K/Akt pathway. These findings offer potential therapeutic targets for future treatments.

Clinical atlas of rectal cancer highlights the barriers and insufficient interventions underlying the unfavorable outcomes in older patients.

Background: Aging confers an increased risk of developing cancer, and the global burden of cancer is cumulating as human longevity increases. Providing adequate care for old patients with rectal cancer is challenging and complex. Method: A total of 428 and 44,788 patients diagnosed with non-metastatic rectal cancer from a referral tertiary care center (SYSU cohort) and the Surveillance Epidemiology and End Results database (SEER cohort) were included. Patients were categorized into old (over 65 years) and young (aged 50-65 years) groups. An age-specific clinical atlas of rectal cancer was generated, including the demographic and clinicopathological features, molecular profiles, treatment strategies, and clinical outcomes. Results: Old and young patients were similar in clinicopathological risk factors and molecular features, including TNM stage, tumor location, tumor differentiation, tumor morphology, lymphovascular invasion, and perineural invasion. However, old patients had significantly worse nutritional status and more comorbidities than young patients. In addition, old age was independently associated with less systemic cancer treatment (adjusted odds ratio 0.294 [95% CI 0.184-0.463, P < 0.001]). We found that old patients had significantly worse overall survival (OS) outcomes in both SYSU (P < 0.001) and SEER (P < 0.001) cohorts. Moreover, the death and recurrence risk of old patients in the subgroup not receiving chemo/radiotherapy (P < 0.001 for OS, and P = 0.046 for time to recurrence [TTR]) reverted into no significant risk in the subgroup receiving chemo/radiotherapy. Conclusions: Although old patients had similar tumor features to young patients, they had unfavorable survival outcomes associated with insufficient cancer care from old age. Specific trials with comprehensive geriatric assessment for old patients are needed to identify the optimal treatment regimens and improve unmet cancer care. Study registration: The study was registered on the research registry with the identifier of researchregistry 7635.

Genomic G-quadruplex folding triggers a cytokine-mediated inflammatory feedback loop to aggravate inflammatory diseases.

DNA G-quadruplex is a non-canonical secondary structure that could epigenetically regulate gene expression. To investigate the regulating role of G-quadruplex, we devised an integrating method to perform the algorithm profiling and genome-wide analysis for the dynamic change of genomic G-quadruplex and RNA profiles in rat nucleus pulposus cells by inducing G-quadruplex folding with multiple stabilizers. A group of genes potentially regulated by G-quadruplex and involved in the inflammation process has been identified. We found that G-quadruplex folding triggers inflammation response by upregulating inflammatory cytokines, which could promote G-quadruplex folding in a manner of positive feedback loop. Moreover, we confirmed that G-quadruplex is a marker indicating elevated inflammatory status and G-quadruplex folding facilitates the development of inflammatory diseases using in vivo intervertebral disc degeneration models. The crosstalk between G-quadruplex and inflammatory cytokines plays a vital role in regulating inflammation-derived diseases, which may provide new insights into the blocking target.

Circular RNA hsa_circ_0083756 promotes intervertebral disc degeneration by sponging miR-558 and regulating TREM1 expression.

OBJECTIVES: Intervertebral disc degeneration (IVDD) is a leading cause of low back pain. Circular RNAs (circRNAs) have been demonstrated to exert vital functions in IVDD. However, the role and mechanism of hsa_circ_0083756 in the development of IVDD remain unclear. MATERIALS AND METHODS: RT-qPCR was performed to detect expressions of hsa_circ_0083756, miR-558 and TREM1 in nucleus pulposus (NP) tissues and cells. CCK8 assay, flow cytometry, TUNEL assay, RT-qPCR and WB were used to clarify the roles of hsa_circ_0083756 in NP cells proliferation and extracellular matrix (ECM) formation. Bioinformatics analyses, dual-luciferase reporter gene experiment, RNA immunoprecipitation (RIP) assay and FISH assay were performed to predict and verify the targeting relationship between hsa_circ_0083756 and miR-558, as well as that between miR-558 and TREM1. Ultimately, the effect of hsa_circ_0083756 on IVDD was tested through anterior disc-puncture IVDD animal model in rats. RESULTS: hsa_circ_0083756 was upregulated in degenerative NP tissues and cells. In vitro loss-of-function and gain-of-function studies suggested that hsa_circ_0083756 knockdown promoted, whereas hsa_circ_0083756 overexpression inhibited NP cells proliferation and ECM formation. Mechanistically, hsa_circ_0083756 acted as a sponge of miR-558 and subsequently promoted the expression of TREM1. Furthermore, in vivo study indicated that silencing of hsa_circ_0083756 could alleviate IVDD in rats. CONCLUSIONS: hsa_circ_0083756 promoted IVDD via targeting the miR-558/TREM1 axis, and hsa_circ_0083756 may serve as a potential therapeutic target for the treatment of IVDD.

Grem1 accelerates nucleus pulposus cell apoptosis and intervertebral disc degeneration by inhibiting TGF-ß-mediated Smad2/3 phosphorylation.

Intervertebral disc degeneration (IVDD) is a main cause of low back pain, and inflammatory factors play key roles in its pathogenesis. Gremlin-1 (Grem1) was reported to induce an inflammatory response in other fields. This study aimed to investigate the mechanisms of Grem1 in the degenerative process of intervertebral discs. Dysregulated genes were determined by analyzing microarray profiles. The expression of Grem1 in 17 human disc samples (male:female = 9:8) and rat models (n = 5 each group) was measured by western blotting (WB), real-time quantitative PCR (RT-qPCR), and immunohistochemistry (IHC). The regulatory effects of Grem1 on apoptosis were examined using siRNAs, flow cytometry, immunofluorescence (IF), and WB. The therapeutic effect was evaluated by locally injecting specific Grem1 siRNA into IVDD rats. The expression of Grem1 was significantly increased in human degenerative intervertebral discs; furthermore, the expression of Grem1 positively correlated with the level of intervertebral disc degeneration. Grem1 was significantly overexpressed in tumor necrosis factor (TNF)-α-induced degenerative NP cells. Apoptosis in degenerative NP cells transfected with siRNA targeting Grem1 was significantly lower than that in the control group. Specific Grem1 siRNA markedly repressed the development of IVDD in surgery-induced IVDD rats. These results indicated that the expression of Grem1 was positively correlated with the severity of intervertebral disc degeneration, and Grem1 siRNA could inhibit Grem1-induced apoptosis and extracellular matrix alterations by mediating the TGF-ß/Smad signaling pathway. This study may provide a therapeutic strategy for alleviating inflammation-induced apoptosis associated with intervertebral disc degeneration.

Visfatin promotes intervertebral disc degeneration by inducing IL-6 expression through the ERK/JNK/p38 signalling pathways.

Visfatin reportedly induces the expression of proinflammatory cytokines. Severe grades of intervertebral disc disease (IVDD) exhibit higher expression of visfatin than mild ones. However, the direct relationship between visfatin and IVDD remains to be elucidated. This study aimed to clarify whether stimulation of visfatin in IVDD is mediated by IL-6. To investigate the role of visfatin in IVDD, a rat model of anterior disc puncture was established by injecting visfatin or PBS using a 27-gauge needle. Results revealed an obvious aggravation of the histological morphology of IVDD in the visfatin group. On treating human NP cellswith visfatin, the levels of collagenII and aggrecan decreased and those of matrix metallopeptidase 3 and IL-6 gradually increased. A rapid increase in ERK, JNK, and p38 phosphorylation was also noted after visfatin treatment. Compared to those treated with visfatin alone, NP cells pretreated with ERK1/2, JNK, and p38 inhibitors or siRNA targeting p38, ERK, and JNK exhibited a significant suppression of IL-6. Our data represent the first evidence that visfatin promotes IL-6 expression in NP cells via the JNK/ERK/p38-MAPK signalling pathways. Further, our findings suggest epidural fat and visfatin as potential therapeutic targets for controlling IVDD-associated inflammation.

Role of the miR-133a-5p/FBXO6 axis in the regulation of intervertebral disc degeneration.

OBJECTIVE: Low back pain is a leading cause of disabilities worldwide, and intervertebral disc degeneration (IVDD)-related disorders have been recognised as one of the main contributors. Nevertheless, the underlying mechanism has not yet been fully understood. The aim of this study was to investigate the role of the miR-133a-5p/FBXO6 axis in the regulation of IVDD. METHODS: RT-qPCR, WB and IHC were performed to assess the expression of FBXO6 in human IVD tissues. Nucleus pulposus (NP) cells were treated with IL-1ß to induce IVDD cellular model. Silence of FBXO6 was achieved using specific siRNAs. CCK-8 assay, flow cytometry, TUNEL assay, RT-qPCR and WB were used to evaluate the role and mechanism of FBXO6 in the process of IVDD. Online tools, GSE datasets and RT-qPCR were used to search the candidate miRNAs targeting FBXO6. The direct binding sites between FBXO6 and miR-133a-5p were further verified by a dual luciferase assay. RT-qPCR, WB and rescue experiments were conducted to identify the regulatory function of miR-133a-5p on the expression of aggrecan, collagen Ⅱ, MMP3, ADAMTS5, IL-6 and COX2. In addition, the role of the NF-κB pathway in regulating miR-133a-5p was studied using lentiviral shRNA, WB and RT-qPCR. RESULTS: Results showed that FBXO6 mainly expressed in the NP tissue of IVD and the expression of FBXO6 decreased with the process of IVDD as well as under IL-1ß stimulation. The silence of FBXO6 led to the decreased expression of aggrecan and collagen Ⅱ and the increased expression of MMP3, ADAMTS5, IL-6 and COX2, which further induced the degeneration of NP cells. The bioinformatic analysis showed that miR-133a-5p was the candidate miRNA targeting FBXO6. miR-133a-5p was upregulated in IVDD tissues and significantly inhibited the expression of FBXO6. The inhibition of miR-133a-5p ameliorated the acceleration of IVDD induced by the silence of FBXO6 in vitro. Moreover, it was demonstrated that IL-1ß regulated the expression of the miR-133a-5p/FBXO6 axis via the NF-κB pathway in NP cells. CONCLUSION: miR-133a-5p was upregulated by IL-1ß to aggravate intervertebral disc degeneration via sponging FBXO6. Inhibiting miR-133a-5p expression or rescuing FBXO6 expression may be promising strategies for the treatment of IVDD. THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE: This study suggests that the miR-133a-5p/FBXO6 axis could regulate NP cells proliferation, apoptosis, synthesis and degradation of extracellular matrix, which provides a promising therapeutic target and strategy for the treatment of IVDD.

Effects of rosuvastatin and zoledronic acid in combination on the recovery of senile osteoporotic vertebral compression fracture following percutaneous vertebroplasty.

OBJECTIVES: This study analyzed the effects of rosuvastatin and zoledronic acid in combination on patient recovery following percutaneous vertebroplasty (PVP) that was performed to treat senile osteoporotic vertebral compression. METHODS: Senile patients with osteoporotic vertebral compression fracture (n = 120) were included in this retrospective study, and they were classified into two groups. Those in the control group (n = 60) were treated with PVP + caltrate and those in the observation group (n = 60) received this treatment with combined zoledronic acid and rosuvastatin. Between-group comparisons were made at both pre- and post-treatment regarding bone density, type I procollagen peptide (CTX) and bone-specific alkaline phosphatase (BAP) levels, visual analog scale (VAS) score, Oswestry Disability Index (ODI) score, and adjacent centrum refracture. RESULTS: Bone density was higher and BAP and CTX levels as well as ODI and VAS scores were lower at post-treatment in the observation group compared with the control group. The refracture rate in the observation group was lower compared with the control group. CONCLUSION: Treatment with a combination of rosuvastatin and zoledronic acid following PVP can improve the condition of senile osteoporotic vertebral compression fracture and patient's functional status, and it can also alleviate pain.

Effect of surgical factors on the augmentation of cement-injectable cannulated pedicle screw fixation by a novel calcium phosphate-based nanocomposite.

Bone cement-augmented pedicle screw system demonstrates great efficacy in spinal disease treatments. However, the intrinsic drawbacks associated with clinically used polymethylmethacrylate (PMMA) cement demands for new bone cement formulations. On the basis of our previous studies, a novel injectable and biodegradable calcium phosphate-based nanocomposite (CPN) for the augmentation of pedicle screw fixation was systematically evaluated for its surgical feasibility and biomechanical performance by simulated and animal osteoporotic bone models, and the results were compared with those of clinical PMMA cement. ASTM-standard solid foam and open-cell foam models and decalcified sheep vertebra models were employed to evaluate the augmentation effects of CPN on bone tissue and on the cement-injected cannulated pedicle screws (CICPs) placed in osteoporotic bone. Surgical factors in CICPs application, such as injection force, tapping technique, screw diameter, and pedicle screw loosening scenarios, were studied in comparison with those in PMMA. When directly injected to the solid foam model, CPN revealed an identical augmentation effect to that of PMMA, as shown by the similar compressive strengths (0.73 ± 0.04 MPa for CPN group vs. 0.79 ± 0.02 MPa for PMMA group). The average injection force of CPN at approximately 40-50 N was higher than that of PMMA at approximately 20 N. Although both values are acceptable to surgeons, CPN revealed a more consistent injection force pattern than did PMMA. The dispersing and anti-pullout ability of CPN were not affected by the surgical factors of tapping technique and screw diameter. The axial pullout strength of CPN evaluated by the decalcified sheep vertebra model revealed a similar augmentation level as that of PMMA (1351.6 ± 324.2 N for CPN vs. 1459.7 ± 304.4 N for PMMA). The promising results of CPN clearly suggest its potential for replacing PMMA in CICPs augmentation application and the benefits of further study and development for clinical uses.