Enhancer RNAs (eRNAs) are noncoding RNAs that synthesized at active enhancers. eRNAs have important regulatory characteristics and appear to be significant for maintenance of cell identity and information processing. Series of functional eRNAs have been identified as potential therapeutic targets for multiple diseases. Nevertheless, the role of eRNAs on intervertebral disc degeneration (IDD) is still unknown yet. Herein, we utilized the nucleus pulposus samples of patients and identified a key eRNA (LINC02569) with the Arraystar eRNA Microarray. LINC02569 mostly locates in nucleus and plays an important role in the progress of IDD by activating nuclear factor kappa-B (NF-κB) signaling pathway. We used a cationic polymer brush coated carbon nanotube (oCNT-pb)-based siRNA delivery platform that we previously designed, to transport LINC02569 siRNA (si-02569) to nucleus pulposus cells. The siRNA loaded oCNT-pb accumulated in nucleus pulposus cells with lower toxicity and higher transfection efficiency, compared with the traditional siRNA delivery system. Moreover, the results showed that the delivery of si-02569 significantly alleviated the inflammatory response in the nucleus pulposus cells via inhibiting P65 phosphorylation and preventing its transfer into the nucleus, and meanwhile alleviated cell senescence by decreasing the expression of P21. Altogether, our results highlight that eRNA (LINC02569) plays important role in the progression of IDD and could be a potential therapeutic target for alleviation of IDD.

Intervertebral disc degeneration (IDD) has been considered as the primary pathological mechanism that underlies low back pain. Understanding the molecular mechanisms underlying human IDD is imperative for making strategies to treat IDD-related diseases. Herein, we report the molecular programs, lineage progression patterns, and paths of cellular communications during the progression of IDD using single-cell RNA sequencing (scRNA-seq) on nucleus pulposus (NP) cells from patients with different grades of IDD undergoing discectomy. New subtypes of cells and cell-type-specific gene signatures of the metabolic homeostatic NP cells (Met NPC), adhesive NP cells (Adh NPC), inflammatory response NP cells (IR NPC), endoplasmic reticulum stress NP cells (ERS NPC), fibrocartilaginous NP cells (Fc NPC), and CD70 and CD82+ progenitor NP cells (Pro NPC) were identified. In the late stage of IDD, the IR NPC and Fc NPC account for a large proportion of NPC. Importantly, immune cells including macrophages, T cells, myeloid progenitors, and neutrophils were also identified, and further analysis showed that significant intercellular interaction between macrophages and Pro NPC occurred via MIF (macrophage migration inhibitory factor) and NF-kB signaling pathways during the progression of IDD. In addition, dynamic polarization of macrophage M1 and M2 cell subtypes was found in the progression of IDD, and gene set functional enrichment analysis suggested a significant role of the macrophage polarization in regulating cell metabolism, especially the Pro NPC. Finally, we found that the NP cells in the late degenerative stage were mainly composed of the cell types related to inflammatory and endoplasmic reticulum (ER) response, and fibrocartilaginous activity. Our results provided new insights into the identification of NP cell populations at single-cell resolution and at the relatively whole-transcriptome scale, accompanied by cellular communications between immune cells and NP cells, and discriminative markers in relation to specific cell subsets. These new findings present clues for effective and functional manipulation of human IDD-related bioremediation and healthcare.

Supplemental Digital Content is available in the text This in vitro experimental study demonstrated that not only structural disruption, but also a single impact injury of the endplate without structural impairment could also initiate intervertebral disc degeneration (IDD), which might be mediated by activation of Piezo1 induced inflammation and abnormal energy metabolism of intervertebral disc cells. Study Design. In vitro experimental study. Objective. To establish an axial impact injury model of intervertebral disc (IVD) and to investigate if a single impact injury without endplate structural disruption could initiate intervertebral disc degeneration (IDD), and what is the roles of Piezo1 in this process. Summary of Background Data. Although IDD process has been confirmed to be associated with structural failures such as endplate fractures, whether a single impact injury of the endplates without structural disruption could initiate IDD remains controversial. Previous studies reported that Piezo1 mediated inflammation participated in the progression of IDD induced by mechanical stretch; however, the roles of Piezo1 in IVD impact injury remain unknown. Methods. Rats spinal segments were randomly assigned into Control, Low, and High Impact groups, which were subjected to pure axial impact loading using a custom-made apparatus, and cultured for 14 days. The degenerative process was investigated by using histomorphology, real-time Polymerase Chain Reaction(PCR), western-blot, immunofluorescence, and energy metabolism of IVD cell. The effects of Piezo1 were investigated by using siRNA transfection, real-time PCR, western-blot, and immunofluorescence. Results. The discs in both of the impact groups presented degenerative changes after 14 days, which showed significant up-regulation of Piezo1, NLRP3 inflammasome, the catabolic (MMP-9, MMP-13), and pro-inflammatory gene (IL-1β) expression than that of the control group (P < 0.05), accompanied by significantly increased release of ATP, lactate, nitric oxide (NO), and glucose consumption of IVD cells at first 7 days. Silencing Piezo1 reduced the activation of NLRP3 inflammasome and IL-1β expression in the nucleus pulposus induced by impact injury. Conclusion. It demonstrated that not only fracture of the endplate but also a single impact injury without structural impairment could also initiate IDD, which might be mediated by activation of Piezo1 induced inflammation and abnormal energy metabolism of IVD cells. Level of Evidence: N/A.

Tissue engineering is a promising approach for the treatment of chronic lower back pain (LBP) caused by intervertebral disc degeneration (IDD) resulting from degeneration and inflammation of annulus fibrosus (AF) tissue. However, scaffold with an anti-inflammatory effect on AF cells has not been reported. In this study, we fabricated a polylactide-glycolide (PLGA)/poly-ε-caprolactone (PCL)Zdextran (DEX) composite membrane loaded with plastrum testudinis extract (PTE), a Traditional Chinese Medicine herbal extract, via electrospinning. The membranes were characterized by mechanical measurements and scanning electron microscopy (SEM). Using anin vitroinflammation model induced by interleukin (IL)-1β, the cytocompatibility and anti-inflammatory effects of the composites were investigated by CCK-8 assay and flow cytometry. Potential regulatory mechanisms were examined by RT-qPCR and Western blotting. The results showed that the P10P8D2 (PLGA 10 g, PCL 8 g, DEX 2 g) composite nanofiber membrane exhibited the most uniform diameter distribution, best mechanical properties, a moderate degradation rate, and the best cytocompatibility characteristics. The optimal concentration of PTE was 120µg/mL. Importantly, P10P8D2 combined with PTE exhibited anti-inflammatory and cell proliferation promotion effects. Moreover, the NF-κBB/NLRP3/IL-βsignaling pathway was inactivated. Our findings suggested that the nanofiber membrane composed of P10P8D2 and PTE has anti-inflammatory and pro-proliferation effects on AF cells. It may provide an effective strategy for AF tissue regeneration.

Aim: To explore the potential functions and mechanism of N6.methyladenosine (m6A) abnormality of RNAs in nucleus pulposus from the intervertebral disc degeneration (IDD). Materials & methods: We performed rat model, m6A epitranscriptomic microarray, bioinformatics analysis and metabolomics. Results: In IDD, most of the differentially methylated RNAs showed a significant demethylation situation. The competing endogenous RNA network LOC102555094/miR-431/GSK-3β combining downstream Wnt pathway were identified in bioinformatics analysis. For metabolomics, activation of Wnt pathway led to reprogramming of glucose metabolism and enzyme activation of PKM2. Finally, quantitative real-time PCR and methylated RNA immunoprecipitation coupled with quantitative real-time PCR revealed the positive correlation between demethylation of LOC102555094 and expression of both FTO and ZFP217. Conclusion: LOC102555094 might be demethylated by ZFP217, activating FTO and LOC102555094/miR-431/GSK-3β/Wnt played a crucial role in IDD.

Background ： It has been acknowledged that the intervertebral disc degeneration(IDD) is associated with an aberrant cell-medicated response to structural failures, such as vertebral burst fracture, radial fissures, and endplate fracture. However, whether a momentary impact injury of the endplates without structural disruption, is sufficiently to initiate disc degeneration remains elusive. This study was to further evolve an in vitro momentary impact injury model of IDD and to investigate if a momentary impact load of the endplates without structural disruption could initiate IDD. Methods. Rat spinal segments (from L1/2 to L5/6, n=54) were harvested and randomly assigned into three groups: Control (n=18), Low Impact (12 J/cm 3 , n=18) and High Impact (25 J/cm 3 , n=18). Samples in both of the impact groups were subjected to axial momentary impact load using a custom-made apparatus, and cultured for 14 days. The degenerative process was investigated by using histomorphology and real-time PCR. Results: The discs in both of the impact groups showed significant degenerative changes at 14 days, both of which showed much higher histological scores and up-regulation of the catabolic (MMP-9, MMP-13) genes transcription than that of the control group ( P ＜0.05). The discs with endplate fracture compared to that with intact endplate also showed strongly up-regulated catabolic (MMP-9, MMP-13) genes transcription, and more significant degenerative changes based on the histological scoring ( P ＜0.05). Conclusion: This study demonstrated that a momentary impact load (12 J/cm 3 ) on the spinal segments of the rats could initiate IDD at 14 days after injury and not only endplate fracture but also a momentary impact injury without structural disruption could also promote IDD.

ObjectiveTo compare the effectiveness of platelet‐rich plasma (PRP), autologous blood (AB), and corticosteroid injections in patients with lateral epicondylitis.Type of StudyNetwork meta‐analysis.Literature SurveyRandomized controlled trials (RCTs) that compared any two forms of injections among PRP, AB, and corticosteroid for the treatment of lateral epicondylitis were searched from inception to 30 November 2018, on PubMed, Embase, and Cochrane library.MethodologyTwo researchers independently selected and assessed the quality of RCTs with the Cochrane Risk of Bias Tool. All relevant data from the included studies were extracted and heterogeneity was checked by Cochran's Q test and inconsistency statistic (I2). Publication bias was evaluated by constructing contour‐enhanced funnel plots. Stata 15 software was applied for pairwise meta‐analysis and network meta‐analysis. To explore the efficacy between different follow‐up periods, we considered the duration within 2 months to be short term, whereas 2 months or more was considered long term.SynthesisTwenty RCTs (n = 1271) were included in this network meta‐analysis. According to ranking probabilities, corticosteroid ranked first for visual analog score (VAS) (surface under the cumulative ranking [SUCRA] = 90.7), modified Nirschl score (82.9), maximum grip strength (69.5), modified Mayo score (MMS) (77.9), and Patient‐Related Tennis Elbow Evaluation (PRTEE) score (93.3) for the short‐term period. For the long‐term period, PRP ranked first for VAS (94.3), pressure pain threshold (99.8), Disabilities of Arm Shoulder and Hand (DASH) score (75.2), MMS (88.2), and the PRTEE score (81.8).ConclusionPRP was associated with more improvement in pain intensity and function in the long term than were the comparators. However, in the short term, corticosteroids were associated with the most improvement.