Circular RNA GRB10 as a competitive endogenous RNA regulating nucleus pulposus cells death in degenerative intervertebral disk

Zhang

Deng

et al. 2020

Preprint

BackgroundIntervertebral disc degeneration (IDD) can induce profound global socioeconomic burdens. Recent studies have suggested that circular RNAs might have crucial functions in the progression of IDD. The purpose of this study was to identify a specific circular RNA and to investigate its regulatory mechanism in IDD.MethodsCircGLCE was selected after microarray analyses and was further analysed by RT-qPCR and FISH. After silencing CircGLCE, its function was assessed with RT-qPCR, immunofluorescence analysis and flow cytometry. Based on Sanger sequencing, miR-587 was identified as a direct target of CircGLCE, and it was further examined with RNA pulldown assays, RT-qPCR, dual luciferase assays and FISH. After silencing CircGLCE or miR-587, western blotting, immunofluorescence analysis, and flow cytometry were conducted. STAP1 was assessed by RT-qPCR and luciferase assay, and experiments with silenced and overexpressed miR-587 were performed. A rescue experiment was also included. In an IDD rat model, the in vivo effects of overexpressing CircGLCE on IDD were analysed with imaging techniques, TUNEL staining, FISH, western blotting, H&E staining and immunohistochemistry.ResultsCircGLCE was found to stably exist in the cytoplasm of nucleus pulposus (NP) cells. It was downregulated in IDD. Knockdown of CircGLCE promoted apoptosis and induced the expression of matrix-degrading enzymes in NP cells. CircGLCE served as a miR-587 sponge in NP cells. Inhibiting miR-587 counteracted the IDD-enhancing effect caused by silencing CircGLCE. STAP1 served as the miRNA target that mediated the functions of miR-587. Overexpressing CircGLCE alleviated IDD in vivo.ConclusionsCircGLCE attenuates IDD by regulating the apoptosis of NP cells and by regulating ECM degradation through the targeting of miR-587/STAP1. CircGLCE may be a potential therapeutic target for IDD treatments.

Section: Discussionmentioning

confidence: 99%

CircGLCE alleviates intervertebral disc degeneration by regulating apoptosis and matrix degradation through the targeting of miR-587/STAP1

Zhang

Deng

et al. 2020

Preprint

“…Mechanistically, reduced circ‐GRB10 levels were found to increase the availability of miR‐328‐5p and thereby repressing ERBB2 (v‐erb‐b2 erythroblastic leukaemia viral oncogene homolog 2; a target of miR‐328‐5p). These data suggested that deregulation of circ‐GRB10/miR‐328‐5p/ERBB2 pathway was involved in IDD development . Restoring the expression of circ‐GRB10 or enhancing the ERBB2 signalling might thus serve a new therapeutic approach for the IDD.…”

Section: Functional Roles Of Specific Circrnas In Iddmentioning

confidence: 93%

“…and miR-328-5p were one of the most significantly negatively correlated circRNA-miRNA pairs 44. By RT-qPCR, the investigators verified the significant downregulation of circ-GRB10 and upregulation of miR-328-5p as well as their negative correlation in an independent set of IDD NP samples.…”

mentioning

confidence: 83%

“…Guo and colleagues reanalysed Liu et al's microarray datasets and showed that circ‐GRB10 (Growth Factor Receptor‐Bound Protein 10) and miR‐328‐5p were one of the most significantly negatively correlated circRNA‐miRNA pairs . By RT‐qPCR, the investigators verified the significant downregulation of circ‐GRB10 and upregulation of miR‐328‐5p as well as their negative correlation in an independent set of IDD NP samples.…”

Section: Functional Roles Of Specific Circrnas In Iddmentioning

confidence: 97%

“…Guo and colleagues reanalysed Liu et al's 30 pathway was involved in IDD development. 44 Restoring the expression of circ-GRB10 or enhancing the ERBB2 signalling might thus serve a new therapeutic approach for the IDD.…”

Section: Circ-grb10mentioning

confidence: 99%

See 2 more Smart Citations

Circular RNAs in nucleus pulposus cell function and intervertebral disc degeneration

et al. 2019

Cell Proliferation

Intervertebral disc degeneration (IDD) is a common cause of low back pain, which inflicts more global disability than any other condition. Although IDD was deemed to be a natural process that comes with ageing, a growing body of evidence suggested that both genetic and environmental factors could modify the development of IDD. In this connection, aberrant function of nucleus pulposus cells has been implicated in IDD pathogenesis. Circular RNAs are a novel class of endogenous non‐coding RNAs that play crucial regulatory roles in diverse cellular processes. Recently, deregulation of circRNAs in nucleus pulposus cells was found to functionally participate in IDD development. In this review, we summarize the current knowledge regarding the deregulation of circRNAs in IDD in relation to their actions on nucleus pulposus cell functions, including cell proliferation, apoptosis and extracellular matrix synthesis/degradation. The potential clinical utilities of circRNAs as therapeutic targets for the management of IDD are also discussed.

Inflammation‐dependent downregulation of miR‐194‐5p contributes to human intervertebral disc degeneration by targeting CUL4A and CUL4B

Journal Cellular Physiology

Han

Deng

et al. 2019

Inflammation is one of the major causes of intervertebral disc degeneration (IDD).Emerging evidence has revealed that increase in the levels of pro-inflammatory cytokines, such as interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-α), can activate a variety of signaling pathways, eventually resulting in IDD. Here, we show that the two cullin family genes, CUL4A and CUL4B, but not other cullins, are specifically overexpressed in IDD samples compared with healthy controls, and the CUL4A and CUL4B levels are positively correlated with the severity of IDD. In vitro analyses in human osteoblast cells (hFOB1.19), nucleus pulposus cells (hNPCs), and annulus fibrosus cells (hAFCs) indicated that treatment with IL-6 and TNF-α can increase CUL4A and CUL4B levels. By performing a microRNA-based microarray analysis, we found a set of microRNAs (miRNAs) that were differentially expressed in IDD samples compared with samples from healthy controls. Of these miRNAs, miR-194-5p, was significantly downregulated in IDD samples and could bind to the three prime untranslated regions (3′-UTRs) of both CUL4A and CUL4B, thereby downregulating their expression. The in vitro overexpression or downregulation of miR-194-5p, with a miR-194-5p-mimic or with anti-miR-194-5p, can cause the repression or induction of both CUL4A and CUL4B, respectively. Interestingly, treatment with IL-6 and TNF-α inhibitors in primary hNPCs and hAFCs that were isolated from patients with IDD led to the downregulation of CUL4A and CUL4B. Together, these findings provide insight into how the inflammation-dependent downregulation of miR-194-5p contributes to the pathogenesis of IDD, which may aid in the development of new therapeutic approaches for IDD by directly targeting miR-194-5p or CUL4A and CUL4B. K E Y W O R D S CUL4A, CUL4B, inflammation, intervertebral disc degeneration (IDD), microRNA (miRNA), miR-194-5p Additional supporting information may be found online in the Supporting Information section at the end of the article. How to cite this article: Chen Z, Han Y, Deng C, et al. Inflammation-dependent downregulation of miR-194-5p contributes to human intervertebral disc degeneration by targeting CUL4A and CUL4B.