Dataset Information

The c-Jun signaling pathway has a protective effect on nucleus pulposus cells in patients with intervertebral disc degeneration

ABSTRACT: Among a range of diverse clinical symptoms, intervertebral disc degeneration (IDD) contributes mostly to the onset of lower back pain. The present study aimed to investigate the effects of c-Jun on nucleus pulposus (NP) cells of IDD and its regulation on molecular mechanisms. Intervertebral disc (IVD) tissues were collected from patients suffering from IDD disease, and NP cells were subsequently isolated and cultured. By overexpressing c-Jun in NP cells, expression levels of mRNAs and proteins of IDD-related genes and inflammatory cytokines were subjected to reverse transcription-quantitative PCR, western blot and ELISA assays. Additional transforming growth factor-? (TGF-?) antibodies were administrated to suppress the function of TGF-?. Cell proliferation and apoptosis were determined via Cell Counting Kit-8 and TUNEL assays, respectively. The results demonstrated that the overexpression of c-Jun robustly upregulated both mRNA and protein expression of TGF-?, TIMP metallopeptidase inhibitor 3, aggrecan and collagen type II alpha 1 chain and simultaneously downregulated the expression of the inflammatory cytokines TNF-?, interleukin (IL)-1?, IL-6 and IL-17. Furthermore, following c-Jun overexpression, survival rates of NP cells were increased while apoptosis rates were decreased. However, the addition of a TGF-? antibody significantly promoted apoptosis and restricted cell survival, which differed from the results of the c-Jun overexpression group. The present study hypothesized therefore that c-Jun may positively regulate TGF-? expression within NP cells of IDD, which could promote the proliferation of IDD-NP cells and accelerate cell viability via reducing apoptosis and the inflammatory response.

SUBMITTER: Lei M

PROVIDER: S-EPMC7523272 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Background contextLow back pain, affecting nearly 40% of adults, mainly results from intervertebral disc degeneration (IVDD), while the pathogenesis of IVDD is still not fully elucidated. Recently, some researches have revealed that necroptosis, a programmed necrosis, participated in the progression of IVDD, nevertheless, the underlying mechanism remains unclear.PurposeTo study the mechanism of necroptosis of Nucleus Pulposus (NP) cells in IVDD, focusing on the role of MyD88 signaling.Study designThe expression and co-localization of necroptotic indicators and MyD88 were examined in vivo, and MyD88 inhibitor was applied to determine the role of MyD88 signaling in necroptosis of NP cells in vitro.MethodsHuman disc specimens were collected from patients receiving diskectomy for lumbar disc herniation (LDH) or traumatic lumbar fractures after MRI scanning. According to the Pfirrmann grades, they were divided into normal (Grades 1, 2) and degenerated groups (4, 5). Tissue slides were prepared for immunofluorescence to assess the co-localization of necroptotic indicators (RIP3, MLKL, p-MLKL) and MyD88 histologically. The combination of TNFα, LPS and Z-VAD-FMK was applied to induce necroptosis of NP cells. Level of ATP, reactive oxygen species (ROS), live-cell staining and electron microscope study were employed to study the role of MyD88 signaling in necroptosis of NP cells.ResultsIn vivo, the increased expression and co-localization of necroptotic indicators (RIP3, MLKL, p-MLKL) and MyD88 were found in NP cells of degenerated disc, while very l low fluorescence intensity in tissue of traumatic lumbar fractures. In vitro, the MyD88 inhibitor effectively rescued the necroptosis of NP cells, accompanied by increased viability, ATP level, and decreased ROS level. The effect of MyD88 inhibition on necroptosis of NP cells was further confirmed by ultrastructure of mitochondria shown by Transmission Electron Microscope (TEM).ConclusionOur results indicated that the involvement of MyD88 signaling in the necroptosis of NP cells in IVDD, which will replenish the pathogenesis of IVDD and provide a novel potential therapeutic target for IVDD.

Project description:Intervertebral disc (IVD) degeneration is a complicated physiological change involving cellular senescence, inflammation and the degradation of the extracellular matrix. Long non?coding RNAs (lncRNAs) have been identified as new players in IVD degeneration. The present study aimed to identify lncRNAs implicated in IVD degeneration via the regulation of cellular senescence. In the present study, nucleus pulposus (NP) cells isolated from moderately degenerated IVD tissues exhibited a senescent phenotype with increased senescence rates, detected by senescence?associated ??galactosidase (SA???gal) staining, and reduced growth and migratory abilities. Microarray and target prediction analyses identified 353 differentially expressed lncRNAs, and 251 cis? and 2,170 trans?acting targets in degenerated NP cells. Bioinformatic analyses revealed that these predicted targets were enriched in the regulation of response to DNA damage stimulus, positive regulation of cell cycle processes and interferon?? production. In addition, a network of the top 10 upregulated and top 10 downregulated lncRNA targets was constructed, and two trans?acting targets, C?C motif chemokine ligand 5 (CCL5) and polyribonucleotide nucleotidyltransferase 1 (PNPT1) involved in aging or senescence, and their corresponding lncRNAs, lnc?ST8SIA5?1:2 and lnc?HRK?2:1, were identified. Reverse transcription?quantitative PCR validation demonstrated that the two targets and two candidate lncRNAs were significantly upregulated in degenerated NP cells. Overexpression of lnc?HRK?2:1, with validated higher expression levels, in normal NP cells induced a senescent phenotype, with enhanced rates of senescence detected by SA???gal staining in cells, decreased growth and migratory abilities and improved expression levels of CCL5 and PNPT1. Collectively, these results suggested that upregulation of lnc?HRK?2:1 prompted NP cell senescence in IVD degeneration, which may be associated with increased expression levels of CCL5 and PNPT1.

Dataset Information

The c-Jun signaling pathway has a protective effect on nucleus pulposus cells in patients with intervertebral disc degeneration

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets