Puerarin modulates proliferation, inflammation and ECM metabolism in human nucleus pulposus mesenchymal stem cells via the lncRNA LINC01535
Ontology highlight
ABSTRACT:
SUBMITTER: Cui P
PROVIDER: S-EPMC11253265 | biostudies-literature | 2024 Jun
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Puerarin (PUR), an 8-C-glucoside of daidzein extracted from Pueraria plants, is closely related to autophagy, reduced reactive oxygen species (ROS) production, and anti-inflammatory effects, but its effects on human nucleus pulposus mesenchymal stem cells (NPMSCs) have not yet been identified. In this study, NPMSCs were cultured in a compression apparatus to simulate the microenvironment of the intervertebral disc under controlled pressure (1.0 MPa), and we found that cell viability was decreased and apoptosis level was gradually increased as compression duration was prolonged. After PUR administration, apoptosis level evaluated by flow cytometry and caspase-3 activity was remitted, and protein levels of Bas as well as cleaved caspase-3 were decreased, while elevated Bcl-2 level was identified. Moreover, ATP production detection, ROS, and JC-1 fluorography as well as quantitative analysis suggested that PUR could attenuate intercellular ROS accumulation and mitochondrial dysfunction. Besides, the rat tail compression model was utilized, which indicated that PUR could restore impaired nucleus pulposus degeneration induced by compression. The PI3K/Akt pathway was identified to be deactivated after compression stimulation by western blot, and PUR could rescue the phosphorylation of Akt, thus reactivating the pathway. The effects of PUR, such as antiapoptosis, cell viability restoration, antioxidation, and mitochondrial maintenance, were all counteracted by application of the PI3K/Akt pathway inhibitor (LY294002). Summarily, PUR could alleviate compression-induced apoptosis and cell death of human NPMSCs in vitro as well as on the rat compression model and maintain intracellular homeostasis by stabilizing mitochondrial membrane potential and attenuating ROS accumulation through activating the PI3K/Akt pathway.
| S-EPMC7201526 | biostudies-literature
Project description:Intervertebral disc degeneration (IVDD) stands as a prevalent chronic orthopedic ailment, profoundly impacting patients' well-being due to incapacitating low back pain. Studies have highlighted a close correlation between IVDD and the programmed cell death of nucleus pulposus (NP) cells orchestrated by interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), and caspase-3 (CASP3). Puerarin, renowned for its anti-inflammatory attributes and its influence on IL-1β and TNF-α, emerges as a promising candidate for IVDD treatment. However, the precise mechanism by which it regulates apoptosis via these pathways remains ambiguous. This investigation utilizes bioinformatics to unveil the molecular intricacies of puerarin-mediated apoptosis regulation in IVDD, substantiated by preliminary in vitro experiments. Analysis exposes aberrant expression of pivotal apoptosis-associated proteins (IL-1β, TNF-α, CASP3, CASP8, and BCL2) in IVDD patients, with network pharmacology indicating puerarin's potential efficacy in IVDD treatment by modulating apoptosis and cellular senescence pathways. Further experiments elucidate puerarin's capacity to stimulate NP cell proliferation while inhibiting apoptosis, potentially contributing to IVDD mitigation. Western blot and PCR outcomes reveal escalated expression of apoptosis-related proteins (IL-1β, TNF-α, and CASP3) in lipopolysaccharide-treated NPCs, ameliorated by puerarin intervention. Molecular docking simulations demonstrate favorable binding properties of puerarin with apoptotic proteins, while flow cytometry analysis indicates its ability to diminish NPC apoptosis. These discoveries imply that puerarin might alleviate NPC apoptosis by modulating key targets, thereby potentially ameliorating IVDD. In summary, this study unveils the intrinsic mechanism of puerarin in regulating NPC apoptosis to alleviate IVDD, underscoring its therapeutic promise.
| S-EPMC11632247 | biostudies-literature
Project description:Cells take in, consume, and synthesize nutrients for numerous physiological functions. This includes not only energy production but also macromolecule biosynthesis, which will further influence cellular signaling, redox homeostasis, and cell fate commitment. Therefore, alteration in cellular nutrient metabolism is associated with pathological conditions. Intervertebral discs, particularly the nucleus pulposus (NP), are avascular and exhibit unique metabolic preferences. Clinical and preclinical studies have indicated a correlation between intervertebral degeneration (IDD) and systemic metabolic diseases such as diabetes, obesity, and dyslipidemia. However, a lack of understanding of the nutrient metabolism of NP cells is masking the underlying mechanism. Indeed, although previous studies indicated that glucose metabolism is essential for NP cells, the downstream metabolic pathways remain unknown, and the potential role of other nutrients, like amino acids and lipids, is understudied. In this literature review, we summarize the current understanding of nutrient metabolism in NP cells and discuss other potential metabolic pathways by referring to a human NP transcriptomic dataset deposited to the Gene Expression Omnibus, which can provide us hints for future studies of nutrient metabolism in NP cells and novel therapies for IDD.
| S-EPMC9801222 | biostudies-literature
Project description:Intervertebral disc degeneration is accompanied by elevated levels of inflammatory cytokines that have been implicated in disease etiology and matrix degradation. While the effects of inflammatory stimulation on disc cell metabolism have been well-studied, their effects on cell biophysical properties have not been investigated. The hypothesis of this study is that inflammatory stimulation alters the biomechanical properties of isolated disc cells and volume responses to step osmotic loading. Cells from the nucleus pulposus (NP) of bovine discs were isolated and treated with either lipopolysaccharide (LPS), an inflammatory ligand, or with the recombinant cytokine TNF-α for 24 hours. We measured cellular volume regulation responses to osmotic loading either immediately after stimulation or after a 1 week recovery period from the inflammatory stimuli. Cells from each group were tested under step osmotic loading and the transient volume-response was captured via time-lapse microscopy. Volume-responses were analyzed using mixture theory framework to investigate two biomechanical properties of the cell, the intracellular water content and the hydraulic permeability. Intracellular water content did not vary between treatment groups, but hydraulic permeability increased significantly with inflammatory treatment. In the 1 week recovery group, hydraulic permeability remained elevated relative to the untreated recovery control. Cell radius was also significantly increased both after 24 hours of treatment and after 1 week recovery. A significant linear correlation was observed between hydraulic permeability and cell radius in untreated cells at 24 hours and at 1-week recovery, though not in the inflammatory stimulated groups at either time point. This loss of correlation between cell size and hydraulic permeability suggests that regulation of volume change is disrupted irreversibly due to inflammatory stimulation. Inflammatory treated cells exhibited altered F-actin cytoskeleton expression relative to untreated cells. We also found a significant decrease in the expression of aquaporin-1, the predominant water channel in disc NP cells, with inflammatory stimulation. To our knowledge, this is the first study providing evidence that inflammatory stimulation directly alters the mechanobiology of NP cells. The cellular biophysical changes observed in this study are coincident with documented changes in the extracellular matrix induced by inflammation, and may be important in disease etiology.
| S-EPMC4061011 | biostudies-literature
Project description:BackgroundThe mechanisms underlying M2 macrophage polarization induced by nucleus pulposus (NP) cells are unclear. The effects that M2-polarized macrophages have on NP cells are also controversial.MethodsTranscriptome sequencing was performed to detect the gene change profiles between NP cells from ruptured intervertebral disc (IVD) and normal IVD. The main difference on biological activities between the two cell groups were analyzed by GO analysis and KEGG analysis. Virus transduction, flow cytometry, immunofluorescence, RT-PCR, western blot, CCK-8, TUNEL staining, and AO/EB staining were performed to explore the interactions between NP cells and RAW264.7 macrophages. Statistics were performed using SPSS26.Results801 upregulated and 276 downregulated genes were identified in NP cells from ruptured IVD in mouse models. According to GO and KEGG analysis, we found that the differentially expressed genes (DEG) were dominantly enriched in inflammatory response, extracellular matrix degradation, blood vessel morphogenesis, immune effector process, ossification, chemokine signaling pathway, macrophage activation, etc. CX3CL1 was one of the top 20% DEG, and we confirmed that both NP tissue and cells expressed remarkably higher level of CX3CL1 in mouse models (p < 0.001∗). Besides, we further revealed that both the recombinant CX3CL1 and NP cells remarkably induced M2 polarization of RAW264.7 (p < 0.001∗), respectively, while this effect was significantly reversed by si-CX3CL1 or JMS-17-2 (p < 0.001∗). Furthermore, we found that M2 macrophages significantly decreased the apoptosis rate (p < 0.001∗) and the catabolic gene levels (p < 0.001∗) of NP cells, while increased the viability, proliferation as well as the anabolic gene levels of NP cells (p < 0.01∗).ConclusionsVia regulating CX3CL1/CX3CR1 pathway, NP cells can induce the M2 macrophage polarization. M2 polarized macrophages can further promote NP cell viability, proliferation, and anabolism, while inhibit NP cell apoptosis and catabolism.
| S-EPMC10234370 | biostudies-literature
Project description:Gene therapy provides an ideal potential treatment for intervertebral disk degeneration by delivering synthetic microRNAs (miRNAs) to regulate the gene expression levels. However, it is very challenging to deliver miRNAs directly, which leads to inactivation, low transfection efficiency, and short half-life. Here, Agomir is loaded in hydrogel to construct a gene-hydrogel microenvironment for regulating the synthesis/catabolism balance of the tissue extracellular matrix (ECM) to treat degenerative diseases. Agomir is a cholesterol-, methylation-, and phosphorothioate-modified miRNA, which can mimic the function of miRNA to regulate the expression of the target gene. Agomir874 that mimics miRNA874 is synthesized to down regulate the expression of matrix metalloproteinases (MMPs) in nucleus pulposus (NP). At the same time, a polyethylene glycol (PEG) hydrogel is synthesized through Ag-S coordination of 4-arm PEG-SH and silver ion solution, which has injectable, self-healing, antimicrobial, degradable, and superabsorbent properties and matches perfectly with the mechanism of intervertebral disk. By delivering Agomir-loaded PEG-hydrogel to a degenerative intervertebral disk, a gene-hydrogel microenvironment is constructed in situ, which reduces the expression of MMPs, regulates the synthesis/catabolism balance of ECM in the NP of the intervertebral disk, and improves the tissue microenvironment regeneration.
| S-EPMC6947697 | biostudies-literature
Project description:Purpose:Mechanical loading plays a vital role in the progression of intervertebral disc (IVD) degeneration, but little is known about the effect of compression loading on human nucleus pulposus-derived mesenchymal stem cells (NP-MSCs). Thus, this study is aimed at investigating the effect of compression on the biological behavior of NP-MSCs in vitro. Methods:Human NP-MSCs were isolated from patients undergoing lumbar discectomy for IVD degeneration and were identified by immunophenotypes and multilineage differentiation. Then, cells were cultured in the compression apparatus at 1.0?MPa for different times (0?h, 24?h, 36?h, and 48?h). The viability-, differentiation-, and differentiation-related genes (Runx2, APP, and Col2) and colony formation-, migration-, and stem cell-related proteins (Sox2 and Oct4) were evaluated. Results:The results showed that the isolated cells fulfilled the criteria of MSC stated by the International Society for Cellular Therapy (ISCT). And our results also indicated that compression loading significantly inhibited cell viability, differentiation, colony formation, and migration. Furthermore, gene expression suggested that compression loading could downregulate the expression of stem cell-related proteins and lead to NP-MSC stemness losses. Conclusions:Our results suggested that the biological behavior of NP-MSCs could be inhibited by compression loading and therefore enhanced our understanding on the compression-induced endogenous repair failure of NP-MSCs during IVDD.
| S-EPMC6196892 | biostudies-other
Project description:The precise role of nucleus pulposus cell proliferation in the pathogenesis of intervertebral disc degeneration remains to be elucidated. Recent findings have revealed that microRNAs, a class of small noncoding RNAs, may regulate cell proliferation in many pathological conditions. Here, we showed that miR-21 was significantly upregulated in degenerative nucleus pulposus tissues when compared with nucleus pulposus tissues that were isolated from patients with idiopathic scoliosis and that miR-10b levels were associated with disc degeneration grade. Moreover, bioinformatics target prediction identified PTEN as a putative target of miR-21. miR-21 inhibited PTEN expression by directly targeting the 3'UTR, and this inhibition was abolished through miR-21 binding site mutations. miR-21 overexpression stimulated cell proliferation and AKT signaling pathway activation, which led to cyclin D1 translation. Additionally, the increase in proliferation and cyclin D1 expression induced by miR-21 overexpression was almost completely blocked by Ly294002, an AKT inhibitor. Taken together, aberrant miR-21 upregulation in intervertebral disc degeneration could target PTEN, which would contribute to abnormal nucleus pulposus cell proliferation through derepressing the Akt pathway. Our study also underscores the potential of miR-21 and the PTEN/Akt pathway as novel therapeutic targets in intervertebral disc degeneration.
| S-EPMC3975380 | biostudies-literature
Project description:The inflammatory-associated factors interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) are widely reported to be associated with intervertebral disc (IVD) degeneration (IVDD). N-acetyl-5-methoxytryptamine (melatonin) is a natural hormone secreted by the pineal gland which has been shown to participate in several physiological and pathological progresses, such as aging, anti-inflammation, anti-apoptosis and autophagy regulation. However, the effects of melatonin on IVD remain unclear. In the present study, we treated human nucleus pulposus cells (NPCs) with melatonin and discovered that melatonin could modulate extracellular matrix (ECM) remodeling induced by IL-1β by enhancing collagen II and aggrecan expression levels and by downregulating matrix metalloproteinase-3 (MMP-3) levels. These findings were verified by western blot and immunofluorescence assays. Intraperitoneal injection of melatonin mitigated IVDD in the rat tail puncture model. X-ray and magnetic resonance imaging (MRI), as well as hematoxylin-eosin (H&E), Safranine O-Green, Alcian blue and Celium red staining methods were adopted to evaluate IVDD grades, the structural integrity of nucleus pulposus (NP) and annulus fibrosus (AF) and the damage and calcification of the cartilage endplate. Melatonin reduced inflammatory cell aggregation and the release of the inflammatory factors IL-1β, IL-6, TNF-α as determined by immunohistochemistry. In conclusion, the present study demonstrated that melatonin could modulate ECM remodeling by IL-1β in vitro and attenuate the IVDD and induction of inflammation in a rat tail puncture model in vivo. The data demonstrated that melatonin may contribute to the restoration processs of IVD following damage and may be used as a potential novel therapy for IVDD.
| S-EPMC6914432 | biostudies-literature
Project description:BackgroundIntervertebral disc degeneration (IVDD) is regarded as the leading cause of low back pain, resulting in disability and a heavy burden on public health. Several studies have unveiled that long noncoding RNAs (lncRNAs) play a key role in the pathogenesis and progression of IVDD. In this study, we aimed to investigate the biological function and latent molecular mechanism of the lncRNA FAM83H antisense RNA 1 (FAM83H-AS1) in IVDD development.MethodsFirstly, we established an IVDD model in rats using advanced glycation end products (AGEs) intradiscal injection. Subsequently, gain-of-function assays were conducted to investigate the role of FAM83H-AS1 in the progression of IVDD. Bioinformatics analysis, RNA pull down assay and rescue experiments were employed to shed light on the molecular mechanism underlying FAM83H-AS1 involving in IVDD.ResultsOur findings verified that AGEs treatment aggravated IVDD damage, and FAM83H-AS1 was downregulated in the IVDD group. Additionally, overexpression of FAM83H-AS1 contributed to the growth of nucleus pulposus (NP) cells and ameliorated IVDD injury. It was revealed that FAM83H-AS1 possessed the speculated binding sites of miR-22-3p. More importantly, we confirmed that FAM83H-AS1 functioned as a sponge of miR-22-3p in IVDD. Lastly, we demonstrated that miR-22-3p mediated the impact of FAM83H-AS1 on cell proliferation, ECM degradation, and inflammation.ConclusionsOur study indicated that FAM83H-AS1 relieved IVDD deterioration through sponging miR-22-3p, and provides novel insights into the mechanisms underlying FAM83H-AS1 in IVDD progression.
| S-EPMC7729348 | biostudies-literature