Emerging role and therapeutic implication of mTOR signalling in intervertebral disc degeneration

Intervertebral disc degeneration (IDD), an important cause of chronic low back pain (LBP), is considered the pathological basis for various spinal degenerative diseases. A series of factors, including inflammatory response, oxidative stress, autophagy, abnormal mechanical stress, nutritional deficiency, and genetics, lead to reduced extracellular matrix (ECM) synthesis by intervertebral disc (IVD) cells and accelerate IDD progression. Mammalian target of rapamycin (mTOR) is an evolutionarily conserved serine/threonine kinase that plays a vital role in diverse degenerative diseases. Recent studies have shown that mTOR signalling is involved in the regulation of autophagy, oxidative stress, inflammatory responses, ECM homeostasis, cellular senescence, and apoptosis in IVD cells. Accordingly, we reviewed the mechanism of mTOR signalling in the pathogenesis of IDD to provide innovative ideas for future research and IDD treatment.     

Effects of pulsed electromagnetic field on intervertebral disc cell apoptosis in rats

Despite numerous studies on pulsed electromagnetic field (PEMF) application, its effects of PEMF on intervertebral disc (IVD) have not yet been investigated in vivo. Accordingly, the effects of PEMF upon IVD in rats were evaluated through molecular surveys. Rats were divided into six groups: Group I and II were exposed to low and high frequency of PEMF (LF and HF, respectively). Group III and IV underwent induced disc degeneration and were exposed to low and high frequency of PEMF (LF/IDD and HF/IDD, respectively). Group V underwent induced disc degeneration (IDD), and group VI was control. The values of caspase 3, Bax, Bcl-2 and β-actin band density, as cell apoptotic markers, were obtained from band densitometry. Our results showed that the value of cleaved caspase-3 of cells and Bax/Bcl-2 ratio in IDD group increased significantly compared to the control group (p?p?相似文献   

lncRNA HOTAIR upregulates autophagy to promote apoptosis and senescence of nucleus pulposus cells

Intervertebral disc degeneration (IDD) is a complex and chronic disease that involves disc cell senescence, death, and extracellular matrix (ECM) degradation. HOTAIR, a long non-coding RNA (lncRNA) is reportedly associated with autophagy, whereas autophagy is shown to promote IDD. However, how it affects nucleus pulposus (NP) cells, the primary component of intervertebral discs is still unclear. We hypothesized that HOTAIR promotes NP cell apoptosis and senescence through upregulating autophagy. Thus, silencing HOTAIR should inhibit autophagy and exert a therapeutic effect on IDD. Our in vitro experiments in human NP cells revealed that HOTAIR expression positively correlated with IDD grade, and overexpression enhanced autophagy. Autophagy inhibition via 3-methyladenine reversed HOTAIR stimulatory effects on apoptosis, senescence, and ECM catabolism, while the AMP-activated protein kinase (AMPK) inhibitor Compound C suppressed HOTAIR-induced autophagy through regulating AMPK/mTOR/ULK1 pathways. Our in vivo experiment then illustrated that silencing HOTAIR ameliorates IDD in rats. Collectively, we demonstrated that HOTAIR stimulates autophagy to promote NP cell apoptosis, senescence, and ECM catabolism. Therefore, silencing HOTAIR has the potential to become a treatment option for IDD.     

The influence of serum,glucose and oxygen on intervertebral disc cell growth <Emphasis Type="Italic">in vitro</Emphasis>: implications for degenerative disc disease

Introduction  

The avascular nature of the human intervertebral disc (IVD) is thought to play a major role in disc pathophysiology by limiting nutrient supply to resident IVD cells. In the human IVD, the central IVD cells at maturity are normally chondrocytic in phenotype. However, abnormal cell phenotypes have been associated with degenerative disc diseases, including cell proliferation and cluster formation, cell death, stellate morphologies, and cell senescence. Therefore, we have examined the relative influence of possible blood-borne factors on the growth characteristics of IVD cells in vitro.     

Retracted: HO-1 overexpression alleviates senescence by inducing autophagy via the mitochondrial route in human nucleus pulposus cells

Intervertebral disc degeneration (IDD) is closely associated with aging. Our previous studies have confirmed that heme oxygenase-1 (HO-1) can inhibit nucleus pulposus (NP) cell apoptosis. However, whether or not HO-1 is involved in NP cell senescence and autophagy is unclear. Our results indicated that HO-1 expression was reduced in IDD tissues and replicative senescent NP cells. HO-1 overexpression using a lentiviral vector reduced the NP cell senescence level, protected mitochondrial function, and promoted NP cell autophagy through the mitochondrial pathway. Autophagy inhibitor 3-MA pretreatment reversed the anti-senescent and protective effects on the mitochondrial function of HO-1, which promoted the degradation of the extracellular matrix (ECM) in the intervertebral disc. In vivo, HO-1 overexpression inhibited IDD and enhanced autophagy. In summary, these results suggested that HO-1 overexpression alleviates NP cell senescence by inducing autophagy via the mitochondrial route.     

Cell death in intervertebral disc degeneration

Degeneration of intervertebral disc (IVD) is mainly a chronic process of excessive destruction of the extracellular matrix (ECM), and also is thought to be the primary cause of low back pain. Presently, however, the underlying mechanism of IVD degeneration is still not elucidated. Cellular loss from cell death has been believed to contribute to the degradation of ECM and plays an important role in the process of IVD degeneration, but the mechanisms of cell death in degenerated IVD remain unclear. Apoptosis, a very important type of IVD cell death, has been considered to play a crucial role in the process of degeneration. Autophagy, a non-apoptosis death type of programmed cell death, has been considered extensively involved in many pathological and physiological processes, including the degenerative diseases. Thus, the research on cell death in IVD degeneration has become a new focus recently. In this review, by analyzing the available literature pertaining to cell death in IVD and discussing the inducing factors of IVD degeneration, NP cells and ECM in IVD degeneration, apoptotic signal transduction pathways involved in IVD cell death, the relationship of cell death with IVD degeneration and potential therapeutic strategy for IVD degeneration by regulating cell death, we conclude that different stimuli induce cell death in IVD via various signal transduction pathways, and that cell death may play a key role in the degenerative process of IVD. Regulation of cell death could be a potential and attractive therapeutic strategy for IVD degeneration.     

Programmed cell death in intervertebral disc degeneration

Intervertebral disc (IVD) degeneration is largely a process of destruction and failure of the extracellular matrix (ECM), and symptomatic IVD degeneration is thought to be one of the leading causes of morbidity or life quality deterioration in the elderly. To date, however, the mechanism of IVD degeneration is still not fully understood. Cellular loss from cell death in the process of IVD degeneration has long been confirmed and considered to contribute to ECM degradation, but the causes and the manners of IVD cell death remain unclear. Programmed cell death (PCD) is executed by an active cellular process and is extensively involved in many physiological and pathological processes, including embryonic development and human degenerative diseases. Thus, the relationship between PCD and IVD degeneration has become a new research focus of interest in recent years. By reviewing the available literature concentrated on PCD in IVD and discussing the methodology of detecting PCD in IVD cells, its inducing factors, the relationship of cell death to ECM degradation, and the potential therapy for IVD degeneration by modulation of PCD, we conclude that IVD cells undergo PCD via different signal transduction pathways in response to different stimuli, that PCD may play a role in the process of IVD degeneration, and that modulation of PCD might be a potential therapeutic strategy for IVD degeneration.     

Compression-induced senescence of nucleus pulposus cells by promoting mitophagy activation via the PINK1/PARKIN pathway

The current research aimed to explore the possible relationship between PINK1/PARKIN-mediated mitophagy and the compression-induced senescence of nucleus pulposus cells (NPCs). Therefore, the stages of senescence in NPCs were measured under compression lasting 0, 24 and 48 hours. The mitophagy-related markers, autophagosomes and mitochondrial membrane potential were tested to determine the levels of PINK1/PARKIN-mediated mitophagy under compression. The PINK1 and PARKIN levels were also measured by immunohistochemistry of human and rat intervertebral disc (IVD) tissues taken at different degenerative stages. A specific mitophagy inhibitor, cyclosporine A (CSA) and a constructed PINK1-shRNA were used to explore the relationship between mitophagy and senescence by down-regulating the PINK1/PARKIN-mediated mitophagy levels. Our results indicated that compression significantly enhanced the senescence of NPCs in a time-dependent manner. Also, PINK1/PARKIN-mediated mitophagy was found to be activated by the extended duration of compression on NPCs as well as the increased degenerative stages of IVD tissues. After inhibition of PINK1/PARKIN-mediated mitophagy by CSA and PINK1-shRNA, the senescence of NPCs induced by compression was strongly rescued. Hence, the excessive degradation of mitochondria in NPCs by mitophagy under continuous compression may accelerate the senescence of NPCs. Regulating PINK1/PARKIN-mediated mitophagy might be a potential therapeutic treatment for IVD degeneration.     

Spermidine promotes nucleus pulposus autophagy as a protective mechanism against apoptosis and ameliorates disc degeneration

Spermidine has therapeutic effects in many diseases including as heart diastolic function, myopathic defects and neurodegenerative disorders via autophagy activation. Autophagy has been found to mitigate cell apoptosis in intervertebral disc degeneration (IDD). Accordingly, we theorize that spermidine may have beneficial effects on IDD via autophagy stimulation. In this study, spermidine's effect on IDD was evaluated in tert‐butyl hydroperoxide (TBHP)‐treated nucleus pulposus cells of SD rats in vitro as well as in a puncture‐induced rat IDD model. We found that autophagy was actuated by spermidine in nucleus pulposus cells. In addition, spermidine treatment weakened the apoptotic effects of TBHP in nucleus pulposus cells. Spermidine increased the expression of anabolic proteins including Collagen‐II and aggrecan and decreased the expression of catabolic proteins including MMP13 and Adamts‐5. Additionally, autophagy blockade using 3‐MA reversed the beneficial impact of spermidine against nucleus pulposus cell apoptosis. Autophagy was thus important for spermidine's therapeutic effect on IDD. Spermidine‐treated rats had an accentuated T2‐weighted signal and a diminished histological degenerative grade than vehicle‐treated rats, showing that spermidine inhibited intervertebral disc degeneration in vivo. Thus, spermidine protects nucleus pulposus cells against apoptosis through autophagy activation and improves disc, which may be beneficial for the treatment of IDD.     

Partial reprogramming strategy for intervertebral disc rejuvenation by activating energy switch

Rejuvenation of nucleus pulposus cells (NPCs) in degenerative discs can reverse intervertebral disc degeneration (IDD). Partial reprogramming is used to rejuvenate aging cells and ameliorate progression of aging tissue to avoiding formation of tumors by classical reprogramming. Understanding the effects and potential mechanisms of partial reprogramming in degenerative discs provides insights for development of new therapies for IDD treatment. The findings of the present study show that partial reprogramming through short‐term cyclic expression of Oct‐3/4, Sox2, Klf4, and c‐Myc (OSKM) inhibits progression of IDD, and significantly reduces senescence related phenotypes in aging NPCs. Mechanistically, short‐term induction of OSKM in aging NPCs activates energy metabolism as a “energy switch” by upregulating expression of Hexokinase 2 (HK2) ultimately promoting redistribution of cytoskeleton and restoring the aging state in aging NPCs. These findings indicate that partial reprogramming through short‐term induction of OSKM has high therapeutic potential in the treatment of IDD.     

CircSEMA4B targets miR-431 modulating IL-1β-induced degradative changes in nucleus pulposus cells in intervertebral disc degeneration via Wnt pathway

Intervertebral disc (IVD) degeneration (IDD), characterized by elevated levels of proinflammatory mediators, increased Aggrecan and collagen degradation, and increased degradation of extracellular matrix (ECM), has been widely regarded as a significant contributor to low back pain. Genetics are significant factors contribute to IDD. Based on previous data, circular RNA SEMA4B (circSEMA4B) is down-regulated in IDD specimens; herein, we demonstrated circSEMA4B overexpression could attenuate the effect of IL-1β on nucleus pulposus cell (NPC) proliferation, senescence, and ECM and Aggrecan degradation in IDD via Wnt signaling. Moreover, miR-431, a direct target of circSEMA4B, could bind to the 3′UTR of SFRP1 or GSK-3β, two inhibitory regulators of Wnt signaling, to inhibit their expression thus playing a role similar to the activator of Wnt signaling in NPCs. The effect of circSEMA4B knockdown on NPCs was partially reversed by miR-431 inhibition; circSEMA4B serves as a miR-431 sponge to compete with SFRP1 or GSK-3β for miR-431 binding, thus inhibiting IL-1β-induced degenerative process in NPCs through Wnt signaling. Rescuing circSEMA4B expression in NPCs in IDD might present a potential strategy for IDD improvement.     

miR-424-5p regulates apoptosis and cell proliferation via targeting Bcl2 in nucleus pulposus cells

ABSTRACT

miRNAs play an important role in the pathogenesis of intervertebral disc degeneration (IDD). The role and the underlying mechanism of miR-424-5p in human nucleus pulposus (NP) are still unknown. We aimed to explore the role of miR-424-5p in IDD.

Real-time PCR was used to detect the expression of miR-424-5p and Bcl2 in IDD tissues and idiopathic scoliosis tissues. Human NP cells were used in our study. MTT and Hoechst apoptosis assays were used to detect the proliferation and apoptosis of NP cells, respectively. Western blotting assays were used to detect the expression levels of Bcl-2, cleaved caspase-3, cleaved caspase-9, caspase-3 and caspase-9 in degenerative NP cells. A luciferase reporter assay was applied to confirm the relationship between miR-424-5p and Bcl2.

Our results showed that the expression of miR-424-5p was increased and Bcl2 was decreased in degenerative NP cells. miR-425-5p expression was negatively correlated with Bcl2 expression in IDD tissues. Suppression of miR-424-5p using an inhibitor increased Bcl2 expression at both the mRNA and protein levels, and it promoted cell viability and inhibited apoptosis. Furthermore, the levels of cleaved caspase-3 and cleaved caspase-9 were downregulated in miR-424-5p-silenced NP cells. Interestingly, we found that silencing miR-424-5p increased p62 expression at both the mRNA and protein levels. Finally, a luciferase reporter assay verified the binding of the miR-424-5p and the 3’UTR of Bcl2.

These results suggested that silencing miR-424-5p suppressed NP cell apoptosis by upregulating Bcl2. Therefore, miR-424-5p might be a novel target for IDD therapies.     

Down‐regulation of insulin‐like growth factor binding protein 5 is involved in intervertebral disc degeneration via the ERK signalling pathway

It is obvious that epigenetic processes influence the evolution of intervertebral disc degeneration (IDD). However, its molecular mechanisms are poorly understood. Therefore, we tested the hypothesis that IGFBP5, a potential regulator of IDD, modulates IDD via the ERK signalling pathway. We showed that IGFBP5 mRNA was significantly down‐regulated in degenerative nucleus pulposus (NP) tissues. IGFBP5 was shown to significantly promote NP cell proliferation and inhibit apoptosis in vitro, which was confirmed by MTT, flow cytometry and colony formation assays. Furthermore, IGFBP5 was shown to exert its effects by inhibiting the ERK signalling pathway. The effects induced by IGFBP5 overexpression on NP cells were similar to those induced by treatment with an ERK pathway inhibitor (PD98059). Moreover, qRT‐PCR and Western blot analyses were performed to examine the levels of apoptosis‐related factors, including Bax, caspase‐3 and Bcl2. The silencing of IGFBP5 up‐regulated the levels of Bax and caspase‐3 and down‐regulated the level of Bcl2, thereby contributing to the development of human IDD. Furthermore, these results were confirmed in vivo using an IDD rat model, which showed that the induction of Igfbp5 mRNA expression abrogated the effects of IGFBP5 silencing on intervertebral discs. Overall, our findings elucidate the role of IGFBP5 in the pathogenesis of IDD and provide a potential novel therapeutic target for IDD.     

The involvement of interleukin-1 and interleukin-4 in the response of human annulus fibrosus cells to cyclic tensile strain: an altered mechanotransduction pathway with degeneration

Introduction  

Recent evidence suggests that intervertebral disc (IVD) cells derived from degenerative tissue are unable to respond to physiologically relevant mechanical stimuli in the 'normal' anabolic manner, but instead respond by increasing matrix catabolism. Understanding the nature of the biological processes which allow disc cells to sense and respond to mechanical stimuli (a process termed 'mechanotransduction') is important to ascertain whether these signalling pathways differ with disease. The aim here was to investigate the involvement of interleukin (IL)-1 and IL-4 in the response of annulus fibrosus (AF) cells derived from nondegenerative and degenerative tissue to cyclic tensile strain to determine whether cytokine involvement differed with IVD degeneration.     

几种椎间盘退变动物模型的建立和应用

椎间盘退变是一种年龄相关的退行性疾病,是引起下腰痛的主要因素,严重影响病人的生活质量,并显著增加家庭的经济负担。目前,缺少椎间盘退变的有效干预和治疗手段,部分原因是其发病机制尚未阐明。椎间盘退变动物模型的构建对于阐明该疾病的病理机制至关重要。椎间盘退变是一个复杂的过程,受机械应力、结构损伤、生物化学与基因表达等多种因素的影响。本文总结了应用异常机械应力、结构损伤、生物化学或化学诱导和基因敲除等方式构建的椎间盘退变动物模型。生物力学是维持椎间盘稳态的重要因素,异常的机械应力会导致椎间盘退变。同时,椎间盘退变常伴随结构性损伤,椎间盘结构破坏也会导致椎间盘发生退变。此外,生物化学或化学诱导和关键基因敲除也会导致椎间盘退变。本文按照造成异常机械应力的因素将机械应力模型分为加压模型和失稳模型;按照椎间盘结构将结构损伤模型分为髓核与纤维环损伤模型和软骨终板损伤模型。总结了生物化学或化学诱导模型以及新型的基因敲除模型。讨论了不同类型椎间盘退变动物模型的可能应用和局限性。     

Computation models simulating notochordal cell extinction during early ageing of an intervertebral disc

Lower back pain due to intervertebral disc (IVD) degeneration is a prevalent problem which drastically affects the quality of life of millions of sufferers. Healthy IVDs begin with high populations of notochordal cells in the nucleus pulposus, while by the second stage of degeneration, these cells will be replaced by chondrocyte-like cells. Because the IVD is avascular, these cells rely on passive diffusion of nutrients to survive. It is thought that this transition in cell phenotype causes the shift of the IVD's physical properties, which impede the flow of nutrients. Our computational model of the IVD illustrates its ability to simulate the evolving chemical and mechanical environments occurring during the early ageing process. We demonstrate that, due to the insufficient nutrient supply and accompanying changes in physical properties of the IVD, there was a resultant exponential decay in the number of notochordal cells over time.     

Genistein protects intervertebral discs from degeneration via Nrf2-mediated antioxidant defense system: An in vitro and in vivo study

Oxidative stress has been reported to be closely associated with the development of intervertebral disc degeneration (IDD). IDD is one of the major causes of low back pain. Genistein (GES), one of the main isoflavones of soybean, has been shown to exert multiple biological functions on different diseases. Here, we tested the therapeutic potential of GES for IDD. In vitro experiments, we confirmed GES was nontoxic to rat nucleus pulposus cells (NPCs) within the concentration of 100 μM. Furthermore, GES was able to suppress apoptosis in tert-butyl hydroperoxide (TBHP)-treated NPCs. In the aspect of extracellular matrix (ECM), GES not only reduced metalloproteinase-13 (MMP-13) and a disintegrin-like and MMP thrombospondin type 1 motif 5 expression, but also increased aggrecan and type II collagen levels. Also, we found GES might rescue TBHP-induced NPCs degeneration by enhancing Nrf2-mediated antioxidant defense system. Silencing Nrf2 partly abolished the protective effects of GES on apoptosis and ECM disruption in TBHP-treated NPCs. Correspondingly, GES ameliorated IDD in a rat model by preserving morphology of degenerative intervertebral discs and promoting Nrf2 expression. To sum up, our study suggests that GES exerts protective effects in NPCs against degeneration and reveals the underlying mechanism of GES on Nrf2 activation in NPCs.     

Regulating the fate of stem cells for regenerating the intervertebral disc degeneration

Lower back pain is a leading cause of disability and is one of the reasons for the substantial socioeconomic burden. The etiology of intervertebral disc (IVD) degeneration is complicated, and its mechanism is still not completely understood. Factors such as aging, systemic inflammation, biochemical mediators, toxic environmental factors, physical injuries, and genetic factors are involved in the progression of its pathophysiology. Currently, no therapy for restoring degenerated IVD is available except pain management, reduced physical activities, and surgical intervention. Therefore, it is imperative to establish regenerative medicine-based approaches to heal and repair the injured disc, repopulate the cell types to retain water content, synthesize extracellular matrix, and strengthen the disc to restore normal spine flexion. Cellular therapy has gained attention for IVD management as an alternative therapeutic option. In this review, we present an overview of the anatomical and molecular structure and the surrounding pathophysiology of the IVD. Modern therapeutic approaches, including proteins and growth factors, cellular and gene therapy, and cell fate regulators are reviewed. Similarly, small molecules that modulate the fate of stem cells for their differentiation into chondrocytes and notochordal cell types are highlighted.