Immunohistochemistry of chondromodulin-I in the human intervertebral discs with special reference to the degenerative changes

The expression of the matrix protein chondromodulin-I has been studied in human intervertebral discs of 101 people using immunohistochemical analyses. The purpose of this report is to present data on the metabolic changes that were found to occur in the chondrocytes of intervertebral discs during development and aging. Chondromodulin-I was highly expressed during the gestational period and gradually decreased after maturation. It was detected in both the extracellular matrix and chondrocytes in the zone of hypertrophic cartilage, the zone of proliferative cartilage and the zone of resting cartilage in fetal discs. It was also present in the annulus fibrosus, nucleus pulposus and end-plate cartilage in mature discs. In degenerative discs, chondromodulin-I immunoreactivity tended to be elevated in the remaining chondrocytes. Our findings suggest that the expression of the protein is developmentally regulated and upregulated through a defense mechanism against the degenerative processes of the aged intervertebral disc.     

Collagen stability and cross-linking in normal and kyphoscoliotic mouse intervertebral discs

Intervertebral discs of the cervical-thoracic region of the spine of BDL mice which are homozygous for the ky gene mutation undergo degeneration. Discs from these mice have a normal collagen content and undergo normal collagen cross linking prior to the appearance of degenerative changes. The major reducible collagen cross-link formed in discs of these mice and in normal CBA strain mice is hydroxylysino-5-ketonorleucine. These results and other previous results indicate that the discs in the ky mouse develop degenerative disease due to an extrinsic factor rather than to an intrinsic abnormality of their extracellular matrix. The extrinsic factor has been identified as spinal muscle atrophy.     

Nucleotomy reduces the effects of cyclic compressive loading with unloaded recovery on human intervertebral discs

The first objective of this study was to determine the effects of physiological cyclic loading followed by unloaded recovery on the mechanical response of human intervertebral discs. The second objective was to examine how nucleotomy alters the disc?s mechanical response to cyclic loading. To complete these objectives, 15 human L5-S1 discs were tested while intact and subsequent to nucleotomy. The testing consisted of 10,000 cycles of physiological compressive loads followed by unloaded hydrated recovery. Cyclic loading increased compression modulus (3%) and strain (33%), decreased neutral zone modulus (52%), and increased neutral zone strain (31%). Degeneration was not correlated with the effect of cyclic loading in intact discs, but was correlated with cyclic loading effects after nucleotomy, with more degenerate samples experiencing greater increases in both compressive and neutral zone strain following cyclic loading. Partial removal of the nucleus pulposus decreased the compression and neutral zone modulus while increasing strain. These changes correspond to hypermobility, which will alter overall spinal mechanics and may impact low back pain via altered motion throughout the spinal column. Nucleotomy also reduced the effects of cyclic loading on mechanical properties, likely due to altered fluid flow, which may impact cellular mechanotransduction and transport of disc nutrients and waste. Degeneration was not correlated with the acute changes of nucleotomy. Results of this study provide an ideal protocol and control data for evaluating the effectiveness of a mechanically-based disc degeneration treatment, such as a nucleus replacement.     

Increase of nerve growth factor levels in the human herniated intervertebral disc: can annular rupture trigger discogenic back pain?

Introduction

Nerve growth factor (NGF) has an important role in the generation of discogenic pain. We hypothesized that annular rupture is a trigger for discogenic pain through the action of NGF. In this study, the protein levels of NGF in discs from patients with disc herniation were examined and compared with those from discs of patients with other lumbar degenerative disc diseases.

Methods

Patients (n = 55) with lumbar degenerative disc disease treated by surgery were included. Nucleus pulposus tissue (or herniated disc tissue) was surgically removed and homogenized; protein levels were quantified using an enzyme-linked immunosorbent assay (ELISA) for NGF. Levels of NGF in the discs were compared between 1) patients with herniated discs (herniated group) and those with other lumbar degenerative disc diseases (non-herniated group), and 2) low-grade and high-grade degenerated discs. Patient’s symptoms were assessed using a visual analog scale (VAS) and the Oswestry disability index (ODI); the influence of NGF levels on pre- and post-operative symptoms was examined.

Results

Mean levels of NGF in discs of patients were significantly higher in herniated discs (83.4 pg/mg total protein) than those in non-herniated discs (68.4 pg/mg).No significant differences in levels of NGF were found between low-grade and high-grade degenerated discs. Multivariate analysis, adjusted for age and sex, also showed significant correlation between the presence of disc herniation and NGF levels, though no significant correlation was found between disc degeneration and NGF levels. In both herniated and non-herniated groups, pre-operative symptoms were not related to NGF levels. In the herniated group, post-operative lower extremity pain and low back pain (LBP) in motion were greater in patients with low levels of NGF; no significant differences were found in the non-herniated group.

Conclusions

This study reports that NGF increased in herniated discs, and may play an important role in the generation of discogenic pain. Analysis of patient symptoms revealed that pre-operative NGF levels were related to post-operative residual lower extremity pain and LBP in motion. The results suggest that NGF in the disc is related to pain generation, however, the impact of NGF on generation of LBP varies in individual patients.

Electronic supplementary material

The online version of this article (doi:10.1186/ar4674) contains supplementary material, which is available to authorized users.     

Evidence for the role of G-proteins in flow stimulation of dinoflagellate bioluminescence

Luminescent dinoflagellates respond to flow by the production of light. The primary mechanotransduction event is unknown, although downstream events include a calcium flux in the cytoplasm, a self-propagating action potential across the vacuole membrane, and a proton flux into the cytoplasm that activates the luminescent chemistry. Given the role of GTP-binding (G) proteins in the mechanotransduction of flow by nonmarine cells and the presence of G-proteins in dinoflagellates, it was hypothesized that flow-stimulated dinoflagellate bioluminescence involves mechanotransduction by G-proteins. In the present study, osmotic swelling of cells of the dinoflagellate Lingulodinium polyedrum was used as a drug delivery system to introduce GDPbetaS, an inhibitor of G-protein activation. Osmotically swollen cells produced higher levels of flow-stimulated bioluminescence at a lower threshold of shear stress, indicating they were more flow sensitive. GDPbetaS inhibited flow-stimulated bioluminescence in osmotically swollen cells and in cells that were restored to the isosmotic condition following hypoosmotic treatment with GDPbetaS. These results provide evidence that G-proteins are involved in the mechanotransduction of flow in dinoflagellates and suggest that G-protein involvement in mechanotransduction may be a fundamental evolutionary adaptation.     

Mechanotransduction as a major driver of cell behaviour: mechanisms,and relevance to cell organization and future research

How do cells process environmental cues to make decisions? This simple question is still generating much experimental and theoretical work, at the border of physics, chemistry and biology, with strong implications in medicine. The purpose of mechanobiology is to understand how biochemical and physical cues are turned into signals through mechanotransduction. Here, we review recent evidence showing that (i) mechanotransduction plays a major role in triggering signalling cascades following cell–neighbourhood interaction; (ii) the cell capacity to continually generate forces, and biomolecule properties to undergo conformational changes in response to piconewton forces, provide a molecular basis for understanding mechanotransduction; and (iii) mechanotransduction shapes the guidance cues retrieved by living cells and the information flow they generate. This includes the temporal and spatial properties of intracellular signalling cascades. In conclusion, it is suggested that the described concepts may provide guidelines to define experimentally accessible parameters to describe cell structure and dynamics, as a prerequisite to take advantage of recent progress in high-throughput data gathering, computer simulation and artificial intelligence, in order to build a workable, hopefully predictive, account of cell signalling networks.     

Extradural Corticosteroid Injection in Management of Lumbar Nerve Root Compression

The effect of extradural corticosteroid injection in patients with nerve root compression syndromes associated with degenerative disease of the lumbar intervertebral discs was assessed in a double-blind controlled trial on 100 consecutive inpatients assigned by random allocation to treatment and control groups. Assessment during admission and at three months revealed statistically highly significant differences in respect of relief of pain and resumption of normal occupation in favour of the group treated by extradural injection. This treatment seems to be a valuable adjunct to the management of lumbar nerve root compression syndromes associated with degenerative disc disease.     

哈萨克族成人腰稚间盘高度的放射学测量及其临床意义

目的：为人工椎间盘的设计提供形态学依据。方法：对56例哈萨克族成人腰椎（L）间盘高度进行放射学测量。结果：56例哈萨克族L1-2椎间盘高度男女性之间差异无统计学意义（P〉0．05）,L3-5椎间盘高度男、女性之间差异有统计学意L（P〈0．005-0．001）;哈萨克族与汉族腰椎间盘高度之间差异均有统计学意义（P〈0．005）。结论：哈萨克族腰椎间盘高度均大于汉族,临床上可通过对腰椎间盘间高度的测量,为人工椎间盘假体设计提供参数．     

Moderate mechanical stimulation rescues degenerative annulus fibrosus by suppressing caveolin-1 mediated pro-inflammatory signaling pathway

Mechanical loading can induce or antagonize the extracellular matrix (ECM) synthesis, proliferation, migration, and inflammatory responses of annulus fibrosus cells (AFCs), depending on the loading mode and level. Caveolin-1 (Cav1), the core protein of caveolae, plays an important role in cellular mechanotransduction and inflammatory responses. In the present study, we presented that AFCs demonstrated different behaviors when subjected to cyclic tensile strain (CTS) for 24 h at a magnitude of 0%, 2%, 5% and 12%, respectively. It was found that 5% CTS had positive effects on cell proliferation, migration and anabolism, while 12% CTS had the opposite effects. Besides, cells exposed to interleukin-1β stimulus exhibited an increase expression in inflammatory genes, and the expression of these genes decreased after exposure to moderate mechanical loading with 5% CTS. In addition, 5% CTS decreased the level of Cav1 and integrin β1 and exhibited anti-inflammatory effects. Moreover, the expression of integrin β1 and p-p65 increased in AFCs transfected with Cav1 plasmids. In vivo results revealed that moderate mechanical stimulation could recover the water content and morphology of the discs. In conclusion, moderate mechanical stimulation restrained Cav1-mediated signaling pathway and exhibited anti-inflammatory effects on AFCs. Together with in vivo results, this study expounds the underlying molecular mechanisms on the effect of moderate mechanical stimulation on intervertebral discs (IVDs) and may provide a new therapeutic strategy for the treatment of IVD degeneration.     

Mechanotransduction in adipocytes

Obesity is widely recognized as a major public health problem due to its strong association with a number of serious chronic diseases including hyperlipidemia, hypertension, type II diabetes and coronary atherosclerotic heart disease. During the development of obesity, the positive energy balance involves recruitment of new adipocytes from preadipocytes in adipose tissue, which have proliferated and differentiated. Given that cells in adipose tissues are physiologically exposed to compound mechanical loading: tensile, compressive and shear strains/stresses, which are caused by bodyweight loads as well as by weight-bearing, it is important to determine whether the adipose conversion process is influenced by mechanical stimulations. In this article we provide a comprehensive review of the experimental studies addressing mechanotransduction in adipocytes, as well as of mathematical and computational models that are useful for studying mechanotransduction in adipocytes or for quantifying the responsiveness of adipocytes to different types of mechanical loading. The new understanding that adipogenesis is influenced by mechanical stimulations has the potential to open new and important research paths, driven by mechanotransduction, to explore mechanisms as well as treatment approaches in obesity and related conditions.     

The Effect of Nucleus Pulposus Crosslinking and Glycosaminoglycan Degradation on Disc Mechanical Function

Altered mechanical loading, secondary to biochemical changes in the nucleus pulposus, is a potential mechanism in disc degeneration. An understanding of the role of this altered mechanical loading is only possible by separating the mechanical and biological effects of early nucleus pulposus changes. The objective of this study was to quantify the mechanical effect of decreased glycosaminoglycans (GAG) and increased crosslinking in the nucleus pulposus using in vitro rat lumbar discs. Following initial mechanical testing the discs were injected according to the four treatment groups: PBS control, chondroitinase-ABC (ChABC) for GAG degradation, genipin (Gen) for crosslinking, or a combination of chondroitinase and genipin (ChABC+Gen). After treatment the discs were again mechanically tested, followed by histology or biochemistry. Neutral zone mechanical properties were changed by approximately 20% for PBS, ChABC, and ChABC+Gen treatments (significant only for PBS in a paired comparison). These trends were reversed with genipin crosslinking alone. With ChABC treatment the effective compressive modulus increased and the GAG content decreased; with the combination of ChABC+Gen the mechanics and GAG content were unchanged. Degradation of nucleus pulposus GAG alters disc axial mechanics, potentially contributing to the degenerative cascade. Crosslinking is unlikely to contribute to degeneration, but may be a potential avenue of treatment.     

Animal models of regenerative medicine for biological treatment approaches of degenerative disc diseases

Degenerative disc disease (DDD) is a painful, chronic and progressive disease, which is characterized by inflammation, structural and biological deterioration of the intervertebral disc (IVD) tissues. DDD is specified as cell-, age-, and genetic-dependent degenerative process that can be accelerated by environmental factors. It is one of the major causes of chronic back pain and disability affecting millions of people globally. Current treatment options, such as physical rehabilitation, pain management, and surgical intervention, can provide only temporary pain relief. Different animal models have been used to study the process of IVD degeneration and develop therapeutic options that may restore the structure and function of degenerative discs. Several research works have depicted considerable progress in understanding the biological basis of disc degeneration and the therapeutic potentials of cell transplantation, gene therapy, applications of supporting biomaterials and bioactive factors, or a combination thereof. Since animal models play increasingly significant roles in treatment approaches of DDD, we conducted an electronic database search on Medline through June 2020 to identify, compare, and discuss publications regarding biological therapeutic approaches of DDD that based on intradiscal treatment strategies. We provide an up-to-date overview of biological treatment strategies in animal models including mouse, rat, rabbit, porcine, bovine, ovine, caprine, canine, and primate models. Although no animal model could profoundly reproduce the clinical conditions in humans; animal models have played important roles in specifying our knowledge about the pathophysiology of DDD. They are crucial for developing new therapy approaches for clinical applications.     

Mechanical profiling of intervertebral discs

Despite recent advances in imaging diagnostic technology and additional treatment options our ability to prevent or inhibit discogenic back pain has not drastically improved. The challenge of linking early degenerative patterns to dysfunction and pain remains. Using a novel material testing device designated the tissue diagnostic instrument (TDI) we measured the local stiffness and strain energy absorption in the radial direction of 13 intact intervertebral discs; effectively generating a mechanical profile of each disc. Prior to measuring mechanical properties, an MR image was taken of each spine segment and the discs were radiologically scored according to the Pfirrmann scale. After testing, a sagittal portion of each L1–L2 disc was excised from each of four spines for histology. No significant correlations were found between Pfirrmann grade and mechanical data. However, polarized light microscopy images of disc sections indicated correlations between local tissue modulus measured with the TDI and the clarity and density of lamellar striations.     

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.     

Damage accumulation location under cyclic loading in the lumbar disc shifts from inner annulus lamellae to peripheral annulus with increasing disc degeneration

It is difficult to study the breakdown of lumbar disc tissue over several years of exposure to bending and lifting by experimental methods. In our earlier published study we have shown how a finite element model of a healthy lumbar motion segment was used to predict the damage accumulation location and number of cyclic to failure under different loading conditions. The aim of the current study was to extend the continuum damage mechanics formulation to the degenerated discs and investigate the initiation and progression of mechanical damage. Healthy disc model was modified to represent degenerative discs (Thompson grade III and IV) by incorporating both geometrical and biochemical changes due to degeneration. Analyses predicted decrease in the number of cycles to failure with increasing severity of disc degeneration. The study showed that the damage initiated at the posterior inner annulus adjacent to the endplates and propagated outwards towards its periphery in healthy and grade III degenerated discs. The damage accumulated preferentially in the posterior region of the annulus. However in grade IV degenerated disc damage initiated at the posterior outer periphery of the annulus and propagated circumferentially. The finite element model predictions were consistent with the infrequent occurrence of rim lesions at early age but a much higher incidence in severely degenerated discs.     

Intervertebral disc viscoelastic parameters and residual mechanics spatially quantified using a hybrid confined/in situ indentation method

With advancing age, injury, musculoskeletal pathology or a combination of these, a degenerative cascade of biomechanical, biochemical, and nutritional alterations diminish the intervertebral discs' ability to maintain its structure and function. While the biomechanics of isolated disc tissues has been investigated across this degenerative spectrum, none have attempted to retain the in situ disc-endplate morphology during compressive tissue characterization. The objective of this study was to spatially quantify the viscoelastic parameters of the intervertebral disc throughout degeneration, including the as yet unreported residual stress/strain. This required the development of a hybrid confined/in situ indentation methodology, which preserves the disc structural morphology. At four locations of the disc (anterior-AF, right and left lateral AF, and NP) stress-relaxation tests were performed using the hybrid confined/in situ indentation method, which utilizes the vertebral endplate as the porous indenter tip. This method allows the endplate to remain interwoven with the disc tissue, retaining its native orientation. Healthy disc tissue exhibited significantly higher residual stress values compared to both moderate and severe degeneration in all locations (p<0.0156). Furthermore, the equilibrium stress at 15% strain (stress relaxation) was significantly diminished with advancing disc degeneration (p<0.0241). The equilibrium viscoelastic parameters show healthy discs encounter higher forces at the same strain level, and are able to maintain this force, where degenerated discs are unable to maintain this force throughout time. This morphology-conserved method provides insight into the spatial compressive mechanical properties of the intervertebral disc across the degeneration spectrum and will aid in modeling these tissue changes.