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.

Spontaneous mineralization of the nucleus pulposus (NP) has been observed in cases of intervertebral disc degeneration (IDD). Inflammatory cytokines have been implicated in mineralization of multiple tissues through their modulation of expression of factors that enable or inhibit mineralization, including TNAP, ANKH or ENPP1. This study examines the underlying factors leading to NP mineralization, focusing on the contribution of the inflammatory cytokine, TNF, to this pathologic event. We show that human and bovine primary NP cells express high levels of ANKH and ENPP1, and low or undetectable levels of TNAP. Bovine NPs transduced to express TNAP were capable of matrix mineralization, which was further enhanced by ANKH knockdown. TNF treatment or overexpression promoted a greater increase in mineralization of TNAP-expressing cells by downregulating the expression of ANKH and ENPP1 via NF-κB activation. The increased mineralization was accompanied by phenotypic changes that resemble chondrocyte hypertrophy, including increased RUNX2 and COL10A1 mRNA; mirroring the cellular alterations typical of samples from IDD patients. Disc organ explants injected with TNAP/TNF- or TNAP/shANKH-overexpressing cells showed increased mineral content inside the NP. Together, our results confirm interactions between TNF and downstream regulators of matrix mineralization in NP cells, providing evidence to suggest their participation in NP calcification during IDD.

Genetic tools such as the Cre-Lox reporter system are powerful aids for tissue-specific cell tracking. For example, it would be useful in examining intervertebral disc (IVD) cell populations in normal and diseased states. A Cre recombinase and its recognition site, loxP have been adapted from the bacteriophage for use in genetic manipulation. The reporter mice used here express the red fluorescent protein, tdTomato with flanking LoxP sites (Rosa26 TdTomato mice). We compared two different Collagen type II (Col2) promoter constructs that drive Cre-recombinase expression in mice: (a) Col2-Cre, which allows constitutive Cre-recombinase expression under the control of the Col2 promoter/enhancer and (b) Col2-CreER, which contains a shorter promoter/enhancer region than Col2-Cre, but has human estrogen binding elements that bind tamoxifen, resulting in Cre-recombinase expression. The goal of the study is to characterize Cre-recombinase distribution pattern in Col2-Cre and Col2-CreER mice using tdTomato as reporter in the spine. The expression patterns of these two mice were further compared with Col2 gene expression in the native mouse NP and AF tissues by real-time PCR. We crossed Col2-Cre mice or Col2-CreER mice with the tdTomato reporter mice, and compared the tdTomato expression patterns. Col2-CreER/tdTomato mice were injected with tamoxifen at postnatal day 7 to activate the Cre-recombinase. TdTomato in the constitutively active Col2-Cre mice was detected in the nucleus pulposus (NP), the entire annulus fibrosus (AF), and in cartilaginous endplate and growth plate cells in the lower lumbar and coccygeal spine. In contrast, when Col2-CreER activity was induced by tamoxifen at P7, tdTomato was limited to the inner AF, and was absent from the NP. We have described the differences in Col2 reporter gene expression, in Col2-Cre/tdTomato and Col2-Cre-ER/tdTomato mouse IVD. The information provided here will help to guide future investigations of IVD biology.

The intervertebral disc (IVD) is composed of the external annulus fibrosus (AF) and the inner gel-like center, the nucleus pulposus (NP). The elastic NP can function to relieve stress and maintain IVD function by distributing hydraulic pressure evenly to annulus and endplate. Degeneration of the NP, which leads to increased death of NP cells, the loss of proteoglycan (PG), and aberrant gene expression, may result in an overall alteration of the biomechanics of the spinal column and cause low back pain. Recent advances in biological therapy strategies that target therapy at the regeneration of degenerated and damaged NP have been investigated in in vitro and in vivo studies and demonstrated promising outcomes. In this article, we review recent studies of biological approaches for NP regeneration.

The articles regarding NP regeneration using biomaterials, stem cells, and gene vectors were identified in PubMed databases.

Stem cell-mediated cell therapy demonstrates the potential to restore the function and structure of the NP. The viral or non-viral vectors encoding functional genes may generate a therapeutic effect when they are introduced into grafted cells or native cells in the NP. Biomaterial scaffolds generate an initial permissive environment for cell growth and allow the remodeling of scaffolds in the regeneration process. Biomaterial scaffolds provide structural support for NP regeneration and serve as a carrier for stem cell and gene vector delivery.

Though recent studies advance the body of knowledge needed to treat degenerated discs, many challenges need to be overcome before the application of these approaches can be successful clinically.

The intervertebral disc (IVD) is the most avascular and acellular tissue in the body and therefore prone to degeneration. During IVD degeneration, the balance between anabolic and catabolic processes in the disc is deregulated, amongst others leading to alteration of extracellular matrix production, abnormal enzyme activities and production of pro-inflammatory substances like cytokines. The established treatment strategy for IVD degeneration consists of physiotherapy, pain medication by drug therapy and if necessary surgery. This approach, however, has shown limited success. Alternative strategies to increase and prolong the effects of bioactive agents and to reverse the process of IVD degeneration include the use of delivery systems for drugs, proteins, cells and genes. In view of the specific anatomy and physiology of the IVD and depending on the strategy of the therapy, different delivery systems have been developed which are reviewed in this article.

The healthy intervertebral disc (IVD) fulfils the essential function of load absorption, while maintaining multi-axial flexibility of the spine. The interrelated tissues of the IVD, the annulus fibrosus, the nucleus pulposus, and the cartilaginous endplate, are characterised by their specific niche, implying avascularity, hypoxia, acidic environment, low nutrition, and low cellularity. Anabolic and catabolic factors balance a slow physiological turnover of extracellular matrix synthesis and breakdown. Deviations in mechanical load, nutrient supply, cellular activity, matrix composition and metabolism may initiate a cascade ultimately leading to tissue dehydration, fibrosis, nerve and vessel ingrowth, disc height loss and disc herniation. Spinal instability, inflammation and neural sensitisation are sources of back pain, a worldwide leading burden that is challenging to cure. In this review, advances in cell and molecular therapy, including mobilisation and activation of endogenous progenitor cells, progenitor cell homing, and targeted delivery of cells, genes, or bioactive factors are discussed.

Lumbar discectomy is the surgical procedure most frequently performed for patients suffering from low back pain and sciatica. Disc herniation as a consequence of degenerative or traumatic processes is commonly encountered as the underlying cause for the painful condition. While discectomy provides favourable outcome in a majority of cases, there are conditions where unmet requirements exist in terms of treatment, such as large disc protrusions with minimal disc degeneration; in these cases, the high rate of recurrent disc herniation after discectomy is a prevalent problem. An effective biological annular repair could improve the surgical outcome in patients with contained disc herniations but otherwise minor degenerative changes. An attractive approach is a tissue-engineered implant that will enable/stimulate the repair of the ruptured annulus. The strategy is to develop three-dimensional scaffolds and activate them by seeding cells or by incorporating molecular signals that enable new matrix synthesis at the defect site, while the biomaterial provides immediate closure of the defect and maintains the mechanical properties of the disc. This review is structured into (1) introduction, (2) clinical problems, current treatment options and needs, (3) biomechanical demands, (4) cellular and extracellular components, (5) biomaterials for delivery, scaffolding and support, (6) pre-clinical models for evaluation of newly developed cell- and material-based therapies, and (7) conclusions. This article highlights that an interdisciplinary approach is necessary for successful development of new clinical methods for annulus fibrosus repair. This will benefit from a close collaboration between research groups with expertise in all areas addressed in this review.

Closure and biological repair of anulus fibrosus (AF) defects in intervertebral disc diseases is a therapeutic challenge. The aim of our study was to evaluate the anabolic properties of bioactive factors on cartilaginous matrix formation by AF cells. Human AF cells were harvested from degenerated lumbar AF tissue and expanded in monolayer culture. AF cell differentiation and matrix formation was initiated by forming pellet cultures and stimulation with hyaluronic acid (HA), human serum (HS), fibroblast growth factor-2 (FGF-2), transforming growth factor-β3 (TGF-β3) and TGF-β3/FGF-2 for up to 4 weeks. Matrix formation was assessed histologically by staining of proteoglycan, type I and type II collagens and by gene expression analysis of typical extracellular matrix molecules and of catabolic matrix metalloproteinases MMP-2 and MMP-13. AF cells, stimulated with HS, FGF-2 and most pronounced with TGF-β3 or TGF-β3/FGF-2 formed a cartilaginous matrix with significantly enhanced expression of matrix molecules and of MMP-13. Stimulation of AF cells with TGF-β3 was accompanied by induction of type X collagen, known to occur in hypertrophic cartilage cells having mineralizing potential. HA did not show any chondro-inductive characteristics. These findings suggest human serum, FGF-2 and TGF-β3 as possible candidates to support biological treatment strategies of AF defects.

Although understanding of the biologic basis of intervertebral disk (IVD) degeneration is rapidly advancing, the unique IVD environment presents challenges to the development and delivery of biologic treatments. Acceleration of cellular senescence and apoptosis in degenerative IVDs and the depletion of matrix proteins have prompted the development of treatments based on replacing IVD cells using various cell sources. However, this strategy has not been tested in animal models. IVD degeneration and associated pain have led to interest in pathologic innervation of the IVD and ultimately to the development of percutaneous devices to ablate afferent nerve endings in the posterior annulus. Degeneration leads to changes in the expression of matrix protein, cytokines, and proteinases. Injection of growth factors and mitogens may help overcome these degenerative changes in IVD phenotype, and these potential treatments are being explored in animal studies. Gene therapy is an elegant method to address changes in protein expression, but efforts to apply this technology to IVD degeneration are still at early stages.

Bone morphogenetic protein-7 (BMP-7) was found to stimulate the synthesis of proteoglycans (PGs) and collagen type II. To increase the biological function of the nucleus pulposus (NP) cells, the Ad-hBMP-7 vector was also successfully constructed and transfected NP cells. However, the disadvantages of adenovirus limit the usefulness of the Ad-hBMP7 vector for clinical application. The rAAV2 vector has empirical advantages, especially for clinical use, to transfer exogenous genes into cells. The purpose of this study was to first determine whether a rAAV2-hBMP-7 vector could be used to transfect canine NP cells and effect on the biological functions of canine NP cells. The canine NP cells transfected by the rAAV-BMP7 were assessed semi-quantitatively for BMP-7 expression with real-time PCR and westernbloting. Aggrecan and collagens type I and II secreted by the NP cells were qualitatively assessed at 4, 7, and 14 days post-transfection in the transfection and control groups. We found that rAAV2 can successfully transfer the hBMP-7 gene into canine NP cells. NP cells transfected by the rAAV-hBMP-7 vector express hBMP-7 for at least 14 days. At 7 and 14 days, the expressed hBMP-7 promotes a remarkable and significant accumulation of both proteoglycans (42% and 77% higher than non-transfected cells) (p<0.05) and collagen type II (63% and 94% higher than non-transfected cells) (p<0.05). Thus, we could speculate that the rAAV-based gene delivery technique promotes the expression of proteoglycans and collagen type II of nucleus pulposus cells. Moreover, this technique may be applicable for the future treatment of degenerative disc disease.

Intervertebral disc degeneration is a primary cause of low back pain and has a high societal cost. The pathological mechanism by which the intervertebral disc degenerates is largely unknown. Cell-based therapy especially using bone marrow mesenchymal stem cells as seeds for transplantation, although still in its infancy, is proving to be a promising, realistic approach to intervertebral disc regeneration. This article reviews current advances regarding regeneration potential in both the in vivo and vitro studies of bone marrow mesenchymal stem cell-based therapy and discusses the up-to-date regeneration mechanisms of stem cell transplantation for treating intervertebral disc degeneration.