Abstract

IntroductionThe decreased disc height characteristic of intervertebral disc (IVD) degeneration has often been linked to low back pain, and thus regeneration strategies aimed at restoring the disc extracellular matrix and ultimately disc height have been proposed as potential treatments for IVD degeneration. One such therapy under investigation by a number of groups worldwide is the use of autologous mesenchymal stem cells (MSCs) to aid in the regeneration of the IVD extracellular matrix. To date, however, the optimum method of application of these cells for regeneration strategies for the IVD is unclear, and few studies have investigated the direct injection of MSCs alone into IVD tissues. In the present article, we investigated the survival and phenotype of human MSCs, sourced from aged individuals, following injection into nucleus pulposus (NP) tissue explant cultures.MethodsHuman MSCs extracted from bone marrow were expanded in monolayer culture and, after labelling with adenoviral vectors carrying the green fluorescent protein transcript, were injected into NP tissue explants (sourced from bovine caudal discs) and maintained in culture for 2, 7, 14 and 28 days post injection. Following fixation and paraffin embedding, cell viability was assessed using in situ hybridisation for polyA-mRNA and using immunohistochemistry for caspase 3. Immunohistochemistry/fluorescence for aggrecan, Sox-9 and types I, II and X collagen together with Alizarin red staining was employed to investigate the MSC phenotype and matrix formation.ResultsMSCs were identified in all injected tissue samples and cell viability was maintained for the 4 weeks investigated. MSCs displayed cellular staining for Sox-9, and displayed cellular and matrix staining for aggrecan and type II collagen that increased during culture. No type I collagen, type X collagen or Alizarin red staining was observed at any time point.ConclusionsMSCs from older individuals differentiate spontaneously into chondrocyte-like NP cells upon insertion into NP tissue in vitro, and thus may not require additional stimulation or carrier to induce differentiation. This is a key finding, as such a strategy would minimise the level of external manipulation required prior to insertion into the patient, thus simplifying the treatment strategy and reducing costs.

Highlights

  • The decreased disc height characteristic of intervertebral disc (IVD) degeneration has often been linked to low back pain, and regeneration strategies aimed at restoring the disc extracellular matrix and disc height have been proposed as potential treatments for IVD degeneration

  • Evidence from studies investigating the pathogenesis of IVD degeneration illustrates that IVD degeneration originates from the nucleus pulposus (NP), where both type II collagen and proteoglycan synthesis and content decrease [8,9], the NP is the area of the disc that is targeted by a number of groups worldwide for regeneration strategies [10,11]

  • Ad-green fluorescent protein (GFP)-labelled mesenchymal stem cells (MSCs) were identified in all tissue samples in which Ad-GFP-infected MSCs were injected and cells were observed in the vicinity of the injection site at all time points post injection

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Summary

Introduction

The decreased disc height characteristic of intervertebral disc (IVD) degeneration has often been linked to low back pain, and regeneration strategies aimed at restoring the disc extracellular matrix and disc height have been proposed as potential treatments for IVD degeneration. One such therapy under investigation by a number of groups worldwide is the use of autologous mesenchymal stem cells (MSCs) to aid in the regeneration of the IVD extracellular matrix. We have shown that IVD cells derived from a degenerate disc display an abnormal phenotype, with increased catabolic and decreased anabolic activity, and are not the ideal cell type to stimulate regeneration, and could even lead to accelerated degeneration of the treated IVD [18,19,20,21,22,23,24]

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