Abstract
IntroductionEarly degenerative changes in the nucleus pulposus (NP) are observed after the disappearance of notochordal cells (NCs). Thus, it has been suggested that NCs play an important role in maintaining the NP and may have a regenerative potential on other cells of the NP. As the number of resident NP cells (NPCs) decreases in a degenerating disc, mesenchymal stromal (stem) cells (MSCs) may be used for cell supplementation. In this study, using cells of one species, the regenerative potential of canine NCs was assessed in long-term three-dimensional coculture with canine NPCs or MSCs.MethodsCanine NCs and canine NPCs or MSCs were cocultured in alginate beads for 28 days under hypoxic and high-osmolarity conditions. Cell viability, cell morphology and DNA content, extracellular matrix production and expression of genes related to NC markers (Brachyury, KRT18) and NP matrix production (ACAN, COL2A1, COL1A1) were assessed after 1, 15 and 28 days of culture.ResultsNCs did not completely maintain their phenotype (morphology, matrix production, gene expression) during 28 days of culture. In cocultures of NPCs and NCs, both extracellular matrix content and anabolic gene expression remained unchanged compared with monoculture groups, whereas cocultures of MSCs and NCs showed increased glycosaminoglycan/DNA. However, the deposition of these proteoglycans was observed near the NCs and not the MSCs. Brachyury expression in the MSC and NC coculture group increased in time. The latter two findings indicate a trophic effect of MSCs on NCs rather than vice versa.ConclusionsNo regenerative potential of canine NCs on canine NPCs or MSCs was observed in this study. However, significant changes in NC phenotype in long-term culture may have resulted in a suboptimal regenerative potential of these NCs. In this respect, NC-conditioned medium may be better than coculture for future studies of the regenerative potential of NCs.Electronic supplementary materialThe online version of this article (doi:10.1186/s13075-015-0569-6) contains supplementary material, which is available to authorized users.
Highlights
Degenerative changes in the nucleus pulposus (NP) are observed after the disappearance of notochordal cells (NCs)
No regenerative potential of canine NCs on canine Nucleus pulposus cell (NPC) or mesenchymal stromal cells (MSC) was observed in this study
Future studies We propose that NC-conditioned medium, produced in culture conditions that support the NC phenotype, is a better alternative for evaluating the regenerative effect of NCs on NPC or MSCs, as it overcomes the drawbacks of loss of NC phenotype and bidirectional intercellular communication in culture
Summary
Degenerative changes in the nucleus pulposus (NP) are observed after the disappearance of notochordal cells (NCs). Degeneration of the IVD involves the transition from a gelatinous to a fibrotic NP [3] This change is associated with a decreased ability of the NP to convert compressive forces into evenly distributed tensile stresses in the surrounding annulus fibrosus (AF), with consequent degeneration of the AF [4]. Both the resident cells and extracellular matrix of the NP undergo major changes in this degenerative process. The healthy gelatinous NP is rich in proteoglycans that keep the tissue hydrated, whereas in the degenerated NP, matrix contains less proteoglycans, different collagen types and more matrix-degrading enzymes [5]
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