Fate of notochord descendent cells in the intervertebral disc

Abstract

Back pain is a prevalent musculoskeletal disorder affecting populations worldwide. Degeneration of the intervertebral discs (IVD) is indicated to be one of the main etiologic factors of back pain. The loss of proteoglycan and consequently dehydration and reduction of swelling of the intervertebral disc core, the nucleus pulposus (NP), is one of the earliest degenerative events. The replacement of large vacuolated notochordal cells by the smaller chondrocyte-like cells in the NP coincide with the onset of IVD degeneration, the loss of the notochordal cells was therefore postulated to be a cause of the degeneration and associated with ageing .

To date, the true origin of the smaller chondrocyte-like cells was still under dispute. In this study, Egfp reporter was constitutively and permanently activated in the notochord via specific Cre expression by the Foxa2mNE enhancer under the recombination system in the Foxa2mNE-cre; Z/EG double transgenic mice. The notochord descendent cells were therefore labeled with green fluorescent protein (GFP) and their localization and expression characteristic were tracked during development, maturation, ageing of mouse tail IVD. To further investigate the cellular changes during degeneration, disc degeneration in the murine disc was induced by puncture. The GFP labeled notochord cells were demonstrated to be entrapped and remained in the NP of the murine disc. With the validation of degeneration by the multichromatic FAST staining, a grading system based on altered sulfated glycosaminoglycan content and cellular organization specialized to murine disc was proposed. The segregation of the notochordal NP cell mass segregation was found to be a common pathway of both age-related and puncture-induced degeneration. While apoptosis and loss of NP cells was presented in puncture-induced disc degeneration, invasion of lamellae cells was only found in severely degenerated discs. Consistent with the findings of the murine puncture model, increased expression of Col2a1 and Col1a1 were demonstrated in punctured discs by ISH, which the notochordal NP cells were showing a remarkable elevation in expression. The data supports a chondrogenic differentiation of the endogenous notochordal cells instead of invasion of chondrocyte-like cells into the NP. Three NP markers identified as a common genes from microarray of rat and human NP, (Carbonic anhydrase3 (Car3), UDP-N-acetyl-alpha-D-galactosamine:polypeptide; N-acetylgalactosaminyltransferase 3 (Galnt3) and HOP homeobox (Obl)), were found specifically co-localized with the notochord descendent cells in all ages and different time-points after puncture, suggesting them as novel but reliable markers for notochordal NP cells. This study provides direct evidence of the origin of NP cells in aged and degenerated IVDs using transgenic mice, and thus enhances the understanding of the role of notochordal cells in the progression of IVD degeneration.