Transcriptional profiling of bovine intervertebral disc cells: implications for identification of normal and degenerate human intervertebral disc cell phenotypesReport as inadecuate




Transcriptional profiling of bovine intervertebral disc cells: implications for identification of normal and degenerate human intervertebral disc cell phenotypes - Download this document for free, or read online. Document in PDF available to download.

Arthritis Research and Therapy

, 12:R22

First Online: 11 February 2010Received: 14 August 2009Revised: 13 November 2009Accepted: 11 February 2010

Abstract

IntroductionNucleus pulposus NP cells have a phenotype similar to articular cartilage AC cells. However, the matrix of the NP is clearly different to that of AC suggesting that specific cell phenotypes exist. The aim of this study was to identify novel genes that could be used to distinguish bovine NP cells from AC and annulus fibrosus AF cells, and to further determine their expression in normal and degenerate human intervertebral disc IVD cells.

MethodsMicroarrays were conducted on bovine AC, AF and NP cells, using Affymetrix Genechip Bovine Genome Arrays. Differential expression levels for a number of genes were confirmed by quantitative real time polymerase chain reaction qRT-PCR on bovine, AC, AF and NP cells, as well as separated bovine NP and notochordal NC cells. Expression of these novel markers were further tested on normal human AC, AF and NP cells, and degenerate AF and NP cells.

ResultsMicroarray comparisons between NP-ACandAF and NP-AC identified 34 NP-specific and 49 IVD-specific genes respectively that were differentially expressed ≥100 fold. A subset of these were verified by qRT-PCR and shown to be expressed in bovine NC cells. Eleven genes SNAP25, KRT8, KRT18, KRT19, CDH2, IBSP, VCAN, TNMD, BASP1, FOXF1 and FBLN1 were also differentially expressed in normal human NP cells, although to a lesser degree. Four genes SNAP25, KRT8, KRT18 and CDH2 were significantly decreased in degenerate human NP cells, while three genes VCAN, TNMD and BASP1 were significantly increased in degenerate human AF cells. The IVD negative marker FBLN1 was significantly increased in both degenerate human NP and AF cells.

ConclusionsThis study has identified a number of novel genes that characterise the bovine and human NP and IVD transcriptional profiles, and allows for discrimination between AC, AF and NP cells. Furthermore, the similarity in expression profiles of the separated NP and NC cell populations suggests that these two cell types may be derived from a common lineage. Although interspecies variation, together with changes with IVD degeneration were noted, use of this gene expression signature will benefit tissue engineering studies where defining the NP phenotype is paramount.

AbbreviationsA2Mα-2-macroglobulin

ACarticular cartilage

ACANaggrecan

AFannulus fibrosus

ANOVAanalysis of variance

ANXA4annexin A4

AQP1aquaporin

BASP1brain abundant: membrane attached signal protein 1

CDH2N-cadherin

CEPcartilaginous end plates

COL2A1type II collagen

COL10A1collagen type X, alpha 1

COMPcartilage oligomeric matrix protein

DSCdesmocollin 2

ECMextracellular matrix

FBLN1fibulin 1

fdrfalse discovery rate

FOXF1forkhead box F1

FOXF2forkhead box F2

GLUT1glucose transporter type 1

GPCglypican

HandEhematoxylin and eosin

HIF1hypoxia inducible factor 1 isoforms

HSPShigh salt precipitation solution

IBSPintegrin-binding sialoprotein

IVDintervertebral disc

KRTcytokeratin

LBPlow back pain

MGPmatrix Gla protein

MMPmatrix metalloproteinase

MSCsmesenchymal stem cells

NCnotochord

NCAM1neural cell adhesion molecule

NPnucleus pulposus

PCAprincipal component analysis

PGproteoglycan

PPLRprobability of positive log-ratio

PTNpleiotrophin

qRT-PCRquantitative real-time polymerase chain reaction

RINRNA integrity number

SNAP25synaptosomal-associated protein, 25 kDa

SOSTDC1sclerostin domain containing 1

Tbrachyury homolog

TNFAIP6tumor necrosis factor, alpha-induced protein 6

TNMDtenomodulin

VCANversican

VEGFvascular endothelial growth factor

VIMvimentin.

Electronic supplementary materialThe online version of this article doi:10.1186-ar2929 contains supplementary material, which is available to authorized users.

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Author: Ben M Minogue - Stephen M Richardson - Leo AH Zeef - Anthony J Freemont - Judith A Hoyland

Source: https://link.springer.com/







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