Dataset Information

Differential gene expression of the intermediate and outer interzone layers of developing articular cartilage in murine embryos.

ABSTRACT: Nascent embryonic joints, interzones, contain a distinct cohort of progenitor cells responsible for the formation of the majority of articular tissues. However, to date the interzone has largely been studied using in situ analysis for candidate genes in the context of the embryo rather than using an unbiased genome-wide expression analysis on isolated interzone cells, leaving significant controversy regarding the exact role of the intermediate and outer interzone layers in joint formation. Therefore, in this study, using laser capture microdissection (three biological replicates), we selectively harvested the intermediate and outer interzones of mouse embryos at gestational age 15.5 days, just prior to cavitation, when the differences between the layers should be most profound. Microarray analysis (Agilent Whole Mouse Genome Oligo Microarrays) was performed and the differential gene expression between the intermediate interzone cells and outer interzone cells was examined by performing a two-sided paired Student's t-test and pathway analysis. One hundred ninety-seven genes were differentially expressed (? 2-fold) between the intermediate interzone and the outer interzone with a P-value ? 0.01. Of these, 91 genes showed higher expression levels in the intermediate interzone and 106 were expressed higher in the outer interzone. Pathway analysis of differentially expressed genes suggests an important role for inflammatory processes in the interzone layers, especially in the intermediate interzone, and hence in joint and articular cartilage development. The high representation of genes relevant to chondrocyte hypertrophy and endochondral ossification in the outer interzone suggests that it undergoes endochondral ossification.

SUBMITTER: Jenner F

PROVIDER: S-EPMC4120811 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature

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Differential gene expression of the intermediate and outer interzone layers of developing articular cartilage in murine embryos.

Jenner Florien F IJpma Arne A Cleary Mairead M Heijsman Daphne D Narcisi Roberto R van der Spek Peter J PJ Kremer Andreas A van Weeren René R Brama Pieter P van Osch Gerjo J V M GJ

Stem cells and development 20140616 16

Nascent embryonic joints, interzones, contain a distinct cohort of progenitor cells responsible for the formation of the majority of articular tissues. However, to date the interzone has largely been studied using in situ analysis for candidate genes in the context of the embryo rather than using an unbiased genome-wide expression analysis on isolated interzone cells, leaving significant controversy regarding the exact role of the intermediate and outer interzone layers in joint formation. There ...[more]

PMID: 24738827

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Project description:Articular cartilage (AC) is a specialized connective tissue that covers the ends of long bones and facilitates the load-bearing of joints. It consists of chondrocytes distributed throughout an extracellular matrix and organized into three zones: superficial, middle, and deep. Nuclear magnetic resonance (NMR) techniques can be used to characterize this layered structure. In this study, devoted to a better understanding of the NMR response of this complex tissue, 20 specimens excised from femoral and tibial cartilage of bovine samples were analyzed by the low-field single-sided NMR-MOUSE-PM10. A multiparametric depth-wise analysis was performed to characterize the laminar structure of AC and investigate the origin of the NMR dependence on depth. The depth dependence of the single parameters T1, T2, and D has been described in literature, but their simultaneous measurement has not been fully exploited yet, as well as the extent of their variability. A novel parameter, α, evaluated by applying a double-quantum-like sequence, has been measured. The significant decrease in T1, T2, and D from the middle to the deep zone is consistent with depth-dependent composition and structure changes of the complex matrix of fibers confining and interacting with water. The α parameter appears to be a robust marker of the layered structure with a well-reproducible monotonic trend across the zones. The discrimination of cartilage zones was reinforced by a multivariate principal component analysis statistical analysis. The large number of samples allowed for the identification of the smallest number of parameters or their combination able to classify samples. The first two components clustered the data according to the different zones, highlighting the sensitivity of the NMR parameters to the structural and compositional variations of AC. Using two parameters, the best result was obtained by considering T1 and α. Single-sided NMR devices, portable and low-cost, provide information on NMR parameters related to tissue composition and structure.

Project description:BackgroundArticular cartilage undergoes an important maturation process from neonate to adult that is reflected by alterations in matrix protein organization and increased heterogeneity of chondrocyte morphology. In the horse, these changes are influenced by exercise during the first five months of postnatal life. Transcriptional profiling was used to evaluate changes in articular chondrocyte gene expression during postnatal growth and development.MethodsTotal RNA was isolated from the articular cartilage of neonatal (0-10 days) and adult (4-5 years) horses, subjected to one round of linear RNA amplification, and then applied to a 9,367-element equine-specific cDNA microarray. Comparisons were made with a dye-swap experimental design. Microarray results for selected genes (COL2A1, COMP, P4HA1, TGFB1, TGFBR3, TNC) were validated by quantitative polymerase chain reaction (qPCR).ResultsFifty-six probe sets, which represent 45 gene products, were up-regulated (p < 0.01) in chondrocytes of neonatal articular cartilage relative to chondrocytes of adult articular cartilage. Conversely, 586 probe sets, which represent 499 gene products, were up-regulated (p < 0.01) in chondrocytes of adult articular cartilage relative to chondrocytes of neonatal articular cartilage. Collagens, matrix-modifying enzymes, and provisional matrix non-collagenous proteins were expressed at higher levels in the articular cartilage of newborn foals. Those genes with increased mRNA abundance in adult chondrocytes included leucine-rich small proteoglycans, matrix assembly, and cartilage maintenance proteins.ConclusionDifferential expression of genes encoding matrix proteins and matrix-modifying enzymes between neonates and adults reflect a cellular maturation process in articular chondrocytes. Up-regulated transcripts in neonatal cartilage are consistent with growth and expansion of the articular surface. Expression patterns in mature articular cartilage indicate a transition from growth to homeostasis, and tissue function related to withstanding shear and weight-bearing stresses.

Dataset Information

Differential gene expression of the intermediate and outer interzone layers of developing articular cartilage in murine embryos.

Publications

Differential gene expression of the intermediate and outer interzone layers of developing articular cartilage in murine embryos.

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