Differentially Expressed MicroRNAs in Chondrocytes from Distinct Regions of Developing Human Cartilage

Abstract

There is compelling in vivo evidence from reports on human genetic mutations and transgenic mice that some microRNAs (miRNAs) play an important functional role in regulating skeletal development and growth. A number of published in vitro studies also point toward a role for miRNAs in controlling chondrocyte gene expression and differentiation. However, information on miRNAs that may regulate a specific phase of chondrocyte differentiation (i.e. production of progenitor, differentiated or hypertrophic chondrocytes) is lacking. To attempt to bridge this knowledge gap, we have investigated miRNA expression patterns in human embryonic cartilage tissue. Specifically, a developmental time point was selected, prior to endochondral ossification in the embryonic limb, to permit analysis of three distinct populations of chondrocytes. The location of chondroprogenitor cells, differentiated chondrocytes and hypertrophic chondrocytes in gestational day 54–56 human embryonic limb tissue sections was confirmed both histologically and by specific collagen expression patterns. Laser capture microdissection was utilized to separate the three chondrocyte populations and a miRNA profiling study was carried out using TaqMan® OpenArray® Human MicroRNA Panels (Applied Biosystems®). Here we report on abundantly expressed miRNAs in human embryonic cartilage tissue and, more importantly, we have identified miRNAs that are significantly differentially expressed between precursor, differentiated and hypertrophic chondrocytes by 2-fold or more. Some of the miRNAs identified in this study have been described in other aspects of cartilage or bone biology, while others have not yet been reported in chondrocytes. Finally, a bioinformatics approach was applied to begin to decipher developmental cellular pathways that may be regulated by groups of differentially expressed miRNAs during distinct stages of chondrogenesis. Data obtained from this work will serve as an important resource of information for the field of cartilage biology and will enhance our understanding of miRNA-driven mechanisms regulating cartilage and endochondral bone development, regeneration and repair.

SOX9 and the many facets of its regulation in the chondrocyte lineage

“…Sequence analysis indicates that the 3′UTR of the human SOX9 mRNA features recognition sites for six distinct miRNAs: miR-101, miR-1/205, miR-590/590-3p, miR-145, miR-300, and miR-384-5p (Figure 4A). None of these miRNAs figures among the miRNAs detected in human embryonic growth plate cartilage (36), but miR-145 was reported expressed in vivo in bovine articular cartilage and in vitro in human and mouse mesenchymal stem cells, articular chondrocytes and chondrosarcoma cells (37–40). Its binding to the SOX9 RNA was shown to reduce the level of SOX9 protein.…”

MiR-26a modulates extracellular matrix homeostasis in cartilage

“…Several transcriptome screening approaches identified miRNAs expressed in cartilage, but their function remains mostly unknown [10]. Here, we used a different strategy to identify and define the function of a novel miRNA expressed in cartilage.…”

“…The expression was normalized to the expression of miR-92. This miRNA is one of the most abundantly expressed miRNAs in growth plate cartilage [10] and was constantly expressed in the proliferative, prehypertrophic and hypertrophic zones when analyzing similar amounts of isolated and transcribed microdissected material from growth plate cartilage. The fold change was calculated with the delta-delta C(T) method [20].…”

MicroRNAs in orthopaedic research: Disease associations, potential therapeutic applications, and perspectives

“…183 Among the differentially-expressed miRNA, miR-140 and miR-181a/d were found to be significantly higher in the healing fractures. This finding makes sense because miR-140 and miR-181 are known to be highly expressed in hypertrophic cartilage tissue 131,134 and endochondral fracture repair occurs via formation of a hypertrophic cartilage callus.…”