Versican gene expression in human articular cartilage and comparison of mRNA splicing variation with aggrecan

Abstract

The chondrocytes in human articular cartilage from subjects of all ages express mRNAs for both of the aggregating proteoglycans aggrecan and versican, although the level of expression of versican mRNA is much lower than that of aggrecan mRNA. Aggrecan shows alternative splicing of the epidermal growth factor (EGF)-like domain within its C-terminal globular region, but there is no evidence for a major difference in situ in the relative expression of this domain with age. At all ages studied from birth to the mature adult, a greater proportion of transcripts lacked the EGF domain. The relative proportions of the two transcripts did not change upon culture and passage of isolated chondrocytes. In contrast, the neighbouring complement regulatory protein (CRP)-like domain was predominantly expressed irrespective of age, but cell culture did result in variation of the splicing of this domain. Versican possesses two EGF-like domains and one CRP-like domain, but at all ages the three domains were predominantly present in all transcripts. This situation persisted upon culture and passage of the chondrocytes. Thus, unlike aggrecan, the versican expressed by human articular cartilage does not appear to undergo alternative splicing of its C-terminal globular region, either in cartilage in situ or in chondrocytes in culture.

Fibronectin regulates proteoglycan production balance in transforming growth factor-β1-induced chondrogenesis

“…Versican is a large chondroitin sulfate proteoglycan expressed by human fibroblasts. The expression levels of versican are much lower than those of aggrecan in human articular cartilage, although the chondrocytes in human articular cartilage express mRNA for both aggrecan and versican (20). Bovine microarray data revealed that the aggrecan expression in articular cartilage is higher than that in the annulus fibrosus, whereas versican expression in articular cartilage is lower than that in the annulus fibrosus (21).…”

Alternative Splicing in the Aggrecan G3 Domain Influences Binding Interactions with Tenascin-C and Other Extracellular Matrix Proteins

“…Alternative splicing of the EGF and SCR modules in the G3 domain is well established (22)(23)(24)(25)(26) and could have an effect on the interactions of the C-type lectin repeat. Although there are binding studies using full-length proteoglycan (17) or recombinant G3 domains (14,15,20), no thorough investigation of the effects of alternative splicing in the G3 domain has been published.…”

“…The other modules of the G3 domains are subject to alternative splicing (22)(23)(24)(25)(26). The expression of the SCR module is variable, but in humans this module is usually present, regardless of age (26). The EGF1 and EGF2 modules are expressed to a lower extent (25-28 and 5-8%, respectively, in humans) (22).…”

“…The C-type lectin module is constitutively expressed and is the mediator of all the G3 interactions listed above. The other modules of the G3 domains are subject to alternative splicing (22)(23)(24)(25)(26). The expression of the SCR module is variable, but in humans this module is usually present, regardless of age (26).…”

Limitations of using aggrecan and type X collagen as markers of chondrogenesis in mesenchymal stem cell differentiation

“…The 30 cycles of PCR determined to be in the linear range for GAPDH, aggrecan, and types I and II collagens, included denaturation (958C, 45 s), annealing (588C, 45 s), and extension (728C, 60 s). 20 Similarly, the 30 cycles for type X collagen PCR included denaturation (958C, 60 s), annealing (508C, 60 s), and extension (728C, 5 min). 21 Amplified products were analyzed by electrophoresis on 2% agarose gels and visualized by ethidium bromide staining.…”