Identification of novel osteochondrosis— Associated genes

Abstract

During the early stages of articular osteochondrosis, cartilage is retained in subchondral bone, but the pathophysiology of this condition of growing humans and domestic animals is poorly understood. A subtractive hybridization study was undertaken to compare gene expression between the cartilage of early experimentally induced equine osteochondrosis lesions and control cartilage. Of the many putative differentially expressed genes identified, eight were confirmed by quantitative PCR analysis as differentially expressed, in addition to those already known to be associated with early lesions. Genes encoding vacuolar H þ -ATPase V 0 subunit d 2 (ATP6V0D2), cathepsin K, integrin-binding sialoprotein, integrin aV, low density lipoprotein receptor-related protein 4, lumican, osteopontin, and thymosin b4 (TMSB4) were expressed at higher levels in lesions than in control cartilage. These genes included 34 genes not previously identified in cartilage. Some genes identified as associated with early lesions are known chondrocyte hypertrophyassociated genes, and in transmission electron microscopy studies normal hypertrophic chondrocytes were observed in lesions. Differential expression of ATP6V0D2 and TMSB4 in the cartilage of early naturally occurring osteochondrosis lesions was confirmed by immunohistochemistry. These results identify novel osteochondrosis-associated genes and provide evidence that articular osteochondrosis does not necessarily result from failure of chondrocytes to undergo hypertrophy. Most bones grow through the process of endochondral ossification, in which chondrocytes play a central role. Growth cartilage is comprised of chondrocytes arranged in zones that correspond to the stages of an organized program of sequential biological events. A zone of resting chondrocytes blends into a zone of proliferative chondrocytes and then a zone of hypertrophic chondrocytes, which ultimately undergo physiological death. Within the zone of hypertrophy, the cartilage matrix surrounding individual chondrocytes is partially degraded, leaving behind cartilage remnants that form vertical struts onto which bone matrix is deposited by invading osteoblasts (reviewed by Mackie et al., 2011 1 ). The processes of chondrocyte proliferation, differentiation, and hypertrophy are tightly regulated by a variety of growth factors, hormones, transcription factors, and components of the cartilage matrix. Many of these factors have been characterized, but it is likely that more remain to be identified. The process of physiological death undergone by chondrocytes is less well characterized, and there has been a debate in the literature as to whether it occurs through apoptosis or a non-apoptotic mode of death.2,3 Moreover, the chondrocytes in growth cartilage express factors capable of regulating the behavior of cells in the invading ossification front, including vascular endothelial cells, osteoclasts, and osteoblasts.

Genetic risk factors for osteochondrosis in various horse breeds

“…The majority of the significantly differently expressed genes encode extracellular proteins or proteins with an extracellular matrix turnover function (i ntegrin αV, lumican, osteopontin, IBS, ATP6V0D2, cathepsin K ) while thymosin β4 had a role in the cell signalling pathways and LRP4 ( lowdensity lipoprotein receptor‐related protein ) interacts with the Wnt‐pathway. These results lead to the hypothesis that OC is due to a fundamental defect that induces a cartilage retention in subchondral bone . Austbø et al .…”

“…The majority of the significantly differently expressed genes encode extracellular proteins or proteins with an extracellular matrix turnover function (integrin aV, lumican, osteopontin, IBS, ATP6V0D2, cathepsin K) while thymosin b4 had a role in the cell signalling pathways and LRP4 (lowdensity lipoprotein receptor-related protein) interacts with the Wntpathway. These results lead to the hypothesis that OC is due to a fundamental defect that induces a cartilage retention in subchondral bone [57]. Austbø et al [58] found an altered expression of TLK2 (tousled-like kinase 2) and an unknown gene (CD465746.1) in cartilage samples of hock joints of Standardbred foals, obtained by breeding parents with OC-hock, and compared with OC-non-affected foals.…”

lncRNAs: function and mechanism in cartilage development, degeneration, and regeneration

“…LncRNA growth arrest-specific 5 (GAS5) had been firstly reported as a tumor inhibitor in renal cell carcinoma [91]. GAS5 was also upregulated in osteoarthritis compared to normal tissues [92,93]. Song and his colleagues [27] found that extraneous GAS5 could upregulate the expressions of MMPs such as MMP-2, MMP-3, MMP-9, MMP-13, and ADAMTS-4.…”

“…LncRNA-MSR is a Tymosinβ-4 (TMSB4) pseudogene, which is a subclass of lncRNA. TMSB4 could inhibit the polymerization of actin filament and induce the expression of metalloproteinase in chondrocytes [92]. LncRNA-MSR acts as a ceRNA through competitively binding to miR-152 to upregulate the TMSB4 expression, which leads to the overexpression of metalloproteinase and disorganization of cytoskeleton and degeneration of ECM [31].…”

The vacuolar H+ ATPase V0 subunit D2 is associated with chondrocyte hypertrophy and supports chondrocyte differentiation

“…There was some ATP6V0D2 staining of discrete cytoplasmic structures that were not labelled by either of these markers. WD Repeat and socs box-containing 2 + 1: Genes selected as novel or poorly characterised cartilage genes identified in a previous subtractive hybridisation study (Mirams et al, 2016). 2: 'no' -not differentially expressed between zones of growth cartilage; '+' -more highly expressed in zone of hypertrophic chondrocytes than in other zones; '-' -less highly expressed in zone of hypertrophic chondrocytes than in other zones.…”