Expression profiles of genes involved in apoptosis and selenium metabolism in articular cartilage of patients with Kashin–Beck osteoarthritis

Kashin–Beck disease (KBD) is a special type of endemic osteoarthritis. It has been suggested that alterations in selenium metabolism and apoptosis play a role in KBD. However, the underlying molecular mechanism remains largely unclear. We performed a microarray analysis using RNA isolated from cartilages of KBD patients and healthy controls, through Significance Analysis of Microarray (SAM) software. Functional gene networks and crucial molecules associated with differentially expressed genes were investigated via Ingenuity Pathway Analysis (IPA) and hub gene analysis. Quantitative real-time PCR was used to check the validation of chip test. We identified 52 up-regulated apoptosis-related genes and 26 down-regulated selenium-related genes between KBD and controls, and these genes associated with the “MYC-mediated apoptosis signaling pathway”. We confirmed the results from array studies with quantitative real-time PCR analysis. Our results suggest that abnormal regulation of selenium metabolism and apoptosis through the MYC mediated signaling pathway contributes to the pathogenesis of KBD, but the relationship between apoptosis gene and selenium gene was not found.     

Pathways related to mitochondrial dysfunction in cartilage of endemic osteoarthritis patients in China

In this paper, we present the first evidence of differences in the mitochondria-related gene expression profiles of adult articular cartilage derived from patients with Kashin-Beck disease and normal controls. The expression of 705 mitochondria-related genes was analyzed by mitochondria-related gene expression analysis and ingenuity pathways analysis. Mitochondria-related gene expression analysis identified 9 up-regulated genes in Kashin-Beck disease based on their average expression ratio. Three canonical pathways involved in oxidative phosphorylation, apoptosis signaling and pyruvate metabolism were identified, which indicate the involvement of mitochondrial dysfunction in the pathogenesis of Kashin-Beck disease.     

Trans-omics pathway analysis suggests that eQTLs contribute to chondrocyte apoptosis of Kashin–Beck disease through regulating apoptosis pathway expression

Kashin–Beck disease (KBD) is a serious osteoarthropathia, mainly characterized by excessive chondrocyte necrosis and apoptosis. The molecular signaling pathways underlying KBD excessive chondrocyte apoptosis remain unclear, leading to a lack of effective medical interventions now. To clarify whether expression quantitative trait loci (eQTLs) contribute to excessive chondrocyte apoptosis of Kashin–Beck disease through regulating the expression of apoptosis pathways. We conducted a genome-wide eQTLs based pathway association analysis of KBD using Affymetrix Human SNP Array 6.0 in 1717 Chinese Han subjects. PLINK software was used for genome-wide association study (GWAS) of KBD. A modified gene set enrichment algorithm was applied for pathway association analysis based on GWAS results. The KBD-associated pathways were compared with abnormally expressed pathways in KBD articular cartilage, identified by microarray study of KBD. We identified 4 eQTLs pathways, which were not only significantly associated with KBD, but also abnormally expressed in KBD articular cartilage, including REACTOME_INTRINSIC_PATHWAY_FOR_APOPTOSIS (P = 0.008), MAHAJAN _RESPONSE_TO_IL1A_UP (P = 0.010), KEGG_PEROXISOME (P = 0.005) and MARKS_HDAC_TARGETS_UP (P = 0.006). Our results suggest that eQTLs contributed to KBD excessive chondrocyte apoptosis through regulating the expression of apoptosis related pathways. This study provides novel insight into the genetic susceptibility and therapeutic rationale of KBD.     

Proteomic changes in articular cartilage of human endemic osteoarthritis in China

Kashin-Beck disease (KBD) is a chronic endemic osteochondropathy with unclear pathogenesis. It is a degenerative disease similar to osteoarthritis, but with different manifestations of cartilage damage. The aim of this investigation was to show the protein changes in KBD cartilage and to identify the candidate proteins in order to understand the pathogenesis of the disease. Proteins were extracted from the media of primary cell cultures of KBD and normal chondrocytes, and separated by two-dimensional fluorescence difference gel electrophoresis (2-D DIGE). MALDI-TOF/TOF analysis revealed statistically significant differences in 27 proteins from KBD chondrocyte cultures, which consisted of 17 up-regulated and ten down-regulated proteins. The results were further validated by Western blot analysis. The proteins identified are mainly involved in cellular redox homeostasis and stress response (MnSOD, Hsp27, Peroxiredoxin-1, and Cofilin-1), glycolysis (PGK-1, PGM-1, α-enolase), and cell motility and cytoskeletal organization (Actin, Calponin-2, and Keratin). These KBD-associated proteins indicate that cytoskeletal remodeling, glycometabolism, and oxidative stress are abnormal in KBD articular cartilage.     

Invited review: the mitochondrion in osteoarthritis

In a variety of tissues, cumulative oxidative stress, disrupted mitochondrial respiration, and mitochondrial damage promote aging, cell death, and ultimately, functional failure and degeneration. Because articular cartilage chondroyctes are highly glycolytic, mitochondrially mediated pathogenesis has not been previously applied in models for pathogenesis of osteoarthritis (OA), a cartilage degenerative disease that increases markedly in aging. However, chondrocyte mitochondria respire in vitro and they demonstrate swelling and changes in number in situ in the course of OA. Normal chondrocyte mitochondrial function is hypothesized to critically support adenosine triphosphate (ATP) reserves in functional stressed chondrocytes during OA evolution. In this model, disruption of chondrocyte respiration by nitric oxide, a mediator markedly up-regulated in OA cartilage, is centrally involved in chondrocyte functional compromise. Furthermore, mitochondrial dysfunction can mediate several specific pathogenic pathways implicated in OA. These include oxidative stress, inadequacy of chondrocyte biosynthetic and growth responses, up-regulated chondrocyte cytokine-induced inflammation and matrix catabolism, increased chondrocyte apoptosis, and pathologic cartilage matrix calcification. In addition, the direct, sublethal impairment of chondrocyte mitochondrial ATP synthesis in vitro decreases matrix synthesis and increases matrix calcification ('disease in a dish'). The weight of evidence reviewed herein strongly supports chondrocyte mitochondrial impairment as a mediator of the establishment and progression of OA.     

Gene set enrichment analysis for non-monotone association and multiple experimental categories

Background  

Recently, microarray data analyses using functional pathway information, e.g., gene set enrichment analysis (GSEA) and significance analysis of function and expression (SAFE), have gained recognition as a way to identify biological pathways/processes associated with a phenotypic endpoint. In these analyses, a local statistic is used to assess the association between the expression level of a gene and the value of a phenotypic endpoint. Then these gene-specific local statistics are combined to evaluate association for pre-selected sets of genes. Commonly used local statistics include t-statistics for binary phenotypes and correlation coefficients that assume a linear or monotone relationship between a continuous phenotype and gene expression level. Methods applicable to continuous non-monotone relationships are needed. Furthermore, for multiple experimental categories, methods that combine multiple GSEA/SAFE analyses are needed.     

The effect of cyclical compressive loading on gene expression in articular cartilage

Osteoarthritis (OA) develops as a consequence of articular cartilage degeneration possibly initiated by excessive or abnormal loading of the joint, and potentially mediated through a proteinase/proteinase inhibitor imbalance. We have shown previously that physiological loads (0.5 MPa, 1 Hz, 3 hour) elicit increased expression and activation of the matrix metalloproteinases (MMPs) in articular cartilage explants in vitro. The objective of this study was to identify mechanically-regulated genes involved in the observed induction of MMP expression and enhanced activation. Differential RNA Display (DRD) was used to identify mechanically-regulated genes by comparing DRD products derived from loaded and unloaded cartilage. One gene up-regulated in cartilage after 10, 30 and 60 minute loading revealed 83% homology with Mus musculus thymosin beta_4 which is known to induce MMP gene expression. The identification of mechanically regulated genes will greatly enhance our understanding of matrix turnover providing an exciting future in elucidating the role of mechanically-regulated genes in the development of OA.     

PAGE: Parametric Analysis of Gene Set Enrichment

Background  

Gene set enrichment analysis (GSEA) is a microarray data analysis method that uses predefined gene sets and ranks of genes to identify significant biological changes in microarray data sets. GSEA is especially useful when gene expression changes in a given microarray data set is minimal or moderate.     

Exome sequencing identified FGF12 as a novel candidate gene for Kashin-Beck disease

The objective of this study was to identify novel causal genes involved in the pathogenesis of Kashin-Beck disease (KBD). A representative grade III KBD sib pair with serious skeletal growth and development failure was subjected to exome sequencing using the Illumina Hiseq2000 platform. The detected gene mutations were then filtered against the data of 1000 Genome Project, dbSNP database, and BGI inhouse database, and replicated by a genome-wide association study (GWAS) of KBD. Ninety grade II or III KBD patients with extreme KBD phenotypes and 1627 healthy controls were enrolled in the GWAS. Affymetrix Genome-Wide Human SNP Array 6.0 was applied for genotyping. PLINK software was used for association analysis. We identified a novel 106T>C at the 3′UTR of the FGF12 gene, which has not been reported by now. Sequence alignment observed high conversation at the mutated 3′UTR+106T>C locus across various vertebrates. In the GWAS of KBD, we detected nine SNPs of the FGF12 gene showing association evidence (P value?<?0.05) with KBD. The most significant association signal was observed at rs1847340 (P value?=?1.90?×?10^?5). This study suggests that FGF12 was a susceptibility gene of KBD. Our results provide novel clues for revealing the pathogenesis of KBD and the biological function of FGF12.     

6天龄与10天龄大鼠的睾丸组织的基因差异表达

通过基因芯片技术,利用Roche-NimbleGen公司制作的大鼠12×135K全基因组表达谱芯片,对日龄为6d和10d的大鼠睾丸组织进行全基因组表达差异分析。结果显示:具有2倍以上的差异表达基因有4298个,其中表达上调的基因共1878个,表达下调的基因共2420个。这些差异表达的基因中有3154个基因具有基因本体注释,参与了154个生物学通路。进一步分析表明具有8倍以上差异表达的基因有13个,这些基因参与了生物学过程、细胞组分和分子功能等基因本体分类,进一步选择3个差异表达的基因,LOC686076、Cxcl6和Trib3,做了实时定量RT-PCR检测。其结果趋势与芯片数据一致。因此,我们初步认为精原干细胞的发生与增殖在大鼠早期的发育过程中已经有大量的基因参与,是一个多基因协调表达的过程。