title: Down-regulated HS6ST2 in osteoarthritis and Kashin-Beck disease inhibits cell viability and influences expression of the genes relevant to aggrecan metabolism of human chondrocytes.
abstract: OBJECTIVE
  Primary OA and Kashin-Beck disease (KBD) show similar pathological changes in articular cartilage. The objective was to screen differentially expressed genes between OA and normal cartilage, confirm the candidate gene expression among OA, KBD and normal cartilage, and then clarify its role in vitro.
  
  
  METHODS
  Differentially expressed genes in OA cartilage were screened by suppression subtractive hybridization (SSH) and verified by real-time quantitative PCR (Q-PCR) analysis. Heparan sulphate 6-O-sulphotransferase 2 (HS6ST2) expression was identified by Q-PCR and immunohistochemistry. After suppressing HS6ST2 by RNA interference in C28/I2 human chondrocyte line, the effects were analysed through determining the cell viability by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), the aggrecan contents by toluidine blue staining and the mRNA expression levels of SRY-type high mobility group box 9 (SOX9), AGC1, MMP3, a disintegrin and metalloproteinase domain with thrombospondin motifs 4 (ADAMTS4) and ADAMTS5 by Q-PCR.
  
  
  RESULTS
  HS6ST2 in the reverse subtraction library was identified as a down-regulated gene in OA and KBD at both mRNA and protein levels. The percentage of safranion O staining area was correlated positively with the percentage of HS6ST2-positive chondrocytes in OA and KBD cartilage. After HS6ST2-specific short interfering RNA (siRNA) transfection to C28/I2 cells, the cell viability was inhibited significantly, and the mRNA expression levels of SOX9 and AGC1 were reduced markedly, while MMP3 expression was increased significantly. CONCLUSION; HS6ST2 down-regulation was identified in both OA and KBD cartilage. The findings first suggest that HS6ST2 may participate in the pathogenesis of OA and KBD by influencing aggrecan metabolism.
authors.affiliation: Department of Genetics and Molecular Biology, Xi'an Jiaotong University College of Medicine, Xi'an, Shaanxi, People's Republic of China.
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.forename: Bo
authors.initials: B
authors.lastname: Zhong
authors.forename: Jian
authors.initials: J
authors.lastname: Sun
authors.forename: Junling
authors.initials: J
authors.lastname: Cao
authors.forename: Juan
authors.initials: J
authors.lastname: Tian
authors.forename: Nannan
authors.initials: N
authors.lastname: Zhong
authors.forename: Wenxiang
authors.initials: W
authors.lastname: Zhao
authors.forename: Lifang
authors.initials: L
authors.lastname: Tian
authors.forename: Peng
authors.initials: P
authors.lastname: Xu
authors.forename: Dawei
authors.initials: D
authors.lastname: Guo
authors.forename: Xichi
authors.initials: X
authors.lastname: Ju
authors.forename: Wei
authors.initials: W
authors.lastname: Ma
authors.forename: Meng
authors.initials: M
authors.lastname: Li
authors.forename: Weikun
authors.initials: W
authors.lastname: Hou
authors.forename: Shemin
authors.initials: S
authors.lastname: Lu
mesh_terms: D051722:ADAM Proteins / Q000378:metabolism; D000328:Adult; D000368:Aged; D053674:Aggrecans / Q000378:metabolism*; D000704:Analysis of Variance; D002358:Cartilage, Articular / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000502:physiology; D019902:Chondrocytes / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D015536:Down-Regulation; D005260:Female; D015870:Gene Expression; D006801:Humans; D057767:Kashin-Beck Disease / Q000235:genetics*; D008297:Male; D019278:Matrix Metalloproteinase 3 / Q000378:metabolism; D008875:Middle Aged; D010003:Osteoarthritis / Q000235:genetics*; D010641:Phenotype; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D055758:SOX9 Transcription Factor / Q000378:metabolism; D015238:Sulfotransferases / Q000235:genetics*; D014162:Transfection
chemical_list: C508319:ACAN protein, human; D053674:Aggrecans; D018076:DNA, Complementary; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D055758:SOX9 Transcription Factor; C527932:SOX9 protein, human; C485757:HS6ST2 protein, human; D015238:Sulfotransferases; D051722:ADAM Proteins; D019278:Matrix Metalloproteinase 3