title: Gene expression of types I, II, and VI collagen, aggrecan, and chondroitin-6-sulfotransferase in the human annulus: in situ hybridization findings.
abstract: BACKGROUND
  Within each lamellar bundle in the annulus, disc cells produce a complex and sophisticated architectural organization which acts to meet the unique biomechanical needs of the disc. How cells coordinate expression of genes throughout the disc is an important but as yet poorly understood process. For the annulus, such coordination probably involves cell-cell communication as well as growth factor and mechanoreceptor signaling to appropriately maintain the disc extracellular matrix (ECM) for the prevention of annular tears.
  
  
  OBJECTIVE
  To determine the percentage and patterns of gene expression for types I, II, and VI collagen, aggrecan, and chondroitin-6-sulfotransferase in the human annulus.
  
  
  METHODS
  Human annulus specimens were obtained from surgical subjects and a control donor in a study approved by the authors' Human Subjects Institutional Review Board.
  
  
  METHODS
  Four Thompson grade II, three grade III, and four grade IV annulus specimens were evaluated with in situ hybridization to determine gene expression.
  
  
  METHODS
  The percentages of cells in the human annulus expressing type I, II, and VI collagen, aggrecan, and chondroitin-6-sulfotransferase.
  
  
  METHODS
  In situ hybridization, a technique with high temporal and spatial resolution, was used to detect gene expression of types I, II, and VI collagen, aggrecan, and chondroitin-6 sulfotransferase in cells in adjacent sections of annulus from discs with Thompson grades of II, III, and IV.
  
  
  RESULTS
  Overall, 30.8% of cells expressed aggrecan, 38.4% type I collagen, 45.6% type II collagen, 48.1% type VI collagen, and 57.7% chondroitin-6-sulfotransferase. An important finding was that adjacent cells could be expressing, or not expressing, the gene of interest. These data could not have been gained from other global molecular techniques such as microarray analysis or reverse transcription polymerase chain reaction (RT-PCR). Information on gene expression by individual disc cells is important to better understand disc matrix homeostasis, the pathogenesis of disc degeneration, and to formulate potential biologic therapies for disc degeneration.
  
  
  CONCLUSIONS
  This in situ hybridization study revealed the important finding that adjacent cells differ in their gene expression patterns for specific genes. Factors that could contribute to this difference in adjacent cell gene expression include cellular heterogeneity within the annulus, the presence of senescent cells with altered gene expression, and/or loss of coordinated disc cell function as a result of disruption of cell-cell communication.
authors.affiliation: Department of Orthopaedic Surgery, Orthopaedic Research Biology, Cannon Research 304, Carolinas Medical Center, Charlotte, NC 28232, USA. helen.gruber@carolinashealthcare.org
authors.forename: Helen E
authors.initials: HE
authors.lastname: Gruber
authors.forename: Jane
authors.initials: J
authors.lastname: Ingram
authors.forename: Kelly
authors.initials: K
authors.lastname: Leslie
authors.forename: Edward N
authors.initials: EN
authors.lastname: Hanley
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D053674:Aggrecans / Q000096:biosynthesis*; D003094:Collagen / Q000096:biosynthesis* / Q000235:genetics; D024042:Collagen Type I / Q000096:biosynthesis; D024043:Collagen Type II / Q000096:biosynthesis; D024142:Collagen Type VI / Q000096:biosynthesis; D005260:Female; D015870:Gene Expression; D006801:Humans; D017403:In Situ Hybridization; D007403:Intervertebral Disc / Q000166:cytology / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D012333:RNA, Messenger / Q000032:analysis; D015238:Sulfotransferases / Q000096:biosynthesis*
chemical_list: D053674:Aggrecans; D024042:Collagen Type I; D024043:Collagen Type II; D024142:Collagen Type VI; D012333:RNA, Messenger; D003094:Collagen; D015238:Sulfotransferases; C019893:chondroitin 4-sulfotransferase