title: Probing the microenvironmental conditions for induction of superficial zone protein expression.
abstract: OBJECTIVE
  To determine the in vitro conditions which promote expression of superficial zone protein (SZP).
  
  
  METHODS
  Chondrocytes from 6-month-old calves were expanded in monolayer culture and the expression of SZP in alginate bead and monolayer culture was quantified with quantitative real time-polymerase chain reaction (qRT-PCR) and immunostaining. The effect of oxygen tension on SZP expression was determined by qRT-PRC analysis of cells cultured in two dimension (2D) and three dimension (3D) under hypoxic (1% pO2) or normoxic (21% pO2) conditions. Finally, to examine the effect of cyclic tensile strain on expression of SZP in 2D and 3D cultures, chondrocytes encapsulated in alginate beams or seeded on type I collagen coated polydimethylsiloxane (PDMS) chambers were subjected to 5% strain at 1 Hz, 2 h/day for 4 days or 2 h at the fourth day of culture and mRNA levels were quantified.
  
  
  RESULTS
  Bovine chondrocytes in monolayer showed a drastic decrease in SZP expression, similar in trend to the commonly reported downregulation of type II collagen (Col2). Chondrocytes embedded in alginate beads for 4 days re-expressed SZP but not Col2. SZP expression was higher under normoxic conditions whereas Col2 was upregulated only in alginate beads under hypoxic conditions. Cyclic mechanical strain showed a tendency to upregulate mRNA levels of SZP.
  
  
  CONCLUSIONS
  A microenvironment encompassing a soft encapsulation material and 21% oxygen is sufficient for fibroblastic chondrocytes to re-express SZP. These results serve as a guideline for the design of stratified engineered articular cartilage and suggest that microenvironmental cues (oxygen tension level) strongly influence the pattern of SZP expression in vivo.
authors: Cartilage Engineering + Regeneration Laboratory, ETHZ, Schafmattstrasse 22, 8093 Zürich, Switzerland.
authors: R
authors: R
authors: Mhanna
authors: E
authors: E
authors: Öztürk
authors: P
authors: P
authors: Schlink
authors: M
authors: M
authors: Zenobi-Wong
mesh_terms: D000818:Animals; D002358:Cartilage, Articular / Q000166:cytology / Q000378:metabolism*; D002417:Cattle; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002478:Cells, Cultured; D060833:Cellular Microenvironment / Q000235:genetics* / Q000502:physiology; D019902:Chondrocytes / Q000378:metabolism*; D024043:Collagen Type II / Q000235:genetics* / Q000378:metabolism; D000860:Hypoxia / Q000235:genetics* / Q000378:metabolism; D007150:Immunohistochemistry; D011509:Proteoglycans / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000032:analysis*; D060888:Real-Time Polymerase Chain Reaction; D013314:Stress, Mechanical; D023822:Tissue Engineering / Q000379:methods; D000099158:Proteoglycan 4
chemical_list: D024043:Collagen Type II; D011509:Proteoglycans; D012333:RNA, Messenger; C498836:PRG4 protein, Bos taurus; D000099158:Proteoglycan 4
keywords: Chondrocytes dedifferentiation; Mechanical strain; Microenvironment; Oxygen tension; Superficial zone protein