Anabolic response of HAC to different oxygen percentages during the expansion and differentiation Phase I. (A - B) Safranin O and type II collagen immunohistochemical stainings of representative tissues generated by human articular chondrocytes (HAC) expanded at 19% (I and III) or 5% (II and IV) oxygen and further cultured in pellets at 19% (I and II) or 5% (III and IV) oxygen. Bar = 100 μm. (C) Quantification of glycosaminoglycans (GAG) accumulated normalized to the amount of DNA. (D - E) Real time reverse transcription-polymerase chain reaction analysis of the expression of type II collagen and aggrecan mRNA by HAC cultured in pellets at 19% and 5% O2. Levels are expressed as fold of difference from ribosomal 18S. For the gene expression analysis only expansion at 19% O2 was considered. Values are mean ± SD of measurements obtained from three independent experiments. * = significantly different from the 19% O2.