Figure 6
(A-H) Immuno-labeling of 5 μm-sections of pellets with anti-collagen types I and II. All photomicrographs represent low (4x objectives) images. Pellets were cultured for three weeks in chondrogenic medium under low (3%) or normal (21% O2) oxygen tension. Images were taken from pellets derived from BM73. Scale bar is 100 μm. (I) Real-time PCR analysis of cDNA derived from pellet cultures of normoxia- and hypoxia-expanded BMSCs. Pellet cultures were performed under normoxic or hypoxic conditions for three weeks in the presence of defined serum-free chondrogenic factors as described in the Materials and Methods text. Real-time PCR analysis was via SYBR Green detection. Presented data represent the mean ± standard error of pellets pooled from four donors (that is, BM68, BM73, BM74 and BM84) in triplicate (N = 4, n = 12 per experimental group). Data represent mean ± standard error of triplicate pellets; one-way ANOVA with Tukey's post hoc test: n.s. (not significant), * = P < 0.05 and *** = P < 0.0001. Gene expression is presented as relative mRNA level normalized to mRNA expression of human β-actin; y-axis. ANOVA, analysis of variance; BMSCs, bone marrow mesenchymal stromal cells; PCR, polymerase chain reaction.