Figure 3. Validation of differential gene expression profiles determined by microarray analysis of aRNA derived from Schmid and Cog mouse growth plate hypertrophic zones.

(A) qPCR performed in triplicate on the aRNA samples described in Figure 2, using selected markers of the unfolded protein response (UPR) and endoplasmic reticulum associated degradation pathway, including (i) BiP, (ii) Calr, (iii) Derl2, (iv) Derl3, (v) Edem1, (vi) ERdj4, (vii) Erp72, (viii) Fgf21, and (ix) Luman. Expression profiles are expressed as fold difference for homozygous Schmid (Schmid) or Tg^cog (Cog) compared with wildtype (Wt), with profiles determined by the microarray analyses described in Figure 2 shaded dark grey, and profiles determined by qPCR shaded light grey. Error bars indicate standard deviation around the mean. (B) In situ analyses performed on 7 day old Wt and Schmid tibial growth plate cryosections using digoxigenin-labelled riboprobes specific for (i,ii) Col10a1 as well as novel markers of the UPR including (iii,iv) Armet, (v,vi) Creld2, (vii,viii) Fgf21, (ix,x) Luman, (xi,xii) Steap1, (xiii,xiv) Syvn1, and (xv,xvi) Wfs1. Dashed lines demarcate approximate growth plate zone boundaries: R – Resting Zone, P – Proliferative Zone, H – Hypertrophic Zone. Boxes inset show magnified representative areas of the hypertrophic zones, to highlight the extent of riboprobe hybridization in these zones. Scale bars = 500 µm.