The CPs from bIPE and bRPE cells cultured on a plastic substratum (P) and on mitomycin-treated 3T3 fibroblasts (3T3) are summarized in
Table 4. For both bIPE and bRPE cells cultured on plastic or on 3T3 feeder layers
PEDF and
ZO1 belong to the group of high-copy genes (CP = 20–30), showing the lowest CP values, corresponding to the highest gene expression levels.
KRT8 and
CD86 belong to the group of low copy genes (CP = 30–40), whereas
BEST,
KRT18, and
RPE65 belong to the group of absolute low-copy genes (CP = 40–50). For these genes no differences in CPs were observed between cells cultured on plastic or on 3T3 feeder layers. Both on plastic and on mitomycin-treated 3T3 fibroblasts, the CPs for bIPE and bRPE cells were similar for all analyzed genes, except for
CRALBP and
CD86. CRALBP, a low-copy gene in cultured bIPE cells (
P = 36.9 ± 1.4; 3T3 = 39.3 ± 1.5), is expressed at higher levels than in bRPE cells (
P = 44.0 ± 2.0; 3T3 = 44.5 ± 3.0), whereas
CD86 is expressed at higher levels in bRPE cells (
P = 32.5 ± 4.4; 3T3 = 31.3 ± 1.6) than in bIPE cells (
P = 40.0 ± 2.3; 3T3 = 40.4 ± 3.0).
To analyze, whether cultivation on mitomycin-treated 3T3 fibroblasts has an effect on gene expression levels, we calculated the 2
−ΔΔCP values (
Table 4,
Fig. 5). Based on the 2
−ΔΔCP values, gene expression of
PEDF was upregulated in the bIPE and bRPE cells cultured on mitomycin-treated 3T3 fibroblasts. Statistical analysis of the corresponding ΔCT data revealed that the difference approached significance at the 5% level (bRPE:
F = 12.86,
P = 0.0697; bIPE:
F = 17.05,
P = 0.0540). Gene expression of
ZO1 was also upregulated and in bRPE cells the upregulation was significant at the 5% level (bRPE:
F = 22.24,
P = 0.0421). In bIPE cells gene expression of
RPE65 was downregulated (
F = 11.89,
P = 0.0748) when the cells were cultured on mitomycin-treated 3T3 fibroblasts.