Validation of genome-wide data.
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Three genes were selected: Programmed Cell Death 8 (PDCD8) which demonstrated more ribosome loading in IPF; Cofilin 2 (CFL2) which showed more ribosome loading in controls; and Fucosyltransferase 10 (FUT10) which displayed no change in ribosome loading between the two cell types. A–C. Polyribosome shift using qRT-PCR. Shown is the quantity of mRNA normalized to a spike in standard, as a function of position in the sucrose gradient. D. Total RNA levels. The total RNA was normalized to a “spike in” standard and actin. E–F. Steady state protein levels of PDCD8, CFL2 and FUT10. Cells (passage 5) were seeded on type I collagen matrices and examined for protein expression using actin as a loading control E. Western blotting from the same cell lines as in (A–D). F. Steady state levels of 6 additional primary myofibroblast lines (3 IPF; and 3 control).