Journal: American Journal of Respiratory and Critical Care Medicine

Article Title: Distinct Cancer-Promoting Stromal Gene Expression Depending on Lung Function

doi: 10.1164/rccm.201801-0080OC

Figure Lengend Snippet: Cancer-associated lung stromal proteomes depend on FEV1pp. (A) Non–cancer-containing stromal samples collected from donors with or without lung cancer across a range of lung-function values were subjected to quantitative proteomics and polysome profiling. This allowed for characterization of gene expression at multiple levels (total mRNA, translational efficiency [after adjusting changes in polysome-associated mRNA levels for changes in total mRNA], and protein). (B) Histograms of P values for changes in protein levels in an ANCOVA in which protein levels are related to age (not shown), cancer (not shown), FEV1pp, and the CFI (n = 58). The dotted line indicates the frequency of these P values when the same analysis was performed on a randomly sampled data set. (C and D) Quantified Western blots with antibodies that detect CAV1 (C) and SFXN3 (D) normalized to vinculin for a subset of stromal samples with and without cancer across a range of FEV1pp values (n = 15). Shown are regression lines with individual samples indicated by dots. Dotted lines indicate 95% confidence intervals for regressions of protein expression in subjects with cancer (red) and subjects without cancer (black) versus FEV1pp. The P value for the CFI is also indicated together with the FEV1pp intersection (blue tick mark along the x axis). ANCOVA = analysis of covariance; CFI = cancer status–FEV1pp interaction; COPD = chronic obstructive pulmonary disease; FEV1pp = FEV1 percent of the predicted value; LC MS/MS = liquid chromatography with tandem mass spectrometry.

Article Snippet: Vinculin (ab18058; Abcam) was used as a loading control along with full-length protein controls for CAV1 and SFXN3 (ag8049 and ag7310, respectively; Proteintech).

Techniques: Quantitative Proteomics, Gene Expression, Western Blot, Expressing, Liquid Chromatography with Mass Spectroscopy, Liquid Chromatography, Mass Spectrometry

Journal: American Journal of Respiratory and Critical Care Medicine

Article Title: Distinct Cancer-Promoting Stromal Gene Expression Depending on Lung Function

doi: 10.1164/rccm.201801-0080OC

Figure Lengend Snippet: Distinct cancer-associated activation of gene expression programs downstream of mTOR signaling and fibrotic ECM depending on FEV1pp. (A) Heatmaps of estimated fold changes (subjects with cancer vs. subjects without cancer, using slopes and intersections from Figure 3) associated with poor (FEV1pp 30) or relatively normal (FEV1pp = 90) lung function. The analysis was performed separately on translational efficiency and protein levels. (B) Cumulative distributions of coefficients for the CFI as quantified from translational efficiency data (n = 32). Subsets of mRNAs previously identified as showing mTOR-sensitive translation or fibrotic ECM-induced translation are compared with background coefficients (i.e., all detected mRNAs not in a subset). A Wilcoxon P value for a difference between the subset and the background is indicated. A shift toward positive interaction coefficients indicates more efficient translation in adjacent stroma of patients with cancer and a high FEV1pp (i.e., similar to CAV1 in Figure 1C), whereas a shift toward negative interaction coefficients indicates more efficient translation in patients with cancer and a low FEV1pp (i.e., similar to SFXN3 in Figure 1D). CFI = cancer status–FEV1pp interaction; ECM = extracellular matrix; FEV1pp = FEV1 percent of the predicted value; IPF = idiopathic pulmonary fibrosis.

Article Snippet: Vinculin (ab18058; Abcam) was used as a loading control along with full-length protein controls for CAV1 and SFXN3 (ag8049 and ag7310, respectively; Proteintech).

Techniques: Activation Assay, Gene Expression