Fig 2: Compression-induced TNC secretion depends on TGF-β receptor and ERK pathways. TNC secreted by HBE cells at 24 h post-compression, in the presence or absence of U0126 or SB431542. Representative Western blot of three independent experiments shows the detection of basolateral secretion of TNC. Transferrin was detected as a loading control (A). Secreted TNC were detected by ELISA in the basolateral conditioned media (B) and in the apical washes (C) (mean ± SEM, 3 non-asthma donors). **** p < 0.0001, significantly different from vehicle control; $$$$ p < 0.0001, significantly different from vehicle with pressure, analyzed by two-way ANOVA with Bonferroni’s post-hoc test. Each symbol represents each of the three donors. The open symbols represent control condition, and the closed symbols represent compressed condition in non-asthmatic HBE cells (blue symbols; B,C).

Fig 3: Mechanical compression induces mRNA expression and secretion of TNC from HBE cells. TNC mRNA expression was measured at 3 h and 24 h post-compression of non-asthmatic cells (mean ± SEM, 4 donors) (A) and asthmatic cells (mean ± SEM, 4 donors) (B). TNC mRNA expression was normalized to GAPDH. The amount/well of TNC secreted in the basolateral conditioned media was measured by ELISA (mean ± SEM, 4 non-asthma and 4 asthma donors) (C). ** p < 0.01, *** p < 0.001, **** p < 0.0001, significantly different from time-matched no pressure control, # p < 0.05 significantly different between non-asthma and asthma, analyzed by two-way ANOVA with Bonferroni’s post-hoc test. Each symbol represents each of the four donors. The open symbols represent control condition, and the closed symbols represent compressed condition in both non-asthmatic (blue symbols; A,C) and asthmatic HBE cells (red symbols; B,C).

Fig 4: Mechanical compression induces the release of EVs containing TNC from HBE cells. Representative Western blots of four independent experiments show that CD9, tissue factor, YKL-40, and TNC were detected in the whole conditioned media with a greater abundance in conditioned media from compressed cells. YKL-40 was detected in the non-EV fraction, while CD9 and tissue factor were detected in the EV fraction. TNC was detected in both the non-EV and the EV fractions.

Fig 5: Compression-induced TNC mRNA expression depends on TGF-β receptor and ERK pathways. Expression plots showing normalized gene expression levels of TNC at 3 h (A) and 24 h (B) in compressed HBE cells in the absence or presence of either U0126 or SB431542 (mean ± SD, 6 non-asthma donors). TNC mRNA expression was validated by qRT-PCR and normalized to GAPDH (mean ± SEM, 3 non-asthma donors) (C). * p < 0.05, **** p < 0.0001, significantly different from time-matched no pressure control, $$ p < 0.01, $$$$ p < 0.0001, significantly different from vehicle with pressure, analyzed by two-way ANOVA with Bonferroni’s post-hoc test. Each symbol represents each of the six (A,B) or four (C) donors. The open symbols represent control condition, and the closed symbols represent compressed condition in non-asthmatic HBE cells (blue symbols; A–C).