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. 2015 Jan;185(1):55–68. doi: 10.1016/j.ajpath.2014.08.027

Figure 3.

Figure 3

Role of urokinase-type plasminogen activator (uPA) in ATII cell epithelial–mesenchymal transitions after bleomycin (BLM) injury. A: ATII cells isolated from mice deficient in uPA expression were treated with PBS, BLM, BLM+CSP, or BLM+CP in vitro. The lysates were tested for the expression of collagen-I, α-SMA, E-cadherin, ZO-1, and β-actin antigens by Western blot analysis. The lysates from uPA-deficient ATII cells treated with TGF-β were used as a control for comparison. The individual bands were quantitated and normalized to the corresponding values of β-actin loading controls and the fold changes in proteins are presented as a bar graph. Differences between treatments versus control are statistically significant. B: Mice deficient in uPA expression were exposed to saline or BLM as described in Figure 2D. Twenty-four hours after initial exposure to BLM, mice were i.p. injected with or without caveolin-1 scaffolding domain peptide (CSP) or control peptide (CP). Seventy-two hours after initiation of BLM injury, ATII cells were isolated and the lysates were evaluated for changes in the expression of collagen-I, α-SMA, E-cadherin, ZO-1, and β-actin by Western blot analysis. The densities of individual bands were normalized against the corresponding densities of β-actin protein, and the fold changes in proteins are presented as a bar graph. C: Total RNA isolated from ATII cells of uPA-deficient mice exposed to saline, BLM, BLM+CSP, or CP were analyzed for changes in the expression of E-cadherin, collagen-I, ZO-1, and α-SMA mRNA levels by real-time PCR. The data were normalized with corresponding levels of β-actin mRNA. The fold changes of mRNA are presented as a bar graph. D: The lung homogenates of WT and uPA-deficient mice exposed to saline, BLM, BLM+CSP, or BLM+CP were tested for active TGF-β by Western blot analysis. E: The bronchoalveolar lavage fluids (BALF) of WT and uPA-deficient mice exposed to saline, BLM, BLM+CSP, or BLM+CP were analyzed for active TGF-β byELISA. ∗∗∗∗P < 0.0001 versus control (A, D, E, and F).