Morphologic Damage of Rat Alveolar Epithelial Type II Cells Induced by Bile Acids Could Be Ameliorated by Farnesoid X Receptor Inhibitor Z-Guggulsterone In Vitro

Abstract

Objective. To determine whether bile acids (BAs) affect respiratory functions through the farnesoid X receptor (FXR) expressed in the lungs and to explore the possible mechanisms of BAs-induced respiratory disorder. Methods. Primary cultured alveolar epithelial type II cells (AECIIs) of rat were treated with different concentrations of chenodeoxycholic acid (CDCA) in the presence or absence of FXR inhibitor Z-guggulsterone (GS). Then, expression of FXR in nuclei of AECIIs was assessed by immunofluorescence microscopy. And ultrastructural changes of the cells were observed under transmission electron microscope and analyzed by Image-Pro Plus software. Results. Morphologic damage of AECIIs was exhibited in high BAs group in vitro, with high-level expression of FXR, while FXR inhibitor GS could attenuate the cytotoxicity of BAs to AECIIs. Conclusions. FXR expression was related to the morphologic damage of AECIIs induced by BAs, thus influencing respiratory functions.

Figure 1

Representative micrographs of FXR immunostaining of CDCA-treated primary cultured AECIIs. Rabbit polyclonal anti-FXR antibody demonstrated the presence of FXR in nuclei of AECIIs. And the fluorescence intensity on AECIIs treated with GS was much lower than that without GS. Magnification bar: 50 μm.

Figure 2

The morphology of primary cultured AECIIs of rat after treatment of CDCA and the FXR blocker GS under transmission electron microscope. In Group 1 (control), Group 5 (40 μM GS, MIX), and Group 9 (40 μM GS, PRE), AECIIs were in good condition. Large numbers of LBs were observed, with dark color and ring-like arrangement (×15000, ×15000, and ×20000, (a), (e), and (i)). In Group 2 (25 μM CDCA), mitochondria had slight swelling, with some LBs demonstrating vacuolization (×15000, (b)). In Group 3 (100 μM CDCA), the number of LBs reduced significantly. Mitochondria swelled seriously and showed balloon-like change (×15000, (c)). In Group 4 (200 μM CDCA), fewer LBs were presented, showing vacuole-like deformity. Microvilli on the surface had disappeared (×15000, (d)). In Groups 6–8 and Groups 10–12 (25, 100, and 200 μM CDCA with 40 μM GS, MIX, and PRE), the number of LBs markedly increased compared to Groups 2–4, with clear lamellar and cellular structure (×20000, ×20000, ×15000, ×20000, ×20000, and ×15000, (f), (g), (h), (j), (k), and (l)). Arrow 1: microvillus; arrow 2: lamellar body; arrow 3: mitochondria.

Figure 3

Effects of different concentrations of CDCA on the sectional area ratio of LBs to cytoplasm of primary cultured AECIIs in the absence or presence of the FXR blocker GS. CDCA was found to dose-dependently reduce the sectional area ratio of LBs to cytoplasm of AECIIs. However, such effects were entirely reversed by FXR inhibitor GS. ^∗ P < 0.01.

Figure 4

Possible mechanisms of BAs-induced respiratory disorder.