Effects of Endoplasmic Reticulum (ER) Stress on Epithelial Injury and Fibrosis in Alveolar Epithelial Type II Cell (AT2)‐Specific Grp78 Knockout Mice

Abstract

To study the causal relationship between unfolded protein response (UPR) activation/ER stress and pulmonary fibrosis, we investigated epithelial injury and fibrosis in Grp78‐CKO1 mice (generated by crossing SFTPC‐rtTA, tetO‐cre and Grp78flox/flox mice) with AT2‐specific knockout (KO) of Grp78. Doxycycline (Dox)‐induced KO of the Grp78 allele resulted in decreased Grp78 mRNA by ~50% in AT2 cells and increased mRNA expression of ER stress markers (i.e., Grp94 (~16‐fold), Atf‐6 (~40‐fold), calreticulin (Crt, ~18‐fold) and CHOP (~2.7‐fold) in whole lung. Despite increased vimentin and collagen expression, Grp78‐CKO1 lungs did not display overt fibrosis, although Sircol assay demonstrated augmentation (~30%) of bleomycin‐induced collagen deposition. Since Grp78 was reduced only 50% in Grp78‐CKO1 mice, Grp78‐CKO2 mice were generated by crossing SftpccreERT2/creERT2 and Grp78flox/flox mice. Grp78 expression in AT2 cells was reduced by >80% in tamoxifen treated mice, with increased mRNA expression of Grp94 (~8‐fold), Atf‐6 (~4‐fold), Crt (~5‐fold) and especially high induction of CHOP (~16‐fold). Hematoxylin/eosin and trichrome staining demonstrated overt fibrosis in lungs of Grp78‐CKO2 mice. These results indicate that 1) ER stress‐mediated epithelial cell injury plays an important role in fibrogenesis and 2) development of fibrosis may be dependent on the degree of ER stress and activation of pro‐apoptotic UPR signaling.