Alveolar Epithelial Cells Undergo Epithelial-to-Mesenchymal Transition in Response to Endoplasmic Reticulum Stress

Abstract

Expression of mutant surfactant protein C (SFTPC) results in endoplasmic reticulum (ER) stress in type II alveolar epithelial cells (AECs). AECs have been implicated as a source of lung fibroblasts via epithelial-to-mesenchymal transition (EMT); therefore, we investigated whether ER stress contributes to EMT as a possible mechanism for fibrotic remodeling. ER stress was induced by tunicamyin administration or stable expression of mutant (L188Q) SFTPC in type II AEC lines. Both tunicamycin treatment and mutant SFTPC expression induced ER stress and the unfolded protein response. With tunicamycin or mutant SFTPC expression, phase contrast imaging revealed a change to a fibroblast-like appearance. During ER stress, expression of epithelial markers E-cadherin and Zonula occludens-1 decreased while expression of mesenchymal markers S100A4 and ␣-smooth muscle actin increased. Following induction of ER stress, we found activation of a number of pathways, including MAPK, Smad, ␤-catenin, and Src kinase. Using specific inhibitors, the combination of a Smad2/3 inhibitor (SB431542) and a Src kinase inhibitor (PP2) blocked EMT with maintenance of epithelial appearance and epithelial marker expression. Similar results were noted with siRNA targeting Smad2 and Src kinase. Together, these studies reveal that induction of ER stress leads to EMT in lung epithelial cells, suggesting possible crosstalk between Smad and Src kinase pathways. Dissecting pathways involved in ER stress-induced EMT may lead to new treatment strategies to limit fibrosis. Idiopathic pulmonary fibrosis (IPF)3 is a fatal lung disease characterized by extensive architectural distortion and progressive parenchymal fibrosis. A substantial body of work supports the concept that alveolar epithelial cells (AECs) play an essential role in the development of IPF, but the specific mechanisms linking AECs to lung fibrosis are not well understood (1). Genetic studies have indicated that mutations in the gene encoding surfactant protein C (SFTPC) can lead to familial interstitial pneumonia, which is the familial form of IPF (2, 3). Mutations in the carboxyl terminal region of the gene result in a product that cannot be processed normally in type II AECs, leading to accumulation of misfolded prosurfactant protein C in the endoplasmic reticulum (ER), ER stress, and activation of the unfolded protein response (UPR) (4). In addition, our group and others (5, 6) have reported that ER stress and UPR activation are found in the alveolar epithelium in lung biopsies from patients with familial interstitial pneumonia in the absence of SFTPC mutation as well as sporadic IPF. Taken together, available data indicate that ER stress may be important in the pathogenesis of IPF; however, the means by which ER stress contributes to lung fibrosis is not currently identified.With ER stress, UPR pathways are activated as a means to help the cell abrogate the untoward effects of protein accumulation in the ER. The three arms of the UPR include processes designed to attenuate protein transla...

Endoplasmic Reticulum Stress Induces Epithelial-Mesenchymal Transition through Autophagy via Activation of c-Src Kinase

“…We found that both TM and TG induced c-Src phosphorylation and PP2, a c-Src kinase inhibitor attenuated effects of TM- or TG-induced EMT, suggesting that activated c-Src kinase mediated ER stress-induced EMT in tubular epithelial cells. These results were compatible with previous reports in alveolar epithelial cells and thyroid cells [7,8]. We also found that c-Src kinase was involved in ER stress-induced autophagy, which was compatible with a previous report that Src was involved in zVAD-induced autophagic cell death in murine fibrosarcoma cells [26].…”

“…Recently, ER stress and UPR pathways are emerging as important determinants of fibrotic remodeling in a number of internal organs [2,3,4,5] through induction of EMT [6,7,8,9]. In kidney, it has been reported that ER stress by cyclosporine and TG in tubular epithelial cells induces EMT [21].…”

“…However, the mechanisms linking ER stress to EMT are incompletely understood. Tanjore et al [7] reported that siRNA targeting of IRE-1 was able to block TM-induced EMT in alveolar epithelial cells by attenuation of TM-induced Smad2 and Src phosphorylation, which were involved in ER stress-induced EMT. They suggested that the IRE-1 arm of the UPR appeared to be involved in activating these signaling pathways.…”

“…Src activation in ER stress-induced EMT was reported in non-renal cells [7,8]. However, it is unclear whether c-Src activation mediates ER stress-induced EMT in renal tubular epithelial cells.…”

“…UPR may either promote cell survival, or if the ER stress is chronic or excessive may lead to apoptosis by inducing proapoptotic transcription factor CHOP [1]. In addition to effects on apoptosis, ER stress and UPR pathways have been suggested as important determinants of fibrotic remodeling in a number of internal organs, including the lungs, liver, gastrointestinal tract, heart and kidney [2,3,4,5] through induction of epithelial-mesenchymal transition (EMT) [6,7,8,9]. …”