Novel alveolar epithelial cell differentiation markers in lung injury and repair

Objective: The alveolar epithelium, consisting of mainly alveolar epithelial type 1 (AT1) and type 2 (AT2) cells, represents a major site of tissue destruction in idiopathic pulmonary fibrosis. Several studies indicate that adult AT2 cells are able to self-renew and exert progenitor function for AT1 cells upon alveolar injury in vivo. However, cell differentiation pathways enabling this plasticity are poorly understood. Here, we used the primary culture of murine AT2 cells as model system to identify novel proteins and pathways involved in epithelial transdifferentiation.

Methods/Results: Expression profiles of primary transdifferentiating AT2 cells were analyzed applying 2D gel electrophoresis and mass spectrometry. Beside others, we found enolase 1 (ENO1) to be upregulated, whereas carbonyl reductase 2 (CBR2) was decreased in transdifferentiating AT2 cells, as further confirmed by quantitative RT-PCR analysis and immunoblotting. This was accompanied by reduction in AT2 cell derived pro surfactant protein C (proSPC) expression and increased AT1 cell T1a expression, as well as an activation of the Wnt/β-catenin pathway. We applied a lung tissue culture model of murine precision cut lung slices ex vivo to further analyze transdifferentiation in the 3D natural spatial lung environment. We observed Wnt/β-catenin signal activation and alveolar epithelial cell transdifferentiation upon lung tissue cultures ex vivo. Interestingly, the inhibition of Wnt/β-catenin signaling in cultured AT2 resulted in decreased expression of ENO1 and T1a and stabilization of CBR2. In an in vivo model of lung fibrosis, which exhibits activated Wnt/β-catenin signaling, decreased expression of CBR2 and proSPC correlated in AT2 cells, whereas ENO1 along with T1a expression was increased.

Conclusion: Proteomic analysis revealed novel proteins differentially expressed in differentiating AT2 cells. Interestingly, newly identified proteins were regulated by β-catenin in vitro and in experimental fibrosis in vivo, suggesting a role in epithelial repair processes upon lung injury.