Objective: To study the in vitro effect and mechanism of Napsin A gene transfection into type II alveolar epithelial cells on pulmonary fibrosis.
Methods: A recombinant lentiviral plasmid PLJM1-Napsin A was constructed and transfected into human type II alveolar epithelial cell line A549. The model of pulmonary fibrosis was established by the in vitro stimulation of A549 cells by transforming growth factor beta-1 (TGF-β1). The morphological changes were observed continuously under inverted microscopy. The proliferation of transgenic and non-transgenic cells was detected by MTT. To observe the degree of epithelial-mesenchymal transition (EMT) by TGF-β1 intervening A549 cells, the expressions of E-cadherin and fibronectin were determined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. Lastly the protein expression of focal adhesion kinase (FAK) was detected by Western blot to investigate the mechanism.
Results: The result of sequencing the recombinant lentiviral plasmid PLJM1-Napsin A was the same as the design sequence. Napsin A mRNA and protein were expressed in transgenic A549 cells (P<0.01). The model of pulmonary fibrosis was established successfully based on the morphology of transformed interstitial cell. As compared with the control group, the proliferation rate of transgenic cells decreased significantly (P<0.05). The mRNA and protein expression of E-cadherin significantly decreased in the model of pulmonary fibrosis (P<0.01), while the expression of fibronectin markedly increased (P<0.01). But the change rate of transgenic cells decreased (P<0.01, P<0.05). The expression of FAK was significantly elevated after the stimulation of TGF-β1 (P<0.01). But the upward trend of the transgenic cells was smaller as compared with the control group (P<0.01).
Conclusion: Pulmonary fibrosis may be suppressed by the transfection of Napsin A gene into type II alveolar epithelial cells. And the mechanism may be through the inhibition of integrin signal transduction.