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. 2020 Sep 4;8:819. doi: 10.3389/fcell.2020.00819

FIGURE 2.

FIGURE 2

PINK1 promotes mitophagy, which releases mtDNA that helps drive cyclic stretching-induced inflammation and injury. (A,B) Lung epithelial cells were treated with Pink1 siRNA, cDNA or empty vector and exposed to cyclic stretching (CS) at 20% tension for 4 h. As expected, Western blot and RT-qPCR showed decreased PINK1 protein and mRNA expression in Pink1-deficient lung epithelial cells, but increased PINK expression in cells treated with Pink1 cDNA. (C) RT-qPCR was simultaneously performed to assess the cell-free mtDNA-79 copies. (D) RT-qPCR was simultaneously performed to assess the cell-free mtDNA-230 copies. (E) Transmission electron microscopy was performed to assess cell injury ultrastructurally (magnification ×20000). Red arrows indicate the autophagosomes. (F–H) Enzyme-linked immunosorbent assays were used to assess the levels of IL-1β, IL-6, and TNF-α in the culture medium. (I) MTT assay was used to examine the viability of cells. Experiments were performed in triplicate. aP < 0.05 vs. control group; bP < 0.05 vs. 20% CS group; cP < 0.05 vs. Pink1 cDNA + 20% CS group; dP < 0.05 vs. Pink1 empty vector + 20% CS group.