PKCδ/midkine pathway drives hypoxia-induced proliferation and differentiation of human lung epithelial cells

Epithelial cells are key players in the pathobiology of numerous hypoxia-induced lung diseases. The mechanisms mediating such hypoxic responses of epithelial cells are not well characterized. Earlier studies reported that hypoxia stimulates protein kinase C (PKC)delta activation in renal cancer cells and an increase in expression of a heparin-binding growth factor, midkine (MK), in lung alveolar epithelial cells. We reasoned that hypoxia might regulate MK levels via a PKC delta-dependent pathway and hypothesized that PKC delta -driven MK expression is required for hypoxiainduced lung epithelial cell proliferation and differentiation. Replication of human lung epithelial cells (A549) was significantly increased by chronic hypoxia (1% O-2) and was dependent on expression of PKC delta. Hypoxia-induced proliferation of epithelial cells was accompanied by translocation of PKC delta from Golgi into the nuclei. Marked attenuation in MK protein levels by rottlerin, a pharmacological antagonist of PKC, and by small interfering RNA-targeting PKC delta, revealed that PKC delta is required for MK expression in both normoxic and hypoxic lung epithelial cells. Sequestering MK secreted into the culture media with a neutralizing antibody reduced hypoxia-induced proliferation demonstrating that an increase in MK release from cells is linked with epithelial cell division under hypoxia. In addition, recombinant MK accelerated transition of hypoxic epithelial cells to cells of mesenchymal phenotype characterized by elongated morphology and increased expression of mesenchymal markers, alpha-smooth muscle actin, and vimentin. We conclude that PKC delta/MK axis mediates hypoxic proliferation and differentiation of lung epithelial cells. Manipulation of PKC delta and MK activity in epithelial cells might be beneficial for the treatment of hypoxia-mediated lung diseases.