EGF stimulates growth by enhancing capacitative calcium entry in corneal epithelial cells.

摘要: In rabbit corneal epithelial cells (RCEC), we determined whether capacitative calcium entry (CCE) mediates the mitogenic response to epidermal growth factor, EGF. [Ca2+]i was measured with single-cell fluorescence imaging of fura2-loaded RCEC. EGF (5 ng/ml) maximally increased 4.4-fold. Following intracellular store (ICS) depletion in calcium-free medium 10 microM cyclopiazonic acid (CPA) (endoplasmic reticulum ATPase inhibitor), addback elicited plasma membrane Ca2+ influx as a result activation operated channel (SOC) activity. Based on Mn2+ quench measurements fura2 fluorescence, 5 ng/ml enhanced such 2.3-fold, whereas Rp-cAMPS (protein kinase A inhibitor) plus it by 5.3-fold. contrast, SOC blocked 100 2-aminoethyldiphenylborate (2-APB, store-operated inhibitor). During exposure either 50 UO126 (MEK-1/2 or forskolin (adenylate cyclase activator), failed affect [Ca2+]i. RT-PCR detected gene expression of: 1) transient receptor potential (TRP) protein isoforms 1, 3, 4, 6 and 7; 2) IP3R 1-3. Immunocytochemistry, conjunction confocal immunogold electron microscopy, localization TRP4 expression. Inhibition CCE 2-APB and/or CPA, eliminated 2.5-fold increase [3H]-thymidine incorporation induced Taken together, RCEC induces through its stimulation Erkl/2 activity, PKA suppresses these effects may be component which is stimulated ICS depletion.