The Effect And Mechanism Of Sunitinib On Proliferation And Apoptosis In HaCat Cells Induced By EGF

PurposeTo investigate the influence and internal mechanism of receptor tyrosine kinase inhibitor sunitinib on proliferation and apoptosis in HaCaT cells (HaCaT/E) induced by EGF.Method1. Tested the effects of proliferation of EGF in HaCaT cell by cell count.2. The influence of proliferation of sunitinib in HaCaT/E cells was detected by MTT assay.3. The influence of the cell cycle of sunitinib in HaCaT/E cells was detected by Flow Cytometry Analysis. Western Blot tested the change of protein expression level of cyclinDl and cyclinEl, causing by sunitinib in HaCaT/E cells.4. The influence of apoptosis of sunitinib in HaCaT/E cells was detected by Flow Cytometry Analysis.Western Blot tested the change of protein expression level of apoptosis-related protein cleaved-PARP causing by sunitinib in HaCaT/E cells.The change of apoptotic morphology caused by sunitinib in HaCaT/E cells was observated by electron microscope.5. Western Blot tested the influence of Stat3signaling pathway in sunitinib-treated HaCaT/E cells.Results1. EGF increased cell number of HaCaT cells in a time-dependent manner. During the cultivation of3days, the number of EGF-treated(10ng/mL) HaCaT cells was significantly higher than the control group at all time points (P<0.05). 2. Results of MTT showed that the sunitinib restrained proliferation of HaCaT/E cells in a concentration and time dependent patterns (P<0.05).3. Sunitinib decreased the proportion of cell cycle HaCaT/E cells in S phase in a concentration-and time dependent patterns. Western Blot results showed that sunitinib reduced the expression of cyclin D1and cyclin E1in HaCaT/E cells (P<0.05).4. Sunitinib induced HaCaT/E cells apoptosis in a concentration and time dependent patterns, The expression of apoptosis related proteins cleaved-PARP quantity increased with the concentration of sunitinib raised (P<0.05). Electron microscopy observed that HaCaT/E cells which treated by sunitinib after24h appeared a series of cell apoptosis include volume smaller, nuclear chromatin edge set, concentration, cytoplasmic vacuolation.5. Western Blot results showed that sunitinib pretreatment could significantly inhibit the activation of phosphorylated Stat3in HaCaT/E cells which induced by EGF (P<0.05).Conclusion1. Sunitinib can inhibit HaCaT/E cells’ proliferation, induce HaCaT/E cells’ apoptosis.2. Sunitinib mediates the proliferation and apoptosis of HaCaT/E cells probably by influencing the expression of cell cycle and apoptosis related proteins through Stat3signaling pathways.