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The Effect and Mechanism of Sunitinib on Proliferation and Apoptosis in HaCat Cells Induced by EGF

Author: LiuYingKe
Tutor: ChenXiang
School: Central South University
Course: Clinical
Keywords: receptor tyrosine kinase inhibitor sunitinib HaCaT cell proliferation apoptosis
CLC: R96
Type: Master's thesis
Year: 2013
Downloads: 2
Quote: 0
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Abstract


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.

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