Dissertation > Medicine, health > Oncology > Skin tumors

The Expression and Regulation of the PI3K/Akt Signaling Pathway in Hemangioma Vascular Endothelial Cell in Vitro

Author LiTangJiang
Tutor YuSong
School Zunyi Medical College,
Course Pediatric surgery
Keywords hemangioma vascular endothelial cell cell culture PI3K/Akt signalpathways cell cycle apoptosis regulation
CLC R739.5
Type Master's thesis
Year 2012
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ObjectiveTo observe the expression of PI3Kp110catalytic subunit and p-Akt, Akt in cultured vascular endothelial cells, and its regulation mean while explored the effect of PI3K/Akt signal pathway in the different phases of children hemangioma.Methods1. The hemangioma vascular endothelial cells which came from the sterile hemangioma were cultured by using the tissue culture method and established hemangioma endothelial cell lines, observing the proliferation characteristics, drawing the growth curve.The hemangioma vascular endothelial cell was identified by the immunohistochernistry technique observing the expression of the Factor Ⅷantigen which only existed in endothelial cells.2. Synchronized cultured endothelial cells, PI3Kp110, p-Akt and Akt of them were detected by Western blotting mean while cell cycle and cell apoptosis were examined by flow cytometry technique,then analysis the level of PI3Kp110, p-Akt and Akt expression and the correlation in hemangioma vascular endothelial cell cycle and apoptosis.3. When the cells were in the logarithmic phase,the culture medium was exchanged to serum-free medium to make the cell synchronization in48hours.25umol/L LY294002,10nmol/L rapamycin and1.0g/L triamcinolone acetonide were added to observed the effects on the expressions of PI3Kp110, Akt and p-Akt, the cell cycle distribution and cell apoptosis with the hemangioma vascular endothelial cell when the cells were cultured after24hours,and then the cell apoptosis,cell cycle related of PI3Kp110, p-Akt and Akt expression level of vascular endothelial cells were analyzed.Results1. The hemangioma vascular endothelial cells migrated from the tissue to the postcultural after four or five days, only a small number of cells spread after one week adherent growth, growth and division after two weeks, three weeks after the cells began to grow fast, and around, the cells covered the bottom plate.Inverted phase contrast microscope and found that the isolated endothelial cultured epithelial cell morphology, part of the cells show cricoid arrangement, and has the slabs of appearance.Cultured cells of Factor Ⅷ related antigen was positive.2. The cultured hemangioma vascular endothelial cells expressed p110, p-Akt and Akt protein in vitro.3. After LY294002and rapamycin acts on the cultured hemangioma vascular endothelial cells for24hour, the protein level of p110, p-Akt were lower than control group, control group p110(0.37±0.04), p-Akt(0.22±0.02), intervention group LY294002p110(0.19±0.01),p-Akt(0.09±0.02),the difference was significant(P

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