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The Effects of Prostaglandin E2 on Cell Invasion in HuCCT1 Cells and the Related Mechanisms

Author: WangYiPin
Tutor: LengJing
School: Nanjing Medical University
Course: Pathology
Keywords: Cholangiocarcinoma Attack PGE2 EP1 receptor MMP2 Ca2 / PKC
CLC: R735.8
Type: Master's thesis
Year: 2011
Downloads: 30
Quote: 0
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Abstract


Background: Prostaglandin E 2 (prostaglandin E 2 , PGE 2 ) is arachidonic acid through the cyclooxygenase -2 (COX -2) catalyzed metabolism, PGE 2 and the cell surface, mainly through four kinds of EP receptors (EP1, EP2, EP3 and EP4 receptor) binding, activation of downstream signal transduction pathways that promote tumor cell proliferation, invasion and metastasis. Invasion and metastasis of tumor cells involved in the process matrix and basement membrane composed of extracellular matrix degradation of ECM. MMP (matrix metalloproteinase, MMPs) are a class can degrade ECM, promote tumor cell diffusion transfer enzymes, matrix metalloproteinase 2 (MMP2) in the malignant phenotype of tumor cells is particularly critical. Recent studies have shown that a large number, MMP2 and liver cancer invasion and metastasis very close relationship. Early experiments confirmed that both HCC cell surface expression of four kinds of EP receptors, and in hepatocellular carcinoma cells PGE 2 through EP1 receptor can significantly enhance the ability of invasion and metastasis of cancer cells, so the study of this basis to continue to explore PGE 2 on human cholangiocarcinoma cell invasion ability of HuCCT1. Objective: a study PGE 2 on cholangiocarcinoma cell invasion ability of HuCCT1. (2) investigate the PGE 2 and EP1 receptor signaling pathway cholangiocarcinoma HuCCT1 cell MMP2 expression and activity. 3 To explore EP1 receptor signaling pathway in PGE 2 -induced cholangiocarcinoma cell invasion HuCCT1 role in the process. Methods: 1. Conventional methods bile duct cells cultured cells as well as cancer cells do not express HuCCT1 EP receptor human embryonic kidney cell line HEK293. 2. PGE 2 or EP1 receptor agonist (inhibitors) processing HuCCT1 cells, Transwell assay its effect on cell invasion. 3. PGE 2 or EP1 receptor agonist (inhibitor) treatment HuCCT1 cells, RT-PCR assay MMP2 mRNA level changes; zymography assay in cell culture supernatant MMP2 activity changed. 4 using plasmid transfection experiments with methods to select stable expression strain stably expressed in HEK293 cells EP1 receptor protein, EP1 receptor for further study of MMP2. 5. PKC inhibitor or calcium chelator treatment HuCCT1 cells, RT-PCR assay MMP2 mRNA level changes; zymography assay in cell culture supernatant MMP2 activity changes; Transwell cell invasion assay variation. Results: 1. 5μM PGE 2 , 5μM EP1 receptor agonist 17-PT-PGE 2 HuCCT1 cells 18h after treatment, Transwell invasion assay found than in the control cells groups were increased by 81.18% (P lt; 0.05), 100% (P lt; 0.01). 2. 5μM PGE 2 , 5μM EP1 receptor agonist 17-PT-PGE 2 HuCCT1 cells 24h after treatment, compared with control group, MMP2 mRNA expression levels were an increase of 89.14% (P lt; 0.01), 163.89% (P lt; 0.01); MMP2 activity were increased 69.11% (P lt; 0.05), 117.65% (P lt; 0.01). 3. 5μM PGE 2 , 5μM EP1 receptor agonist 17-PT-PGE 2 Processing Stable transfection of HEK293 cells EP1R-pcDNA3 after 24h, MMP2 enzyme activity than control group increased 36.07% (P lt; 0.05), 61.59% (P lt; 0.05). 4. 10μM EP1 receptor inhibitor sc-51322 treated HuCCT1 cells, MMP2 mRNA levels and activity of MMP2 than PGE 2 treatment group decreased by 47.74% (P lt; 0.05), 84.58% ( P lt; 0.01). 5. 5μM PKC inhibitor BIS-1, 10μM calcium chelator BAPTA-AM treatment HuCCT1 cells, with 17-PT-PGE 2 treatment group, MMP2 mRNA levels were decreased by 44.17% (P lt; 0.05), 34.42% (P lt; 0.05); MMP2 activity decreased by 70.95% (P lt; 0.05), 71.82% (P lt; 0.05). 6. 10μM sc-51322, 5μM BIS-1 and 10μM BAPTA-AM treatment HuCCT1 cells 18h after the invasion of cells than in the control group were reduced by 59.77% (P lt; 0.01), 64.19% (P lt; 0.01), 53.55 % (P lt; 0.01). Conclusions: PGE 2 HuCCT1 significantly enhanced cholangiocarcinoma cell invasion, this regulation may be through EP1 receptor / Ca 2 / PKC signaling pathway and increased expression of MMP2 activity to be achieved.

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CLC: > Medicine, health > Oncology > Gastrointestinal Cancer > Gallbladder, bile duct cancer
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