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All-trans retinoic acid on human lung adenocarcinoma A549 cell line and the expression of cyclooxygenase-2

Author: QinYan
Tutor: DengShuZuo
School: Luzhou Medical College
Course: Internal Medicine
Keywords: Lung Cancer Lung adenocarcinoma All-trans retinoic acid Cyclooxygenase-2
CLC: R734.2
Type: Master's thesis
Year: 2010
Downloads: 20
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Abstract

Objective: In vitro studies retinoid derivatives - all-trans retinoic acid (all-trans retinoic acid, ATRA) on human lung adenocarcinoma A549 cell line to explore the ATRA on human lung adenocarcinoma A549 cell line and cyclooxygenase -2 (Cyclooxygenase-2, COX-2) expression and its possible mechanism. Methods: (1) develop lung adenocarcinoma A549 cells: the lung adenocarcinoma A549 cells were seeded in 25ml flask, containing 10% fetal bovine serum RPMI1640 medium in 5% CO2, 37 ℃ incubator for culture. (2) in logarithmic growth phase cultured lung adenocarcinoma A549 cells used in the experiment, cells treated with ATRA at a time into 24 hours, 48 hours and 72 hours three time periods, each period and then the drug concentration divided 1 × 10-7mol / L (group 2), 1 × 10-6mol / L (group 3), 1 × 10-5mol / L (group 4), 1 × 10-4mol / L (group 5 ) Four drug concentration group and a control group without drug (group 1) were five groups of 10 test holes. Subject to the concentration of drug in each experimental cell at each experimental time period, with four MTT method (MTT method) were measured, were detected in each experimental group and control group cells absorbance values calculated at each time period each experimental concentrations of drugs on cell growth inhibition rate. (3) to select the best drug action time (72 hours) and the experimental drug concentration by immunocytochemistry SP (streptavidin-peroxidase) method for the determination cytoplasmic expression of COX-2 and cultivate the same time does not make dosing cells control group, the experimental group compared with the control group within the cytoplasm of COX-2 expression whether there are differences. (4) to select the best drug action time (72 hours) and the experimental drug concentration, enzyme-linked immunosorbent assay (enzyme linked immunosorbent assay, ELISA) determination of cell supernatant of COX-2 content change, cultivating the same time Cells without drug as control group, the experimental group compared with the control cells supernatant levels of COX-2 whether there are differences. (5) to select the best drug action time (72 hours) and the experimental drug concentration, cell morphology was observed by transmission electron microscopy changes, training time is not the same dosing cells as control group, the experimental group compared with the control group, whether cell morphology differences. Results: (1) when the concentration of ATRA for different experimental lung adenocarcinoma A549 cells after 24 hours: 1 × 10-7mol / L concentration group, 1 × 10-6mol / L concentration group were compared with the control group, the difference was not statistically Significance (P gt; 0.05), indicating that this effect of two concentrations of ATRA cells for 24 hours, no significant inhibition of cell; 1 × 10-5mol / L concentration group, 1 × 10-4mol / L concentration group and control group respectively. , the difference was statistically significant (P lt; 0.05), indicating that this two concentrations of ATRA on cells significantly inhibited; adjacent comparison between the experimental groups, the difference was not statistically significant (P gt; 0.05); Comprehensive , and with the effect of different concentrations of ATRA lung adenocarcinoma A549 cells after 24 hours, when it reaches a certain concentration of the drug on cell inhibition. (2) When the concentration of ATRA for different experimental lung adenocarcinoma A549 cells after 48 hours: 1 × 10-7mol / L concentration compared with the control group, the difference was not statistically significant (P gt; 0.05), indicating that this concentration of ATRA effector cells 48 hours after cells had no significant inhibition; 1 × 10-6mol / L concentration group, 1 × 10-5mol / L concentration group and 1 × 10-4mol / L concentration group were compared with the control group, the difference was statistically significant (P lt; 0.05), indicating that three concentrations of ATRA on cell significantly inhibited; adjacent comparison between the experimental groups, the difference was statistically significant (P lt; 0.05); summary, with effect of different concentrations of ATRA lung adenocarcinoma A549 cells for 48 hours, when a certain concentration of the drug inhibited the cell, and with the increasing concentration of the more significant inhibition. (3) When the concentration of ATRA for different experimental lung adenocarcinoma A549 cells after 72 hours: 1 × 10-7mol / L concentration group, 1 × 106mol / L concentration group, 1 × 10-5mol / L concentration group, and 1 × 10-4mol / L concentration group were compared with the control group, the difference was statistically significant (P lt; 0.05), shows that the concentration of ATRA on cells was inhibited; adjacent comparison between the experimental groups, the difference was statistically significance (P lt; 0.05); summary, experiment with different concentrations of ATRA lung adenocarcinoma A549 cells 72 hours later, with the experimental group concentration, the inhibitory effect more obvious. (4) Select the different experimental concentrations of ATRA lung adenocarcinoma A549 cells after 72 hours, showing that COX-2 expression in the cytoplasm in the fall, and with the experimental group concentration increased, COX-2 expression decreased even more significantly, with the control group, the difference was statistically significant (P lt; 0.05), indicates that COX-2 may be involved in the process of ATRA on cell inhibition. (5) to select different concentrations of ATRA experimental lung adenocarcinoma A549 cells 72 hours after each experimental group were seen in the supernatant of A549 cells decreased levels of COX-2, and with the experimental group concentration increased, COX-2 content decreased even more significantly, compared with the control group, the difference was statistically significant (P lt; 0.05), adjacent to the experimental groups, the difference was statistically significant (P lt; 0.05), further indicates that COX-2 may be involved in ATRA on inhibition of the process. (6) to select different concentrations of ATRA lung adenocarcinoma A549 cells after 72 hours, visible under the electron microscope concentrations of each experimental lung adenocarcinoma A549 cells showed clear apoptotic trend cell morphology compared with the control group significantly different. Further illustrate ATRA indeed lung adenocarcinoma A549 cells was inhibited. Conclusion: ATRA can inhibit the growth of A549 cells, and its mechanism may inhibit cell growth by inhibiting the expression of COX-2 cells is achieved for the clinical application of ATRA chemoprevention of lung cancer provides a new theoretical basis.

All-trans retinoic acid on human lung adenocarcinoma A549 cell line and the expression of cyclooxygenase-2

Abstract

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