Experimental Study On The Regulatory Effect Of α7-acetylcholine Receptor On Cell Proliferation, Apoptosis And Invasion Observing Neural Infiltration In Cholangiocarcinoma

Objective:To study effect of a7AchR agonist nicotine and antagonist α-BTX on cell proliferation, apoptosis and invasion of bile duct carcinoma cell line QBC939, and Preliminary Probe on the regulatory effect and related Mechanism of a7-acetylcholine receptor on cell proliferation and invasion.Methods:1.MTT assay was carried out to determine the role of a7AchR agonist nicotine and antagonist α-BTX in proliferation of QBC939cells.2.transwell invasion model was used to determine effect of a7AchR agonist nicotine and antagonist α-BTX in invasion of QBC939cells.3. Detect the effect of nicotine and α-BTX pretreatment on the survive ability of cholangiocarcinoma cells when applied with5-FU by using MTT and Flat cloning formation experiment.4.RT-PCR was used to determine the effect of a7AchR agonist nicotine and antagonist α-BTX in the expressing of PTEN of QBC939cells.Results:With the stimulate concentration of nicotine increased, the ability of proliferation was significant enhanced in48h and72h (48h:F=80.86, P<0.05;72h: F=61.49P<0.05).The effect of promoting proliferation was time-dependent, The effect was more significant in48h and72h than24h (F=0.49, P>0.05), but also48h>72h.Meanwhile,contrast with other experiment group, the quantity of QBC939cell crossed Matrigel was significantly increased with nicotine treatment (P<0.05). a7AchR antagonist α-BTX significantly inhibit pro-tumor phenotype of nicotine. Applied with5-FU,for various centrations nicotine (10-3g/L,10-4g/L,10-5g/L), the survive rate of QBC939was128%,124%,118%, while α-BTX stimulating group and combined stimulation was92%,94%,93%,92%.And that cloning formation ability of nicotine-exposed (6.2±0.40) is significantly improve than α-BTX stimulating group (3.2±0.20), combined stimulation (3.2±0.20) and control group (3.4±0.33).the expresstion of PTENmRNA of nicotine-exposed group, α-BTX stimulating group,combined stimulation control group is respectively0.24±0.055,0.6±0.062,0.66±0.050,0.62±0.03, The differences between nicotine-exposed group and other experiment group, were statistical significance (p<0.05),but the effect of α-BTX stimulating group, combined stimulation and control group were similar to each other (P>0.05). Conclusion:Nicotine can significantly enhanced QBC939cell proliferation and invasion, and signicantly decrease the chemotherapy sensitivity for Cholangiocarcinoma.nicotine exert the effect of promoting tumor via a7nicotine acetylcholine receptor in vitro. The effect of anti-apoptosis of nicotine was closely related to the express of PTEN.