布比卡因对胃癌细胞抑制作用的机制研究

Inhibitory effect and the mechanism of bupivacaine on gastric cancer cells

目的探讨布比卡因对胃癌细胞增殖、迁移、侵袭的影响及其对长链非编码RNA小核仁RNA宿主基因(LncRNA SNHG16)/微小RNA-302c-3p(miR-302c-3p)分子轴的调控作用。方法体外培养人胃癌细胞AGS,细胞中分别加入3个剂量(50,100,200μmol·L^(-1))的布比卡因处理细胞(命名为低、中、高3个剂量实验组),分别将pcDNA、pcDNA-SNHG16转染至AGS细胞随后加入高剂量布比卡因处理细胞(命名为第1转染组、第2转染组)。噻唑蓝法、Transwell实验用于检测AGS细胞增殖、迁移及侵袭,实时荧光定量-PCR法用于分析SNHG16和miR-302c-3p基因表达水平,通过双荧光素酶报告实验确定SNHG16和miR-302c-3p基因之间靶向关系。结果空白对照组和低、中、高3个剂量实验组的细胞存活率分别为(99.21±5.45)%,(83.98±11.87)%,(59.78±4.93)%,(41.00±5.45)%;这4组的SNHG16基因的表达水平分别为0.98±0.05,0.70±0.06,0.49±0.07,0.30±0.06;这4组的miR-302c-3p基因的表达水平分别为0.98±0.04,1.26±0.11,1.65±0.12,2.29±0.31;这4组的迁移细胞数分别为(104.56±3.57),(84.56±6.86),(60.44±10.78),(36.11±5.42)个;这4组的侵袭细胞数分别为(117.89±7.66),(84.67±6.78),(63.56±6.48),(28.89±4.20)个。SNHG16可竞争性结合miR-302c-3p。第1转染组、第2转染组细胞存活率分别为(41.73±13.11)%,(90.15±5.83)%;miR-302c-3p的表达水平分别为0.99±0.06,0.47±0.17;迁移细胞数分别为(36.89±8.34),(74.78±8.63)个;侵袭细胞数分别为(28.67±4.87),(79.11±10.56)个。上述指标:3个剂量实验组与空白对照组比较差异均有统计学意义(均P<0.05),且有剂量组依赖性;第2转染组与第1转染组比较差异均有统计学意义(均P<0.05)。结论布比卡因可通过下调SNHG16基因的表达而上调miR-302c-3p基因的表达进而减弱胃癌细胞增殖、迁移及侵袭能力。

Objective To explore the effect of bupivacaine on cell proliferation, migration and invasion in gastric cancer and its regulation on the molecular axis of long noncoding RNA small nucleolar RNA host gene(LncRNA SNHG16)/microRNA-302 c-3 p(miR-302c-3p). Methods Culture human gastric cancer cells AGS in vitro,add different three doses(50,100,200 μmol·L;)of bupivacaine-treated cells(named experimental-L, experimental-M,experimental-H groups, respectively);transfect pcDNA, pcDNA-SNHG16 into AGS cells and then add high dose bupivacaine treated cells( named transfection-1 group, transfection-2 group). MTT and Transwell chamber experiments were used to detect the proliferation, migration and invasion of AGS cells. The expression of SNHG16 and miR-302 c-3 p gene were detected by real time fluorescence quantitative-PCR method.The dual luciferase report experiment was used to detect the targeting relationship between SNHG16 and miR-302 c-3 p gene.Results The cell survival rates of blank control group, experimental-L,experimental-M,experimental-H groups were(99. 21 ± 5. 45)%,(83. 98 ± 11. 87)%,(59. 78 ± 4. 93)%,(41. 00 ± 5. 45)%, respectively;the expression levels of SNHG16 gene in the 4 groups were 0. 98 ± 0. 05, 0. 70 ± 0. 06, 0. 49 ± 0. 07, 0. 30 ± 0. 06,respectively;the expression levels of miR-302 c-3 p gene in the 4 groups were 0. 98 ± 0. 04, 1. 26 ± 0. 11,1. 65 ± 0. 12, 2. 29 ± 0. 31, respectively;the numbers of migrating cells in the 4 groups were(104. 56 ± 3. 57),(84. 56 ± 6. 86),(60. 44 ± 10. 78),(36. 11 ± 5. 42),respectively;the number of invading cells in the 4 groups were(117. 89 ± 7. 66),(84. 67 ± 6. 78),( 63. 56 ± 6. 48) and( 28. 89 ± 4. 20), respectively.SNHG16 gene can competitively bind miR-302 c-3 p gene. The cell survival rates of the transfection-1 group and transfection-2 group were(41. 73 ± 13. 11)% and(90. 15 ± 5. 83)%, respectively;the expression levels of miR-302 c-3 p gene were0. 99 ± 0. 06, 0. 47 ± 0. 17,respectively;the numbers of migrating cells were(36. 89 ± 8. 34) and(74. 78 ± 8. 63)cell,respectively;the numbers of invasive cells were( 28. 67 ± 4. 87) and( 79. 11 ± 10. 56) cell, respectively.Comparison between the three dose experimental groups and the blank control group,the differences of the factors were statistically significant( all P< 0. 05). The transfection-2 groups was compared with transfection-1 groups,the difference of the factors were statistically significant( all P< 0. 05).Conclusion Bupivacaine could reduce the proliferation, migration and invasion of gastric cancer cells by down-regulating the expression of SNHG16 and up-regulating the expression of miR-302 c-3 p.