| 4-DAMP抑制A375黑色素瘤增殖作用及机制研究 |
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| 作者单位： | ;1.哈尔滨医科大学药理教研室(省部共建科技部国家重点实验室);2.海南医学院第一临床学院;3.海南医学院附属第一医院麻醉科;4.海南医学院药学院 |
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| 摘 要： | 目的:研究毒蕈碱型M_3受体拮抗剂(4-diphenylacetoxy-N-methylpiperidine-methiodide,4-DAMP)对黑色素瘤增殖的抑制作用,从而探讨M_3受体拮抗剂在抗肿瘤领域的发展前景及意义。方法:利用MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide]检测肿瘤细胞活力;EdU荧光染色观察细胞增殖情况。成年雄性BALB/c裸小鼠分为对照组、5-氟尿嘧啶(5-Fluorouracil, 5-Fu)组(20 mg·kg~(-1)·d~(-1))及4-DAMP组(2 mg·kg~(-1)·d~(-1)),建立荷瘤模型。连续给药21 d后处死小鼠,计算抑制率、脾指数。实时定量PCR (quantitative reverse-transcription polymerase chain reaction, qRT-PCR)检测A375细胞系中M3R、MIA(melanoma inhibitory activity)、SP-1 (transcription factor Sp1)、Lnc-SPRY4-IT1(SPRY4 Intronic Transcript 1)的基因表达。Western Blot检测ERK1/2(extracellular signal-regulated kinase 1/2)蛋白含量的表达。结果:MTT及EdU均表明,4-DAMP可明显抑制A375细胞的活力(P<0.01)。动物实验结果表明,5-Fu组与4-DAMP组肿瘤体积及质量均明显减小(P<0.05);H&E染色可见细胞间隙变大,结缔组织纤维增多。qRT-PCR结果表明,4-DAMP或转染si-M3R均可降低细胞内MIA、SP-1以及LncRNA-SPRY4-IT1的表达。Western Blot结果表明,4-DAMP或转染si-M3R后均可降低细胞内ERK1/2的表达。结论:M_3受体拮抗剂4-DAMP在细胞及动物实验中均能抑制黑色素瘤增殖发挥抗肿瘤功效,降低肿瘤细胞内MIA、SP-1的mRNA和LncRNA-SPRY4-IT1的表达,同时抑制了ERK1/2的表达。
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| 关 键 词： | A375细胞 黑色素瘤 4-DAMP 增殖 |
Inhibitory effect of M_3 receptor antagonist 4-DAMP on melanoma proliferation of A375 cell line |
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| Affiliation: | Department of Pharmacology(State-Province Key Laboratories of Biomedicine and Pharmaceutics),Harbin Medical University,150081 Harbin,Heilongjiang Province;Shandong Provincial Third Hospital,250000 Jinan,Shandong;Department of Pharmacology(State-Province Key Laboratories of Biomedicine and Pharmaceutics),Harbin Medical University,150081 Harbin,Heilongjiang Province;The First Clinical College,Hainan Medical College,571199 Haikou,Hainan;Department of Anesthesiology,The First Affiliated Hospital of Hainan Medical College,570102 Haikou,Hainan;Hainan Medical College,571199 Haikou,Hainan |
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