肉桂醛对黑素瘤A375细胞侵袭能力的影响

Effect of Cinnamic Aldehyde on Invasion of Human Melanoma A375 Cells

目的探讨肉桂醛对体外二维和三维培养的人黑素瘤细胞株A375侵袭能力的影响。方法 A375细胞在体外二维培养后,在培养基中加入不同浓度(0,10,20和40μmol/L)肉桂醛,24h后细胞划痕实验检测细胞迁移,Transwell实验检测细胞侵袭情况。在组织工程皮肤基础上加入A375细胞,培养第5天开始在培养基中加入肉桂醛20μmol/L。在培养第15天固定组织工程皮肤标本,HE染色观察A375细胞生长和侵袭情况,免疫组织化学观察PCNA和MMP-9表达情况。二维和三维培养的A375细胞经肉桂醛处理后,ELISA法检测NF-κB活化情况。结果肉桂醛在二维培养条件下能抑制A375细胞的迁移,也能抑制A375细胞的侵袭,抑制强度和肉桂醛浓度成正比,各浓度试剂作用差异具有统计学意义(P均<0.05)。肉桂醛能抑制三维培养模型内黑素瘤A375细胞的增殖和侵袭,肉桂醛处理前后黑素瘤细胞PCNA和MMP-9的表达差异也具有统计学意义(P均<0.01)。肉桂醛能抑制二维和三维培养的A375细胞的NF-κB活化,各浓度间作用差异具有统计学意义(P均<0.05)。结论肉桂醛能显著抑制体外二维和三维培养模型中黑素瘤细胞的侵袭。抑制NF-κB活化可能是其作用机制之一。

Objective To investigate the effect of cinnamic aldehyde on invasion of A375 cells in two-dimensional （2D） and three-dimensional （3D） cuhure. Methods A375 cells in two-dimensional culture were treated with cinnamic aldehyde of different concentration （0,10,20 and 40μM） for 24 hours. And then to measure the migration of melanoma A375 cell by in vitro wound closure assay, and to assess the invasiveness of them by Transwell assay. Melanoma A375 cells were inoculated onto the surface of tissue-engineered skin and cultured at the air- liquid interface. From day 5, cinnamic aldehyde at a concentration of 20 μmol/L was added to the culture me- dium. Skin samples cultured for fifteen days were stained with H. E for the observation of growth and invasion of A375 cells. The immunohistoehemical staining of proliferating cell nuclear antigen （PCNA） and matrix met- alloproteinase （MMP）-9 was carried out separately. After the human A375 melanoma cells were treated with cinnamic aldehyde in 2D and 3D culture,the NF-kB activation in A375 cells was detected with ELISA. Results Cinnamie aldehyde inhibited A375 cells migration to scratch wounds and reduced the invasion of A375 cells substantially in a concentration-dependent manner （ P 〈 0.05 ）. HE staining showed cinnamic aldehyde inhibited the proliferation and invasion of melanoma cells in the 3D culture model. The expression levels of PCNA and MMP-9 in melanoma cells were significantly different between control and CA-treated skin equivalents （P 〈 0. 01 ）. Cinnamic aldehyde suppressed NF-kB activation in human A375 melanoma cells in 2D and 3D culture in a concentration-dependent manner （ P 〈 0.05 ）. Conclusion Cinnamic aldeyde significantly prevents invasion of melanoma A375 cells in 2D and 3D culture model. NF-kB inactivation may be one of the mechanisms of cinnamic aldehyde effects on A375 melanoma cells.