采用三维模型探索香烟烟雾提取物(CSE)对A549细胞的影响

Using 3-D Model to Study Cigarette Smoke Extract(CSE) Interaction with A549 Cells

本研究采用D-型和L-型自组装短肽作为纳米纤维支架材料进行细胞三维培养体系平台,探索香烟烟雾提取物(CSE)诱导人类Ⅱ型肺泡上皮(A549)细胞氧化损伤后,考察其细胞行为在二维和三维微环境下的差异。A549细胞分别在二维和三维环境中用不同浓度的CSE溶液诱导24 h、48 h或72 h之后,进行细胞增殖活力,乳酸脱氢酶(LDH),细胞周期,细胞凋亡检测,以及运用吖啶橙/溴化乙锭(AO/EB)和4',6-二脒基-2-苯基吲哚二盐酸盐(DAPI)染色观察A549细胞形态。结果表明,与二维环境相比,在三维环境中细胞活力更高,LDH释放量较少,细胞凋亡减少。细胞的周期分布检测表明,随着CSE浓度的增大,发生G1期阻滞;当15%,50%CSE作用时,与二维环境相比,三维培养环境中的G1期细胞比例更低,G2期和S期更高,G1期阻滞减弱。上述结果均表明D-型和L-型肽手性自组装短肽能够构建出适于A549细胞生长、增殖的三维微环境,并能模拟细胞胞外基质,对细胞形态、增殖、周期及凋亡等有重要影响。

In the study,we used D-form and L-form self-assembling peptide nanofiber scaffolds as three-dimensional cell culture system to study the cigarette smoke extract（CSE） induced by human type Ⅱ alveolar epithelial（A549） cells after oxidative damage,and compared the difference of cell behaviors in two dimensional and three dimensional microenvironment.A549 cells in 2-dimensional and 3-dimensional environment were exposed to various concentrations of CSE for 24 h,48 h or 72 h.Then proliferation ability,cell cycle,lactate dehydrogenase（LDH）,and cell apoptosis were measured and the morphology of the cells was observed using AO-EB and DAPI staining.The results showed that A549 cell viability was higher,LDH activity was lower and cell apoptosis reduced in 3-D than that in 2-D.The results of cell cycle distribution showed G1 phase arrest occurred with the increase of the concentration of CSE solution.However,exposed to 10%,15% CSE,the cell proportion in G1 phase was lower,while those in S phase and G2 phase were higher in 3-D compared with 2-D,meanwhile G1 phase arrest decreased.The results indicated that D-form and L-form self-assembling peptides were able to construct a 3-D microenvironment that was suitable for the growth and proliferation of A549 cells,which could mimic the cellular extracellular matrix and play a key role to affect cell behaviors including cell morphology,proliferation,cycle and apoptosis.