Suppression of fibrinolytic activity, cell adhesion, and viability of Lewis Lung Carcinoma cells by nanodot arrays

• Main Authors: Li, Shuang-Hua, 李順華
• Other Authors: Huang, Guewha Steven
• Format: Others
• Language: en_US
• Published: 2010
• Online Access: http://ndltd.ncl.edu.tw/handle/65502029754325378484

碩士 === 國立交通大學 === 材料科學與工程學系奈米科技碩博士班 === 99 === Biomaterials play an important role in bioengineering applications. To evaluate cellular response to a nanoscaled surface, Mouse Lewis Lung Carcinoma (LL/2) cells were cultured onto Nanodot arrays with dot diameters ranging from 10 to 200-nm. The Nanodot arrays were fabricated by anodic aluminum oxide (AAO) processing on TaN-coated wafers. The size-dependent effect of Nanodot arrays on cell growth, corresponding to cell proliferation, cell adhesion, cytoskeletal organization, and gene expression is the main point of our study. Optimized growth occurred when LL/2 cells were cultured on nanodot arrays with dot size at 50-nm, on which maximum viability was maintained even when the cell density reached saturation. Nanodots of 100 and 200-nm prevented viable growth of LL/2 cells with 34.7% and 46.71% reduction at 72 hours, respectively. Cells seeded on 50-nm nanodots showed cell morphology with largest surface area, more extended lamellipodia, and fastest growth rate. Apoptosis-like growth was observed on 100 and 200-nm nanodots with significant reduction in the surface area. Immunostaining was performed against vinculin and actin filaments indicated that 50-nm nanodots promote cell adhesion and cytoskeletal organization. 100-nm nanodots retarded the formation of focal adhesion while 200-nm nanodots inhibited the organization of cytoskeleton. Then, we carry out an in vitro gene function study using quantitative Real-Time RT-PCR. Levels of mRNAs encoding Chemokines (CCL-2/MCP-1 and CCL-3/MIP-1α), Cytokines, (IL-10, TNF-α, and IL-6) and Adhesion molecules (VEGF) increased significantly on the 100 and 200-nm nanodots. The mRNA encoding fibrinolysis molecule (PAI-1) has high expression on 50-nm nanodots. The most cells seeded on 100 and 200-nm nanodots induced inflammatory responses, oxidative stress, and aopotosis. Finally, utilizing Western blot analyzed protein expression. High expression of vinculin and PAI-1 were occurred on 50-nm diameter nanodots. It indicated that 50-nm nanodots have good focal adhesion and suppress fibrinolysis with cells. The results presented here illustrate the ability of nanodot arrays to modulate the growth of LL/2 cells is size-dependent. Optimized growth with the best viability, morphology, adhesion, and imperceptible fibrinolysis occurred at size of 50-nm. Retardation of growth and lamellipodia extension were observed when the dot size was larger than 100-nm. The important of a surface characterization may have a significant effect on cellular behavior. Our device will serve as a convenient and fast tool for tissue engineering and cancer treatment.