NMR-BASED METABOLOMICS OF KIDNEY (HEK 293T) CELLS CULTURED ON SAM-COATED INDIUM TIN OXIDE (ITO)

Human embryonic kidney 293T (HEK 293T) cells have been used in organoid formation and specialized applications within organoid systems; however, they often suffer from loose adherence, which limits their applicability. To improve the HEK 293T cellular adhesion and proliferation and to provide insights into the pathways involved in adhesion, HEK 293T cells were cultured on tissue culture plastic and glass substrates with surfaces modified with various self-assembled monolayers (SAMs). The ITO-MPS SAM-coated substrate gave the most promising improved cell adherence and proliferation results. 1H NMR spectroscopy was used to analyze the metabolomics present in the media with and without ITO-MPS-SAM coated substrates for a period of 120 hours. The confocal microscopy images are consistent with the findings from NMR and provide a visual confirmation of the enhanced cellular environment. The metabolomic analysis yielded thirty metabolites, including two promoters (ATP and glucose-6-phosphate) and three modulators (tyramine, 5-hydroxytryptophan, and acetate) of adhesion, which were only significantly present
on the fifth day of culture.

人胚肾293T（HEK 293T）细胞已被应用于类器官构建及类器官系统内的特定研究场景，但该细胞常存在贴壁不牢的缺陷，极大限制了其应用范围。为改善HEK 293T细胞的黏附与增殖能力，并解析其黏附相关的分子通路机制，本研究将HEK 293T细胞接种于经不同自组装单分子层（self-assembled monolayers，SAMs）修饰的组织培养塑料基质与玻璃基质上。其中，经ITO-MPS SAM修饰的基质，其细胞黏附与增殖的改善效果最为理想。针对培养时长120小时的、分别添加与未添加ITO-MPS SAM修饰基质的细胞培养基，采用氢谱核磁共振（1H NMR）光谱法对其中的代谢组成分进行分析。共聚焦显微镜成像结果与核磁共振分析结果高度吻合，为细胞培养微环境的优化提供了可视化佐证。本次代谢组分析共鉴定出30种代谢物，其中包含2种黏附促进因子（ATP与葡萄糖-6-磷酸）及3种黏附调节因子（酪胺、5-羟色氨酸与乙酸盐），上述代谢物仅在培养第5天时显著检出。