Multimodal Imaging Unveils the Impact of Nanotopography on Cellular Metabolic Activities

Nanoscale surface topography is an effective approach in modulating cell-material interactions, significantly impacting cellular and nuclear morphologies, as well as their functionality. However, the adaptive changes in cellular metabolism induced by the mechanical and geometrical microenvironment of the nanotopography remain poorly understood. In this study, we investigated the metabolic activities in cells cultured on engineered nanopillar substrates by using a label-free multimodal optical imaging platform. This multimodal imaging platform, integrating two photon fluorescence (TPF) and stimulated Raman scattering (SRS) microscopy, allowed us to directly visualize and quantify metabolic activities of cells in 3D at the subcellular scale. We discovered that the nanopillar structure significantly reduced the cell spreading area and circularity compared to flat surfaces. Nanopillar-induced mechanical cues significantly modulate cellular metabolic activities with variations in nanopillar geometry further influencing these metabolic processes. Cells cultured on nanopillars exhibited reduced oxidative stress, decreased protein and lipid synthesis, and lower lipid unsaturation in comparison to those on flat substrates. Hierarchical clustering also revealed that pitch differences in the nanopillar had a more significant impact on cell metabolic activity than diameter variations. These insights improve our understanding of how engineered nanotopographies can be used to control cellular metabolism, offering possibilities for designing advanced cell culture platforms which can modulate cell behaviors and mimic natural cellular environment and optimize cell-based applications. By leveraging the unique metabolic effects of nanopillar arrays, one can develop more effective strategies for directing the fate of cells, enhancing the performance of cell-based therapies, and creating regenerative medicine applications.