Research

Photoresponsive substrates

Dynamic substrates are chemically functionalized substrates whose chemical and physical characteristics can be controlled by an application of external stimuli, such as heat, voltage and light. We have developed for the first time a photoresponsive dynamic substrate (J. Am. Chem. Soc. 2004). The substrate bears cell-repellent polymers or proteins via photocleavable 2-nitrobenzyl ester, hence its photoirradiation reaction converts the surface from a state preventing cell adhesion to a state promoting cell adhesion. We applied this substrate for patterned co-culturing of heterotypic cells (Langmuir 2008) and cell migration navigation (J. Am. Chem. Soc. 2007, Colloids Surf. B, 2012). Several modifications in the surface design were made from the original substrate (Chem. Lett. 2008, Phys. Chem. Chem. Phys. 2011, Sci. Technol. Adv. Mater. 2011). We also studied the impact of the surface chemistry on the switchability (Langmuir 2013).

Collective cell migration

Collective cell migration plays critical roles in both physiological and pathological processes. Basically, epithelial cells migrate collectively, whereas mesenchymal cells prefer to migrate as individuals. However, in some spatiotemporally limited situations of life, some cells aggressively ignore this rule. For example, the change in the collective characteristics via epithelial-mesenchymal transition (EMT) or vice versa (MET) is essential in embryonic development and morphogenesis. Also, cancer metastasis can be considered as the loss of the collective feature upon escaping from the original tissue and its retrieval to settle down and form new colony at other tissues. So far, various internal and external factors have been shown to change collective characteristics in vitro and in vivo, however the contribution of cellular niches, composed of surrounding cells and extracellular matrices, is still unclear. To address this issue, we are developing new materials that allow us to engineer cellular nano-, micro-, and mechanical environments and explore the regulation mechanisms of migration collectivity with the help of imaging and biochemical approaches. In our recent paper, we found that spatiotemporal maturation of the cell-cell interactions is important for acquisition of collective characteristics (Biomaterials 2012). We are also interested in building a new model of collective migration based on experimental data and mathematical calculation.

Photoactivatable nanoparticles

Administration of drugs and biological compounds at high spatiotemporal resolutions in living system is important for the analysis of the intertwined inter- and intracellular signal transduction networks. One promising approach for this purpose is using caged compounds, whose activities are suppressed by covalent linkage of a photocleavable protecting group but are restored upon flash photolysis of the photolabile group to release the original biological compounds. However, there are only a limited number of commercially available products, and their de novo construction remains a great challenge, particularly to biological scientists, as it requires precise design, complex synthesis and purification. To overcome this limitation, we have developed colloidal gold nanoparticles (GNPs) bearing photocleavable reactive functional groups and poly(ethylene glycol) (PEG) brushes on the particle surface. Under this molecular design, the GNP together with PEG function as bulky caging groups that suppress the activity of the cargo. In addition, the GNP facilitates the synthesis and purification of caged compounds as a solid phase support. As a proof of concept, we synthesized a caged compound for histamine (J. Am. Chem. Soc. 2009). These materials are useful platforms for the caged compounds synthesis and provide new methodology for engineering and exploring cellular functions.

Fluorescence imaging of protein conformational change in living cells

Proteins involved in intracellular signaling dynamically change their conformation upon activation. We synthesized fluorescent molecules that can be selectively attached to a peptide sequence in living cells. By genetically engineering a protein of interest to have the peptide sequence and labeled it with the fluorescent molecule, we developed a method for real-time imaging of protein conformational change in living cells. Based on this method, we succeeded in visualizing agonist-induced conformational dynamics of a cytosolic protein (calmodulin, Anal. Chem. 2001) and a membrane protein (β2 adrenergic receptor, Biophys. Biochem. Res. Commun. 2008) in living cells.