Lectin-Micropatterned Biointerfaces Regulate Cytoskeletal Organization and Viscoelastic Phenotypes of Bladder Cancer Cells

Abstract

Altered glycosylation is a hallmark of cancer progression and provides opportunities to modulate cell adhesion through glycan–lectin interactions. In this study, we investigate how lectin-mediated adhesion and substrate micropatterns influence cytoskeletal organization and viscoelastic properties of bladder cancer cells. Bladder cancer cell lines─HCV29, 5637, and T24 cells─representing different malignancy stages were cultured on homogeneous and micropatterned lectin-functionalized substrates. Lectins recognizing N-glycans (Dolichos biflorus agglutinin, DBA; Wheat germ agglutinin, WGA) and fucosylated glycans (Lotus tetragonolobus lectin, LTL) were used to selectively position bladder cancer cells via glycan-dependent adhesion. Cell morphology and cytoskeletal organization were analyzed using fluorescence microscopy, while nanomechanical and rheological properties were quantified by atomic force microscopy (AFM) using nanoindentation and stress-relaxation experiments. All studied lectins showed affinity for HCV29, 5637, and T24 cells. HCV29 and T24 cells displayed favorable interactions with N-glycan-binding lectins, whereas 5637 cells showed reduced sensitivity to lectin chemistry. Lectin-striped micropatterns induced pronounced morphological changes, including cell elongation and cytoskeletal polarization. AFM measurements demonstrated that both surface chemistry and surface confinement modulate viscoelastic phenotype of cells, with an increase in apparent Young’s modulus observed for cells growing on lectin stripes. Viscoelastic parameters such as the equilibrium elastic modulus and Deborah number increased for 5637 and T24 cells, indicating a shift toward more solid-like behavior. We found that lectin affinity alone did not determine cell positioning or mechanical adaptation. Instead, cell-line–specific responses emerged from the combined effects of initial adhesion cues, geometric confinement, and subsequent adaptive cellular processes. Our findings show that lectin-based micropatterning provides a biologically relevant modulating cell organization and probing the mechanobiological behavior of bladder cancer cells, promising a framework for developing automated biomechanical assays and mechanophenotypic biomarkers relevant to cancer research.