Microneedle-Based Analysis Reveals Polarity-Dependent Spatial Regulation of Macrophage Phagocytosis

Phagocytosis and migration in macrophages share key regulators, including Rho family GTPases; however, whether phagocytic membrane extension generates a transient, whole-cell polarity that coordinates migration and spatial prioritization of engulfment remains unclear. Here, we investigated the spatiotemporal coupling between membrane extension and cell migration using opsonized microneedles, which enable controlled stimulation together with long-range membrane extension and backtracking dynamics. During single-needle stimulation, membrane extension was tightly coupled to directional migration, whereas membrane retraction showed weaker coupling. In sequential stimulation with two microneedles, ongoing phagocytosis suppressed competing membrane extension at spatially opposite locations, and a reversal in migration direction was accompanied by initiation of membrane extension toward the second needle. Third-needle experiments further revealed a polarized spatial distribution of phagocytic responsiveness across the cell surface. Consistently, uniform stimulation with multiple opsonized microbeads demonstrated sequential, one-at-a-time engulfment even under near-simultaneous target attachment. These results support a model in which phagocytic membrane extension establishes transient, whole-cell polarity that spatially gates engulfment and coordinates whole-cell migration. The microneedle manipulation platform provides a powerful approach for dissecting the spatiotemporal regulation of phagocytosis and for understanding macrophages as adaptive living micromachines integrating mechanical inputs, transient polarity formation, and sequential target processing.