Facile calcium ion-regulated grafting of dense and highly stretched hyaluronan for selective mediation of cancer cells rolling under high-speed flow.

Abstract

The development of materials that selectively mediate the rolling of cancer cells is important for the high-throughput enrichment of high-speed cancer cells. Here we constructed a dense and stretched low molecular weight hyaluronic acid (HA_9.6k)-modified surface to selectively promote the rolling of CD44-high cancer cells. The HA surface (calcium ion-regulated HA_9.6k surface, Ca-rHA) was fabricated via a calcium ion-regulated method, where calcium ion incorporation induced the shrink of HA_9.6k chains to achieve the highest reported grafting density of about 2.73 ± 0.20 × 10⁴ HA chains μm^-2. Upon the removal of calcium ions, the dense HA_9.6k chains switched to a highly stretched conformation. The high density and flexibility of Ca-rHA bearing abundant binding sites enhanced the rolling of CD44-high cancer cells and reduced the velocity of cells from 1389 µm s^-1 to 99 µm s^-1 (7%), comparable to that of the physiological rolling event and outperforming traditional grafting-to HA and E-selectin, without causing phenotypic changes. When processing complex samples under high-speed flow, Ca-rHA selectively mediated the rolling of cancer cells and enriched their ratio to peripheral blood mononuclear cells from 1:1 to 15:1. As the only reported artificial biomaterial capable of selectively mediating the rolling of cancer cells under a physiological high-speed flow, Ca-rHA holds promise in enriching intact cells for downstream analysis in the clinics by encouraging the surface-cell contacts. STATEMENT OF SIGNIFICANCE: The development of materials that selectively mediate the rolling of cancer cells is important for the high-throughput enrichment of cancer cells rolling under high-speed flow, yet is less reported. To selectively promote the rolling of cancer stem cell marker CD44-high cancer cells, a surface with dense and stretched low molecular weight hyaluronic acid (HA_9.6k) was constructed. With Ca²⁺ regulation, HA_9.6k chains shrank to achieve the highest reported grafting density of 2.73 ± 0.20 × 10⁴ chains μm^-2 and further switched to a highly stretched conformation after the removal of Ca²⁺ ions. As the only reported artificial biomaterial capable of selectively mediating the rolling of cancer cells under a physiological high-speed flow, this Ca²⁺-regulated HA_9.6k surface holds promise in enriching intact cells for downstream analysis in the clinics.