Mechanics and kinetics of sickled red blood cells in sickle cell disease

Abstract

Abstract: Sickle cell disease (SCD) is a hereditary blood disorder affecting millions worldwide, characterized by the polymerization of mutant hemoglobin under low oxygen conditions. This polymerization stiffens red blood cells (RBCs), reducing their oxygen affinity and dramatically altering their mechanical properties, which contributes to widespread tissue and organ complications. Despite its high morbidity and mortality, treatment options for SCD remain limited, partly due to an incomplete understanding of the biophysical mechanisms underlying disease progression and the absence of reliable biomarkers that reflect these mechanisms.A critical gap in SCD research has been the inability to simultaneously measure the mechanical and oxygenation properties of individual RBCs across the full range of physiological oxygen tensions. To address this, we developed a novel microfluidic platform that enables high-throughput, single-cell measurements of RBC deformability and oxygen saturation under controlled shear stress and oxygen gradients. Using this system, we uncovered a previously unrecognized bi-modal distribution of RBC mechanics and oxygen saturation directly linked to hemoglobin polymerization. Our findings demonstrate that RBCs containing polymerized hemoglobin exhibit an elastic shear modulus up to two orders of magnitude higher than healthy cells and show decreased oxygen affinity. Remarkably, the stiffness of these polymerized cells remains oxygen-independent, although their prevalence increases with decreasing oxygen tension. These insights bridge a critical knowledge gap by linking cellular-level biophysical changes to systemic disease manifestations and open new avenues for mechanistically informed biomarkers of disease severity. Sickle hemoglobin polymerization is an incredibly dynamic process, heavily influenced by cellular and environmental parameters. To study its kinetics, we developed a microfluidic platform to track individual RBCs flowing through dynamic oxygen gradients, enabling real-time observation of oxygen desaturation and hemoglobin polymerization kinetics. This unique capability allows us to quantitatively characterize the thermodynamics and kinetics of intracellular polymerization, extending our insight into this fundamental process in disease pathophysiology. These advances provide a mechanistic foundation for multiscale understanding of SCD, informing therapeutic target identification and evaluation. This work represents a significant step toward translating biophysical insights into improved clinical management and treatment development for sickle cell disease.