Quantifying the biophysical properties of stomatocytes in health and disease.

Hereditary stomatocytosis (HSt) comprises red blood cell (RBC) disorders characterized by cup-shaped erythrocytes that share a nominal morphology yet respond oppositely to splenectomy: the procedure is often curative in overhydrated HSt (OHS) and intermediate phenotypes but can precipitate life-threatening thromboembolism in dehydrated HSt (DHS/xerocytosis). This "splenectomy paradox" persists because clinicians cannot predict which patients will benefit. A central obstacle is that RBC biomechanics is governed by several partly independent parameters-membrane shear modulus (μ), bending rigidity (kc), surface-to-volume ratio (S/V), and cytoplasmic viscosity (ηcyto)-that existing assays capture only piecemeal. Here we present an in silico dissipative particle dynamics (DPD) study that constructs a control discocyte (CTR-RBC) and three stomatocyte models (ST-RBC1-3) at fixed membrane area (A0 = 132.9 μm2) and decreasing volume (V = 109.7, 101.5, 89.8 fL), spanning the OHS-to-DHS hydration range. Lacking direct membrane-mechanics data for HSt, we anchor the models to published HSt morphometry, to mechanical measurements on healthy and simulated stomatocytic cells, and to matched brightfield imaging of cup-shaped RBCs used solely as a morphological reference. Tracing this single parameter set through five mechanically orthogonal assays, we find that interendothelial-slit (IES) traversal is geometry-dominated: overhydrated ST-RBC1 requires roughly an order of magnitude higher critical pressure than CTR-RBC, whereas dehydrated ST-RBC3 passes freely. ST-RBC3 nonetheless suppresses membrane tank-treading and raises low-shear whole-blood viscosity by ∼25% at physiological haematocrit, comparable to the hyperviscosity of Gaucher-disease blood. A funnel-obstacle chip amplifies these differences into a label-free centerline-offset signal predicted to separate all four RBC types (∼4.5σ between the extreme phenotypes). Together, these results unite single-cell mechanics, splenic filtration, and hemorheology in one framework, offer a mechanical account of the OHS-versus-DHS splenectomy paradox, and point toward a microfluidic route to pre-operative risk stratification in HSt.