Pero, Eleonora (2025) Blood Cell Dynamics as a Mechanobiological Marker of Disease: Insights into Thrombosis and Cystic Fibrosis. [Tesi di dottorato]

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Abstract

Blood represents a uniquely complex and informative biological fluid. Circulating throughout the entire body, it continuously integrates mechanical, biochemical, and inflammatory cues from every organ and tissue. This systemic connectivity makes blood an exceptional window into human health. Among circulating cells, red blood cells (RBCs) and platelets are particularly sensitive to changes in their microenvironment. The dynamic behavior of RBCs (how they deform and interact under flow) encodes information about membrane integrity, cytoskeletal structure, and ion-channel activity. Pathological alterations, whether hereditary or acquired, leave measurable mechanical fingerprints. Similarly, the ability of platelets to marginate, adhere to the vessel wall, and form aggregates during hemostatic process reflects the delicate balance between biological signaling, cellular mechanics and hemodynamic forces. Hence, the dynamics of blood cells under flow can offer insight into both vascular and systemic dysfunction. Building on this concept, the present Ph.D. thesis investigates the mechanobiology of blood cells as a marker of disease, focusing on two distinct pathological contexts: thrombosis and cystic fibrosis. The first part of this thesis investigates the flow-dependent morphology of platelet aggregates. It reflects a complex interplay of physical and biological drivers and is increasingly recognized as a potential early marker of coagulation disorders. Current approaches to evaluating aggregate morphology are largely qualitative and often overlook the influence of hemodynamic parameters. To address this limitation, the study proposes target metrics and a standardized framework for the quantitative characterization of platelet aggregate morphology. This approach combines microfluidics, confocal microscopy, and advanced image analysis. Applied to healthy subjects, this study can be extended to build a reference dataset for comparisons with thrombotic conditions and related disorders. Microfluidic experiments reveal that shear rate governs aggregate alignment along the flow direction and modulates occlusive potential, while hematocrit enhances substrate coverage and stabilizes thrombus adhesion. Complementary resolved three-dimensional numerical simulations based on lattice-Boltzmann, immersed-boundary, and finite-element methods reveal how vascular geometry and local hemodynamics modulate platelet margination. These results provide a mechanistic link between blood dynamics and thrombus architecture. The second part of the thesis introduces for the first time a quantitative evaluation of RBC deformability in individuals with cystic fibrosis (CF). CF is a disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. CFTR encodes for a chloride channel that, beyond epithelial tissue, is also expressed on RBC membrane. RBC deformability is measured with osmotic gradient ektacytometry (OGE) and compared with healthy controls. Results show a significant altered RBC deformability profile in CF individuals indicative of marked cellular dehydration. Treatment with CFTR modulators restored deformability to near-normal levels, mirroring improvements in clinical parameters and thus highlighting the sensitivity of RBC deformability to ion-channel function. Remarkably, deformability alterations were also detected in CFTR-related disorders, suggesting a potential diagnostic application for challenging or borderline phenotypes. Overall, this work highlights the potential of blood-cell dynamics as a biophysical probe for disease detection and therapeutic monitoring.

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