Point-of-Care Monitoring of Red Blood Cell Adhesion As a Marker of Disease Severity and Treatment Response in Sickle Cell Disease

Abnormal adhesion of blood cells to endothelium are pathophysiologically central to Sickle Cell Disease (SCD). Despite recent advances in identifying and targeting cellular adhesion in SCD, knowledge of abnormal cellular adhesion has not been integrated into routine clinical care or trial design, due to a requirement for complicated custom-designed systems, highly trained personnel, and extensive sample manipulation. The SCD Biochip is a novel point-of-care (POC) microfluidic assay that allows rapid, preprocessing-free, and standardized interrogation of red blood cell (RBC) adhesion to endothelium components (fibronectin, FN, or laminin, LN) in whole blood.

After a complete blood count as a standard of care, 15 µL of surplus whole blood was injected, without processing, on to FN- or LN-functionalized Biochips at or above the physiological shear stresses of post-capillary venules (1-5 dyne/cm²). Serial quantitative and qualitative evaluations of RBC adhesion and deformability, using standardized protocols, were performed on >100 SCD adult subjects with correlative clinical data.

Greater RBC adherence per unit area to FN or LN was seen in HbSS compared with compound heterozygous SCD. HbSS samples with a low fetal hemoglobin (HbF) displayed greater adhesion to FN or LN than did samples with a high HbF (8% HbF as the cutoff). RBC adhesion was also significantly greater in samples from HbSS patients with higher LDH levels compared to those with lower LDH levels (500 IU/L as the cutoff). Qualitative analyses revealed deformable and non-deformable HbSS-containing RBCs. Deformable RBCs adhere less to FN when exposed to higher shear stresses, and the most tightly adherent cells were non-deformable RBCs. At highest shear stress (50 dyne/cm²), 75±17% of adhered HbSS RBCs were non-deformable (n=9). Overall, there was a significant association between adhered non-deformable RBCs (%) and serum LDH levels (n=21, Pearson correlation coefficient of 0.79, p<0.005).

Longitudinal analyses performed on individual subjects ≥1 month apart showed stable RBC adhesion to FN or LN being treated for SCD with transfusions or hydroxyurea (stable levels of HbA or HbF). In two patients monitored over 4 months, adhesion to FN or LN dropped >60% from baseline after episodes of transfusion (elevated levels of HbA) or initiation of hydroxyurea (elevated levels of HbF).

The SCD Biochip evaluates, simply and with small sample volumes, complex adhesion properties, which reflect clinical phenotypes, including hemoglobin composition, hemolysis, and treatment status. Applied serially and under varied clinical scenarios, this adaptable POC technology will yield a more precise characterization of abnormal adhesive events in a given individual and a more accurate assessment of response to therapy overall.