More interestingly, we observed an inverse relationship between hypoxic WBV and RBC adhesion to LN in hypoxia, a similar behavior as in normoxic conditions (Pearson’s correlation coefficient = -0.6, p=0.03, one-way ANOVA). The hypoxic viscosity results showed that WBV of control blood samples (HbAA, N=3) remained relatively unchanged, while the HbSS samples (N=10) became more viscous under hypoxia (p=0.007, student’s t-test). Consequently, we integrated a micro gas exchanger to our microfluidic system, as we have previously described, in order to probe the change in WBV as well as RBC adhesion to LN from normoxic to hypoxic conditions. Hypoxia plays a crucial role in SCD pathophysiology as it leads to the formation of abnormally adhesive and stiff red blood cells due to hemoglobin polymerization. These findings suggest that WBV and RBC adhesion may play distinct roles in the pathophysiology of homozygous SCD. Interestingly, subjects with a lower WBV exhibited significantly greater RBC adhesion to LN compared to those with a higher WBV (mean adhesion ± SD: 957☗67 per fov vs 452☓31 per fov, p<0.05, Student’s t-test, fov: field of view). Our results showed that individuals with homozygous SCD (HbSS) had significantly higher WBV compared to healthy controls when the hematocrit of the samples were adjusted to 50% (4.8☐.41 cP vs 4.15☐.07 cP, p4 cP, N=21). The blood samples with a volume of 15 μl were first injected into the microchannels at a shear stress of 1 dyne/cm 2, and then non-adherent cells were removed by flowing a buffer solution (1% BSA in PBS) at the same shear stress. RBC adhesion assay – The adhesion tests were performed in laminin-functionalized microfluidic channels. The conversion from mean blood velocity to microfluidic WBV was carried out using a standard curve generated by using clinically measured viscosity values (piston style viscometer). The images were then cross-correlated to compute the mean flow velocities in a given region of interest, which was a function of fluid viscosity. Micro PIV for velocity measurements – Whole blood samples were injected into rectangular microfluidic devices (4 mm x 0.05 mm) under constant physiological pressure (20 mm.Hg), and high resolution images of the blood flow were acquired via an inverted microscope (Olympus IX83). A total number of 10 samples from healthy subjects (HbAA), 14 samples from subjects with HbSC, and 29 samples from subjects with HbSS were tested. Subject clinical information, including medical treatments and previous comorbidities, were acquired after patients had provided a written consent from the Adult Sickle Cell Clinic at University Hospitals Cleveland Medical Center (UHCMC) in Cleveland, Ohio. To assess whole blood viscosity (WBV) of subjects with sickle cell disease (SCD) using a high-throughput microfluidic system integrated with a micro particle image velocimetry (PIV) technique under normoxic or hypoxic conditions, and to examine its clinical associations and impact.īlood collection – Whole blood samples from de-identified healthy donors and SCD subjects (≥18) were collected in EDTA (Ethylenediaminetetraacetic acid) containing vacutainers based on an Institution Review Board (IRB) approved protocol. Further, for the first time, we report both WBV and RBC adhesion levels simultaneously using this microfluidic system. Here, we describe a microfluidic platform integrated with the micro particle image velocimetry (PIV) technique to quantify WBV using unprocessed whole blood samples from individuals with SCD. Although the separate effects of these factors on WBV determination have been well documented, a more comprehensive approach that takes into account all these factors in a patient-specific fashion is needed to better understand the role of WBV in SCD pathophysiology. It can be influenced by a variety of factors including plasma viscosity, RBC deformability, hematocrit, and plasma protein levels. Whole blood visocity (WBV) is a pivotal biophysical parameter in many cardiovascular diseases and hematological disorders such as sickle cell disease (SCD). 1Case Western Reserve University, Cleveland Heights, OHģCase Western Reserve University, Cleveland, OH
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