Imaging flow cytometry documents incomplete resistance of human sickle F-cells to ex vivo hypoxia-induced sickling

Fertrin, Kleber Yotsumoto; Beers, Eduard J. van; Samsel, Leigh; Mendelsohn, Laurel; Saiyed, Rehan; Nichols, James S.; Hepp, David A.; Brantner, Christine A.; Daniels, Mathew P.; McCoy, J. Philip; Kato, Gregory J.

doi:10.1182/blood-2014-03-559054

Cited by 6 publications

(4 citation statements)

References 4 publications

(1 reference statement)

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“…These results also imply that when investigating the influence of HbF, the average concentration of HbF in a cell population is less important than the HbF content in individual RBCs (53). This interpretation is supported by an ex vivo study showing incomplete resistance of F cells in hypoxia-induced sickling (56).…”

Section: Discussionmentioning

confidence: 63%

Kinetics of sickle cell biorheology and implications for painful vasoocclusive crisis

Diez-Silva

Kato

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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We developed a microfluidics-based model to quantify cell-level processes modulating the pathophysiology of sickle cell disease (SCD). This in vitro model enabled quantitative investigations of the kinetics of cell sickling, unsickling, and cell rheology. We created short-term and long-term hypoxic conditions to simulate normal and retarded transit scenarios in microvasculature. Using blood samples from 25 SCD patients with sickle hemoglobin (HbS) levels varying from 64 to 90.1%, we investigated how cell biophysical alterations during blood flow correlated with hematological parameters, HbS level, and hydroxyurea (HU) therapy. From these measurements, we identified two severe cases of SCD that were also independently validated as severe from a genotype-based disease severity classification. These results point to the potential of this method as a diagnostic indicator of disease severity. In addition, we investigated the role of cell density in the kinetics of cell sickling. We observed an effect of HU therapy mainly in relatively dense cell populations, and that the sickled fraction increased with cell density. These results lend support to the possibility that the microfluidic platform developed here offers a unique and quantitative approach to assess the kinetic, rheological, and hematological factors involved in vasoocclusive events associated with SCD and to develop alternative diagnostic tools for disease severity to supplement other methods. Such insights may also lead to a better understanding of the pathogenic basis and mechanism of drug response in SCD.sickle cell anemia | vasoocclusion | capillary obstruction ratio | cell deformability | Aes-103

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Section: Discussionmentioning

confidence: 63%

Kinetics of sickle cell biorheology and implications for painful vasoocclusive crisis

Diez-Silva

Kato

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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100

134

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“…This method could be applied to other hemoglobins with appropriate antibodies. Moreover, combining quantitative measurements of different hemoglobins and the total hemoglobin per RBC, would inform about their relative amount and concentration, which is of interest for an anti‐polymerization effect, using the corpuscular Hb concentration (MCHC) per RBC 21,28 . However, the latter requires an individual volume measurement which is difficult to obtain when the shape is not regular as in SCD.…”

Section: Discussionmentioning

confidence: 99%

Individual red blood cell fetal hemoglobin quantification allows to determine protective thresholds in sickle cell disease

et al. 2020

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Polymerization of the sickle hemoglobin (HbS) is a key determinant of sickle cell disease (SCD), an inherited blood disorder. Fetal hemoglobin (HbF) is a major modulator of the disease severity by both decreasing HbS intracellular concentration and inhibiting its polymerization. However, heterocellular distribution of HbF is common in SCD. For HbS polymerization inhibition, the hypothesis of an "HbF per red blood cell (HbF/RBC) threshold" requires accurate measurement of HbF in individual RBC. To date, HbF detection methods are limited to a qualitative measurement of RBC populations containing HbF-the F cells, which are variable. We developed

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“…further evaluated the concentration of intracellular HbF on morphology of individual sickle RBCs. 57 Deoxygenated sickle RBCs are first fluorescently labeled against HbF and then imaged using IFC which determines the HbF expression and morphology of the cells. 57 Automated classification of RBC morphologies can also be accomplished by standardizing acquisition of high-resolution microscopic images of individual RBCs using microfluidic devices, and then automatically classifying RBC shape using standard decision-tree-based algorithms or novel approaches based on deep learning (Figure 3).…”

Section: Rbc Morphologymentioning

confidence: 99%

“…57 Deoxygenated sickle RBCs are first fluorescently labeled against HbF and then imaged using IFC which determines the HbF expression and morphology of the cells. 57 Automated classification of RBC morphologies can also be accomplished by standardizing acquisition of high-resolution microscopic images of individual RBCs using microfluidic devices, and then automatically classifying RBC shape using standard decision-tree-based algorithms or novel approaches based on deep learning (Figure 3). 40,58 These image analysis techniques can also be applied to a variety of traditional protocols (such as basic peripheral blood smears and sickling assays) as well as emerging microfluidic technologies that investigate RBC sickling behavior under hypoxia.…”

Section: Rbc Morphologymentioning

confidence: 99%

Blood rheology biomarkers in sickle cell disease

Rab

Shevkoplyas

et al. 2020

Exp Biol Med (Maywood)

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Sickle cell disease (SCD) is the most common inherited blood disorder, affecting approximately 100,000 patients in the U.S. and millions more worldwide. Patients with SCD experience a wide range of clinical complications, including frequent pain crises, stroke, and early mortality, all originating from a single-point mutation in the β-globin subunit. The RBC changes resulting from the sickle mutation lead to a host of rheological abnormalities that diminish microvascular blood flow, and produce severe anemia due to RBC hemolysis, and ischemia from vaso-occlusion initiated by sticky, rigid sickle RBCs. While the pathophysiology and mechanisms of SCD have been investigated for many years, therapies to treat the disease are limited. In addition to RBC transfusion, there are only two US Food and Drug Administration (FDA)-approved drugs to ameliorate SCD complications: hydroxyurea (HU) and L-glutamine (Endari™). The only curative therapy currently available is allogeneic hematopoietic stem cell transplantation (HSCT), which is generally reserved for individuals with a matched related donor, comprising only 10–15% of the total SCD population. Potentially curative advanced gene therapy approaches for SCD are under investigation in ongoing clinical trials. The ultimate goal of any curative treatment should be to repair the hemorheological abnormalities caused by SCD, and thus normalize blood flow and prevent clinical complications. Our mini-review highlights a set of key hemorheological biomarkers (and the current and emerging technologies used to measure them) that may be used to guide the development of novel curative and palliative therapies for SCD, and functionally assess outcomes. Impact statement Severe impairment of blood rheology is the hallmark of SCD pathophysiology, and one of the key factors predisposing SCD patients to pain crises, organ damage, and early mortality. As novel therapies emerge to treat or cure SCD, it is crucial that these treatments are functionally evaluated for their effect on blood rheology. This review describes a comprehensive panel of rheological biomarkers, their clinical uses, and the technologies used to obtain them. The described technologies can produce highly sensitive measurements of the ability of current treatments to improve blood rheology of SCD patients. The goal of curative therapies should be to achieve blood rheology biomarkers measurements in the range of sickle cell trait individuals (HbAS). The use of the panel of rheological biomarkers proposed in this review could significantly accelerate the development, optimization, and clinical translation of novel therapies for SCD.

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Imaging flow cytometry documents incomplete resistance of human sickle F-cells to ex vivo hypoxia-induced sickling

Cited by 6 publications

References 4 publications

Kinetics of sickle cell biorheology and implications for painful vasoocclusive crisis

Kinetics of sickle cell biorheology and implications for painful vasoocclusive crisis

Individual red blood cell fetal hemoglobin quantification allows to determine protective thresholds in sickle cell disease

Blood rheology biomarkers in sickle cell disease

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