Therapeutic Efficacy of Semisynthetic Supra Perfusion Resuscitation Fluids, EAF Peg Alb and EAF Peg Hb, Are Differentiated By Their Cerebral Effects in Animal Models of Sickle Cell Disease

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 773-773
Author(s):  
Craig A Branch ◽  
Min-Hui Cui ◽  
Sangeetha Thangaswamy ◽  
Nicholas Branch ◽  
Seetharama Acharya
Keyword(s):  
Blood Flow ◽  
Sickle Cell Disease ◽  
Oxygen Delivery ◽  
Therapeutic Efficacy ◽  
Systemic Circulation ◽  
Cerebral Oxygen ◽  
Cremaster Muscle ◽  
Cell Disease ◽  
The Brain

Abstract Background: Extension Arm Facilitated (EAF) PEG Alb and EAF PEG Hb are low viscosity semisynthetic hybrid biopolymers which are isoviscous with conventional colloidal plasma expanders but are distinguished from them because they are supra perfusion resuscitation fluids (SPF's). These SPF's have longer half-life, are pseudoplastic and facilitate the production of NO in vivo by increasing shear thinning of RBC's. We recently tested two SPF's, EAF-P5K6 Alb and P3K6 Hb in WT mice, and in two Tg models of Sickle Cell Disease (SCD): the Berkley mouse (BERK), which is a severe anemic model exhibiting a high impairment of systemic blood flow, and in the NY1DD mouse which only exhibits extensive blood flow impairment when challenged with hypoxia followed by reoxygenation. Here we present a comparison of the systemic and cerebral effects of the EAF PEGgylated SPF's. Methods: A single intraperitoneal 10% top-load dose of either drug was given to WT, NY1DD or BERK mice. In NY1DD mice SPF's were administered after hypoxia at the beginning of reoxygenation (8% for 18 hours), while SPF's were given to WT or BERK mice under normoxia conditions. Three hours after the administration of drug, in vivo intra-vital microscopic observation of post-capillary venules in cremaster muscle was performed. In a separate group of WT and BERK animals, we employed MRI to examine the therapeutic efficacy of a single dose of the same SPF's by measuring cerebral blood flow (CBF) and sufficiency of cerebral oxygen delivery (B OLD MRI R esponse to a brief period of H yperO xia, BRHO) serially following treatment. Results: In NY1DD mice, EAF P5K6 Alb significantly attenuated hopoxia reoxygenation induced impairment of cremaster blood flow and associated vaso-occlusion, while EAF P3K6 Hb completely neutralized the experimentally induced sickle crisis. In BERK mice, both SPF's had comparable effects: the chronic state of vaso-occluison as observed in the cremaster muscle was eliminated completely by EAF P3K6-Hb. In MRI experiments in WT mice, both drug candidates resulted in increases in CBF, which resolved over 1 week. The increased CBF was accompanied by decreased BRHO consistent with a pseudo 'luxury perfusion' afforded by the accentuated delivery of oxygen. On the other hand, when BERK mice were treated with EAF P5K6 Alb or EAF P3K6 Hb, CBF trended lower, but with the Alb SPF, BRHO increased, and the Hb SPF, BHRO was unchanged, suggesting that the slightly reduced CBF led to increased O2 deficiency with the PEG-Alb, but not with the PEG-Hb. Conclusion : In WT mice, SPF's increase CBF in the brain where the facility to modify NO production is intact, resulting in over delivery of oxygen as confirmed by reductions in deoxy-Hb levels by BROH imaging, confirming supraperfusionary properties of the SPF's. In SCD animals, both SPF's attenuate muscle vaso-occlusion and restore blood flow. In addition, in experimentally induced sickle crisis (NY1DD), EAF P3K6 Hb maintained O2 level in the plasma and attenuate depolymerization of deoxyHb. In the severely anemic BERK mouse, EAF P5K6-Alb slightly attenuated CBF, likely due to reduced cerebral perfusion pressure (CPP), while O2 extraction increased suggesting that reduced CBF was detrimental to cerebral oxygen delivery. This effect was remediated when EAF P3K6-Hb is administered, which afforded additional oxygen to offset the losses due to reduced CBF. EAF P3K6 Hb led to slightly reduced CBF in NY1DD and BERK mice to levels approaching that obtained after administering EAF P5K6 Alb, but without inducing further oxygen debt. EAF P3K6 Hb appears to be the choice agent as this SPF facilitates increased delivery of O2 to hypoxic tissues thereby neutralizing painful crisis, and protects the brain from further ischemic insults. The influence of SCD on CBF by MRI is opposite to the decrease in blood flow observed in the systemic circulation. The infusion of SFA's increased flow in the systemic circulation, but reduced CBF in a disease dependent fashion. These divergent responses suggest the need for oxygen supplementation when developing SCD therapeutics. In particular, these studies suggest that high oxygen affinity PEG-Hb may have increased the therapeutic efficacy of this SPF by preventing the complete deoxygenation of HbS in the RBC. An antioxidant conjugated to the SFP, such as quercetin, could attenuate the hypoxia reoxygenation induced acute crisis and improve the efficacy of SCD therapeutics. Disclosures No relevant conflicts of interest to declare.

scholarly journals Assessment of Functional Shunting in Patients with Sickle Cell Disease

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 121-121
Author(s):  
Liza Afzali-Hashemi ◽  
Lena Vaclavu ◽  
John C Wood ◽  
Aart J Nederveen ◽  
Henk J.M.M. Mutsaerts ◽  
...  
Keyword(s):  
Blood Flow ◽  
Sickle Cell Disease ◽  
Oxygen Delivery ◽  
Research Funding ◽  
Negative Association ◽  
Cell Disease ◽  
Oxygen Utilization ◽  
Arteriovenous Shunting ◽  
Sagittal Sinus ◽  
The Brain

Abstract Introduction Silent cerebral infarcts (SCI) are common in patients with sickle cell disease (SCD) and thought to be caused by a mismatch between oxygen delivery and consumption. The mechanism underlying insufficient oxygen utilization is related to severity of anemia, and paradoxically, to the elevated cerebral blood flow (CBF) observed in SCD patients. CBF is elevated as a compensatory mechanism to maintain oxygen delivery, but high CBF levels can result in rapid transit of blood through the brain capillaries, limiting offloading of oxygen to the tissue; a process called arteriovenous shunting. One way to assess functional arteriovenous shunting is to use noncontrast perfusion MRI techniques in which we can assess the signal intensity of an endogenous blood tracer when it reaches the sagittal sinus. This venous signal (VS) reflects the amount of labeled blood that has not exchanged with the brain parenchyma. Under normal physiological conditions, the VS intensity will increase approximately proportionally with CBF as we expect only some of the water to exchange with tissue as it flows by. However, it is unknown whether functional shunting scales with CBF only, or whether other hemodynamic processes play a role in patients with SCD. We hypothesize that, under pathophysiological conditions such as in SCD patients, more labeled blood may pass unexchanged through the capillaries, which results in higher VS. In the present study, we investigated functional shunting by quantifying VS and assessed its association with hemodynamic, demographic and laboratory parameters in both pediatric and adult SCD patients, and controls. In addition, VS-CBF relationship was studied by further increasing CBF after a vasodilatory challenge. Methods We included 28 children (mean age 12.7 ± 2.3, 9 F) and 38 adults (mean age 32.1 ± 11.2, 14 F) with SCD (HbSS and HbS), and 10 healthy race-matched adult controls (mean age 36.4 ± 15.9, 4 F). For the CBF and VS measurements, pseudo-continuous arterial spin labelling (pCASL) data were acquired using 3T MRI. We segmented the ASL blood pool in the sagittal sinus to determine a common region of interest for each group. We used these images as masks to calculate average VS. Notably, for the comparison between children and adults the ratio between VS to gray matter CBF was used (VGR) instead of the VS, to take into account higher CBF in children. To get more insight into the oxygen utilization, oxygen extraction fraction (OEF) and cerebral metabolic rate of oxygen (CMRO 2) were calculated. In adult participants acetazolamide (ACZ) was used as a vasodilatory challenge. The hematologic laboratory parameters hemoglobin (Hb) and LDH were used as markers of anemia and hemolysis, respectively. Results VS as a marker of cerebral shunting was higher in both adult and pediatric patients with SCD as compared to controls (p<0.01) and increased after ACZ administration in all groups (p<0.01) (Fig. 1A). VS was significantly associated with CBF both before (R 2=0.59, p<0.001) and after ACZ administration (R 2=0.57, p<0.001) in all groups (Fig. 1B). To test the impact of demographics and hematological parameters on the presence of shunting, VS was corrected for CBF (VS CBF) and the residuals were used in multiple linear regression analysis against age, sex, blood flow velocity in the brain feeding arteries, participant groups, hydroxyurea, Hb and LDH. Cerebral shunting, reflected by VS CBF showed significant association with Hb (Fig. 1C). In addition, we added OEF, CMRO 2 and ACZ condition as additional parameters in adults and used a linear mixed model to accommodate the repeated measures dependencies. A negative association between the level of cerebral shunting (VS CBF) and CMRO 2 was found (β=-0.79, p<0.001) in all groups (Fig. 1C), and in adult patients with SCD, CMRO 2, Hb (β=-14.2 p<0.001) and LDH (β=0.13, p=0.002) were significant predictors of VS CBF. Conclusion Our results show that the VS in the sagittal sinus on ASL images can be used to assess functional arteriovenous shunting in the brain. Given its negative association with CMRO 2 in combination with the negative association with hemoglobin and positive correlation with LDH, this functional shunting seems to reflect pathophysiologic shunting related to higher disease severity. Future studies will focus on the relation between functional shunting and the prevalence of SCI, investigating its link to aberrant capillary oxygen exchange in SCD. Figure 1 Figure 1. Disclosures Vaclavu: Philips Healthcare: Research Funding. Biemond: GBT: Honoraria, Research Funding, Speakers Bureau; Novo Nordisk: Honoraria; Novartis: Honoraria, Research Funding, Speakers Bureau; Celgene: Honoraria; Sanquin: Research Funding.

scholarly journals A Meta-Analytic Comparison of Cerebral Blood Flow As Measured By MRI in Children with Sickle Cell Disease Versus Healthy Controls

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1391-1391
Author(s):  
Melanie E. Fields ◽  
Kristin Guilliams ◽  
Dustin Ragan ◽  
Lauren H. Yaeger ◽  
Monica L. Hulbert ◽  
...  
Keyword(s):  
Blood Flow ◽  
Cerebral Blood Flow ◽  
Sickle Cell Disease ◽  
Literature Search ◽  
Meta Analysis ◽  
Cerebral Oxygen ◽  
Healthy Controls ◽  
Cell Disease ◽  
Older Children ◽  
Study Heterogeneity

Abstract Stroke and its neurocognitive sequelae are devastating complications of sickle cell disease (SCD). Cerebral blood flow (CBF), a measure of tissue specific blood flow in the brain, is a key part of cerebral metabolism, and can provide insight into the pathophysiology of the neurologic complications of SCD. Previous work suggests that CBF is elevated in SCD patients compared to healthy controls (HC). Elevated CBF likely represents a compensatory mechanism to maintain cerebral oxygen delivery and metabolism in the setting of decreased arterial oxygen content secondary to chronic anemia. Previous studies are limited by small sample size and variations in imaging technique, making meta-analysis an alluring method for obtaining large numbers of participants to determine and compare population norms. Therefore, we performed a systematic meta-analysis to establish normal CBF values as measured by arterial spin labeled perfusion MR imaging (ASL) in pediatric HC and SCD populations, hypothesizing that CBF would be elevated in children with SCD when compared to a HC cohort. A systematic literature search was performed in 02/2014 for the HC cohort, and 06/2014 for the SCD cohort in Medline, Embase, and CINAHL. Whole brain cerebral blood flow (WB-CBF) measurements obtained by ASL in participants from 1 - 18 years (yr) were necessary for inclusion in both cohorts, and a diagnosis of hemoglobin SS or hemoglobin S-Beta thalassemia null for inclusion in the SCD cohort. ASL is the preferred imaging modality for CBF in pediatrics because ASL non-invasively measures CBF without the use of an exogenous contrast agent or ionizing radiation exposure. Data was excluded from the HC cohort if a medical disorder known to affect CBF was reported. Analysis included determination of mean WB-CBF values weighted by the inverse variance with 95% confidence intervals (CI) stratified by mean age, which was categorized as 1-9 yr and 9-18 yr secondary to known CBF changes throughout development. Statistical heterogeneity, or between-study variability, was assessed with Cochrane's Χ2 and I2. An I2 value greater than 30% was used as the threshold for analysis with a random-effects model. A funnel plot of the effect estimates versus the sample size for each individual study was used to assess for publication bias. Metaregression was used to determine the contribution of covariates to between-study heterogeneity in measurements of WB-CBF. All analyses were performed with STATA 13.1 (College Station, Texas) using the METAN software package. The literature search produced 155 articles for HC patients, and 12 articles for SCD patients; 8 publications met the inclusion and exclusion criteria for the HC cohort (89 children) and 4 publications for SCD cohort (92 children). Between the ages of 1 – 9 yr, mean WB-CBF in HC children was 67.6 ml/100 g/min (95% CI 37.9, 97.2), and mean WB-CBF in children with SCD was 117.0 ml/100 g/min (95%CI 48.7, 185.4). Between the ages of 10 – 18 yr, mean WB-CBF in HC children was 60.8 ml/100 g/min (95% CI 37.7, 83.9), and mean WB-CBF in children with SCD was 68.4 ml/100 g/min (95% CI 50.5, 86.3) (figures 1 and 2). The difference between the two cohorts is not statistically significant because of overlapping CI. The CochraneÕs X2 was significant with a p < 0.001 in all 4 analysis, with the I2 for each analysis ranging from 93.1% - 98.9%, indicating significant between-study heterogeneity. Mean age, ASL labeling technique, MRI vendor, and magnet strength, were not significant predictors of between-study heterogeneity within the HC cohort after controlling for multiple comparisons with the Bonferroni correction. Metaregression analysis of the SCD cohort could not be performed secondary to insufficient number of studies. Visual inspection of funnel plots did not provide evidence of publication bias. Mean WB-CBF as determined by meta-analysis in younger SCD patients is higher than mean WB-CBF in HC children, with little difference seen in older children. These findings suggest that younger children with SCD require increased compensation via elevated CBF to maintain cerebral oxygen metabolism when compared to older children. The significant between-study heterogeneity and wide CI illustrate a need to standardize the ASL technique for CBF measurement within pediatric HC and SCD populations, and the importance of including an age-matched control population when measuring CBF in SCD children. Figure 1. Figure 1. Figure 2. Figure 2. Disclosures No relevant conflicts of interest to declare.

scholarly journals Hairy Platelet-Derived Extracellular Vesicles Promote Lung Vaso-Occlusion in Sickle Cell Disease

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 958-958
Author(s):  
Prithu Sundd ◽  
Maritza Ann Jimenez ◽  
Margaret F. Bennewitz ◽  
Tomasz Brzoska ◽  
Egemen Tutuncuoglu ◽  
...  
Keyword(s):  
Blood Flow ◽  
Sickle Cell Disease ◽  
Extracellular Vesicles ◽  
Human Blood ◽  
Nlrp3 Inflammasome ◽  
Cell Disease ◽  
Caspase 1 ◽  
Neutrophil Aggregation

Abstract Background: Acute chest syndrome (ACS) is a type of acute lung injury and the leading cause of mortality in Sickle Cell Disease (SCD). Current treatments for ACS are primarily supportive, and there is a critical need for rescue therapies. ACS is often a sequela of acute systemic vaso-occlusive crisis and preceded by thrombocytopenia. However, the role of platelets in the pathogenesis of ACS remains largely unknown. Methods: We used our validated model of vaso-occlusive crisis in transgenic, humanized SCD mice, which is triggered by intravenous challenge with nanogram levels of the TLR4 ligand, lipopolysaccharide (LPS). Platelet-neutrophil aggregates and blood flow in the lung microcirculation was visualized in real time in vivo, using multi-photon-excitation microscopy of intact lung in live SCD mice. SCD or control human blood was perfused through microfluidic channels in vitro and neutrophil-platelet aggregation was visualized using fluorescence microscopy. Platelet derived extracellular vesicles were characterized using nanoparticle tracking and biochemical approaches. Results: We have made a novel finding that the arrest of blood flow and injury in the lung is secondary to blockade of pulmonary arterioles by platelet-neutrophil aggregates. Using in vitro microfluidic studies, we confirmed that platelet-neutrophil aggregation is higher in LPS-treated SCD patient blood compared with healthy controls, and this correlates with increased numbers of platelet-derived extracellular vesicles (EVs) that express IL-1β. Our studies also reveal that platelet-neutrophil aggregation in pulmonary arterioles of SCD mice is associated with an increase in peripheral blood levels of platelet-derived EVs containing IL-1β. Remarkably, inhibition of TLR4 or TLR4/NLRP3-inflammasome activated caspase-1, or inhibition of IL-1β signaling, attenuated release of platelet EVs and platelet-neutrophil aggregation in the lung arterioles of SCD mice in vivo and SCD human blood in vitro . Conclusions: TLR4 and NLRP3-inflammasome-mediated caspase-1 activation in platelets during vaso-occlusive crisis leads to release of IL-1β-containing EVs into the circulation. These circulating platelet EVs promote platelet-neutrophil aggregation in pulmonary arterioles, which results in arrest of blood flow in the lung, leading to ACS. Therapeutic inhibition of TLR4/NLRP3-caspase-1 signaling in platelets or IL-1β signaling is a potential therapy for ACS in SCD patients. Acknowledgments: This study was supported by 1R01HL128297-01 (P.S.) and VMI startup funds (P.S.). Disclosures No relevant conflicts of interest to declare.

scholarly journals Treatment of sickle cell disease by increasing oxygen affinity of hemoglobin

Blood ◽  
2021 ◽  
Author(s):  
Eric R Henry ◽  
Belhu Metaferia ◽  
Quan Li ◽  
Julia Harper ◽  
Robert B Best ◽  
...  
Keyword(s):  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Oxygen Delivery ◽  
Cell Damage ◽  
Oxygen Affinity ◽  
Red Cell ◽  
Cell Disease ◽  
Oxygen Dissociation ◽  
Dissociation Curves

The issue of treating sickle cell disease with drugs that increase hemoglobin oxygen affinity has come to the fore with the FDA approval in 2019 of voxelotor, the only anti-sickling drug approved since hydroxyurea in 1998. Voxelotor reduces sickling by increasing the concentration of the non-polymerizing, high oxygen affinity R (oxy) conformation of HbS. Treatment of sickle cell patients with voxelotor increases Hb levels and decreases indicators of hemolysis, but with no indication as yet that it reduces the frequency of pain episodes. Here we use the allosteric model of Monod, Wyman, and Changeux to simulate whole blood oxygen dissociation curves and red cell sickling in the absence and presence of voxelotor under the in vivo conditions of rapid oxygen pressure decreases. Our modeling agrees with experiments using a new robust assay, which shows the very large, expected decrease in sickling from the drug. The modeling indicates, however, that the increase in oxygen delivery from reduced sickling is largely offset by the increase in oxygen affinity. The net result is that the drug increases overall oxygen delivery only at the very lowest oxygen pressures. Reduction of sickling does, however, mitigate against red cell damage and explains the observed decrease in hemolysis. More importantly, our modeling of in vivo oxygen dissociation, sickling, and oxygen delivery suggests that drugs that increase fetal hemoglobin or decrease MCHC, should be more therapeutically effective than drugs that increase oxygen affinity.

scholarly journals Impairment of Cerebrovascular Hemodynamics in Patients With Severe and Milder Forms of Sickle Cell Disease

2021 ◽  
Vol 12 ◽  
Author(s):  
Liza Afzali-Hashemi ◽  
Koen P. A. Baas ◽  
Anouk Schrantee ◽  
Bram F. Coolen ◽  
Matthias J. P. van Osch ◽  
...  
Keyword(s):  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Oxygen Delivery ◽  
Vascular Reactivity ◽  
Fetal Hemoglobin ◽  
Cerebrovascular Reactivity ◽  
Healthy Controls ◽  
Cell Disease ◽  
The Brain ◽  
Cerebrovascular Hemodynamics

In patients with sickle cell disease (SCD), cerebral blood flow (CBF) is elevated to counteract anemia and maintain oxygen supply to the brain. This may exhaust the vasodilating capacity of the vessels, possibly increasing the risk of silent cerebral infarctions (SCI). To further investigate cerebrovascular hemodynamics in SCD patients, we assessed CBF, arterial transit time (ATT), cerebrovascular reactivity of CBF and ATT (CVRCBF and CVRATT) and oxygen delivery in patients with different forms of SCD and matched healthy controls. We analyzed data of 52 patients with severe SCD (HbSS and HbSβ0-thal), 20 patients with mild SCD (HbSC and HbSβ+-thal) and 10 healthy matched controls (HbAA and HbAS). Time-encoded arterial spin labeling (ASL) scans were performed before and after a vasodilatory challenge using acetazolamide (ACZ). To identify predictors of CBF and ATT after vasodilation, regression analyses were performed. Oxygen delivery was calculated and associated with hemoglobin and fetal hemoglobin (HbF) levels. At baseline, severe SCD patients showed significantly higher CBF and lower ATT compared to both the mild SCD patients and healthy controls. As CBFpostACZ was linearly related to CBFpreACZ, CVRCBF decreased with disease severity. CVRATT was also significantly affected in severe SCD patients compared to mild SCD patients and healthy controls. Considering all groups, women showed higher CBFpostACZ than men (p &lt; 0.01) independent of baseline CBF. Subsequently, post ACZ oxygen delivery was also higher in women (p &lt; 0.05). Baseline, but not post ACZ, GM oxygen delivery increased with HbF levels. Our data showed that baseline CBF and ATT and CVRCBF and CVRATT are most affected in severe SCD patients and to a lesser extent in patients with milder forms of SCD compared to healthy controls. Cerebrovascular vasoreactivity was mainly determined by baseline CBF, sex and HbF levels. The higher vascular reactivity observed in women could be related to their lower SCI prevalence, which remains an area of future work. Beneficial effects of HbF on oxygen delivery reflect changes in oxygen dissociation affinity from hemoglobin and were limited to baseline conditions suggesting that high HbF levels do not protect the brain upon a hemodynamic challenge, despite its positive effect on hemolysis.

scholarly journals MetAP2 Inhibition Modifies Hemoglobin S (HbS) to Delay Polymerization and Improve Blood Flow in Sickle Cell Disease

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2260-2260
Author(s):  
Melanie Demers ◽  
Sarah Sturtevant ◽  
Kevin Guertin ◽  
Dipti Gupta ◽  
Kunal Desai ◽  
...  
Keyword(s):  
Blood Flow ◽  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Whole Blood ◽  
Research Funding ◽  
Oxygen Affinity ◽  
Enzyme Kinetic ◽  
Erythroid Cells ◽  
Cell Disease

Dilution of HbS with non-sickling hemoglobin or hemoglobin with increased oxygen affinity is clinically beneficial in sickle cell disease. Aldehydes, including 5-HMF, tucaresol or GBT440, modify the N-terminus of HbS by reversible covalent imine formation generating modified forms of HbS that resist polymerization under low oxygen concentrations. In contrast to reversible imine formation by aldehydes, we hypothesize that stable modification of HbS will result from N-terminal retention of the initiator methionine (iMet) and subsequent N-terminal acetylation of the iMet (acetyl-iMet). MetAP2 is the methionine aminopeptidase able to cleave iMet from Val1 on α-globin and βS-globin as the unfolded N-terminal peptides emerge from the ribosome. Enzyme kinetic studies with pure MetAP2 and N-terminal octapeptides showed that βS-globin peptide is a 5-fold better substrate than α-globin peptide. Lentiviral shRNA knock-down of MetAP2 in differentiating erythroid HUDEP cells in vitro confirmed that α-globin is more extensively modified than βS-globin, consistent with the enzyme kinetic data. Selective MetAP2 inhibitors used to treat cultured human erythroid cells (HUDEP and PBMC derived CD34+) and Townes SCD mice in vivo confirmed that both α-globin and βS-globin domains of HbS are extensively modified by N-terminal iMet and acetyl-iMet. N-terminal retention of iMet and subsequent acetylation creates a mixture of modified HbS tetramers with combined modifications on both globins. Cation exchange chromatography separated nine different modified HbS variants from unmodified HbS as identified by LCMS. Purified samples of HbS modified by N-terminal iMet and acetyl-iMet had increased oxygen affinity as measured by decreased P50. Modified HbS containing the acetyl-iMet-βS-globin were found to have delayed polymerization under complete hypoxia (sodium metabisulfite triggered hypoxia in 1.8 M phosphate). Two modified HbS variants were further purified for X-ray crystallography studies (βS-globin / iMet-α-globin and acetyl-iMet-βS-globin / iMet-α-globin). Oxyhemoglobin structures of both modified HbS variants were in the R2-state previously described in structures of aldehyde modified HbS. This R2-state stabilizes the oxygenated R-state of HbS from conversion to the deoxygenated T-state that initiates HbS polymerization in sickle RBC. Treatment by selective irreversible covalent or reversible MetAP2 inhibitors resulted in high levels of HbS modification (>75%) in cultured erythroid cells (HUDEP and CD34+ cells). Dose dependent modification of HbS was observed in Townes sickle cell mouse blood RBC in vivo with total modification of HbS approaching 50%. In whole blood ex vivo studies, modification of HbS decreased RBC sickling under hypoxia (4% O2) and significantly increased the affinity of RBC for oxygen (decreased P50). Blood samples from MetAP2 inhibitor treated mice were analyzed for single-cell O2 saturation of the RBC and for the fractional flow velocity drop in whole blood rheology under decreasing partial oxygen pressures. In blood from vehicle treated sickle mice, a low-saturation peak of deoxy-HbS was observed in 7.8% O2, in contrast to blood from MetAP2 inhibitor-treated mice where the low-saturation peak was only observed in 6.4% O2. Similarly, in an assay of O2 dependent blood flow rheology, the half-maximum fractional velocity drop occurred at 5% O2 in control blood decreasing to 2% O2 in MetAP2 inhibitor treated blood. Our studies show that MetAP2 inhibition results in retention of iMet on βS-globin and α-globin and allows further acetylation of the retained iMet to create a mixture of N-terminal modified HbS tetramers. These modified HbS variants resist polymerization and RBC sickling under conditions of low O2 by delaying HbS polymerization and increasing O2 affinity. Our data suggests that MetAP2 may warrant further study as a potential therapeutic target for sickle cell disease. Disclosures Demers: Sanofi: Employment. Sturtevant:Sanofi: Employment. Guertin:Sanofi: Employment. Gupta:Sanofi: Employment. Desai:Sanofi: Employment. Vieira:Sanofi: Employment. Hicks:Sanofi: Employment. Ismail:Sanofi: Employment. Safo:Sanofi: Consultancy, Research Funding; Virginia Commonwealth University: Patents & Royalties. Wood:Sanofi: Consultancy, Research Funding. Higgins:Sanofi: Consultancy, Research Funding. Light:Sanofi: Employment.

scholarly journals Regulation of Brain Blood Flow and Oxygen Delivery in Elite Breath-Hold Divers

2014 ◽  
Vol 35 (1) ◽  
pp. 66-73 ◽  
Author(s):  
Christopher K Willie ◽  
Philip N Ainslie ◽  
Ivan Drvis ◽  
David B MacLeod ◽  
Anthony R Bain ◽  
...  
Keyword(s):  
Blood Flow ◽  
Oxygen Delivery ◽  
Blood Gases ◽  
Arterial Oxygen ◽  
Breath Hold ◽  
Cerebral Oxygen ◽  
Arterial Oxygen Content ◽  
Arterial Blood ◽  
End Tidal ◽  
The Brain

The roles of involuntary breathing movements (IBMs) and cerebral oxygen delivery in the tolerance to extreme hypoxemia displayed by elite breath-hold divers are unknown. Cerebral blood flow (CBF), arterial blood gases (ABGs), and cardiorespiratory metrics were measured during maximum dry apneas in elite breath-hold divers ( n=17). To isolate the effects of apnea and IBM from the concurrent changes on ABG, end-tidal forcing (‘clamp’) was then used to replicate an identical temporal pattern of decreasing arterial PO2 (PaO2) and increasing arterial PCO2 (PaCO2) while breathing. End-apnea PaO2 ranged from 23  to 37 mm Hg (30±7 mm Hg). Elevation in mean arterial pressure was greater during apnea than during clamp reaching +54±24% versus 34±26%, respectively; however, CBF increased similarly between apnea and clamp (93.6±28% and 83.4±38%, respectively). This latter observation indicates that during the overall apnea period IBM per se do not augment CBF and that the brain remains sufficiently protected against hypertension. Termination of apnea was not determined by reduced cerebral oxygen delivery; despite 40% to 50% reductions in arterial oxygen content, oxygen delivery was maintained by commensurately increased CBF.

scholarly journals A model for the regulation of cerebral oxygen delivery

1998 ◽  
Vol 85 (2) ◽  
pp. 554-564 ◽  
Author(s):  
Fahmeed Hyder ◽  
Robert G. Shulman ◽  
Douglas L. Rothman
Keyword(s):  
Blood Flow ◽  
Oxygen Delivery ◽  
Current Model ◽  
Oxygen Extraction Fraction ◽  
Physiological Data ◽  
Cerebral Oxygen ◽  
Oxygen Diffusivity ◽  
Wide Range ◽  
Capillary Bed

On the basis of the assumption that oxygen delivery across the endothelium is proportional to capillary plasma[Formula: see text], a model is presented that links cerebral metabolic rate of oxygen utilization ([Formula: see text]) to cerebral blood flow (CBF) through an effective diffusivity for oxygen (D) of the capillary bed. On the basis of in vivo evidence that the oxygen diffusivity properties of the capillary bed may be altered by changes in capillary[Formula: see text], hematocrit, and/or blood volume, the model allows changes in D with changes in CBF. Choice in the model of the appropriate ratio of Ω ≡ (ΔD/D)/(ΔCBF/CBF) determines the dependence of tissue oxygen delivery on perfusion. Buxton and Frank ( J. Cereb. Blood Flow. Metab. 17: 64–72, 1997) recently presented a limiting case of the present model in which Ω = 0. In contrast to the trends predicted by the model of Buxton and Frank, in the current model when Ω > 0, the proportionality between changes in CBF and[Formula: see text] becomes more linear, and similar degrees of proportionality can exist at different basal values of oxygen extraction fraction. The model is able to fit the observed proportionalities between CBF and[Formula: see text] for a large range of physiological data. Although the model does not validate any particular observed proportionality between CBF and[Formula: see text], generally values of ([Formula: see text]/[Formula: see text])/(ΔCBF/CBF) close to unity have been observed across ranges of graded anesthesia in rats and humans and for particular functional activations in humans. The model’s capacity to fit the wide range of data indicates that the oxygen diffusivity properties of the capillary bed, which can be modified in relation to perfusion, play an important role in regulating cerebral oxygen delivery in vivo.

Abstract T P370: Blood Transfusions in Pediatric Sickle Cell Disease Normalize Cerebral Blood Flow While Maintaining Constant Cerebral Oxygen Metabolism

Stroke ◽  
2015 ◽  
Vol 46 (suppl_1) ◽  
Author(s):  
Andria L Ford ◽  
Hongyu An ◽  
Kristin Guilliams ◽  
Melanie Fields ◽  
Cihat Eldeniz ◽  
...  
Keyword(s):  
Blood Flow ◽  
Oxygen Delivery ◽  
Stroke Risk ◽  
Oxygen Metabolism ◽  
Blood Transfusions ◽  
Compensatory Mechanism ◽  
Cerebral Oxygen ◽  
Cell Disease ◽  
Cerebral Oxygen Metabolism ◽  
Pediatric Sickle Cell Disease

Background: Chronic blood transfusions (Tx) reduces stroke risk in pediatric sickle cell disease (SCD). Cerebral blood flow (CBF) is elevated in SCD, likely representing a compensatory mechanism to maintain cerebral oxygen metabolism (CMRO2) in the setting of reduced arterial oxygen content (CaO2) from chronic anemia. When exhausted compensatory mechanisms are unable to meet oxygen demands, stroke ensues. We measured MR-derived CBF and oxygen extraction fraction (OEF) pre- and post-Tx, hypothesizing that Tx ‘resets’ the CBF baseline by increasing CaO2 via increased hemoglobin (Hb), while maintaining cerebral oxygen delivery and metabolism. Methods: SCD children on chronic Tx were enrolled in a prospective, observational MRI study. MR-CBF and MR-OEF were acquired before and 2 hours after exchange Tx. MR-CBF and MR-OEF were measured using pseudocontinuous arterial spin labelling and a novel asymmetric spin echo sequence, respectively. CaO2 =1.35 x [Hb] x SaO2. CMRO2 = CaO2 x CBF x OEF. Results: Two SCD children underwent MRI pre- and post-Tx (six more are anticipated prior to ISC). For subject #1 (18 yo F with overt stroke), mean global CBF was 128 and 98 ml/min/100g pre- and post-Tx, respectively, indicating a 24% CBF reduction. For subject #2 (6 yo F with elevated transcranial Doppler velocities), mean global CBF was 189 and 129 ml/min/100g pre- and post-Tx, respectively, a 32% CBF reduction (Fig). Both Hb and CaO2 were increased after Tx, resulting in unchanged oxygen delivery (CaO2 x CBF) post-Tx. Moreover, OEF and CMRO2 were not significantly different pre- and post-Tx, consistent with our hypothesis that CBF increases to maintain oxygen delivery. Conclusions: Elevated CBF is likely a compensatory mechanism to maintain constant oxygen delivery in SCD children who have chronically low CaO2. In our subjects, Tx improved CaO2, allowing CBF to normalize. This reduced hemodynamic stress likely contributes to the lower stroke risk in chronically transfused SCD children.

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