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The widespread adoption of high-calorie, high-fat, high-sucrose diets (HFHSD) has become a global health concern, particularly due to their association with cardiovascular diseases and metabolic disorders. These comorbidities increase susceptibility to severe outcomes from viral infections and trauma, with trauma-related incidents significantly contributing to global mortality rates. This context underscores the critical need for a reliable blood supply. Recent research has focused on high molecular weight (MW) polymerized human hemoglobin (PolyhHb) as a promising alternative to red blood cells (RBCs), showing encouraging outcomes in previous studies. Given the overlap of metabolic disorders and trauma-related health issues, it is crucial to assess the potential toxicity of PolyhHb transfusions, particularly in models that represent these vulnerable populations. This study evaluated the effects of PolyhHb exchange transfusion in guinea pigs that had developed metabolic disorders due to a 12-week HFHSD regimen. The guinea pigs, underwent a 20 % blood volume exchange transfusion with either PolyhHb or the lower molecular weight polymerized bovine hemoglobin, Oxyglobin. Results revealed that both PolyhHb and Oxyglobin transfusions led to liver damage, with a more pronounced effect observed in HFHSD-fed animals. Additionally, markers of cardiac dysfunction indicated signs of cardiac injury in both the HFHSD and normal diet groups following the Oxyglobin transfusion. This study highlights how pre-existing metabolic disorders can exacerbate the potential side effects of hemoglobin-based oxygen carriers (HBOCs). Importantly, the newer generation of high MW PolyhHb showed lower cardiac toxicity compared to the earlier generation low MW PolyhHb, known as Oxyglobin, even in models with pre-existing endothelial and metabolic challenges., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Pedro Cabrales reports financial support was provided by US Department of Defense. Pedro Cabrales reports financial support was provided by National Institutes of Health. Andre F. Palmer is an inventor on U.S. patent application PCT/US2022/041743. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Background: Preserved Ratio Impaired Spirometry (PRISm) is defined as FEV 1 <80% predicted, FEV 1 /FVC ≥0.70. PRISm is associated with respiratory symptoms and co-morbidities. Our objective was to discover novel genetic signals for PRISm and see if they provide insight into the pathogenesis of PRISm and associated co-morbidities., Methods: We undertook a genome-wide association study (GWAS) of PRISm in UK Biobank participants (Stage 1), and selected SNPs reaching genome-wide significance for replication in 13 cohorts (Stage 2). A combined meta-analysis of Stage 1 and Stage 2 was done to determine top SNPs. We used cross-trait Linkage Disequilibrium score regression to estimate genome-wide genetic correlation between PRISM and pulmonary and extra-pulmonary traits. Phenome-wide association studies of top SNPs was performed., Results: 22 signals reached significance in the joint meta-analysis, including four signals novel for lung function. A strong genome-wide genetic correlation (r g ) between PRISm and spirometric COPD (r g =0.62, p-value <0.001) was observed, and genetic correlation with type II diabetes (r g =0.12, p-value 0.007). PheWAS showed that 18 of 22 signals were associated with diabetic traits and 7 with blood pressure traits., Discussion: This is the first GWAS to successfully identify SNPs associated with PRISm. Four of the signals; rs7652391 (nearest gene MECOM ), rs9431040 ( HLX ), rs62018863 ( TMEM114 ) and rs185937162 ( HLA-B ) have not been described in association with lung function before, demonstrating the utility of using different lung function phenotypes in GWAS. Genetic factors associated with PRISm are strongly correlated with risk of both other lung diseases and extra-pulmonary co-morbidity., (Copyright ©The authors 2023.)

Cerebral malaria (CM) is a severe manifestation of malaria that commonly occurs in children and is hallmarked by neurologic symptoms and significant Plasmodium falciparum parasitemia. It is currently hypothesized that cerebral hypoperfusion from impaired microvascular oxygen transport secondary to parasitic occlusion of the microvasculature is responsible for cerebral ischemia and thus disease severity. Animal models to study CM, are known as experimental cerebral malaria (ECM), and include the C57BL/6J infected with Plasmodium berghei ANKA (PbA), which is ECM-susceptible, and BALB/c infected with PbA, which is ECM-resistant. Here we sought to investigate whether changes in oxygen (O 2 ) delivery, O 2 flux, and O 2 utilization are altered in both these models of ECM using phosphorescence quenching microscopy (PQM) and direct measurement of microvascular hemodynamics using the cranial window preparation. Animal groups used for investigation consisted of ECM-susceptible C57BL/6 (Infected, n = 14) and ECM-resistant BALB/c (Infected, n = 9) mice. Uninfected C57BL/6 (n = 6) and BALB/c (n = 6) mice were included as uninfected controls. Control animals were manipulated in the exact same way as the infected mice (except for the infection itself). C57BL/6 ECM animals at day 6 of infection were divided into two cohorts: Early-stage ECM, presenting mild to moderate drops in body temperature (>34 < 36 °C) and Late-stage ECM, showing marked drops in body temperature (<33 °C). Data taken from new experiments conducted with these animal models were analyzed using a general linear mixed model. We constructed three general linear mixed models, one for total O 2 content, another for total O 2 delivery, and the third for total O 2 content as a function of convective flow. We found that in both the ECM-susceptible C57BL/6J model and ECM-resistant BALB/c model of CM, convective and diffusive O 2 flux along with pial hemodynamics are impaired. We further show that concomitant changes in p50 (oxygen partial pressure for 50% hemoglobin saturation), only 5 mmHg in the case of late-stage CM C57BL/6J mice, and O 2 diffusion result in insufficient O 2 transport by the pial microcirculation, and that both these changes are required for late-stage disease. In summary, we found impaired O 2 transport and O 2 affinity in late-stage ECM, but only the former in either early-stage ECM and ECM-resistant strains., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Thyroid hormones play a critical role in regulation of multiple physiological functions and thyroid dysfunction is associated with substantial morbidity. Here, we use electronic health records to undertake a genome-wide association study of thyroid-stimulating hormone (TSH) levels, with a total sample size of 247,107. We identify 158 novel genetic associations, more than doubling the number of known associations with TSH, and implicate 112 putative causal genes, of which 76 are not previously implicated. A polygenic score for TSH is associated with TSH levels in African, South Asian, East Asian, Middle Eastern and admixed American ancestries, and associated with hypothyroidism and other thyroid disease in South Asians. In Europeans, the TSH polygenic score is associated with thyroid disease, including thyroid cancer and age-of-onset of hypothyroidism and hyperthyroidism. We develop pathway-specific genetic risk scores for TSH levels and use these in phenome-wide association studies to identify potential consequences of pathway perturbation. Together, these findings demonstrate the potential utility of genetic associations to inform future therapeutics and risk prediction for thyroid diseases., (© 2023. Springer Nature Limited.)

The pathophysiologic mechanism of sickle cell disease (SCD) involves polymerization of deoxygenated haemoglobin S (HbS), leading to red blood cell (RBC) sickling, decreased RBC deformability, microvascular obstruction, haemolysis, anaemia and downstream clinical complications. Pharmacological increase in the concentration of oxygenated HbS in RBCs has been shown to be a novel approach to inhibit HbS polymerization and reduce RBC sickling and haemolysis. We report that GBT021601, a small molecule that increases HbS-oxygen affinity, inhibits HbS polymerization and prevents RBC sickling in blood from patients with SCD. Moreover, in a murine model of SCD (SS mice), GBT021601 reduces RBC sickling, improves RBC deformability, prolongs RBC half-life and restores haemoglobin levels to the normal range, while improving oxygen delivery and increasing tolerance to severe hypoxia. Notably, oral dosing of GBT021601 in animals results in higher levels of Hb occupancy than voxelotor and suggests the feasibility of once-daily dosing in humans. In summary, GBT021601 improves RBC health and normalizes haemoglobin in SS mice, suggesting that it may be useful for the treatment of SCD. These data are being used as a foundation for clinical research and development of GBT021601., (© 2023 The Authors. British Journal of Haematology published by British Society for Haematology and John Wiley & Sons Ltd.)

Direct measurement of cardiac pressure-volume (PV) relationships is the gold standard for assessment of ventricular hemodynamics, but few innovations have been made to "multi-beat" PV analysis beyond traditional signal processing. The Prony method solves the signal recovery problem with a series of dampened exponentials or sinusoids. It achieves this by extracting the amplitude, frequency, dampening, and phase of each component. Since its inception, application of the Prony method to biologic and medical signal has demonstrated a relative degree of success, as a series of dampened complex sinusoids easily generalizes to multifaceted physiological processes. In cardiovascular physiology, the Prony analysis has been used to determine fatal arrythmia from electrocardiogram signals. However, application of the Prony method to simple left ventricular function based on pressure and volume analysis is absent. We have developed a new pipeline for analysis of pressure volume signals recorded from the left ventricle. We propose fitting pressure-volume data from cardiac catheterization to the Prony method for pole extraction and quantification of the transfer function. We implemented the Prony algorithm using open-source Python packages and analyzed the pressure and volume signals before and after severe hemorrhagic shock, and after resuscitation with stored blood. Each animal (n = 6 per group) underwent a 50% hemorrhage to induce hypovolemic shock, which was maintained for 30 min, and resuscitated with 3-week-old stored RBCs until 90% baseline blood pressure was achieved. Pressure-volume catheterization data used for Prony analysis were 1 s in length, sampled at 1000 Hz, and acquired at the time of hypovolemic shock, 15 and 30 min after induction of hypovolemic shock, and 10, 30, and 60 min after volume resuscitation. We next assessed the complex poles from both pressure and volume waveforms. To quantify deviation from the unit circle, which represents deviation from a Fourier series, we counted the number of poles at least 0.2 radial units away from it. We found a significant decrease in the number of poles after shock (p = 0.0072 vs. baseline) and after resuscitation (p = 0.0091 vs. baseline). No differences were observed in this metric pre and post volume resuscitation (p = 0.2956). We next found a composite transfer function using the Prony fits between the pressure and volume waveforms and found differences in both the magnitude and phase Bode plots at baseline, during shock, and after resuscitation. In summary, our implementation of the Prony analysis shows meaningful physiologic differences after shock and resuscitation and allows for future applications to broader physiological and pathophysiological conditions., Competing Interests: Declaration of Competing Interest All authors must disclose any financial and personal relationships with other people or organisations that could inappropriately influence (bias) their work. Examples of potential conflicts of interest include employment, consultancies, stock ownership, honoraria, paid expert testimony, patent applications/registrations, and grants or other funding., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Rationale: Eosinophils are associated with airway inflammation in respiratory disease. Eosinophil production and survival is controlled partly by interleukin-5: anti-interleukin-5 agents reduce asthma and response correlates with baseline eosinophil counts. However, whether raised eosinophils are causally related to chronic obstructive pulmonary disease (COPD) and other respiratory phenotypes is not well understood., Objectives: We investigated causality between eosinophils and: lung function, acute exacerbations of COPD, asthma-COPD overlap (ACO), moderate-to-severe asthma and respiratory infections., Methods: We performed Mendelian randomisation (MR) using 151 variants from genome-wide association studies of blood eosinophils in UK Biobank/INTERVAL, and respiratory traits in UK Biobank/SpiroMeta, using methods relying on different assumptions for validity. We performed multivariable analyses using eight cell types where there was possible evidence of causation by eosinophils., Measurements and Main Results: Causal estimates derived from individual variants were highly heterogeneous, which may arise from pleiotropy. The average effect of raising eosinophils was to increase risk of ACO (weighted median OR per SD eosinophils, 1.44 (95%CI 1.19 to 1.74)), and moderate-severe asthma (weighted median OR 1.50 (95%CI 1.23 to 1.83)), and to reduce forced expiratory volume in 1 s (FEV 1 )/forced vital capacity (FVC) and FEV 1 (weighted median estimator, SD FEV 1 /FVC: -0.054 (95% CI -0.078 to -0.029), effect only prominent in individuals with asthma)., Conclusions: Broad consistency across MR methods may suggest causation by eosinophils (although of uncertain magnitude), yet heterogeneity necessitates caution: other important mechanisms may be responsible for the impairment of respiratory health by these eosinophil-raising variants. These results could suggest that anti-IL5 agents (designed to lower eosinophils) may be valuable in treating other respiratory conditions, including people with overlapping features of asthma and COPD., Competing Interests: Competing interests: MDT and LVW receive funding from GSK for collaborative research projects outside of the submitted work. IH has funded research collaborations with GSK, Boehringer Ingelheim and Orion., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY. Published by BMJ.)

Lung-function impairment underlies chronic obstructive pulmonary disease (COPD) and predicts mortality. In the largest multi-ancestry genome-wide association meta-analysis of lung function to date, comprising 580,869 participants, we identified 1,020 independent association signals implicating 559 genes supported by ≥2 criteria from a systematic variant-to-gene mapping framework. These genes were enriched in 29 pathways. Individual variants showed heterogeneity across ancestries, age and smoking groups, and collectively as a genetic risk score showed strong association with COPD across ancestry groups. We undertook phenome-wide association studies for selected associated variants as well as trait and pathway-specific genetic risk scores to infer possible consequences of intervening in pathways underlying lung function. We highlight new putative causal variants, genes, proteins and pathways, including those targeted by existing drugs. These findings bring us closer to understanding the mechanisms underlying lung function and COPD, and should inform functional genomics experiments and potentially future COPD therapies., (© 2023. The Author(s).)

For the past thirty years, hemoglobin-based oxygen carriers (HBOCs) have been under development as a red blood cell substitute. Side-effects such as vasoconstriction, oxidative injury, and cardiac toxicity have prevented clinical approval of HBOCs. Recently, high molecular weight (MW) polymerized human hemoglobin (PolyhHb) has shown positive results in rats. Studies have demonstrated that high MW PolyhHb increased O 2 delivery, with minimal effects on blood pressure, without vasoconstriction, and devoid of toxicity. In this study, we used guinea pigs to evaluate the efficacy and safety of high MW PolyhHb, since like humans guinea pigs cannot produce endogenous ascorbic acid, which limits the capacity of both species to deal with oxidative stress. Hence, this study evaluated the efficacy and safety of resuscitation from severe hemorrhagic shock with high MW PolyhHb, fresh blood, and blood stored for 2 weeks. Animals were randomly assigned to each experimental group, and hemorrhage was induced by the withdrawal of 40% of the blood volume (BV, estimated as 7.5% of body weight) from the carotid artery catheter. Hypovolemic shock was maintained for 50 min. Resuscitation was implemented by infusing 25% of the animal's BV with the different treatments. Hemodynamics, blood gases, total hemoglobin, and lactate were not different before hemorrhage and during shock between groups. The hematocrit was lower for the PolyhHb group compared to the fresh and stored blood groups after resuscitation. Resuscitation with stored blood had lower blood pressure compared to fresh blood at 2 h. There was no difference in mean arterial pressure between groups at 24 h. Resuscitation with PolyhHb was not different from fresh blood for most parameters. Resuscitation with PolyhHb did not show any remarkable change in liver injury, inflammation, or cardiac damage. Resuscitation with stored blood showed changes in liver function and inflammation, but no kidney injury or systemic inflammation. Resuscitation with stored blood after 24 h displayed sympathetic hyper-activation and signs of cardiac injury. These results suggest that PolyhHb is an effective resuscitation alternative to blood. The decreased toxicities in terms of cardiac injury markers, vital organ function, and inflammation following PolyhHb resuscitation in guinea pigs indicate a favorable safety profile. These results are promising and support future studies with this new generation of PolyhHb as alternative to blood when blood is unavailable., (© 2022. The Author(s).)

Infection with COVID-19 has resulted in over 276,000 deaths in the United States and over 1.5 million deaths globally, with upwards of 15% of patients requiring hospitalization. Severe COVID-19 infection is, in essence, a microvascular disease. This contention has been emphasized throughout the course of the pandemic, particularly due to the clinical manifestation of severe infection. In fact, it has been hypothesized and shown in particular instances that microvascular function is a significant prognosticator for morbidity and mortality. Initially thought to be isolated to the pulmonary system and resulting in ARDS, patients with COVID-19 have been observed to have acute cardiac, renal, and thrombolytic complications. Therefore, severe COVID-19 is a vascular disease that has systemic implications. The objective of this review is to provide a mechanistic background for the microvascular nature of severe COVID-19 infection, with a particular emphasis on dysfunction of the endothelial glycocalyx and nitric oxide mediated pathogenesis., Competing Interests: Declaration of Competing Interest There are no potential conflict of interest, real or perceived, by the authors., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Electronic health record (EHR) databases provide rich, longitudinal data on interactions with healthcare providers and can be used to advance research into respiratory conditions. However, since these data are primarily collected to support health care delivery, clinical coding can be inconsistent, resulting in inherent challenges in using these data for research purposes., Methods: We systematically searched existing international literature and UK code repositories to find respiratory disease codelists for asthma from January 2018, and chronic obstructive pulmonary disease and respiratory tract infections from January 2020, based on prior searches. Medline searches using key terms provided in article lists. Full-text articles, supplementary files, and reference lists were examined for codelists, and codelists repositories were searched. A reproducible methodology for codelists creation was developed with recommended lists for each disease created based on multidisciplinary expert opinion and previously published literature., Results: Medline searches returned 1126 asthma articles, 70 COPD articles, and 90 respiratory infection articles, with 3%, 22% and 5% including codelists, respectively. Repository searching returned 12 asthma, 23 COPD, and 64 respiratory infection codelists. We have systematically compiled respiratory disease codelists and from these derived recommended lists for use by researchers to find the most up-to-date and relevant respiratory disease codelists that can be tailored to individual research questions., Conclusion: Few published papers include codelists, and where published diverse codelists were used, even when answering similar research questions. Whilst some advances have been made, greater consistency and transparency across studies using routine data to study respiratory diseases are needed., Competing Interests: CM, HW, MM, LD, AM, CI, MA, EV, EOR, ATW, PWS have nothing to declare. JKQ reports grants from AUK-BLF, The Health Foundation, MRC, grants and personal fees from AZ, BI, GSK, Bayer, grants from Chiesi, outside the submitted work. AS reports grants from AUK-BLF and HDR UK., (© 2022 MacRae et al.)

Ischemia-reperfusion injury increased vascular permeability, resulting in fluid extravasation from the intravascular compartment into the tissue space. Fluid and small protein extravasation lead to increased interstitial fluid pressure and capillary collapse, impairing capillary exchange. Polymerized human serum albumin (PolyHSA) has an increased molecular weight (MW) compared with unpolymerized human serum albumin (HSA) and can improve intravascular fluid retention and recovery from ischemia-reperfusion injury. To test the hypothesis that polymerization of HSA can improve recovery from ischemia-reperfusion injury, we studied how exchange transfusion of 20% of the blood volume with HSA or PolyHSA immediately before reperfusion can affect local ischemic tissue microhemodynamics, vascular integrity, and tissue viability in a hamster dorsal window chamber model. Microvascular flow and functional capillary density were maintained in animals exchanged with PolyHSA compared with HSA. Likewise, exchange transfusion with PolyHSA preserved vascular permeability measured with extravasation of fluorescently labeled dextran. The intravascular retention time of the exchanged PolyHSA was significantly longer compared with the intravascular retention time of HSA. Lastly, the viability of tissue subjected to ischemia-reperfusion injury increased in animals exchanged with PolyHSA compared with HSA. Therefore maintenance of microvascular perfusion, improvement in vascular integrity, and reduction in tissue damage resulting from reperfusion with PolyHSA suggest that PolyHSA is a promising fluid therapy to improve outcomes of ischemia-reperfusion injury. NEW & NOTEWORTHY Polymerized human serum albumin reduced reperfusion injury and preservers microvascular hemodynamics. Polymerized human serum albumin reduces fluid extravasation and prevents fluid extravasation. Consequently, the tissue viability of ischemic tissue is preserved by polymerized human serum.

Therapeutic agents that increase the Hb affinity for oxygen (O 2 ) could, in theory, lead to decreased O 2 release from Hb and impose a hypoxic risk to tissues. In this study, GBT1118, an allosteric modifier of Hb affinity for O 2 , was used to assess the impact of increasing Hb affinity for O 2 on brain tissue oxygenation, blood pressure, heart rate, O 2 delivery, and tolerance to hypoxia in Townes transgenic sickle cell disease (SCD) mice. Brain oxygenation and O 2 delivery were studied during normoxia and severe hypoxic challenges. Chronic treatment with GBT1118 increased Hb affinity for O 2 , reducing the Po 2 for 50% HbO 2 saturation (P50) in SCD mice from 31 mmHg to 18 mmHg. This treatment significantly reduced anemia, increasing hematocrit by 33%, improved cardiac output (CO), and O 2 delivery and extraction. Chronically increasing Hb affinity for O 2 with GBT1118 preserved cortical O 2 tension during normoxia, improved cortical O 2 tension during hypoxia, and increased tolerance to severe hypoxia in SCD mice. Independent of hematological changes induced by chronic treatment, a single dose of GBT1118 significantly improved tolerance to hypoxia, highlighting the benefits of increasing Hb affinity for O 2 and consequently reducing sickling of RBCs in blood during hypoxia in SCD. NEW & NOTEWORTHY Chronic pharmacologically increased hemoglobin affinity for oxygen in sickle cell disease mice alleviated hematological consequences of sickle cell disease, increasing RBC half-life, hematocrit, and hemoglobin concentration, while also decreasing reticulocyte count. Additionally, chronically increased hemoglobin affinity for oxygen significantly improved survival as well as cortical tissue oxygenation in sickle cell disease mice during hypoxia, suggesting that oxygen delivery and utilization is improved by increased hemoglobin affinity for oxygen.

Treatment-resistant depression (TRD) is a major contributor to the disability caused by major depressive disorder (MDD). Primary care electronic health records provide an easily accessible approach to investigate TRD clinical and genetic characteristics. MDD defined from primary care records in UK Biobank (UKB) and EXCEED studies was compared with other measures of depression and tested for association with MDD polygenic risk score (PRS). Using prescribing records, TRD was defined from at least two switches between antidepressant drugs, each prescribed for at least 6 weeks. Clinical-demographic characteristics, SNP-based heritability (h 2 SNP ) and genetic overlap with psychiatric and non-psychiatric traits were compared in TRD and non-TRD MDD cases. In 230,096 and 8926 UKB and EXCEED participants with primary care data, respectively, the prevalence of MDD was 8.7% and 14.2%, of which 13.2% and 13.5% was TRD, respectively. In both cohorts, MDD defined from primary care records was strongly associated with MDD PRS, and in UKB it showed overlap of 71-88% with other MDD definitions. In UKB, TRD vs healthy controls and non-TRD vs healthy controls h 2 SNP was comparable (0.25 [SE = 0.04] and 0.19 [SE = 0.02], respectively). TRD vs non-TRD was positively associated with the PRS of attention deficit hyperactivity disorder, with lower socio-economic status, obesity, higher neuroticism and other unfavourable clinical characteristics. This study demonstrated that MDD and TRD can be reliably defined using primary care records and provides the first large scale population assessment of the genetic, clinical and demographic characteristics of TRD., (© 2021. The Author(s).)

Background: Hemolysis releases toxic cell-free hemoglobin (Hb), heme, and iron, which overwhelm their natural scavenging mechanisms during acute or chronic hemolytic conditions. This study describes a novel strategy to purify a protein cocktail containing a comprehensive set of scavenger proteins for potential treatment of hemolysis byproducts., Study Design and Methods: Tangential flow filtration was used to purify a protein cocktail from Human Cohn Fraction IV (FIV). A series of in vitro assays were performed to characterize composition and biocompatibility. The in vivo potential for hemolysis byproduct mitigation was assessed in a hamster exchange transfusion model using mechanically hemolyzed blood plasma mixed with the protein cocktail or a control colloid (dextran 70 kDa)., Results: A basis of 500 g of FIV yielded 62 ± 9 g of a protein mixture at 170 g/L, which bound to approximately 0.6 mM Hb, 1.2 mM heme, and 1.2 mM iron. This protein cocktail was shown to be biocompatible in vitro with red blood cells and platelets and exhibits nonlinear concentration dependence with respect to viscosity and colloidal osmotic pressure. In vivo assessment of the protein cocktail demonstrated higher iron transport to the liver and spleen and less to the kidney and heart with significantly reduced renal and cardiac inflammation markers and lower kidney and hepatic damage compared to a control colloid., Discussion: Taken together, this study provides an effective method for large-scale production of a protein cocktail suitable for comprehensive reduction of hemolysis-induced toxicity., (© 2021 AABB.)

Fluid resuscitation following severe inflammation-induced hypoperfusion is critical for the restoration of hemodynamics and the prevention of multiorgan dysfunction syndrome during septic shock. Fluid resuscitation with commercially available crystalloid and colloid solutions only provides transient benefits, followed by fluid extravasation and tissue edema through the inflamed endothelium. The increased molecular weight (M.W.) of polymerized human serum albumin (PolyHSA) can limit fluid extravasation, leading to restoration of hemodynamics. In this prospective study, we evaluated how fluid resuscitation with PolyHSA impacts the hemodynamic and immune response in a lipopolysaccharide (LPS) induced endotoxemia mouse model. Additionally, we evaluated fluid resuscitation with PolyHSA in a model of polymicrobial sepsis induced by cecal ligation and puncture (CLP). Resuscitation with PolyHSA attenuated the immune response and improved the maintenance of systemic hemodynamics and restoration of microcirculatory hemodynamics. This decrease in inflammatory immune response and maintenance of vascular wall shear stress likely contributes to the maintenance of vascular integrity following fluid resuscitation with PolyHSA. The sustained restoration of perfusion, decrease in pro-inflammatory immune response, and improved vascular integrity that results from the high M.W. of PolyHSA indicates that a PolyHSA based solution is a potential resuscitation fluid for endotoxic and septic shock.

Easy access to high-calorie and fat-dense fast food has resulted in unhealthy dietary and lifestyle changes worldwide, which affects both developed and developing economies. This predisposes populations to a considerable number of metabolic and inflammatory conditions, such as diabetes, nonalcoholic fatty liver disease (NAFLD), and cardiovascular disease (CVD). Guinea pigs have been proposed as a model to study high-fat diet-induced metabolic disease due to their similar antioxidant metabolism and lipid profile to humans, and their susceptibility to atherosclerosis and endothelial disease. This study aims to evaluate cardiovascular and metabolic disorders induced by high-fat high-sucrose diet (HFHSD) in guinea pigs. Two to three-week-old male guinea pigs were fed a normal diet (ND) or HFHSD for 12 wk. Guinea pigs fed a HFHSD developed glucose intolerance, dyslipidemia, and liver, cardiac, and kidney damage. However, hypertension, dysautonomia, endothelial disease, and obesity were absent in these HFHSD guinea pigs. Taken together, these results show that guinea pigs fed a HFHSD are a nonobese model of metabolic disorders, resulting in important cardiac damage. Moreover, our findings suggest that NAFLD may be an important risk factor for diet-induced CVD. NEW & NOTEWORTHY In this study, we show a new animal model for diet-induced disease metabolic disorders without obesity in guinea pigs. Moreover, results suggest a strong relation between liver disease and increased cardiovascular risks.

Hemorrhagic shock (HS) is a severe life-threatening condition characterized by loss of blood volume and a lack of oxygen (O 2 ) delivery to tissues. The objective of this study was to examine the impact of manipulating Starling forces in the microcirculation during HS to increase microvascular perfusion without restoring blood volume or increasing O 2 carrying capacity. To decrease interstitial tissue pressure, we developed a non-contact system to locally apply negative pressure and manipulate the pressure balance in capillaries, while allowing for visualization of the microcirculation. Golden Syrian hamsters were instrumented with dorsal window chambers and subjected to a controlled hemorrhaged of 50% of the animal's blood volume without any fluid resuscitation. A negative pressure chamber was attached to the dorsal window chamber and a constant negative pressure was applied. Hemodynamic parameters (including microvascular diameter, blood flow, and functional capillary density [FCD]) were measured before and during the four hours following the hemorrhage, with and without applied negative pressure. Blood flow significantly increased in arterioles during negative pressure. The increase in flow through arterioles also improved microvascular perfusion as reflected by increased FCD. These results indicate that negative pressure increases flow in the microcirculation when fluid resuscitation is not available, thus restoring blood flow, oxygen delivery, and preventing the accumulation of metabolic waste. Applying negative pressure might allow for control of microvascular blood flow and oxygen delivery to specific tissue areas., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Microvascular fluid exchange is primarily dependent on Starling forces and both the active and passive myogenic response of arterioles and post-capillary venules. Arterioles are classically considered resistance vessels, while venules are considered capacitance vessels with high distensibility and low tonic sympathetic stimulation at rest. However, few studies have investigated the effects of modulating interstitial hydrostatic pressure, particularly in the context of hemorrhagic shock. The objective of this study was to investigate the mechanics of arterioles and functional capillary density (FCD) during application of negative tissue interstitial pressure after 40% total blood volume hemorrhagic shock. In this study, we characterized systemic and microcirculatory hemodynamic parameters, including FCD, in hamsters instrumented with a dorsal window chamber and a custom-designed negative pressure application device via intravital microscopy. In large arterioles, application of negative pressure after hemorrhagic shock resulted in a 13 ± 11% decrease in flow compared with only a 7 ± 9% decrease in flow after hemorrhagic shock alone after 90 minutes. In post-capillary venules, however, application of negative pressure after hemorrhagic shock resulted in a 31 ± 4% decrease in flow compared with only an 8 ± 5% decrease in flow after hemorrhagic shock alone after 90 minutes. Normalized FCD was observed to significantly improve after application of negative pressure after hemorrhagic shock (0.66 ± 0.02) compared to hemorrhagic shock without application of negative pressure (0.50 ± 0.04). Our study demonstrates that application of negative pressure acutely improves FCD during hemorrhagic shock, though it does not normalize FCD. These results suggest that by increasing the hydrostatic pressure gradient between the microvasculature and interstitium, microvascular perfusion can be transiently restored in the absence of volume resuscitation. This study has significant clinical implications, particularly in negative pressure wound therapy, and offers an alternative mechanism to improve microvascular perfusion during hypovolemic shock., (© 2021 The Authors. Physiological Reports published by Wiley Periodicals LLC on behalf of The Physiological Society and the American Physiological Society.)

Traumatic brain injury (TBI) is often accompanied by hemorrhage, and treatment of hemorrhagic shock (HS) after TBI is particularly challenging because the two therapeutic treatment strategies for TBI and HS often conflict. Ischemia/reperfusion injury from HS resuscitation can be exaggerated by TBI-induced loss of autoregulation. In HS resuscitation, the goal is to restore lost blood volume, while in the treatment of TBI the priority is focused on maintenance of adequate cerebral perfusion pressure and avoidance of secondary bleeding. In this study, we investigate the responses to resuscitation from severe HS after TBI in rats, using fresh blood, polymerized human hemoglobin (PolyhHb), and lactated Ringer's (LR). Rats were subjected to TBI by pneumatic controlled cortical impact. Shortly after TBI, HS was induced by blood withdrawal to reduce mean arterial pressure (MAP) to 35-40 mmHg for 90 min before resuscitation. Resuscitation fluids were delivered to restore MAP to ~ 65 mmHg and animals were monitored for 120 min. Increased systolic blood pressure variability (SBPV) confirmed TBI-induced loss of autoregulation. MAP after resuscitation was significantly higher in the blood and PolyhHb groups compared to the LR group. Furthermore, blood and PolyhHb restored diastolic pressure, while this remained depressed for the LR group, indicating a loss of vascular tone. Lactate increased in all groups during HS, and only returned to baseline level in the blood reperfused group. The PolyhHb group possessed lower SBPV compared to LR and blood groups. Finally, sympathetic nervous system (SNS) modulation was higher for the LR group and lower for the PolyhHb group compared to the blood group after reperfusion. In conclusion, our results suggest that PolyhHb could be an alternative to blood for resuscitation from HS after TBI when blood is not available, assuming additional testing demonstrate similar favorable results. PolyhHb restored hemodynamics and oxygen delivery, without the logistical constraints of refrigerated blood.

Background: Hemoglobin (Hb)-based oxygen (O 2 ) carriers (HBOCs) are being developed as alternatives to red blood cells and blood when these products are unavailable. Clinical trials of previous HBOC generations revealed side effects, including hypertension and vasoconstriction, that were not observed in preclinical studies. Large molecular weight (MW) polymerized bovine Hb (PolybHb) represents a new class of HBOC with promising results. We evaluated the safety profile of PolybHb after an exchange transfusion (ET) in guinea pigs (GPs). This study compares changes in indices of cardiac, inflammatory, and organ function after ET with high (R-state) and low (T-state) O 2 affinity PolybHb with high MW., Study Design and Methods: Guinea pigs underwent a 20% ET with PolybHb. To assess the implication of PolybHb ET on the microcirculation, hamsters instrumented with a dorsal window chamber were subjected to a similar volume ET., Results: T and R-state PolybHb did not induce significant alterations in cardiac function. T-state PolybHb induced mild vasoconstriction shortly after transfusion, while R-state did not have acute effects on microvascular tone., Conclusion: Large MW PolybHbs were found to be safe and efficacious in increasing O 2 carrying capacity and the O 2 affinity of the PolybHb did not affect O 2 delivery or extraction by tissues in relevant preclinical models. In conclusion, these results suggest that both T-state and R-state PolybHb are safe and do not impair O 2 delivery. The results are encouraging and support further evaluation of high MW PolybHbs and their future feasibility compared to allogenic blood in a trauma model., (© 2020 AABB.)

Amelioration of immune overactivity during sepsis is key to restoring hemodynamics, microvascular blood flow, and tissue oxygenation, and in preventing multi-organ dysfunction syndrome. The systemic inflammatory response syndrome that results from sepsis ultimately leads to degradation of the endothelial glycocalyx and subsequently increased vascular leakage. Current fluid resuscitation techniques only transiently improve outcomes in sepsis, and can cause edema. Nitric oxide (NO) treatment for sepsis has shown promise in the past, but implementation is difficult due to the challenges associated with delivery and the transient nature of NO. To address this, we tested the anti-inflammatory efficacy of sustained delivery of exogenous NO using i.v. infused NO releasing nanoparticles (NO-np). The impact of NO-np on microhemodynamics and immune response in a lipopolysaccharide (LPS) induced endotoxemia mouse model was evaluated. NO-np treatment significantly attenuated the pro-inflammatory response by promoting M2 macrophage repolarization, which reduced the presence of pro-inflammatory cytokines in the serum and slowed vascular extravasation. Combined, this resulted in significantly improved microvascular blood flow and 72-h survival of animals treated with NO-np. The results from this study suggest that sustained supplementation of endogenous NO ameliorates and may prevent the morbidities of acute systemic inflammatory conditions. Given that endothelial dysfunction is a common denominator in many acute inflammatory conditions, it is likely that NO enhancement strategies may be useful for the treatment of sepsis and other acute inflammatory insults that trigger severe systemic pro-inflammatory responses and often result in a cytokine storm, as seen in COVID-19., (Copyright © 2020. Published by Elsevier Inc.)

Cisplatin is a promising therapeutic for the treatment of non-small cell lung cancer (NSCLC). Unfortunately, a significant portion of NSCLC patients relapse due to cisplatin chemoresistance. This chemoresistance is thought to be primarily associated with hypoxia in the tumor microenvironment. Administration of hemoglobin (Hb)-based oxygen (O 2 ) carriers (HBOCs) is a promising strategy to alleviate hypoxia in the tumor, which may make cisplatin more effective. In this study, we administered a high O 2 affinity, relaxed state (R-state) polymerized hemoglobin (PolyHb) to three different NSCLC cell lines cultured in vitro and implanted in vivo into healthy mice. The R-state PolyHb administered in this study is unable to deliver O 2 unless under severe hypoxia which significantly limits its oxygenation potential. In vitro sensitivity studies indicate that the administration of PolyHb increases the effectiveness of cisplatin under hypoxic conditions. Additional animal studies revealed that co-administration of PolyHb with cisplatin attenuated tumor growth without alleviating hypoxia. Analysis of reactive O 2 species production in the presence of hypoxic culture indicates that exogenous ROS production by oxidized PolyHb may the mechanism of chemosensitization. This ROS mechanism, coupled with oxygenation, may be a potential chemosensitizing strategy for use in NSCLC treatment., Competing Interests: CONFLICTS OF INTEREST Authors have no conflicts of interest to declare., (Copyright: © 2020 Lucas et al.)

Background: Hemoglobin (Hb)-based oxygen carriers (HBOCs) have been proposed as alternatives to blood for decades. Previous studies demonstrated that large molecular diameter HBOCs based on polymerized bovine Hb (PolybHb) attenuate Hb side-effects and toxicity. The objective of this study was to test the safety and efficacy of tense state PolybHb after long-term storage., Methods and Results: PolybHb was subjected to diafiltration to remove low molecular weight (< 500 kDa) species and stored for 2 years. PolybHb was studied in parallel with blood, collected from rats and stored leukodepleted under blood bank conditions for 3 weeks. Rats were hemorrhaged and resuscitated to 90% of the blood pressure before the hemorrhage with fresh blood, stored blood, fresh PolybHb, or 2-year-stored PolybHb. Hemorrhagic shock impaired oxygen delivery and cardiac function. Resuscitation restored blood pressure and cardiac function, but stored blood required a significantly larger transfusion volume to recover from shock compared with fresh blood and PolybHb (fresh and stored). Stored blood transfusion elevated markers of organ damage compared with all other groups., Conclusions: These studies indicate that large molecular diameter PolybHb is as efficacious as fresh blood in restoring cardiac function and confirm the lack of degradation of PolybHb's safety or efficacy during long-term storage.

Alternatives to blood for use in transfusion medicine have been investigated for decades. An ideal alternative should improve oxygen (O 2 )-carrying capacity and O 2 delivery and support microvascular blood flow. Previous studies have shown that large-molecular diameter hemoglobin (Hb)-based oxygen carriers (HBOCs) based on polymerized bovine Hb (PolybHb) reduce the toxicity and vasoconstriction of first-generation HBOCs by increasing blood and plasma viscosity and preserving microvascular perfusion. The objective of this study was to examine the impact of PolybHb concentration and therefore O 2 -carrying capacity and solution viscosity on resuscitation from hemorrhagic shock in rats. PolybHb was diafiltered on a 500-kDa tangential flow filtration (TFF) module to remove low-molecular weight (MW) PolybHb molecules from the final product. Rats were hemorrhaged and maintained in hypovolemic shock for 30 min before transfusion of PolybHb at 10 g/dL (PHB10), 5 g/dL (PHB5), or 2.5 g/dL (PHB2.5) concentration, to restore blood pressure to 90% of the animal's baseline blood pressure. Resuscitation restored blood pressure and cardiac function in a PolybHb concentration-dependent manner. Parameters indicative of the heart's metabolic activity indicated that the two higher PolybHb concentrations better restored coronary O 2 delivery compared with the low concentration evaluated. Markers of organ damage and inflammation were highest for PHB10, whereas PHB5 and PHB2.5 showed similar expression of these markers. These studies indicate that a concentration of ~5 g/dL of PolybHb may be near the optimal concentration to restore cardiac function, preserve organ function, and mitigate the toxicity of PolybHb during resuscitation from hemorrhagic shock. NEW & NOTEWORTHY Large-molecular diameter polymerized bovine hemoglobin avoided vasoconstriction and impairment of cardiac function during resuscitation from hemorrhagic shock that was seen with previous hemoglobin-based O 2 carriers by increasing blood viscosity in a concentration-dependent manner. Supplementation of O 2 -carrying capacity played a smaller role in maintaining cardiac function than increased blood and plasma viscosity.

Polymerized hemoglobin (PolyHb) is a promising hemoglobin (Hb)-based oxygen carrier currently undergoing development as a red blood cell substitute. Unfortunately, commercially developed products are composed of low-molecular-weight (LMW) PolyHb molecules, which extravasate, scavenge nitric oxide, and result in vasoconstriction and hypertension. The naturally occurring Hb-scavenging species haptoglobin (Hp), combined with the purified heme-scavenging species apohemoglobin (apoHb), is a potential candidate to alleviate the pressor effect of PolyHb. This study evaluated the protective activity of administering the apoHb-Hp complex to mitigate the vasoactive response induced by the transfusion of LMW PolyHb. Hp binding to PolyHb was characterized in vitro. The effectiveness of apoHb-Hp administration on reducing the vasoconstriction and pressor effects of PolyHb was assessed by measuring systemic and microcirculatory hemodynamics. Transfusion of LMW PolyHb to vehicle control pretreated animals increased mean arterial pressure while decreasing arteriole diameter and functional capillary density. However, transfusion of LMW PolyHb to apoHb-Hp pretreated animals prevented changes in mean arterial pressure, heart rate, arteriole diameter, blood flow, and functional capillary density relative to before transfusion. These results indicate that the increased size of PolyHb after binding to the apoHb-Hp complex may help compartmentalize PolyHb in the vascular space and thus reduce extravasation, nitric oxide scavenging, and toxicity responsible for vasoconstriction and systemic hypertension., (© 2020 by The American Society of Hematology.)

Hemoglobin (Hb)-based oxygen (O 2 ) carriers (HBOCs) have been developed as an alternative to red blood cells (RBCs) for use in transfusion medicine. HBOCs have many benefits over RBCs; however, previous generations of HBOCs failed in clinical trials due to unanticipated cardiotoxicity. These problems likely originated from vasoconstriction, hypertension, oxidative stress, and the presence of low-molecular-weight (MW) Hb species in the HBOC formulation. Therefore, the objective of this study is to compare the toxicity of small-MW Polymerized bovine Hb (SPolyHb) to large-MW Polymerized bovine Hb (LPolyHb) in guinea pigs, since they lack the ability to synthesize vitamin C and are more sensitive to oxidative stress than other preclinical animal models. The two PolyHbs used in this study have similar molecular diameters (72 and 69 nm, respectively), but the SPolyHb included approximately 15% Hb polymers with MW below 256 kDa, which were significantly removed from LPolyHb. Solutions were injected as a hypervolemic (topload) infusion of 10% of the blood volume into animals. SPolyHb caused a 50% elevation in mean arterial pressure (MAP) from the baseline, while LPolyHb caused only a small increase in MAP. Both PolyHbs also increased markers of organ damage and tissue and systemic inflammation compared to controls. SPolyHb caused significant changes in tissue function and vital organ toxicity markers compared to LPolyHb, specifically markers related to kidney, liver, and lung injury and systemic inflammation and iron transport by the reticuloendothelial system. LPolyHb had a longer half-life than SPolyHb, which correlates with observations made in the reticuloendothelial and iron transport systems. These studies indicate that the molecular size of PolyHb determines vasoactivity, circulation time, mechanism of elimination, toxicity, and inflammation induced by its infusion.

The microcirculation is a complex network of vessels ranging from as large as 100 μm to as small as 5 μm. This complex network is responsible for the regulation of oxygen to the surrounding tissues and ensures metabolite washout. With a more complete understanding of the microcirculation's physiologic and pathologic tendencies, engineers can create new solutions to combat blood pathologies and shock-related diseases. Over the last number of decades a grown interest in the microcirculation has resulted in the development of fundamental techniques to quantify the microvasculature flow and the release of oxygen to tissues., Competing Interests: Disclosure The authors have nothing to disclose., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background: Resuscitation from hemorrhagic shock (HS) by blood transfusion restores oxygen (O2) delivery and provides hemodynamic stability. Current regulations allow red blood cells (RBCs) to be stored and used for up to 42 days. During storage, RBCs undergo many structural and functional changes. These storage lesions have been associated with adverse events and increased mortality after transfusion, increasing the need for improved RBC storage protocols. This study evaluates the efficacy of anaerobically stored RBCs to resuscitate rats from severe HS compared with conventionally stored RBCs., Methods and Results: Rat RBCs were stored under anaerobic, anaerobic/hypercapnic, or conventional conditions for a period of 3 weeks. Hemorrhage was induced by controlled bleeding, shock was maintained for 30 min, and RBCs were transfused to restore and maintain blood pressure near the prhemorrhage level. All storage conditions met current regulatory 24-h posttransfusion recovery requirements. Transfusion of anaerobically stored RBCs required significantly less RBC volume to restore and maintain hemodynamics. Anaerobic or anaerobic/hypercapnic RBCs restored hemodynamics better than conventionally stored RBCs. Resuscitation with conventionally stored RBCs impaired indices of left ventricular cardiac function, increased hypoxic tissue staining and inflammatory markers, and affected organ function compared with anaerobically stored RBCs., Conclusions: Resuscitation from HS via transfusion of anaerobically stored RBCs recovered cardiac function, restored hemodynamic stability, and improved outcomes.

The mechanical properties and deformability of Red Blood Cells (RBCs) are important determinants of blood rheology and microvascular hemodynamics. The objective of this study is to quantify the mechanical properties and wall shear stress experienced by the RBC membrane during capillary plug flow in vivo utilizing high speed video recording from intravital microscopy, biomechanical modeling, and computational methods. Capillaries were imaged in the rat cremaster muscle pre- and post-RBC transfusion of stored RBCs for 2-weeks. RBC membrane contours were extracted utilizing image processing and parametrized. RBC parameterizations were used to determine updated deformation gradient and Lagrangian Green strain tensors for each point along the parametrization and for each frame during plug flow. The updated Lagrangian Green strain and Displacement Gradient tensors were numerically fit to the Navier-Lame equations along the parameterized boundary to determined Lame's constants. Mechanical properties and wall shear stress were determined before and transfusion, were grouped in three populations of erythrocytes: native cells (NC) or circulating cells before transfusion, and two distinct population of cells after transfusion with stored cells (SC1 and SC2). The distinction, between the heterogeneous populations of cells present after the transfusion, SC1 and SC2, was obtained through principle component analysis (PCA) of the mechanical properties along the membrane. Cells with the first two principle components within 3 standard deviations of the mean, were labeled as SC1, and those with the first two principle components greater than 3 standard deviations from the mean were labeled as SC2. The calculated shear modulus average was 1.1±0.2, 0.90±0.15, and 12 ± 8 MPa for NC, SC1, and SC2, respectively. The calculated young's modulus average was 3.3±0.6, 2.6±0.4, and 32±20 MPa for NC, SC1, and SC2, respectively. o our knowledge, the methods presented here are the first estimation of the erythrocyte mechanical properties and shear stress in vivo during capillary plug flow. In summary, the methods introduced in this study may provide a new avenue of investigation of erythrocyte mechanics in the context of hematologic conditions that adversely affect erythrocyte mechanical properties., (Copyright © 2020 Jani, Lucas, Jani, Munoz, Williams, Ortiz, Yalcin and Cabrales.)

Acclimatization to hypoxia or high altitude involves physiological adaptation processes, to influence oxygen (O 2 ) transport and utilization. Several natural products, including aromatic aldehydes and isothiocyanates stabilize the R-state of hemoglobin (Hb), increasing Hb-O 2 affinity and Hb-O 2 saturation. These products are a counter intuitive therapeutic strategy to increase O 2 delivery during hypoxia. 5-Hydroxymethylfurfural (5-HMF) is well known Amadori compound formed during the Maillard reaction (the non-enzymatic browning and caramelization of carbohydrate-containing foods after thermal treatment), with well documented effects in Hb-O 2 affinity. This study explores the therapeutic potential of 5-HMF on left ventricular (LV) cardiac function (LVCF) during hypoxia. Anesthetized Golden Syrian hamsters received 5-HMF i.v., at 100 mg/kg and were subjected to stepwise increased hypoxia (15, 10, and 5%) every 30 min. LVCF was assessed using a closed chest method with a miniaturized conductance catheter via continuous LV pressure-volume (PV) measurements. Heart hypoxic areas were studied using pimonidazole staining. 5-HMF improved cardiac indices, including stroke volume (SV), cardiac output (CO), ejection fraction (EF), and stroke work (SW) compared to the vehicle group. At 5% O 2 , SV, CO, EF, and SW were increased by 53, 42, 33, and 51% with 5-HMF relative to vehicle. Heart chronotropic activity was not statistically changed, suggesting that differences in LV-CF during hypoxia by 5-HMF were driven by volume dependent effects. Analysis of coronary blood flow and cardiac muscle metabolism suggest no direct pharmacological effects from 5-HMF, therefore these results can be attributed to 5-HMF-dependent increase in Hb-O 2 affinity. These studies establish that naturally occurring aromatic aldehydes, such as 5-HMF, produce modification of hemoglobin oxygen affinity with promising therapeutic potential to increase O 2 delivery during hypoxic hypoxia., (Copyright © 2019 Lucas, Ao-ieong, Williams, Jani, Muller, Yalcin and Cabrales.)

This paper implements logistic regression models (LRMs) and feature selection for creating a predictive model for recovery form hemorrhagic shock (HS) with resuscitation using blood in the multiple experimental rat animal protocols. A total of 61 animals were studied across multiple HS experiments, which encompassed two different HS protocols and two resuscitation protocols using blood stored for short periods using five different techniques. Twenty-seven different systemic hemodynamics, cardiac function, and blood gas parameters were measured in each experiment, of which feature selection deemed only 25% of the them as relevant. The reduced feature set was used to train a final logistic regression model. A final test set accuracy is 84% compared to 74% for a baseline classifier using only MAP and HR measurements. Receiver operating characteristics (ROC) curve analysis and Cohens kappa statistics were also used as measures of performance, with the final reduced model outperforming the model, including all parameters. Our results suggest that LRMs trained with a combination of systemic hemodynamics, cardiac function, and blood gas parameters measured at multiple timepoints during HS can successfully classify HS recovery groups. Our results show the predictive ability of traditional and novel hemodynamic and cardiac function features and their combinations, many of which had not previously been taken into consideration, for monitoring HS. Furthermore, we have devised an effective methodology for feature selection and shown ways in which the performance of such predictive models should be assessed in future studies.

Oxygen plays a central role in cardiac energy metabolism. At high altitude where the ambient oxygen level is low, we found EDNRB is associated with human hypoxia adaptation. Our subsequent study in global heterozygous knockout mice (Ednrb -/+ ) revealed that cardiac function was conserved in these mice when exposed to extreme hypoxia. The major goal of this study was (i) to determine the functional role of cardiomyocyte EdnrB in maintaining cardiac function under hypoxic stress and (ii) to validate the phenotypes we detected in Ednrb -/+ mice using EDNRB blockers. Unlike the global knockouts, cardiac-specific heterozygote (EdnrB flox/+ ) and homozygote (EdnrB flox/flox ) EdnrB knockout mice were phenotypically normal. When treated with graded low levels of oxygen (10% and 5% O 2 ), both EdnrB flox/+ and EdnrB flox/flox were hypoxia tolerant. The cardiac indexes at 10% and 5% O 2 for EdnrB flox/+ were significantly higher and lactate levels were significantly lower when compared to the cre-negative controls (P < 0.05). Simultaneously, mice treated with BQ-788 (EDNRB-specific blocker) had a significantly higher cardiac index (P < 0.005) and significantly lower lactate levels (P < 0.0001) than in control mice. A similar result was obtained with mice treated with Bosentan (non-specific). These data indicate that a lower level or complete lack of EdnrB in the cardiomyocytes significantly improves cardiac performance under extreme hypoxia, a novel role of cardiomyocyte EdnrB in the regulation of cardiac function. Furthermore, this rescue under extreme hypoxia can also be achieved using EDNRB-specific pharmacological agents, e.g., BQ-788. This systematically confirms, both genetically and pharmacologically, the protective role of a lower EDNRB under extreme hypoxia stress., Key Messages: Under normal condition, cardiomyocytes-specific EdnrB knockout mice, both heterozygote and homozygote, are phenotypically normal. Under hypoxic condition, a lower level or complete deletion of cardiomyocyte EdnrB conserves cardiac function by maintaining high cardiac index. Similarly, mice treated with both specific (BQ-788) and non-specific (Bosentan) EDNRB blockers are tolerant to hypoxia by maintaining better cardiac function. The oxygen perfusion under extreme hypoxia is better in the mice with lower EDNRB, as depicted by lower lactate level at 5% oxygen. Our current study systematically confirms, both genetically and pharmacologically, the protective role of a lower EDNRB under extreme hypoxia stress. Overall, it supports our hypothesis that studies on human hypoxia adaptation provide new insight to common disease pathogenesis and treatments.

In 2013, nearly 15 million units of banked blood were transfused in the United States of America alone. Blood shortages are expected to increase globally. Donated blood is not equal due to differences in quality and deterioration rate. There are no methods to detect time-dependent biochemical and biophysical changes of red blood cells (RBCs) or the deterioration rate of donated RBCs. Nine randomly selected RBC units collected by the San Diego Blood Bank were examined for interdonor variability over six weeks of storage. In vitro RBC quality was assessed weekly by conventional biochemical tests including free Hb, K+, ATP, P50, 2,3 DPG, lactate, and pH. Deformability was measured via cell filtration. Briefly, the RBC suspension (10% Hct), was forced through a 5.0-μm pore membrane (106 mm 2 ) at various flow rates. No interdonor variability in biochemical or mechanical parameters was observed at baseline. Interdonor variability in biochemical properties (free Hb, K+, ATP, P50, 2,3 DPG, lactate, and pH) was observed after 14 days of storage. However, significant differences from baseline in RBC mechanical properties (i.e., filterability) were observed as early as 7 days into storage at the lowest flow rates and after 28 days of storage at all flow rates. There was a net decrease in filterability over time for all donors, but the rate at which filterability decreased (i.e., deterioration rates) was different when comparing individual donors. Changes in all biochemical parameters were significant different between donors. These data suggest that filterability is more sensitive to changes in blood quality than conventional biochemical parameters.

Background: Small rodent models are routinely used to evaluate the safety and efficacy of blood transfusions. Limited comprehensive literature exists about effect of different storage solutions in rat red blood cells (RBCs) characteristics. RBCs undergo time dependent biochemical and biophysical changes during storage known as hypothermic storage lesions (HSLs)., Objective: This study evaluates the effects of RBC additive solutions (AS) during storage of rat RBCs., Methods: Blood was leukoreduced and stored as per manufacturer instructions at 4°C up to 42-days. Three solutions, CPDA-1; AS-1; and AS-7 (SOLX), were evaluated. Biochemical parameters measured included extracellular K+, pH, hemolysis, 2,3-diphosphoglycerate (2,3-DPG), oxygen affinity, ATP, and lactate. Mechanical properties measured included RBC deformability, elongation index (EI), RBC membrane shear elastic modulus (SEM), mean corpuscular volume (MCV), viscosity, and aggregability., Results: There were no differences in biochemical or mechanical parameters at baseline or after one week of storage. However, after two weeks, AS-7 preserved biochemical and mechanical properties as compared to CPDA-1 and AS-1. Changes were observed to be significant after 14-days of storage. AS-7 prevented extracellular K+ increase, reduced acidosis, showed lower hemolysis, preserved ATP and 2,3-DPG levels (consequently oxygen affinity), and reduced lactate. AS-7, when compared to CPDA-1 and AS-1, prevented the reduction in RBC deformability and was found to preserve the EI at multiple shear stresses, the membrane SEM, the aggregability and viscosity., Discussion: Rat RBCs stored with AS-7 presented reduced changes in biochemical and mechanical parameters, when compared with rat RBCs stored in CPDA-1 and AS-1, after as early as two weeks of storage.

New Findings: What is the central question of this study? The purpose of the present study was to determine the effects of exercise-induced haemoconcentration and hyperthermia on blood viscosity. What is the main finding and its importance? Exercise-induced haemoconcentration, increased plasma viscosity and increased blood aggregation, all of which increased blood viscosity, were counterbalanced by increased red blood cell (RBC) deformability (e.g. RBC membrane shear elastic modulus and elongation index) caused by the hyperthermia. Thus, blood viscosity remained unchanged following prolonged moderate-intensity exercise in the heat. Previous studies have reported that blood viscosity is significantly increased following exercise. However, these studies measured both pre- and postexercise blood viscosity at 37 °C even though core and blood temperatures would be expected to have increased during the exercise. Consequently, the effect of exercise-induced hyperthermia on mitigating change in blood viscosity may have been missed. The purpose of this study was to isolate the effects of exercise-induced haemoconcentration and hyperthermia and to determine their combined effects on blood viscosity. Nine subjects performed 2 h of moderate-intensity exercise in the heat (37 °C, 40% relative humidity), which resulted in significant increases from pre-exercise values for rectal temperature (from 37.11 ± 0.35 to 38.76 ± 0.13 °C), haemoconcentration (haematocrit increased from 43.6 ± 3.6 to 45.6 ± 3.5%) and dehydration (change in body weight = -3.6 ± 0.7%). Exercise-induced haemoconcentration significantly (P < 0.05) increased blood viscosity by 9% (from 3.97 to 4.33 cP at 300 s(-1)), whereas exercise-induced hyperthermia significantly decreased blood viscosity by 7% (from 3.97 to 3.69 cP at 300 s(-1)). When both factors were considered together, there was no overall change in blood viscosity (from 3.97 to 4.03 cP at 300 s(-1)). The effects of exercise-induced haemoconcentration, increased plasma viscosity and increased red blood cell aggregation, all of which increased blood viscosity, were counterbalanced by increased red blood cell deformability (e.g. red blood cell membrane shear elastic modulus and elongation index) caused by the hyperthermia. Thus, blood viscosity remained unchanged following prolonged moderate-intensity exercise in the heat., (© 2015 The Authors. Experimental Physiology © 2015 The Physiological Society.)

New Findings: What is the central question of this study? The aim was to evaluate the effect of perfusion pressure on blood flow in small arterioles. The hypothesis was that blood flow regulates the thickness of the red-cell-free layer and, therefore, blood flow determines blood apparent viscosity and local vascular resistance in vascular networks with limited myogenic or metabolic regulation of blood flow. What is the main finding and its importance? Reduced perfusion pressures lowered volumetric flow rates and increased local vascular resistance, due to increased blood apparent viscosity. Thus, the local vascular resistance of small arterioles with limited myogenic or metabolic regulation of blood flow, appeared to be determined by changes in blood rheology rather than blood vessel diameter. The study of blood flow regulation is important to understand and resolve pathological conditions. As blood is a complex non-Newtonian multiphase system, the foundations of blood rheological properties have been obtained mostly in viscometers. However, blood rheological behaviour in vivo depends on the concentration of red blood cells (RBCs), their mechanical properties and the RBC hydrodynamics, including RBC migration away from the vessel wall in shear flow. This migration promotes the formation of a RBC-depleted zone, or cell-free layer (CFL), which reduces the apparent viscosity of blood. We hypothesize that perfusion pressure determines blood apparent viscosity in microvessels, as shear rate affects axial migration of RBCs by influencing the CFL thickness. In this study, we analysed the effects of perfusion pressure on blood flow in individual arterioles within the rat cremaster muscle preparation. Perfusion pressures to this microvascular bed were controlled by occlusions of the iliac artery using a pressure cuff. Blood flow measurements were obtained from direct measurements of blood flow velocity profile, as well as determination of CFL thickness using intravital microscopy. Our results indicate that perfusion pressure determines shear rates and the CFL thickness and its variations. In addition, blood flow reduction increased local vascular resistance by augmenting blood apparent viscosity rather than vascular hindrance. In conclusion, blood rheology could act as an intrinsic mechanism to further limit blood flow to tissue with limited myogenic and metabolic responses at low perfusion pressures., (© 2015 The Authors. Experimental Physiology © 2015 The Physiological Society.)