Cellular Hemoglobin Research Articles

Evaluation of inflammatory-thrombosis panel as a diagnostic tool for vascular Behçet's disease.

Vascular Behçet's disease (VBD) is prevalent in 40% of BD, but lacks laboratory biomarker for timely diagnosis. We aimed to establish a diagnostic panel for discerning VBD and non-VBD patients and identify hemostatic-thrombotic markers most related to VBD pathogenesis using machine learning algorithm. A total of 338 BD patients comprising 123 VBD and 215 non-VBD were enrolled. Twenty-six clinical and laboratory features selected from LassoCV were included in multiple classifier to choose the optimal model for VBD differentiation. The Shapley Additive exPlanations (SHAP) was employed to interpret the contribution of model features for VBD prediction. Logistic regression analysis and nomogram were conducted to screen risk factors of VBD. Inflammatory (neutrophils%, NK cells, IL-6), hematological (hemoglobin, hemoglobin distribution width (HDW)) and thrombosis (activated partial thromboplastin clotting time (APTT), D-dimer) parameters were elevated in VBD. Then we chose top contributors from XGBoost model and performed ten-fold cross validation, the diagnostic accuracy of which exceeded 0.90. Utilizing SHAP method, we identified higher incidence of arterial thrombosis or aneurysm and deep vein thrombosis, upregulated NK cell count, HDW, APTT and D-dimer, downregulated reticulocyte%, B cell count, red blood cell distribution width, cellular hemoglobin (CH) and TNF-α would ultimately generate the phenotype of VBD. Severity, hemoglobin, mean corpuscular hemoglobin, CH, HDW, APTT and D-dimer were found as potential risk factors for vascular outcomes among BD. Our study developed a well-performed model leveraging clinical and laboratory parameters for differentiating VBD. Inflammatory and thrombotic risk factors are potential contributors to VBD. Key Points • Inflammatory (neutrophils%, NK cells, IL-6), hematological (HGB, HDW) and thrombosis (APTT, D-dimer) parameters were elevated in VBD. • We firstly developed an inflammatory-thrombosis model as a diagnostic tool for VBD. • HGB, MCH, CH, HDW, APTT and D-dimer are potential risk factors for VBD.

Overview on Hereditary Spherocytosis Diagnosis.

Hereditary spherocytosis (HS) is a congenital haemolytic disorder, resulting from plasma membrane protein deficiency of red blood cells (RBCs). Typical pathological signs are anemia, jaundice, and splenomegaly; in newborns, jaundice is the main symptom. This study focused on the state of art about the HS diagnosis, from traditional to innovative methods, including diagnostic algorithms that can be applied for pediatric and adult patients, for different laboratory diagnostic levels. The first erythrocyte parameters used for HS diagnosis were the mean corpuscular hemoglobin concentration (MCHC), mean corpuscular volume (MCV), and red blood cell distribution width (RDW); nowadays new parameters are used in blood cell counter. Advia analyzers (Siemens Medical Solutions) supply the hyper-dense cell percentage (% Hyper), which reflects the red blood cells hyperchromia. Sysmex instruments (i.e. XT-4000i, XE-5000, XN-Series) provide the MicroR, that is the percentage of erythrocytes smaller than 60 fL, Hypo-He, which is the percentage of erythrocytes with a content of hemoglobin less than 17 pg and % Hyper-He, which represents the percentage of RBC with cellular hemoglobin content higher than 49 pg. CELL-DYN Sapphire (Abbott Diagnostics) introduced the HPR parameter (% HPR), which represents the erythrocytes with hemoglobin > 410 g/L. Beckman Coulter instruments supply the mean sphered corpuscular volume (MSCV), which is the average volume of all erythrocytes, including mature erythrocytes and reticulocytes. Other reference tests for screening and diagnosis of HS are the acidified glycerol lysis test (AGLT), the eosin-5-maleimide (EMA) binding test and genetic testing by next-generation sequencing. The diagnostic workup of hereditary spherocytosis could be improved thanks to all the available tests, including new molecular tools. However, it requires synergy between clinicians and laboratory staff, evaluating clinical manifestations, all available data related to the disease and the prognosis to fill the diagnostic gaps in the near future.

EPIDEMIOLOGY OF BOVINE FASCIOLIASIS: PREVALENCE AND HAEMATO-BIOCHEMICAL ALTERATIONS IN KATSINA ABATTOIR NORTH WESTERN NIGERIA

Bovine fascioliasis, caused by liver flukes Fasciola hepatica and Fasciola gigantica, significantly affects cattle health and production globally. This study evaluated the prevalence and haemato-biochemical changes associated with bovine fascioliasis in 134 cattle at Katsina Central Abattoir. Using the sedimentation technique, faecal samples revealed a 3.73% infection rate, with females showing a higher prevalence (4.55%) than males (2.17%), though not statistically significant (P > 0.05). The age group 2-3 years had the highest prevalence (25.00%), with significant differences among age groups (P < 0.05). Among breeds, Wadara had the highest prevalence (20.00%) compared to White Fulani (4.93%), with a significant difference (P < 0.05). Medium-conditioned cattle showed the highest prevalence (6.67%) versus good-conditioned ones (1.37%) with significant difference (P < 0.05). Infected cattle exhibited significant reductions (P < 0.05) in packed cell volume (PCV), haemoglobin (Hb), and total erythrocyte count (RBC), as well as decreases in white blood cell count (WBC), mean cellular volume (MCV), and mean cellular haemoglobin concentration (MCHC). Biochemical analysis indicated elevated alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), and total bilirubin (TB) levels (P < 0.05), with reduced albumin (ALB), total protein (TP), and conjugated bilirubin (CB) levels (P < 0.05). These results highlight the significant impact of bovine fascioliasis on cattle health in Katsina State, emphasizing the need for effective control strategies.

Osmotic gradient ektacytometry - a novel diagnostic approach for neuroacanthocytosis syndromes.

The unique red blood cell (RBC) properties that characterize the rare neuroacanthocytosis syndromes (NAS) have prompted the exploration of osmotic gradient ektacytometry (Osmoscan) as a diagnostic tool for these disorders. In this exploratory study, we assessed if Osmoscans can discriminate NAS from other neurodegenerative diseases. A comprehensive assessment was conducted using Osmoscan on a diverse group of patients, including healthy controls (n = 9), neuroacanthocytosis syndrome patients (n = 6, 2 VPS13A and 4 XK disease), Parkinson's disease patients (n = 6), Huntington's disease patients (n = 5), and amyotrophic lateral sclerosis patients (n = 4). Concurrently, we collected and analyzed RBC indices and patients' characteristics. Statistically significant changes were observed in NAS patients compared to healthy controls and other conditions, specifically in osmolality at minimal elongation index (Omin), maximal elongation index (EImax), the osmolality at half maximal elongation index in the hyperosmotic part of the curve (Ohyper), and the width of the curve close to the osmolality at maximal elongation index (Omax-width). This study represents an initial exploration of RBC properties from NAS patients using osmotic gradient ektacytometry. While specific parameters exhibited differences, only Ohyper and Omax-width yielded 100% specificity for other neurodegenerative diseases. Moreover, unique correlations between Osmoscan parameters and RBC indices in NAS versus controls were identified, such as osmolality at maximal elongation index (Omax) vs. mean cellular hemoglobin content (MCH) and minimal elongation index (EImin) vs. red blood cell distribution width (RDW). Given the limited sample size, further studies are essential to establish diagnostic guidelines based on these findings.

Low Initial Cell Density Promotes the Differentiation and Maturation of Human Pluripotent Stem Cells into Erythrocytes.

Human pluripotent stem cell (hPSC)-derived red blood cells (RBCs) possess great potential for compensating shortages in transfusion medicine. For better RBC generation from hPSCs, we compared the cell seeding density in the embryoid body formation-based hPSC induction protocol. In the selection of low- and high-density inoculation conditions, we found that low-density culture performed better in the final RBC product with more cell output and increased average cellular hemoglobin content. An elaborate study using flow cytometry demonstrated that low inoculation density promoted endothelial-to-hematopoietic transition, followed by improved hematopoietic progenitor formation and erythrocyte generation. The improved transformation from glycolysis to mitochondrial oxidation and reduced apoptosis might be responsible for this effect. Hints from RNA sequencing suggested that molecules involved in microenvironment interaction and metabolic regulation might respond for the different developmental potential. The possible mediators between outer message and intracellular response could be the nutrition sensors FOXO, PRKAA1 (AMPK), and MTOR genes. It is possible that low inoculation density triggered metabolic regulation signals, promoted mitochondrial oxidation, and resulted in enhanced cell amplification and hematopoietic differentiation. The low cell culture density will improve RBC generation from hPSCs.

A novel formula of herbal extracts regulates growth performance, antioxidant capacity, intestinal microbiota and resistance against Aeromonas veronii in largemouth bass (Micropterus salmoides)

In this study, a novel herbal extracts formula (HE) comprising Scutellaria baicalensis, Taraxacum mongolicum, Biancaea sappan and Pulsatilla chinensis extract with a weight ratio of 3.0: 1.8: 3.0: 1.7, was selected based on bacteriostatic test in vitro. Subsequently, a 30-day feeding trial was conducted to evaluate the effect of dietary HE (0, 0.05, 0.10, 0.50 and 1.00%, respectively) inclusion on the performance of largemouth bass (Micropterus salmoides). The results unveiled that dietary HE 0.10% and HE 0.50% inclusion notably increased the specific growth rate (SGR) and antioxidant enzyme (SOD and GPX) activities in the intestine and liver of fish (p < 0.05). Additionally, a significantly greater height of villi and width of villi were observed in the HE 0.50% and HE 0.10% groups, respectively (p < 0.05). Conversely, histological examination indicated that dietary HE1.00% inclusion led to discernible injury to the intestines, with certain villi displaying breakage and shedding. Hematological parameters including white blood cells (WBC) and red blood cells (RBC) were significantly increased, while the erythrocyte indices including average cellular hemoglobin (MCH), and average cellular hemoglobin concentration (MCHC) remarkably decreased in fish fed HE inclusion compared to the basal diet. The 16S rRNA sequence analysis demonstrated that Cetobacterium, Aeromonas and Plesiomonas were the three dominant species in the intestines regardless of diet. Additionally, the LEfSe analysis revealed that Aeromonas was the most affected taxon, which significantly decreased in the HE 0.10%, HE 0.50% and HE 1.00% groups (p < 0.05), while the Cetobacterium content remarkably increased in HE 0.10% and HE 0.50% groups (p < 0.05). Finally, the highest survival rate of fish infected with Aeromonas veronii HZ012 was observed in the group supplemented with 0.50% HE. Collectively, our study revealed that the supplementation of HE at a proportion of 0.10–0.50% could lead to a promotion of growth performance, maintenance of liver and intestinal health, as well as a protective effect against A. veronii infection in largemouth bass.

Relationships Between Markers of Iron Status and Hematological Parameters in Patients With Sickle Cell Disease.

Based on the relationship between the intracellular concentration of sickle hemoglobin S (HbS) and the delay that occurs prior to the onset of sickling following deoxygenation, targeting the intracellular HbS concentration is a recognized therapeutic approach for sickle cell disease (SCD). We and others have shown that restricting iron by dietary or pharmacologic means improves hematologic parameters, inflammation, and organ damage in mouse models of SCD. Clinical evidence corroborating these findings is confined to case reports and small case series studies, none of which account for treatment or α-thalassemia. We hypothesize that increased transferrin saturation is associated with increased mean cellular hemoglobin concentration (MCHC) which in turn is associated with decreased red cell counts and worsening anemia. To investigate this hypothesis, we examined the relationships between transferrin saturation and MCHC with each of the parameters that define MCHC in sickle patients (HbSS without α-thalassemia) and healthy volunteers (HVs). Results indicate that transferrin saturation and MCHC are positively correlated with each other in sickle patients and HV. In patients with SCD, MCHC and transferrin saturation are negatively correlated with RBC count and are not correlated with hemoglobin, whereas each is positively associated with HV. Transferrin saturation and MCHC are each positively correlated with the hemolysis marker, lactate dehydrogenase. These observations support a model where increased transferrin saturation contributes to higher intracellular HbS concentrations with subsequent increases in sickling and hemolysis in sickle patients, suggesting that pharmacologic approaches to decrease serum iron may provide a therapeutic approach for patients with SCD. Trial Registration: This study was registered with ClinicalTrials.gov identifiers: NCT00011648, NCT00081523, and NCT04817670.

Clinical Utility of Cellular Hemoglobin as a Compatible Marker to the Saline Replacement Method for Hemoglobin Measurement in Lipemic Samples

Clinical Utility of Cellular Hemoglobin as a Compatible Marker to the Saline Replacement Method for Hemoglobin Measurement in Lipemic Samples

Role of inflammation in determining the severity of COVID-19 infection in patients with diabetes: A comparative study.

There is a need to consider the geographical origins when studying the association between COVID-19 and the comorbid conditions. To examine the role of inflammation in determining the severity of COVID-19 among hospitalized patients with diabetes and compare these roles with those who does not have diabetes. A cross sectional comparative design was used with a convenience sample of 352 patients. Samples were collected from hospitalized patients with COVID-19 who were divided into 2 groups (diabetes and non-diabetes). Data regarding results of selected inflammatory markers and sociodemographic were collected. The severity of COVID-19 differed significantly between the diabetes and non-diabetes groups (Chi square = 25.58 P < .05). There was significant difference in the mean scores of neutrophil counts, monocyte count, Basophil count, erythrocyte sedimentation rate, partial thromboplastin time, C-creative protein, platelets, white blood cells, and mean cellular hemoglobin center between those with and those without diabetes. The diabetes were shown more increased in the predictors and severity of the COVID-19 disease. However, neutrophil to lymphocyte ratio, neutrophil count, and age were the significant predictors of the severity level of COVID-19 among patients with diabetes. In conclusion, our study addressed the influence of having diabetes among hospitalized patients with moderate and severe COVID-19 infection. The results showed that severity of COVID-19 infection was affected by diabetes where those with diabetes had more tendency to suffer from the severe form of the disease rather that the moderate level.

A-144 Three-wavelength Polarization Imaging Method for the Analysis of Individual Human Blood Cells With Dye-Enhanced Dispersion of Dilution Buffer

Abstract Background Performing basic hematology analysis in a near-patient setting enables more rapid clinical decision making in the process of diagnosis. Traditional blood analyzers in central labs use a complex combination of sensing principles and chemical modification of the sample to derive the necessary parameters for a complete blood count (CBC). To reduce cost and maintenance, varying approaches have been employed to determine a CBC with a single-step measurement technique. One common option is the use of optical microscopy to deliver highly reliable results, examples are fluorescence microscopy, digital holographic microscopy, or spatial light interference microscopy. However, most of these technologies lack a proper determination of cellular-level RBC parameters. Therefore, a non-interferometric, quantitative-phase measurement technique was developed to derive relevant CBC parameters from a single measurement, comparable to a standard lab analyzer. Methods The optical setup employed by this method enables the parallel acquisition of brightfield and differential interference contrast (DIC) images by polarization-sensitive cameras. By using three cameras with individual optical filters, quantitative phase and absorption information can be calculated, which enables the determination of mean cellular volume (MCV), mean cellular hemoglobin (MCH), and mean cellular hemoglobin concentration (MCHC) for red blood cells (RBCs). The whole-blood sample is diluted in a buffer containing a non-cell-penetrating dye to enhance optical refraction contrast and therefore precision in the dependent calculated quantities. The sample is then introduced to a glass flow cell, and the cells are allowed to sediment to the bottom for imaging. Machine learning is applied to the derived images to classify unstained native white blood cells (WBCs) to yield a 5-part differential, which distinguishes between neutrophil, monocyte, lymphocyte, eosinophil and basophil. The algorithm is trained by imaging FACS-validated, purified WBC subgroups. All blood samples are measured on an ADVIA® 2120 Hematology System for control. Results A one-step, optical CBC hematology system based on quantitative phase and absorption image analysis was developed. The correlations for MCV and MCHC compared to a standard lab CBC analyzer were 0.77 and 0.84. The accuracy for the 5-part differential WBC classification was approximately 0.8. To enable a visual inspection of the classification results, a pseudo-colored image of each cell was generated and uploaded in a review tool. Conclusion With a more-refined, less-expensive optical setup, a high-quality CBC analyzer suitable for use at the point of care can be derived with the presented method. A possible simplification could be the use of low-cost but high-resolution Fourier ptychography microscopy (FPM) technology. Imaging of blood samples on a single-cell level offers the potential for the analysis of even more morphological parameters, for example, disease-related changes in the cell membrane. Also, the adhesion behavior of the WBCs in the glass flow cell could indicate cell activation status and therefore give information about infection processes. *Trademark disclaimer: “ADVIA, and all associated marks are trademarks of Siemens Healthcare Diagnostics, Inc. or its affiliates. All other trademarks and brands are the property of their respective owners.”

Empowering frail older adults: multicomponent elastic-band exercises and BCAA supplementation unleash physical health and preserve haematological biomarkers.

The effectiveness of Branched Chain Amino Acids (BCAAs) supplementation on enhancing exercise performance in both young and older adults remains a topic of debate. Recent research suggests that BCAAs combined with regular exercise might have an impact on human erythropoiesis, blood dynamics, and iron homeostasis. Given the increasing longevity of the global population, it is crucial to investigate the potential benefits of BCAA supplementation and regular exercise as non-pharmacological interventions for improving the overall health of frail older adults. To assess the influence of a 40-week multicomponent exercise intervention (MEP) combined BCCA supplementation on the haematological indicators of frail older adults (83-93 years old) residing in nursing homes. A prospective, naturalistic, controlled clinical trial employing an intervention-washout-intervention was conducted for this purpose. The study included four experimental groups: MEP plus BCAA supplementation (MEP + BCAA, n = 8), MEP only (n = 7), BCAA supplementation only (n = 7), and control group non exercising (CG, n = 13). Fried's physical frailty (PF) protocol was employed to stratify the participants. Additionally, the assessment included the evaluation of nutritional status, comorbidities, and anthropometric measurements. Among the several haematological markers examined, only mean cellular Haemoglobin Concentration (MCH) [F = 4.09; p < 0.03] and Mean Cell haemoglobin Concentration (MCHC) [F = 10, 323; p < 0,0001] showed significant effects of time group. Our findings demonstrate that a long-term intervention with BCAA plus MEP did not lead to significant alterations in the haematological profile. An 8-week withdrawal from interventions did not affect the frailty status in the MEP and MEP + BCAA groups, whereas the control group exhibited an increase in PF status. The findings, demonstrating the potential pro-immune effect and maintenance of MCH and MCHC levels, highlight the relevance of incorporating exercise and nutritional strategies to promote healthy aging. This study contributes to the achievement of the United Nations Sustainable Development Goals 3 (good health and well-being) and 10 (reduced Inequalities) for all.

Relationship Between the Coma Scores and Hemogram Parameters Measured by New-generation Devices.

To investigate the relationship between the coma scores-Glasgow coma scale (GCS), Sequential Organ Failure Assessment (SOFA), and Acute Physiological and Chronic Health Assessment (APACHE-II)-in intensive care unit (ICU) patients and the percentage of macrocytosis (%MAC), immature granulocyte (IG), cellular haemoglobin concentration (cHGB), nucleated red blood cell (NRBC), nucleated red cell/ white cell ratio (NR/W), hyperchromic ratio (%HPR), and platelet distribution width (PDW) values. A descriptive comparative study. Place and Duration of the Study:Medicine Faculty, University ofHarran,Turkiye, from December 2020 to May 2022. The hemogram parameters of the patient groups with a GCS of 3-8 (n=51) and a GCS of 9-15 (n=43) and a control group of 55 healthy volunteers were measured using the new-generation hemogram autoanalyzer AlinityHQ (Abbott, USA). These parameters were compared with the coma scores (GCS, SOFA, and APACHE-II) of the patients. There was a statistically significant difference in IG, %MAC and PDW values (p-values were 0.025, 0.011, and 0.004, respectively) and an inverse correlation with GCS scores (correlation coefficients were -0.247, -0.264, and -0.297, respectively) was observed. There was also a significant correlation between the SOFA scores and %HPR and cHGB (correlation coefficients were 0.234, -0.358, p-values were 0.025, 0.001, respectively), and the APACHE-II scores and NRBC and NR/W values (correlation coefficients were -0.270, -0.247, p values were 0.009, 0.017, respectively). While other haematological parameters other than PDW were not associated with coma scores, parameters measured using new-generation haematological devices (%MAC, IG, cHGB, NRBC, NR/W, and %HPR) were found to be associated with estimated coma scores. These parameters can therefore be used as simple, rapid prognostic biomarkers and assist researchers in the development of new scoring models. ICU, Hyper, Coma, Sofa, Apache.

Response to Hemostatic Complications During Neonatal Extracorporeal Membrane Oxygenation: Roller Pump and Centrifugal Pump Driven Circuits.

To the Editor: Vermeer et al.1 recently published a case series and in vitro analysis of acute oxygenator failure manifested as a rise in preoxygenator pressure without evidence of gas transfer failure. As practitioners in the field who have fielded similar questions from colleagues describing similar issues, we applaud them for describing this problem that has been informally discussed in meetings and discussion boards for several years. The debate in neonatal extracorporeal life support (ECLS) between roller and centrifugal pumps has a long history.2 While the focus has primarily been on sources of increased hemolysis, the acute failure of the ECLS circuit by oxygenator clogging has received less attention. Vermeer et al.1 began using the RotaFlow in neonatal ECLS because of supply shortages and the availability and experience with adult ECLS. After a disturbing trend of complications, they embarked on a formal assessment of three systems (roller pump, RotaFlow, and PedIVAS) and associated failure rates in the neonatal population. The authors described anticoagulation, heat generation, and overall blood velocity as plausible mechanisms generating the cellular debris that led to the clogging phenomenon. In discussing the velocity, they came very close to what we would consider to be the fundamental problem inherent in the design of any centrifugal pump, which is their best efficiency point (BEP). The BEP is the point at which energy imparted to spinning the impeller generates the most forward bulk flow from the pump. The RotaFlow and PedivAS were designed to operate at 5 and 1.5 L/min, respectively. Schöps et al.3 used computational fluid dynamics to describe zones of recirculation, which do not contribute to the bulk flow of the blood, within adult devices operated at 1 L/min or less. Consequently, blood remains in the pump shear field for an extended time, significantly increasing the risk for hemolysis, which was demonstrated in vitro. Vermeer et al.1 have now provided in vivo evidence for the in silico and in vitro data provided by Schöps et al.3 However, we believe there is still more to the story. Several other factors are likely at play in neonatal ECLS, including cellular fragility4 and hemoglobin greater than 12–13 g/dl, which has been shown to be an independent factor leading to blood damage for neonatal ECLS regardless of pump type.5 A shrinking pool of devices and trained healthcare providers has led to the utilization of devices for the greatest number of patients (adults) in the most vulnerable population (neonates), and these are conditions for which these devices were never designed. Even the PediVAS, which is a pediatric device, is designed to operate at flow nearly three times that required for neonatal ECLS. We believe the study described by Vermeer et al.,1 Schöps et al.,3 Jenks et al.,5 and others in this space are important starting points for the ultimate discussion that a true neonatal ECLS centrifugal blood pump needs to be developed. We hope that such study continues and drives manufacturers to consider the needs of this population, ultimately allowing clinicians to more safely provide the life-saving support that this population needs.

Could homocysteine represent a negative acute phase reactant in canine infections-a pilot study?

Homocysteine (Hcy) was investigated as the biomarker of cardiac, renal, and gastrointestinal disorders in dogs. Data about low Hcy concentrations in the systemic inflammatory response syndrome raised a hypothesis that Hcy in dogs is a negative acute-phase reactant. This survey compared Hcy concentrations, serum amyloid A (SAA), and the routine laboratory parameters between healthy (HD, N=6) and dogs with inflammation of different extent: mild ( dirofilariosis (DIR), N=31), moderate (babesiosis (BAB), N=12), and severe (pyometra (PYO), N=8). The BAB and PYO groups had lower Hcy er than HD. Also, the levels in the PYO group were below those in the DIRO group. SAA had the inverse pattern. Across the groups, Hcy and SAA levels correlated negatively (ρ = -0.502, P&lt;0.001). Hcy and SAA correlated with the erythrocyte count, hematocrit, hemoglobin and mean cellular hemoglobin concentrations, and neutrophil count, with correlations being positive for Hcy and negative for SAA. Among all dogs, hemoglobin was the only independent predictor of Hcy concentration. Hcy levels in canine infections, decreased as acute-phase reaction (APR) intensified. Also, they were related with the hematology changes accompanying the APR. Further studies will establish the clinical potential of these alterations.

Biological Variation and Reference Change Value of Routine Hematology Measurands in a Population of Managed Bottlenose Dolphins (Tursiops truncatus).

Hematological analyses are particularly useful in assessing a dolphin's health status. However, the creation of appropriate reference intervals for this species is difficult due to the low number of reference individuals. The implementation of individual reference intervals (iRIs) allows researchers to overcome this limitation and, moreover, also consider the within-individual variability. The aims of this study were (1) to evaluate the biological variations in some hematological measurands, including erythrocytes (RBC), hematocrit (Hct), mean cellular volume and hemoglobin content (MCV and MCHC, respectively), RBC distribution width (RDW), leukocytes (WBC), and platelets (PLT); and (2) to calculate the index of individuality (IoI) and reference change value (RCV), which enable the production of iRIs, in healthy managed bottlenose dolphins. Seven dolphins were included, and the results of six hematological exams were analyzed for each animal. Analytical imprecision (CVa), within-dolphin variation (CVi), and between-dolphins variations (CVg) were calculated, and the IoI and RCV were derived for each measurand. All the hematological measurands had intermediate IoI except WBC, for which Iol was low. The calculated RCV ranged from 10.33% (MCV) to 186.51% (WBC). The results reveal that the majority of hematological measurands have an intermediate level of individuality in dolphins, and thus the application of iRIs is appropriate. The calculated RCV can also be applied to other managed dolphins and could be useful in interpreting serial CBC exams.

Blood Variation Implicates Respiratory Limits on Elevational Ranges of Andean Birds.

AbstractThe extent to which species ranges reflect intrinsic physiological tolerances is a major question in evolutionary ecology. To date, consensus has been hindered by the limited tractability of experimental approaches across most of the tree of life. Here, we apply a macrophysiological approach to understand how hematological traits related to oxygen transport shape elevational ranges in a tropical biodiversity hot spot. Along Andean elevational gradients, we measured traits that affect blood oxygen-carrying capacity-total and cellular hemoglobin concentration and hematocrit, the volume percentage of red blood cells-for 2,355 individuals of 136 bird species. We used these data to evaluate the influence of hematological traits on elevational ranges. First, we asked whether the sensitivity of hematological traits to changes in elevation is predictive of elevational range breadth. Second, we asked whether variance in hematological traits changed as a function of distance to the nearest elevational range limit. We found that birds showing greater hematological sensitivity had broader elevational ranges, consistent with the idea that a greater acclimatization capacity facilitates elevational range expansion. We further found reduced variation in hematological traits in birds sampled near their elevational range limits and at high absolute elevations, patterns consistent with intensified natural selection, reduced effective population size, or compensatory changes in other cardiorespiratory traits. Our findings suggest that constraints on hematological sensitivity and local genetic adaptation to oxygen availability promote the evolution of the narrow elevational ranges that underpin tropical montane biodiversity.

SARS-CoV-2 Altered Hemorheological and Hematological Parameters during One-Month Observation Period in Critically Ill COVID-19 Patients.

Hematological and hemorheological parameters are known to be altered in COVID-19; however, the value of combined monitoring in order to deduce disease severity is only scarcely examined. A total of 44 acute SARS-CoV-2-infected patients (aCOV) and 44 age-matched healthy controls (Con) were included. Blood of aCOV was sampled at admission (T0), and at day 2 (T2), day 5 (T5), day 10 (T10), and day 30 (T30) while blood of Con was only sampled once. Inter- and intra-group differences were calculated for hematological and hemorheological parameters. Except for mean cellular volume and mean cellular hemoglobin, all blood cell parameters were significantly different between aCOV and Con. During the acute disease state (T0-T5), hematological and hemorheological parameters were highly altered in aCOV; in particular, anemic conditions and increased immune cell response/inflammation, oxidative/nitrosative stress, decreased deformability, as well as increased aggregation, were observed. During treatment and convalescence until T30, almost all abnormal values of aCOV improved towards Con values. During the acute state of the COVID-19 disease, the hematological, as well as the hemorheological system, show fast and potentially pathological changes that might contribute to the progression of the disease, but changes appear to be largely reversible after four weeks. Measuring RBC deformability and aggregation, as well as oxidative stress induction, may be helpful in monitoring critically ill COVID-19 patients.

Dietary Iron Restriction Alters Microbiota-Host Crosstalk and Protects Against Vaso-Occlusion and Organ Damage in Sickle Cell Disease Mice

Introduction: Sickle cell disease (SCD) is the most commonly inherited monogenic hemoglobinopathy and results from a mutation in the β-globin gene (HbS), leading to red blood cell (RBC) sickling, vaso-occlusive episodes (VOE), and organ damage. Chronic hemolysis, inflammation, and repeated red blood cell transfusions in SCD can disrupt iron homeostasis. Patients who receive multiple blood transfusions develop iron-overload, and another subpopulation of SCD patients manifest iron deficiency. SCD mice exhibit increased iron absorption via upregulation of the intestinal transcription factor hypoxia inducible factor-2a(HIF-2a) leading to the development of iron overload. Disruption of intestinal HIF-2a or limiting iron intake by dietary iron restriction in SCD mice, significantly reduced systemic iron overload, lowered cellular sickle hemoglobin, and consequently improved hemolysis and ameliorated anemia. Our lab previously demonstrated that microbial signals from the gut also participated in initiation of VOE and organ damage in SCD mice. To elucidate connections between dietary iron, the microbiota-gut axis, and SCD pathogenesis, we hypothesize that dietary iron restriction may reduce microbiota load and gut permeability, and contributes to amelioration of acute and chronic complications in SCD mice. Methods: SCD mice were generated by transplanting bone marrow cells from Berkeley SCD (Tg[Hu-miniLCRα1GγAγδβS]Hba−/− Hbb−/−) mice into 8-week old, lethally-irradiated C57BL/6J mice. Immediately after transplantation, SCD mice were fed either a regular-iron diet (Ctrl) containing 185 parts per million (ppm) iron or an otherwise identical iron-restricted diet (IRD) containing 3 ppm iron. VOE and organ damage analysis were conducted 3-4 months after treatments. Results: IRD resulted in a reduced organ iron concentration in SCD compared to Ctrl group(0.6±0.1 v.s. 4.0±0.7ug iron/mg liver tissue; P=0.001), confirming that dietary iron restriction reverses organ iron deposition in SCD mice. SCD mice treated with IRD exhibited reduced MCHC (19.7±0.3 v.s. 20.1±0.2 pg; P=0.006) and serum indirect bilirubin (0.4±0.04 v.s. 0.7±0.07 mg/dL; P=0.001) indicating a decrease in cellular sickle hemoglobin and hemolysis. Furthermore, IRD reduced the fraction of aged neutrophils by >50% in SCD mice which corresponded with improved VOE parameters (RBC velocity: 2.0±0.2 v.s. 1.6±0.1mm/s; P=0.01; blood flow rate: 314±33 v.s. 230±13pL/s; P=0.01; leukocyte adhesion: 10±1 v.s. 14±1cells/100um vein; P=0.01; IRD v.s. Ctrl). Importantly, IRD led to prolonged survival after TNF-a challenge in SCD mice (Fig1). IRD resulted in a dramatic reduction in lung leukocyte infiltration, indicating significantly reduced lung inflammation in SCD mice (Fig2A). Compared to Ctrl liver samples, IRD resulted in significantly less liver necrosis, leukocyte infiltration and fibrosis (Fig2B). We further hypothesized that, in addition to reducing iron concentration in organs, IRD ameliorates organ damage by preventing translocation of microbial compounds. Microbiota load in IRD treated SCD mice were decreased to 25% of Ctrl level. IRD inhibited translocation of FITC-Dextran by 5-fold compared to Ctrl suggesting functionally improved gut permeability. We detected significantly decreased TLR2 ligands in serum of IRD compared to Ctrl mice (0.31±0.04 v.s. 0.46±0.05 TLR2 ligand activities O.D. 650nm; P=0.04) suggesting ligands from gram-positive bacteria (TLR2 agonists) may play a role in triggering SCD complications. Conclusion: Our study demonstrates first time that dietary iron restriction reduces VOE severity and alleviates chronic organ damage in SCD mice. These findings provide strong evidence for the therapeutic potential of manipulating iron homeostasis and the gut microbiome to ameliorate SCD pathophysiology. Many treatments under development focus on lowering the systemic iron concentration to relieve disease complications, and our data suggest that iron-induced changes in microbiota load and gut integrity are related and novel therapeutic targets. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Results of Randomized Comparative Efficacy Study of Iron Deficiency Anemia Management in Children with Iron III Hydroxide Polymaltose Complex and Iron Sulfate

Background. The guidelines on selection between iron preparations, iron sulfate (IS) and iron III hydroxide polymaltose complex (HPC), for iron deficiency anemia (IDA) management in pediatrics are contradictory. Objective. The aim of the study is to compare efficacy and safety of iron (III) HPC and IS for IDA treatment in children. Materials and methods. Randomization of children (aged from 1 month to 18 years) with IDA into 2 therapeutic groups was implemented in 2019-2020. on pediatric districts of children’s city outpatient's clinic: IS dosage of 3 mg/kg/day and iron (III) HPC dosage of 5 mg/kg/day. Hemogram monitoring was performed every 14 days. The efficacy was compared according to the rate of Hb level increase, erythrocytes indices, and serum ferritin (SF) level at the recorded moment of Hb normalization. For the safety — the rate of adverse effects. Results. 65 children with IDA were included in the study and randomized into 2 groups, the median age was 2.3 years (1st; 3rd quartile = 1.1; 4.3). Hb level increase was comparable in both groups (0.34 (0.23; 0.48) g/L/day and 0.24 (0.21; 0.30) g/L/day, р = 0.891). IS is more effective than iron (III) HPC in terms of average red blood cell volume by the time of Hb normalization (83.5 (80.0; 86.7) fl against 80.7 (79.0; 82.0) fl, р = 0.006), and mean cellular hemoglobin content (28.9 (SD = 2.0) pg against 27.4 (SD = 1.8) pg, р = 0.004). The timing of SF levels normalization did not differ. Adverse effects to iron III HPC treatment occurred 5.5 times more frequently than to IS (p = 0.0001). Conclusion. The efficacy of IS and iron III HPC at standard doses is comparable. The advantage in tolerability and recovery of erythrocytic indices justifies the feasibility of using IS preparations in the first line of treatment for children with IDA.

PI-10: DIETARY IRON RESTRICTION PROTECTS AGAINST VASO-OCCLUSION AND ORGAN DAMAGE IN SICKLE CELL DISEASE

Purpose: Sickle cell disease (SCD) is the most commonly inherited monogenic hemoglobinopathy and results from a mutation in the β-globin gene (HbS), leading to red blood cell (RBC) sickling, vaso-occlusive episodes (VOE), and organ damage. Chronic hemolysis, inflammation, and repeated red blood cell transfusions in SCD can disrupt iron homeostasis. Patients who receive multiple blood transfusions develop iron-overload, and another subpopulation of SCD patients manifest iron deficiency. SCD mice exhibit increased iron absorption via upregulation of the intestinal transcription factor hypoxia inducible factor-2a(HIF-2a) leading to the development of iron overload. Disruption of intestinal HIF-2a or limiting iron intake by dietary iron restriction in SCD mice, significantly reduced systemic iron overload, lowered cellular sickle hemoglobin as reflected by reduced RBC mean corpuscular hemoglobin concentration (MCHC), and consequently improved hemolysis and ameliorated anemia. Understanding iron homeostasis is therefore critical to SCD pathogenesis and holds the promise for identifying novel approaches to treat SCD. Materials and methods: SCD mice were generated by transplanting bone marrow cells from Berkeley SCD (Tg[Hu-miniLCRα1GγAγδβS]Hba−/− Hbb−/−) mice into 8-week old, lethally-irradiated C57BL/6J mice. Immediately after transplantation, SCD mice were fed either a regular-iron diet (Ctrl) containing 185 parts per million (ppm) iron or an otherwise identical iron-restricted diet (IRD) containing 3 ppm iron. VOE and organ damage analysis were done 3-4 months after treatments. Results: IRD resulted in a reduced organ iron concentration (Fig-1A) in SCD compared to Ctrl group, confirming that dietary iron restriction ameliorates organ iron deposition in SCD mice. SCD mice treated with IRD exhibited reduced MCHC (Fig-1B) and serum indirect bilirubin(IDBILC) (Fig-1C) indicating a decrease in cellular sickle hemoglobin and hemolysis. Furthermore, IRD reduced the fraction of aged neutrophils by >50% in SCD mice (Fig-1D), which corresponded with improved VOE parameters. Specifically, intravital microscopy (IVM) demonstrated elevated mean centerline RBC velocity and blood flow rate and decreased leukocyte adhesion in IRD compared to Ctrl mice (Fig-1E-G). Importantly, IRD led to prolonged survival after TNF-a challenge in SCD mice (Fig-1H). IRD resulted in a dramatic reduction in lung leukocyte infiltration, indicating significantly reduced lung inflammation in SCD mice (Fig-2A). Compared to Ctrl liver samples, IRD resulted in significantly less liver necrosis, leukocyte infiltration and fibrosis (Fig-2B,C), decreased serum liver enzymes (ALT, AST; Fig-2D), and reduced levels of total and direct bilirubin (TBILC, DBILC; Fig-2D).We further hypothesized that, in addition to reducing iron concentration in organs, IRD ameliorates organ damage by preventing translocation of microbial compounds.IRD inhibits translocation of FITC-Dextran by 5-fold compared to Ctrl suggesting functionally improved gut permeability (Fig-2E). We detected significantly decreased TLR2 ligands in serum of IRD mice (Fig-2F) suggesting ligands from gram-positive bacteria (TLR2 agonists) may play a role in triggering SCD complications. Conclusion: Our study demonstrates for the first time that dietary iron restriction reduces VOE severity and alleviates chronic organ damage in SCD mice. Novel approaches that target iron availability/absorption in the gut, the gut microbiome and aged neutrophil crosstalk and gut barrier integrity, provide an important area of further investigation with the potential of improving hemolysis, VOE, and organ damage in SCD patients.(A)The bone-marrow/spleen/liver Non-heme iron content. (B)MCHC, (C)IDBILC,(D)aged neutrophils percentage in Ctrl/IRD SCD mice. IVM parameters (E) (Vrbc) (F) blood flow rate (J) the number of adherent leukocytes and (J) survival rate after TNF-a injection(A)Leukocyte infiltration in lung and (B)liver tissue. (C)Liver necrosis, leukocyte infiltration, and fibrosis. (D)Liver ALT and AST, TBILC, and DBILC from sera. (E)serum FITC-Dextran and (F) TLR2 ligands activity in Ctrl/IRD SCD mice The authors do not declare any conflict of interest