Total Hb Concentration Research Articles

Study on the Effects of Acupuncture with the "Yizhi Tiaoshen" Acupoint Formula on Blood Oxygen Metabolism and Neural Function in Key Brain Regions of AD Rats.

Exploring the effects of acupuncture at the "Yizhi Tiaoshen" acupoint on blood oxygen metabolism and neurological function changes in the brain regions of AD model rats. The AD model was replicated by intraperitoneal injection of D-galactose combined with bilateral hippocampal CA1 injection of Okadaic acid (OA). Thirty rats with successfully replicated model were selected through Morris water maze experiment and randomly divided into model group, donepezil hydrochloride group, and acupuncture group, with 10 rats in each group. After treatment, fNIRs were used to detect changes in Oxy Hb, Deoxy Hb, and Total Hb in the cerebral cortex of rats in each group, in order to evaluate the neurological function changes in key brain areas. The escape latency of the donepezil hydrochloride group and the acupuncture group was shortened, the number of crossings through the original platform increased, and the duration of stay in the quadrant where the original platform was located was prolonged. Based on fNIRs detection, the main differential channels of blood oxygen metabolism in AD rats were identified as 2-2 and 8-7, corresponding to the prefrontal and parietal lobes, respectively. The concentrations of Oxy Hb and Total Hb were significantly increased in both treatment groups, while the concentration of Deoxy Hb was significantly decreased. Acupuncture with the "Yizhi Tiaoshen" acupoint formula and donepezil hydrochloride can improve the learning and memory function of AD rats, and its mechanism may be related to improving blood oxygen metabolism in the prefrontal and parietal regions and protecting neuronal function.

Biomarker dosimetry of acute low level of thermal neutrons and radiation adaptive response effect on rats

In this paper, we demonstrated the biological effects of acute low-dose neutrons on the whole body of rats and investigated the impact of that level of neutron dose to induce an in vivo radio-adaptive response. To understand the radio-adaptive response, the examined animals were exposed to acute neutron radiation doses of 5 and 10 mSv, followed by a 50 mSv challenge dose after 14 days. After irradiation, all groups receiving single and double doses were kept in cages for one day before sampling. The electron paramagnetic resonance (EPR) method was used to estimate the radiation-induced radicals in the blood, and some hematological parameters and lipid peroxidation (MDA) were determined. A comet assay was performed beside some of the antioxidant enzymes [catalase enzyme (CAT), superoxide dismutase (SOD), and glutathione (GSH)]. Seven groups of adult male rats were classified according to their dose of neutron exposure. Measurements of all studied markers are taken one week after harvesting, except for hematological markers, within 2 h. The results indicated lower production of antioxidant enzymes (CAT by 1.18–5.83%, SOD by 1.47–17.8%, and GSH by 11.3–82.1%). Additionally, there was an increase in red cell distribution width (RDW) (from 4.61 to 25.19%) and in comet assay parameters such as Tail Length, (from 6.16 to 10.81 µm), Tail Moment, (from 1.17 to 2.46 µm), and percentage of DNA in tail length (DNA%) (from 9.58 to 17.32%) in all groups exposed to acute doses of radiation ranging from 5 to 50 mSv, respectively. This emphasizes the ascending harmful effect with the increased acute thermal neutron doses. The values of the introduced factor of radio adaptive response for all markers under study reveal that the lower priming dose promotes a higher adaptation response and vice versa. Ultimately, the results indicate significant variations in DNA%, SOD enzyme levels, EPR intensity, total Hb concentration, and RDWs, suggesting their potential use as biomarkers for acute thermal neutron dosimetry. Further research is necessary to validate these measurements as biodosimetry for radiation exposure, including investigations involving the response impact of RAR with varied challenge doses and post-irradiation behavior.

Clinical, seroepidemiological and hematological study of Buffaloes Babesiosis in Basrah, Iraq

Objective: Babesiosis is one of the important diseases in cattle and buffaloes, the current study conducting to diagnoses and evaluate the infection by B.pigeamina in buffaloes during the period of November 2022 to September 2023. Methods: The study included 210 animals of both sexes and varied ages. The blood sampling from 220 water buffaloes (Bubalus Bubalis) in the north of Basrah governorate during the period of November 2022 to September 2023; the samples collected from both sexes and different ages ranged between 1 to 5 years. At these ten months 20 blood samples was randomly collected each month to compared with the control 10 animals. Results: The blood smear from infected buffaloes show the parasite clearly while the clinical signs reported as: babesiosis confirm 16% (32 affected animals) represented by fever, emaciation, anorexia, low milk production, jaundice of mucous membrane, corneal opacity and hemoglobinuria. The subclinical infected buffaloes which represented 41% (82 affected animals) appear healthy with no clinical signs. Also indicate that were no significant differences (P less than 0.05) between the infected and control group. The hematological parameters show significant differences (P less than 0.05) in RBCs count, total WBC count, deferential LC, Hb concentration and PCV in the infected buffaloes compare to the control animals while there were no significant differences (P less than 0.05) between the subclinical and control buffaloes except the RBCs count. The serological examination revealed to significant differences (P less than 0.05) in the infected group in posit to the control especially the liver enzymes and kidney function test enzymes which include: AST, ALT, ALP, BUN, creatinine and TP. Moreover, According the season and months the study detects the high infection rate in the summer while there were decrease in the infection rate in the mild or cold weather months. Moreover, the study indicates the high infection percentage in females compare to the male sex. Conclusion: Babesiosis is very important disease in Buffaloes in north of Basrah and there are no significant differences in infection rates between male and female.

Reversal of Rocuronium-Induced Muscle Relaxation with Sugammadex Enhances Oxygen Metabolism in Skeletal Muscle.

We measured changes in blood flow and oxygenation in the brachioradialis muscle using near-infrared spectroscopy (NIRS) during reversal of rocuronium-induced muscle relaxation with administration of sugammadex in patients (n=25) under general anaesthesia, to investigate whether reversal of muscle relaxant-induced muscle relaxation increases oxygen metabolism in skeletal muscle under general anaesthesia. NIRS measurements, including oxy-haemoglobin (Hb), deoxyHb, total Hb concentration, tissue oxygen index, and various cardiopulmonary parameters, were recorded at four timepoints: T0 (baseline), 3 min before sugammadex administration; T1, immediately before sugammadex administration; T2, at complete recovery of muscle contractility; and T3, 3 min after T2. All measured values at each timepoint were compared using multiple comparison tests. The median values (quartile deviation; QD) (μmol/L) of oxyHb and deoxyHb at T0, T1, T2, and T3 were 0, -0.01 (0.14), -1.15 (0.54), and -1.52 (0.54), and 0, 0.11 (0.06), 0.86 (0.5), and 1.36 (0.61), respectively. The levels of oxyHb were significantly lower and those of deoxyHb were significantly higher at T2 and T3 when compared to those at T1, respectively (P<0.01). There were no significant changes in other measurements. These results suggest that reversal of rocuronium-induced muscle relaxation by sugammadex slightly increases oxygen metabolism in the brachioradialis muscle. This study might support the clinical finding that administration of neuromuscular blockers decreases whole body oxygen consumption in patients receiving mechanical ventilation under general anaesthesia.

Myelodysplastic Syndrome Red Blood Cell Physiology Abnormalities and Their Correlation with Disease Risk and Therapeutic Interventions

PV and KT as well as VP and MA contributed equally Introduction: Myelodysplastic syndromes (MDS) are clonal hematopoietic disorders characterized by ineffective hematopoiesis, different degree of cytopenias, and increased risk of transformation to AML. Anemia constitutes a main clinical manifestation contributing to increased morbidity and mortality. It is currently managed with red blood cell (RBC) transfusions, erythropoiesis stimulating agents (EPO), and recently,with luspatercept. The latter enhances late stage erythropoiesis and is indicated for Ring Sideroblasts (RS) MDS resistant to EPO. Several lines of evidence suggest that MDS pathophysiology is associated with structural and functional distortions of mature red cell (RBCs), such as hemolysis that in rare instances include acquired lesions like hemoglobinopathies and membranopathies.These abnormalities add variability to the clinical MDS phenotypes and could serve as biomarkers for early diagnosis or monitoring. However, they have been scarcely investigated. Patients and methods: We examined 40 MDS patients and 12 healthy sex- and age-matched controls. According to the IPSS-Rrisk stratification system, 8 patients were High Risk (HR) and 32 patients Low Risk (LR)MDS, of whom 8 under EPO (EPO-LR) and 4 under luspatercept (luspatercept-LR) treatment. None of them had received RBC transfusions in the past three months. Following full blood count analyses and RBC fractionations, we measured hemolysis (basal, osmotic, mechanical, and oxidative), eryptosis and oxidative stress, as well as the antioxidant capacity of the plasma through spectrophotometry,fluorometry and flow cytometry. Results: Compared to controls, the cohort of MDS patients was characterized by anemia (Hb 10.7 vs 13.6g/dL) and significantly higher HDWand RDW (17.5 vs. 13.4%) indexes. RBCs exhibited signs of eryptosis (RBC calcium concentration 7063±1443 vs. 6027±1541 RFU/mg; PS surface exposure 0.61±0.45 vs. 0.33±0.17%) and oxidative stress (higher metHb/oxyHb ratio; membrane-bound hemichromes 45.0±15.3 vs. 30.2±13.3). About 18% of the patients presented spontaneous hemolysis. Clinical severity correlated with the levels of cytopenias, and oxidative hemolysis. Anemia correlated positively with the age, and the levels of membrane-bound hemichromes, eryptosis and intracellular reactive oxygen species (ROS). According to ROC analysis, the combination of RDW, metHb/oxyHb ratio, and total Hb concentration is predictive of MDS, across all IPSS-R categories(area under the curve: 0.908; Figure 1). We then examined the subgroups of MDS patients. Apart from the RDW, the LR-MDS subgroup exhibited on average normal RBC features, in opposite to the more anemic and thrombocytopenicHR-MDS subgroup that showed pathologically increasedresistance to osmotic hemolysis, intracellular calcium (7946±1917 vs. 6026±1541RFU), and metHb/oxyHb ratio (18.5±7.3 vs. 10.4±3.9%), when compared to controls. Of note, similar abnormalities were observed in the EPO subgroup that further showed reticulocytosis, PS externalization, and membrane-bound hemichromes(p&amp;lt;0.05) Compared to the non-treated subgroup, the EPO-LR had lower WBC counts, but higher RDW, PS exposure (0.75±0.44 vs. 0.41±0.26%) and resistance to osmotic stress, aspreviously reported in other EPO-treated patients.Luspatercept-LR had higher WBC and PLT counts but exhibited EPO-like effects on RBCs, including pathological resistance to osmotic/mechanical lysis, ROS concentration (1347±453 vs. 960±160 RFU), Hb oxidation, and PS exposure (1.05±0.60 vs. 0.41±0.26%), compared to the non-treated LR subgroup. Conclusion: MDS RBCs are highly heterogeneous in terms of volume and Hb concentration and present signs of oxidative stress and eryptosis. Consequently, these easily accessible markers could be an accessory tool to the MDS diagnosis.In the context of ineffective hematopoiesis and increased apoptosis of precursor erythroid cells,the LR-MDS patients exhibit minimal RBC abnormalities. On the other hand, the expansion of BM blasts and the lower level of apoptosis in HR-MDS seem to affect RBCs in a hemoglobinopathy-like manner in terms of Hb oxidation, eryptosis signaling, and membrane ion channels. EPO and luspatercept treated LR patients exhibit improved cytopenias but worse RBC physiology most probably attributed to clinical factors like RS phenotype.

Observing temporal variation in hemolysis through photoacoustics with a low cost LASER diode based system

Patients under hemolytic condition need continuous monitoring of lysis as depletion of Red Blood Cells (RBC) and the presence of antioxidant free hemoglobin (Hb) in excess amount due to hemolysis lead to severe deterioration of their health. Out of many modalities, Photoacoustics (PA) offers real time information noninvasively from deep lying blood vessels since Hb is the strongest chromophore in mammalian blood and the PA response of blood varies with the amount of Hb present. During hemolysis, total Hb content in blood however remains unchanged, thus, questions the use of PA in hemolysis detection. In this report, a hypothesis that the amplitude of the PA signal would not change with the amount of lysis is framed and tested by applying osmotic shock to the RBCs in hypotonic environment and the PA response is recorded over time using a low cost NIR based PA system. The experimental outcome indicates that PA amplitude falls off as lysis progresses in course of time consequently rejecting the hypothesis. The decaying PA response also carries the signature of RBC swelling during the early phase of lysis. The PA measurement can detect hemolysis as low as 1.7%. These findings further advocate transforming this NIR-PA system into a portable, noninvasive patient care device to monitor hemolysis in-vivo.

Serum vascular endothelial growth factor affects tissue fluid accumulation and is associated with deteriorating tissue perfusion and oxygenation in severe sepsis: a prospective observational study

BackgroundPositive fluid balance and tissue fluid accumulation are associated with adverse outcomes in sepsis. Vascular endothelial growth factor (VEGF) increases in sepsis, promotes vascular permeability, and may affect tissue fluid accumulation and oxygenation. We used near-infrared spectroscopy (NIRS) to estimate tissue hemoglobin (Hb) oxygenation and water (H2O) levels to investigate their relationship with serum VEGF levels.Material and methodsNew-onset severe sepsis patients admitted to the intensive care unit were enrolled. Relative tissue concentrations of oxy-Hb ([HbO2]), deoxy-Hb ([HbR]), total Hb ([HbT]), and H2O ([H2O]) were estimated by near-infrared spectroscopy (NIRS) for three consecutive days and serum VEGF levels were measured. Comparisons between oliguric and non-oliguric patients were conducted and the correlations between variables were analyzed.ResultsAmong 75 eligible patients, compared with non-oliguric patients, oliguric patients were administrated more intravascular fluids (median [IQR], 1926.00 [1348.50–3092.00] mL/day vs. 1069.00 [722.00–1486.75] mL/day, p < 0.001) and had more positive daily net intake and output (mean [SD], 1,235.06 [1303.14] mL/day vs. 313.17 [744.75] mL/day, p = 0.012), lower [HbO2] and [HbT] over the three-day measurement (analyzed by GEE p = 0.01 and 0.043, respectively) and significantly higher [H2O] on the third day than on the first two days (analyzed by GEE p = 0.034 and 0.018, respectively). Overall, serum VEGF levels were significantly negatively correlated with [HbO2] and [HbT] (rho = − 0.246 and − 0.266, p = 0.042 and 0.027, respectively) but positively correlated with [H2O] (rho = 0.449, p < 0.001). Subgroup analysis revealed a significant correlation between serum VEGF and [H2O] in oliguric patients (rho = 0.532, p = 0.003). Multiple regression analysis determined the independent effect of serum VEGF on [H2O] (standardized coefficient = 0.281, p = 0.038).ConclusionsIn severe sepsis, oliguria relates to higher positive fluid balance, lower tissue perfusion and oxygenation, and progressive tissue fluid accumulation. Elevated serum VEGF is associated with worsening tissue perfusion and oxygenation and independently affects tissue fluid accumulation.

Apohemoglobin-Haptoglobin Complex Prevents Microvascular Dysfunction in Sickle Cell Mice: Potential Therapeutic Approach for Vaso-Occlusive Crisis

Background: Sickle cell disease (SCD) is associated with episodes of vaso-occlusive events (VOE) caused by the effects of hemoglobin S (HbS) polymerization in red blood cells (RBCs) and resultant hemolysis. In the acute setting, treatment for VOE is limited to supportive care such as hydration and aggressive pain management as no specific therapies can alter the course of acute VOE. The apohemoglobin-haptoglobin (apoHb-Hp) complex is a two-protein formulation that is capable of scavenging and neutralizing the toxic effects of cell-free hemoglobin (Hb) and heme released from the hemolysis of polymerized HbS RBCs. The primary objective of this study was to evaluate the microcirculatory effects of apoHb-Hp during hypoxia-induced VOE. Methods: The effects of apoHb-hp was studied in a mouse model of SCD using sickle cell transgenic mice (HbSS-Towne's mice, and compared to mice expressing normal human Hb (HbAA-Towne's mice). Each group consisted of 11 mice, they were initially kept in normal conditions (21% O2) for an hour followed by a hypoxic state (7% O2) for another hour and finally brought back to normal conditions (21% O2) for an hour before being sacrificed. ApoHb-hp (60 mg/mL) and Hp (85 mg/mL) were injected intravenously to yield equimolar total heme and Hb binding capacity between treatments prior to hypoxia exposure. During baseline, hypoxia and normoxia systemic parameters (mean arteriole pressure and heart rate), blood gases (pO2,A, pO2,v, sO2,A, sO2,v), hematological properties (total Hb concentration, hematocrit, and concentration of lactate) and microcirculatory hemodynamics (velocity, diameter, flow and functional capillary density) were measured. After the mice were sacrificed a count of present macrophages, neutrophils and lymphocytes were quantified in the pulmonary tissue. Results: There was no notable changes in blood gas concentrations with any of the treatments. ApoHb-Hp treatment improved microcirculatory hemodynamics of HbSS mice by preventing hypoxia-induced vasoconstriction and reduction of blood flow in arterioles <40µm (P=0.818, P<0.0001), arterioles >40 µm (P>0.9999, P<0.0001) and venules 20<x<60 µm (P=0.150, P=0.330) compared to HbAA mice. Furthermore, the functional capillary density saw favorable recovery to baseline when HbSS mice were treated with apoHb-Hp seeing a 36% drop initially, 50% drop after 24 hours, 35% drop after 48 hours, and 27% drop after 72 hours. For comparison, HbSS mice that didn't receive treatment saw a 50% drop initially, 77% drop after 24 hours, 80% drop after 48 hours, and 72% drop after 72 hours when compared to baseline. Conclusion: These results suggest a role for ApoHb-Hp complex in alleviating microvascular changes occurring during hypoxia mediated acute vaso-occlusion in SCD and merits further study for development as a potential therapeutic. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Methylation of the suppressor of Cytokine Signaling 1 Gene (SOCS1) in Philadelphia-negative myeloproliferative neoplasms.

Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway activation is initiated by mutations in the JAK2 gene. This activation is in turn, a vital pathogenetic mechanism in myeloproliferative neoplasms (MPNs). However, several factors affect the pathogenesis of MPNs other than the JAK2 gene mutations, such as the downregulation of cytokine signaling (SOCS) proteins, which are potent inhibitors of the JAK/STAT pathway. Therefore, we hypothesized that the regulation of SOCS protein system might be a possible pathogenetic mechanism of MPNs through activating the JAK/STAT pathway. Our study aimed to investigate the status of the Suppressors of cytokine signaling 1 (SOCS1) in 125 MPNs specimens at the level of mutated points. The acquired mutations, aberrant expression, and/or CpG island hypermethylation of SOCS1 were analyzed among Philadelphia-negative myeloproliferative neoplasm patients. SOCS1 was identified in 20.0% of all patients with Philadelphia-negative myeloproliferative neoplasm. At the diagnosis, the prevalence of methylation was 41.0% for Polycythaemia Vera (PV), 27.7% for Essential Thrombocythaemia (ET), and 6.6% for Primary Myelofibrosis (PMF). The methylation was not detected in 20 healthy adult people. A significant association was found between disease groups (p=.077). The presence of methylated SOCS1 was found to be significantly correlated with age (p=.005), total RBCs count (p=.019), hemoglobin (Hb) concentration (p=.002), and Hematopoietic cell transplant (HCT) (p=.007) in PV patients. However, the presence of methylated SOCS1 was found to be significantly associated with age (p=.012), total RBCs count (p=.022), Hb concentration (p=.024), HCT (p=.033), and platelets count (p=.037) in ET patients. Moreover, the presence of methylated SOCS1 was significantly associated with Hb concentration (p=.046) and HCT (p=.040) in PMF patients. We concluded that the activation of the JAK/STAT signaling pathway in alternative or with JAK2 mutations leads to SOCS1 hypermethylation, which could represent a potential therapeutic target.

Increased tissue water in patients with severe sepsis affects tissue oxygenation measured by near-infrared spectroscopy: a prospective, observational case-control study.

BackgroundTissue oedema affects tissue perfusion and interferes with the monitoring of tissue oxygenation in patients with severe sepsis. However, the underlying mechanisms remain unclear. We used a wireless near-infrared spectroscopy (NIRS) device that transmits tri-wavelength light to quantify tissue haemoglobin (Hb) and water (H2O) content. We estimated tissue H2O in severe sepsis patients and healthy controls, compared their difference, and investigated the correlation of tissue H2O with systemic haemodynamics and its impact on tissue oxygenation.MethodsSeventy-seven adult patients with new-onset severe sepsis admitted to the intensive care unit within 72 h and 30 healthy volunteers (controls) were enrolled. The NIRS device was placed on the participant’s leg to estimate the relative tissue concentrations of oxy-Hb ([HbO2]), deoxy-Hb ([HbR]), total Hb ([HbT]), and H2O ([H2O]) at rest for three consecutive days. Two-sample t-test or Mann-Whitney U test, chi-square test, and generalised estimating equations (GEEs) were used for comparisons.ResultsIn severe sepsis patients, the [H2O] in the anterior tibia was higher [mean (standard deviation, 95% confidence interval), 10.57 (3.37, 9.81–11.34) vs. 7.40 (1.89, 6.70–8.11)] and the [HbO2], [HbT], and tissue Hb oxygen saturation (StO2) were lower [0.20 (0.01, 0.20–0.20) vs. 0.22 (0.01, 0.22–0.23), 0.42 (0.02, 0.42–0.43) vs. 0.44 (0.02, 0.44–0.45), and 47.25% (1.97%, 46.80–47.70%) vs. 49.88% (1.26%, 49.41–50.35%), respectively] than in healthy controls in first-day measurements. GEE analysis revealed significant differences in [H2O], [HbO2], [HbT], and StO2 between groups over three consecutive days (all P≤0.001). In addition, [HbO2] and StO2 levels gradually decreased over time in the patient group. A negative correlation was observed between [H2O] and [HbO2] and StO2, which became more obvious over time (day 1: r=−0.51 and r=−0.42, respectively; both P<0.01; day 3: r=−0.67 and r=−0.63, respectively, both P<0.01). Systolic arterial pressure was positively related to [H2O] (r=0.51, P<0.05, on day 1) but was not associated with tissue oxygenation parameters.ConclusionsNIRS can be used to quantify tissue H2O. Severe sepsis patients have increased tissue H2O, which responds to changes in arterial blood pressure and affects tissue oxygenation.

Apohemoglobin‐haptoglobin (Apohb‐Hp) Complex as a Vasculo‐protective Agent in Hypoxia Induced Vaso‐occlusion in Sickle Cell Disease

Sickle Cell Disease is associated with episodes of vaso‐occlusive crises (VOCs) caused by hemoglobin S (HbS) polymerization and resultant microvascular endothelial dysfunction due to hemolysis, inflammation, defects in nitric oxide metabolism and oxidative stress. In the acute setting, care for VOC involves supportive care including hydration and aggressive pain management. There are no specific therapies known to alter the course of the disease during an acute VOC. Apohemoglobin‐haptoglobin (Apohb‐Hp) complex is a two‐protein formulation that scavenges free hemoglobin (Hb) as well as free heme and has been shown to have a role in alleviating vasoconstriction and hypertension induced by cell free Hb1. The objective of this study was to evaluate microcirculatory effects of Apohb‐Hp during hypoxia induced vaso occlusion in sickle cell mice.We used intravital microscopy on the dorsal skin fold to observe the microcirculatory effects of Apo Hb‐Hp, haptoglobin (Hp), vehicle control in HbSS‐Townes mice (6‐8 week and ~25 g) and Hb AA mice. Each group consisted of 11 mice, they were initially kept in normal conditions (21% O2) for an hour followed by a hypoxic state (7% O2) for another hour and finally brought back to normal conditions (21% O2) for an hour before being sacrificed. In some cases, some mice were followed for 72 hours before being sacrificed. ApoHb‐Hp and Hp were injected intravenously at a concentration of 59 mg/mL and 86 mg/mL respectively at a single dosing quantity of 150 mg/kg. The Hb binding capacity of Apohb‐Hp was 15 mg/mL and the heme binding capacity was 0.5 mg/mL, the Hb binding capacity of Hp was 30 mg/mL with no heme binding capacity. During baseline, hypoxia and normoxia systemic parameters (mean arteriole pressure and heart rate), blood gases (pO2,A, pO2,v, sO2,A­, sO2,v), hematological properties (total Hb concentration, hematocrit, and concentration of lactate) and microcirculatory hemodynamics (velocity, diameter, flow and functional capillary density) were measured. After the mice were sacrificed a count of present macrophages, neutrophiles and lymphocytes were quantified in the plasmaThere was no notable changes in blood gas concentrations with any of the treatments. Apohb‐hp treatment treated animals showed an improvement in microcirculatory dynamics. Apohb‐Hp treated animals showed a statically higher diameter in arterioles (&lt;40 um and &gt;40 um) as well as venules with a diameter between 20‐60 um when compared to controls and hp treated mice, suggesting that treatment prevented vasoconstriction in arterioles. Apohb‐hp treatment also improved flow rates in arterioles and venules when compared to the control and hp treated mice. Furthermore, the functional capillary density of the mice followed for 72 hours after hypoxia showed that the mice treated with apohb‐hp had a greater number of functional capillaries compared to the control and hp treated group. Lastly, the amount macrophages, neutrophiles and lymphocytes decreased when treated with apohb‐hp illustrating another promising result as VOCs are generally associated with an active immune response.These results suggest a role for ApoHb‐Hp complex in alleviating microvascular changes occurring during hypoxia mediated acute vaso‐occlusion in SCD and merits further study for development as a potential therapeutic.

Hematological, serum biochemical, and histopathological evaluation following sub-chronic administration of Halea ciliata in experimental rats

Background: This study Background: Halea ciliata (Mitragyna ciliata Aubrev. Et Pellegr) also known as Abura leaves are common among the Yoruba groups of Nigeria for wrapping Kola nuts. The plants have been used traditionally to treat wide variety of conditions, however, there is limited information and literature reports concerning the toxicity and safety of this plant. The present study was conducted to evaluate the sub-chronic toxicity of the crude methanolic extract of Halea ciliata in Wistar rats. Methods: The methanolic extract of Halea ciliata was administered orally at doses of 200 and 400 mg/kg bw /day respectively for 3 weeks to healthy Wistar rats. At the end of the experimental period, certain haematological, serum biochemical and histopathological parameters were evaluated. Results: The activities of ALP and concentrations of total proteins, sodium, potassium, RBC count, PCV, PLT, and HB were not significantly (p&gt;0.05) different among all the groups, including the control. On the other hand, a significant (p&lt;0.05) increase in the WBC counts serum concentrations of urea and creatinine were observed when compared with the control group. A significant (p&lt;0.05) difference between the 200 mg/kg dose treatment group was observed when compared with the control group with respect to AST, and ALT activities. However, there was no difference between the control group and the highest treatment group of 400 mg/kg, with respect to ALT and AST. Conclusion: Administration of crude methanol extract of Halea ciliata at doses of 200 and 400 mg/kg BW significantly altered the serum urea and creatinine in rats. However, rats dosed with the same extracts at 400 mg/kg were devoid of biochemical anomalies in liver serum biomarkers when compared with the control counterpart. This study, therefore, suggested the use of Halea ciliata is relatively safe for the liver but its effect on kidney markers calls for further investigation.

Effects of paraquat dichloride on adult male wistar rat. an approach in the toxicity of body weights and hematological tissues

The main objective of the experiment is aimed to examine the impact of paraquat dichloride on the body weights of adult male Wistar rats as well as in their haematological tissues. The total number of male Wistar rats was 48 and was divided into 4 groups of 12 each. The 1st group (Test 1) served as control. The other three groups served as experimental groups in which the paraquat dichloride was administered. The experimental groups which are 2nd group (Test 2), 3rd group (Test 3) and 4th group (Test 4) received doses of paraquat dichloride of 10 mg/kg b.wt/per oral/day, 20 mg/kg b.wt/per oral/day and 40 mg/kg b.wt/per oral/day respectively. The experiment was carried out for a time duration of 42 days. The mean values of body weights, erythrocyte indices, total erythrocyte count (TEC), total leucocyte count (TLC) and Hb concentrations were significantly (P&lt;0.05) decreased in Test 2, 3 and 4 rats, excluding packed cell volume (PCV), which in comparison to the control groups. Outcomes and observations from hematological parameters and bone marrow revealed the toxicity of paraquat dichloride to hematological tissues resulting in anemia and bone marrow hypoplasia which eventually leads to failure of the bone marrow and extensively hematopoietic system of adult male Wistar rats following long term exposure and high concentration.

Clinical Results of the Drepaglobe Trial for Sickle Cell Disease Patients

▪In sickle cell disease (SCD), the β6 Glu→Val substitution leads to sickle hemoglobin (HbS) polymerization and red blood cell (RBC) sickling. Transplantation of autologous, genetically modified hematopoietic stem/progenitor cells (HSPCs) represents a promising therapeutic option for patients lacking a compatible donor.We previously designed a lentiviral vector (βAS3 LV) expressing a potent anti-sickling βAS3 globin and demonstrated its safety and efficacy in SCD patient cells (Weber et al., 2018). In vitro and in vivo preclinical studies demonstrated the safety and efficacy of a gene therapy (GT) protocol based on the efficient transduction of plerixafor-mobilized SCD HSPCs by βAS3 LV.Based on these results a Phase 1/2 clinical trial (NCT03964792) started in January 2020. Three homozygous SCD patients (P1, P2 and P3) were recruited. Before GT, patients suffered from severe SCD symptoms that were not controlled despite of a strict adherence to the treatment. Plerixafor-mobilized HSPCs were transduced achieving a VCN of 0.9±0.2 in liquid culture, 1.6±0.3 in BFU-E and 0.8±0.2 in CFU-GM. The patients were hypertransfused to reach HbS levels <30%. After busulphan-based myeloablative conditioning, they received a drug product 6 to 8.1x 10 6 per kg.Here, for the 3 patients we report the follow-up of 18 (P1), 10 (P2) and 6 (P3) months. No drug-related severe adverse events were observed. Importantly, no sign of clonal hematopoiesis was detected by NGS sequencing performed on the graft before and after transduction. Despite of the similar VCN in the drug product, gene marking in peripheral blood mononuclear cells was variable and ranged between 0.2 and 0.6, as measured at the last follow-up (0.4 in P1, 0.6 in P2 and 0.2 in P3). These results pinpoint the difficulty to estimate the gene marking, self-renewal and engraftment potential of HSCs in the grafts.P1 and P2 clinically benefit from the GT treatment despite they presented non-severe vaso-occlusive crises (VOCs) not requiring hospitalization (two for P1 at month 6 and 14 and one in P2 at month 8 post-GT). Despite the rapid resolution of these episodes, P1 started hydroxyurea at month 10 and stopped it at month 16. P2 initiated a phlebotomy program to treat liver iron overload and decrease viscosity. Intravascular hemolysis improved in P1 and P2, as shown by a decrease in plasma hemoglobin and heme, and an increase in hemopexin (HPX). Interestingly, in P1 a mild worsening occurred at the time of VOC. In P1 and P2, total bilirubin, a marker of extravascular hemolysis, decreased post-GT. Deformability, measured using osmoscan, improved in P1 and to a lesser extent in P2; however, both still exhibited some degree of dehydration compared to normal RBCs (Fig. 1). Adhesion to thrombospondin decreased after GT with an increase during VOC in P1 or prior the VOC in P2 (Fig. 1). In both patients, markers of inflammation (e.g., C reactive protein) and reticulocyte counts decreased upon GT. An in vitro sickling assays was performed for P1 and showed significant improvement, reaching a frequency of sickling cells similar to SCD heterozygous carriers (from 88% before GT to 45% 6 months post-GT).The GT treatment was not effective in P3 due to the modest intake and rapid decline of gene modified cells.In P1 and P2, who were no longer on transfusions, HbAS3 levels correlated with the VCN (2.1 g/dl for P1 and 3.3 g/dl for P2) and the therapeutic Hb accounted for 21.5%, and 28.7% of the total hemoglobins, respectively. For P3, who remained transfusion-dependent, this frequency was <3% due to the low VCN and the presence of the transfusion-derived Hb. Notably, total Hb concentration was only 8.1 g/dl in P2 before the treatment and reached 11.4 g/dl at the last follow-up, suggesting a better survival of RBCs derived from genetically modified HSPCs. Furthermore, in P1 and P2, HbAS3 levels were significantly higher in mature RBCs compared to reticulocytes, indicating a survival advantage of βAS3 globin-expressing cells.Overall, these clinical data indicate a variable efficacy of the DREPAGLOBE GT treatment, which likely depends on the extent of gene marking achieved in HSCs in vivo and on the engraftment capability of genetically modified HSCs in SCD patients' bone marrow. Single-cell RNA-seq analysis of HSPCs and evaluation of the bone marrow niche in SCD patients will aid to define critical parameters for achieving successful outcomes in GT clinical trials for SCD. [Display omitted] DisclosuresJoseph: bluebird bio: Consultancy. El Nemer: Hemanext: Consultancy. Bartolucci: INNOVHEM: Other: Co-founder; AGIOS: Consultancy; GBT: Consultancy; Bluebird: Consultancy, Research Funding; Jazz Pharma: Other: Lecture fees; Fabre Foundation: Research Funding; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Lecture fees, Steering committee, Research Funding; F. Hoffmann-La Roche Ltd: Consultancy; Hemanext: Consultancy; Addmedica: Consultancy, Other: Lecture fees, Research Funding; Emmaus: Consultancy. Cavazzana: Smart Immune: Other: co-founder.

Inhibition of Type-2 Diabetes Mellitus Development by Sophocarpine through Targeting PPARy-Regulated Gene Expression.

Diabetes mellitus (DM), a metabolic disorder, is the causes of oxidative stress leading to complications in micro- and macro-vascular system. The present study investigated sophocarpine for anti-diabetic potential in vivo in mice model. Sophocarpine administration to diabetic mice significantly (p < 0.05) attenuated glucose content in the plasma. The diabetes mediated lowering of GSH, ceruloplasmin and vitamin E was prevented in mice plasma by sophocarpine administration. Sophocarpine significantly (p < 0.05) reversed diabetes mediated suppression of insulin level and total Hb content in the mice plasma. In sophocarpine administrated diabetic mice C-peptide level was elevated and glycosylated hemoglobin content was suppressed significantly (p < 0.05) relative to diabetic group. Administration of sophocarpine significantly (p< 0.05) repressed diabetes mediated increase in TG and TC levels in dose-based manner. Administration of sophocarpine exhibited preventive role against diabetes mediated pathological damage to pancreas in the mice. Sophocarpine administration to diabetic mice repressed PPARγ recruitment significantly (p < 0.05) in dose-dependent manner. Sophocarpine prevents oxidative stress mediated pancreatic damage through increase in vitamin E, GSH and C-peptide levels, Moreover, the PPARγ activity was down-regulated, LDL-c content lowered and HDL-c level elevated in diabetic mice by sophocarpine. Therefore, sophocarpine may be developed for treatment of diabetes, however, further in vivo studies need to confirm the same.

Evaluation of Biochemical, Hematological and Antioxidant Properties in Mice Exposed to a Triherbal (&amp;lt;i&amp;gt;Nigella sativa, Carica papaya and Boswellia sacra&amp;lt;/i&amp;gt;) Formular

Nigella sativa, Carica papaya and Boswellia sacra are medicinal plants in the commonly used in folkloric medicine due to the presence of its immense therapeutic properties. Fifty (50) female albino mice weighing between 15-22g were divided into five groups of 10 mice each. Animal in group 1 served as control group and were administered distilled water while animal in group 2 were given 2ml of cisplatin (orally). Animal in group 3-5 were given orally; 100 mg/kg (low dose), 200 mg/kg (medium dose) and 400 mg/kg (high dose) of triherbal preparation. The feeding regimens lasted for 28 days. After 28 days, mammary gland and blood samples were collected for haematological and antioxidant analysis. The triherbal formula decreased the GSH and MDA levels of mice treated with 100 mg/kg and 400 mg/kg doses compare to control. The measurement of total protein content, SOD and CAT increased in treated animals compared to control. However, RBC (Red Blood Cell) counts significantly decreased in the low, medium and high dose groups (0.95±0.08, 6.57±0.08 and 3.55±0.55 x 106 cells/mm3 respectively) compared to control (7.34±0.40) at P<0.05. Also, significant decreases (P<0.05) in the level of the total WBC (White Blood Cell) count, platelet count, PCV (Packed Cell Volume) and Hb (haemoglobin) concentration were observed. The decreases were dose dependent. The MCH (Mean Corpuscular Haemoglobin) and MCHC (Mean Corpuscular Haemoglobin Concentration) except MCV (Mean Corpuscular Volume) significantly decreased in treated group only. The triherbal formulation exhibited significant antioxidant activities showing increased levels of SOD, CAT and Protein content due to activation of the enzyme involve in detoxification of free radicals and decreased in the level of GSH and MDA due to accumulation of peroxides and H2O2. Also, decreased in haematological parameters due to the presence of phytochemicals such as phenol, resins, saponins, sterols, tannis and terpenes in the triherbal formula. Therefore, it has potential to induce haematotoxicity hence consumption of high concentrations should be discouraged.

Direct and Label-Free Determination of Human Glycated Hemoglobin Levels Using Bacteriorhodopsin as the Biosensor Transducer

Glycated hemoglobin (HbA1c) levels are an important index for the diagnosis and long-term control of diabetes. This study is the first to use a direct and label-free photoelectric biosensor to determine HbA1c using bacteriorhodopsin-embedded purple membranes (PM) as a transducer. A biotinylated PM (b-PM) coated electrode that is layered with protein A-oriented antibodies against hemoglobin (Hb) readily captures non-glycated Hb (HbA0) and generates less photocurrent. The spectra of bacteriorhodopsin and Hb overlap so the photocurrent is reduced because of the partial absorption of the incident light by the captured Hb molecules. Two HbA0 and HbA1c aptasensors that are prepared by conjugating specific aptamers on b-PM coated electrodes single-step detect HbA0 and HbA1c in 15 min, without cross reactivity, with detection limits of ≤0.1 μg/mL and a dynamic range of 0.1–100 μg/mL. Both aptasensors exhibit high selectivity and long-term stability. For the clinical samples, HbA0 concentrations and HbA1c levels that are measured with aptasensors correlate well with total Hb concentrations and the HbA1c levels that are determined using standard methods (correlation gradient = 0.915 ± 0.004 and 0.981 ± 0.001, respectively). The use of these aptasensors for diabetes care is demonstrated.

Determination of Methemoglobin in Hemoglobin Submicron Particles Using NMR Relaxometry.

Methemoglobin (MetHb) is a hemoglobin (Hb) derivative with the heme iron in ferric state (Fe3+), unable to deliver oxygen. Quantification of methemoglobin is a very important diagnostic parameter in hypoxia. Recently, novel hemoglobin microparticles (Hb-MP) with a narrow size distribution around 700 nm, consisting of cross-linked Hb were proposed as artificial oxygen carriers. The cross-linking of Hb by glutaraldehyde (GA) generates a certain amount of MetHb. Due to the strong light scattering, quantitative determination of MetHb in Hb-MP suspensions by common spectrophotometry is not possible. Here, we demonstrate that 1H2O NMR relaxometry is a perfect tool for direct measurement of total Hb and MetHb concentrations in Hb-MP samples. The longitudinal relaxation rate 1/T1 shows a linear increase with increasing MetHb concentration, whereas the transverse relaxation rate 1/T2 linearly increases with the total Hb concentration. In both linear regressions the determination coefficient (R2) is higher than 0.99. The method does not require time-consuming pretreatment or digestion of the particles and is not impaired by light scattering. Therefore, it can be established as the method of choice for the quality control of Hb-MP and similar hemoglobin-based oxygen carriers in the future.

Protective effects of extracts of lichen Dirinaria consimilis (Stirton) D.D. Awasthi in bifenthrin- and diazinon-induced oxidative stress in rat erythrocytes in vitro

The intoxication of insecticides such as bifenthrin and diazinon has been reported to generate free radicals, and thereby alter the antioxidant defense system in erythrocytes. The present study is aimed to investigate the protective effects of acetone (DA) and methanolic (DM) extracts of lichen Dirinaria consimilis against bifenthrin and diazinon toxicity in rats’ erythrocytes in vitro. Rats’ erythrocytes were exposed to bifenthrin and diazinon, individually and also in combination with DA or DM at 1 ppm for 3 h at 37 ˚C. By using spectrophotometric methods, all the samples were estimated for changes in hemoglobin (Hb) concentration, malondialdehyde (MDA) levels, and enzyme [Superoxide dismutase (SOD), catalase (CAT), and glutathione S-transferases (GST)] activities. The outcomes showed that both the insecticides were highly toxic to rats’ erythrocytes. Among all groups, both the insecticides and DA exposed groups showed very low levels of MDA content, and GST activity in rats’ erythrocytes, when compared to the control. Besides, DA groups pretreated with both insecticides showed significant improvement of total Hb concentration, SOD, and CAT activities, with respect to the control. Hence, the present results indicate that the extracts of D. consimilis act as an antioxidant agent that reduces oxidative stress burden in insecticides toxicity.

Total haemoglobin - a reference measuring system for improvement of standardisation.

Background Total haemoglobin (Hb) concentration in blood belongs to the most requested measurands, and the HiCN method (hemiglobincyanide) is accepted as a reference. Although the reaction principle is clearly characterised, measurement conditions and settings are not consistently defined, some of them influencing the results. An improvement of standardisation is the object. Methods After method optimization, measurement results between different calibration laboratories (CL) were compared with each other and also with results of the National Metrology Institute of Germany (PTB), with target values of certified reference material, within the RELA scheme, and to >1500 results from routine laboratories. Results Overall deviations between three CLs were ≤0.5% (n = 24 samples) in a measurement range of 20 g/L to 300 g/L. A CV of 0.4% was determined in pooled blood (1 year long-term imprecision, 99.0%-101.1% recovery of the mean). For selected measurements (n = 4 samples) the PTB participated without significant differences to three CLs, and no significant differences were observed comparing CLs to certified values of reference materials. The expanded measurement uncertainty (probability 95%) was estimated as 1.1%. Conclusions A reference measuring system, comprising measuring instruments and other devices, including reagents and supply, to generate reference measurement values for total Hb concentration of high accuracy and low measurement uncertainty is presented. Measurement parameters are investigated and defined. The reference measuring system is ready to offer service to EQA providers and to the IVD industry for certifying control materials or calibrators.