Total Hb Research Articles

Role of neutrophil-lymphocyte ratio as a potential marker to differentiate between pulmonary tuberculosis and community-acquired pneumonia in children

Background: This is often creating challenge to differentiate pulmonary tuberculosis (TB) with community acquired pneumonia (CAP) particularly in children. The neutrophil-lymphocyte count ratio (NLR) as an indicator of inflammation has been used in various infectious diseases. The aim of this study was to investigate the role of NLR as a possible marker in differentiating pulmonary TB from CAP.Methods: In this retrospective record based analytic study, conducted from July 2020 to Dec 2021, patient aged 2 months to 18 years diagnosed as pulmonary TB or CAP were enrolled in this study i.e., 155 children after exclusion. The diagnostic ability of NLR in differentiating between pulmonary Tb and CAP were assessed.Results: Out of 155 children, 85 (54.8%) had CAP and 70 (45.1%) had pulmonary TB. Mean values of NLR were significantly lower in patients with TB than in pneumonia (3.16±1.9 vs 5.05±1.2, p<0.000). Total WBC, neutrophil, Hb and CRP were also significantly lower in patients with pulmonary TB. On the other hand, lymphocyte count was significantly higher in children with pulmonary TB in compare to children with pneumonia (p=0.04).Conclusions: The NLR can be used as a useful simple laboratory marker in differentiating children with Pulmonary TB from CAP, especially in resource limited high TB disease burden countries.

Fluorescence/electrochemiluminescence approach for instant detection of glycated hemoglobin index

The percentage of glycated hemoglobin (HbA1c) in total hemoglobin (Hb) is an important index for the diagnosis of Type II diabetes (T2D) because it reflects the long-term glucose level in blood. Herein, employing a one-pot co-reduction approach using glutathione (GSH) as structure-directing agent, a cluster-like AuAg nanoparticle (AuAg NPs) material was synthesized, therefore an electrochemiluminescence (ECL) aptamer-sensor for HbA1c detection was developed based on functionalized electrode with this material. Meanwhile, the quantitative determination of total Hb was realized based on the quenching effect of Hb on the fluorescence (FL) of luminol. Under compatible conditions, the results of both indexes can be satisfactorily acquired. This multimodal detection system has a good linear response toward Hb from 0.1 to 2.5 μM and HbA1c from 0.005 to 0.5 μM. The blood test proves this strategy is capable of accurate Hb and HbA1c detection, thus to obtain the percentage of HbA1c in total Hb (HbA1c%), which has the potential application for clinical diagnosis of diabetes mellitus.

Effect of Iron Deficiency Anemia on Glycated Albumin Levels: A Comparative Study in Nondiabetic Subjects with Iron Deficiency Anemia.

Objective Glycated hemoglobin A1c (HbA1c), used for monitoring glycemia control, is altered in iron deficiency anemia (IDA). Glycated albumin (GA) is considered an alternate biomarker to HbA1c. However, effect of IDA on GA needs to be studied. Materials and Methods Thirty nondiabetic cases with IDA and 30 healthy controls were included. Fasting plasma glucose (FPG), creatinine, urea, albumin, total protein, ferritin, iron, unsaturated iron binding capacity, hemoglobin (Hb), HbA1c, complete hemogram, and GA were estimated. Transferrin saturation and total iron binding capacity (TIBC) were calculated. Statistical analysis was done using unpaired two-tailed t -test/Mann-Whitney U -test and Pearson's correlation/Spearman-rank correlation, as appropriate. Results Total protein, albumin, Hb, iron, ferritin, and transferrin saturation were significantly lower while FPG, GA, TIBC, and HbA1c were significantly higher in cases compared to controls. HbA1C and GA have a significant negative correlation with iron, transferrin saturation, and ferritin. Significant negative correlations of GA with albumin ( r = -0.754; p < 0.001) and Hb ( r = -0.435; p = 0.001) and that of HbA1c with albumin ( r = -0.271; p = 0.03) and Hb ( r = -0.629; p < 0.001) while significant positive correlation of Hb with albumin ( r = 0.395; p = 0.002) and HbA1c with FPG ( r = 0.415; p = 0.001) were observed. Conclusion Low albumin levels increase plasma protein glycation, including albumin. Hence, elevated GA levels indicate false elevation of GA in scenario of lowered albumin observed in IDA, similar to HbA1c. Thus, using GA in diabetes mellitus with IDA should be avoided or used with caution to prevent potentially inappropriate treatment intensification and risk of hypoglycemia.

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.

PERSISTENT ALBUMINURIA IN A LARGE PROSPECTIVE COHORT OF CHILDREN WITH SICKLE CELL ANEMIA: RATES AND RISK FACTORS

Sickle Cell Anemia (SCA) modifies kidney structure and function. Progressive reduction in kidney function begins at very young stages of life with abnormally increased glomerular filtration rate, followed by albuminuria in late childhood. Albuminuria has been associated with poor prognosis. However, albuminuria is often intermittent in individuals with SCA, and its progression over time needs to be better understood. We aimed to investigate the progression of albuminuria and its risk factors in a large prospective cohort of children with SCA. We recruited steady-state pediatric patients with SCA at ‘Fundação Hemominas'outpatient clinic, Minas Gerais, Brazil. Random-spot urine specimens were collected during routine visits. Albuminuria was defined by urinary albumin/creatinine ratio ≥30 mg/g of creatinine. Participants were prospectively categorized by the presence or absence of albuminuria at baseline and on the follow-up urine test. Participants with albuminuria at baseline and on the follow-up urine test were considered to have persistent albuminuria. Those with no albuminuria at baseline or on the follow-up urine tests were considered to have no albuminuria. Participants with no albuminuria at baseline who developed albuminuria on the follow-up test were considered progressors. Those with albuminuria at baseline but none on follow-up were considered remitters. Baseline steady-state laboratory data were obtained from medical records. Baseline kidney injury biomarkers in urine were measured using multilplex xMAP® assays. Univariate and multivariate analyses were performed using Cox regression. From 572 children included in the cohort [287 (50.2%) males], 112 (19.6%; 95% CI:16.3%–22.8%) had albuminuria at enrollment; mean age was 12.9±4.6 years. A total of 246 participants (43%) had two samples and were included in the longitudinal analysis. The median follow-up was 2.1 (6m–4y) years, providing 467.9 patient-years. One hundred eight-seven (76%) participants were receiving intensification therapy (HU, chronic blood transfusion, or both). Out of 246 participants, 169 (68.7%) presented no albuminuria, 13 (5.3%) were progressors, 19 (7.7%) remitters, and 45 (18.3%; 95% CI 13.5%–23.1%) had persistent albuminuria. The incidence of persistent albuminuria was 9.6/100 patient-years (95% CI 6.9–12.3). In cox multivariate analysis, persistent albuminuria was associated with older age (HR = 1.1, 95% CI 1.02–1.19; p = 0.018), lower total Hb (HR = 0.75, 95% CI 0.57–0.98; p = 0.040), and higher indirect bilirubin (HR = 1.3, 95% CI 1.11–1.58; p = 0.002) and kidney injury molecule-1 levels (HR = 1.007, 95% CI 1.001–1.013; p = 0.020). This study provides important findings on the natural history of albuminuria during childhood of individuals with SCA. The prevalence of persistent albuminuria was similar to previously published studies in children with SCA (13% to 30%). Our findings replicated the association of persistent albuminuria with aging, anemia, and higher levels of hemolysis and renal tubular injury markers. These features may allow early identification of individuals at highest risk for persistent albuminuria, enabling therapeutic interventions to prevent the decline of renal function. This study showed a high frequency of persistent albuminuria in children with SCA. Age, anemia, and biomarkers of hemolysis and renal tubular injury were significantly associated with persistent albuminuria.

REFRACTORY HYPOXEMIA: A CASE OF DAPSONE-INDUCED METHEMOGLOBINEMIA IN A PATIENT WITH COVID-19

REFRACTORY HYPOXEMIA: A CASE OF DAPSONE-INDUCED METHEMOGLOBINEMIA IN A PATIENT WITH COVID-19

Angelica archangelica and Ginkgo biloba Extracts Recover Functional Blood Hemoglobin Derivatives in Rabbits Exposed to High Altitude.

Shortage of oxygen is a common condition for residents of high-altitude (HA) areas. In mammals, hemoglobin (Hb) has four derivatives: oxyhemoglobin (Hb-O2), carboxyhemoglobin (Hb-CO), sulfhemoglobin (Hb-S), and methemoglobin (Met-Hb). In HA areas, aberrant physiological performance of blood hemoglobin is well-established. The study aimed to investigate the influence of 30 days of HA residence on rabbits' total Hb, Hb derivatives, Hb autooxidation rate, and antioxidant enzymes in comparison to low-altitude control rabbits. Further, the study aimed to investigate the effect of antioxidant-rich Angelica archangelica and/or Ginkgo biloba extracts on the same parameters in HA-resident rabbits. Rabbits subjected to 30 days of HA residence were compared to low-altitude control rabbits. HA-residence rabbits were then orally administered 0.11 g/kg b.wt. of Angelica archangelica and/or Ginkgo biloba extract for 14 days. Hb derivatives and Hb autooxidation rate were measured spectrophotometrically. Antioxidant enzymes were estimated using specialized kits. Compared to low-altitude rabbits, 30-day HA-residence rabbits showed a noticeable increase (p<0.05) in Hb-O2 and Hb-CO concentration. In addition, Met-Hb concentration, autooxidation rate of Hb molecules, and activities of superoxide dismutase (SOD) and glutathione peroxidase (GPx) exhibited a remarkable increase in HA-residence rabbits (p<0.01), reflective of rapid ROS generation. In HA-residence rabbits, both individual and combined treatment with antioxidant-rich extracts for 14 days resulted in recovery to near-normal functional levels of Hb-O2 and Met-Hb, Hb autooxidation rate, and activities of SOD and GPx, while only combined treatment led to Hb-O2 recovery. The findings suggest that functional Hb levels may be recovered by oral administration of A. archangelica, G. biloba, or combined treatments. In conclusion, oxidative stress due to living in HA areas may be avoided by supplementation with natural antioxidants.

A Comparative Study on Pathological Changes in the Small Intestine of Sheep and Goat Experimentally Infected with Trichostrongylus colubriformis.

BackgroundTrichostrongylus colubriformis, also called hairworm, is a genus of parasitic roundworm affecting gastro-intestinal tracts of a ruminant. Gross and microscopic lesion characterizations and comparing its effect in the small intestine of sheep and goats experimentally infected with T. colubriformis were undertaken in the study.MethodsDuring the study period, 13 sheep and 14 goats were included in the experiment. The larvae of T. culibriformis were obtained from abattoirs and larvae were recovered by Bearmann techniques. The infective larvae of T. culibriformis (L3) as a single dose of 10,000 per-animal was administered orally to infected groups of sheep and goats. Blood was collected for hematological and serum biochemical analysis. Tissues for gross and histopathologic lesions characterization were collected from killed infected animals at 56 days.ResultsFrom the infected group, the total recovered mean worm burden was recorded as higher in goats (P<0.05) than sheep, with an establishment rate of 50.16% and 34.46%, respectively. The total mean PCV, Hb, and albumin values recorded in the infected groups of sheep and goats were significantly (P<0.05) lower than non-infected control of both animal groups. In goats, the total serum protein was significantly (P<0.05) lower in the infected group than the non-infected control group. Gross lesions found were enteritis with petechial hemorrhages, edema, hyperemia, and mucosal slough, which were marked in the duodenum (62.69%) and jejunum (33.33%) in sheep and 47.05% duodenum and 45.09% jejunum in goats. The microscopic lesions developed by T. colubriformis were subtotal villus atrophy, hemorrhage, straightened and elongated dilated crypts, loss of epithelium, mucosal erosion, and infiltration of inflammatory cells.ConclusionThe present study showed that T. colubriformis infection caused physiological and pathological changes of the small-intestine in sheep and goats, with more severe infection in goats than sheep, although they were under the same management condition.

P-028: CIRCULATING CD34+CD235A+ CELL COUNT CORRELATES WITH LEVELS OF HBF IN SICKLE CELL ANEMIA PATIENTS UNDER HYDROXYUREA TREATMENT

Purpose: Studying stress erythropoiesis (SE) in sickle cell anemia (SCA) is difficult because of the rigor in accessing bone marrow samples. However, the characterization of circulating cell populations reflecting SE is feasible and critical, especially in the perspectives of gene therapy. Materials and methods: Clinical and laboratory data were collected from 49 subjects with SCA (SS-genotype) under chronic transfusion (CT, n = 15), hydroxyurea therapy (HC, n = 20), or under both treatments (CT/HC, n = 14) at the Blood Center of Ribeirão Preto - Brazil. The mean age was 35.22 y/o (20-64), and 53.06% were males. Six patients from each group, under steady-state, and five healthy donors (HD group) were selected for downstream analysis of PBMCs. Briefly, 5 mL of blood were collected in EDTA-evacuated tubes and processed immediately with Ficoll-Paque PLUS (Cytiva). 106 live cells were stained with anti-CD235a, -CD123, -CD36, and -CD34 (stemness and erythroid maturation markers) BD Biosciences antibodies and analyzed by FACS in a BD FACSCanto II. FACS analysis was conducted in the FlowJo software v.11, using conventional gating strategies and dimensionality reduction machine learning algorithms. Complete blood and reticulocyte count, liver function, hemoglobin profiling, and FACS data were plotted for statistical comparison of mean ranks using Kruskal-Wallis, Dunn’s multiple comparisons, or Spearman’s rank-order correlation tests. Data analysis was performed in statistical software GraphPad Prism v.6.0 and open-source statistical programming language R v.4.0.2, using packages ‘Hmisc’ and ‘corrplot’ for correlation analysis and visualization of results. Significance levels were considered when p<0.05 in which ‘*’, ‘**’, ‘***’ and ‘****’ indicate p<0.05; p<0.01, p<0.001, and p<0.0001, respectively. Results: Subjects under HC had overall higher median Hb, HCt, MCV, MCH, HBS, and HBF while CT groups presented higher RDW. The number of nucleated RBCs (nRBC) was similar among groups and the ratio of RBCHb/RetHb increased under HC treatment. Also, levels of plasma-free hemoglobin (calculated based on total and cellular Hb) were lower in HC. Reticulocytosis was higher in CT and hemolysis parameters appeared discretely lower in HC (Table 1). FACS analysis revealed that patients had higher circulating CD34+ cell counts. Levels of CD235+ cells inside the PBMC compartment were higher in HC patients. Also, CD34+CD235a+ population counts were significantly higher in HC than in CT and CT/HC, representing more than 15% of the circulating CD34+ compartment in some patients. Up to 60% of the CD34+CD235a+ were also CD36+ (Figure 1 A). In contrast, CD34+CD123+ were increased among CT and CT/HC groups. CD235a+ and CD34+CD235a+ populations positively correlated with HbF levels in the HC group, whilst its dose did not associate with any blood count or hemoglobin parameters (Figure 1 B). Interestingly, the same population counts positively correlated with age among non-HC takers. Conclusion: These results reaffirm the capacity of HC treatment to improve hematological parameters in SCA. Notwithstanding, HC presented increased levels of SE cell populations than the CT groups, reflecting different mechanisms of SE onset between CT and HC treated patients. Financial Support: CAPES, CTC/FAPESP-(2013/08135-2); INCTC-(465539/2014-9).Complete blood count and hemolysis biomarkers of HC and CT / HC groups compared with CT, calculated using Kruskal-Wallis test. ‘*’, ‘**’, ‘***’ and ‘****’ indicate p?0.05; p?0.01, p?0.001, and p?0.0001, respectively.A) Levels of CD34+ and CD34+CD235a+ cell populations. B) Correlogram of stress erythropoiesis cell FACS data versus hematological parameters of hydroxyurea treatment groups. Levels of stress cells are positively correlated with levels of HbF. The authors do not declare any conflict of interest

Co-occurrence of oculocutaneous albinism type 2 and mild sickle cell disease explained by HbS/βthal genotype in an individual from the Democratic Republic of Congo

Oculocutaneous albinism type 2 (OCA2) is a pigmentation disorder characterized by hypopigmentation of the skin, hair and eyes and ocular features. Sickle cell disease (SCD) is caused either by homozygosity of the beta globin gene variant c.20A > T/p.Glu6Val giving rise to severe anemia or by combined abnormal hemoglobins (HbS/βthal) leading to mild SCD. We report a 45 years old female patient from the Democratic Republic of Congo affected with these two disorders. She presented with creamy white skin and numerous pigmented patches called dendritic freckles, nystagmus, foveal hypoplasia grade 2, photophobia and very poor visual acuity. Sequencing of the OCA2 gene identified the common exon 7 deletion and a new pathogenic variant c.1444A > C/p.Thr482Pro. She had mild SCD with a total Hb level of 101 g/l. Hbβ sequencing identified variants c.20A > T giving rise to HbS and c.315 + 1 G > A characteristic of β-thalassemia. A heterozygous 3.7 kb deletion of the α globin gene was also found. The combined Hbβ/α globin genotype explains the mild SCD phenotype. Co-occurrence of OCA2 and SCD raises the question whether the patient's phenotype simply results from the addition of the two diseases' phenotypes or whether interaction between the two diseases modulates the phenotype of each other.

Association Between Rental Assistance Programs and Hemoglobin A1c Levels Among US Adults

Programs that provide affordable and stable housing, such as federal rental assistance, may be associated with improved mean blood glucose levels and related diabetes outcomes. To assess whether 2 different types of federal rental assistance programs are associated with glycated hemoglobin A1c (HbA1c) levels among middle-aged and older US adults. This cohort study used data from the National Health and Nutrition Examination Survey (NHANES) linked with US Department of Housing and Urban Development records of rental assistance participation. Adults aged 45 years or older who were receiving 2 types of rental assistance (project-based housing or housing vouchers) at the time of the NHANES interview and those who would receive rental assistance within the subsequent 2 years (waitlist group) were included. Data were collected from January 1999 to December 2016 and analyzed in October 2021. Rental assistance participation, including project-based housing (subsidized housing developments including public housing) and housing vouchers (tenant-based subsidies for private market housing). The primary outcome was continuous HbA1c level, a common measure of blood glucose reflecting diabetes control. Linear regression was used to estimate the association between the 2 rental assistance programs and HbA1c level. Logistic regression was used to assess the association between rental assistance programs and HbA1c cut points (prediabetes: 5.7% to ≤6.5%; diabetes: >6.5%; uncontrolled diabetes: ≥9% [to convert to proportion of total Hb, multiply by 0.01]). Analyses used weights created by the National Center for Health Statistics that adjust for linkage eligibility. Among 1050 adults in the study (41.6% aged ≥65 years; 70.1% female), 795 were receiving rental assistance at time of the NHANES interview (450 lived in project-based housing, and 345 had housing vouchers), and 255 received rental assistance within 2 years after the interview. Participants in project-based housing had lower HbA1c levels compared with individuals in the waitlist group (β, -0.290; 95% CI, -0.599 to 0.020), but the difference was not significant. No significant differences in HbA1c levels were found between those receiving housing vouchers and those in the waitlist group (β, 0.051; 95% CI, -0.182 to 0.284). Receiving project-based housing was associated with a reduced likelihood of uncontrolled diabetes (-3.7 percentage points; 95% CI, -7.0 to -0.0 percentage points) compared with being in the waitlist group. In this cohort study of a nationally representative sample of US adults, living in project-based, federally subsidized housing was associated with a reduced likelihood of uncontrolled diabetes. The findings suggest that affordable housing programs may be associated with improved diabetes outcomes.

P1465: INITIAL SAFETY AND EFFICACY STUDY OF RM-001, AUTOLOGOUS HBG1/2 PROMOTER-MODIFIED CD34+ HEMATOPOIETIC STEM AND PROGENITOR CELLS, IN TRANSFUSION-DEPENDENT ΒETA-THALASSEMIA

Background: Natural mutations that cause hereditary persistence of fetal hemoglobin (HPFH) alleviate the symptoms of β-thalassemia. It is now known that small natural deletions and mutations in the γ- globin gene (HBG1/HBG2) promoters disrupt the binding of the transcriptional repressors BCL11A, causing HPFH. Using gene editing to mimic these mutations should reactivate γ- globin in patients with transfusion-dependent β-thalassemia (TDT) and ameliorate the debilitating effects of mutations in β-globin. Aims: To induce HbF production in erythrocytes, we used ex vivo CRISPR-Cas9–based gene editing to modify HBG1/HBG2 promoter in hematopoietic stem and progenitor cells (HSPCs), producing RM-001. We initiated initial safety and efficacy study of RM-001 in TDT patients. Here, we present initial results from the first 2 patients treated with RM-001. Methods: Patients (6–35 y of age) with TDT receiving packed red blood cell (pRBC) transfusions of ≥100 mL/kg/y or ≥10 units/y in the previous 2 y were eligible. Peripheral CD34+ HSPCs were collected by apheresis after mobilization with G-CSF and plerixafor. CD34+ cells were edited with CRISPR-Cas9 using a guide RNA specific for the binding site of BCL11A on the HBG1/HBG2 promoter. Prior to RM-001 DP infusion, patients received myeloablative conditioning with Busulfan from day-7 to day-3. Patients were monitored for stem cell engraftment/hematopoietic recovery, adverse events (AEs), Hb production, HbF and F-cell expression, pRBC transfusion requirements. Results: Data presented here for the first 2 TDT patients treated with RM-001. Both patients have β0/β0 genotype (CD17/CD41-42, CD41-42/CD41-42), with an annualized packed red blood cell (pRBC) transfusion history of 39 units/y and 54.8 units/y over 2 y prior to consent, respectively. Both patients received a single dose of RM-001 cells, ceased pRBC transfusions within 1 month after RM-001 infusion and remained transfusion-free through the reported period. The first patient (CD17/CD41-42, 9y) achieved neutrophil and platelet engraftment on Day(D+)19 and D+21; the other (CD41-42/CD41-42, 13y) achieved neutrophil and platelet engraftment on D+16 and D+21, respectively. No serious adverse event (SAE) occurred. The safety profile was generally consistent with busulfan myeloablation and autologous hematopoietic stem cell transplantation. Total Hb, HbF and F-cell percentage increased over time in both patients (Figure). Both patients received 6 units of pRBC before transfusion independent (TI) achieved (Hb stably ≥ 90g/L), with last pRBC transfusion on D+15 and D+28, respectively. The first patient achieved TI on D+28 and the other achieved TI on D+39. At 3 month post-RM-001 infusion, total Hb exceed 110 g/L in both patients, with 97-98 g/L HbF. Approximately 89% peripheral erythrocytes are F-cells at 3 month post-RM-001 infusion (Figure). Data will be updated for the presentation. Image:Summary/Conclusion: The first 2 patients treated with RM-001 demonstrated successful engraftment and both have been transfusion free within two month. Although both patients have β0/β0 genotype, they ceased pRBC transfusions within 1 month after RM-001 infusion and remained transfusion-free through the reported period. This is the first-in-human study of autologous HBG1/2 promoter-modified CD34+ HSPCs (RM-001) in TDT and both patients were treated successfully without SAE. Our studies provide novel and initial clinical evidence supporting the safety and efficacy of RM-001 for treating TDT.

Abstract LB057: Bevacizumab leaves a photoacoustic fingerprint in breast cancer mouse models

Abstract Background: Photoacoustic tomography (PAT) provides a direct readout of tumor haemoglobin (Hb) concentration and oxygenation. This readout could be used to provide an early indication of response to anti-angiogenic drugs, thus optimizing the management of these therapies. Experimental Procedures: Two cohorts of nude Balb/c were inoculated with either estrogen-dependent MCF-7 (n=7) or estrogen-independent MDA-MB-231 (n=19) cells. Mice were randomly split into Control (Ctrl) and Bevacizumab (Bev,10 mg/Kg, IP, weekly) groups. PAT was performed weekly, starting just before enrolment. Oxy- and deoxy-Hb were quantified in the tumour and reference areas. Total Hb (THb) and O2 saturation (SO2) were calculated. Blood samples and tissues were collected after imaging. Hypoxia (Hypoxyprobe and CA-IX) and vascular density/maturity (CD31 and ASMA) were analyzed by immunohistochemistry. Circulating levels of human and mouse vascular endothelial growth factor (hVEGF and mVEGF) and Hb were measured. Summary of New Data: Reflecting clinical observations, the MCF-7 Bev group and most of the mice from the MDA-MB-231 Bev group (8/11) showed no improvement in survival with Bev treatment. All resistant tumours (Bev MCF-7 and Bev MDA-MB-231 non-responders (Bev-NR)) showed an increase in hVEGF production (Ctrl vs Bev; 73.0513.06 vs. 497103.9 for MCF-7; 247.984.81 vs. 330.4 85.97 for Bev NR). MDA-MB-231 responders (Bev-R) showed a significant decrease in hVEGF (247.984.81 vs. 80.53 26.13). These results indicate that VEGF was only successfully sequestered in a small subset of animals (3/11) and that VEGF overload might be a resistance mechanism. At the final time point, PAT showed no difference between Ctrl and Bev-NR in THb for either group (THbMDA-MB-231 7.2±1.3 vs. 5.5±1.2). THb was found significantly increased in the Bev-R group by PAT (13.9±3.3 p-valuevs.Crtl=0.0339; p-valuevs.Bev=0.0072) and biochemical measurements. SO2 showed a slight but significant decrease between Ctrl and Bev-NR (0.58±0.03 vs. 0.48±0.01). A dramatic decrease was seen in the Bev-R group (0.31±0.07) and significantly different from Ctrl and Bev-NR. SO2 was sensitive to early changes, SO2 values increased in Ctrl and Bev-NR 48h after enrolment (p-values paired t-test pCtrl= 0.0134; pNR=0.0268) while Bev-R shows a trend (p=0.0649). Analysis of SO2MSOT over time shows a significant (p&amp;lt;0.0001) change in slope at 3 weeks after enrolment in the Bev-R group compared to either of the Ctrl or Bev-NR groups. Our results indicate that PAT SO2 can differentiate responders at very early stages of the treatment. Histologically, we observed fewer blood vessels but better structured in Bev-NR than in Bev-R, which showed higher vascular density and also higher levels of hypoxia makers. Statement of the Conclusions: Our results postulates PAT as a low-cost, label-free and non-invasive candidate to monitor response to Bevacizumab over time. Our results also indicate that tumours with functional vasculature may be resistant to Bevacizumab treatment. Citation Format: Isabel Quiros-Gonzalez, Michal R. Tomaszewski, Monika A. Golinska, Emma Brown, Laura Ansel-Bollepalli, Lina Hacker, Dominique-Laurent Couturier, Rosa M. Sainz, Sarah E. Bohndiek. Bevacizumab leaves a photoacoustic fingerprint in breast cancer mouse models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB057.

Non-invasive metabolic profiling of inflammation in joints and entheses by multispectral optoacoustic tomography.

To explore the metabolic characteristics of arthritis and enthesitis using multispectral opto-acoustic tomography (MSOT), a technology using near-infrared multispectral laser to stimulate tissues and detect the emitted acoustic energy, enabling non-invasive quantification of tissue components in vivo based on differential absorbance at multiple wavelengths. We performed a cross-sectional study in patients with RA or PsA and healthy controls (HCs). Participants underwent clinical, ultrasonographic and MSOT examination of MCP and wrist joints as well as the entheses of the common extensor tendon at the lateral humeral epicondyles and of the patellar, quadriceps and Achilles tendon. MSOT-measured haemoglobin (Hb), oxygen saturation, collagen and lipid levels were quantified and scaled mean differences between affected and unaffected joints and entheses were calculated as defined by clinical examination or ultrasonography using linear mixed effects models. We obtained 1535 MSOT and 982 ultrasonography scans from 87 participants (34 PsA, 17 RA, 36 HCs). Entheseal tenderness was not associated with significant metabolic changes, whereas enthesitis-related sonographic changes were associated with increased total Hb, oxygen saturation and collagen content. In contrast, the presence of arthritis-related clinical and sonographic findings showed increased Hb levels, reduced oxygen saturation and reduced collagen content. Synovial hypertrophy was associated with increased lipid content in the joints. MSOT allows determination of distinct metabolic differences between arthritis and enthesitis in a non-invasive setting in humans in vivo.

The addition of oral Tranexamic acid to knee arthroplasty patients does not further improve blood loss: a double blinded randomized control trial.

Perioperative intravenous (IV) TA has become routine in knee and hip arthroplasty. Less evidence exists on the administration of oral TA in the post- operative period. Our study aims to identify the efficacy and safety of combined perioperative IV and post-operative oral TA on blood loss and Hemoglobin (Hb) drop compared to perioperative IV TA alone. Patients undergoing primary elective knee arthro- plasty at our institution were invited to participate in the study (n=50). A computer-generated randomisation sequence was created online (www.randomization. org), with an allocation ratio of 1:1 and a block size of 50. Group A received perioperative IV TA alone and post-operative oral TA (n= 26) and Group B received perioperative IV TA plus 48 hours additional oral placebo (n= 24). Day 3 total blood loss and Hb drop was calculated. Continuous, normally distributed data (total blood loss) was compared utilising using one-way analysis of variance with post hoc Tukey test. Continuous skewed data (Hb drop) was compared using the Kruskal-Wallis test. P <0.05 was considered statistically significant. Group A demonstrated a trend in decreased total blood loss that was close to statistical significance ( p = 0.072). No difference in Hb drop was identified between the 2 groups. Increased nausea was also observed in Group A. The administration of oral TA to post-operative knee arthroplasty patients does not improve further blood loss compared to patients receiving perioperative IV TA pre-operatively and at wound closure.

Post hoc analysis of a long-term safety extension study: Responses to siltuximab in idiopathic multicentric Castleman disease patients receiving on-label dosing.

e19586 Background: Siltuximab is the only FDA- and EMA-approved therapy for idiopathic multicentric Castleman disease. Its long-term efficacy and safety were established in a 6-year extension study featuring patients who responded to treatment or had stable disease in a phase 1 or phase 2 trial. However, dosage could be adjusted at physician discretion; of 60 patients, 33 remained on the on-label dosing of 11 mg/kg administered every 3 weeks. To assess the safety and efficacy of on-label dosing, we performed a post hoc analysis in patients who maintained this dosing. Methods: A mixed-effects model was used to analyze longitudinal measurements for tumor response, C-reactive protein (CRP), hemoglobin (Hb), erythrocyte sedimentation rate (ESR), fibrinogen, albumin, and platelet counts at multiple time points. Adverse events (AEs) were also assessed. Greenhouse–Geisser correction was used to ensure that the assumption of sphericity was met for the mixed-effects model. Tukey post hoc correction was applied to determine significant differences between specific time points. Results: Of the 33 patients on 3-week dosing, one transitioned to receiving siltuximab every 6 weeks for 25 cycles but restarted 3-weekly dosing due to suspected progressive disease (PD). Disease control was observed in 31 of the 33 patients (93.9%) at their last on-study assessment (complete response [CR] in 9 [27.3%]). Quantitative analyses were performed on the 32 patients who maintained 3-weekly dosing until their last on-study assessment. Siltuximab led to sustained mean levels of CRP, Hb, ESR, fibrinogen, albumin, and platelet counts for the duration of the study. In the 19 patients with residual IL-6 activity at screening (CRP levels &gt; 4 mg/L) there was an overall reduction in CRP from baseline at multiple time points. This group showed a trend towards improved total Hb levels, which increased by 9.43% (adjusted p = 0.048) at cycle 31, and some demonstrated an improvement in ESR, fibrinogen, and albumin levels. All patients reported AEs, mostly grade 1 or 2, with no deaths. At point of enrolment into the extension study, the three most common AEs were hypertension (36.4%), fatigue (42.4%), and pain (51.5%). Conclusions: Our findings show that long-term on-label dosage of siltuximab sustains an improved inflammatory profile and disease control.[Table: see text]

Vascular myogenic control of tibial bone blood flow in young healthy individuals

Bone health is critically dependent on adequate blood flow supplied by its extensive vascular network. Without adequate perfusion to provide oxygen and essential nutrients critical for bone metabolism, nearly all skeletal functions are compromised. However, despite the importance of blood flow to bone, regulation of bone blood flow is poorly understood in humans. The broad hypothesis of the current work is that the primary regulators of blood flow to bone are not different from other vascular beds. We have recently shown that sympathetic innervation of the bone vasculature has an active role in controlling bone blood flow in young healthy individuals. This work furthers our previous findings by investigating additional key bone blood flow regulators ‐ myogenic vasodilation and vasoconstriction. While vascular myogenic control has been investigated in soft tissue, to our knowledge there has been no research in the bone vasculature in humans, with only a handful of animal studies.In young healthy individuals (N=8), we characterized myogenic control of tibial blood flow via novel custom‐made near infrared spectroscopy (NIRS). Tibial blood flow via NIRS was measured as oxy‐, deoxy‐, and total hemoglobin (Hb). We employed reactive hyperemia and leg dependency to assess myogenic vasodilation and vasoconstriction in the whole calf and in the tibia. The magnitude of maximum myogenic vasodilation was defined by the peak reactive blood flow response to sustained ischemia. Arterial flow to the lower leg was occluded by rapidly inflating a pneumatic thigh cuff above the knee to 220 mmHg in supine subjects. Tibial blood flow was assessed via NIRS and whole leg blood flow velocity in the popliteal artery of the same leg was measured via Doppler ultrasound during 1 min of rest, 10 min of cuff inflation, and 5 min of recovery. Myogenic vasoconstriction was assessed via leg dependency, i.e., during two levels of increased transmural pressure achieved by lowering the leg below heart level. Only tibial blood flow was assessed during 5 min in supine position, 5 min in seated upright position, and 5 min in seated position with legs lowered at 90deg knee flexion.During cuff occlusion, there was a time dependent decrease in bone oxyHb accompanied by an increase in deoxyHb indicating lack of flow with continued metabolism. Tibial total Hb (sum of oxy‐ and deoxy‐Hb) did not change significantly during cuff inflation, but increased rapidly and markedly with cuff release, far surpassing baseline values when flow was restored. A similar overshoot response was seen in whole leg blood flow with cuff release. Thus, the response of tibial flow to reactive hyperemia follows the same behavior as the whole limb, indicating an active myogenic vasodilator response in bone similar to other vascular beds. During leg dependency, despite increases in arterial perfusion pressure at both levels (ΔP, 25.8 + 3.67, and 40.0 + 3.02 mmHg), there was no change in tibial flow (total Hb, ‐0.39 + 7.40, and ‐0.29 + 16.2 μM). Hence, unchanged tibial flow despite increased pressure indicates a compensatory vasoconstrictor response in the tibia. Our results indicate that similar to other vascular beds, myogenic control plays an active role in regulating the bone vasculature in young healthy individuals. Characterizing blood flow regulation in bone constitutes the basis of defining its role in overall bone health and its contribution to numerous bone loss conditions.

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.

Discrimination Between Healthy Eyes and Those With Mild Glaucoma Damage Using Hemoglobin Measurements of the Optic Nerve Head.

The Laguna ONhE, a software that measures the hemoglobin (Hb) concentration of the optic nerve head (ONH) from fundus photographs, demonstrated good accuracy in discriminating healthy eyes from eyes with mild glaucoma. The aim was to evaluate Hb concentration of the optic nerve to distinguish between healthy eyes and eyes with mild glaucoma. Eyes from patients with mild primary open angle glaucoma (MD > -6 dB) (n=58) and from healthy subjects (n=64) were selected. Retinal nerve fiber layer thickness measurements of all eyes were acquired with optical coherence tomography. Optic disc photographs were also obtained, and the images were analyzed using the Laguna ONhE software, which measures the amount of Hb in 24 sectors of the ONH. The software also calculates the Glaucoma Discriminant Function (GDF), an index that expresses the chance of the ONH being compatible with glaucoma. Areas under the receiver operating characteristic curve and sensitivities at fixed specificities of 90% and 95% of each Laguna ONhE parameter were calculated. The mean retinal nerve fiber layer thickness and vertical cup/disc ratio of the control and glaucoma groups were 90.0±10.6 μm versus 66.28±9.85 μm ( P <0.001) and 0.5±0.09 versus 0.65±0.09 ( P <0.001), respectively. Total Hb (67.9±4.45 vs. 62.89±4.89, P <0.001) and GDF (11.57±15.34 vs. -27.67±20.94, P <0.001) were significantly higher in the control group. The Hb concentration was also significantly higher in 21 of the 24 sectors in the control group compared with the glaucoma group ( P <0.05). The GDF had the largest areas under the receiver operating characteristic curve (0.93), with 79.3% sensitivity at a fixed specificity of 95%. Measurements of optic nerve Hb concentration using a colorimetry photographic device demonstrated good accuracy in discriminating healthy eyes from eyes with mild glaucoma. Further studies are need to understand vascular factors implicated in the development of glaucoma.