Differences In Red Blood Cells Research Articles

Durable and Robust Fetal Globin Induction without Anemia in Rhesus Monkeys Following Autologous Hematopoietic Stem Cell Transplant with BCL11A Erythroid Enhancer Editing

Elevated fetal hemoglobin (HbF, α2γ2) levels are clinically beneficial for patients with β-hemoglobinopathies. Editing of the erythroid-specific BCL11A enhancer induces HbF, inhibiting sickling and restoring globin chain balance in erythroid cells derived from hematopoietic stem and progenitor cells (HSPCs) from SCD and β-thalassemia patients respectively, without detectable genotoxicity or adverse effects on hematopoietic stem cell (HSC) function (Wu Y, Nat Med, 2019). Here, we sought to evaluate engraftment and HbF induction potential of erythroid-specific BCL11A enhancer edited CD34+ HSPCs in a non-human primate transplantation model in which hemoglobin switching is conserved. We targeted the erythroid-specific +58 DNAse I hypersensitive site of BCL11A, which has identical human and rhesus sequences at the spacer and protospacer adjacent motif (PAM) of the potent #1617 sgRNA. Ribonucleoprotein complex (RNP) composed of 3x-NLS SpCas9 protein and either BCL11A enhancer targeting (#1617) or AAVS1 targeting sgRNA was electroporated into rhesus CD34+ HSPCs (n=3). Following erythroid differentiation, substantial γ-globin expression (54-77%, p<0.01) was observed in BCL11A edited cells (81-85% indels) as compared to 19-25% and 15-24% for non-electroporated and AAVS1 edited cells, respectively, with no significant difference in red blood cell (RBC) enucleation efficiency (44-47%) among groups. We tested BCL11A enhancer editing with autologous HSC transplant in two cohorts, with two macaques per cohort. For cohort 1, we performed competitive engraftment of BCL11A enhancer and AAVS1 edited HSPCs to test long-term reconstitution. For cohort 2, we evaluated BCL11A enhancer editing alone to evaluate HbF induction and hematopoietic reconstitution. For each cohort, purified CD34+ HSPCs were electroporated with RNP one day after G-CSF and plerixafor mobilization and cultured for two days prior to cryopreservation. HSPCs were thawed and infused following 2×5 Gy total body irradiation. For cohort 1 (n=2, ZL25 and ZL22, 1.34-1.39×106 CD34+ HSPCs/kg), we observed reduced indel frequencies (8-41%) at early post-infusion time points compared to cell products (18-49%), suggesting indels in unfractionated HSPCs may overestimate those in engrafting cells and/or hematopoietic ablation was incomplete. From weeks 6 to 83, stable indel frequencies were detected in both BCL11A (~3-18%) and AAVS1 (~10-45%), suggesting no selective advantage for BCL11A enhancer edited, AAVS1 edited, or non-edited HSCs. For cohort 2 (BCL11A enhancer editing alone (n=2, ZM17 and ZM26, 1.78-6.06×106 CD34+ cells/kg), cell products showed improved editing with ~95% indels and ~65-78% γ-globin protein after in vitro erythroid culture. Animals engrafted with typical kinetics and displayed stable indel ratios up to 28 weeks post-transplantation. A significant correlation was detected between γ-globin level and indel frequency comparing all 4 transplanted animals and unedited controls (R2=0.76, p<0.01). In both edited and unedited animals γ-globin levels peaked in the first two months after transplantation and subsequently declined and plateaued. In ZM17 (~70% BCL11A enhancer indels at ~24 weeks), ~12% γ-globin was observed in peripheral blood (PB) at last measurement (compared to 0.5% γ-globin in RBC prior to transplant). In the same animal, editing ranged from 78-81% across all PB and bone marrow (BM) lineages (excluding CD3+ T-cells with 63% indels), including B-lymphoid, myeloid, erythroid, and HSPCs (in particular including 78% indels in CD71+ CD45- erythroblasts). Hemoglobin, hematocrit, and reticulocyte counts and peripheral smear appearance were all normal, suggesting no erythroid toxicity. Colony-forming ability of BM-derived mononuclear cells was similar in edited and control animals. In summary, we evaluated the clinical potential of autologous BCL11A erythroid enhancer editing in rhesus macaques. BCL11A enhancer edited HSCs can persist for at least 83 weeks post-transplant and provide therapeutic levels of HbF in peripheral RBCs without anemia or other apparent hematologic toxicity. Furthermore, these results emphasize input CD34+ HSPC dose and conditioning intensity as critical variables that influence gene editing following autologous HSCT. Overall, these findings support BCL11A erythroid enhancer genome editing as a promising strategy for therapeutic HbF induction. Disclosures Weiss: GlaxoSmithKline: Consultancy; Cellarity INC: Consultancy; Esperian: Consultancy; Beam Therapeutics: Consultancy; Rubius INC: Consultancy.

Laparoscopic transperitoneal adrenalectomy v.s. posterior retroperitoneal adrenalectomy in the aspect of complete blood count

Introduction: Minimally invasive surgery is the gold standard for treatment of benign adrenal tumours up to 7–8 cm in diameter. Nowadays, two surgical approaches are performed routinely – lateral transperitoneal adrenalectomy (LTA) and posterior retroperitoneal adrenalectomy (PRA). Both methods are recognized as safe and beneficial for a patient, what can also prove the statistical analysis in the perioperative period. Aim: The aim of our work was to compare lateral transperitoneal adrenalectomy with posterior retroperitoneal adrenalectomy in the aspect of complete blood count. M aterial and Methods: Patients, who underwent videoscopic adrenalectomy in our Department from 02.2015 to 06.2018, were prospectively included to the study. The inclusion criteria was the tumour size smaller than 8cm in abdominal CT. Primary neoplasms were excluded from the study. All surgieres were carried out by one surgeon. Patients were randomized to PRA or LTA. The blood samples were collected - a day before the surgery, and a day, a week, and a month after the surgery. We assessed the rate of leucocytes, red blood cells, hemoglobin and platelets in the peripheral blood. Additionally, the volume of intraoperative blood loss was measured. Results: 77 videoscopic adrenalectomies were carried out, 33 (42,9%) – LTA, 44 (57,1%) – PRA. The average age of the patients constituted- LTA – 61.2± 8.3, PRA – 59.3±10.2. Tumour size in LTA – 4 cm, in PRA – 4.1 cm. Adrenal gland adenomas, basing on postoperative histopathological findings, were most commonly removed tumours in our study: LTA – 82%, PRA – 70%. Intraoperative blood loss was not significant in both groups. We found differences in red blood cells, leucocytes, hemoglobin and platelets rates in each group, without statistically siginificant difference in both groups. Conclusion: Basing on our work, both minimally invasive adrenalectomies are not only effective but also safe. There were not any significant differences in both groups in the aspect of complete blood count. Both approaches – PRA and LTA can be routinely performed as a treatment of benign adrenal lesions and the choice of the approach depends on surgeon’s experience.

Association of long term exposure to outdoor volatile organic compounds (BTXS) with pro-inflammatory biomarkers and hematologic parameters in urban adults: A cross-sectional study in Tabriz, Iran

This study aimed to compare the hematologic variables and pro-inflammatory biomarkers in urban adults living in Tabriz, Iran, facing various levels of outdoor volatile organic compounds (VOCs). Of all 219 people (212 male and 7 female), 71 were from the low traffic area and 148 were from high traffic and industrial areas. To validate the exposure levels, 93 air samples were taken to determine the target VOCs (benzene, toluene, xylenes, and styrene collectively called BTXS) concentrations in the studied areas. ANOVA and Tukey's tests were used for statistical analysis. Based on the results, significant differences were observed between the mean concentrations of BTXS with the following order of abundance: industrial > high traffic > low traffic. The Considerable decrease was observed in red blood cells (RBCs), hemoglobin, hematocrit, and eosinophils of 0.324 ( × 106/μL), 0.57 g/dL, 1.87%, and 0.17 ( × 103/μL), respectively in industrial area participants as compared to the low traffic area. However, a significant increase was observed in white blood cell count (WBC), neutrophils number, neutrophils percent, TNF-α and INF-γ of 0.88 ( × 103/μL), 0.80 ( × 103/μL), 3.53%, 34.2 ng/mL, and 40.06 ng/mL, respectively in the same groups. The comparison of low and high traffic areas showed significant differences in RBC (p = 0.034), tumor necrosis factor alpha (TNF-α) (p < 0.001), and interferon gamma (INF-γ) (p < 0.001). On the contrary, no significant difference was observed in TNF-α and INF-γ among the high traffic and industrial areas. In conclusion, the results showed that the samples from high traffic and industrial areas were regularly exposed to higher values of BTXS due to traffic and industrial pollutants as compared to the samples residing in low traffic regions. Based on the results living in both high traffic and industrial regions can increase adverse effects on hematologic parameters and pro-inflammatory cytokines.

Repeatability of regional pulmonary functional metrics of Hyperpolarized 129 Xe dissolved-phase MRI.

MRI of hyperpolarized 129 Xenon (HP 129 Xe) is increasingly utilized for investigating pulmonary function. The solubility of HP 129 Xe in lung tissue, blood plasma (Barrier), and red blood cells (RBC), with unique chemical shifts, enables spectroscopic imaging of potential imaging biomarkers of gas exchange and microstructural pulmonary physiology. To quantify global average and regional repeatability of Barrier:gas, RBC:gas, and RBC:Barrier ratios derived from dissolved-phase 129 Xe imaging and their dependence on intervisit changes in lung inflation volume. Prospective. Fourteen healthy volunteers. One subject was unable to complete the study resulting in 13 subjects for analysis (eight female, five male, ages 24-69, 53.8 ± 13.9). 1.5T. Subjects were imaged using a 3D radial 1-point Dixon method to separate Barrier and RBC component signals, at two different timepoints, with ~1 month between visits. RBC:Gas, Barrier:Gas, and RBC:Barrier measures were compared across time and with pulmonary function tests (PFTs). Repeatablilty was quantified using Bland-Altman plots, coefficient of repeatability, coefficient of variation (CV), and intraclass correlation coefficients (ICCs). Dependence of imaging measures on PFTs and lung volume was evaluated using Spearman and Pearson correlation coefficients, respectively. Statistical significance was determined by F-test for intraclass correlations, and t-test for Spearman correlations and regression. Mean RBC:Gas, Barrier:Gas, and RBC:Barrier had CVs of 19.2%, 20.0%, and 11.5%, respectively, and had significant ICCs, equal to 0.78, 0.79, and 0.92, respectively. Intervisit differences in RBC:Barrier were significantly correlated with intervisit differences in DLCO (r = 0.93, P = 0.007). Significant correlations with intervisit lung volume differences and intervisit differences in mean RBC:Gas (r = -0.73, P = 0.005) and Barrier:Gas (r = -0.69, P = 0.009) were found. Three commonly used 129 Xe MRI-based measures of gas-exchange show good repeatability, particularly the Barrier:RBC ratio, which did not depend on lung inflation volume and was strongly associated with intervisit changes in DLCO . 1 Technical Efficacy: Stage 1 J. Magn. Reson. Imaging 2019;50:1182-1190.

Non-Occlusive Retinal Vascular Inflammation and Role of Red Blood Cell Deformability in Birdshot Chorioretinopathy

ABSTRACTPurpose: To investigate differences in red blood cell (RBC) deformability between birdshot chorioretinopathy (BCR) subjects and matched controls, and to postulate its relationship with lack of vascular occlusion in BCR.Methods: In a single center, prospective, non-randomized mechanistic study, blood samples were collected from eight healthy controls and nine BCR patients, and subjected to biochemical and hematological tests, as well as RBC indices assessment using dual-beam optical tweezers.Results: The mean age of the controls was 52.37 ± 10.70 years and BCR patients was 53.44 ± 12.39 years. Initial cell size (Io) for the controls was 8.48 ± 0.25 μm and 8.87 ± 0.31 μm for BCR RBCs (p = 0.014). The deformability index (DI) for the controls was 0.066 ± 0.02 and that for BCR RBCs was 0.063 ± 0.03 (p = 0.441).Conclusion: There was no statistically significant difference in DI between RBCs from BCR and healthy controls. This may explain the rare occurrence of retinal vascular occlusion despite the underlying vasculitic pathophysiology of BCR.

Valuation of three different anaesthetic protocols on complete blood count and biochemical parameters on Wistar rats

The objective of the current study was to determine the impact of three different anaesthetic protocols on blood and biochemical parameters. Eighteen female Wistar rats (HsdOla:WI) 3 months old, weighting 197.09 ± 7.39 g were used. Baseline blood collection was performed in all animals from the lateral coccygeal vein for evaluation of glucose. The animals were then randomly allocated to receive one of three different anaesthetic protocols: dexmedetomidine/ketamine intramuscularly (0.25 mg/kg and 50 mg/kg respectively), or isoflurane 0.2 ml on cotton inside a syringe case, or isoflurane administered by vaporiser (5% induction and 2-3% maintenance of anaesthesia, delivered in oxygen flow 1 L/min). Blood samples were collected from caudal vena cava for complete blood count and biochemical analysis, while the lung and the liver were harvested for histological evaluation. Comparison between groups, as far as complete blood count parameters is concerned, revealed statistical significant differences in red blood cells, haematocrit and haemoglobin values with lower values being observed in vaporiser anaesthesia group. Furthermore, within group comparison revealed statistical significant differences for glucose in all three groups. No evidence of inflammatory, degenerative neoplastic or toxicity related lesions were observed during histological evaluation of the lung and liver.

Efficacy and safety of Danggui Buxue Decoction in combination with western medicine treatment of anemia for renal anemia: a systematic review and meta-analysis.

To evaluate the efficacy and safety of Danggui Buxue Decoction for renal anemia when combined with western medicine treatment of anemia. Electronic searching Medline, Embase, Web of Science, Cochrane Library, Chinese BioMedical Literature Database (CBM), China National Knowledge Infrastructure (CNKI), WanFang data, Chinese Sci-tech periodical full-text database (VIP). Randomized controlled trials reported results of efficacy and safety of Danggui Buxue Decoction in combination with western medicine treatment of anemia for renal anemia. The "risk of bias assessment tool (Version 5.1.0)" of Cochrane Handbook was applied to assess the quality of included trials and RevMan 5.3 software was used for data analysis. A total of 111 studies was retrieved, seven studies including 460 cases were included, the methodological quality of included trials was poor. The result of meta-analysis demonstrated that there was no difference in hemoglobin (Hb) [weighted mean differences (WMD) =-8.75, 95% confidence interval (CI): (-18.64, 1.13), P=0.08], whereas the subgroup analysis showed the difference was significant when the ratio of Radix Astragali to Radix Angelicae Sinensis was 5:1 [WMD =-16.27, 95% CI: (-28.73, -3.80), P=0.01], increase of Hb was more effective in experimental group than control group and the difference was not significant when the ratio of Radix Astragali to Radix Angelicae Sinensis was 5≠1 [WMD =-0.57, 95% CI: (-4.52, 3.39), P=0.78]. There were significant differences in red blood cell (RBC) [WMD =-0.49, 95% CI: (-0.69, -0.28), P<0.00001], hematocrit (HCT) [WMD =-1.92, 95% CI: (-3.15, -0.69), P=0.002] and clinical efficacy [odd ratio (OR) =0.30, 95% CI: (0.13, 0.69), P=0.004] between Danggui Buxue Decoction combination group and control group, the experimental group was better than control group. There was no adverse event reported in the experimental group. Danggui Buxue Decoction in combination with conventional western medicine (CWM) for renal anemia might be superior to CWM alone and there was no adverse event in the experimental group, it might be more effective when the ratio of Radix Astragali to Radix Angelicae Sinensis was 5:1. However, the quality of included studies was not high, and less attention was paid to the safety, high quality randomized controlled trials are needed to further confirm the findings.

Establishing Reference Ranges of Hematological Parameters from Malian Healthy Adults.

IntroductionMeasurement of immuno-hematological parameters has been historically helpful in the diagnosis and treatment monitoring of many infectious diseases and cancers. However, these parameters have not yet been established in many developing countries where patient care strongly relies on such low-cost tests. This study describes the immuno-hematological parameter ranges for Malian healthy adults.MethodsA cross sectional study was conducted from August 2004 to May 2013. We included 213 healthy volunteers (173 male and 40 female), aged between 18–59 years. Median, 2.5 and 97.5 percentile ranges for each immuno-hematological parameter are presented.ResultsIn our study population, the hematological parameters’ ranges were mostly different to the universal established ranges. We found in our population a Median white blood cell (WBC) count of 5200 cells/μL [3237.5–11900], Red Blood Cell (RBC) count of 4.94 10^6 [3.56–6.17], hemoglobin (Hb) of 14.2 g/dL [12.2–17.38], platelet count (Plt) of 275 10^3/μL [145.4–614.4], lymphocytes 2050/μL [1200–3800], neutrophils 2200/μL [1040–6220]; monocytes 200/μL [100–660]; eosinophils 131/μL [0–1026]; CD4 902 cells/μL [444–1669] and CD8 485 cells/μL [0–1272]. We found significant gender differences in RBC, Hb level and MPV. However, RBC and Hb were higher in males median values compared to females (median values) (p<0.001), whereas the Mean platelet volume lower values (MPV) in males than females (P<0.047). The hemoglobin level for some West African countries (Mali, Burkina Faso, Togo, and Nigeria) ranged from 13.5 to 15.1 g/dL for males and 12 to 13 g/dL for females. However in East and Southern Africa, the values were anywhere from 14.1 to 16.1 for males and 11.2 to 14.4 for females.ConclusionOur data may help physicians to better define hematological abnormalities in patients. They may also be used to define new “normal hematological values” in Malian population or in the whole West African population.

Sex-Related Differences in Hematological Parameters and Organosomatic Indices of Oreochromis niloticus Exposed to Aflatoxin B1 Diet.

A 24-week feeding experiment was conducted to assess whether males and females of Oreochromis niloticus exhibit differences in their hematological responses and organosomatic indices to dietary AFB1 contamination. Triplicate groups of O. niloticus (initial body weight: 24.1 ± 0.6 g) were fed with four diets (Diets 1 to 4) containing 0, 20, 200, and 2,000 μg AFB1 kg−1. A significant decrease (P < 0.05) in hemoglobin (Hb), red blood cells (RBC), and hematocrit (Hct) was observed in AFB1 exposure groups, with the lowest levels recorded in the 2000 μg AFB1 kg−1 treatment. A significant increase in mean white blood cells (WBC), neutrophils, and lymphocytes was observed in AFB1 exposure groups. No sex-related differences in RBC, WBC, lymphocytes, monocytes, and neutrophils levels were observed. However, hemoglobin and hematocrit values for female O. niloticus were significantly lower than those for male O. niloticus. Organosomatic indices showed that the relative liver, kidney, and spleen weights were significantly higher (P < 0.05) in the AFB1 supplemented group than in the control group. However, the effect of aflatoxin on organosomatic indices does not depend on sex but rather depends on the dose of aflatoxin in the diet. These results provide useful information for monitoring changes in the health status of male and female O. niloticus.

Erythroid-Specific Variant of the Nuclear Exportin Xpo7 Conserved Only in Mammals May Explain Functional Differences Between Mammalian Definitive and Lower Vertebrate (or Primitive) Erythropoiesis

Exportin 7 (Xpo7) was previously found to be crucial for late murine erythroid nuclear maturation when its knockdown significantly inhibited chromatin condensation and ultimate enucleation in a primary mouse erythroblast culture [1]. Additionally, Xpo7 was found to have an erythroid-specific isoform, transcribed from an erythroid-specific transcription start site (TSS) in the first intron of Xpo7 (named Xpo7-1B in contrast to the ubiquitously expressed variant -1A) [1]. Xpo7-1B differs from Xpo7-1B in 8 amino acid residues in the first exon and 9 residues in the 5th exon, and these differences are not conserved either in lower vertebrates or in primitive mouse erythropoiesis. It is intriguing that both lower vertebrate and primitive mammalian erythroblasts do not enucleate.In order to understand the in vivo function of Xpo7 and determine if these differences in the two isoforms confer differential functions to the proteins or are simply regulatory, we examined the definitive fetal liver erythropoiesis (considered adult "stress erythropoiesis") and adult steady state erythropoiesis of transgenic mouse models lacking either Xpo7-1A, Xpo7-1B, or both. We had previously knocked down Xpo7 in zebrafish using morpholinos (zebrafish only express the Xpo7-1A homologue) and found no effect on adult erythroid differentiation or proliferation. Adult Xpo7-1A knockout (KO) mice are born in normal Mendelian ratios, are not anemic, and express normal Xpo7-1B only in the blood. Closer examination reveals that fetal liver erythroid proliferation is actually higher in Xpo7-1A KO mice than either heterozygous (HET) or wild type (WT) while differentiation is normal. The double KO (Xpo7-1AB KO) appears to be embryonic lethal as only 4 live knockout pups were born out of 71 live births and even these 4 pups appear to be partial chimeras. We characterized the Xpo7-1AB HET mice in more detail and fetal liver erythroblasts of HET mice showed normal proliferation and differentiation compared to WT, and adults showed no differences in red blood cell (RBC) counts, hematocrit or hemoglobin. No complete live Xpo7-1AB KO has been born so we are now evaluating embryos at E3, E9.5, and E14.5. Given the possibility of embryonic lethality, in order to evaluate the erythroid-specific isoform Xpo7-1B, we have crossed a mouse floxed for the Xpo7-1B TSS/1st exon with a tamoxifen-inducible Cre-recombinase mouse. These mice have been given tamoxifen after reaching adulthood and we are currently evaluating their hematopoietic parameters. We have also set up HET-HET timed matings and administered tamoxifen to pregnant females to evaluate fetal liver erythropoiesis in Xpo7-1B HET, WT, and KO embryos.Not only will this work further our understanding of the process of global chromatin condensation during erythropoiesis, data from this in vivo study may reveal differences in erythropoiesis between mammals and lower vertebrates, which may also reflect intrinsic differences in mammalian primitive versus definitive erythropoiesis. Based on our in vitro knockdown studies, we anticipate that the phenotype of the Xpo7-1B KO mice may be similar to congenital dyserythropoietic anemia (CDA), a human disorder involving abnormalities in erythroid progenitor chromatin condensation and nuclear membrane substructure, so examination of these mice may also uncover a novel mechanism for this form of ineffective erythropoiesis.[1] Hattangadi, SM, et al. Blood (2014) 124, 1931-40. DisclosuresNo relevant conflicts of interest to declare.

Effects of Age, Breed and Sex on Haematological Parameters of Growing Omani Goat Breeds

Sixty seven healthy growing goat kids of three Omani breeds, born and raised in Sultan Qaboos University Agricultural Experiment Station, were used to study the effects of age, breed and sex on haematological parameters. Blood samples were collected at several times from the first to 25 weeks to study the following parameters: Red Blood Cells (RBCs), White Blood Cells (WBCs), Packed Cell Volume (PCV), Haemoglobin (Hb), Mean Cell Volume (MCV), Mean Cell Haemoglobin Concentration (MCHC) and Mean Cell Haemoglobin (MCH). At birth, MCV and MCH levels were significantly higher in Batina (BAT) kids than both Jabal Akhdar (JA) and Dhofari (DOF), but there were no breed differences in the rest of the haematological values. There were significant breed differences in RBC, MCH, WBC, MCV at different ages, with Hb and PCV being highly significant at all ages. All blood parameters changed with time but showed different patterns in all breeds. There was no effect of sex on RBC, MCHC, PCV, Hb or WBC, but female kids had significantly higher MCH and MCV values at week 12. This study indicates that haematology parameters in Omani goat kids could be affected by age, breed and sex.

Comparison of serum cytokine levels between dogs with multicentric lymphoma and healthy dogs

In humans, multiple cytokines have been linked to the development of lymphoma, and are relevant biomarkers for response to chemotherapy and prognosis. In contrast, only a few circulating cytokines have been studied in dogs with lymphoma. We prospectively enrolled thirty-one dogs newly diagnosed with multicentric lymphoma. Immunophenotype was determined by flow cytometry in all dogs, separating them into 2 subgroups: B cell lymphoma (n=21) and T cell lymphoma (n=10). Nineteen healthy dogs were enrolled in the control group. Circulating cytokine concentrations were measured using a commercial canine multiplex magnetic bead-based assay which included Interleukin-2 (IL-2), IL-6, IL-7, IL-8, IL-10, IL-15, IL-18, Granulocyte Macrophage Colony-Stimulating Factor (GM-CSF), Tumor Necrosis Factor-α (TNF-α), Interferon γ (IFN-γ), IFN-γ induced Protein-10 (IP-10), Keratinocyte Chemoattractant-like (KC-like), and Monocyte Chemoattractant Protein-1 (MCP-1). The serum levels of each cytokine were first compared between the lymphoma and control groups, and then between the B cell lymphoma, T cell lymphoma, and control groups. There was no significant difference between the lymphoma and healthy control groups regarding sex, age and weight. MCP-1, IL-6, and IL-10 were significantly higher in dogs with lymphoma compared to healthy dogs (p<0.01, p=0.01 and p=0.03, respectively). MCP-1 and IL-10 were significantly higher in the B cell lymphoma group than in the healthy group (p=0.01, p=0.01, respectively). MCP-1 and IL-6 levels were significantly higher in the T cell lymphoma group than in the healthy group (p=0.02, p<0.01, respectively). IL-6 was significantly higher in the T cell lymphoma group than in the B cell lymphoma group (p=0.03). Significant differences among the groups were found for IL-15 and KC-like, but they were affected by age and/or sex. There were no significant differences in serum IL-2, IL-7, IL-8, IL-18, GM-CSF, TNF-α, IFN-γ, and IP-10 between any of the groups. Significant differences in red blood cell, white blood cell, neutrophil, lymphocyte and monocyte counts were also found between the different groups of dogs. Our data showed different serum cytokine and peripheral blood cell profiles between dogs with lymphoma and healthy dogs, and between dogs with B cell and T cell lymphoma. Further study is necessary to investigate the role of these cytokines in lymphoma pathogenesis, response to treatment, and prognosis, and the influence of age, sex and blood cell counts on their expression.

Failure to Achieve Recommended ω‐3 DHA Intake During Pregnancy: Nutrition Education Needed

Dietary Guidelines for Americans 2010 recommends pregnant or breastfeeding women consume 8–12 oz of seafood/week, translating to an average DHA+EPA intake of 200–250 mg/day. In an on‐going study of overweight pregnant women in Louisiana, we assessed dietary intake and fatty acid status of red blood cells (RBC) mid‐pregnancy. Women (n=10, 20–34 years of age) entered the study at 17–20 weeks of pregnancy (age: n=5 ≥25 y, n=5 &lt;25 y; ethnicity: n=5 African American, n=5 White; SES: n=5 low SES, n=5 med/high SES). Dietary intake was assessed by repeated (n=7) 24‐hour dietary recalls using the University of Minnesota Nutrient Data System for Research (NDSR). Red blood cell fatty acid concentrations were determined with gas chromatography at the time of study entrance. The relative weight percentage (wt%) of each RBC fatty acid identified was determined from the retention time. On average, women consumed 93±59 mg DHA+EPA/day via diet; n=1 achieved the recommended intake of 200 mg/day. Taking prenatal supplementation into consideration, the average DHA+EPA intake was 199±96 mg/day; n=5 achieved the recommended intake of 200 mg/day. While all enrolled women (n=10) consumed prenatal supplements during pregnancy, not all prenatal supplements contained DHA+EPA (n=1). On average, prenatal supplements provided an additional 121±77 mg DHA+EPA/day. Including prenatal supplement consumption, the average ω‐6:ω‐3 ratio was 1.01±0.78. There was a significant difference in DHA+EPA intake between the low SES and med/high SES women, (P=0.01); low SES had higher intake (255±70 mg/day) than med/high SES (115±78 mg/day). Ethnicity and age did not significantly affect DHA+EPA intake (P≥0.3). The top 5 dietary sources of DHA+EPA were crawfish, chicken, shrimp, eggs, and crab. Other seafood sources commonly consumed included catfish, oysters, and tilapia. In terms of fatty acid status, the wt% of RBC DHA was 9.25±1.3 (range: 7.33–11.05) and RBC EPA was 0.14±0.03 (range: 0.1–0.22). There were no significant differences in RBC DHA+EPA status for SES, age, or ethnicity (P≥0.2). Furthermore, there was no correlation between DHA+EPA intake and RBC concentrations (r=−0.06). The latter observation may be explained, in part, by the fact that DHA is preferentially transferred across the placenta to meet fetal demands during gestation. These data point to a need for nutrition education for pregnant women and women of childbearing ages regarding the importance of consuming fish varieties that are highly enriched in DHA+EPA.Support or Funding InformationFunded in part by: LSU Undergraduate Research Grant (C Thaxton); USDA AFRI 2012‐67017‐19293 (CJL‐K); and LSU AgCenter.

Massive Transfusion Protocols: Indications, Ratios and Mortality in the Non-Trauma Setting

Background: Many institutions have implemented massive transfusion protocols (MTPs) to prevent hemodilution and to restore normal coagulation function, with the ultimate goal of controlling hemorrhage and reducing complications. Our institution issues two different MTPs: trauma (T-) and non-trauma (NT-). T-MTPs have a 1:1 ratio for red blood cell (RBC):plasma issued by the blood bank, whereas NT-MTPs have a 1.8:1 ratio. Appropriate blood product ratios, indications and patient outcomes in the NT-MTP setting are not well studied. To determine how various MTP parameters impact 24-hour patient mortality, we retrospectively reviewed MTP activations at our large academic urban medical center.Methods: All activated MTPs over a 3-year period (2012-2014) were reviewed. Data was collected from blood bank quality assurance and inpatient electronic medical records. NT-MTPs were sub-classified into indication by type of hemorrhage. All statistical analyses (binary logistic regression, Kruskal-Wallis) were performed using STATA version 11. A p value of <0.05 was considered significant.Results: From 2012-2014, there were 177 MTP activations for 167 patients, of which 98 were male (59%) and 69 were female (41%). The average age of all patients was 56 years, with a range of 7 months to 95 years. Trauma patients (mean age 40 years) tended to be younger than non-trauma patients (mean age 60 years). Refer to Table 1 for types of hemorrhage and ratios of blood products transfused. Thirty-eight patients (22.8%) died within 24 hours of MTP activation, including 10 (30.3%) of the trauma patients and 28 (21.7%) of the non-trauma patients (Figure 1). Mortality did not vary significantly by type of hemorrhage or by ratio of RBC:plasma transfused, including patients receiving no plasma. For each additional RBC unit transfused, patients had a higher chance of dying (odds ratio [OR] 1.17; p=0.002, confidence interval 1.1-1.3) within 24 hours, after controlling for number of platelet, plasma, and cryoprecipitate units received. The overall median RBC:plasma ratio transfused was 1.7 (interquartile range [IQR] 1.3-2); T-MTPs had a median ratio of 1.4 (IQR 1.1-1.9) and NT-MTPs had a median ratio of 1.7 (IQR 1.3-2.1). There was no significant difference in RBC:plasma ratios clinicians transfused for different types of hemorrhage, despite the blood bank issuing different ratios to the clinicians for T-MTPs and NT-MTPs. The total number of RBCs, platelet units, and plasma units transfused did not differ by type of hemorrhage. In all MTPs, transfusion of platelets did not have a significant impact on 24-hour survival.Conclusions: We found that only the number of RBC units transfused had a significant association (OR 1.17) with 24-hour mortality during an MTP. The RBC:plasma ratio, number of platelets or plasma, and use of platelets during an MTP did not affect 24-hour mortality. The number of RBC units transfused most likely reflects the clinician's assessment of the severity of the situation, and does not imply that the RBCs affected the 24-hour mortality. We found the ratio of products issued during an MTP was not what was actually transfused to patients, indicating that clinicians were not transfusing according to protocol. Specifically, for NT-MTPs overall, the RBC:plasma ratio transfused was lower than what the blood bank issued, indicating clinicians are choosing to infuse more plasma despite a lack of evidence in the non-trauma setting. Prospective randomized trials comparing different RBC:plasma:platelet ratios in NT-MTPs are warranted.Table 1Blood Product Ratios Transfused by Type of HemorrhageType of HemorrhageNumber of Patients (%)RBC:Plasma - Median (IQR)RBC:Platelets - Median (IQR)Overall167 (100%)1.7 (1.3-2)5 (4-7)Trauma33 (20%)1.4 (1.1-1.9)5.5 (3.7-8.4)Postoperative50 (30%)1.6 (1.3-1.8)5 (3.5-6)Gastrointestinal29 (17%)2 (1.7-2)5 (4-7)Intraoperative20 (12%)1.6 (1.3-2.7)6.2 (3.3-7.5)Abdominal11 (7%)2 (1.3-3.3)6 (4-7)Vascular9 (5%)1.3 (1-2)5 (2-5)Obstetrical8 (5%)2.2 (2-2.3)5 (4-7)Central Nervous System4 (2%)1.8 (1.2-4)4.2 (4-4.3)Pulmonary2 (1%)22Superficial Soft Tissue1 (0.5%)1N/A [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Antigen Density Impacts RBC Survival and Antigen Modulation Following Incompatible RBC Transfusion

Background: While chronic transfusion therapy can signficantly reduce morbidity and mortality in patients with hemoglonbinopathies, transfusion is not without risk. Differences in red blood cell (RBC) antigen distributions between donors and recipients can result in the development of RBC alloantibodies, which can make it difficult, if not impossible, to find compatible blood. As a result, heavily alloimmunized individuals experience greater morbidity and mortality than those who do not possess RBC-induced alloantibodies. While incompatible RBC transfusion can result in fatal hemolytic transfusion reactions (HTRs), these adverse events are not the invariable outcome of incompatible transfusion. Although a variety of factors likely contribute to the likilihood of developing a HTR following incompatible RBC transfusion, variability in the density of the donor antigen may contribute to different outcomes. Using a novel model that utilizes RBC donors with different levels of the KEL antigen, we sought to directly examine the impact of antigen density on RBC clearance following incompatible RBC transfusion.Methods: Wild type C57BL/6 mice (WT) were passively immunized with anti-KEL serum, generated following immunization of WT recipients with KEL RBCS, 2 hours prior to incompatible KEL RBC transfusion. RBC collected from previously generated KEL transgenic mice that express high (KELhi), intermediate (KELmed), and low (KELlo) levels of human KEL glycoprotein antigen, were labeled with chloromethylbenzamido 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI), while WT RBC were labeled with fluorescently distinct 3,3'-dihexadecyloxacarbocyanine perchlorate (DiO) to facilitate detection post-transfusion. Passively immunized recipient WT mice were transfused with labeled KEL RBC to WT RBC. Percent KEL RBC survival was measured using flow cytometric analysis at various time points post transfusion by comparing the DiI to DiO ratio. Bound antibody, complement and the KEL antigen were assessed by staining cells with anti-Ig, anti-C3 and anti-KEL followed by flow cytometric analysis.Results: Transfusion of RBCs expressing low, intermediate or high levels of the KEL antigen into non-immunized mice failed to result in any detectable alteration in RBC survival or KEL expression. However, transfusion of KELlo, KELmed and KELhi RBCs into anti-KEL immunized recipients resulted in differential clearance of each population, with KELhi experiencing the most clearance, followed by KELmed and KELlo. Similarly, the level of detectable C3 and bound antibody on KEL positive cells likewise correlated with the initial level of KEL antigen expression, with more C3 and IgG being detected on the surface of KELhi following transfusion into immunized recipients, than KELmed or KELlo. However, KELhi RBCs also experience the most significant alterations in KEL antigen levels following antibody engagement. Regardless of the KEL positive RBC transfused, each population developed a relatively rapid state of resistance to additional anti-KEL-mediated RBC removal.Conclusions: Our results indicate that lower antigen densities may be less efficient in not only facilitating rapid clearance of RBCs following incompatible transfusion, but also antibody-induced antigen modulation. While KELhi RBCs displayed the greatest degree of clearance and antigen modulation, each population developed resistance to antibody-mediate removal, suggesting that once RBC antigen levels fall below a certain threshold, clearance discontinues, regardless of the initial levels of antigen prior to incompatible transfusion. Taken together, these results suggest that antigen density may provide an additional feature of RBC transfusion that may impact the outcome of incompatible transfusion in heavily alloimmunized recipients. DisclosuresNo relevant conflicts of interest to declare.

Hemorheological Approach for Early Detection of Chronic Kidney Disease and Diabetic Nephropathy in Type 2 Diabetes.

Hemorheologic alterations or changes in blood viscosity have been suggested to play a role in the pathogenesis of microvascular complications in diabetes. We measured various hemorheologic parameters in type 2 diabetes patients at different stages of chronic kidney disease (CKD) and assessed their possible role as early markers of diabetic nephropathy and renal insufficiency. One hundred-five patients with type 2 diabetes were divided into four groups according to glomerular filtration rate (GFR), which represents the kidney function. Hemorheologic parameters, including erythrocyte deformability, fibrinogen/elongation index (EI), and aggregation index (AI) were measured using a microfluidic hemorheometer, and critical shear stress (CSS) was measured using a microfluidic technique. Various metabolic parameters were assessed from fasting blood samples, and the urine albumin-to-creatinine ratio (ACR) was calculated from first morning voided urine. There were significant differences in red blood cell (RBC) deformability, AI, CSS, fibrinogen/EI, and ACR among patients in different stages of CKD (all P<0.05). RBC deformability and fibrinogen/EI significantly differed between normal (GFR >90 mL/min/1.73 m(2)) and CKD stage 2 (GFR 60-90 mL/min/1.73 m(2)) patients, whereas there was no such difference in ACR. In multiple regression analysis, fibrinogen/EI under a moderate shear stress of 3 Pa was an independent predictor of GFR (β=-0.328, P<0.05). Also, AI, CSS, and fibrinogen/EI were significantly different among patients at different stages of diabetic nephropathy, with a significant difference in fibrinogen/EI between normal and microalbuminuric patients (all P<0.05). RBC deformability and fibrinogen/EI are sensitive parameters measured via point-of-care testing for detecting erythrocyte alterations in early CKD and nephropathy in patients with type 2 diabetes. Further studies are warranted to verify their use as screening tools for diabetic nephropathy and renal impairment.

No increase in blood transfusions during liver transplantation since the withdrawal of aprotinin

The aims of this study were to determine whether the withdrawal of aprotinin (APRO) led to an increased bleeding risk in patients undergoing orthotopic liver transplantation (OLT). A retrospective analysis compared consecutive patients undergoing OLT and treated with aprotinin (APRO group; n = 100) with a group in which aprotinin was not used (no-APRO group; n = 100). Propensity score matching was then performed for each group to identify 2 matched cohorts. Patients were matched by their primary diagnoses and Model for End-Stage Liver Disease scores. This resulted in 2 matched cohorts with 55 patients in each group. None of the patients in the APRO group had significant fibrinolysis. In the no-APRO group, 23.6% of the patients developed fibrinolysis (P < 0.003). Tranexamic acid was used in 61.5% of the patients (n = 8) in the no-APRO group in whom lysis was present, and this resolved the fibrinolysis in all but 1 of these patients. There were no differences in red blood cell, fresh frozen plasma, platelet concentrate, or cryoprecipitate transfusions between the 2 groups. In conclusion, we have shown a significant increase in the prevalence of fibrinolysis during OLT since the withdrawal of APRO. However, there has been no increase in transfusion requirements.

Effect of whole egg powder on growth performance, blood cell counts, nutrient digestibility, relative organ weights, and meat quality in broiler chickens

A 5-week experiment using 660 male hatchling ROSS 308 broilers was conducted to investigate the effect of whole egg powder (EP) on the growth performance, blood cell counts, nutrient digestibility, relative organ weights, and meat quality in broiler chickens. Broilers were randomly allotted to four dietary treatments consisting of a typical basal diet and that diet with 1%, 2% or 3% EP substituted, isocalorically and isonitrogenously. During d 1–25, and d 1–35, BWG was linearly enhanced in broilers fed EP (P=0.04). Between d 1 and 25, and for the overall period to d 35, FCR was linearly increased (P=0.01) by adding EP. No significant effect of EP was observed on apparent digestibility of dry matter or nitrogen among treatments but energy digestibility was linearly enhanced by inclusion of EP (P=0.04).The white blood cell count was linearly higher (P=0.04) in birds fed with EP but there were no differences in red blood cells or percent lymphocytes. The drip loss of breast meat after 24h, 48h, and 72h of refrigerated storage was linearly reduced 50–60% by feeding EP (P=0.01). In conclusion, inclusion of up to 3% whole egg powder in diets of broilers was shown to improve growth performance to d 35, energy digestibility and decreased drip loss of breast meat.

Hematological parameters of pike Esox lucius in relation to different ages and seasons

The purpose of this study was to assess of some hematological parameters of Esox lucius collected from Anzali wetland from fall 2010 to summer 2011. Variations in hematological parameters such as hematocrit (Ht), hemoglobin concentration (Hb), erythrocyte (RBC) and leukocyte (WBC) counts, mean cell volume (MCV), mean cell hemoglobin (MCH), mean cell hemoglobin concentration (MCHC), lymphocytes, monocytes, neutrophils, and eosinophils of fish were compared according to age and seasonal differences. Analysis of variance showed significant differences in values of white blood cell (WBC), red blood cell (RBC), MCV, MCH, lymphocytes, monocytes, and eosinophils between ages. Statistical analysis revealed significant differences in RBC and WBC between all seasons. However, there were no significant differences in Ht, Hb, MCHC, MCH, MCV, neutrophils, lymphocytes, monocytes, and eosinophils between all seasons. Results of current study may be helpful in establishing a hematological data bank for this species including standard values of blood parameters.

Blood group typing in five Afghan populations in the North Hindu‐Kush region: implications for blood transfusion practice

Blood incompatibility arises from individual and ethnic differences in red blood cell (RBC) antigen profiles. This underlines the importance of documenting RBC antigen variability in various ethnic groups. Central Asia is an area with a long and complex migratory history. The purpose of this article is to describe key antigen frequencies of Afghan ethnic groups in the Hindu-Kush region of Afghanistan as a basis for improving blood transfusion practices in that area. The key ABO, Rh and Kell antigens were investigated in five Afghan populations. In order to depict accurately the blood group gene diversity in the area, DNA from eight additional Pakistani populations were included, and the entire sample set screened using two multiplex polymerase chain reactions sensitive for 17 alleles in 10 blood group genetic systems (MNS, Kell, Duffy, Kidd, Cartwright, Dombrock, Indian, Colton, Diego and Landsteiner-Wiener). Phenotype and allele frequencies fell within the ranges observed in Western European and East Asian populations. Occurrence of DI*01, IN*01, LW*07 and FY*02N.01 and prevalence of ABO*B were consistent with migratory history as well as with putative environmental adaptation in the subtropical environment Hindu-Kush region. These findings expand the current knowledge about key antigen frequencies. Regarding occurrence of viral markers, further blood transfusion in the region requires rigorous typing.