Animal Hemoglobins Research Articles

Coadministration of PEGylated apohemoglobin and haptoglobin can limit vascular dysfunction in the microcirculation and prevent acute inflammation.

Unfortunately, during pathological conditions resulting in chronic hemolysis cell-free hemoglobin (Hb) is released into the circulation that releases free heme, resulting in several complications. One approach to prevent these toxicities is the administration of supplemental scavenger proteins, haptoglobin (Hp) and hemopexin (Hpx). The goal of this body of work is to objectively measure the levels of vascular reactivity and inflammatory profiles after an infusion of acellular hemoglobin in animals that were given a coadministration of PEGylated human apohemoglobin (PEG-apoHb), a hemopexin (Hpx)-mimetic that can scavenge free heme from hemoglobin, together with human plasma-derived Hp that can scavenge dimerized Hb. Using intravital microscopy, Golden Syrian hamsters instrumented with a dorsal window chamber were used to evaluate the in vivo effects of four experimental groups that were then challenged with a hypovolemic injection (10% of the animal's blood volume) of human Hb (hHb, 5 g/dL). The four experimental groups consisted of: 1) lactated Ringer's solution (control), 2) PEG-apoHb only, 3) Hp only, and 4) PEG-apoHb + Hp. The microvascular hemodynamics (diameter and flow) in arterioles and venules were recorded at baseline, 20 min after treatment, and 20 min after hHb challenge. Systemic parameters (blood pressure and heart rate), blood gases (pH, Pco2, and Po2), blood parameters (Hb concentration and hematocrit), and multiorgan functionality/inflammation were also measured. Our results suggest that coadministration of PEG-apoHb + Hp as a booster before the infusion of acellular hemoglobin significantly prevented vasoconstriction in the microcirculation, significantly increased the number of functional capillaries, and significantly reduced inflammation.NEW & NOTEWORTHY Coadministration of PEGylated human apohemoglobin (PEG-apoHb)-a hemopexin (Hpx) mimetic that can scavenge free heme-and human plasma-derived haptoglobin (Hp) that can scavenge hemoglobin (Hb), reduces microcirculatory dysfunction and cardiac and kidney inflammation in a Hb-challenge model.

Non-invasive estimation of in vivo optical properties and hemodynamic parameters of domestic animals: a preliminary study on horses, dogs, and sheep.

Biosensors applied in veterinary medicine serve as a noninvasive method to determine the health status of animals and, indirectly, their level of welfare. Near infrared spectroscopy (NIRS) has been suggested as a technology with this application. This study presents preliminary in vivo time domain NIRS measurements of optical properties (absorption coefficient, reduced scattering coefficient, and differential pathlength factor) and hemodynamic parameters (concentration of oxygenated hemoglobin, deoxygenated hemoglobin, total hemoglobin, and tissue oxygen saturation) of tissue domestic animals, specifically of skeletal muscle (4 dogs and 6 horses) and head (4 dogs and 19 sheep). The results suggest that TD NIRS in vivo measurements on domestic animals are feasible, and reveal significant variations in the optical and hemodynamic properties among tissue types and species. In horses the different optical and hemodynamic properties of the measured muscles can be attributed to the presence of a thicker adipose layer over the muscle in the Longissimus Dorsi and in the Gluteus Superficialis as compared to the Triceps Brachii. In dogs the absorption coefficient is higher in the head (temporalis musculature) than in skeletal muscles. The smaller absorption coefficient for the head of the sheep as compared to the head of dogs may suggest that in sheep we are indeed reaching the brain cortex while in dog light penetration can be hindered by the strongly absorbing muscle covering the cranium.

Spectroscopic study of methylene blue in vivo: effects on tissue oxygenation and tumor metabolism

Methylene blue (MB) is a promising photosensitizer (PS) for the treatment of pathological neoplasms, since it has both photodynamic activity (under laser irradiation) and redox and catalytic properties (in the absence of light). In the framework of this work, using spectroscopic methods, the effect of intravenous administration of MB on tissue oxygenation of hemoglobin in small animals in vivo in tumor and normal tissues was analyzed. The influence of MB on cell metabolism was analyzed. It has been shown that the use of MB promotes an increase in oxygen consumption by the tumor, and also leads to a shift in metabolism towards oxidative phosphorylation. It was shown that the use of MB contributes to an increase in oxygen consumption by the tumor, and also leads to a shift in metabolism towards oxidative phosphorylation.

Possible Protective Effect of Spanish Black Radish on Hematological Parameters Aganist Benzene - Induced Leukemia in Rats

Background: The Spanish black radish, which is grown all over the world and is full of useful nutrients, helps in the therapy of many illnesses. Objective: The aim of this study was to evaluate the possible protective effect of Spanish black radish extracts against benzene-induced leukemia in rats. Materials and Methods: A total of 35 rats were purchased from the animal house at the College of Pharmacy, University of Karbala, Iraq. Benzene was used for the induction of leukemia in experimental animals. An extract of black radish was set after mixing up 50 g of the finely powdered plant material with two solvents (500 mL of 100% ethanol or water). After leukemia induction, rats were randomly divided into seven groups: first group, negative control group; second group, positive control group; third group, (lack radish aqueous extract 100 mg/kg; fourth group, black radish aqueous extract 200 mg/kg; fifth group, black radish ethanol extract 100 mg/kg; sixth group, black radish ethanol extract 200 mg/kg; and seventh group, cyclophosphamide 5 mg/kg. Results: Leukemia induction resulted in a severe increase in the total leukocyte count and a severe decrease in red blood cells, hemoglobin, and platelet levels of the experimental animals. However, there were no significant differences between the negatively treated group and the benzene-treated group regarding platelet count. Posttreatment with Spanish black radish extracts, especially the aqueous plant extract, shows an improvement in blood profiles compared to the benzene-treated group. Conclusion: Our study indicates that Spanish black radish might be a promising natural, antileukemic source.

Influence of plant extracts on insulin resistance in experimental diabetes mellitus

Objective. The aim of the study was to evaluate the effect of medicinal plant extracts on insulin sensitivity in the model of diabetes mellitus.Material and Methods. Experiments were carried out using 110 outbred white male rats. Diabetes mellitus was induced by the administration of streptozotocin to animals receiving food with a high fat content for the formation of insulin resistance. The animals were injected with metformin, rosiglitazone, and aqueous extracts of nettle, dandelion, burdock, and Galega officinalis. The rats were divided into two groups during pharmacotherapy: group 1 continued to receive fat-enriched diet; group 2 received a regular diet with 8% fat content. The blood levels of glucose, glycated hemoglobin, triglycerides, and high-density lipoprotein cholesterol were measured. The metabolic index was calculated to assess insulin resistance.Results. The extracts of nettle and burdock reduced the concentration of glucose and glycated hemoglobin in animals with a model of diabetes mellitus treated with fat-enriched food. Dandelion extract did not change the level of fasting glucose in blood whereas it lowered the concentration of glycohemoglobin. The extracts of nettle, Galega officinalis, and dandelion reduced blood concentrations of glucose in animals receiving normal diet. The extracts of all plants reduced the level of glycated hemoglobin. The concentration of triglycerides normalized regardless of the diet when plant extracts were introduced. The concentrations of high-density lipoprotein cholesterol increased in both diets in animals treated with Galega officinalis and dandelion extracts. The metabolic index in rats receiving food with 8%-fat content did not differ from that of intact animals after therapy with nettle and Galega officinalis extracts. Dandelion extract alleviated insulin resistance in animals continuing to receive a diet enriched with fats.Conclusion. The extracts of nettle, Galega officinalis, burdock, and dandelion improved carbohydrate and lipid metabolism in diabetes mellitus caused by the administration of streptozotocin to rats to the same extent as metformin and rosiglitazone. Extracts of nettle and Galega officinalis alleviated insulin resistance in animals receiving a normal diet; dandelion extract alleviated insulin resistance in animals receiving fat-enriched food.

Identification and Tetramer Structure of Hemin-Binding Protein SPD_0310 Linked to Iron Homeostasis and Virulence of Streptococcus pneumoniae.

ABSTRACTIron and iron-containing compounds are essential for bacterial virulence and host infection. Hemin is an important supplement compound for bacterial survival in an iron-deficient environment. Despite strong interest in hemin metabolism, the detailed mechanism of hemin transportation in Gram-positive bacteria is yet to be reported. The results of our study revealed that the homologous proteins of SPD_0310 were significantly conservative in Gram-positive bacteria (P < 0.001), and these proteins were identified as belonging to an uncharacterized protein family (UPF0371). The results of thermodynamic and kinetic studies have shown that SPD_0310 has a high hemin-binding affinity. Interestingly, we found that the crystal structure of SPD_0310 presented a homotetramer conformation, which is required for hemin binding. SPD_0310 can interact with many hemin-binding proteins (SPD_0090, SPD_1609, and GAPDH) located on the cell surface, which contributes to hemin transfer to the cytoplasm. It also has a high affinity with other iron transporters in the cytoplasm (SPD_0226 and SPD_0227), which facilitates iron redistribution in cells. More importantly, the knockout of the spd_0310 gene (Δspd_0310) resulted in a decrease in the iron content and protein expression levels of many bacterial adhesion factors. Moreover, the animal model showed that the Δspd_0310 strain has a lower virulence than the wild type. Based on the crystallographic and biochemical studies, we inferred that SPD_0310 is a hemin intermediate transporter which contributes to iron homeostasis and further affects the virulence of Streptococcus pneumoniae in the host. Our study provides not only an important theoretical basis for the in-depth elucidation of the hemin transport mechanism in bacteria but also an important candidate target for the development of novel antimicrobial agents based on metal transport systems.IMPORTANCE Iron is an essential element for bacterial virulence and infection of the host. The detailed hemin metabolism in Gram-positive bacteria has rarely been studied. SPD_0310 belongs to the UPF0371 family of proteins, and results of homology analysis and evolutionary tree analysis suggested that it was widely distributed and highly conserved in Gram-positive bacteria. However, the function of the UPF0371 family remains unknown. We successfully determined the crystal structure of apo-SPD_0310, which is a homotetramer. We found that cytoplasmic protein SPD_0310 with a special tetramer structure has a strong hemin-binding ability and interacts with many iron transporters, which facilitates hemin transfer from the extracellular space to the cytoplasm. The results of detailed functional analyses indicated that SPD_0310 may function as a hemin transporter similar to hemoglobin in animals and contributes to bacterial iron homeostasis and virulence. This study provides a novel target for the development of antimicrobial drugs against pathogenic Gram-positive bacteria.

EFFECT OF ENERGY DRINK ON LIGAND FORMS OF HEMOGLOBIN

The article is devoted to the study of the effect of energy drink on ligand forms of hemoglobin in experimental animals. According to our results, the consumption of energy drink causes significant changes in both the total level of hemoglobin and its ligand forms. In particular, we have established a reduction of oxyhemoglobin and accumulation of dyshemoglobin: methemoglobin, sulfhemoglobin and carboxyhemoglobin, which in turn affects the oxygen homeostasis of the body, the development of tissue hypoxia and can cause structural and functional disorders in the body under such conditions.

Effect of Foliage Dressing in Nano-Potassium Molybdate on Daily Gain and Antioxidant Function in Grazing the Chinese Merino Sheep.

The study was implemented for exploring influences of nano-K2MoO4 on the daily gain and antioxidant function of Chinese merino sheep in the native pasture, Xinjiang Uygur Autonomous Region, Northwest of China. Eighty of the sheep, weight of (45.56 ± 2.35) kg, were randomly distributed to the tested areas for 90 days, 20 sheep/group. The findings showed that the contents of Mo and N in the forage of applying nano-K2MoO4 were extremely higher than those in the control pastures (P <0.01). The daily gain in the fertilized groups were remarkably lower than that in the control group (P <0.01). The harvest of forage in the fertilized pastures were significantly higher than those in control (P <0.01). The contents of Mo in blood and liver in grazing the Chinese merino sheep were extremely higher than those from the control group (P <0.01). The contents of Cu in blood and liver in grazing the Chinese merino sheep were extremely lower than those in the unfertilized pastures (P <0.01). The levels of hemoglobin (Hb), blood platelet (PLT), and erythrocyte count (RBC) in animals from the fertilized pastures were extremely lower than those in the control pastures (P < 0.01). The activities of SOD, GSH-Px, and CAT in serum were significantly lower than those in group C. The serum MDA levels were significantly higher than those in the control group (P < 0.01). In conclusion, the application of nano-K2MoO4 in pastures can greatly improve the yield of forage, but strikingly decreased the daily gain and antioxidant function in grazing the Chinese merino sheep.

Ethanol Extract of Brassica Rapa Attenuate the Complete Freund’s Adjuvant-Induced Inflammation in Rats

Turnip is an ancient edible plant belongs to family Braaicacaceae. Traditionally it is used for the treatment of various disorders such as diabetes, hepatotoxicity, ulcer, and anti-inflammatory activity. Therefore the current investigation was attempted to investigate the anti-inflammatory activity of Brassica rapa root extract against complete fluids adjuvant induced inflammation in experimental animals. The Ethanol extract of Brassica rapa was prepared by a hot extraction procedure and phytochemical analysis of different extract was done by different chemical tests. Daily oral treatment of indomethacin, and extract for twenty-one days after three days of CFA administration significantly decrease the paw volume, hepatic biomarkers as well as ameliorated the level of a hematological parameter such as hemoglobin, RBC, WBC, ESR and bodyweight of experimental animals. From the result of the current investigation, it can be concluded that the Brassica rapa extract possesses anti-inflammatory activity. Further study is required to explore the mechanism responsible for its anti-inflammatory activity.

Improved Hemodynamic Stability in Experimental Trauma/Hemorrhagic Shock by Red Blood Cell‐containing Resuscitation Fluids Secondary to Preserved Diastolic Blood Pressure

ObjectiveFluid resuscitation with blood products is standard of care in the current management of trauma/hemorrhagic shock (T/HS). However, the availability of whole blood is rare in civilian prehospital care, and thus blood component therapy and other fluid replacement options are required. This study sought to differentiate and compare the effects of lactated Ringer’s solution (LR), plasma, washed red blood cells (RBC), and whole blood (Control) as resuscitation fluids for hemodynamic restoration after T/HS.MethodsWister male rats (400g) were randomly divided into four groups: LR, Plasma, RBC, and Control, anesthetized with isoflurane and subjected to experimental T/HS. A mid‐line laparotomy incision was made, and blood was slowly withdrawn to maintain a mean arterial blood pressure (MAP) of 35–40 mmHg for 90 minutes. After the 90‐min shock period, intravenous fluids corresponding to the experimental group were started with the goal of achieving a MAP ≥ 65 mmHg, the clinical target for adequate pressure. Animals were monitored for 120 minutes following start of reperfusion. Vital signs were continuously measured, as well as arterial blood gases, lactate, and hemoglobin.ResultsMAP in animals that received RBCs was similar to that of the whole‐blood Control group (70.4 ± 4.0 mmHg vs 83.0 ± 5.8 mmHg, RBC vs Control, NS), and significantly greater than MAPs of Plasma (59.9 ± 4.4 mmHg, p=0.03) and LR (45.6 ± 4.8 mmHg, p&lt;0.001) groups. As systolic blood pressure (SBP) was not significantly different between RBC, Plasma and Control groups, the higher MAP seen in the RBC group compared to Plasma was due to increased diastolic blood pressure (DBP) (55.5 ± 3.1 mmHg vs 38.6 ± 3.2 mmHg, RBC vs Plasma, p=0.01). The RBC and Control groups were able to achieve a higher MAP with decreased volume resuscitation percentage (RBC: 50.7 ± 13.6%; Control: 39.3 ± 5.1%) compared to the Plasma and LR groups (81.1 ± 4.4% and 190.5 ± 10.6%, respectively). Animals given blood products also displayed significantly lower lactate levels compared to those given LR (Control: 2.3 ± 1.2 mM; Plasma: 4.8 ± 0.9 mM, RBC: 4.3 ± 1.3 mM, vs LR: 10.7 ± 3.5 mM, p=0.004 all groups compared to LR). Ending hemoglobin in RBC animals was significantly greater than in other groups (p&lt;0.001 compared to all groups). However, blood gas analysis revealed that the RBC group also had the lowest O2 saturation (88.3 ± 0.5% vs. LR: 93.9 ± 4.7%, Plasma: 94.8 ± 0.6%, WB: 93.5 ± 4.5%), yet PaO2 was comparable between groups.ConclusionIn our model of experimental T/HS, reperfusion fluids carrying red blood cells substantially improved hemodynamic stability by returning the mean arterial blood pressure to survivable values. The efficacy of RBC and whole blood resuscitation may be largely the result of improvements in diastolic blood pressure, suggesting increased contribution from systemic vascular resistance in these groups.Support or Funding InformationDOD W81XWH‐17‐2‐0047

5-Aminolevulinic Acid Improves Water Content and Reduces Skin Wrinkling

5-Aminolevulinic acid (ALA) is a natural amino acid that is used as a raw material for the biosynthesis of red-blood-cell hemoglobin in humans and animals. ALA is the subject of research in a wide range of human health care fields, including skin care and medicine. This study investigated whether and to what extent cosmetics containing ALA (skin lotions and creams) improved facial-skin condition in women. As such, a double-blind controlled experiment was conducted among 45 women aged 47.0 ± 1.72 years. Participants were divided into two intervention groups, including those who used cosmetics containing ALA (C-ALA, n = 22) and those who used cosmetics without ALA (W-ALA, n = 23). Specifically, participants were instructed to use their assigned cosmetics twice per day (morning and evening) after washing their faces before each application. Inner skin condition and skin-water content were evaluated via VISIA-Evolution and Multi Display Devices at baseline and after four and eight weeks of use. The experiment lasted for a total of eight weeks. A baseline comparison at week four showed that skin moisture content increased significantly for the C-ALA group (p = 0.021). Further, wrinkling significantly decreased at week four among participants in the C-ALA group who were evaluated as having many wrinkles at baseline (p = 0.034). These findings suggest that cosmetics containing ALA have moisturizing effects and reduce wrinkling caused by dryness.

The Management of Acute Blood Loss With Hemoglobin-Based Oxygen Carriers in Jehovah’s Witness Patients: A Case Series and Review of Literature

Abstract Managing anemia and acute blood loss in critical patients has long been reliant on the judicious use of red blood cell transfusions. The primary goal of blood transfusion is the efficient delivery of oxygen to end organs, especially the brain. Provided that appropriate guidelines are followed, allogenic blood transfusions are considered a safe and effective therapeutic option. Some patients refuse or decline blood transfusions due to cultural and/or religious beliefs, with the most well known being followers of Jehovah’s Witnesses (JWs). For this patient population, hemoglobin-based oxygen carriers (HBOCs) are a potential therapeutic option. Hemoglobin-based oxygen carriers are infusible oxygen-carrying fluids prepared from purified human or animal hemoglobin, which do not need to be refrigerated or crossmatched. We describe our experience at our institution where three JW patients were administered HBOCs. As HBOCs are not approved by the Food and Drug Administration and undergoing clinical trials, a compassionate use protocol was used to obtain HBOC-201 or Hemopure (Biopure Corporation). We explored factors attributing to overall response to treatment and laboratory effects of HBOC use. The first patient (24-year-old male, recurrent gastrointestinal bleed) received 8 units over a period of 2 weeks of HBOC. Pretransfusion hemoglobin and reticulocyte counts of 3.8 g/dL and 18.15% and posttransfusion counts were 12.8 g/dL and 16.97, respectively, with stable vital signs and renal function. Patient 2 (49-year-old female, chronic renal failure) received 8 units of Hemopure over 1 week. Before HBOC and erythropoietin administration, her hemoglobin and reticulocyte count were 4.2 g/dL and 0%, respectively, with stable vital signs and renal function. Post, her hemoglobin levels increased to 11.4 g/dL with a reticulocyte count of 16.0%. Unfortunately, the third patient (61-year-old female) expired secondary to complications from her initially diagnosed pulmonary embolism and massive stroke. She had received two units of HBOC and experienced a rise in hemoglobin (2.9 to 4.1 g/dL) and reticulocyte count (0% to 2.7%) with stable vitals during administration. Rapid resuscitation along with HBOC administration correlated with improved recovery in two of the three patients and yet none experienced hypertensive events. Younger age (24 and 49 years old) and their comorbid conditions are two factors that could explain the positive response to administration of HBOCs. Hemoglobin and hematocrit levels were often inaccurate as well as platelet counts, electrolytes, and liver enzymes. Coagulation tests (partial thromboplastin time, prothrombin time) were also affected by HBOC use. Overall, the utilization of HBOCs was beneficial in the hemodynamic management of two of our JW patients who refused allogenic blood transfusions. Additional research is needed to further understand the effectiveness of HBOCs on this patient population.

The high affinity of small-molecule antioxidants for hemoglobin

Hemoglobin has previously been shown to display ascorbate peroxidase and urate peroxidase activity, with measurable Michaelis-Menten parameters that reveal a particularly low Km for ascorbate as well as for urate – lower than the respective in vivo concentrations of these antioxidants in blood. Also, direct detection of a hemoglobin-ascorbate interaction was possible by monitoring the 1H-NMR spectrum of ascorbate in the presence of hemoglobin. The relative difference in structures between ascorbate and urate may raise the question as to exactly what the defining structural features would be, for a substrate that binds to hemoglobin with high affinity. Reported here are Michaelis-Menten parameters for hemoglobin acting as peroxidase against a number of other substrates of varying structures – gallate, caffeate, rutin, 3-hydroxyflavone, 3,6-dihydroxyflavone, quercetin, epicatechin, luteolin – all with high affinities (some higher than those of physiologically-relevant redox partners of Hb – ascorbate and urate). Moreover, this high affinity appears general to animal hemoglobins. 1H-NMR and 13C-NMR spectra reveal a general pattern wherein small hydrophilic antioxidants appear to all have their signals affected, presumably due to binding to hemoglobin. Fluorescence and calorimetry measurements confirm these conclusions. Docking calculations confirm the existence of binding sites on hemoglobin and on myoglobin for ascorbate as well as for other antioxidants. Support is found for involvement of Tyr42 in binding of three out of the four substrates investigated in the case of hemoglobin (including ascorbate and urate, as blood-contained relevant substrates), but also for Tyr145 (with urate and caffeate) and Tyr35 (with gallate).

Conservation of Binding of Llama and Other Animals’ Hemoglobins to Haptoglobins Across Species

The plasma protein haptoglobin binds hemoglobin released from lysed erythrocytes. It causes removal of the free hemoglobin, thus preventing pathological oxidation of cells. Hemoglobin higher level structures are well conserved across animal species but primary structures can be as little as 50% homologous. Because of these differences, the question arose as to what extent hemoglobins can bind to haptoglobins from different species. The charge properties and binding to three genetic variants of human haptoglobin were compared by non-denaturing agarose gel electrophoresis, with llama, human, dog, pig, horse and goat hemoglobins. In this study, it was reported for the first time that llama and alpaca hemoglobins differed in electrophoretic mobility from hemoglobins from several animal species and humans. Llama hemoglobin was more positively charged than the other mammalian hemoglobins. Electrophoretic mobility changes of the animal hemoglobins in the presence of human plasma and two different purified human haptoglobin genetic variants suggested that hemoglobins from all animals in this study could bind all three genetic variants of human haptoglobin. In all cases, the llama hemoglobin-haptoglobin samples had lesser mobility than those of the other mammals. This study showed that the binding sites on hemoglobin and haptoglobin for each other have been evolutionarily conserved despite differences in primary structure and marked difference in the charge of llama hemoglobin from the other animal species and humans.

Cross reaction and forensic comparison of blood testing done by private and public sector laboratories

Blood evidence consists of one of the most important forensic pieces. However, it is not always easy to identify a stain as blood. The best and fastest way of doing so, is by using rapid screening derived from immunochromatographic tests to spot hemoglobin. However, cross reaction between species might be an issue since hemoglobin of other animals could be present, especially considering crimes against fauna, outdoor crime scenes with an easy access for animals, and human and domestic animal trauma cases. According to the particular structure in each country there are private and/or public laboratories working in this field. Methods and resources from private and public sectors are different, and procedures also differ from country to country. The main objective of our work was to validate the presence of human blood using interspecific samples and the criteria defined by each lab using rapid blood detection test kits from the public sector related to forensic analysis in Brazil and from the private sector in Germany. As a secondary goal, we have evaluated sensibility, the Hook Effect and pH effect in some commercially available kits. Our results showed that the tests carried out are suitable for the detection of human blood in forensic samples using methods commonly employed in the public and private sectors.

Llama Hemoglobin Binding to Heterologous Haptoglobins

The plasma protein haptoglobin binds hemoglobin released from lysed erythrocytes. The complex is internalized through the macrophage membrane protein CD163 thus preventing pathological oxidation of biological molecules and tissues. Hemoglobin primary structures can be as little as 50% homologous among species. Haptoglobin also shows structural variation among species and a genetic polymorphism is found in humans and not in other mammals. The goal of this study was to examine the extent to which hemoglobins can bind to haptoglobins from different species. The llama is adapted to high altitude and its hemoglobin might thus differ from those of many other mammals. By using non‐denaturing agarose gel electrophoresis, the charge properties and binding to human haptoglobin was compared for llama, human, dog, pig, horse and goat hemoglobins.Llama hemoglobin was found to be more positively charged than the other mammalian hemoglobins. All hemoglobins tested showed a band of increased anodal mobility when incubated with human plasma which suggested formation of hemoglobin‐haptoglobin complexes. For one plasma sample there was only one new band whereas for a second sample two new anodal hemoglobin bands were formed. This suggested the ability of the different species hemoglobins to bind to different genetic polymorphisms of human haptoglobin. When the different hemoglobins were incubated with purified haptoglobin variants Hp 1:1 and Hp 2:2, one new anodal hemoglobin‐containing band was seen. The new band in the Hp 1:1 samples had higher mobility than those in the Hp 2:2 samples. In all cases the llama hemoglobin‐haptoglobin samples had lesser mobility than those of the other mammals. Because incubation with one of the plasma samples produced two new bands rather than one as with the Hp 1:1 and Hp2:2 it is surmised that this plasma sample had the Hp 2:1 variant and that all animal hemoglobins tested can bind the three genetic variants of human haptoglobin although this variation is regarded to be absent in non‐human species. It is concluded that the binding sites on hemoglobin and haptoglobin for each other have been evolutionarily conserved despite differences in primary structure and marked difference in the charge of llama hemoglobin.Support or Funding InformationThe research was supported by Furman University Research Experience for Undergraduates 2016 session, NSF grant NSF CHE‐1460806 – awarded to K. Buchmueller and T. Hanks at Furman University and by Albany State University, Center for Undergraduate Research

Clinical, hematologic, and biochemical findings in cattle infected with lumpy skin disease during an outbreak in southwest Iran.

This study was performed to determine the clinical, hematologic, and biochemical findings in animals affected with lumpy skin disease (LSD) in southwest Iran. Sixty cattle with LSD were included in this study and compared with 20 healthy ones as the control group. The disease was diagnosed based on clinical examination and confirmed by polymerase chain reaction analysis of the blood samples. The major observed clinical signs included skin nodules, fever, enlarged lymph nodes, and edema. In hematologic assessment, the average numbers of leukocytes, lymphocytes, eosinophils, erythrocytes, and platelets, as well as the average level of hemoglobin in the infected animals were significantly lower than in the control group. Biochemical experiments showed that the serum glucose, total and direct bilirubin, aspartate aminotransferase, and creatine phosphokinase activities in the infected group were significantly elevated. LSD also caused a significant reduction in the levels of serum creatinine, albumin, and iron. In total, LSD was associated with an overall decline in different blood cell types and significant changes in serum biochemical profile. These alterations could be related to the inflammatory disease processes and injuries in various organs, especially the liver. Hematologic and biochemical profiles can be utilized to better understand different aspects of LSD pathogenesis and ultimately improve its prognostic, management, and treatment methods.

Non-anemic Iron Deficiency from Birth to Weaning Does Not Impair Growth or Memory in Piglets.

Early iron deficiency is associated with impaired (cognitive) development, the severity of which depends on the timing and duration of the under-supply of iron. To design effective treatment and prevention strategies for iron deficiency in humans, suited animal models are needed. In an earlier study (Antonides et al., 2015b) we separated 10 pairs of piglets from their mothers within a few days after birth and reared one sibling with artificial iron-deficient (ID) and the other with balanced control milk until weaning. ID piglets grew slower and showed poorer reference memory (RM) performance than their controls in a spatial holeboard task, even weeks after iron repletion. One putative intervening factor in that study was pre-weaning maternal deprivation. In an attempt to refine the piglet iron-deficiency model, we assessed whether piglets reared by sows, but withheld iron supplementation, can serve as animal model of iron deficiency. As sow milk is inherently ID, piglets normally receive a prophylactic iron injection. Ten pairs of piglets were housed with foster sows until weaning (4 weeks). One sibling per pair was randomly assigned to the control group (receiving iron dextran injections: 40 mg iron per kilogram body mass on days 3 and 10), the other to the ID group. From weaning, all pigs were fed a balanced commercial diet. Blood samples were taken in week 1, 3.5, 6, and 12. Pre-weaning blood iron values of ID piglets were lower than those of controls, but recovered to normal values after weaning. Hemoglobin of ID piglets did not reach anemic values. Hematocrit and hemoglobin of ID animals did not decrease, and serum iron even increased pre-weaning, suggesting that the piglets had access to an external source of iron, e.g., spilled feed or feces of the foster sows. Growth, and spatial memory assessed in the holeboard from 10 to 16 weeks of age, was unaffected in ID pigs. We conclude that sow-raised piglets are not a suitable model for iron-deficiency induced cognitive deficits in humans. Based on our previous and the present study, we conclude that growth and memory are only impaired in piglets that suffered from pre-weaning anemia.

Catalase and Glutathione Peroxidase in Dogs Naturally Infected by Leishmania infantum

Background: Canine leishmaniasis (CanL) is caused by an obligatory intracellular parasite of Leishmania genus that affects organs and tissues. Several studies evaluate the role of reactive oxygen species (ROS) in the pathogenesis of many diseases. The overproduction of ROS on infectious diseases can induce an imbalance between oxidants and antioxidants at cellular or systemic level. Thus, the aim of this study was to evaluate the activity of antioxidant enzymes in CanL.Materials, Methods &amp; Results: Females (n = 17) and males (n = 10), at different ages and with different weight, were selected for this study. Dogs were divided into two groups according classical clinical signs and sorological test to CanL. Animals were considered infected based on indirect immunofluorescent assay and ELISA titration ≥ 1:40. Group B (n = 15) composed by positive dogs to CanL from Zoonosis Control Center of Fortaleza (Ceará, Brazil) and group A (n = 12) was composed by dogs from private kennel that were serologically negative to L. infantum and had absence of clinical signs to CanL. Blood sample were collected for evaluation of hematological and biochemical parameters and glutathione peroxidase (GPx) and catalase (CAT) enzymatic activity. Data were analyzed by Student’s t-test and Pearson correlation coefficient (P &lt; 0.05). Total proteins (TP, mg/dL) and alkaline phosphatase (ALP, U/L) were increased (P &lt; 0.05) on group B (8.2 ± 1.2; 165.4 ± 46.4) when compared to group A (6.5 ± 1.1; 109.1 ± 38.3), respectively. Hemoglobin (Hb; g/dL) and hematocrit (Hct; %) were decreased (P &lt; 0.05) on Group B (14.7 ± 1.8; 48.2 ± 5.7) when compared to group A (16.5 ± 1.3; 52.1 ± 2.4), respectively. Group B presented CAT (U/g Hb) and GPx (mU/mg Hb) lower (189.4 ± 90.4; 3,609.6 ± 1,569.1) than group A (326.6 ± 104.5; 5,055.6 ±1,569.1), respectively (P &lt; 0.001). Positive correlation was observed between RBC and CAT; however, it was not significant.Discussion: Organisms require a good defense system in order to revert the overproduction of free radicals and consequently the injuries caused by them. This is possible through the production of antioxidant agents, which act on oxidative prevention and on tissue and cellular regeneration, by taking the reduced glutathione (GSH), superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) enzymes in the frontline. Erythrocyte changes promoted by CanL suggest possible correlation between anemia and the appearance of clinical signs, which in many cases is not seen. Erythrocytes contain SOD, CAT and GPx enzymes, thus, changes in these cells will reflect on the activity of these enzymes. In our results only CAT showed positive correlation with erythrocyte count, however it was not significant. GPx activity was lower (P &lt; 0.001) in infected dogs than control group. This result agrees with another study, which showed a decrease in GPx levels in CanL, although it was not significant. However, it was found a positive correlation (P &lt; 0.001) between erythrocytes and GPx activity and between hemoglobin and GPx activity in animals with leishmaniasis. These results suggest that the reduction in detoxification activity can be related to the decrease in erythrocyte count and that the GPx activity depends on the control mechanism of the antioxidant system in CanL. Furthermore, this result could be associated with decrease of blood cell count in animals with CanL, once GPx is an erythrocyte enzyme, which plays an important role in hemoglobin protection against oxidative damage. This study was carried out in naturally infected dogs with L. infantum. In conclusion, CAT and GPx activities are relate to oxidative stress induced by L. infantum infection and can be used as biomarkers on CanL.

A Comparison of Animal Haemoglobins using High-Performance Liquid Chromatography (HPLC)

Automated cation-exchange HPLC is now considered the cornerstone diagnostic method in many laboratories for two reasons. Firstly, unlike other methodologies it detects variant haemoglobin’s and measures haemoglobin’s A, A 2 and F percentages in a single procedure. Secondly it is suitable for larger workloads in that it is now fully automated [1-5] . Haemoglobin is most generally characterised as consisting of two α and two non- α dimers that couple to form a tetramer in certain physiological conditions. This with its own haem group in a ‘pocket’ is then formed to make up a haemoglobin molecule. This study was undertaken in order to document these different animal haemoglobin’s using HPLC in samples obtained from animal species in both the Kruger National Park and the Pretoria Zoo, with the proviso of setting up a comprehensive atlas of these particular animal haemoglobin’s, which could be used in a clinical diagnostic setting [7, 9, 16, 18] . HPLC analysis was carried out on the following adult species: Gorilla, chimpanzee, chacma baboon, vervet monkey, lion, leopard, white rhino, giraffe, wild dog, buffalo, blue and black wildebeest, Nile crocodile, eland, pigmy hippopotamus and cheetah. The findings indicated that each species had its own unique chromatogram when compared to that of humans, with regards to: Retention times (i.e. the time from application to elution from the column), their characteristic percentage and in some cases, in the shape of the peak or curve produced [6-16] . Analysis of these animal haemoglobin’s produced chromatograms with three or more significant peaks (Fig 1). Mostly these peaks corresponded to the α -globin chain, the β -globin chain and the haem unit. As confirmed in previous studies [9, 16, 17] there were some species that had as many as four distinct chromatographic peaks (Fig 2). The retention times produced from the 17 animal species examined in this study can then serve as profiles, which could be used to compile a comprehensive atlas of their unique haemoglobin chromatograms. It could also be used to identify each species where each has its own unique chromatogram. This atlas, similar to that compiled for the human haemoglobin’s [28] could then be used in a clinical setting [16] where abnormalities in the HPLC retention times may be detected if the animal is in any way compromised.