Functionally distinct haemoglobins of the cryopelagic Antarctic teleost Pagothenia borchgrevinki

Pagothenia borchgrevinki , has a higher haemoglobin concentration than other Antarctic notothenioids and the high oxygen capacity may correlate with the relatively active mode of life of this fish. The fish has five haemoglobins (Hb C, Hb 0, Hb 1, Hb 2 and Hb 3) with Hb 1 accounting for 70–80% of the total, and Hb C being present in trace amounts. Hb 1 and Hb 2 are functionally similar in terms of Bohr and Root effects. Hb 3 has a weaker Bohr effect than Hb 1 and Hb 2, and the Root effect is similar to that of Hb 1. Hb 0 has a strong Bohr effect and the Root effect is enhanced to a larger extent by the physiological effectors chlorides and phosphates than that of the other components with the exception of Hb C. The heats of oxygenation are lower than those of temperate fish haemoglobins. Temperature variations may have a different effect on the functional properties of each haemoglobin, and chloride and phosphates may play an important role in the conformational change between the oxy and deoxy structures. The complete amino acid sequences of Hb 1 and Hb 0, as well as partial N-terminal or internal sequences of the other haemoglobins, have been established. The high multiplicity of functionally distinct haemoglobins indicates that P. borchgrevinki , has a specialized haemoglobin system.     

Changes in the nucleotide triphosphate/haemoglobin and nucleotide triphosphate/red cell ratios of rainbow trout, Salmo gairdneri Richardson, subjected to prolonged starvation and bleeding

The nucleotide triphosphate/haemoglobin (NTP/Hb) and nucleotide triphosphate/red cell (NTP/cell) ratios of rainbow trout increased during prolonged starvation. A decline was noted in blood lactic acid concentration. Red cell count, haemoglobin concentration and haematocrit also declined. Changes in mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH) and mean corpuscular haemoglobin concentration (MCHC) were found not to be significant. The NTP/Hb and NTP/cell ratios of both fed and starved trout rose seven days following a 15% reduction in blood volume by cardiac puncture. A rise in whole blood NTP concentration was found only in the bleeding response of fed animals. No significant change was noted in blood lactic acid concentration. The decline in haematocrit was significant only in the starved group. In both groups, however, red cell count and blood haemoglobin concentration fell. MCV rose whereas MCHC declined in all bled animals. Changes in MCH were not significant in either group. Negative correlations were noted between red cell count and both the NTP/Hb and NTP/cell ratios and between haemoglobin concentration and the NTP/Hb ratio. Positive correlations were seen between the two ratios and between red cell count and haemoglobin concentration.     

Alpha-thalassemia in Puglia. II. Neonatal screening for Bart's hemoglobin

550 blood specimens from the umbilical cord of newborn babies from Apulia have been screened in order to detect Hb Bart's. The electrophoresis of the haemoglobin by Cellogel (Tris Glycine pH 8.6) carried out on the 550 specimens have revealed in 43 variable quantities of Hb Bart's: 34 cases (6,18%) showed non measurable quota, whereas in the other 9 cases (1,63%) Hb Bart's varied from 2,19% to 26%. The haemoglobin biosynthesis "in vitro" of the baby presenting 26% of Hb Bart's has been reported.     

Macromolecular isoforms of Daphnia magna haemoglobin

The haemoglobin (Hb) of Daphnia magna acclimated to different oxygen conditions was sampled, and in its natively assembled state it was separated by chromatofocusing. The Hb isoforms were analysed for their subunit composition under denaturating conditions by two-dimensional gel electrophoresis. The Hb system is suggested to consist of three predominant Hb aggregates, which are characterised by a specific subunit composition and synthesised in response to different ambient oxygen conditions. In normoxia, a dominant Hb aggregate (DmHbI) with a pI of 4.4-4.6 was composed of subunits B, C, E, F and G. In severe hypoxia, a different dominant Hb isoform (DmHbIII) with a pI of 5.7-5.9 was composed of subunits A, B, C, D, E and F. Further analyses in moderate hypoxia provided evidence for a third Hb isoform (DmHbII) composed of subunits B, C, D, E and F. Sequence alignment and homology modelling of the tertiary structure of the D. magna Hb domains 1 and 2 revealed functionally relevant substitutions of amino acid residues at positions B10, E7 and E11, which determine the functional properties of D. magna haemoglobin in terms of haem contact, oxygen binding and affinity. Both domains are predicted to possess the common haemoglobin fold, but helices C and D are not properly formed, and helix G is interrupted by a short coil.     

Familial Polycythaemia Caused by a New Haemoglobin Variant: Hb Heathrow, β 103 (G5) Phenylalanine Leucine

A new variant of haemoglobin A (Hb A) with a high affinity for oxygen has been found in an English family. Five members are affected and all are polycythaemic. This variant (Hb Heathrow) is the first of this class to be found in this country and has the same electrophoretic mobility as Hb A. It was discovered only by measuring the oxygen affinity of the patients'' red cells. This emphasizes the need for measuring the oxygen affinity of haemoglobin in patients with polycythaemia if other clinical and haematological features associated with polycythaemia rubra vera are absent.     

Frequency of hemoglobin abnormalities and hemoglobinopathies in Mozambique peoples (comparison between leprosy and non-leprosy subjects]

In 588 bloodsamples of negride natives from Mo?ambique, preferably Chuabo and Macua, haemoglobin analyses were performed. In 21 cases an increase of Hb A2 was found, indicating the presence of heterozygous beta-thalassaemia, in one case the changes in Hb-analysis were typical for beta-delta-thalassaemia, 18 samples could be shown to contain Hb S, typical for the heterozygous sickle cell trait. Futhermore in 7 cases Hb A2' was found. In two bloodsamples haemoglobin variants were observed, which according to their electrophoretical mobility were assumed to represent Hb D in one case, and Hb G in the other. In the Chuabo population the frequency of the thalassaemia gene was found to be more than twice as high as in the Macua population. In non-lepers Hb S was observed with a remarkable higher incidence than in lepers.     

The haemoglobin system of the mudfish, Labeo capensis: adaptations to temperature and hypoxia

The effect of temperature and hypoxic acclimation on the haemoglobin system and intraerythrocytic organic phosphate concentrations in the South African mudfish, Labeo capensis, have been investigated. Exposure to hypoxia or increased temperature raised haemoglobin concentration and decreased NTP/Hb ratio. Temperature acclimation did not effect the oxygenation characteristics of the haemolysate or haemoglobin multiplicity, as evident from isoelectric focussing experiments that showed one cathodic (Hb I) and three anodic haemoglobins (Hb II, III and IV). Oxygen equilibria of the isolated components showed a smaller Bohr effect and lower temperature and organic phosphate sensitivities in the cathodic than in the anodic haemoglobins. Unlike the trout and eel haemoglobin systems, both the anodic and cathodic haemoglobins from L. capensis exhibited sensitivity to organic phosphates but the effect was smaller in the latter. The results indicate that oxygen transport in mudfish blood is supported by variations in the red cell organic phosphate\haemoglobin ratio and the functional differentiation between anodic and cathodic haemoglobins.     

Difference between the Hb C variants in Brahman and in indigenous Southern African cattle breeds

Haemoglobin polymorphism in Brahman cattle and seven Southern African cattle breeds was investigated by means of starch gel electrophoresis. A difference was found between the migration rates of the Hb C of Brahman cattle and that of the indigenous Southern African cattle breeds, showing that these are actually two separate variants. We suggest that the faster migrating type, which occurs in Brahman cattle, should be called Hb C, while the slower migrating type of the Southern African breeds, should be called Hb I.
A comparison of the migration of the α and β chains of all the variants we encountered, including foetal haemoglobin, was carried out by means of starch gel electrophoresis in urea at acid and alkaline pH levels. Evidence was found of a difference in mobility of the non-a chains of haemoglobin A, B, C, I and F at different pH levels while no difference was detected in the migration rate of their respective α chains. These results confirm the theory that genetic variation is restricted to the non-α chain of bovine haemoglobin.
Progeny studies on 225 families confirm that the Hb I variant is allelic to Hb A and Hb B variants. The gene frequencies have been calculated on the basis that Hb I and Hb C are allelic at the β locus. The distribution of the different phenotypes is in accordance with this theory, assuming that the populations obey Hardy-Weinberg equilibrium.     

Two new sheep haemoglobins, one of which is replaced by haemoglobin C in anaemia

Two new haemoglobin variants, provisionally named Hb G and Hb H, were found during a survey of 295 Welsh Mountain cross-bred sheep. Both haemoglobins appear to be beta chain variants controlled by genes allelic to those for the common forms Hb A and Hb B. Studies on an anaemic Hb AH and an Hb AG type sheep showed that Hb G, like Hb A, is replaced by Hb C in anaemia whereas Hb H, like Hb B, is not replaced.     

Physical, biochemical and functional characterization of haemoglobin from three strains of Artemia

The brine shrimp, Artemia, an inhabitant of coastal and inland salterns, encounter fluctuations in the salinity which in turn influences the oxygen availability of their habitat. Hence, experiments were performed to analyze variations in haemoglobin structure and patterns of three strains of Artemia from South India and also to reflect the effect of varying oxygen levels in their habitat. Haemoglobins were purified on a DEAE-Sephadex column and haemoglobin types were analyzed by comparing their relative mobility on a non-denaturing medium. Furthermore, their molecular masses were determined by gel filtration in Sepharose column and by dodecylsulfate polyacrylamide gel electrophoresis. Results clearly reveal the presence of three distinct extracellular haemoglobins Hb I, Hb II and Hb III in Tuticorin strain while the other strains displayed only trails or the complete absence of Hb III and Hb II. Estimated molecular masses of these haemoglobins are 235,000-250,000 Da. Denaturation of the reduced and alkylated haemoglobins revealed apparently one polypeptide chain with a molecular mass of 124,000 Da. Upon denaturing gel electrophoresis of native haemoglobin Hb II, it was found that the 124,000 Da, polypeptide was cleaved specifically into two unequally-sized fragments of 50,400 and 79,800 Da. With regard to oxygen affinity, Hb III has a very high affinity for oxygen, an almost negligible Bohr effect and a good physiological adaptation to temperature changes. By combining the three haemoglobins in different proportions Artemia strains must be able to withstand diverging environmental conditions. In particular, the absence of Hb III in Puthalam and its occurrence as a faint band in Thamaraikulam could be correlated to the oxygen levels of their habitats.     

Glucose-6-Phosphate Dehydrogenase Deficiency and Haemoglobin Drop after Sulphadoxine-Pyrimethamine Use for Intermittent Preventive Treatment of Malaria during Pregnancy in Ghana – A Cohort Study

Background

Sulphadoxine-Pyrimethamine (SP) is still the only recommended antimalarial for use in intermittent preventive treatment of malaria during pregnancy (IPTp) in some malaria endemic countries including Ghana. SP has the potential to cause acute haemolysis in G6PD deficient people resulting in significant haemoglobin (Hb) drop but there is limited data on post SP-IPTp Hb drop. This study determined the difference, if any in proportions of women with significant acute haemoglobin drop between G6PD normal, partial deficient and full deficient women after SP-IPTp.

Methods and Findings

Prospectively, 1518 pregnant women who received SP for IPTp as part of their normal antenatal care were enrolled. Their G6PD status were determined at enrollment followed by assessments on days 3, 7,14 and 28 to document any adverse effects and changes in post-IPTp haemoglobin (Hb) levels. The three groups were comparable at baseline except for their mean Hb (10.3 g/dL for G6PD normal, 10.8 g/dL for G6PD partial deficient and 10.8 g/dL for G6PD full defect women).The prevalence of G6PD full defect was 2.3% and 17.0% for G6PD partial defect. There was no difference in the proportions with fractional Hb drop ≥ 20% as compared to their baseline value post SP-IPTp among the 3 groups on days 3, 7, 14. The G6PD full defect group had the highest median fractional drop at day 7. There was a weak negative correlation between G6PD activity and fractional Hb drop. There was no statistical difference between the three groups in the proportions of those who started the study with Hb ≥ 8g/dl whose Hb level subsequently fell below 8g/dl post-SP IPTp. No study participant required transfusion or hospitalization for severe anaemia.

Conclusions

There was no significant difference between G6PD normal and deficient women in proportions with significant acute haemoglobin drop post SP-IPTp and lower G6PD enzyme activity was not strongly associated with significant acute drug-induced haemoglobin drop post SP-IPTp but a larger study is required to confirm consistency of findings.     

Effects of confinement stress and additional zinc treatment on some blood parameters in the dogfish Scyliorhinus canicula

Several blood parameters were studied in the dogfish Scyliorhinus canicula subjected to different confinement conditions (transport and maintenance) and zinc contamination. Confinement stress significantly decreased erythrocyte counts (RBCC), haematocrit (Ht), haemoglobin (Hb), leucocrit (Lt), mean corpuscular haemoglobin (MCH) and mean corpuscular haemoglobin concentration (MCHC). Mean corpuscular volume (MCV) did not change with this stress and plasma glucose concentration significantly increased. In fish subjected to confinement stress conditions, Zn treatment significantly decreased Hb, MCH and MCHC and plasma glucose concentration. Ht and MCV did not change and Lt and RBCC significantly increased. Results are discussed related to other stress effects and effects of zinc treatment on fish confined in non-stressful conditions.     

The influence of haemoglobin types on reproduction and production in Hungarian Merino sheep

Fewer Hb A and AB lambs were born than expected in the investigated Hungarian Merino population but the factors responsible for these differences could not be identified. The Hb A and AB breeding ewes were not inferior to their B type herd mates with respect to average daily weight gain (when lambs), reproductive performance (prolificacy) and survival rate. Their reproductive performance was, in some cases, even better than that of the ewes with other haemoglobin types. The haemoglobin types cannot be used as markers when selecting for higher prolificacy.     

Haematological studies on the prewintering and wintering frog, Rana esculenta

1. A study of the haematology of the frog Rana esculenta including erythrocyte count (RBC), haemoglobin content (Hb), haematocrit (HCT), mean cell volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC) and erythrocyte size as a function of prewinter and winter was made. 2. The RBC count and Hb were significantly higher in contrast to MCV and MCH values during prewinter in both sexes. 3. The surface area to volume ratio was higher in prewinter whereas the length to width ratio (eccentricity) of the cytosome and nucleus was significantly higher during winter in both sexes. 4. Sexual differences in the erythrocyte count, Hb content and the surface area to volume ratio were also observed. 5. The physiological significance of these observations are reported for Rana esculenta.     

Postnatal changes in 2,3-diphosphoglycerate (2,3-DPG) content of calf erythrocytes

In 82 calves, 22 adult cows and 10 fetuses the 2,3-DPG level was determined in the erythrocytes. The lowest level was found in cows (1.13 microM/g haemoglobin, on the average), and in calves aged 35 days or more. In the foetuses the mean 2,3-DPG concentration in the erythrocytes was 4.98 microM/g Hb and it was higher that that in the erythrocytes of cows, the oldest foetuses and calves during the first two days of postnatal life. During 5 weeks of postnatal life the changes taking place in 2,3-DPG concentration could be divided into two periods: period I or the period of increase covering the first 6 days of life, with a characteristic rise in the concentration of this component from 1.37 to 15.80 microM/g Hb, and period II or the period of decrease lasting from the 6th to the 35th day of life. In period II two phases could be discerned, the first phase lasting from the 6th to the 10th day with a steep fall of 2,3-DPG level from 15.80 to 4.58 microM/g Hb, and the second phase from the 10th to the 35th day of life in which the level of 2,3-DPG reached slowly the value found in adult cows. A comparison of oxygen affinity of haemoglobin in calves aged 6 days, which was composed of about 80% of fetal haemoglobin and about 20% of adult haemoglobin, and in adult cows, which contained exclusively adult haemoglobin, showed that the oxygen-binding capacity of haemoglobin was lower in calves.     

Surprising differences in the respiratory protein of insects: A spectroscopic study of haemoglobin from the European honeybee and the malaria mosquito

Only recently it was discovered that haemoglobin (Hb) belongs to the standard gene repertoire of insects, although their tracheal system is used for respiration. A classical oxygen-carrying function of Hb is only obvious for hexapods living in hypoxic environments. In other insect species, including the common fruit fly Drosophila melanogaster, the physiological role of Hb is yet unclear. Here, we study recombinant haemoglobin from the European honeybee Apis mellifera (Ame) and the malaria mosquito Anopheles gambiae (Aga). Spectroscopic evidence shows that both proteins can be classified as hexacoordinate Hbs with a strong affinity for the distal histidine. AgaHb1 is proposed to play a role in oxygen transport or sensing based on its multimeric state, slow autoxidation, and small but significant amount of five-coordinated haem in the deoxy ferrous form. AmeHb appears to behave more like vertebrate neuroglobin with a complex function given its diversified distribution in the genome.     

Haematological parameters in a neotropical fish, Corydoras paleatus (Jenyns, 1842) (Pisces, Callichthyidae), captured from pristine and polluted water

We report normal ranges of haematological indices in healthy Corydoras paleatus from an unpolluted area. Haematological parameters studied include: erythrocyte counts (Er), haematocrit (Ht), haemoglobin concentration (Hb), mean cell volume (MCV), mean cell haemoglobin (MCH) and mean cell haemoglobin concentration (MCHC). Normal red blood parameters did not change according to maturation stages, sex or seasons. Then, we compared them with those coming from fish captured in a site polluted by sewage. Fish exposed to pollution presented significantly higher values of Er, Ht, Hb, MCH and MCHC than those captured in an unpolluted area. Discriminant analysis showed that Hb is a key parameter to point out differences between populations exposed to different environmental conditions. We suggest that haematological values of C. paleatus, registered during this study, could be used as biomarkers in future works evaluating the incidence of environmental stress on fish as well as pointing out changes in the water quality.     

Haemoglobin of the antarctic fish Pagothenia bernacchii. Amino acid sequence, oxygen equilibria and crystal structure of its carbonmonoxy derivative.

The Antarctic fish Pagothenia bernacchii has one major haemoglobin, Hb1 (over 95% of the total blood content). Hb1 has a strong alkaline Bohr effect and at low pH exhibits the reduced ligand affinity and co-operativity that comprise the Root effect. We have determined the complete amino acid sequence of P. bernacchii Hb1 and also the structure of its carbonmonoxy derivative by X-ray crystallography, to a resolution of 2.5 A. The crystallographic R-factor of the refined structure is 18%. The three-dimensional structure of this fish haemoglobin is similar to that of human haemoglobin A, with a root-mean-square difference in main-chain atom positions of 1.4 A after superimposition of the two structures, despite only 48% homology of their amino acid sequences (including insertion of a single residue in the CD region of the fish alpha-chain). Large structural differences occur only at the N and C termini of both the alpha- and beta-chains. Neither these nor other smaller structural differences provide any obvious explanation of the Root effect of this or other fish haemoglobins.     

Inositol tripyrophosphate: a new membrane permeant allosteric effector of haemoglobin

Nine inositol tripyrophosphate (ITPP) salts have been synthesized. Their ability to act as allosteric effectors of haemoglobin (Hb) has been measured in vitro with free Hb and whole blood. All the synthesized compounds bound to free Hb and were also able to cross, to a certain extent, the plasma membrane of the red blood cells (RBCs) in whole blood samples, lowering the affinity of Hb for oxygen. The oxy-haemoglobin dissociation curves were significantly shifted towards higher values of oxygen partial pressures, both for free Hb and for intracellular Hb in whole blood.