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Introduction: The mini-implant, temporary anchorage devices (TADS), are now a common method of treatment in Orthodontics with versatility, minimal invasiveness and the relationship between costs and benefits that they offer even today. Skeletal anchorage has, to a large degree, replaced conventional anchorage in situations where anchorage is considered either critical, insufficient, or likely to result in undesirable side effects such as vertical displacements generated by inter-maxillary force systems., Objectives: The objective of this study is to carry out a review about the factors that seem affect the success or failure rate of orthodontic mini-implants. A computerized literature review was performed by searching the MEDLINE database (Entrez PubMed, www.ncbi.nlm.nih.gov), Google Scholar, Scopus, Cochrane Central Register of Controlled Trials, Isi Web of Knowledge until March 2016 . The main subject heading "orthodontics" was combined with these keywords mini-implant, mini-screw, micro-implants, mini-implant success rate, mini-implant failure rate, skeletal anchorage, temporary anchorage device (TADS). In the selection process, abstracts were initially read independently by two researchers to identify potentially eligible full text papers which were then retrieved and assessed in order to decide on the final inclusion., Conclusions: The clinical success of orthodontic anchorage by mini-implants depends on the stability of the miniscrews used for fixation. For good stability, the application site must provide bone of good quantity and quality. We can reasonably assume that the stability of the anchorage of the mini-implants could be optimized by selecting a position with particular characteristics of quality and quantity of bone, in relation to cortical and total mandibular and jaw bone thickness. These expected informations are important because they indicates that the bone quality and quantity are significant when considering an implant placement site, but also that there are other confounding factors influencing the success rate.

Objectives: The work has the objective to analyze the literature on the degradation of the adhesive interface. In particular the study is focused on the role of the metalloproteinase in the hydrolytic degradation of collagen matrix in the bonded interface. The survey will concern also the latest innovations to improve and increase the link between dentin and the restorative materials through the MMPs inhibitors., Methods: The research has been carried out in the MEDLINE database by choosing keywords as "metalloproteinases" and "dentin bond" and "degradation". In vitro studies were included in the research, excluding studies with no human and deciduous teeth. Language was limited to English., Results: The collagenolytic enzymes in mineralized dentin have been demonstrated to have an important role in dental hard tissue pathologies, including the degradation of the hybrid layer., Conclusion: The preservation of the collagen matrix integrity is a key issue in the attempts to improve the dentin bonding durability.

Unlabelled: The occlusal surface is the most affected area by dental caries and the sealing of the pits and of the fissures has been found, in time, the preventive method most effective trying to counteract the onset. Currently, the WHO considers it as a primary preventive measure, in other words one of the most effective and least invasive available to ensure the complete protection and the total preservation of the occlusal carious by the phenomenon., Purpose: THE AIM OF THIS WORK HAS BEEN TO PERFORM A SYSTEMATIC REVIEW OF THE LITERATURE ON CLINICAL TRIALS OF DIFFERENT SEALING MATERIALS, IN ORDER TO: compare their individual characteristics, highlight the reliability and the long-term efficacy and identify the most significant variables, both technological and clinics, in order to declare whether or not the success of this method prior., Materials and Methods: The research has been carried out in the MEDLINE database by choosing keywords as "sealants" and "follow up". Only studies published in the last thirteen years have been considered and have been evaluated only types of scientific articles that fall within the definition of Anglo-Saxon "Clinical Trial" and "Controlled Clinical Trial", excluding all experimental works in vitro, case-reports, meta-analyzes and literature reviews. Have been also considered only scientific papers on patients between the ages of 0 and 18 years., Results: Out of 29 studies, evaluating a total of 2900 individuals (aged between 2.5 and 17 years), 7411 seals made by using resin-based sealants (RB Sealants), modified glass ionomer sealants (RMGI) and compomer sealants have been analyzed. The best retention capacity of the material in time has been obtained from the use of RB Sealants compared to RMGI, demonstrating retention values much lower with partial loss of material at a distance of one year from the clinic. The compomers demonstrate retention values intermediates. The incidence of caries in a year is negligible for all sealants application., Conclusions: In terms of retention, resin-based sealants (RB Sealants) are the materials that give more guarantees of success at 12 months, while in the same period there haven't been significant differences in caries prevention of disease among the various classes sealing materials analyzed.

One of the most important factors implicated in preterm birth (PTB) is acute genitourinary tract infection. The bacteria causing chronic periodontal inflammation include Gram-negative rods and anaerobes similar to those found in women with bacterial vaginosis. The aim of this prospective study is to investigate the relationship between oral and vaginal microflora and preterm low birth weight. Real-time polymerase chain reaction was used to detect both the presence and level of six periodontitis-related species: Aggregatibacter actinomycetemcomitans (Aa), Porphyromonas gingivalis (Pg), Tannerella forsythia (Tf), Treponema denticola (Td), Fusobacterium nucleatum ssp(Fn), and Prevotella intermedia (Pi) for both oral samples of subgingival plaque and cervical samples, obtained from 80 patients, during gynaecological examinations. The more representative oral pathogen (less than 60 percent) species in oral samples of preterm and term group were Tf, Td, and Fn. 24.4 percent of pregnant women presented periodontal pathogens in vaginal swab; the most representative species with a percentage over 0.1 percent of total bacteria in genital tract of preterm group were Tf, Td, and Piwith a positive correlation (less than 0.5). The presence of the bacterium T. denticolain the vagina, regardless of the amount, adversely affects preterm delivery.

Objective: Elastomeric materials play an important role in the orthodontic practice, including the retraction force to move teeth into extraction sites, closing diastemas, selective shifting of the midline and generalized space closure. Frictional resistance and ligating strength of archwire-bracket-ligature complex occurs during utilization of elastomeric and metallic ligatures when orthodontic forces are applicated. The aim of this study was to analyze elastic deformation of three types of elastomeric ligatures, after clinical use., Material and Methods: ELASTOMERIC LIGATURES: ring-shape, transparent, latex ligatures (Leone® S.p.A.), ring-shape, grey, polyurethane ligatures (Micerium® S.p.A.) and grey, polyurethane, Slide low-friction ligatures (Leone® S.p.A.). A total of 9 orthodontic patients undergoing fixed orthodontic therapy were selected. Three specimens were applied, one for each types of ligature, inside the oral cavity of each subject. Samples were kept in the oral cavity for 28 days, ligating 0.16 X 0.22 inches stainless steel archwires to stainless steel premolars brackets (Leone® S.p.A., Sesto Fiorentino, FI, Italy) for Bidimensional technique. After the pre-established time, the systems of ligature were removed and washed. Control group consisted of 9 unused specimens of each ligation type. Each elastomeric ligature was observed under the scanning electron microscope (SEM) to determine variations in size. The archwire-bracket-ligature complex was also analyzed., Results: Transparent O-ring ligatures showed significant volumetric and structural changes. The external rounded shape was rather maintained, while the internal shape tended to appear square. Both external and internal diameter significantly increased (p<0.005 and p<0.0001 respectively) while the thickness decreased ( p<0,005) when analyzed with t-test. Polyurethane ring-shape ligatures retained the initial ring design. Both external and internal diameter increased (p<0.0001), while the thickness remained almost unchanged. The internal border was more squared, and showed jagged edges with continuous and irregular extroversions. Grey, polyurethane Slide low-friction ligatures showed a reduced dimensional change. There was a slight increase in two dimensions, length and width, (14-16%) (p<0.05 and p<0.001) while there was a not significant decrease in thickness (10%)., Conclusion: From SEM analysis of ligature morphology it emerges that latex and polyurethane O-ring ligatures endure significant volumetric and structural changes, after clinical use, index of a greater degree of friction and early loss in functionality. Grey, polyurethane Slide low-friction ligatures presented limited variation in size after clinical use.

Unlabelled: Fixed orthodontic appliances make it difficult to maintain the oral hygiene, resulting in plaque accumulation. Retention of bacterial plaque, represents a risk for white spot lesions and development of periodontal disease., Aim: Purpose of this study was to determine in vivo the retention of plaque on three different elastic ligatures, in comparison with stainless steel ligature, to determine a possible association between type of ligatures and accumulation of microorganisms., Material and Methods: THREE ELASTIC LIGATION SYSTEMS WERE ANALYZED FOR PLAQUE RETENTION: ring-shape, clear, latex ligatures (Leone® Spa), ring-shape, grey, polyurethane ligatures (Micerium® Spa) and grey, polyurethane, Slide low-friction ligatures (Leone® Spa), compared with stainless steel ligatures (Leone® Spa) used as control. Forthy orthodontic patients undergoing fixed orthodontic therapy were selected. A sample for each type of ligature were applied inside the oral cavity of each subject. Samples were kept in the oral cavity for 28 days, ligating 0.16 X 0.22 stainless steel archwire to stainless steel orthodontic premolars brackets. The presence of bacterical slime was quantified by spectrophotometric method (crystal violet-Bouin's fixative) and morphological observations was evaluated by Scanning Electron Microscopy (SEM)., Results: From analysis of bacterical slime emerges that all the elastics showed a low plaque retention, especially if compared to the group of steinless steel ligatures, that presented a greater plaque adhesion, statistically significant compared to the Slide group (r<0.0002) and the two elastic groups (r<0.0001). This study reported no significant difference between the Slide ligatures and the traditional elastic ligatures as regards the retention of plaque. SEM images showed presence of cocci, rods and few filamentous organisms and an interbacterial matrix in all observed samples., Conclusion: Elastomeric ligatures showed a significant lower susceptibility to plaque adhesion, in comparison to the stainless steel of the metallic ligatures. No statistically significant difference was observed among the elastic devices.

Blocks of two porous synthetic hydroxyapatites (HA) with porosity fraction of 30-40 and 50-60 vol%, respectively and a coralline derived porous HA were evaluated in vitro in presence of the osteogenic line MC3T3-E1 and of L929 fibroblasts. The two tested biomaterials did not affect cellular proliferation (MTT test), but the contact inhibited alkaline phosphatase activity. Porous aggregates resulted perfectly biocompatible in the tests performed, since observations performed by light microscopy did not show any cell morphological change, osteoblast presented a stellar shape and typical pseudopodes. SEM observations showed intercellular matrix containing fibers on HA-based porous aggregates.

We describe a simple method by which thin sections ( approximately 100 microm) from modern and archaeological teeth and bones can be obtained. A detailed embedding-cutting-mounting procedure is proposed, suggesting the use of a dental adhesive system, composite resins and conventional embedding resins, with the aims of improving the quality of the sections and substantially reducing the steps and time needed to prepare specimens for histological analysis. The introduction of this dental materials-based system allows an accurate positioning of the sample embedded inside the resin, prevents cracks and distortions of the section during the cutting phase and generally improves mounting sections on slides.

Background: In the oral cavity, many bacteria can only survive by adhering to hard surfaces. The roughness and free energy of these surfaces play an important part in this process. Precision dental alloys may undergo corrosion, but findings show that this does not seem to cause problems of biocompatibility. The release of metallic ions into the oral cavity may both inhibit bacterial growth and influence bacterial adhesion. The object of the present study was to bring to light any possible correlation between corrosion and/or ionic release and bacterial adhesion with regard to 18 different types of dental alloy, both before and after polishing., Methods: Electrochemical analyses were carried out (cyclic potentiodynamic and potentiostatic polarisation tests). Corrodible elements were analysed through Atomic Absorption Spectroscopy of specimens of each alloy. The inhibition of bacterial adhesion and growth was determined using bacteria specific to the oral cavity., Results: All the alloys examined show a tendency towards spontaneous passivation with low values of anodic current. The evaluation of ionic release confirmed the biocompatibility of the tested materials and the solutions conditioned with the alloys did not inhibit bacterial growth. There was no significant bacterial adhesion after polishing. Bacteria adhere to unpolished alloys in a specific manner and are inhibited from doing so in the presence of alloys for gold-resin restorations containing silver and copper., Conclusions: When polished, all the alloys are resistant to in vitro electrochemical corrosion and bacterial adhesion. The possibility cannot be excluded that bacterial adhesion occurs after the materials have been in place in the oral cavity for some time. The alloys which were found to inhibit bacterial attack may be more suitable, while not representing a biological risk for the surrounding tissues.

This article presents a review of the effects of sea salt on gingival tissues. The beneficial effects of sea salt are described.

A large number of methods are now available for the preclinical screening of implantable materials concerning their biocompatibility and their ability to stimulate tissue formation. In vitro techniques represent a very useful tool, since this way we can realistically simulate the biological events which occur in vivo at the bone-implant interface. In the present study scanning electron microscopy and light microscopy observations were performed in order to assess the effect of an hydroxyapatite granulate on cell behaviour and morphology. Uptake of proteins to hydroxyapatite surface has been also investigated by comparing the amounts adsorbed after incubation with bovine serum albumin and bovine pancreaticamilase. According to our preliminary observation cells do not show signs of toxicity or inhibition of cell growth even after 14 days of co-culture with hydroxyapatite. Granules were covered by an uninterrupted cell layer by day seven. Even after two days micrographs show cells anchored and spread over the surface of the underlying granules, with a flattened and stellate shape. Such a morphology indicates a very high cellular activity, suggesting that the interaction with hydroxyapatite seriously increased metabolism. Measurements of protein adsorption on the hydroxyapatite surface show that changes in the size of particles affect the binding of proteins, while, in the case of granular hydroxyapatite, despite changes in size of granules, variations of protein adsorption were not observed, neither in relation to their different isoelectric point. Our preliminary results represent a good example of the opportunities presented by an experimental in vitro model.

The functional properties of fetal bovine hemoglobin have been studied as a function of temperature, chloride and 2,3-diphosphoglycerate (DPG) concentration. The fetal bovine erythrocyte has six times the concentration of the allosteric modulator DPG compared with the adult cell, and yet the oxygen affinity of the fetal hemoglobin still exceeds that of the adult molecule at the respective physiological concentration of DPG and at physiological temperature. We find that the allosteric modulator strongly affects the enthalpy of oxygen for the fetal hemoglobin but not for the adult protein. We propose that this may be an important mechanism for the exchange of heat from mother to fetus. In particular, under stripped conditions the oxygen affinity of fetal bovine Hb is considerably higher than that of the adult hemoglobin. Due to the higher DPG concentration that characterizes fetal bovine erythrocytes this difference is almost abolished in the presence of the respective physiological concentration of DPG and at 20 degrees C. However, on going from 20 degrees C to 37 degrees C, the difference in O2 affinity between the two hemoglobins is restored, as it should if oxygen has to be transferred from maternal to fetal blood, by virtue of the lower overall heat of oxygenation (delta H) displayed by fetal Hb when in the presence of DPG at physiological concentration. This behavior is reminiscent of that of human fetal Hb and outlines the role of temperature and of its interplay with heterotropic ligands in the modulation of hemoglobin function to fully meet the physiological needs of the organism.

Considering the variety of species that depend on hemoglobin for oxygen transport, these molecules must execute their primary function under extreme environmental conditions. Hence, a thermodynamic analysis of oxygen binding with hemoglobins from different species reveals a series of adaptive mechanisms which are based on the thermodynamic connection between the binding of heterotropic effectors and the reaction with oxygen. The examples reported, from fishes to human fetus, illustrate how evolution can alter the structural basis of the heterotropic interactions to optimize the oxygenation-deoxygenation cycle in dependence of the physiological needs of the particular organisms. Moreover they show that a thermodynamic analysis of the reaction with oxygen overcomes the meaning of a detailed structural and functional characterization going deeper into the physiology of the specific organism.

The functional properties of the single haemoglobin (Hb) of Emperor penguin (Aptenodytes forsteri) have been investigated at different temperatures as a function of proton and organic phosphate concentration. The complete amino acid sequence has been established. Comparison with that of human HbA shows 12 substitutions in the contact regions of alpha beta dimers. In addition to overall similarities shared with most of the avian Hbs previously described, this Hb shows significant differences, which could be related to the peculiar behaviour of this penguin. In particular we may consider that: (1) the shape of the Bohr effect curve seems well adapted for gas exchange during very prolonged dives, preserving penguin Hb from a sudden and not controlled stripping of oxygen; (2) the very minor enthalpy change observed at lower pH could be an example of molecular adaptation, through which oxygen delivery becomes essentially insensitive to exposure to the extremely low temperatures of the environment. Moreover, the small alkaline Bohr effect has been found to be only chloride-linked, since the pH dependence of the oxygen affinity is totally abolished in the absence of this ion. These functional characteristics are discussed on the basis of the primary structure of alpha and beta-chains.

The functional and spectroscopic properties of chamois (Rupicapra rupicapra) and steinbock (Capra hircus ibex) haemoglobin (Hb) have been studied with special reference to the action of allosteric effectors and temperature. Moreover, the amino acid sequences of the N-terminal segments of the alpha- and beta-chains have been determined. The present results indicate that chamois and steinbock Hbs display a low affinity for O2, which appears to be modulated in vivo by Cl- ions rather than 2,3-bisphosphoglycerate. The Bohr effect for O2 binding to chamois and steinbock Hb is higher than for reindeer and bovine Hbs, being similar to that of human Hb. Moreover, the temperature-dependence of oxygenation appears intermediate between that of human and reindeer Hbs. E.p.r. and absorption spectroscopic properties of the ferrous nitrosylated derivative of chamois and steinbock Hbs suggest that both haemoproteins are in a low-affinity conformation even in the absence of InsP6. The reduced effect of polyphosphates on the functional and spectroscopic properties of chamois and steinbock Hb agree with amino acid differences in the N-terminal segment of the beta-chains (i.e. the deletion of Val(NA1) and the replacement of His(NA2), present in human Hb, and Gln(NA2), present in horse Hb, by Met). The molecular mechanism modulating the basic reaction of O2 with chamois and steinbock Hb may be linked to specific physiological needs related to the high-altitude habitats of these two animals.

Human fetal hemoglobin is known to display, at 20 degrees C, a lower affinity than human adult hemoglobin for oxygen when both proteins are in the absence of organic phosphates. The physiologically important reverse situation is achieved at 37 degrees C upon addition of 2,3-bisphosphoglycerate (DPG), whose lower effect on fetal hemoglobin is related to some amino acid substitutions present in gamma-chains. However, the difference in oxygen affinity observed at 37 degrees C is not solely due to the different modulation power of DPG with respect to adult and fetal hemoglobins. In fact, the results presented here reveal new aspects linked to the interplay of temperature and organic phosphates. In particular, the lower effect of DPG on fetal hemoglobin renders almost identical the oxygen affinity of the two hemoglobins at 20 degrees C, abolishing the difference observed in the absence of the effector. Successively on going from 20 degrees C to 37 degrees C, by virtue of the lower overall heat of oxygenation (delta H) displayed by fetal hemoglobin when in the presence of DPG, adult hemoglobin shows a lower oxygen affinity, as it should if oxygen has to be transferred from maternal to fetal blood.

Morphologic and metabolic erythrocyte modifications are thought to be the basis of cell removal from circulating blood. A significant role has been ascribed to the immunological network which may remove aged or misshapen erythrocytes through the binding of specific autoantibodies. Along this line recent observations indicate that a senescence antigen appears in consequence of postsynthetic modifications of band 3, one of the most important erythrocyte membrane proteins, which accounts for many functional activities of the red cells. On this basis, we raised a mouse hybridoma anti-band 3 monoclonal antibody (B6 MoAb) of the IgG2a class which monitors band 3 differences among normal red blood cells separated by Percoll density gradient. These differences are outlined by the decrease of B6 MoAb binding to band 3 monomer, the appearance of an 80-90 kDa new band, lighter than band 3, and the increase of low molecular weight fragments in the 4.5 region. The B6 MoAb appears to be very useful in detecting modifications of band 3 since it bind to a 19 kDa Chy-Try fragment estimated to be sensitive to aging.

It has been generally assumed that mammals have blood with a greater temperature sensitivity than ectothermic organisms. Recent results have shown that in some species of mammals, Hb displays a value of overall oxygenation enthalpy (delta H) much less exothermic than that observed for most mammalian hemoglobins, including human adult Hb. In this respect, a very interesting case is represented by porcine blood which shows a modest effect of temperature, the temperature coefficient of its oxygen-dissociation curve being significantly lower than that of human blood. Here we report a detailed functional characterization of pig Hb, which, interpreted on the basis of the amino acid sequence of the alpha and beta chains of the molecule, sheds some light on the molecular basis of the phenomenon.

The effect of temperature on the oxygen-binding properties of hemoglobin (Hb) from ruminants, such as ox, reindeer, musk ox, mouflon and egyptian water buffalo is compared to that of human adult Hb (HbA). A striking difference emerges where in the presence of chloride ions and in the absence of 2,3-diphosphoglycerate [Gri(2,3)P2] a strongly reduced exothermic oxygenation process is observed for all ruminant Hb investigated with respect to HbA. Next, in the presence of physiological concentrations of Gri(2,3)P2, HbA displays a less exothermic oxygenation process, with values tending toward those observed in ruminant Hb [where Gri(2,3)P2 is not a physiological effector and for which the addition of Gri(2,3)P2 has essentially no effect on the oxygenation enthalpy]. Different from HbA, the intrinsically less exothermic oxygen binding seems to be independent of the experimental conditions for ruminant Hb, underlying specific structural characteristics which might be responsible for this feature.

The ligand-binding properties of hemoglobins from two homozygote phenotypes (AA and BB) of water buffalo (Bubalus bubalis) have been characterized by equilibrium and kinetic techniques. In the case of the BB phenotype, the two constituent hemoglobins have been purified and separately analysed. Buffalo hemoglobins display the reduced sensitivity to organic phosphates characteristic of ruminant hemoglobins, their physiological effector probably being the chloride ion. In contrast to the other known hemoglobins from ruminants, all the hemoglobins from the water buffalo display a significant temperature sensitivity, the delta H for oxygen binding in the presence of physiological effectors approaching that of human hemoglobin (delta H = -30.5 kJ/mol O2). This discrepancy with the other ruminant hemoglobins (e.g. ox, delta H = -10.4 kJ/mol O2), whose primary structure is very similar to that of buffalo, hemoglobins might be correlated to the different habitat and phylogenetic history of the two subfamilies (Bos and Bubalus) of Bovidae.

The equilibrium O2-binding properties of the hybrid haemoglobin (Hb) present in vivo in erythrocytes from mule and of its parent Hbs from horse and donkey were compared with special reference to the effect of heterotropic ligands such as Cl-, D-glycerate 2,3-bisphosphate (DPG) and inositol hexakisphosphate. All these Hbs display a decreased effect by polyphosphates, confirming that what has been observed for horse Hb [Giardina, Brix, Clementi, Scatena, Nicoletti, Cicchetti, Argentin & Condò (1990) Biochem. J. 266, 897-900] is common to other equine species, at least from a qualitative standpoint. However, different quantitative aspects can be detected, which can be accounted for by a different role for the two types of chain in characterizing the binding free energy for the various heterotropic effectors. In particular, it is shown that the binding mode of DPG and inositol hexakisphosphate displays different features since long-range effects can be observed clearly for inositol hexakisphosphate but not for DPG. In general terms, in spite of a different intrinsic O2 affinity, the modulation of functional properties by third ligands leads these Hbs to behave, under physiological conditions, similarly to human HbA. It might represent an interesting example of how different species with similar functional needs find different ways to produce a similar functional behaviour.

An extensive set of data relating to the binding of oxygen by haemocyanin from the squid Todarodes sagittatus has been collected under various experimental conditions. The results obtained show that, within the range of physiological pH, the concentration of protons affects mainly the high-affinity state of the molecule without significantly affecting the low-affinity state. As far as the effect of temperature is concerned, the data show a characteristic feature which is very similar to that previously described in the case of haemoglobins from Arctic mammals such as reindeer (Rangifer tarandus) and musk ox. (Ovibos moschatus). The shape of the oxygen equilibrium curve shows strong temperature-dependence, since the overall heat of the binding of oxygen to the low-affinity state of the molecule is strongly exothermic and that to the high-affinity state is very close to zero. The results provide an outline of the intramolecular compromise that, through the interplay of temperature and protons, optimizes the loading and unloading of oxygen under the various environmental conditions experienced by this species of squid.

The spectroscopic (EPR and absorbance) properties of the nitric oxide derivative of ferrous man, horse, buffalo, deer, mouflon, musk ox, ox, and reindeer hemoglobin (HbNO) have been investigated in the absence of any allosteric effector at pH 6.5 (in 0.1 M 2-[N-morpholino]ethanesulphonic acid/NaOH chloride-free buffer system), as well as at 100 K and/or 20 degrees C. Man and horse HbNO show spectroscopic properties that are generally taken as typical of the high affinity state of ferrous tetrameric Hb's; on the other hand, the spectroscopic properties of ruminant (i.e., buffalo, deer, mouflon, musk ox, ox, and reindeer) HbNO are characteristic of the low affinity conformation. These results are in keeping with the functional properties of the mammalian Hb's considered and have been related to the peculiar low oxygen affinity of ruminant Hb's.

Evolution has adopted different strategies to solve the problem of transporting oxygen to respiring tissues, according to needs dictated by the environment. A thermodynamic analysis of haemoglobins of organisms living in extreme polar environments (mammals and fish) provides elegant examples of such adaptations.

Haemocyanin from the lobster Palinurus elephas has been shown to change in its subunit composition according to the time of year. In contrast, in Palinurus mauritanicus, a lobster living at greater depth, no seasonal changes in subunit composition have been observed. The results obtained from a set of experiments performed on some Palinurus mauritanicus acclimatized in an aquarium have clearly indicated that modifications of haemocyanin subunit composition may be involved in the adaptation of arthropods to environmental change.

The functional properties of hemoglobin from Egyptian water buffalo have been characterized as a function of pH, temperature and chloride concentration. Alongside overall similarities shared with ox and Arctic ruminant hemoglobins, hemoglobin from buffalo shows significant differences with respect to the effect of temperature. The results obtained may suggest that the limited effect of temperature on oxygen binding recently reported for ox hemoglobin could be regarded as an interesting case of a reminiscence of a past glacial age.

The primary structures of alpha- and beta-chains of hemoglobin from reindeer (Rangifer tarandus tarandus) were determined. Comparison of the reindeer hemoglobin sequence with those of human and bovine hemoglobins showed 50 and 29 substitutions per alpha beta dimer, respectively. The influence of replacements on the modulation of hemoglobin oxygen affinity by heterothopic ligands and temperature, as well as their importance on the structure-function relationships in hemoglobin are discussed.

Compounds with low molecular weight (haptens) are not usually immunogenic. Thus, an hapten should be conjugated with a carrier protein therefore immunizing an animal, in order to induce a strong immune response and in turn obtain large amount of antibodies. Suitable macromolecules are usually albumins, thyroglobulins, haemocyanins and polylysine. The production of specific antibodies against haptens is important both for assays of hormones and drugs in biological fluids or for therapeutic applications in tumour therapy. Approaches in choice of both carrier protein and conjugation methods will be described.

The functional properties of haemoglobin from the Lesser Rorqual whale (Balaenoptera acutorostrata) have been characterized as a function of the heterotropic effector concentrations and temperature. The results obtained suggest the existence of sophisticated modulation mechanisms based on the interplay of organic phosphates, carbon dioxide, lactate and temperature. These, together with the very small apparent heat of oxygenation (delta H) of oxygen binding, have been physiologically interpreted on the basis of the specific metabolic needs of this diving mammal.

The effect of inositol hexakisphosphate (IHP) on the spectroscopic (EPR and absorbance) properties of the nitric oxide derivative of ferrous horse and bovine hemoglobin (Hb) has been investigated. In the absence of IHP, the nitric oxide derivative of ferrous horse Hb shows spectroscopic properties similar to those of the corresponding derivative of ferrous human Hb that are generally taken as typical of the high affinity state of tetrametric hemoproteins. Similar to human Hb, the addition of IHP to the nitric oxide derivative of ferrous horse Hb induces a transition toward a species characterized by spectral properties typical of the low affinity state of hemoglobins. Nevertheless, the equilibrium constant for IHP binding to the nitric oxide derivative of ferrous horse Hb (= 1.5 x 10(2) M-1) is much lower than that reported for the association of the polyphosphate to the same derivative of ferrous human Hb (greater than 3 x 10(5) M-1). Conversely, the spectroscopic properties of the nitric oxide derivative of ferrous bovine Hb are characteristic of the low affinity state of tetrameric hemoproteins, both in the absence and in the presence of IHP. These results, taken together with the behavior of the nitric oxide derivative of ferrous human Hb, provide further evidence for the peculiar oxygen binding properties of horse and bovine Hb.

The most surprising characteristic of reindeer hemoglobin (Hb) concerns its response to changes in temperature. Thus, the shape of the oxygen-binding curve is strongly temperature dependent due to the difference in the enthalpy of oxygenation between the T and R state of the molecule. In fact, delta H of oxygen binding to the T state is strongly exothermic whereas that of the R state is very close to zero or possibly positive after correction for the heat of oxygen solubilization. Moreover, the allosteric transition T0----R0 has been found to display a negative delta H and a contemporaneous decrease in entropy, a behavior which is precisely the opposite of what has been reported for other hemoglobins. As a whole, reindeer Hb represents a beautiful example of the significance that comparative studies may have in assessing the general validity of the main properties of the hemoglobin molecule.

The functional properties of dromedary hemoglobin have been studied as a function of chloride, polyphosphates and pH and compared with those of human hemoglobin. The two proteins have the same amino acid residues at the anion-binding sites as well as at the level of the groups responsible for the alkaline Bohr effect. Analysis of the experimental data reveals that: (a) intrinsic oxygen affinity and the Bohr effect are very similar for the two proteins; (b) the association equilibrium constants of chloride are substantially higher in the dromedary system, both in the unligated and ligated state; (c) two polyanion-binding sites occur in dromedary oxy and deoxyhemoglobin; (d) association constants of polyphosphates for the higher-affinity binding site (probably in the cavity between beta chains) are comparable for the two proteins under physiological conditions; association constants for the second binding site in dromedary hemoglobin are not affected by pH changes; (e) the dependence of oxygen affinity in dromedary hemoglobin upon chloride concentration is complex, this anion at relatively low concentrations reverses the effect of millimolar polyphosphate; (f) both in stopped-flow and flash photolysis experiments the kinetic behaviour of dromedary hemoglobin is consistent with the equilibrium results. The pronounced sensitivity to solvent composition of the functional properties of dromedary hemoglobin even in the oxy state stresses the potential relevance of this conformation for regulating the oxygen transport in vivo.

1. The oxygen binding properties of the hemoglobin from the Lesser Rorqual, Balaenoptera acutorostrata, has been investigated with respect to the possible effects of organic phosphates on gas transport in arctic environments. 2. The intrinsic oxygen affinity of the hemoglobin is high and strongly modulated by the effects of organic phosphates. 3. In the absence of organic phosphates, the temperature sensitivity of oxygen binding expressed by the heat of oxygenation, delta H, is -16.2 kcal/mol when corrected for the heat of oxygen in solution. 4. In the presence of organic phosphates there is a marked decrease in the temperature sensitivity delta H approximately -5 kcal/mol). 5. This feature is of great importance for oxygen unloading in the flippers and the tail, where the temperature is lower than the trunk of the whale. 6. Furthermore the organic phosphates strongly increase the Bohr coefficient, delta log P50/delta pH, from less than -0.3 in stripped hemoglobin to about -1.5 when the hemoglobin is saturated with P6-inositol. 7. This feature may be of great physiological importance by reducing the CO2 tension and acidosis after a prolonged dive.

Homogeneous components of Wistar rat hemoglobin have been isolated and characterized from the molecular and functional point of view. The O2 equilibrium behaviour of the three main components (HbII, HbIII, HbIVA) has been investigated as a function of pH and organic phosphate concentration. The ligand-binding kinetics of the isolated components have been also studied and are fully consistent with their equilibrium behaviour. It should be remarked that the choice of the system was governed largely by the ability of rat hemoglobins to crystallize very quickly. This almost unique molecular property together with the complete reversibility of the process may allow information to be obtained on the thermodynamics of ligand-linked phase changes.

Some divalent cations (at 10(-3)M concentration) have been shown to act as oxygen-linked ligands. The effect of Ca++ (only on hemoglobin S) and Zn++ (on normal hemoglobin and hemoglobin S) can be traced to a preferential binding of these cations to the oxy- conformation of the proteins; the reverse is true for Ni++, since its presence reduces the oxygen affinity of both hemoglobins. The results suggest: i) the existence of at least two different cation binding sites on hemoglobin; ii) the replacement of glutamic acid by valine in hemoglobins S introduces sufficient structural modification to form a new cation binding site.

The present paper reports data for the effect of pH and D-glycerate 2,3-bisphosphate (D-glycerate-2,3-P2) on the oxygen equilibrium of normal and SH(beta 93)-modified human hemoglobin. At sufficiently high D-glycerate-2,3-P2 concentrations, both oxy and deoxy forms of HbA are saturated with the organic phosphate at all pH values between 6 and 9. Furthermore the difference in the affinity for D-glycerate-2,3-P2 between deoxy and oxy hemoglobin remains constant with pH, implying that the pK values of the Bohr effect groups in deoxy and oxyhemoglobin are not affected by the presence of D-glycerate-2,3-P2 on the hemoglobin molecule. In the hemoglobins modified in position beta 93, the difference in affinity between deoxy and oxy hemoglobin for D-glycerate-2,3-P2 decreases with decrease in pH due mainly to a decrease in the affinity of the deoxy form for D-glycerate-2,3-P2. The effect of the chemical modification appears to be primarily a change in pK of a Bohr group in deoxy hemoglobin which is especially pronounced in the presence of phosphates.

Previous results [(1988) Arct. Med. Res. 47, 83-88] have shown that hemoglobin from reindeer is characterized by a low overall heat of oxygenation. This particular aspect has been investigated further in a series of precise oxygen equilibrium experiments. The results obtained show a peculiar dependence of the temperature effect on the fractional saturation of hemoglobin with oxygen, which could be regarded as a very interesting case of molecular adaptation to extreme environmental conditions.

Perutz & Brunori (1982) proposed that the COOH-terminal His and Ser F9 of the beta-chains of fish and amphibian hemoglobins are responsible for their Root effect and part of their alkaline Bohr effect. Analysis of the kinetics of carbon monoxide binding by hemoglobin from the tadpole of Xenopus laevis supports that model and suggests an explanation for the absence of an alkaline Bohr effect in many aquatic Anura and Urodela.