Immunocastration as an alternative to caponization: evaluation of its effect on body and bone development and on meat color and composition.

Caponization is associated with some morbidity and mortality, which contributes to important economic losses. This practice is executed without any pain relief (neither anesthesia nor analgesia) and can be painful and without consideration of animal welfare. On the other hand, immunocastration accomplished by Improvac and Bovipriva in pigs and cattle represents a noninvasive procedure, and for that reason is regarded as an alternative with improved animal welfare. This study includes 4 experimental groups consisting of capons, slips, roosters, and birds submitted to the Improvac treatment. The administration of Improvac was associated with a considerable reduction in serum testosterone concentration (reduced by 79% compared to average serum testosterone of roosters). Regarding significant differences among experimental groups, birds from the Improvac group were intermediate between capons and slips with respect to abdominal fat pad weight and yield, breast meat water and protein contents, and femur length. Conversely, color parameters such as lightness, redness, and hue angle for Improvac birds were intermediate between roosters and capons. Thus, immunocastration with Improvac could represent an alternative solution to caponization, with considerable improvements in animal welfare.

Iodine intake in Portuguese pregnant women: results of a countrywide study.

BACKGROUND: Iodine is the key element for thyroid hormone synthesis, and its deficiency, even moderate, is harmful in pregnancy, when needs are increased, because of its potential deleterious effects on fetal brain development. In Portugal, no recent data on iodine intake exists. The objective of this countrywide study was to analyze iodine status in pregnant Portuguese women in order to propose adequate measures to the health authorities. SUBJECTS AND METHODS: Using a fast colorimetric method, urine iodine concentration (UIC) was evaluated in 3631 pregnant women followed in 17 maternity hospitals from hinterland and coastal areas in Continental Portugal and the Portuguese islands of Açores and Madeira. RESULTS: Median UIC value was 84.9 µg/l (range 67.6-124.1) in Continental Portugal, 69.5 µg/l in Madeira, and 50.0 µg/l in Açores. The percentage of satisfactory values (>150 µg/l) was 16.8, ranging from 8.8 to 34.1 in the Continent, and being 8.2 in Madeira and 2.3 in Açores. The percentage of values below 50 µg/l was 23.7, ranging from 14.0 to 37.4 in the Continent, 33.7 in Madeira, and 50.0 in Açores. CONCLUSIONS: Our results point to an inadequate iodine intake in pregnant women assisted in most Portuguese maternity hospitals. Considering the potential deleterious effects of inadequate iodine supply in pregnancy, iodine supplementation is strongly recommended in this period of life.

[Diagnosis and therapeutic management of an intraorbital organic foreign body]. / Diagnostic et prise en charge des corps étrangers végétaux intra-orbitaires.

INTRODUCTION: Orbital organic foreign bodies are rare and can present different clinical features. The objective of this report is to show the danger of this type of foreign body, present the imaging data, and suggest a diagnostic approach and therapeutic management. We describe three cases of orbital organic foreign bodies with three different clinical presentations. CASE REPORTS: The first case was a 43-year-old male complaining of a chronic cutaneous fistula of the inferior right eyelid lasting 4 months after an orbital trauma with a wooden object. Two surgeries were necessary to extract the foreign bodies. In the second case, a 37-year-old female with post-traumatic ptosis after a bicycle accident several months before, the imaging exams revealed a fracture of the left orbital ceiling and a superior extraconical foreign body that was removed by a neurosurgery approach. The third case, a 69-year-old male with a right orbit abscess following a trauma with a tree branch had a persistent right orbit inflammation lasting 4 months despite two drainage surgeries and an extraction of an orbital organic foreign body. A third surgery was necessary to completely extract the foreign bodies. DISCUSSION/CONCLUSION: Detecting an orbital organic foreign body is sometimes difficult, especially when the clinical history is unclear, the ophthalmologic exam reveals no abnormalities, or if the patient is referred to the hospital several months after the traumatic event. Orbital organic foreign bodies can lead to potentially serious orbital or intracranial complications. Computed tomography and orbital ultrasound sometimes do not yield an evocative pattern: for instance, in computed tomography, the low density of wood can be misdiagnosed as air. On the other hand, these foreign bodies can persist in the orbit after several surgical explorations because they can easily break or migrate. Magnetic resonance imaging is useful when an orbital organic foreign body is suspected, but the analysis is easier when clinical data are suggestive.

[Contact lens fitting after keratoplasty for keratoconus]. / Adaptation des lentilles de contact sur greffe de cornée pour kératocône.

PURPOSE: The objective of this study was to determine the epidemiological and clinical characteristics of contact lens fitting after corneal graft for keratoconus. Contact lens types, visual acuity and complications were examined. METHODS: A retrospective medical chart review from patients admitted to Pellegrin Hospital at CHU Bordeaux, between January 2001 and December 2006, for contact lens fitting after keratoplasty for keratoconus. RESULTS: This retrospective study involved 39 eyes of 29 patients with an average age of 34.3 years and an average follow-up of 31.3 months. The mean time between grafting and contact lens fitting was 50.9 months. Large-diameter rigid gas permeable contact lenses were the most frequent lenses used. Average visual acuity after contact lens fitting was 8/10. Intolerance to contact lenses was observed in 8 eyes (20.5%) eyes and the complications associated with their use responded well to medical treatment. CONCLUSION: Patients with keratoconus must be alerted to the possible necessity of fitting contact lenses after corneal graft. They can play an important role in the improvement of visual function without significant complications.

Self-perceptions about having an ostomy: a postoperative analysis.

The aim of this study is to identify the self-perceptions of patients at different postoperative phases and settings. The case study involves two ostomy patients: a young woman with a temporary ileostomy, and an older man with a permanent colostomy. Both patients were interviewed while still in the hospital on the third, fifth, and seventh postoperative days, and on the second day of the first and third months after discharge. Four major factors were identified after the patients were analyzed: social support, health and life expectations, physical suffering, and self-care. Family support predominated the social support category for both patients. The results showed that the patients' perceptions about having an ostomy were not influenced by the type of ostomy (i.e., whether the ostomy was temporary or permanent in the in-hospital phase). However, these results/perceptions, primarily those related to health and life expectancy and self-care, changed early in the study. We believe it is important for the healthcare team to know the perceptions patients have about their ostomies and themselves postoperatively. This, in turn, may contribute to systematic and personalized care based on specific patients' demands that vary at different stages of treatment and, most certainly, in different cultures.

Hyperprolactinemia due to big big prolactin is differently detected by commercially available immunoassays.

Macroprolactinemia, i.e. sustained hyperprolactinemia where the predominant circulating form of prolactin (PRL) is of large molecular weight, is a common phenomenon comprising up to one-fourth of all cases of hyperprolactinemia. We measured serum PRL levels by four different immunoassay systems (PROL-CTK, RIAgnost, Delfia, ACS 180) and by the Nb2 bioassay in patients with prolactinomas/idiopathic hyperprolactinemias in whom monomeric PRL was the major species of PRL (n=11, group 1) and in patients with macroprolactinemia (n=12, group 2). In group 1, the results obtained with the different immunoassays and with the Nb2 assay were highly correlated (r=0.945-0.982). On the other hand, big big-PRL (bb-PRL) was differently recognized by the immunoassays, since measured serum PRL values from each patient were highly variable in group 2. RIA-gnost Prolactin and Delfia Prolactin detected bb-PRL similarly and they were highly correlated with each other (r=0.937, p<0.0001). ACS 180 detected bb-PRL somewhat differently from the RIA-gnost and Delfia systems, but likewise most of the patients of group 2 had PRL values above normal. PROL-CTK was the method less influenced by the presence of bb-PRL since most of the subjects with macroprolactinemia had PRL levels either within the normal range or only marginally elevated. From the immunoassays tested, PROL-CTK was the system which was less correlated with the Nb2 bioassay in group 2 (r=0.252; NS). Our experience is that macroprolactinemia is an asymptomatic condition in most of the cases. Therefore, we suggest that the routine measurement of PRL should be done with methods that are only minimally affected by the presence of macroprolactin. Such an approach would obviate the use of extensive, frequently expensive and ultimately useless diagnostic tests that are needed to determine the cause of the hyperprolactinemia.

The structure of an electron transfer complex containing a cytochrome c and a peroxidase.

Efficient biological electron transfer may require a fluid association of redox partners. Two noncrystallographic methods (a new molecular docking program and 1H NMR spectroscopy) have been used to study the electron transfer complex formed between the cytochrome c peroxidase (CCP) of Paracoccus denitrificans and cytochromes c. For the natural redox partner, cytochrome c550, the results are consistent with a complex in which the heme of a single cytochrome lies above the exposed electron-transferring heme of the peroxidase. In contrast, two molecules of the nonphysiological but kinetically competent horse cytochrome bind between the two hemes of the peroxidase. These dramatically different patterns are consistent with a redox active surface on the peroxidase that may accommodate more than one cytochrome and allow lateral mobility.

The surface-charge asymmetry and dimerisation of cytochrome c550 from Paracoccus denitrificans--implications for the interaction with cytochrome c peroxidase.

The implications of the dimeric state of cytochrome c550 for its binding to Paracoccus cytochrome c peroxidase and its delivery of the two electrons required to restore the active enzyme during catalysis have been investigated. The amino acid sequence of cytochrome c550 of Paracoccus denitrificans strain LMD 52.44 was determined and showed 21 differences from that of strain LMD 22.21. Based on the X-ray structure of the latter, a structure for the cytochrome c550 monomer from strain 52.44 is proposed and a dipole moment of 945 debye was calculated with an orientation close to the exposed haem edge. The behaviour of the cytochrome on molecular-exclusion chromatography is indicative of an ionic strength-dependent monomer (15 kDa)/dimer (30 kDa) equilibrium that can also be detected by 1H-NMR spectroscopy. The apparent mass of 50 kDa observed at very low ionic strength was consistent with the presence of a strongly asymmetric dimer. This was confirmed by cross-linking studies, which showed that a cross-linked species of mass 30 kDa on SDS behaved with an apparent mass of 50 kDa on molecular-exclusion chromatography. A programme which carried out and evaluated molecular docking of two monomers to give a dimer generated a most probable dimer in which the monomer dipoles lay almost antiparallel to each other. The resultant dipole moment of the dimer is therefore small. Although this finding calls into question the possibility of preorientation of a strongly asymmetrically charged cytochrome as it collides with a redox partner, the stoichiometry of complex formation with cytochrome c peroxidase as studied by 1H-NMR spectroscopy shows that it is the monomer that binds.

A single histidine is required for activity of cytochrome c peroxidase from Paracoccus denitrificans.

The diheme cytochrome c peroxidase from Paracoccus denitrificans was modified with the histidine-specific reagent diethyl pyrocarbonate. At low excess of reagent, 1 mol of histidine was modified in the oxidized enzyme, and modification was associated with loss of the ability to form the active state. With time, the modification reversed, and the ability to form the active state was recovered. The agreement between the spectrophotometric measurement of histidine modification and radioactive incorporation using a radiolabeled reagent indicated little modification of other amino acids. However, the reversal of histidine modification observed spectrophotometrically was not matched by loss of radioactivity, and we propose a slow transfer of the ethoxyformyl group to an unidentified amino acid. The presence of CN- bound to the active peroxidatic site of the enzyme led to complete protection of the essential histidine from modification. Limited subtilisin treatment of the native enzyme followed by tryptic digest of the C-terminal fragment (residues 251-338) showed that radioactivity was located in a peptide containing a single histidine at position 275. We propose that this conserved residue, in a highly conserved region, is central to the function of the active mixed-valence state.

The affinity and specificity of Ca(2+)-binding sites of cytochrome-c peroxidase from Paracoccus denitrificans.

The binding of Ca2+ to the dihaem cytochrome-c peroxidase from Paracoccus denitrificans was analysed by following perturbations in the visible and 1H-NMR spectra of both haem groups. The enzyme contains at least two types of Ca(2+)-binding site. Site I is occupied in the isolated enzyme, binds Ca2+ with a redox-state-independent Kd of 1.2 microM and accommodates neither Mg2+ nor Mn2+. Site II is unoccupied in dilute solutions of the isolated oxidised enzyme and binds Ca2+ cooperatively with a Kd of 0.52 mM. In the mixed valence form, the binding affinity increases to resemble that of site I. The cooperativity was shown by -Ca2+ binding to site II, the titration of haem methyl 1H-NMR resonances, and a half-of-sites effect observed for modification of an essential histidine with diethylpyrocarbonate. These are all consistent with site II being situated at the interface between two monomers of a dimeric enzyme. Thus the equilibrium of binding to site II is a reflection of the equilibrium for dimerisation and conditions which shift that equilibrium towards the dimer, such as increased ionic strength or high protein concentration, also increase Ca2+ affinity. Binding of Ca2+ to site II is required for formation of the active high spin state at the peroxidatic haem.

Mössbauer characterization of Paracoccus denitrificans cytochrome c peroxidase. Further evidence for redox and calcium binding-induced heme-heme interaction.

Mössbauer and electron paramagnetic resonance (EPR) spectroscopies were used to characterize the diheme cytochrome c peroxidase from Paracoccus denitrificans (L.M.D. 52.44). The spectra of the oxidized enzyme show two distinct spectral components characteristic of low spin ferric hemes (S = 1/2), revealing different heme environments for the two heme groups. The Paracoccus peroxidase can be non-physiologically reduced by ascorbate. Mössbauer investigation of the ascorbate-reduced peroxidase shows that only one heme (the high potential heme) is reduced and that the reduced heme is diamagnetic (S = 0). The other heme (the low potential heme) remains oxidized, indicating that the enzyme is in a mixed valence, half-reduced state. The EPR spectrum of the half-reduced peroxidase, however, shows two low spin ferric species with gmax = 2.89 (species I) and gmax = 2.78 (species II). This EPR observation, together with the Mössbauer result, suggests that both species are arising from the low potential heme. More interestingly, the spectroscopic properties of these two species are distinct from that of the low potential heme in the oxidized enzyme, providing evidence for heme-heme interaction induced by the reduction of the high potential heme. Addition of calcium ions to the half-reduced enzyme converts species II to species I. Since calcium has been found to promote peroxidase activity, species I may represent the active form of the peroxidatic heme.

The kinetics of the oxidation of cytochrome c by Paracoccus cytochrome c peroxidase.

In work that is complementary to our investigation of the spectroscopic features of the cytochrome c peroxidase from Paracoccus denitrificans [Gilmour, Goodhew, Pettigrew, Prazeres, Moura and Moura (1993) Biochem. J. 294, 745-752], we have studied the kinetics of oxidation of cytochrome c by this enzyme. The enzyme, as isolated, is in the fully oxidized form and is relatively inactive. Reduction of the high-potential haem at pH 6 with ascorbate results in partial activation of the enzyme. Full activation is achieved by addition of 1 mM CaCl2. Enzyme activation is associated with formation of a high-spin state at the oxidized low-potential haem. EGTA treatment of the oxidized enzyme prevents activation after reduction with ascorbate, while treatment with EGTA of the reduced, partially activated, form abolishes the activity. We conclude that the active enzyme is a mixed-valence form with the low-potential haem in a high-spin state that is stabilized by Ca2+. Dilution of the enzyme results in a progressive loss of activity, the extent of which depends on the degree of dilution. Most of the activity lost upon dilution can be recovered after reconcentration. The M(r) of the enzyme on molecular-exclusion chromatography is concentration-dependent, with a shift to lower values at lower concentrations. Values of M(r) obtained are intermediate between those of a monomer (39,565) and a dimer. We propose that the active form of the enzyme is a dimer which dissociates at high dilution to give inactive monomers. From the activity of the enzyme at different dilutions, a KD of 0.8 microM can be calculated for the monomerdimer equilibrium. The cytochrome c peroxidase oxidizes horse ferrocytochrome c with first-order kinetics, even at high ferrocytochrome c concentrations. The maximal catalytic-centre activity ('turnover number') under the assay conditions used is 62,000 min-1, with a half-saturating ferrocytochrome c concentration of 3.3 microM. The corresponding values for the Paracoccus cytochrome c-550 (presumed to be the physiological substrate) are 85,000 min-1 and 13 microM. However, in this case, the kinetics deviate from first-order progress curves at all ferrocytochrome c concentrations. Consideration of the periplasmic environment in Paracoccus denitrificans leads us to propose that the enzyme will be present as the fully active dimer supplied with saturating ferrocytochrome c-550.

Spectroscopic characterization of cytochrome c peroxidase from Paracoccus denitrificans.

The cytochrome c peroxidase of Paracoccus denitrificans is similar to the well-studied enzyme from Pseudomonas aeruginosa. Like the Pseudomonas enzyme, the Paracoccus peroxidase contains two haem c groups, one high potential and one low potential. The high-potential haem acts as a source of the second electron for H2O2 reduction, and the low-potential haem acts as a peroxidatic centre. Reduction with ascorbate of the high-potential haem of the Paracoccus enzyme results in a switch of the low-potential haem to a high-spin state, as shown by visible and n.m.r. spectroscopy. This high-spin haem of the mixed-valence enzyme is accessible to ligands and binds CN- with a KD of 5 microM. The Paracoccus enzyme is significantly different from that from Pseudomonas in the time course of high-spin formation after reduction of the high-potential haem, and in the requirement for bivalent cations. Reduction with 1 mM ascorbate at pH 6 is complete within 2 min, and this is followed by a slow appearance of the high-spin state with a half-time of 10 min. Thus the process of reduction and spin state change can be easily separated in time and the intermediate form obtained. This separation is also evident in e.p.r. spectra, although the slow change involves an alteration in the low-spin ligation at this temperature rather than a change in spin state. The separation is even more striking at pH 7.5, where no high-spin form is obtained until 1 mM Ca2+ is added to the mixed-valence enzyme. The spin-state switch of the low-potential haem shifts the midpoint redox potential of the high-potential haem by 50 mV, a further indication of haem-haem interaction.