Translational Research for Orthopedic Bone Graft Development.

Designing biomaterials for bone-substitute applications is still a challenge regarding the natural complex structure of hard tissues. Aiming at bone regeneration applications, scaffolds based on natural collagen and synthetic nanohydroxyapatite were developed, and they showed adequate mechanical and biological properties. The objective of this work was to perform and evaluate a scaled-up production process of this porous biocomposite scaffold, which promotes bone regeneration and works as a barrier for both fibrosis and the proliferation of scar tissue. The material was produced using a prototype bioreactor at an industrial scale, instead of laboratory production at the bench, in order to produce an appropriate medical device for the orthopedic market. Prototypes were produced in porous membranes that were e-beam irradiated (the sterilization process) and then analysed by scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM), dynamic mechanical analysis (DMA), cytotoxicity tests with mice fibroblasts (L929), human osteoblast-like cells (MG63) and human MSC osteogenic differentiation (HBMSC) with alkaline phosphatase (ALP) activity and qPCR for osteogenic gene expression. The prototypes were also implanted into critical-size bone defects (rabbits' tibia) for 5 and 15 weeks, and after that were analysed by microCT and histology. The tests performed for the physical characterization of the materials showed the ability of the scaffolds to absorb and retain water-based solvents, as well as adequate mechanical resistance and viscoelastic properties. The cryogels had a heteroporous morphology with microporosity and macroporosity, which are essential conditions for the interaction between the cells and materials, and which consequently promote bone regeneration. Regarding the biological studies, all of the studied cryogels were non-cytotoxic by direct or indirect contact with cells. In fact, the scaffolds promoted the proliferation of the human MSCs, as well as the expression of the osteoblastic phenotype (osteogenic differentiation). The in vivo results showed bone tissue ingrowth and the materials' degradation, filling the critical bone defect after 15 weeks. Before and after irradiation, the studied scaffolds showed similar properties when compared to the results published in the literature. In conclusion, the material production process upscaling was optimized and the obtained prototypes showed reproducible properties relative to the bench development, and should be able to be commercialized. Therefore, it was a successful effort to harness knowledge from the basic sciences to produce a new biomedical device and enhance human health and wellbeing.

Incorporating sprouted chickpea flour in pasta increases brachial artery flow-mediated dilation.

PURPOSE: Consumption of alternative flours, such as sprouted chickpea flour, has shown increased popularity in recent years. Foods rich in antioxidants have been shown to influence brachial artery flow-mediated dilation (FMD), a non-invasive test of a crucial layer of the artery called the endothelium. Partially replacing the semolina flour in pasta with sprouted chickpea flour (SCF) may acutely affect endothelial function post-digestion. We sought to determine if FMD was higher, lower, or the same post-digestion of pasta made with 60% semolina flour and 40% SCF (SCF40) vs. post-digestion of pasta made with 100% semolina flour (SEM100, i.e., control). METHODS: Trolox equivalent antioxidant capacity (TEAC) analysis was performed on the same flour samples. Healthy participants underwent a screening visit and two randomized controlled meal data collection visits (SCF40 and SEM100). At each data collection visit, participants consumed 255 g of pasta with butter. FMD was assessed 2-3 h after pasta consumption. RESULTS: TEAC results showed that SCF40 (2.031 ± 0.096 mmol trolox/100 g sample) had significantly greater antioxidant capacity than SEM100 (1.736 ± 0.046 mmol trolox/100 g sample; p = 0.02). Twenty-two healthy participants (5 men and 17 women; 26 ± 2 years, 66.6 ± 2.3 kg, BMI = 24 ± 1 kg/m2, SBP = 114 ± 3 mmHg, DBP = 75 ± 2 mmHg, HR = 74 ± 3 BPM) were studied. FMD in the SCF40 condition (10.3% ± 1.2%) was greater than the SEM100 condition (7.9% ± 0.8%, p = 0.02). CONCLUSION: These data suggest that partial substitution with sprouted chickpea flour in place of semolina flour in pasta acutely improves post-digestion FMD, which may be beneficial for cardiovascular health (ClinicalTrials.gov Identifier: NCT03801486).

Dual RNA-Seq of Human Leprosy Lesions Identifies Bacterial Determinants Linked to Host Immune Response.

To understand how the interaction between an intracellular bacterium and the host immune system contributes to outcome at the site of infection, we studied leprosy, a disease that forms a clinical spectrum, in which progressive infection by the intracellular bacterium Mycobacterium leprae is characterized by the production of type I IFNs and antibody production. Dual RNA-seq on patient lesions identifies two independent molecular measures of M. leprae, each of which correlates with distinct aspects of the host immune response. The fraction of bacterial transcripts, reflecting bacterial burden, correlates with a host type I IFN gene signature, known to inhibit antimicrobial responses. Second, the bacterial mRNA:rRNA ratio, reflecting bacterial viability, links bacterial heat shock proteins with the BAFF-BCMA host antibody response pathway. Our findings provide a platform for the interrogation of host and pathogen transcriptomes at the site of infection, allowing insight into mechanisms of inflammation in human disease.

UV-induced hydrogen-atom transfer in 3,6-dithiopyridazine and in model compounds 2-thiopyridine and 3-thiopyridazine.

Monomeric 3,6-dithiopyridazine (3-mercapto- 6(1H)-pyridazinethione) was studied using the matrix-isolation method combined with quantum chemical calculations. The monomers of 3,6-dithiopyridazine, trapped from the gas phase into a low-temperature Ar matrix, were found to adopt the thione-thiol structure. In agreement with this experimental observation, the thione-thiol form was predicted (at the QCISD level) to be more stable by 13.5 kJ mol(-1) and by 39.6 kJ mol(-1) than the dithiol and the dithione tautomers, respectively. Monomers of 3,6-dithiopyridazine isolated in Ar matrixes were then irradiated with broadband UV (λ > 335 nm) light. Upon such irradiation, the thione-thiol form of the compound converted into the dithiol tautomer. The same phototransformation was observed when monochromatic λ = 385 nm laser light was used for irradiation. This allowed a first observation and spectral characterization of the dithiol form of 3,6-dithiopyridazine. Subsequent irradiation of the UV-generated dithiol tautomer with shorter-wavelength UV (λ > 275 nm) light led to partial repopulation of the thione-thiol form. Spectral signatures of the analogous photoreversibility were also found for the phototautomeric transformation in the model compound 3-thiopyridazine. The reliability of the QCISD predictions of relative energies of thiol and thione tautomeric forms was tested on the archetype example of 2-thiopyridine. For this compound, the comparison of the computed relative energy 10.9 kJ mol(-1) with the experimental estimate 10.0 ± 1.5 kJ mol(-1) (both in favor of the thiol form) was more than satisfactory.

Investigação de áreas de risco como metodologia complementar ao controle da leishmaniose visceral canina / Investigation of risk areas as complemental methodology for the control of canine visceral leishmaniasis

Foram investigadas áreas de risco de leishmaniose visceral canina no município de Camaçari, Bahia. Um total de 278 cães distribuídos em 141 residências, pertencentes a 20 áreas de risco investigadas, foi examinado sorologicamente (ELISA). A soroprevalência geral foi 21,7 por cento (56/258) depois da exclusão dos 20 cães usados no início do estudo para delimitar a área. Os resultados respectivos das análises univariada e multivariada dos fatores relacionados à infecção do cão por Leishmania chagasi, a captura e distribuição do vetor na área e a metodologia usada para localizar os focos caninos são discutidos.

Risk areas of canine visceral leishmaniasis in the city of Camaçari, Bahia, Brazil, were investigated. A total of 278 dogs from 141 homes pertaining to 20 investigated risk areas was serologically screened (ELISA). The general seroprevalence was 21.7 percent (56/258) after exclusion of 20 dogs used at the beginning of the survey to limit the study area. The respective results of the univariated and multivariated analysis of factors related to infection of dogs by Leishmania chagasi, to vector distribu-tion pattern in the area and to the methodology used to localize the canine focuses are discussed.

Pressure dependence of ionic polarizabilities in crystals.

A program in Maple V is developed that allows for the calculation of polarizabilities of both anions and cations on the basis of a new method and of the extension of the one developed previously. Furthermore, an alternative way of evaluation of delta r(i)/delta p is given. Values of delta alpha(total)/delta p can be evaluated for a given salt even when experimental data at high pressures are not available.

Effects of dopamine agonists on Cebus apella monkeys with previous long-term exposure to fluphenazine.

Sixty-one weeks after 48 weeks of treatment with fluphenazine decanoate or placebo, 37 socially living Cebus apella monkeys were evaluated for differences in dopaminergic sensitivity by exposure to 0.75 mg/kg, i.m. of amphetamine (AMPH) (indirect agonist) and apomorphine (APOM) (direct agonist). The fluphenazine-treated animals differed (p < or = 0.05) from control animals on some hourly measures of composite behavioral variables (CBVs). Animals exposed to fluphenazine showed a greater decrease in the aggressiveness CBV and a smaller decrease in self- and environment-directed behaviors than placebo animals. CBVs for normal locomotion and directs affiliation showed no significant differences. The fluphenazine-treated group showed greater agonist induction of stereotypic behavior (p < or = 0.01), and larger decreases in prolactin response to AMPH (p < or = 0.05). Our findings indicate that following extended treatment with an antipsychotic there is increased sensitivity to dopamine, as evidenced by stereotypies and possibly hypophyseal responsiveness.

Effects of protein-energy malnutrition on muscarinic receptor density and acetylcholinesterase activity in rat brain.

The effect of protein-energy malnutrition on the muscarinic receptor density as indicated by 3H-N-methylscopolamine binding, and acetylcholinesterase activity was studied in several brain areas (hippocampus, motor area, somatosensory area, and basal ganglia) of adult female rats. Malnutrition tended to cause a decrease in muscarinic receptors in the motor cortex (undernourished 350.0 +/- 33.5 vs. control 410.0 +/- 26.9 fmol/mg protein) and somatosensory cortex (undernourished 357.1 +/- 35.9 vs. control 416.7 +/- 29.4 fmol/mg protein). However, significant decreases in muscarinic receptor occurred in the hippocampus (undernourished 319.2 +/- 31.7 vs. control 403.1 +/- 43.6 fmol/mg protein) and basal ganglia (undernourished 297.0 +/- 11.8 vs. control 401.3 +/- 17.7 fmol/mg protein). No significant differences in acetylcholinesterase activity or protein content were observed between control and undernourished animals in any of the brain areas studied.

Cognitive effects of milacemide and methylphenidate in healthy young adults.

Cognitive effects of the novel glycine prodrug milacemide (400 mg), the catecholaminergic agonist methylphenidate (20 mg), and placebo were evaluated in 48 healthy young adults. Throughout a 6-h drug treatment day, subjects repeatedly performed tests of target-detection vigilance, immediate and delayed verbal free recall, and Buschke Selective Reminding; total free recall and forced-choice recognition tests were administered at the end of the day. Significant improvement in both vigilance reaction time and Selective Reminding Sum Recall was observed in the methylphenidate group. Contrary to expectations, the milacemide group evidenced significant declines in both vigilance perceptual sensitivity and free-recall difference scores (delayed-immediate). Vigilance reaction times significantly decreased over repeat testing in all groups, but only the methylphenidate group differed from placebo. The reaction-time functions for milacemide and placebo were similar, suggesting arousal was not diminished under milacemide and could not account for the cognitive decrements. No significant drug effects obtained for total free recall or recognition performance. Although the glycine prodrug milacemide was ineffective as a cognitive enhancer, the involvement of the NMDA receptor in memory function reported in the literature supports continued exploration of other approaches for manipulating NMDA receptor activity.

Muscarinic receptors and acetylcholinesterase activity in brain of patients with dementia

The present paper describes the determination of muscarinic receptor number ([3H]-N-methylscopolamine binding) and acetylcholinesterase activity in six brain areas (pre-central gyrus, post-central gyrus, hippocampus, caudate nucleus,lentiform nucleus and substantia inominata) of demented patients (diagnosed by screening tests and neurological evaluation) and age-matched controls.These was a significant increase in muscarinic receptors in the hippocampus and substanctia inominata (171.2 and 359.4fmol/mg protein, respectively) of the demented group as compared with controls (123.9 and 219.0 fmol/mg protein, respectively). No changes were observed in pre-and post-central gyrus, while a tendency towards decreased binding was detected in the caudate nucleus and lentiform nucleus of the demented group. Lower acetylcholinesterase activity was also detected in the demented group in all areas studied although the differences were significant only in the post-central gyrus, caudate nucleus and substantia inominata which showed a 49.21 and 25% decrease in enzyme activity respectively. The results are discussed in terms of a compensatory mechanism of presynaptic deficiency such as that occurring in Parkinson's disease

An enkephalin degrading aminopeptidase of human brain preserved during the vertebrate phylogeny.

1. A soluble human brain aminopeptidase which hydrolyses the Tyr-Gly bond of Met-enkephalin and Leu-enkephalin was identified in the brains of the following vertebrates: mammals (Callithrix jacchus and Rattus norvegicus), bird (Gallus domesticus), reptile (Tupinambis teguixin), amphibia (Bufo paracnemis), fish (Sarotherdon niloticus) and elasmobranchy (Galeocerdo cuvieri). 2. The properties of this enzyme are: molecular weight near 100,000 Da, isoelectric point near 4.9, optimum pH near 7.5, activation by dithiothreitol, strong inhibition by Cu2+, Zn2+, Ni2+, puromycin and bacitracin, hydrolysis of enkephalins and basic and neutral aminoacid-beta-naphythylamide substrates. 3. The results indicate the preservation of this human brain aminopeptidase during the course of vertebrate phylogeny.

Muscarinic receptors and acetylcholinesterase activity in brain of patients with dementia.

The present paper describes the determination of muscarinic receptor number ([3H]-N-methylscopolamine binding) and acetylcholinesterase activity in six brain areas (pre-central gyrus, post-central gyrus, hippocampus, caudate nucleus, lentiform nucleus and substantia innominata) of demented patients (diagnosed by screening tests and neurological evaluation) and age-matched controls. There was a significant increase in muscarinic receptors in the hippocampus and substantia innominata (171.2 and 359.4 fmol/mg protein, respectively) of the demented group as compared with controls (123.9 and 219.0 fmol/mg protein, respectively). No changes were observed in pre- and post-central gyrus, while a tendency towards decreased binding was detected in the caudate nucleus and lentiform nucleus of the demented group. Lower acetylcholinesterase activity was also detected in the demented group in all areas studied although the differences were significant only in the post-central gyrus, caudate nucleus and substantia innominata which showed a 49, 21 and 25% decrease in enzyme activity, respectively. The results are discussed in terms of a compensatory mechanism of presynaptic deficiency such as that occurring in Parkinson's disease.

Efficacy of benztropine therapy for drooling.

This study assessed the efficacy of synthetic anticholinergic benztropine and incidence of side-effects in 20 developmentally-disabled patients with severe drooling. The double-blind, placebo-controlled, crossover protocol included one-week baseline, two-week placebo and two-week benztropine conditions (mean dose 3.8 mg). A significant decrease in drooling during the benztropine condition relative to placebo was demonstrated and conservative response rates (calculated by deleting placebo responders), ranged up to 65 to 70 per cent. For patients completing the protocol the incidence of side-effects did not differ across conditions and minor problems such as a dry mouth were eliminated by small dose adjustments. More serious cholinergic side-effects, which resolved within 24 to 48 hours, necessitated discontinuation of the drug in three patients. This study demonstrates that synthetic anticholinergics can provide an important therapeutic alternative to surgical and behavioral therapies for drooling.

Isolation and characterization of human brain aminopeptidase which hydrolysis enkephalin-containing peptides.

A human brain aminopeptidase, which hydrolyses low molecular weight enkephalin-containing peptides (ECPs), was purified to apparent homogeneity from the homogenate of human brain. The enzyme purification involved DEAE-cellulose chromatography and preparative polyacrylamide gel electrophoresis. The purified aminopeptidase hydrolyses only the Tyr1-Gly2 bond of enkephalines and of ECPs. The rate of hydrolysis of Met-enkephalin and Leu-enkephalin was higher than the rate of hydrolysis of ECPs containing 7 to 13 aminoacid residues. Large ECPs such as peptide E and beta-endorphin were not hydrolysed. The molecular weight of this enzyme is about 100,000 daltons, as determined by gel filtration on Sephadex G-200 and by polyacrylamide gel electrophoresis in presence of sodium dodecyl sulfate. The enzyme has an isoelectric point of pH 4.9, is activated by dithiothreitol (DTT) and inhibited by puromycin, bacitracin, p-mercuryacetate, Zn++, Cu++ and Ni++. The optimum pH for enzyme activity is 7.5.

Concurrent on-line analysis of striatal ascorbate, dopamine and dihydroxyphenylacetic acid concentrations by in vivo voltammetry.

A new method for on-line analysis of in vivo differential normal pulse voltammograms is reported. They are fitted by least squares to an expression describing the contribution of different electroactive species and the baseline. Its validity for resolving ascorbate (AA), dopamine (DA) and DOPAC signals is evidenced by both in vitro testing and the changes recorded in the striatum of anesthetized rats following drug treatments having well-known effects on DA release and metabolism. Thus, pargyline and amphetamine treatments respectively increased and decreased DA and DOPAC, whereas they both were increased by haloperidol. AA levels followed those of DA except when haloperidol was given.

Mianserin pharmacokinetics and behavior in hyperkinetic children.

The present study was conducted to derive pediatric mianserin pharmacokinetic parameters, which were compared to those from healthy young adults, and to obtain preliminary information regarding the utility of mianserin for the management of hyperkinesis in children. The sample consisted of six prepubescent children with hyperkinetic behavior disorders who had not responded, or had developed tolerance to, stimulant medication. Mianserin pharmacokinetics were derived from plasma samples obtained over a 36- to 50-hour period following a single oral dose which ranged from 0.28 to 0.72 mg/kg. Children evidenced a significantly faster elimination half-life and a significantly smaller apparent kinetic volume of distribution than did adults, whereas maximum plasma concentration, time to maximum concentration, and apparent oral plasma clearance were similar. Ratings of behavioral deviance were obtained from teachers and parents during placebo and mianserin titration to a maximum dose of 40 mg/day. Although half the children showed some decrease in hyperactivity ratings, the small sample size and high variability of response preclude conclusions regarding the efficacy of mianserin for childhood hyperkinesis. Possible side effects in our sample included akathisia, excitability, insomnia, and migraine-like headache, as well as cardiovascular effects of tachycardia and two instances of minor electrocardiographic change. Our pharmacokinetic findings will be of import should mianserin prove useful for such childhood disorders as depression and/or enuresis.(ABSTRACT TRUNCATED AT 250 WORDS)