Marco Antonio de Oliveira

Haemozoin in Schistosoma mansoni.

a Departamento de Bioquimica Medica, Uni6ersidade Federal do Rio de Janeiro, Cidade Uni6ersitaria, 21941-590, Rio de Janeiro, RJ, Brazil b Di6isao de Quimica, Setor de Quimica Orgânica, CENPES, Petrobras, Cidade Uni6ersitaria, 21949-900, Rio de Janeiro, RJ, Brazil c Centro de Biociencias e Biotecnologia, Uni6ersidade Estadual do Norte Fluminense, Campos dos Goytacazes, RJ, 28015-620, Brazil d Instituto de Biofisica Carlos Chagas Filho, Uni6ersidade Federal do Rio de Janeiro, Cidade Uni6ersitaria, 21941-590, Rio de Janeiro, RJ, Brazil

On the pro-oxidant effects of haemozoin

Haemozoin (Hz) is a haem aggregate produced in some blood‐feeding organisms. There is a general belief that Hz formation would be a protective mechanism against haem toxicity. Here we show that when aggregated into Hz, haem is less deleterious than its free form. When haem was added to phosphatidylcholine (PC) liposomes, there was an intense stimulation of oxygen consumption, which did not occur when Hz was incubated with the same preparation. Evaluation of oxygen radical attack to lipids, by measurement of thiobarbituric acid reactive substances (TBARS), showed significantly lower levels of lipid peroxidation in samples containing PC liposomes incubated with Hz than with haem. However, TBARS production induced by Hz was much higher when using 2‐deoxyribose (2‐DR) as substrate, than with PC liposomes. Spin‐trapping analysis by electron paramagnetic resonance (EPR) of Hz and tert‐butylhydroperoxide (tert‐BuOOH) showed that production of methoxyl and tert‐butoxyl radicals was only slightly reduced compared to what was observed with haem. Interestingly, when large Hz crystals were used in 2‐DR TBARS assays and tert‐BuOOH EPR experiments, the pro‐oxidant effects of Hz were strongly reduced. Moreover, increasing concentrations of Hz did not induce erythrocyte lysis, as occurred with haem. Thus, the reduced capacity of Hz to impose radical damage seems to result from steric hindrance of substrates to access the aggregated haem, that becomes less available to participate in redox reactions.

The beta-3 adrenoceptor agonist, mirabegron relaxes isolated prostate from human and rabbit: new therapeutic indication?

Alpha1 (α1)‐blockers, 5‐alpha reductase and phosphodiesterase type‐5 inhibitors are pharmacological classes currently available for benign prostatic hyperplasia (BPH) treatment. Mirabegron, a beta‐3 adrenoceptor (β3‐AR) agonist has been approved for the therapy of overactive bladder and may constitute a new therapeutic option for BPH treatment. This study is aimed to evaluate the in vitro effects of mirabegron in human and rabbit prostatic smooth muscle.

A Genome-Wide Association Study for Agronomic Traits in Soybean Using SNP Markers and SNP-Based Haplotype Analysis

Mapping quantitative trait loci through the use of linkage disequilibrium (LD) in populations of unrelated individuals provides a valuable approach for dissecting the genetic basis of complex traits in soybean (Glycine max). The haplotype-based genome-wide association study (GWAS) has now been proposed as a complementary approach to intensify benefits from LD, which enable to assess the genetic determinants of agronomic traits. In this study a GWAS was undertaken to identify genomic regions that control 100-seed weight (SW), plant height (PH) and seed yield (SY) in a soybean association mapping panel using single nucleotide polymorphism (SNP) markers and haplotype information. The soybean cultivars (N = 169) were field-evaluated across four locations of southern Brazil. The genome-wide haplotype association analysis (941 haplotypes) identified eleven, seventeen and fifty-nine SNP-based haplotypes significantly associated with SY, SW and PH, respectively. Although most marker-trait associations were environment and trait specific, stable haplotype associations were identified for SY and SW across environments (i.e., haplotypes Gm12_Hap12). The haplotype block 42 on Chr19 (Gm19_Hap42) was confirmed to be associated with PH in two environments. These findings enable us to refine the breeding strategy for tropical soybean, which confirm that haplotype-based GWAS can provide new insights on the genetic determinants that are not captured by the single-marker approach.

Role of a Novel Tetrodotoxin-Resistant Sodium Channel in the Nitrergic Relaxation of Corpus Cavernosum from the South American Rattlesnake Crotalus Durissus Terrificus

INTRODUCTION Coitus in snakes may last up to 28 hours; however, the mechanisms involved are unknown. AIM To evaluate the relevance of the nitric oxide (NO)-cyclic guanosine monophosphate (cGMP)-phosphodiesterase type 5 (PDE5) system in snake corpus cavernosum reactivity. METHODS Hemipenes were removed from anesthetized South American rattlesnakes (Crotalus durissus terrificus) and studied by light and scanning electronic microscopy. Isolated Crotalus corpora cavernosa (CCC) were dissected from the non-spiny region of the hemipenises, and tissue reactivity was assessed in organ baths. MAIN OUTCOME MEASURES Cumulative concentration-response curves were constructed for acetylcholine (ACh), sodium nitroprusside (SNP), 5-cyclopropyl-2-[1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-yl]pyrimidin-4-ylamine (BAY 41-2272), and tadalafil in CCC precontracted with phenylephrine. Relaxation induced by electrical field stimulation (EFS) was also done in the absence and presence of N(ω) nitro-L-arginine methyl ester (L-NAME; 100 µM), 1H-[1, 2, 4] oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 10 µM) and tetrodotoxin (TTX; 1 µM). RESULTS The hemipenes consisted of two functionally concentric corpora cavernosa, one of them containing radiating bundles of smooth muscle fibers (confirmed by α-actin immunostaining). Endothelial and neural nitric oxide synthases were present in the endothelium and neural structures, respectively; whereas soluble guanylate cyclase and PDE5 were expressed in trabecular smooth muscle. ACh and SNP relaxed isolated CCC, with the relaxations being markedly reduced by L-NAME and ODQ, respectively. BAY 41-2272 and tadalafil caused sustained relaxations with potency (pEC(50) ) values of 5.84 ± 0.17 and 5.10 ± 0.08 (N=3-4), respectively. In precontracted CCC, EFS caused frequency-dependent relaxations that lasted three times longer than those in mammalian CC. Although these relaxations were almost abolished by either L-NAME or ODQ, they were unaffected by TTX. In contrast, EFS-induced relaxations in marmoset CC were abolished by TTX. CONCLUSIONS Rattlesnake CC relaxation is mediated by the NO-cGMP-PDE5 pathway in a manner similar to mammals. The novel TTX-resistant Na channel identified here may be responsible for the slow response of smooth muscle following nerve stimulation and could explain the extraordinary duration of snake coitus.

The Evolutionary Implications of Hemipenial Morphology of Rattlesnake Crotalus durissus terrificus (Laurent, 1768) (Serpentes: Viperidae: Crotalinae)

Most amniotes vertebrates have an intromittent organ to deliver semen. The reptile Sphenodon and most birds lost the ancestral penis and developed a cloaca-cloaca mating. Known as hemipenises, the copulatory organ of Squamata shows unique features between the amniotes intromittent organ. They are the only paired intromittent organs across amniotes and are fully inverted and encapsulated in the tail when not in use. The histology and ultrastructure of the hemipenes of Crotalus durissus rattlesnake is described as the evolutionary implications of the main features discussed. The organization of hemipenis of Crotalus durissus terrificus in two concentric corpora cavernosa is similar to other Squamata but differ markedly from the organization of the penis found in crocodilians, testudinata, birds and mammals. Based on the available data, the penis of the ancestral amniotes was made of connective tissue and the incorporation of smooth muscle in the framework of the sinusoids occurred independently in mammals and Crotalus durissus. The propulsor action of the muscle retractor penis basalis was confirmed and therefore the named should be changed to musculus hemipenis propulsor.The retractor penis magnus found in Squamata has no homology to the retractor penis of mammals, although both are responsible for the retraction of the copulatory organ.

Tetrodotoxin-insensitive electrical field stimulation-induced contractions on Crotalus durissus terrificus corpus cavernosum

Reptiles are the first amniotes to develop an intromitent penis, however until now the mechanisms involved in the electrical field stimulation-induced contraction on corpora cavernosa isolated from Crotalus durissus terrificus were not investigated. Crotalus and rabbit corpora cavernosa were mounted in 10 mL organ baths for isometric tension recording. Electrical field stimulation (EFS)-induced contractions were performed in presence/absence of phentolamine (10 μM), guanethidine (30 μM), tetrodotoxin (1 μM and 1mM), A-803467 (10 μM), 3-iodo-L-Tyrosine (1 mM), salsolinol (3 μM) and a modified Krebs solution (equimolar substitution of NaCl by N-methyl–D-glucamine). Immuno-histochemistry for tyrosine hydroxylase was also performed. Electrical field stimulation (EFS; 8 Hz and 16 Hz) caused contractions in both Crotalus and rabbit corpora cavernosa. The contractions were abolished by previous incubation with either phentolamine or guanethidine. Tetrodotoxin (1 μM) also abolished the EFS-induced contractions of rabbit CC, but did not affect EFS-induced contractions of Crotalus CC. Addition of A-803467 (10 μM) did not change the EFS-induced contractions of Crotalus CC but abolished rabbit CC contractions. 3-iodo-L-Tyrosine and salsolinol had no effect on EFS-induced contractions of Crotalus CC and Rabbit CC. Replacement of NaCl by N- Methyl-D-glucamine (NMDG) abolished EFS-induced contractions of rabbit CC, but did not affect Crotalus CC. The presence of tyrosine hydroxylase was identified in endothelial cells only of Crotalus CC. Since the EFS-induced contractions of Crotalus CC is dependent on catecholamine release, insensitive to TTX, insensitive to A803467 and to NaCl replacement, it indicates that the source of cathecolamine is unlikely to be from adrenergic terminals. The finding that tyrosine hydroxylase is present in endothelial cells suggests that these cells can modulate Crotalus CC tone.

Pharmacokinetic and pharmacodynamic evaluation of a nanotechnological topical formulation of lidocaine/prilocaine (nanorap) in healthy volunteers.

Background: Nanorap is a new nanotechnological formulation for topical anesthesia composed of lidocaine (2.5%) and prilocaine (2.5%). This study evaluated the pharmacokinetics of Nanorap. For the determination of lidocaine and prilocaine in human plasma, a new method using high-performance liquid chromatography coupled with tandem mass spectrometry was developed. Nanorap pharmacodynamic (PD) and its physical proprieties were also evaluated. Methods: Nanorap was administered by topical application of 2 g to healthy volunteers, and blood samples were collected for the pharmacokinetics analysis. The drugs were extracted from plasma by liquid–liquid extraction with ether/hexane (80/20, vol/vol). The chromatography separation was performed on a Genesis C18 analytical column 4 &mgr;m (100 × 2.1 mm i.d.) with a mobile phase of methanol/acetonitrile/water (40/30/30, for lidocaine, and 50/30/20, for prilocaine, vol/vol/vol) + 2 mM of ammonium acetate and ropivacaine as internal standard. The drugs were quantified using a mass spectrometer with an electrospray source in the electrospray ionization positive mode configured for multiple reaction monitoring. The PD of Nanorap was evaluated with the use of a visual analog scale. Nanorap was characterized by cryofracture. Results: The chromatography run-time was 5.5 minutes for lidocaine and 3.3 minutes for prilocaine, and the lower limit of quantification was 0.05 ng/mL for both drugs. Mean Cmax was 6.62 and 1.72 ng/mL for lidocaine and prilocaine, respectively. Median Tmax was 6.5 hours for both drugs. Nanocapsules had a mean size of 88 nm and mean drug association of 92.5% and 89% for lidocaine and prilocaine, respectively. The PD study showed that Nanorap has a sufficient analgesic effect (>30% reduction in pain) after 10 minutes of application. Conclusions: A new simple, selective, and sensitive method for determination of lidocaine and prilocaine in human plasma was developed. Nanorap generated safe plasma levels of the drugs and satisfactory analgesic effect.

Burnout among medical students during the first years of undergraduate school: Prevalence and associated factors

Objective To evaluate the prevalence and possible factors associated with the development of burnout among medical students in the first years of undergraduate school. Method A cross-sectional study was conducted at the Barretos School of Health Sciences, Dr. Paulo Prata. A total of 330 students in the first four years of medical undergraduate school were invited to participate in responding to the sociodemographic and Maslach Burnout Inventory-Student Survey (MBI-SS) questionnaires. The first-year group consisted of 150 students, followed by the second-, third-, and fourth-year groups, with 60 students each. Results Data from 265 students who answered at least the sociodemographic questionnaire and the MBI-SS were analyzed (response rate = 80.3%). One (n = 1, 0.3%) potential participant viewed the Informed Consent Form but did not agree to participate in the study. A total of 187 students (187/265, 70.6%) presented high levels of emotional exhaustion, 140 (140/265, 52.8%) had high cynicism, and 129 (129/265, 48.7%) had low academic efficacy. The two-dimensional criterion indicated that 119 (44.9%) students experienced burnout. Based on the three-dimensional criterion, 70 students (26.4%) presented with burnout. The year with the highest frequency of affected students for both criteria was the first year (p = 0.001). Personal attributes were able to explain 11% (ΔR = 0.11) of the variability of burnout under the two-dimensional criterion and 14.4% (R2 = 0.144) under the three-dimensional criterion. Conclusion This study showed a high prevalence of burnout among medical students in a private school using active teaching methodologies. In the first years of graduation, students’ personal attributes (optimism and self-perception of health) and school attributes (motivation and routine of the exhaustive study) were associated with higher levels of burnout. These findings reinforce the need to establish preventive measures focused on the personal attributes of first-year students, providing better performance, motivation, optimism, and empathy in the subsequent stages of the course.