Genetic differentiation and adaptive evolution at reproductive loci in incipient Drosophila species.

Accessory gland proteins (Acps) are part of the seminal fluid of male Drosophila flies. Some Acps have exceptionally high evolutionary rates and evolve under positive selection. Proper interactions between Acps and female reproductive molecules are essential for fertilization. These observations lead to suggestions that fast evolving Acps could be involved in speciation by promoting reproductive incompatibilities between emerging species. To test this hypothesis, we used population genetics data for three sibling species: D. mayaguana, D. parisiena and D. straubae. The latter two species are morphologically very similar and show only incipient reproductive isolation. This system allowed us to examine Acp evolution at different time frames with respect to speciation and reproductive isolation. Comparing data of 14 Acp loci with data obtained for other genomic regions, we found that some Acps show extraordinarily high levels of divergence between D. mayaguana and its two sister species D. parisiena and D. straubae. This divergence was likely driven by adaptive evolution at several loci. No fixed nucleotide differences were found between D. parisiena and D. straubae, however. Nevertheless, some Acp loci did show significant differentiation between these species associated with signs of positive selection; these loci may be involved in this early phase of the speciation process.

Evaluating the efficacy of hydrogen peroxide vapour against foot-and-mouth disease virus within a BSL4 biosafety facility.

An evaluation was made of the efficacy of 35% hydrogen peroxide vapour (HPV) against foot-and-mouth disease virus (FMDV) in a biosafety facility. Biological indicators (BIs) were produced using three serotypes of FMDV, all with a titre of ≥106 TCID50 per ml. Fifteen BIs of each serotype were distributed across five locations, throughout a 30-m3 airlock chamber, producing a total of 45 BIs. Thirty-five percent HPV was generated and applied using a Bioquell vaporization module located in the centre of the chamber. After a dwell period of 40 min, the HPV was removed via the enclosures air handling system and the BIs were collected. The surfaces of the BIs were recovered into Glasgow's modified Eagle's medium (GMEM), cultivated in BHK21 Cl13 cell culture and analysed for evidence of cytopathic effect (CPE). No CPE was detected in any BI sample. Positive controls showed CPE. The experimentation shows that FMDV is susceptible to HPV decontamination and presents a potential alternative to formaldehyde. SIGNIFICANCE AND IMPACT OF THE STUDY: Foot-and-mouth disease virus (FMDV) is an important pathogen in terms of biosafety due to its infectious nature and wide range of host animals, such as cattle, sheep, goats and pigs. Outbreaks of FMDV can have a severe impact on livestock production, causing morbidity, mortality, reduced yields and trade embargoes. Laboratories studying FMDV must possess BSL4 robust bio-decontamination methods to prevent inadvertent release. Formaldehyde has been the primary agent for environmental decontamination, but its designation as a human carcinogen has led to a search for alternatives. This study shows 35% hydrogen peroxide vapour has the potential to be a rapid, effective, residue-free alternative.

Effects of glycerol, equilibration time and antioxidants on post-thaw functional integrity of bovine spermatozoa directly obtained from epididymis.

This work aimed to evaluate the effect of stabilisation times, glycerol concentration, and the catalase and superoxide dismutase supplementation of diluent on parameters of frozen-thawed spermatozoa from epididymis of Nelore bulls: Experiment 1: spermatozoa diluted in Tris-egg yolk with glycerol (3%, 5% or 7%) and stabilisation times (0, 2 or 4 hr at 5°C); Experiment 2: Tris-egg yolk only, Tris-egg yolk with catalase (CAT, 50 or 100 U ml-1 ) or superoxide dismutase (SOD, 50 or 100 U ml-1 ). Frozen-thawed spermatozoa were evaluated for kinetic parameters, plasma membrane and acrosome integrity, mitochondrial activity and IVF capacity. ALH and BCF were affected (p < .05) by glycerol at 3% after 4-hr equilibration time and 7% after 2-hr equilibration time. Glycerol 3% had lower (p < .05) iPM and iAc after 4 hr. Glycerol 5% had greater (p < .05) hPMM after 4 hr and iAc after 2 hr than at 0 hr. SOD 100 U ml-1 had lower (p < .05) linearity and wobble compared to control group. No was observed differences to fertilisation rate (p < .05) among groups. In conclusion, glycerol 5% in Tris-egg yolk extender for 4 hr is suitable for the preservation of sperm kinetics and membrane integrity. CAT (50 and 100 U ml-1 ) or SOD (50-100 U ml-1 ) had no beneficial effects on sperm kinetics, plasma and acrosomal membrane integrity, mitochondrial activity or the capacity for IVF of frozen-thawed spermatozoa from epididymis of Nelore bulls.

Ultrastructure evaluation of goat spermatozoa after freezing in a skim milk-based extender with Trolox supplementation.

The aim of this work was to evaluate the in vitro effect of adding Trolox in freezing extender for goat semen. Ejaculates from five bucks were evaluated, and when approved, the samples were pooled, diluted according to experimental groups [Trolox 0 (control), 30, 60 and 120 nmol ml(-1) ] and frozen in an automated system. Thawed samples (37 °C/30 s) were evaluated for plasma membrane (PMi) and acrosome integrity (Aci), mitochondrial membrane potential (MMP) and sperm kinematics by CASA system. Spermatozoa ultrastructure was evaluated in fresh and post-thawed semen. No significant difference (P > 0.05) was observed among control and Trolox groups in the analyses of PMi, Aci, MMP and CASA in goat spermatozoa after thawing. Samples of 60 and 120 nmol ml(-1) Trolox groups had a higher percentage of cells that had intact plasma membranes in spermatozoa head than in the other groups, although they did not differ (P > 0.05) before being frozen. A higher percentage (P < 0.05) of spermatozoa with intact mitochondria was observed in fresh semen, control and Trolox 60 nmol ml(-1) groups than in the other groups. Addition of Trolox to skim milk extender at 60 nmol ml(-1) ultrastructurally preserves the plasma membrane and mitochondrial sheath integrity in goat spermatozoa after cryopreservation.

Bet v 1--a Trojan horse for small ligands boosting allergic sensitization?

BACKGROUND: Birch pollen allergy represents the main cause of winter and spring pollinosis in the temperate climate zone of the northern hemisphere and sensitization towards Bet v 1, the major birch pollen allergen, affects over 100 million allergic patients. The major birch pollen allergen Bet v 1 has been described as promiscuous acceptor for a wide variety of hydrophobic ligands. OBJECTIVE: In search of intrinsic properties of Bet v 1, which account responsible for the high allergenic potential of the protein, we thought to investigate the effects of ligand-binding on immunogenic as well as allergenic properties. METHODS: As surrogate ligand of Bet v 1 sodium deoxycholate (DOC) was selected. Recombinant and natural Bet v 1 were characterised physico-chemically as well as immunologically in the presence or absence of DOC, and an animal model of allergic sensitization was established. Moreover, human IgE binding to Bet v 1 was analysed by nuclear magnetic resonance (NMR) spectroscopy. RESULTS: Ligand-binding had an overall stabilizing effect on Bet v 1. This translated in a Th2 skewing of the immune response in a mouse model. Analyses of human IgE binding on Bet v 1 in mediator release assays revealed that ligand-bound allergen-induced degranulation at lower concentrations; however, in basophil activation tests with human basophils ligand-binding did not show this effect. For the first time, human IgE epitopes on Bet v 1 were determined using antibodies isolated from patients' sera. The IgE epitope mapping of Bet v 1 demonstrated the presence of multiple binding regions. CONCLUSIONS AND CLINICAL RELEVANCE: Deoxycholate binding stabilizes conformational IgE epitopes on Bet v 1; however, the epitopes themselves remain unaltered. Therefore, we speculate that humans are exposed to both ligand-bound and free Bet v 1 during sensitization, disclosing the ligand-binding cavity of the allergen as key structural element.

Effect of leptin on in vivo goat embryo production.

The aim of this study was to evaluate the effect of leptin administration during superovulation on in vivo goat embryo production. Ten mature does were superovulated with 133 mg follicle-stimulating hormone (FSH) i.m. in six descending doses at 12-h intervals. The goats received 4.8 µg/kg human recombinant leptin s.c. (leptin group, n = 5) or phosphate-buffered saline (PBS) (control group, n = 5) with the first and second FSH doses. The does were mated and subjected to embryo collection by transcervical technique 6 days later. The total number of cells per embryo and the number of cells with fragmented DNA were assessed in selected blastocysts by combining Hoechst 33342 and terminal dUTP nick-end labelling (TUNEL) staining. Plasma concentrations of oestradiol (E2 ) and progesterone (P4 ) were determined by electrochemiluminescence from the day of FSH treatment, on the day of superovulatory oestrus and on the day before embryo collection. Compared with the control group, the does that received leptin had a higher number of transferable embryos (p < 0.005), fewer embryos classified as degenerated (p < 0.001) and fewer TUNEL-positive cells/blastocyst (p < 0.001). The number of transferable embryos was positively correlated with E2 concentrations on day of oestrus (r = 0.562; p < 0.01) and P4 concentrations on the day of embryo collection (r = 0.912; p < 0.001). We concluded that in vivo leptin administration during FSH treatment improved embryo quality and affected ovarian steroidogenesis in superovulated goats.

Sex-Specific Patterns of Cerebral Atrophy and Enlarged Perivascular Spaces in Patients with Cerebral Amyloid Angiopathy and Dementia.

BACKGROUND AND PURPOSE: Cerebral amyloid angiopathy is characterized by amyloid ß deposition in leptomeningeal and superficial cortical vessels. Cognitive impairment is common and may occur independent of concomitant Alzheimer disease neuropathology. It is still unknown which neuroimaging findings are associated with dementia in cerebral amyloid angiopathy and whether they are modulated by sex. This study compared MR imaging markers in patients with cerebral amyloid angiopathy with dementia or mild cognitive impairment or who are cognitively unimpaired and explored sex-specific differences. MATERIALS AND METHODS: We studied 58 patients with cerebral amyloid angiopathy selected from the cerebrovascular and memory outpatient clinics. Clinical characteristics were collected from clinical records. Cerebral amyloid angiopathy was diagnosed on MR imaging on the basis of the Boston criteria. Visual rating scores for atrophy and other imaging features were independently assessed by 2 senior neuroradiologists. RESULTS: Medial temporal lobe atrophy was higher for those with cerebral amyloid angiopathy with dementia versus those cognitively unimpaired (P = .015), but not for those with mild cognitive impairment. This effect was mainly driven by higher atrophy in men with dementia, compared with women with and without dementia (P = .034, P = .012; respectively) and with men without dementia (P = .012). Enlarged perivascular spaces in the centrum semiovale were more frequent in women with dementia versus men with and without dementia (P = .021, P = .011; respectively) and women without dementia (P = .011). CONCLUSIONS: Medial temporal lobe atrophy was more prominent in men with dementia, whereas women showed a higher number of enlarged perivascular spaces in the centrum semiovale. Overall, this finding suggests differential pathophysiologic mechanisms with sex-specific neuroimaging patterns in cerebral amyloid angiopathy.

In vitro and in vivo evaluation of ram sperm frozen in tris egg-yolk and supplemented with superoxide dismutase and reduced glutathione.

The aim of the present study was to evaluate the in vitro and in vivo effect of the addition of superoxide dismutase (SOD) and reduced glutathione (GSH) to ram semen freezing extender. Significant differences (p < 0.05) were detected between groups regarding total motility (TM), straightness (STR) and wobble (WOB), for which the GSH 7 mM group had lesser TM and better STR than the other groups and the GSH 5 and 7 mM groups had higher wobble values than the control, SOD 25 and 100 U/ml groups. The ultrastructural analysis revealed that the acrosome was better preserved after freezing in the SOD 100 U/ml and GSH 2 and 5 mM (p < 0.05) groups than the other groups, whereas mitochondria in both the control group and the 7 mM GSH group suffered the greatest damage. The plasma membrane remained preserved after freezing, regardless of the group. For in vivo fertilization, the SOD group achieved better results than the GSH group (p > 0.05). It can therefore be concluded that the addition of SOD 100 U/ml and GSH 2 and 5 mM preserves the acrosome integrity of frozen ram spermatozoa, while the addition of SOD 100 U/ml to Tris egg-yolk extender offers protection to the membranes of sperm cells after thawing.

Mitogenomic analysis of extant condor species provides insight into the molecular evolution of vultures.

The evolution of large vultures linked to mountainous habitats was accompanied by extreme physiological and behavioral specializations for energetically efficient flights. However, little is known on the genetic traits associated with the evolution of these obligate soaring scavengers. Mitochondrial DNA plays a vital role in regulating oxidative stress and energy production, and hence may be an important target of selection for flight performance. Herein, we characterized the first mitogenomes of the Andean and California condors, the world's heaviest flying birds and the only living representatives of the Vultur and Gymnogyps genus. We reconstructed the phylogenetic relationships and evaluated possible footprints of convergent evolution associated to the life-history traits and distributional range of vultures. Our phylogenomic analyses supported the independent evolution of vultures, with the origin of Cathartidae in the early Paleogene (~ 61 Mya), and estimated the radiation of extant condors during the late Miocene (~ 11 Mya). Selection analyses indicated that vultures exhibit signals of relaxation of purifying selection relative to other accipitrimorph raptors, possibly indicating the degeneration of flapping flight ability. Overall, our results suggest that the extreme specialization of vultures for efficient soaring flight has compensated the evolution of large body sizes mitigating the selection pressure on mtDNA.

Structural biology of membrane-acting peptides: conformational plasticity of anticoccidial peptide PW2 probed by solution NMR.

The bottleneck for the complete understanding of the structure-function relationship of flexible membrane-acting peptides is its dynamics. At the same time, not only the structure but also the dynamics are the key points for their mechanism of action. Our model is PW2, a TRP-rich, cationic peptide selected from phage display libraries that shows anticoccidial activity against Eimeria acervulina. In this manuscript we used a combination of several NMR techniques to tackle these difficulties. The structural features of the membrane-acting peptide PW2 was studied in several membrane mimetic environments: we compared the structural features of PW2 in SDS and DPC micelles, that were reported earlier, with the structure properties in different lipid vesicles and the peptide free in water. We were able to unify the structural information obtained in each of these systems. The structural constraints of the peptide free in water were fundamental for the understanding of plasticity necessary for the membrane interaction. Our data suggested that the WWR sequence is the region responsible for anchoring the peptide to the interfaces, and that this same region displays some degree of conformational order in solution. For PW2, we found that affinity is related to the aromatic region, by anchoring the peptide to the membrane, and specificity is related to the N- and C-termini, which are able to accommodate in the membrane due to its plasticity.

Effect of Ginkgo biloba on the reproductive outcome and oxidative stress biomarkers of streptozotocin-induced diabetic rats.

The aim of the present study was to evaluate the effect of Ginkgo biloba treatment (EGb 761, 200 mg kg-1 day-1) administered from day 0 to 20 of pregnancy on maternal reproductive performance and on the maternal and fetal liver antioxidant systems of streptozotocin-induced diabetic Wistar rats. On day 21 of pregnancy, the adult rats (weighing approximately 250 +/- 50 g, minimum number = 13/group) were anesthetized to obtain maternal and fetal liver samples for superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), and total glutathione (GSH-t) determinations. The uterus was weighed with its contents. The diabetic (G3) and treated diabetic (G4) groups of rats presented significant maternal hyperglycemia, reduced term pregnancy rate, impaired maternal reproductive outcome and fetal-placental development, decreased GSH-Px (G3 = G4 = 0.6 +/- 0.2) and SOD (G3 = 223.0 +/- 84.7; G4 = 146.1 +/- 40.8), and decreased fetal CAT activity (G3 = 22.4 +/- 10.6; G4 = 34.4 +/- 14.1) and GSH-t (G3 = G4 = 0.3 +/- 0.2), compared to the non-diabetic groups (G1, untreated control; G2, treated). For G1, maternal GSH-Px = 0.9 +/- 0.2 and SOD = 274.1 +/- 80.3; fetal CAT = 92.6 +/- 82.7 and GSH-t = 0.6 +/- 0.5. For G2, G. biloba treatment caused no toxicity and did not modify maternal or fetal-placental data. EGb 761 at the nontoxic dose used (200 mg kg-1 day-1), failed to modify the diabetes-associated increase in maternal glycemia, decrease in pregnancy rate, decrease in antioxidant enzymes, and impaired fetal development when the rats were treated throughout pregnancy (21 days).

Metabolomic Analysis Reveals Vitamin D-induced Decrease in Polyol Pathway and Subtle Modulation of Glycolysis in HEK293T Cells.

We combined 1H NMR metabolomics with functional and molecular biochemical assays to describe the metabolic changes elicited by vitamin D in HEK293T, an embryonic proliferative cell line adapted to high-glucose concentrations. Activation of the polyol pathway, was the most important consequence of cell exposure to high glucose concentration, resembling cells exposed to hyperglycemia. Vitamin D induced alterations in HEK293T cells metabolism, including a decrease in sorbitol, glycine, glutamate, guanine. Vitamin D modulated glycolysis by increasing phosphoglycerate mutase and decreasing enolase activities, changing carbon fate without changing glucose consumption, lactate export and Krebs cycle. The decrease in sorbitol intracellular concentration seems to be related to vitamin D regulated redox homeostasis and protection against oxidative stress, and helped maintaining the high proliferative phenotype, supported by the decrease in glycine and guanine and orotate concentration and increase in choline and phosphocholine concentration. The decrease in orotate and guanine indicated an increased biosynthesis of purine and pyrimidines. Vitamin D elicited metabolic alteration without changing cellular proliferation and mitochondrial respiration, but reclaiming reductive power. Our study may contribute to the understanding of the metabolic mechanism of vitamin D upon exposure to hyperglycemia, suggesting a role of protection against oxidative stress.

Implications of protein conformational diversity for binding and development of new biological active compounds.

The new generation of biologically active compounds developed during the 20(th) century relied on knowledge of enzymology and protein structure, and were based initially, on the understanding that protein-protein and small molecule-protein interactions occurred through a lock-and-key mechanism. Later, evidence suggested that this mechanism was usually followed by a conformational change, known as induced fit. Recent studies on protein dynamics, mainly by nuclear magnetic resonance (NMR) relaxation measurements, have shown that proteins are not structured in a unique conformation. Rather, they frequently have regions of conformational diversity. In the present review we will discuss a novel view of binding, put forward in by several research groups in the last 5 to 10 years. In the free state, protein regions displaying conformational diversity exhibit equilibria among pre-existing conformations. In the presence of a ligand, one of these conformations is stabilized, so that the ligand does not need to induce a new conformation. Upon ligand binding there is a population shift toward the bound conformational state. Conformational diversity of binding sites of several proteins has been measured and has important practical as well as thermodynamical consequences: binding sites can be mapped without prior knowledge of the ligand and also evolution of binding sites depends mostly on the free state, occurring at least partially independently of the ligand.

In-Cell NMR spectroscopy: inhibition of autologous protein expression reduces Escherichia coli lysis.

Structural studies by in-cell nuclear magnetic resonance are a developing new field of research, and their objective is to obtain structural information of proteins and other biological macromolecules in the cytoplasm of Escherichia coli cells. The major limitation of in-cell experiments is cell lysis that occurs during the experiments. In this article, we describe how inhibition of autologous expression by rifampicin at a high concentration decreases cell lysis in E. coli. We suggest that rifampicin is acting in the programmed cell death gene system MazEF, which is triggered by stress conditions and ultimately leads to cell lysis.

1H, 13C and 15N resonance assignments and second structure information of Fag s 1: Fagales allergen from Fagus sylvatica.

Fagales allergens belonging to the Bet v 1 family account responsible for the majority of spring pollinosis in the temperate climate zones in the Northern hemisphere. Among them, Fag s 1 from beech pollen is an important trigger of Fagales pollen associated allergic reactions. The protein shares high similarity with birch pollen Bet v 1, the best-characterized member of this allergen family. Of note, recent work on Bet v 1 and its homologues found in Fagales pollen demonstrated that not all allergenic members of this family have the capacity to induce allergic sensitization. Fag s 1 was shown to bind pre-existing IgE antibodies most likely primarily directed against other members of this multi-allergen family. Therefore, it is especially interesting to compare the structures of Bet v 1-like pollen allergens, which have the potential to induce allergic sensitization with allergens that are mainly cross-reactive. This in the end will help to identify allergy eliciting molecular pattern on Bet v 1-like allergens. In this work, we report the (1)H, (15)N and (13)C NMR assignment of beech pollen Fag s 1 as well as the secondary structure information based on backbone chemical shifts.

fd coat protein structure in membrane environments: structural dynamics of the loop between the hydrophobic trans-membrane helix and the amphipathic in-plane helix.

By performing multidimensional solution NMR experiments on micelle samples it was possible to determine the structure of the membrane-bound form of fd coat protein based on short-range distance and dihedral angle constraints using distance geometry and simulated annealing calculations. Its dynamics were described by 15N relaxation measurements (T1, T2, heteronuclear nuclear Overhauser enhancement (NOE)) fitted with the Lipari-Szabo model-free formalism adapted for the transmembrane and in-plane helices of a membrane protein. The overall correlation time of the protein in micelles was found to be approximately 9 ns, and the local motion of each backbone N-H vector was described by an order parameter and an effective correlation time. The 50 residue protein has an amphipathic alpha-helix (residues 7 to 16) and a hydrophobic alpha-helix (residues 27 to 44), which were found to be approximately perpendicular on the basis of NOEs in the residues that connect the two helices. The residues connecting the helices are of particular interest in membrane proteins, and in this case the loop consists of two turns. The relaxation data show the presence of an extra motion in the amphipathic alpha-helix on the nanosecond timescale and additional flexibility of several residues in the loop connecting the two helices.

Ten Years of a National Oral Health Policy in Brazil: Innovation, Boldness, and Numerous Challenges.

Brazil is the only country in the world to propose a universal health care system with the aim of guaranteeing delivery of all levels of health care, free of charge, to a population of over 200 million inhabitants by means of a unified health system ("Sistema Único de Saúde" [SUS]). The national policy of oral health, also known as Smiling Brazil ("Brasil Sorridente"), was implemented in 2004. Oral health was designated as 1 of the 4 priority areas of the SUS, transforming oral health care in Brazil, with the objective that the SUS achieve the integrality of care envisaged at its creation. The aim of this article is to share part of this experience in order to prompt reflection about the inclusion of oral health care in other health care systems around the world. The most significant results of Smiling Brazil can be seen in 3 areas: (1) oral health epidemiological indicators, (2) financial investment and professional development, and (3) the building of an oral health care network throughout the 10 y of the policy. The "Discovery!" article presented here portrays 10 y of evolution; however, it is important to point out that this is a process undergoing construction and that the oral health care network needs to be further expanded, refined, and solidified so that over time and through changes in the political parties in power, Smiling Brazil prevails as a perennial policy and not merely an action by a single government.

Correlation between conformation and antibody binding: NMR structure of cross-reactive peptides from T. cruzi, human and L. braziliensis.

The structure of peptides corresponding to the C-terminal residues from Trypanosoma cruzi (R13), human (H13) and Leishmania braziliensis (A13) ribosomal proteins were determined using nuclear magnetic resonance. Although there is only one amino acid difference between them, the peptides present distinct structures in solution: R13 adopts a random coil conformation while H13 and A13 form a bend. Interaction of these peptides with polyclonal antibodies from chronic Chagas' disease patients and a monoclonal antibody raised against T. cruzi ribosomal P2beta protein was probed by transferred NOE. The results show that the flexibility of R13 is fundamental for the binding to the antibody.

Selectively labeling the heterologous protein in Escherichia coli for NMR studies: a strategy to speed up NMR spectroscopy.

Nuclear magnetic resonance is an important tool for high-resolution structural studies of proteins. It demands high protein concentration and high purity; however, the expression of proteins at high levels often leads to protein aggregation and the protein purification step can correspond to a high percentage of the overall time in the structural determination process. In the present article we show that the step of sample optimization can be simplified by selective labeling the heterologous protein expressed in Escherichia coli by the use of rifampicin. Yeast thioredoxin and a coix transcription factor Opaque 2 leucine zipper (LZ) were used to show the effectiveness of the protocol. The (1)H/(15)N heteronuclear correlation two-dimensional NMR spectrum (HMQC) of the selective (15)N-labeled thioredoxin without any purification is remarkably similar to the spectrum of the purified protein. The method has high yields and a good (1)H/(15)N HMQC spectrum can be obtained with 50 ml of M9 growth medium. Opaque 2 LZ, a difficult protein due to the lower expression level and high hydrophobicity, was also probed. The (15)N-edited spectrum of Opaque 2 LZ showed only the resonances of the protein of heterologous expression (Opaque 2 LZ) while the (1)H spectrum shows several other resonances from other proteins of the cell lysate. The demand for a fast methodology for structural determination is increasing with the advent of genome/proteome projects. Selective labeling the heterologous protein can speed up NMR structural studies as well as NMR-based drug screening. This methodology is especially effective for difficult proteins such as hydrophobic transcription factors, membrane proteins, and others.

The toxic effects of Coleus barbatus B. on the different periods of pregnancy in rats.

Extracts of Coleus barbatus B. have been used in folk medicine to interrupt pregnancy. In order to evaluate if this plant interferes with embryo implantation or with the normal development of the concepts, pregnant Wistar rats were treated with increasing doses (220, 440 and 880 mg/kg per day) of a hydroalcoholic extract of C. barbatus. The rats received the extract by gavage from days 0 to 5 of pregnancy (preimplantation period) or 6 to 15 (organogenic period). Control groups received distilled water during the same periods. The animals were killed at term for the evaluation of maternal and fetal parameters. The results showed that the treatment with 880 mg/kg per day of the extract of C. barbatus before embryo implantation caused delayed fetal development and an anti-implantation effect, which justifies the popular use of this extract with abortive purposes. After embryo implantation delayed development associated with maternal toxicity was observed in the fetuses of the group which received 880 mg/kg per day.