The SC10-RNase promoter displays changes in DNA methylation patterns through pistil development in self-incompatible Nicotiana alata.

In Solanaceae, self-incompatibility is a genetic mechanism that prevents endogamy in plant populations. Expression of the S-determinants, S-RNase, and SLF, is tightly regulated during pistil and pollen development. However, the molecular mechanism of gene expression regulation in S-RNase-based self-incompatibility systems must be better understood. Here, we identified a 1.3 Kbp sequence upstream to the coding region of the functional SC10-RNase allele from the self-incompatible Nicotiana alata, which directs SC10-RNase expression in mature pistils. This SC10-RNase promoter includes a 300 bp region with minimal elements that sustain the SC10-RNase expression. Likewise, a fragment of a transposable element from the Gypsy family of retrotransposons is also present at the -320 bp position. Nevertheless, its presence does not affect the expression of the SC10-RNase in mature pistils. Additionally, we determined that the SC10-RNase promoter undergoes different DNA methylation states during pistil development, being the mCHH methylation context the most frequent close to the transcription start site at pistil maturity. We hypothesized that the Gypsy element at the SC10-RNase promoter might contribute to the DNA methylation remodeling on the three sequence contexts analyzed here. We propose that mCHH methylation enrichment and other regulatory elements in the S-RNase coding region regulate the specific and abundant SC10-RNase expression in mature pistils in N. alata.

Energy and mineral metabolism of peripartum mares and foals of the Quarter Horse breed.

BACKGROUND: Several studies have demonstrated variations in peripartum blood biochemical analytes used to monitor the health status of mares and their foals of different breeds. OBJECTIVES: We aimed to characterize the physiologic changes of peripartum energy and mineral metabolism in healthy Quarter Horse mares and their neonatal foals. METHODS: Blood samples were initially collected from 17 mares on days 60 and 30 prepartum, and from mares and their foals on the day of parturition (after colostrum ingestion), and on days 15, 30, and 60 postpartum. The serum concentrations of calcium, phosphorus, magnesium, total cholesterol (T-Cho), triglyceride, beta-hydroxybutyrate (BHBA), and nonesterified fatty acids (NEFA) were measured using commercial kits. Statistical differences were analyzed using one-way ANOVA followed by Tukey's test (parametric variables) or the Kruskal-Wallis test followed by Dunn's multiple comparison test (nonparametric variables). RESULTS: Fourteen of 17 healthy Quarter Horse mares and their neonatal foals remained in the study until termination. Serum BHBA, T-Cho, and calcium concentrations in mares showed significant differences during the observation period. Serum BHBA concentrations increased from 60 days prepartum to 60 days postpartum. Postpartum serum T-Cho and calcium concentrations significantly decreased until day-30 postpartum. In the foals, NEFA, T-Cho, calcium, and magnesium concentrations significantly differed between sampling times. In the foals, serum NEFA concentrations decreased from parturition until day-60 postpartum, while serum T-Cho, calcium, and magnesium concentrations were lowest at day-30 postpartum. CONCLUSIONS: The temporal changes observed in the blood biochemical analytes of this study could be used to improve the clinical evaluation of periparturient mare and neonatal foal Quarter Horses.

Hypnotic intervention in people with fibromyalgia: A randomized controlled trial.

Fibromyalgia affects the quality of life of the patients, as well as their family. It also affects their social, labor, physical, and psychological dynamics. We aimed to evaluate the effectiveness of audio-recorded hypnosis in ameliorating fibromyalgia symptoms. We enrolled 97 individuals with fibromyalgia (mean age: 45 years) and randomly distributed them to two groups (48 in the experimental group and 47 in the control group). Individuals in both groups maintained their standard pharmacological treatment and continued their usual physical or psychological activities. The experimental group received an audio-recorded hypnosis intervention in the first session; subsequently, they received another audio hypnosis session to use for daily practice for a month. We evaluated the pre- and post-intervention pain intensity, pain interference, fatigue intensity, fatigue interference, depressive symptomatology, and satisfaction with life. We found that the self-administered audio-recorded hypnotic intervention significantly decreased the intensity and interference of pain and fatigue, as well as the depressive symptomatology. Audio-recorded clinical hypnosis techniques could provide an effective, practical, and economical alternative for reducing fibromyalgia-related symptoms.

NaTrxh is an essential protein for pollen rejection in Nicotiana by increasing S-RNase activity.

In self-incompatible Solanaceae, the pistil protein S-RNase contributes to S-specific pollen rejection in conspecific crosses, as well as to rejecting pollen from foreign species or whole clades. However, S-RNase alone is not sufficient for either type of pollen rejection. We describe a thioredoxin (Trx) type h from Nicotiana alata, NaTrxh, which interacts with and reduces S-RNase in vitro. Here, we show that expressing a redox-inactive mutant, NaTrxhSS , suppresses both S-specific pollen rejection and rejection of pollen from Nicotiana plumbaginifolia. Biochemical experiments provide evidence that NaTrxh specifically reduces the Cys155 -Cys185 disulphide bond of SC10 -Rnase, resulting in a significant increase of its ribonuclease activity. This reduction and increase in S-RNase activity by NaTrxh helps to explain why S-RNase alone could be insufficient for pollen rejection.

Brief mindfulness, mental health, and cognitive processes: A randomized controlled trial.

The present study evaluated the impact of a brief mindfulness training program on emotional regulation, cognitive rumination, psychological well-being, and depressive symptoms. This is an experimental study, through a randomized controlled trial. A total of 103 people enrolled on a voluntary basis, of whom 51 were randomly selected to participate in the experimental group and 52 in the control group without intervention (on the waiting list). It is established that a brief, four-session mindfulness training program significantly increases psychological well-being and significantly reduces the symptoms associated with depression; there was a decrease in emotional suppression and in intrusive and deliberate rumination, and an increase in cognitive reappraisal. Implications of this brief mindfulness program on the symptoms associated with depression, psychological well-being, emotional regulation, and cognitive rumination are analyzed, as are the scope and limitations of the study.

Comparative development of staminate and pistillate flowers in the dioecious cactus Opuntia robusta.

KEY MESSAGE: PCD role in unisexual flowers. The developmental processes underlying the transition from hermaphroditism to unisexuality are key to understanding variation and evolution of floral structure and function. A detailed examination of the cytological and histological patterns involved in pollen and ovule development of staminate and pistillate flowers in the dioecious Opuntia robusta was undertaken, and the potential involvement of programmed cell death in the abortion of the sex whorls was explored. Flowers initiated development as hermaphrodites and became functionally unisexual by anthesis. Female individuals have pistillate flowers with a conspicuous stigma, functional ovary, collapsed stamens and no pollen grains. Male individuals have staminate flowers, with large yellow anthers, abundant pollen grains, underdeveloped stigma, style and an ovary that rarely produced ovules. In pistillate flowers, anther abortion resulted from the premature degradation of the tapetum by PCD, followed by irregular deposition of callose wall around the microsporocytes, and finally by microspore degradation. In staminate flowers, the stigma could support pollen germination; however, the ovaries were reduced, with evidence of placental arrest and ovule abortion through PCD, when ovules were present. We demonstrate that PCD is recruited in both pistillate and staminate flower development; however, it occurs at different times of floral development. This study contributes to the understanding of the nature of the O. robusta breeding system and identifies developmental landmarks that contribute to sexual determination in Cactaceae.

Clinical Hypnosis and Music In Breast Biopsy:A Randomized Clinical Trial.

A randomized clinical study was conducted to evaluate the effects on anxiety, depression, stress and optimism levels of an audio-recorded clinical hypnosis intervention and a music session and compare them with a control group in women scheduled for breast biopsy. We analyzed the data of 170 patients with an average age of 47 years, who were randomly assigned to each of the groups. The psychosocial variables were measured in three moments: baseline, which corresponds to the period before the intervention with hypnosis, music or waiting in the room before biopsy; a second measurement after the interventions and a third measurement after the breast biopsy procedure was finished. The results showed a statistically significant reduction in the stress (p < .001, η2p = .06); pain, (p < .01, η2p = .04); anxiety, (p < .001, ƞ2p = .07) and depression, (p < .001, ƞ2p = .05) in hypnosis and music groups compared with the control group. Before biopsy, hypnosis decrease significantly pain and depression levels compared with music, but after biopsy there were no differences between both groups. It is recommended to use audio-recorded hypnosis and music interventions to reduce physical and emotional discomfort during the biopsy procedure and to improve the quality of life of patients with suspected breast cancer.

Screening for selective anticancer activity of plants from Grazalema Natural Park, Spain.

Since plants are an important source of anticancer drugs, we have carried out a random screening for selective anticancer activity of 57 extracts from 45 plants collected in Grazalema Natural Park, an area in the South of Spain of high plant diversity and endemism. Using lung cancer cells (A549) and lung non-malignant cells (MRC-5), we found that several extracts were more cytotoxic and selective against the cancer cells than the standard anticancer agent cisplatin. Five active extracts were further tested in cancer and normal cell lines from other tissues, including three skin cell lines with increasing degree of malignancy. An extract from the leaves of Daphne laureola L. (Thymelaeaceae) showed a striking potency and selectivity on lung cancer cells and leukemia cells; the IC50 values against these cancer cells were approximately 10,000-fold lower than against the normal cells. Daphnane-type diterpene orthoesters may be responsible for this highly selective anticancer activity.

Evaluation of an inexpensive needle test for the diagnosis of phosphorus deficiency and management of phosphorus supplementation for cattle: A multiple case study.

The needle test (NT) is a point-of-care test developed in Brazil to evaluate the Phosphorus (P) status in cattle. Based on bone resistance, the NT is a very inexpensive method which allows the diagnosis of any degree of P deficiency in a fast and simple way in vivo and directly on farm. The NT measures three levels of resistance in the transverse process (TP) of the lumbar vertebrae: a) TP that are impenetrable and result in warping of the needle (P healthy animals); b) TP offering some resistance to the penetration (animals with subclinical P deficiency); and c) TP which has minimal resistance to penetration (clinical P deficiency). This manuscript presents results from a series of case studies to evaluate the hypothesis that the NT could be used to assess P status in cattle and assesses the usefulness of results to support decision making on mineral supplementation strategies for grazing cattle. The NT was able to detect the changes in the resistance patterns of the TP, as there was reduction or elevation of P levels in the mineral mixtures. The NT was useful to assist in decision-making for adoption of mineral supplementation strategies better suited for each farm, helping farmers to save money and avoid unnecessary waste of P.

Bio-economic evaluation of a reduced phosphorus supplementation strategy for a cow-calf system in Brazil: a case study.

Fertility, weight of calves at weaning, and the economic aspects of a breeding herd receiving mineral supplements containing 75 or 12.5 g of phosphorus (P)/kg were measured from 2013 to 2016. No differences in reproduction parameters or weight at weaning were found before and after the adoption of the new scheme of mineral supplementation. Before the study, the annual cost with the formula containing more P was equivalent to 29.3 weaned beef calves; after the P reduction, the annual cost was equivalent to 2.2 to 6.8 weaned calves. After 3 years of supplementation with 12.5 g P/kg no signs of P deficiency were observed. The clinical-nutritional diagnosis of the herd indicated no cause-effect of P content of mineral supplements upon fertility or performance of healthy cows, demonstranting that the adequate forage allowance was enough to meet most P required by the cows.

Preclinical evaluation of an mRNA HIV vaccine combining rationally selected antigenic sequences and adjuvant signals (HTI-TriMix).

BACKGROUND: The development of a prophylactic vaccine against HIV-1 has so far not been successful. Therefore, attention has shifted more and more toward the development of novel therapeutic vaccines. Here, we evaluated a new mRNA-based therapeutic vaccine against HIV-1-encoding activation signals (TriMix: CD40L + CD70 + caTLR4) combined with rationally selected antigenic sequences [HIVACAT T-cell immunogen (HTI)] sequence: comprises 16 joined fragments from Gag, Pol, Vif, and Nef). METHODS: For this purpose, peripheral blood mononuclear cells from HIV-1-infected individuals on cART, lymph node explants from noninfected humans, and splenocytes from immunized mice were collected and several immune functions were measured. RESULTS: Electroporation of immature monocyte-derived dendritic cells from HIV-infected patients with mRNA encoding HTI + TriMix potently activated dendritic cells which resulted in upregulation of maturation markers and cytokine production and T-cell stimulation, as evidenced by enhanced proliferation and cytokine secretion (IFN-γ). Responses were HIV specific and were predominantly targeted against the sequences included in HTI. These findings were confirmed in human lymph node explants exposed to HTI + TriMix mRNA. Intranodal immunizations with HTI mRNA in a mouse model increased antigen-specific cytotoxic T-lymphocyte responses. The addition of TriMix further enhanced cytotoxic responses. CONCLUSION: Our results suggest that uptake of mRNA, encoding strong activation signals and a potent HIV antigen, confers a T-cell stimulatory capacity to dendritic cells and enhances their ability to stimulate antigen-specific immunity. These findings may pave the way for therapeutic HIV vaccine strategies based on antigen-encoding RNA to specifically target antigen-presenting cells.

Development and physical evaluation of Maytenus ilicifolia effervescent granules using factorial design

The medicinal plant Maytenus ilicifolia is a commonly used phytomedicine for the treatment of gastritis. The high dose required and low density of these extracts make necessary a daily intake of several capsules, hindering adherence to the medication. The purpose of this work was to develop a suitable dosage form for the administration of Maytenus ilicifolia using effervescent granules. A 23 factorial design was used to study the physical characteristics of the granules (particle size distribution, repose angle, Carr index, scanning electron microscopy and disintegration time). Moisture stability was also determined. According to the experimental design, granule size is the most important factor in determining the flow characteristics of effervescent granules. In turn, the disintegration time is controlled by the content of sodium bicarbonate present in the effervescent mixture as well as the granule size. The stability of formulations when exposed to moisture is strongly influenced by the percentage of effervescent mixture present in the vegetal granules. Precautions in handling and storage should be taken to ensure the stability of these preparations. The effervescent granules produced from Maytenus ilicifolia met the pharmacopoeial quality parameters, with appropriate mechanical and physical characteristics and proved to be a promising vehicle for plant extracts.

A planta medicinal Maytenus ilicifolia é comumente empregada como fitoterápico no tratamento da gastrite. As elevadas doses requeridas e a baixa densidade dos extratos dessa planta levam à ingestão diária de várias cápsulas do medicamento, dificultando a adesão ao tratamento. A proposta desse trabalho foi desenvolver uma forma farmacêutica adequada para administração de Maytenus ilicifolia usando granulados efervescentes. Um desenho fatorial 23 foi empregado para estudar as características físicas dos granulados (distribuição dos tamanhos de partícula, ângulo de repouso, índice de Carr, microscopia eletrônica de varredura e tempo de desintegração). A higroscopicidade das preparações também foi estudada. De acordo com o desenho experimental, o tamanho de partícula é o fator mais importante para a determinação das características de fluxo dos granulados efervescentes. Em contrapartida, o tempo de desintegração é controlado pelo conteúdo de bicarbonato de sódio presente na mistura efervescente, assim como pelo tamanho do granulado. A estabilidade das formulações quando expostas à umidade é fortemente influenciada pelo percentual de mistura efervescente presente nos granulados. Precauções de manipulação e armazenamento devem ser tomadas para garantir a estabilidade dessas preparações. Os granulados efervescentes produzidos com Maytenus ilicifolia cumprem os requisitos farmacopeicos de qualidade, com adequadas características físicas e mecânicas, provando ser um veículo promissor para extratos vegetais.

NaStEP: a proteinase inhibitor essential to self-incompatibility and a positive regulator of HT-B stability in Nicotiana alata pollen tubes.

In Solanaceae, the self-incompatibility S-RNase and S-locus F-box interactions define self-pollen recognition and rejection in an S-specific manner. This interaction triggers a cascade of events involving other gene products unlinked to the S-locus that are crucial to the self-incompatibility response. To date, two essential pistil-modifier genes, 120K and High Top-Band (HT-B), have been identified in Nicotiana species. However, biochemistry and genetics indicate that additional modifier genes are required. We recently reported a Kunitz-type proteinase inhibitor, named NaStEP (for Nicotiana alata Stigma-Expressed Protein), that is highly expressed in the stigmas of self-incompatible Nicotiana species. Here, we report the proteinase inhibitor activity of NaStEP. NaStEP is taken up by both compatible and incompatible pollen tubes, but its suppression in Nicotiana spp. transgenic plants disrupts S-specific pollen rejection; therefore, NaStEP is a novel pistil-modifier gene. Furthermore, HT-B levels within the pollen tubes are reduced when NaStEP-suppressed pistils are pollinated with either compatible or incompatible pollen. In wild-type self-incompatible N. alata, in contrast, HT-B degradation occurs preferentially in compatible pollinations. Taken together, these data show that the presence of NaStEP is required for the stability of HT-B inside pollen tubes during the rejection response, but the underlying mechanism is currently unknown.

Compatibility and incompatibility in S-RNase-based systems.

BACKGROUND: S-RNase-based self-incompatibility (SI) occurs in the Solanaceae, Rosaceae and Plantaginaceae. In all three families, compatibility is controlled by a polymorphic S-locus encoding at least two genes. S-RNases determine the specificity of pollen rejection in the pistil, and S-locus F-box proteins fulfill this function in pollen. S-RNases are thought to function as S-specific cytotoxins as well as recognition proteins. Thus, incompatibility results from the cytotoxic activity of S-RNase, while compatible pollen tubes evade S-RNase cytotoxicity. SCOPE: The S-specificity determinants are known, but many questions remain. In this review, the genetics of SI are introduced and the characteristics of S-RNases and pollen F-box proteins are briefly described. A variety of modifier genes also required for SI are also reviewed. Mutations affecting compatibility in pollen are especially important for defining models of compatibility and incompatibility. In Solanaceae, pollen-side mutations causing breakdown in SI have been attributed to the heteroallelic pollen effect, but a mutation in Solanum chacoense may be an exception. This has been interpreted to mean that pollen incompatibility is the default condition unless the S-locus F-box protein confers resistance to S-RNase. In Prunus, however, S-locus F-box protein gene mutations clearly cause compatibility. CONCLUSIONS: Two alternative mechanisms have been proposed to explain compatibility and incompatibility: compatibility is explained either as a result of either degradation of non-self S-RNase or by its compartmentalization so that it does not have access to the pollen tube cytoplasm. These models are not necessarily mutually exclusive, but each makes different predictions about whether pollen compatibility or incompatibility is the default. As more factors required for SI are identified and characterized, it will be possible to determine the role each process plays in S-RNase-based SI.

Chemical composition of essential oil of Senecio coincyi, an endemic species of the Central Iberian Peninsula.

The essential oil has been studied of leaves of Senecio coincyi Rouy, an endemic species of Spain restricted to a very small area of the Central Iberian Peninsula. Samples from five locations were obtained by hydrodistillation and extraction and analysed by gas chromatography-mass spectrometry. The main compound was 1-tridecene (28.1 +/- 8.5%). The presence of unsaturated hydrocarbons (1-undecene, 1-dodecene and 1-tridecene) seems to indicate a chemotaxonomic relationship between Senecio coincyi and S. congestus.

[Linear endobronchial ultrasound as the initial diagnostic tool in patients with indications of mediastinal disease]. / La ultrasonografía endobronquial lineal como instrumento de diagnóstico inicial en el paciente con ocupación mediastínica.

BACKGROUND AND OBJECTIVE: Linear endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) has proven useful for sampling mediastinal masses and nodes and for staging lung cancer. The aim of this study was to assess the usefulness of this diagnostic tool in patients with indications of mediastinal disease that could not be diagnosed by noninvasive methods or white light bronchoscopy. PATIENTS AND METHODS: All patients undergoing linear EBUS-TBNA for the diagnosis of mediastinal masses and/or adenopathy at our endoscopy unit were included in the study. Diagnoses obtained by linear EBUS-TBNA or any surgical technique performed after a nondiagnostic EBUS-TBNA were considered as final. RESULTS: In the study population of 128 patients with a mean (SD) age of 62.0 (11.2) years, a total of 294 TBNAs were performed on 12 masses and 282 nodes. Satisfactory samples were obtained in 11 cases (91.7%) from masses and in 233 cases (82.6%) from nodes. Linear EBUS-TBNA was diagnostic, obviating the need for mediastinoscopy in 115 patients (diagnostic sensitivity, 89.8%). The technique confirmed the diagnosis in 85 of the 94 patients with cancer (90.4%), in 8 of the 10 patients with tuberculosis (80.0%), and in the 5 patients with sarcoidosis. CONCLUSIONS: Linear EBUS-TBNA is a useful diagnostic tool in patients with mediastinal disease for whom a pathologic diagnosis is not achieved by noninvasive methods or white light bronchoscopy.

Compartmentalization of S-RNase and HT-B degradation in self-incompatible Nicotiana.

Pollen-pistil interactions are crucial for controlling plant mating. For example, S-RNase-based self-incompatibility prevents inbreeding in diverse angiosperm species. S-RNases are thought to function as specific cytotoxins that inhibit pollen that has an S-haplotype that matches one of those in the pistil. Thus, pollen and pistil factors interact to prevent mating between closely related individuals. Other pistil factors, such as HT-B, 4936-factor and the 120 kDa glycoprotein, are also required for pollen rejection but do not contribute to S-haplotype-specificity per se. Here we show that S-RNase is taken up and sorted to a vacuolar compartment in the pollen tubes. Antibodies to the 120 kDa glycoprotein label the compartment membrane. When the pistil does not express HT-B or 4936-factor, S-RNase remains sequestered, unable to cause rejection. Similarly, in wild-type pistils, compatible pollen tubes degrade HT-B and sequester S-RNase. We suggest that S-RNase trafficking and the stability of HT-B are central to S-specific pollen rejection.

Stylar glycoproteins bind to S-RNase in vitro.

S-RNases determine the specificity of S-specific pollen rejection in self-incompatible plants of the Solanaceae, Rosaceae, and Scrophulariaceae. They are also implicated in at least two distinct types of unilateral interspecific incompatibility in Nicotiana. However, S-RNase itself is not sufficient for most types of pollen rejection, and evidence for its direct interaction with pollen tubes is limited. Thus, non-S-RNase factors also are required for pollen rejection. As one approach to identifying such factors, we tested whether SC10-RNase from Nicotiana alata would bind to other stylar proteins in vitro. SC10-RNase was immobilized on Affi-gel, and binding proteins were analyzed by SDS-PAGE and immunoblotting. In addition to SC10-RNase and a small protein similar to lily chemocyanin, the most prominent binding proteins include NaTTS, 120K, and NaPELPIII, these latter three being arabinogalactan proteins previously shown to interact directly with pollen tubes. We also show that SC10-RNase and these glycoproteins migrate as a complex in a native PAGE system. Our hypothesis is that S-RNase forms a complex with these glycoproteins in the stylar ECM, that the glycoproteins interact directly with the pollen tubes and thus that the initial interaction between the pollen tube and S-RNase is indirect.

S-RNase complexes and pollen rejection.

Biochemical interactions between the pollen and the pistil allow plants fine control over fertilization. S-RNase-based pollen rejection is among the most widespread and best understood of these interactions. At least three plant families have S-RNase-based self-incompatibility (SI) systems, and S-RNases have also been implicated in interspecific pollen rejection. Although S-RNases determine the specificity of SI, other genes are required for the pollen rejection system to function. Progress is being made toward identifying these non-S-RNase factors. HT-protein, first identified as a non-S-RNase factor that was required for SI in Nicotiana alata, has now been implicated in other species as well. In addition, several pistil proteins bind to S-RNase in vitro. One hypothesis is that S-RNase forms a complex with these proteins in vivo that is the active form of S-RNase in pollen rejection.