A disordered region retains the full protease inhibitor activity and the capacity to induce CD8+ T cells in vivo of the oral vaccine adjuvant U-Omp19.

U-Omp19 is a bacterial protease inhibitor from Brucella abortus that inhibits gastrointestinal and lysosomal proteases, enhancing the half-life and immunogenicity of co-delivered antigens. U-Omp19 is a novel adjuvant that is in preclinical development with various vaccine candidates. However, the molecular mechanisms by which it exerts these functions and the structural elements responsible for these activities remain unknown. In this work, a structural, biochemical, and functional characterization of U-Omp19 is presented. Dynamic features of U-Omp19 in solution by NMR and the crystal structure of its C-terminal domain are described. The protein consists of a compact C-terminal beta-barrel domain and a flexible N-terminal domain. The latter domain behaves as an intrinsically disordered protein and retains the full protease inhibitor activity against pancreatic elastase, papain and pepsin. This domain also retains the capacity to induce CD8+ T cells in vivo of U-Omp19. This information may lead to future rationale vaccine designs using U-Omp19 as an adjuvant to deliver other proteins or peptides in oral formulations against infectious diseases, as well as to design strategies to incorporate modifications in its structure that may improve its adjuvanticity.

In vitro cytotoxic potential of extracts from Aristolochia foetida Kunth against MCF-7 and bMECs cell lines.

The aim of this study was to evaluate the cytotoxic potential of Aristolochia foetida Kunth. Stems and leaves of A. foetida Kunth (Aristolochiaceae) have never been investigated pharmacologically. Recent studies of species of the Aristolochiaceae family found significant cytotoxic activities. Hexane, dichloromethane, ethyl acetate and methanol extracts were analyzed by 1H NMR and GC-MS to know the metabolites in each extract. In GC-MS analysis, the main compounds were methyl hexadecanoate (3); hexadecanoic acid (4); 2-butoxyethyl dodecanoate (9); ethyl hexadecanoate (20); methyl octadeca-9,12,15-trienoate (28) and (9Z,12Z,15Z)-octadeca-9,12,15-trienoic acid (40). The results showed a significant reduction in cell viability of the MCF-7 (breast cancer) cell line caused by organic extracts in a dose-dependent manner. The cytotoxicity activity of the dichloromethane extract from the stems (DSE) showed IC50 values of 45.9 µg/mL and the dichloromethane extract of the leaves (DLE) showed IC50 values of 47.3 µg/mL. DSE and DLE had the highest cytotoxic potential in an in vitro study against the MCF-7 cell line and non-tumor cells obtained from the bovine mammary epithelial (bMECs). DSE and DLE induced a loss in mitochondrial membrane potential (ΔΨm) and can cause cell death by apoptosis through the intrinsic pathway in the MCF-7 cell line. DSE and DLE are cytotoxic in cancer cells and cause late apoptosis. Higher concentrations of DSE and DLE are required to induce a cytotoxic effect in healthy mammary epithelial cells. This is the first report of the dichloromethane extract of A. foetida Kunth that induces late apoptosis in MCF-7 cancer cells and may be a candidate for pharmacological study against breast cancer.

Caferana seeds (Bunchosia glandulifera) as a new source of nutrients: Evaluation of the proximal composition, solvent extraction, bioactive compounds, and δ-lactam isolation.

The proximal composition, amino acid, carbohydrate, and volatile profiles of caferana (Bunchosia glandulifera) seeds flour were here assessed. Seeds were also subjected to the following extraction processes: one with pressurized ethanol (PLE) and two with ethanol + supercritical CO2 mixture at different temperatures and pressures (SC1 and SC2). Extracts were characterized in terms of caffeine, total phenolic, and δ-lactam. The characterization of caferana seed and its extracts is unprecedented in terms of carbohydrate and volatiles profiles, besides the δ-lactam identification/isolation. SC2 extract exhibited a higher caffeine (9.3 mg/g) and δ-lactam (29.4 mg/g) content, whereas the PLE extract contained a higher total phenolic amount (3.0 mgGAE/g). Caferana is regionally associated to protective effects on mental health. Its byproduct (seed) revealed to be a promising source of bioactive compounds, and a potential raw material of nutritive extracts and flours that can be incorporated into pharmaceutical, nutraceutical, cosmetic, and food products.

Toxicity evaluation of Eleutherine plicata Herb. extracts and possible cell death mechanism.

Eleutherine plicata has been shown to be a promising medicinal plant, and its activity has been associated with naphthoquinones. The present study aimed at evaluating the cytotoxicity, genotoxicity, and oral toxicity of the ethanol extract (EEEp), dichloromethane fraction (FDMEp) of E. plicata, and isoeleutherin. For the cytotoxicity evaluation, the viability test (MTT) was used. Genotoxicity was accessed through the Comet assay (alkaline version), acute and subacute oral toxicities were also evaluated. The antioxidant capacity of the samples in the wells where the cells were treated with E. plicata was evaluated. Furthermore, the participation of caspase-8 in the possible mechanism of action of isoeleutherin, eleutherin, and eleutherol was also investigated through a docking study. FDMEp and isoeleutherin were cytotoxic, with higher rates of DNA fragmentation observed for FDMEp and isoeleutherin, and all samples displayed higher antioxidant potential than the control. In the acute oral toxicity test, EEEp, FDMEp, and isoeleutherin did not cause significant clinical changes. In the subacute toxicity assay, EEEp and FDMEp also did not cause clinical, hematological, or biochemical changes. The three compounds bound similarly to caspase-8. Despite the results of cytotoxicity, in vitro studies demonstrated that the use of EEEp appears to be safe and cell death may involve its binding to caspase-8.

Determination of chemical structure and anti-Trypanosoma cruzi activity of extracts from the roots of Lonchocarpus cultratus (Vell.) A.M.G. Azevedo & H.C. Lima.

Trypanosoma cruzi is the agent of Chagas disease, an infection that affects around 8 million people worldwide. The search for new anti-T. cruzi drugs are relevant, mainly because the treatment of this disease is limited to two drugs. The objective of this study was to investigate the trypanocidal and cytotoxic activity and elucidate the chemical profile of extracts from the roots of the Lonchocarpus cultratus. Roots from L. cultratus were submitted to successive extractions with hexane, dichloromethane, and methanol, resulting in LCH, LCD, and LCM extracts, respectively. Characterization of extracts was done using 1H-RMN, 13C-RMN, CC and TLC. Treatment of T. cruzi forms (epimastigotes, trypomastigotes, and amastigotes) with crescent concentrations of LCH, LCD, and LCM was done for 72, 48, and 48 h, respectively. After this, the percentage of inhibition and IC50/LC50 were calculated. Benznidazole was used as a positive control. Murine macrophages were treated with different concentrations of both extracts for 48 h, and after, the cellular viability was determined by the MTT method and CC50 was calculated. The chalcones derricin and lonchocarpine were identified in the hexane extract, and for the first time in the genus Lonchocarpus, the presence of a dihydrolonchocarpine derivative was observed. Other chalcones such as isocordoin and erioschalcone B were detected in the dichloromethane extract. The dichloromethane extract showed higher activity against all tested forms of T. cruzi than the other two extracts, with IC50 values of 10.98, 2.42, and 0.83 µg/mL, respectively; these values are very close to those of benznidazole. Although the dichloromethane extract presented a cytotoxic effect against mammalian cells, it showed selectivity against amastigotes. The methanolic extract showed the lowest anti-T. cruzi activity but was non-toxic to peritoneal murine macrophages. Thus, the genus Lonchocarpus had demonstrated in the past action against epimastigotes forms of T. cruzi but is the first time that the activity against infective forms is showed, which leading to further studies with in vivo tests.

Alkaloids of Phaedranassa dubia (Kunth) J.F. Macbr. and Phaedranassa brevifolia Meerow (Amaryllidaceae) from Ecuador and its cholinesterase-inhibitory activity.

Alzheimer's disease is considered the most common cause of dementia and, in an increasingly aging population worldwide, the quest for treatment is a priority. Amaryllidaceae alkaloids are of main interest because of their cholinesterase inhibition potential, which is the main palliative treatment available for this disease. We evaluated the alkaloidal profile and the in vitro inhibitory activity on acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) of bulb alkaloid extract of Phaedranassa dubia and Phaedranassa brevifolia collected in Ecuador. Using gas chromatography coupled to mass spectrometry (GC-MS), we identified typical Amaryllidaceae alkaloids in these species, highlighting the presence of lycorine-type alkaloids in P. dubia and haemanthamine/crinine-type in P. brevifolia. The species P. dubia and P. brevifolia showed inhibitory activities against AChE (IC50 values of 25.48 ± 0.39 and 3.45 ± 0.29 µg.mL-1, respectively) and BuChE (IC50 values of 114.96 ± 4.94 and 58.89 ± 0.55 µg.mL-1, respectively). Computational experiments allowed us to understand the interactions of the alkaloids identified in these samples toward the active sites of AChE and BuChE. In silico, some alkaloids detected in these Amaryllidaceae species presented higher estimated binding free energy toward BuChE than galanthamine. This is the first study about the alkaloid profile and biological potential of P. brevifolia species.

Neuroinflammation and glial cell activation in mental disorders.

Mental disorders (MDs) are highly prevalent and potentially debilitating complex disorders which causes remain elusive. Looking into deeper aspects of etiology or pathophysiology underlying these diseases would be highly beneficial, as the scarce knowledge in mechanistic and molecular pathways certainly represents an important limitation. Association between MDs and inflammation/neuroinflammation has been widely discussed and accepted by many, as high levels of pro-inflammatory cytokines were reported in patients with several MDs, such as schizophrenia (SCZ), bipolar disorder (BD) and major depression disorder (MDD), among others. Correlation of pro-inflammatory markers with symptoms intensity was also reported. However, the mechanisms underlying the inflammatory dysfunctions observed in MDs are not fully understood yet. In this context, microglial dysfunction has recently emerged as a possible pivotal player, as during the neuroinflammatory response, microglia can be over-activated, and excessive production of pro-inflammatory cytokines, which can modify the kynurenine and glutamate signaling, is reported. Moreover, microglial activation also results in increased astrocyte activity and consequent glutamate release, which are both toxic to the Central Nervous System (CNS). Also, as a result of increased microglial activation in MDs, products of the kynurenine pathway were shown to be changed, influencing then the dopaminergic, serotonergic, and glutamatergic signaling pathways. Therefore, in the present review, we aim to discuss how neuroinflammation impacts on glutamate and kynurenine signaling pathways, and how they can consequently influence the monoaminergic signaling. The consequent association with MDs main symptoms is also discussed. As such, this work aims to contribute to the field by providing insights into these alternative pathways and by shedding light on potential targets that could improve the strategies for pharmacological intervention and/or treatment protocols to combat the main pharmacologically unmatched symptoms of MDs, as the SCZ.

Vanadium: History, chemistry, interactions with α-amino acids and potential therapeutic applications.

In the last 30 years, since the discovery that vanadium is a cofactor found in certain enzymes of tunicates and possibly in mammals, different vanadium-based drugs have been developed targeting to treat different pathologies. So far, the in vitro studies of the insulin mimetic, antitumor and antiparasitic activity of certain compounds of vanadium have resulted in a great boom of its inorganic and bioinorganic chemistry. Chemical speciation studies of vanadium with amino acids under controlled conditions or, even in blood plasma, are essential for the understanding of the biotransformation of e.g. vanadium antidiabetic complexes at the physiological level, providing clues of their mechanism of action. The present article carries out a bibliographical research emphaticizing the chemical speciation of the vanadium with different amino acids and reviewing also some other important aspects such as its chemistry and therapeutical applications of several vanadium complexes.

Inherited Xq13.2-q21.31 duplication in a boy with recurrent seizures and pubertal gynecomastia: Clinical, chromosomal and aCGH characterization.

We report on a 16-year-old boy with a maternally inherited ~ 18.3 Mb Xq13.2-q21.31 duplication delimited by aCGH. As previously described in patients with similar duplications, his clinical features included intellectual disability, developmental delay, speech delay, generalized hypotonia, infantile feeding difficulties, self-injurious behavior, short stature and endocrine problems. As additional findings, he presented recurrent seizures and pubertal gynecomastia. His mother was phenotypically normal and had completely skewed inactivation of the duplicated X chromosome, as most female carriers of such duplications. Five previously reported patients with partial Xq duplications presented duplication breakpoints similar to those of our patient. One of them, a fetus with multiple congenital abnormalities, had the same cytogenetic duplication breakpoint. Three of the reported patients shared many features with our proband but the other had some clinical features of the Prader-Willi syndrome. It was suggested that ATRX overexpression could be involved in the major clinical features of patients with partial Xq duplications. We propose that this gene could also be involved with the obesity of the patient with the Prader-Willi-like phenotype. Additionally, we suggest that the PCDH11X gene could be a candidate for our patient's recurrent seizures. In males, the Xq13-q21 duplication should be considered in the differential diagnosis of Prader-Willi syndrome, as previously suggested, and neuromuscular diseases, particularly mitochondriopathies.