Allophylus africanus Stem Bark Extract Modulates the Mitochondrial Apoptotic Pathway in Human Stomach Cancer Cells.

The present work aimed to detail the mechanisms elicited by Allophylus africanus P. Beauv. stem bark extract in human stomach cancer cells and to identify the bioactives underlying the cytotoxicity. MTT reduction and LDH leakage assays allowed characterizing the cytotoxic effects in AGS cells, which were further detailed by morphological analysis using phalloidin and Hoechst 33258. Proapoptotic mechanisms were elucidated through a mitochondrial membrane potential assay and by assessing the impact upon the activity of caspase-9 and -3. The extract displayed selective cytotoxicity against AGS cells. The absence of plasma membrane permeabilization, along with apoptotic body formation, suggested that pro-apoptotic effects triggered cell death. Intrinsic apoptosis pathway activation was verified, as mitochondrial membrane potential decrease and activation of caspase-9 and -3 were observed. HPLC-DAD profiling enabled the identification of two apigenin-di-C-glycosides, vicenin-2 (1) and apigenin-6-C-hexoside-8-C-pentoside (3), as well as three mono-C-glycosides-O-glycosylated derivatives, apigenin-7-O-hexoside-8-C-hexoside (2), apigenin-8-C-(2-rhamnosyl)hexoside (4) and apigenin-6-C-(2-rhamnosyl)hexoside (5). Isovitexin-2″-O-rhamnoside (5) is the main constituent, accounting for nearly 40% of the total quantifiable flavonoid content. Our results allowed us to establish the relationship between the presence of vicenin-2 and other apigenin derivatives with the contribution to the cytotoxic effects on the presented AGS cells. Our findings attest the anticancer potential of A. africanus stem bark against gastric adenocarcinoma, calling for studies to develop herbal-based products and/or the use of apigenin derivatives in chemotherapeutic drug development.

HPLC-DAD-ESI/MSn and UHPLC-ESI/QTOF/MSn characterization of polyphenols in the leaves of Neocarya macrophylla (Sabine) Prance ex F. White and cytotoxicity to gastric carcinoma cells.

Among several extracts from species from Guinea-Bissauan flora, the hydroethanol extract obtained from the leaves of gingerbread plum (Neocarya macrophylla (Sabine) Prance ex F. White.) revealed to be one of the most cytotoxic towards human gastric AGS carcinoma cells. Considering the increasing use of N. macrophylla in the food industry and the abundant biomass of agricultural wastes being generated, the identification of phenolic bioactives has been attained by HPLC-DAD-ESI/MSn and UHPLC-ESI/QTOF/MSn. Twenty-seven phenolic constituents were identified for the first time in the monotypic genus Neosartorya, 5-O-caffeoylquinic acid being detected as the major constituent (4.90 ± 0.20 mg g-1 dry extract). While 15 flavan-3-ols derivatives were determined, the extract is predominantly characterized by the occurrence of quercetin, kaempferol, apigenin and chrysoeriol glycosides. Typical apoptotic changes in gastric adenocarcinoma AGS cells upon exposure to N. macrophylla leaf extract were observed. The apoptotic cell death is mediated by the activation of the mitochondrial pathway, as loss of mitochondrial membrane potential was detected, as well as increased caspase-9 and -3 activities. The industrial relevance of this plant material, along with the data presented here on the potential anticancer effects of N. macrophylla and the efficient extraction of phenolic bioactives using water and ethanol (GRAS substance), calls for further research on the leaves as a potential functional food and/or ingredient.

Activation of caspase-3 in gastric adenocarcinoma AGS cells by Xylopia aethiopica (Dunal) A. Rich. fruit and characterization of its phenolic fingerprint by HPLC-DAD-ESI(Ion Trap)-MSn and UPLC-ESI-QTOF-MS2.

While the fruits of Xylopia aethiopica (Dunal) A. Rich. are important in African countries as a local trade product, their composition remains scarcely investigated. Phenolic fingerprint is herein delivered through HPLC-DAD-ESI(Ion Trap)-MSn and UPLC-ESI-QTOF-MS2 analysis, six cinnamoylquinic acid derivatives and twenty-four flavonoid glycosides being determined, chrysoeriol-7-O-glycosides being the main constituents. A cytotoxicity screening of twenty-eight hydroethanol extracts, obtained from a collection of Guinea-Bissauan plants, against A549 and AGS carcinoma cells, revealed the selective and potent effect towards AGS cells (IC50 = 151 × 10-3 g L-1), upon exposure to the extract from X. aethiopica fruits. Additional experiments demonstrated insignificant effect on LDH release at 151 × 10-3 g L-1, morphological analysis further suggesting induction of apoptosis. Pro-apoptotic effects were confirmed, as the extract enabled the activation of the effector caspase-3, broadening the knowledge on the anticancer mechanisms elicited by the fruits of X. aethiopica. Phenolic constituents might contribute to the cytotoxic effects, particularly via caspase-3 activation. Considering that X. aethiopica fruit is very often referred as an anticancer ingredient in Africa, but mainly the potent cytotoxicity herein recorded, our results call for additional research aiming to identify non-phenolic constituents contributing to the effects and also to further detail the anticancer mechanisms.

Anti-inflammatory properties of Xylopia aethiopica leaves: Interference with pro-inflammatory cytokines in THP-1-derived macrophages and flavonoid profiling.

ETHNOPHARMACOLOGICAL RELEVANCE: Ethnopharmacological surveys on Guinea-Bissauan flora reveal that several species are used to treat or ameliorate the symptomatology of conditions with an inflammatory background. As such, extracts obtained from a series of plants recorded in those surveys were screened for their anti-inflammatory properties, a hydroethanolic extract obtained from the leaves of Xylopia aethiopica (Dunal) A. Rich, (Annonaceae), used on the treatment of headache, muscular pain and rheumatic pain, scoring positively and being further investigated. AIM OF THE STUDY: In order to identify species with anti-inflammatory properties, extracts were screened for their ability to interfere with LPS-induced TNF-α levels. Since significant effects were recorded upon treatment with the extract of the leaves obtained from X. aethiopica, further assays were conducted to elucidate additional mechanisms underlying its anti-inflammatory potential. Since little is known on the chemical composition of the plant, we also aimed to characterise its phenolic profile. MATERIALS AND METHODS: Interference with cytokines was evaluated by ELISA assay, through the quantification of TNF-α and IL-6 levels in the culture medium collected from LPS-activated THP-1-derived-macrophages. Inhibition of 5-lipoxygenase was assessed based on the oxidation of linoleic acid to 13-hydroperoxylinoleic acid. Characterization of the phenolic profile was attained by HPLC-DAD. RESULTS: Evaluation of TNF-α levels in LPS-challenged THP-1 macrophages evidenced a significant inhibition (>90%) upon treatment with the hydroethanolic extract obtained from X. aethiopica leaves at a concentration of 500 µg/mL. Additional anti-inflammatory effects were recorded, including a significant decrease on IL-6 levels at 250 and 500 µg/mL. The extract proved to be active towards 5-LOX, leading to significant inhibition at concentrations ranging from 16 to 250 µg/mL (IC50 = 85 µg/mL). Phenolic profiling allowed the identification and quantitation of eight constituents, including caffeoylquinic acids (1-3), mono-O-glycosylated flavonols (5-8), and the mono-O-glycosyl flavone luteolin-7-O-glucoside (4). The main phenolic constituent, kaempferol-3-O-rutinoside (8), was found to significantly contribute to the anti-inflammatory effects, namely through the inhibition of 5-LOX. However, no effects on the decrease of TNF-α and IL-6 levels caused by this phenolic compound were found. CONCLUSION: The anti-inflammatory effects of X. aethiopica leaves are demonstrated experimentally, thus substantiating its use in folk Medicine. Relevantly, the observed anti-inflammatory properties can stimulate further studies in order to fully unveil the therapeutic potential of the plant, namely as a source of phenolic compounds with a significant ability to interfere with conventional inflammatory targets.

Benzoquinones from Cyperus spp. trigger IRE1α-independent and PERK-dependent ER stress in human stomach cancer cells and are novel proteasome inhibitors.

BACKGROUND: The roots and tubers of several species of the Cyperus genus are used in several parts of the world as foodstuffs and beverages. The genus is rich in several classes of quinones, however their biological properties have not been studied before. PURPOSE: We evaluated the anticancer effect of several benzoquinones isolated from the genus and described their mechanism of action towards cancer cells. METHODS: The most potent molecules were selected according to their effect upon cell viability. The mechanism of cell death was studied by using pharmacological inhibitors of caspases, caspase-3/4/9 activity assays, annexin-V/7-AAD by flow cytometry and intracellular reactive oxygen species and calcium levels through fluorescence spectroscopy. Elucidation of the involvement of distinct branches of the ER stress pathway was pursued by RT-PCR and WB for mRNA and protein expression levels, respectively, as well as pharmacological inhibitors. Proteasome inhibitory activity was assessed by using purified 20S catalytic subunit with the fluorogenic substrate Suc-Leu-Leu-Val-Tyr-AMC. RESULTS: Cytotoxicity studies against cancer cell lines showed that the human gastric cancer cell line AGS was the most susceptible, the most potent molecule, hydroxycyperaquinone, exhibiting an IC50 close to 1 µM. Morphological and biochemical traits suggested that a process of regulated cell death was taking place, which was shown to be intrinsic pathway-independent. Results indicated that benzoquinones exert their toxicity by triggering ER stress, as shown by increased expression of CHOP (mRNA and protein levels), intracellular reactive oxygen species, changes in calcium dynamics and caspase-4 activation. Proteasome inhibition by these molecules is described for the first time. CONCLUSION: Hydroxycyperaquinone is a novel sub-micromolar inhibitor of the 20S catalytic core of the 26S proteasome, causing cell death via IRE1α-independent/PERK-dependent pathways in stomach cancer cells. Its presence in products consumed orally may be of relevance for gastric tumors.

Bioenergetic cues shift FXR splicing towards FXRα2 to modulate hepatic lipolysis and fatty acid metabolism.

OBJECTIVE: Farnesoid X receptor (FXR) plays a prominent role in hepatic lipid metabolism. The FXR gene encodes four proteins with structural differences suggestive of discrete biological functions about which little is known. METHODS: We expressed each FXR variant in primary hepatocytes and evaluated global gene expression, lipid profile, and metabolic fluxes. Gene delivery of FXR variants to Fxr(-/-) mouse liver was performed to evaluate their role in vivo. The effects of fasting and physical exercise on hepatic Fxr splicing were determined. RESULTS: We show that FXR splice isoforms regulate largely different gene sets and have specific effects on hepatic metabolism. FXRα2 (but not α1) activates a broad transcriptional program in hepatocytes conducive to lipolysis, fatty acid oxidation, and ketogenesis. Consequently, FXRα2 decreases cellular lipid accumulation and improves cellular insulin signaling to AKT. FXRα2 expression in Fxr(-/-) mouse liver activates a similar gene program and robustly decreases hepatic triglyceride levels. On the other hand, FXRα1 reduces hepatic triglyceride content to a lesser extent and does so through regulation of lipogenic gene expression. Bioenergetic cues, such as fasting and exercise, dynamically regulate Fxr splicing in mouse liver to increase Fxrα2 expression. CONCLUSIONS: Our results show that the main FXR variants in human liver (α1 and α2) reduce hepatic lipid accumulation through distinct mechanisms and to different degrees. Taking this novel mechanism into account could greatly improve the pharmacological targeting and therapeutic efficacy of FXR agonists.

Effect of Calea serrata Less. n-hexane extract on acetylcholinesterase of larvae ticks and brain Wistar rats.

Acetylcholinesterase (AChE), an enzyme that hydrolyses acetylcholine (ACh) at cholinergic synapses, is a target for pesticides and its inhibition by organophosphates leads to paralysis and death of arthropods. It has been demonstrated that the n-hexane extract of Calea serrata had acaricidal activity against larvae of Rhipicephalus (Boophilus) microplus and Rhipicephalus sanguineus. The aim of the present study was to understand the mechanism of the acaricidal action of C. serrata n-hexane extract are specifically to investigate the in vitro anticholinesterase activity on larvae of R. microplus and in brain structures of male Wistar rats. The n-hexane extract significantly inhibited in vitro acetylcholinesterase activity in R. microplus larvae and rat brain structures. The results confirm that inhibition of acetylcholinesterase is a possible mechanism of action of hexane extract at C. serrata.

Acaricidal properties of the essential oil and precocene II obtained from Calea serrata (Asteraceae) on the cattle tick Rhipicephalus (Boophilus) microplus (Acari: Ixodidae).

Calea serrata Less. (Asteraceae), an endemic species of south Brazil known as "quebra-tudo", is used in Afro-Brazilian religious rituals and in folk medicine for treating liver disorders. Phytochemical studies of the n-hexane extract of this plant demonstrated the presence of precocene II, a benzopyran derivative known for its insecticidal activity. The aim of this work was to isolate this benzopyran and determine the chemical composition of the essential oil of C. serrata and further to evaluate the acaricidal activity of the essential oil and precocene II against the larvae of Rhipicephalus (Boophilus) microplus. The LC(99.9) and LC(50) values obtained with the oil, which presents precocene II and sesquiterpenes, were 3.94 µL/mL and 0.28 µL/mL, respectively. For precocene II this values were 4.25mg/mL and 1.78 mg/mL, respectively. The results indicate a synergistic interaction between the components of the oil and precocene II.

Comemoração dos 5 anos do Programa Remédio Casa UBS Brasília - M'Boi Mirim - Sul / Celebrating five years of the Program Home Remedy UBS Brasília - M'Boi Mirim - South

Em agosto de 2010 foi realizada a comemoração de 5 anos do Programa Remédio em Casa com a presença do Prefeito de São Paulo e representantes de instituições parceiras, da Coordenadoria Regional Sul, Supervisões de Saúde, demais unidades de saúde e população. A UBS Jardim Brasília foi a unidade escolhida, por ser uma unidade completa em suas ações e prevenção à saúde, com enfoque nas doenças crônicas e degenerativas. Estas ações são realizadas pelo Núcleo de Apoio à Saúde da Família - NASF e Estratégia de Saúde da Família - ESF. A Unidade tem um fluxo de atendimento bem estabelecido com a participação da equipe multiprofissional, garantia de consulta no prazo para a renovação das precrições e desenvolve vários grupos educativos como atividades corporais e artesanato (com geração de renda aos participantes). Na UBS Brasília a meta de pacientes ativos no Programa esperada para o ano de 2010 está em 65 por cento(AU)

Acaricidal activity of Calea serrata (Asteraceae) on Boophilus microplus and Rhipicephalus sanguineus.

Calea serrata (Asteraceae) is an endemic Southern Brazilian plant species used for religious and medicinal purposes. Previous study revealed the presence of chromenes, a class of natural compounds that possess insecticidal properties. This study reports the effect of the hexane extract from the aerial parts of this plant on egg hatchability, egg production and mortality rates of newly hatched larvae of the cattle tick Boophilus microplus. Larvae of Rhipicephalus sanguineus, the brown dog tick, received the same treatment. The extract was toxic to the eggs of B. microplus and to the larvae of both B. microplus and R. sanguineus.

Chemical composition and larvicidal properties of the essential oils from Drimys brasiliensis Miers (Winteraceae) on the cattle tick Rhipicephalus (Boophilus) microplus and the brown dog tick Rhipicephalus sanguineus.

The essential oil obtained from leaves and stem barks of the Southern Brazilian native Drimys brasiliensis Miers, a tree with medicinal properties, was analyzed by gas chromatography (GC) and GC/mass spectrometry (MS). The oil was characterized by sesquiterpenoids (66%), cyclocolorenone being the most abundant (30.4%), followed by bicyclogermacrene (11.8%) and alpha-gurjunene (6.0%). Laboratory tests were carried out to determine the toxicity of the essential oil on larvae of the cattle tick Rhipicephalus (Boophilus) microplus and the brown dog tick Rhipicephalus sanguineus by the larval immersion test. It was observed that the oil was lethal, killing 100% of the larvae of both ticks at the doses of 25, 12.5, and 6.25 microl/ml. The lowest dose tested, 3.125 microl/ml, was also toxic, killing 95-98% of the larvae.

Acaricidal properties of extracts from the aerial parts of Hypericum polyanthemum on the cattle tick Boophilus microplus.

Laboratory tests were carried out on larvae and adults of the cattle tick Boophilus microplus to determine the toxicity of n-hexane and crude methanolic extracts of Hypericum polyanthemum (Guttiferae) using the larval immersion test (LIT) and adult immersion test (AIT). In the AIT, the effectiveness of treatment against engorged females was assessed by measuring egg production: the n-hexane extract was found to have a small effect on the egg laying at the highest concentration (19.2% of egg-laying inhibition) whilst the crude methanolic extract did not affect the egg production. For the LIT the n-hexane extracts were highly toxic to the larvae at all the concentrations (100% of mortality). The crude methanolic extract was also toxic to the larvae at higher concentrations killing 100, 96.7, 84.7 and 52.7% at the concentrations of 50, 25, 12.5, and 6.25 mg/ml, respectively, 48 h after the immersion of the acarus.