Acetogenins-Rich Fractions of Annona coriacea Suppress Human Glioblastoma Viability and Migration by Regulating Necroptosis and MMP-2 Activity In Vitro.

Glioblastoma (GBM) is an incurable primary brain tumor with a poor prognosis. Resection, radiation therapy, and temozolomide (TMZ) are insufficient to increase survival, making the treatment limited. Thus, the search for more effective and specific treatments is essential, making plants a promising source for elucidating new anti-glioblastoma compounds. Accordingly, this study investigated the effects of four fractions of hexane and ethyl acetate extract of Annona coriacea Mart., enriched with acetogenins, against GBM cell lines. All four fractions were selectively cytotoxic to GBM cells when compared to TMZ. Moreover, A. coriacea fractions delayed cell migration; reduced cytoplasmic projections, the metalloproteinase 2 (MMP-2) activity; and induced morphological changes characteristic of necroptosis, possibly correlated with the increase in receptor-interacting protein kinase 1 and 3 (RIP-1 and RIP-3), apoptosis-inducing factor (AIF), and the non-activation of cleaved caspase 8. The present findings reinforce that fractions of A. coriacea Mart. should be considered for more studies focusing treatment of GBM.

Correlation of H3K9ac and H4K12ac With Cell Proliferation Marker Ki-67 in Oral Leukoplakia: An Immunohistochemical Study.

This study aimed to analyze the immunohistochemical expression of H3K9ac and H4K12ac in oral leukoplakia (OL) and its association with cell proliferation marker Ki-67 and clinicopathologic data. Paraffin-embedded, formalin-fixed tissue samples from 50 OLs and 15 fragments of the normal oral mucosa (NOM) were submitted to immunohistochemical assay using the streptavidin-biotin-peroxidase method. Quantitative analysis of the antigen-antibody reaction was performed by obtaining integrated optical density (IOD) and the percentage of positive nuclei (PPN) with ImageJ software. OL samples presented higher PPN ( P =0.02) and lower IOD values ( P =0.007) for H4K12ac in comparison to NOM. The area under the receiver operating characteristic curve for PPN and IOD values of H4K12ac immunostaining were 0.70 ( P =0.02) and 0.73 ( P =0.007), respectively. No differences were found between OL and NOM for H3K9ac. Cell proliferation marker Ki-67 had a positive correlation with PPN ( P <0.0001) and IOD ( P =0.0007) for H3K9ac expression and with IOD values ( P =0.002) for H4K12ac expression. The present findings suggest that alterations in the acetylation pattern of H4K12 occur in the early stages of oral carcinogenesis and that both H3K9ac and H4K12ac might have a role in the regulation of epithelial cell proliferation of OL.

Prévalence des hémorragies digestives au cours des vagues de COVID-19 au service de Réanimation Chirurgicale du CHU Joseph Ravoahangy Andrianavalona, Antananarivo, Madagascar / Prevalence of gastrointestinal bleeding during the waves of COVID -19 in the surgical intensive care unit of the CHU Joseph Ravoahangy Andrianavalona,Antananarivo, Madagascar

Introduction : La CoViD-19 est une maladie à « plusieurs visages ¼ qui peut affecter tous les systèmes. La survenue d'hémorragies digestives fait partie des manifestations de cette maladie. L'objectif de cette étude est de présenter les cas d'hémorragies digestives chez des patients infectés par le SARS-CoV-2. Matériels et Méthodes : Une étude rétrospective couvrant la période correspondant aux deux vagues de CoViD-19 à Antananarivo (Madagascar) a été réalisée, plus particulièrement au service de Réanimation Chirurgicale du Centre Hospitalier Universitaire Joseph Ravoahangy Andrianavalona. Résultats : Huit sur 101 patients de 51 à 81 ans, hospitalisés pour CoViD-19, ont présenté une hémorragie digestive dont les manifestations allaient de l'hématémèse au méléna ou une association de ces deux manifestations hémorragiques. Ces patients ont été traités par, entre autres une anticoagulation et une corticothérapie, comme défini dans le protocole national de prise en charge de la CoViD-19, avant l'épisode hémorragique. Aucun patient n'a présenté d'état de choc, l'indice de choc allait de 0,5 à 0,9. Deux patients ont pu bénéficier d'une fibroscopie digestive haute. Le score de Glasgow Blatchford variait de 6 à 13. Parmi ces huit patients, quatre sont décédés. Conclusion : Lors de la prise en charge de la CoViD-19, au vu des manifestations thrombotiques surtout, il faut procéder à une protection au niveau digestif lorsqu'une anticoagulation à titre curatif doit être réalisée. Également cette protection digestive doit être effectuée au-devant de la corticothérapie, laquelle entre dans le cadre du traitement de la CoViD-19. Tout cela pour minimiser le risque de survenue de saignement gastro-intestinal .

Background: CoViD-19 is a "many-faced" disease that can affect all the body organism. The occurrence of gastrointestinal bleeding is one of the manifestations of this disease. The aim of this study was to present cases of gastrointestinal bleeding in patients infected with SARS-CoV-2. Materials and Methods: A retrospective study covering the period corresponding to the two waves of CoViD-19 in Antananarivo (Madagascar) was carried out, more particularly in the Surgical Intensive Care Unit of the University Hospital Joseph Ravoahangy Andrianavalona. Results: Eight out of 101 patients aged 51 to 81, hospitalized for CoViD-19, presented gastrointestinal bleeding, with hematemesis or melena or a combination of these two bleeding manifestations. These patients were treated, among other coagulation and corticosteroid therapy as defined in the national protocol for the management of CoViD-19 before the bleeding episode. None of the patients presented with a shock; the shock index ranged from 0.5 to 0.9. Two patients were able to benefit from an upper digestive fibroscopy. The Glasgow Blatchford score ranged from 6 to 13. Of these eight patients, four died. Conclusion: During the management of CoViD-19, the thrombotic manifestations are treated with curative anticoagulation must be performed, which can cause digestive bleeding. Also, in front of the corticosteroid therapy which is part of the treatment of CoViD-19, also digestive protection must be carried out to minimize the risk of occurrence of gastrointestinal bleeding.

Can the exposure system adopted influence the results of the atrazine toxicity in hepatic tissue of fish?

The widespread use of atrazine, a herbicide used to control weeds, has contributed to the increased contamination of aquatic environments. To assess the toxicological effects of a xenobiotic on a nontarget organism in the laboratory, different models of toxicological exposure systems have been widely used. Therefore, the aim of this study was to evaluate and compare the action of sublethal concentrations of atrazine on the hepatic histology of Oreochromis niloticus, considering two models of exposure: static (where atrazine was only added once) and semi-static (where atrazine was periodically renewed). Fish were exposed to a concentration of 2 ppm atrazine for 15 days, which was verified by high-performance liquid chromatography. The livers were stained with hematoxylin and eosin and histopathological data were collected. In addition, they were submitted to immunohistochemistry for inducible nitric oxide synthase (iNOS). A maximum variation of 45% (static) and 12.5% (semi-static) was observed between the observed and nominal atrazine concentration. Nuclear and cytoplasmic changes were observed in both experimental models. Hepatocytes from the livers of the static system showed a degenerative appearance, while in the semi-static system, intense cytoplasmic vacuolization and necrosis were observed. iNOS positive cells were identified only in macrophages in the hepatocytes of fish in the semi-static system. These results directly showed how the choice of exposure system can influence the results of toxicological tests. However, future analysis investigating the by-products and nitrogen products should be carried out since the histopathological findings revealed the possibility of these compounds serving as secondary contamination routes.

21-Benzylidene digoxin decreases proliferation by inhibiting the EGFR/ERK signaling pathway and induces apoptosis in HeLa cells.

Cardiotonic steroids (CTS) are agents traditionally known for their capacity to bind to the Na,K-ATPase (NKA), affecting the ion transport and the contraction of the heart. Natural CTS have been shown to also have effects on cell signaling pathways. With the goal of developing a new CTS derivative, we synthesized a new digoxin derivative, 21-benzylidene digoxin (21-BD). Previously, we have shown that this compound binds to NKA and has cytotoxic actions on cancer, but not on normal cells. Here, we further studied the mechanisms of actions of 21-BD. Working with HeLa cells, we found that 21-BD decreases the basal, as well as the insulin stimulated proliferation. 21-BD reduces phosphorylation of the epidermal growth factor receptor (EGFR) and extracellular-regulated kinase (ERK), which are involved in pathways that stimulate cell proliferation. In addition, 21-BD promotes apoptosis, which is mediated by the translocation of Bax from the cytosol to mitochondria and the release of mitochondrial cytochrome c to the cytosol. 21-BD also activated caspases-8, -9 and -3, and induced the cleavage of poly (ADP-ribose) polymerase-1 (PARP-1). Altogether, these results show that the new compound that we have synthesized exerts cytotoxic actions on HeLa cells by inhibition of cell proliferation and the activation of both the extrinsic and intrinsic apoptotic pathways. These results support the relevance of the cardiotonic steroid scaffold as modulators of cell signaling pathways and potential agents for their use in cancer.

Hexane partition from Annona crassiflora Mart. promotes cytotoxity and apoptosis on human cervical cancer cell lines.

Cervical cancer is the third most commonly diagnosed tumor type and the fourth cause of cancer-related death in females. Therapeutic options for cervical cancer patients remain very limited. Annona crassiflora Mart. is used in traditional medicine as antimicrobial and antineoplastic agent. However, little is known about its antitumoral properties. In this study the antineoplastic effect of crude extract and derived partitions from A. crassiflora Mart in cervical cancer cell lines was evaluated. The crude extract significantly alters cell viability of cervical cancer cell lines as well as proliferation and migration, and induces cell death in SiHa cells. Yet, the combination of the crude extract with cisplatin leads to antagonistic effect. Importantly, the hexane partition derived from the crude extract presented cytotoxic effect both in vitro and in vivo, and initiates cell responses, such as DNA damage (H2AX activity), apoptosis via intrinsic pathway (cleavage of caspase-9, caspase-3, poly (ADP-ribose) polymerase (PARP) and mitochondrial membrane depolarization) and decreased p21 expression by ubiquitin proteasome pathway. Concluding, this work shows that hexane partition triggers several biological responses such as DNA damage and apoptosis, by intrinsic pathways, and was also able to promote a direct decrease in tumor perimeter in vivo providing a basis for further investigation on its antineoplastic activity on cervical cancer.

Design, synthesis, biological activity and structure-activity relationship studies of chalcone derivatives as potential anti-Candida agents.

Vulvovaginal candidiasis (VVC) affects millions of women around the world every year. Candida albicans is the most frequently isolated pathogen in women and its rapid ability to develop resistance to first and second line therapies has boosted the search for new and effective antifungal agents. In this study, we show the in vitro anti-Candida activity of fifteen synthetic chalcone analogs and their antifungal potential in an in vivo model of VVC. Chalcone 12 showed potent antifungal effects, being able to inhibit the growth of Candida spp. at a concentration of 15.6 µg mL-1. In addition, mechanism of action studies have indicated the ergosterol fungal membrane as the target of this compound. Despite a considerable antifungal activity, the chalcone 12 showed high cytotoxicity in kidney cells lineages. Moreover, this compound was able to reduce Candida-associated virulence, impairing yeast-hyphal transition in C. albicans. An in vivo model of VVC showed that chalcone 12 significantly reduces the fungal load. Taken together, these findings showed that the chalcone 12 is a potent anti-Candida agent in vitro beyond of contribute to improve the fungal infection in a model of CVV. However, it showed low selectivity and high toxicity, suggesting molecular modifications to minimize these proprieties.

Octopus: a platform for the virtual high-throughput screening of a pool of compounds against a set of molecular targets.

Octopus is an automated workflow management tool that is scalable for virtual high-throughput screening (vHTS). It integrates MOPAC2016, MGLTools, PyMOL, and AutoDock Vina. In contrast to other platforms, Octopus can perform docking simulations of an unlimited number of compounds into a set of molecular targets. After generating the ligands in a drawing package in the Protein Data Bank (PDB) format, Octopus can carry out geometry refinement using the semi-empirical method PM7 implemented in MOPAC2016. Docking simulations can be performed using AutoDock Vina and can utilize the Our Own Molecular Targets (OOMT) databank. Finally, the proposed software compiles the best binding energies into a standard table. Here, we describe two successful case studies that were verified by biological assay. In the first case study, the vHTS process was carried out for 22 (phenylamino)urea derivatives. The vHTS process identified a metalloprotease with the PDB code 1GKC as a molecular target for derivative LE&007. In a biological assay, compound LE&007 was found to inhibit 80% of the activity of this enzyme. In the second case study, compound Tx001 was submitted to the Octopus routine, and the results suggested that Plasmodium falciparum ATP6 (PfATP6) as a molecular target for this compound. Following an antimalarial assay, Tx001 was found to have an inhibitory concentration (IC50) of 8.2 µM against PfATP6. These successful examples illustrate the utility of this software for finding appropriate molecular targets for compounds. Hits can then be identified and optimized as new antineoplastic and antimalarial drugs. Finally, Octopus has a friendly Linux-based user interface, and is available at www.drugdiscovery.com.br . Graphical Abstract Octopus: A platform for inverse virtual screening (IVS) to search new molecular targets for drugs.

Comparative histology in the liver and spleen of three species of freshwater teleost

We describe and compare the histology of liver and spleen ofGeophagus brasiliensis (Perciformes), Hypostomus francisci (Siluriformes) and Hoplias aff. malabaricus (Characiformes), tropical freshwater fishes. InG. brasiliensisandH. aff. malabaricusthe hepatocytes were arranged in tubular form whereas in H. franciscithey cord-like. In all species, hepatocytes presented glycogen, but in G. brasiliensis and H. aff. malabaricus they showed strong stained for hemossiderin in the cytoplasm. InG. brasiliensis and H. aff. malabaricus, melanomacrophage centres (MMCs) were associated to hepatic structures and only in G. brasiliensis was observed intrahepatic exocrine pancreas. The spleen, in all species, was characterized by red and white pulp without boundary between the two regions, but only in H. francisci was recorded nodular organization in splenic parenchyma. The G. brasiliensisandH. aff. malabaricuspresented in the white pulp MMCs linked mainly to ellipsoids. Besides, we observed large MMCs in the spleen in relation to liver of G. brasiliensis and H. aff. malabaricus. In liver, highest values of reticular fibers and collagen were observed inG. brasiliensis. In spleen, highest values of reticular fibers and collagen were recorded inH. aff. malabaricusandH. francisci, respectively. Histological differences confirm the hypothesis that the phylogenetic distance is reflected in liver and spleen.(AU)

Nós descrevemos e comparamos a histologia do fígado e do baço de Geophagus brasiliensis (Perciformes), Hypostomus francisci (Siluriformes) e Hoplias aff. malabaricus (Characiformes), peixes neotropicais de água doce. Em G. brasiliensis e H. aff. malabaricus os hepatócitos organizaram-se na forma tubular enquanto que em H. francisci eles apresentaram-se como cordões celulares. Em todas as espécies, os hepatócitos apresentaram glicogênio, mas em G. brasiliensis e H. aff. malabaricus, eles mostraram forte marcação para hemossiderina no citoplasma. Em G. brasiliensis e H. aff. malabaricus, centros melanomacrofágicos (CMMs) foram associados a estruturas hepáticas e somente em G. brasiliensis foi observado pâncreas exócrino intrahepático. O baço, em todas as espécies, foi caracterizado pela polpa vermelha e branca sem limites entre as duas regiões, mas somente em H. francisci foi registrado uma organização nodular no parênquima esplênico. G. brasiliensiseH. aff. malabaricusapresentaram na polpa branca CMMs associados principalmente a elipsoides. Além disso, nós observamos CMMs grandes no baço em relação ao fígado de G. brasiliensis e de H. aff. malabaricus. No fígado, valores altos de fibras reticulares e colágeno foram observado em G. brasiliensis. No baço, valores altos de fibras reticulares e colágeno foram registrados em H. aff. malabaricuseH. francisci, respectivamente. Diferenças histológicas confirmam a hipótese que a distância filogenética está refletida no fígado e no baço.(AU)

Biological variables of Hypostomus francisci (Siluriformes: Loricariidae) from Itapecerica River, Minas Gerais State, Brazil.

Herein we determine for the first time the reproduction parameters and population structure of Hypostomus francisci in the Itapecerica River, São Francisco Basin. A total of 250 specimens was captured quarterly between March 2010 and February 2012. Body weight, total length and weight of the gonads were obtained in the laboratory. Gonad samples were submitted to histological and histochemical techniques. Females with spawning capable ovaries were used to determine the fecundity and relative fecundity. Sex ratio with 1:1.01 (female:male) was observed. Males were more numerous than females for individuals smaller than 170 mm, however the number of females was significantly greater for specimens larger than 330 mm. The length-weight relationship estimated for H. francisci indicates negative-allometric growth. Females spawning capable were observed mostly in November-December-January. Two cohorts of oocytes at a determined time evidencing the development type group-synchronic. The eggs reaching 3.4 mm and the fecundity ranged from 312-1,460 oocytes with an average of 585.81 ± 337.43 oocytes per female. The reproductive parameters and population structure of H. francisci from Itapecerica River suggested that this species showed singular reproductive tactics among congeners.

Volatile compounds of Lamiaceae exhibit a synergistic antibacterial activity with streptomycin

Bacterial infections cause thousands of deaths in the world every year. In most cases, infections are more serious because the patient is already weakened, and often, the bacteria are already resistant to the antibiotics used. Counterparting this negative scenario, the interest in medicinal plants as an alternative to the synthetic antimicrobial drugs is blossoming worldwide. In the present work, we identified the volatile compounds of ethanol extracts of Melissa officinalis, Mentha sp., Ocimum basilicum, Plectranthus barbatus, and Rosmarinus officinalis by gas chromatography/mass spectrometry (GC/MS). Also was evaluated antimicrobial activity of ethanol extracts against 6 bacteria of clinical interest, and was tested the interaction of these extracts with a commercial antibiotic streptomycin. Phytol was a compound identified in all extracts by GC/MS, being majoritary component in Plectranthus barbatus and Rosmarinus officinalis. The Gram-positive bacteria were more sensitive to ethanol extracts, and Plectranthus barbatus and Rosmarinus officinalis were the most active extracts. Ethanol extracts exhibited a synergetic effect with streptomycin. These results encourage additional studies, in order to evaluate the possibilities of using ethanol extracts of Lamiaceae family as natural source for antibacterial activity.

Volatile compounds of Lamiaceae exhibit a synergistic antibacterial activity with streptomycin.

Bacterial infections cause thousands of deaths in the world every year. In most cases, infections are more serious because the patient is already weakened, and often, the bacteria are already resistant to the antibiotics used. Counterparting this negative scenario, the interest in medicinal plants as an alternative to the synthetic antimicrobial drugs is blossoming worldwide. In the present work, we identified the volatile compounds of ethanol extracts of Melissa officinalis, Mentha sp., Ocimum basilicum, Plectranthus barbatus, and Rosmarinus officinalis by gas chromatography/mass spectrometry (GC/MS). Also was evaluated antimicrobial activity of ethanol extracts against 6 bacteria of clinical interest, and was tested the interaction of these extracts with a commercial antibiotic streptomycin. Phytol was a compound identified in all extracts by GC/MS, being majoritary component in Plectranthus barbatus and Rosmarinus officinalis. The Gram-positive bacteria were more sensitive to ethanol extracts, and Plectranthus barbatus and Rosmarinus officinalis were the most active extracts. Ethanol extracts exhibited a synergetic effect with streptomycin. These results encourage additional studies, in order to evaluate the possibilities of using ethanol extracts of Lamiaceae family as natural source for antibacterial activity.

Metallothionein immunolocalization in actinic skin nonmelanoma carcinomas.

Basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) are the most frequent skin cancer. Its pathogeny is linked to genotoxic effects of actinic radiation exposure, especially to ultraviolet wavelength. Metallothionein (MT) is a low-molecular weight protein with high affinity for heavy metal. Its intracellular function has been related to heavy metals and free-radical detoxification, although many studies linked MT to protective action against actinic mutagenesis. In other way, overexpression in malignant tumors has been related to worse prognosis. We aimed to evaluate MT immunohistochemical expression in skin cancer associated to actinic radiation. Twenty-six BCC cases, 20 SCC, and 6 normal skin fragments were investigated. Immunohistochemical assay were performed by streptavidin-biotin-peroxidase technique with standard monoclonal antibody (E9). In normal skin, immunostaining was observed in basal layer of the epithelium. In the epithelium adjacent to tumors, suprabasal layer was also intensely labeled. Mean MT immunostaining indices were 18.5+21.2% for BCC and 69.1+14.4% for SCC. This difference was statistically significant. Higher MT expression in SCC as compared with BCC suggests association with tumoral aggressiveness.