Trypsin inhibitor from Enterolobium contortisiliquum seeds impairs Aedes aegypti development and enhances the activity of Bacillus thuringiensis toxins.

BACKGROUND: Disease vector insects are barriers for human development. The use of synthetic chemicals to control these vectors has caused damage to the environment and contributed to the arising of resistant insect populations. This has led to an increased search for plant-derived molecules with insecticidal activity or that show synergistic effects with known insecticidal substances, such as protease inhibitors. Thus, we aimed to evaluate the effect of Enterolobium contortisiliquum trypsin inhibitor (EcTI) on Aedes aegypti development as well as its effect on insecticidal activity of Bacillus thuringiensis toxins. RESULTS: EcTI showed an apparent molecular mass about of 20 kDa in SDS-PAGE and was able to inhibit in vitro the activity of trypsin and proteases from midgut of Ae. aegypti larvae. EcTI was not able to cause acute toxicity on mosquito larvae even at 1000 µg mL-1 , however it promoted a delay in larval development after prolonged exposure. The zymogram results for EcTI-treated larvae (from 50 to 200 µg mL-1 ) showed an increase of midgut proteases activity as a larvae defense mechanism, however no changes in the enzyme profile was observed. These same concentrations were able to enhance up to three fold the insecticidal activity of B. thuringiensis toxins without causing toxicity to Artemia sp. nauplii, a non-target organism. CONCLUSIONS: The results offer a novel approach by combining EcTI and B. thuringiensis toxins for combating Ae. aegypti larvae. © 2020 Society of Chemical Industry.

Proteomics analysis of zebrafish larvae exposed to 3,4-dichloroaniline using the fish embryo acute toxicity test.

The zebrafish (Danio rerio) is a small teleost fish that is becoming increasingly popular in laboratories worldwide and several attributes have also placed the zebrafish under the spotlight of (eco)toxicological studies. Since the 1990s, international organizations such as ISO and OECD have published guidelines for the use of zebrafish in ecotoxicological assessment of environmental toxicants such as the Fish Embryo Acute Toxicity (FET) test, OECD n° 236 guideline. This protocol uses 3,4-dichloroaniline (DCA), an aniline pesticide whose toxicity to fish species at early life stages is well known, as a positive control. Despite its use, little is known about its molecular mechanisms, especially in the context of the FET test. Therefore, this study aimed to investigate such changes in zebrafish larvae exposed to DCA (4 mg/L) for 96 hours using gel-free proteomics. Twenty-four proteins detected in both groups were identified as significantly affected by DCA exposure, and, when considering group-specific entities, 48 proteins were exclusive to DCA (group-specific proteins) while 248 were only detected in the control group. Proteins modulated by DCA treatment were found to be involved in metabolic processes, especially lipids and hormone metabolism (eg, Apoa1 and Apoa1b and vitelogenins), as well as proteins important for developmental processes and organogenesis (eg, Myhc4, Acta2, Sncb, and Marcksb). The results presented here may therefore provide a better understanding of the relationships between molecular changes and phenotype in zebrafish larvae treated with DCA, the reference compound of the FET test.

Evaluation of seeds ethanolic extracts of Triplaris gardneriana Wedd. using in vitro and in vivo toxicological methods.

Triplaris gardneriana Wedd. is a tree used in folk medicine to treat venereal diseases and inflammation as well as a source of biological compounds with antioxidant capacity. In order to assess the safety of these bioactive compounds, the present study aimed to determine the toxicity of an ethanolic extract of T. gardneriana, (EETg). Toxicological tests included hemolytic activity, toxicity toward the brine shrimp Artemia, cytotoxicity against breast cancer cells (MCF7) and acute oral toxicity in rodents. In addition, toxicogenomics techniques were used to determine genome expression in MCF7 cells exposed to EETg. The results showed that the extract exhibits approximately 60% of hemolytic activity at the highest tested concentration (64 µg/ml) and toxicity against nauplii of Artemia sp. (LC50 of 67.85 µg/ml). Further, EETg appears to be cytotoxic to MCF7 (cell viability reduced to 40% at 250 µg/ml after 24 hr). Genomic data demonstrated differential expression of 14 genes. Data analysis indicated possible altered pathways (e.g., xenobiotic metabolism), possible adverse health risks (e.g., hepatotoxicity), and drugs with similar gene expression profile (e.g., antimicrobials). The investigation provides important information on potentially adverse aspects of EETg, which need to be considered prior to the therapeutic utilization of this plant.Abbreviations: EETg: ethanolic extract of T. gardneriana seeds; MCF7: michigan cancer foundation-7 which refers to a human breast cell line (adenocarcinoma); NGS: next-generation sequencing; edgeR: empirical analysis of digital gene expression data in R; Consensus: consensus path database; FDR: false discovery rate; NCBI: national center for biotechnology information; KEGG: kyoto encyclopedia of genes and genomes; Ingenuity: ingenuity pathway analysis software; CMAP: connectivity map; OECD: organization for economic co-operation and development; HL-60: human promyelocytic leukemia cells; PC3: prostate cancer cells.

Expanding Pertussis Epidemiology in 6 Latin America Countries through the Latin American Pertussis Project.

The Latin American Pertussis Project (LAPP), established in 2009, is a collaboration between the Centers for Disease Control and Prevention, Pan American Health Organization, Sabin Vaccine Institute, and the ministries of health of 6 countries in Latin America. The project goal is to expand understanding of pertussis epidemiology in Latin America to inform strategies for control and prevention. Here we describe LAPP structure and activities. After an initial surveillance evaluation, LAPP activities are tailored to individual country needs. LAPP activities align with Global Health Security Agenda priorities and have focused on expanding laboratory diagnostic capacity, implementing a laboratory quality control and quality assurance program, and providing epidemiologic support to strengthen reporting of pertussis surveillance data. Lessons learned include that ongoing mentoring is key to the successful adoption of new technologies and that sustainability of laboratory diagnostics requires a regional commitment to procure reagents and related supplies.

A Protein Isolate from Moringa oleifera Leaves Has Hypoglycemic and Antioxidant Effects in Alloxan-Induced Diabetic Mice.

Moringa oleifera has been used in traditional medicine to treat diabetes. However, few studies have been conducted to relate its antidiabetic properties to proteins. In this study, a leaf protein isolate was obtained from M. oleifera leaves, named Mo-LPI, and the hypoglycemic and antioxidant effects on alloxan-induced diabetic mice were assessed. Mo-LPI was obtained by aqueous extraction, ammonium sulphate precipitation and dialysis. The electrophoresis profile and proteolytic hydrolysis confirmed its protein nature. Mo-LPI showed hemagglutinating activity, cross-reaction with anti-insulin antibodies and precipitation after zinc addition. Single-dose intraperitoneal (i.p.) administration of Mo-LPI (500 mg/kg·bw) reduced the blood glucose level (reductions of 34.3%, 60.9% and 66.4% after 1, 3 and 5 h, respectively). The effect of Mo-LPI was also evidenced in the repeated dose test with a 56.2% reduction in the blood glucose level on the 7th day after i.p. administration. Mo-LPI did not stimulate insulin secretion in diabetic mice. Mo-LPI was also effective in reducing the oxidative stress in diabetic mice by a decrease in malondialdehyde level and increase in catalase activity. Mo-LPI (2500 mg/kg·bw) did not cause acute toxicity to mice. Mo-LPI is a promising alternative or complementary agent to treat diabetes.

Day and night surgery: is there any influence in the patient postoperative period of urgent colorectal intervention?

BACKGROUND: Medical activity performed outside regular work hours may increase risk for patients and professionals. There is few data with respect to urgent colorectal surgery. The aim of this work was to evaluate the impact of daytime versus nighttime surgery on postoperative period of patients with acute colorectal disease. METHODS: A retrospective study was conducted in a sample of patients with acute colorectal disease who underwent urgent surgery at the General Surgery Unit of Braga Hospital, between January 2005 and March 2013. Patients were stratified by operative time of day into a daytime group (surgery between 8:00 and 20:59) and the nighttime group (21:00-7:59) and compared for clinical and surgical parameters. A questionnaire was distributed to surgeons, covering aspects related to the practice of urgent colorectal surgery and fatigue. RESULTS: A total of 330 patients were included, with 214 (64.8%) in the daytime group and 116 (35.2%) in the nighttime group. Colorectal cancer was the most frequent pathology. Waiting time (p < 0.001) and total length of hospital stay (p = 0.008) were significantly longer in the daytime group. There were no significant differences with respect to early or late complications. However, 100% of surgeons reported that they are less proficient during nighttime. CONCLUSIONS: Among patients with acute colorectal disease subjected to urgent surgery, there was no significant association between nighttime surgery and the presence of postoperative medical and surgical morbidities. Patients who were subjected to daytime surgery had longer length of stay at the hospital.

Assessment of Laparoscopic Skills Performance: 2D Versus 3D Vision and Classic Instrument Versus New Hand-Held Robotic Device for Laparoscopy.

INTRODUCTION AND OBJECTIVES: Laparoscopic surgery has undeniable advantages, such as reduced postoperative pain, smaller incisions, and faster recovery. However, to improve surgeons' performance, ergonomic adaptations of the laparoscopic instruments and introduction of robotic technology are needed. The aim of this study was to ascertain the influence of a new hand-held robotic device for laparoscopy (HHRDL) and 3D vision on laparoscopic skills performance of 2 different groups, naïve and expert. MATERIALS AND METHODS: Each participant performed 3 laparoscopic tasks-Peg transfer, Wire chaser, Knot-in 4 different ways. With random sequencing we assigned the execution order of the tasks based on the first type of visualization and laparoscopic instrument. Time to complete each laparoscopic task was recorded and analyzed with one-way analysis of variance. RESULTS: Eleven experts and 15 naïve participants were included. Three-dimensional video helps the naïve group to get better performance in Peg transfer, Wire chaser 2 hands, and Knot; the new device improved the execution of all laparoscopic tasks (P < .05). For expert group, the 3D video system benefited them in Peg transfer and Wire chaser 1 hand, and the robotic device in Peg transfer, Wire chaser 1 hand, and Wire chaser 2 hands (P < .05). CONCLUSION: The HHRDL helps the execution of difficult laparoscopic tasks, such as Knot, in the naïve group. Three-dimensional vision makes the laparoscopic performance of the participants without laparoscopic experience easier, unlike those with experience in laparoscopic procedures.

Food safety knowledge on the Bt mutant protein Cry8Ka5 employed in the development of coleopteran-resistant transgenic cotton plants.

Insecticidal Cry proteins from Bacillus thuringiensis (Bt) have been exploited in the development of genetically modified (GM) crops for pest control. However, several pests are still difficult to control such as the coleopteran boll weevil Anthonomus grandis. By applying in vitro molecular evolution to the cry8Ka1 gene sequence, variants were generated with improved activity against A. grandis. Among them, Cry8Ka5 mutant protein showed coleoptericidal activity 3-fold higher (LC50 2.83 µg/mL) than that of the original protein (Cry8Ka1). Cry8Ka5 has been used in breeding programs in order to obtain coleopteran-resistant cotton plants. Nevertheless, there is some concern in relation to the food safety of transgenic crops, especially to the heterologously expressed proteins. In this context, our research group has performed risk assessment studies on Cry8Ka5, using the tests recommended by Codex as well as tests that we proposed as alternative and/or complementary approaches. Our results on the risk analysis of Cry8Ka5 taken together with those of other Cry proteins, point out that there is a high degree of certainty on their food safety. It is reasonable to emphasize that most safety studies on Cry proteins have essentially used the Codex approach. However, other methodologies would potentially provide additional information such as studies on the effects of Cry proteins and derived peptides on the indigenous gastrointestinal microbiota and on intestinal epithelial cells of humans. Additionally, emerging technologies such as toxicogenomics potentially will offer sensitive alternatives for some current approaches or methods.

Further evidences for the mode of action of the larvicidal m-pentadecadienyl-phenol isolated from Myracrodruon urundeuva seeds against Aedes aegypti.

Nowadays, dengue fever is considered the most important arbovirosis worldwide and its control is still based upon combating the vector Aedes aegypti. Besides monitoring of mosquito populations resistant to conventional insecticides, the search for new environmentally safe insecticides and conduction of molecular studies focusing on the elucidation of mode of action and possible resistance mechanisms are considered the key for a sustainable management of the mosquito vector. Thus, the present work aimed to assess changes in protein expression of 3rd-instar larvae of Ae. aegypti after exposure to the natural insecticide m-pentadecadienyl-phenol. Bidimensional electrophoresis followed by mass spectrometry resulted in identification of 12 proteins differentially expressed between control and treated groups. Larvae exposed to the toxic compound for 24h showed elevated detoxification response (glutathione-S-transferase), increased levels of stress-related proteins (HSP70) as well as evidence of lysosome stabilization to enable survival. Furthermore, expression of proteins involved in protection of peritrophic membrane and metabolism of lipids indicated systemic effect of toxic effects in treated larvae.

Increased Levels of Antinutritional and/or Defense Proteins Reduced the Protein Quality of a Disease-Resistant Soybean Cultivar.

The biochemical and nutritional attributes of two soybean (Glycine max (L.) Merr.) cultivars, one susceptible (Seridó) and the other resistant (Seridó-RCH) to stem canker, were examined to assess whether the resistance to pathogens was related to levels of antinutritional and/or defense proteins in the plant and subsequently affected the nutritional quality. Lectin, urease, trypsin inhibitor, peroxidase and chitinase activities were higher in the resistant cultivar. Growing rats were fed with isocaloric and isoproteic diets prepared with defatted raw soybean meals. Those on the Seridó-RCH diet showed the worst performance in terms of protein quality indicators. Based on regression analysis, lectin, trypsin inhibitor, peroxidase and chitinase appear to be involved in the resistance trait but also in the poorer nutritional quality of Seridó-RCH. Thus, the development of cultivars for disease resistance may lead to higher concentrations of antinutritional compounds, affecting the quality of soybean seeds. Further research that includes the assessment of more cultivars/genotypes is needed.

Food safety assessment of an antifungal protein from Moringa oleifera seeds in an agricultural biotechnology perspective.

Mo-CBP3 is an antifungal protein produced by Moringa oleifera which has been investigated as potential candidate for developing transgenic crops. Before the use of novel proteins, food safety tests must be conducted. This work represents an early food safety assessment of Mo-CBP3, using the two-tiered approach proposed by ILSI. The history of safe use, mode of action and results for amino acid sequence homology using the full-length and short contiguous amino acids sequences indicate low risk associated to this protein. Mo-CBP3 isoforms presented a reasonable number of alignments (>35% identity) with allergens in a window of 80 amino acids. This protein was resistant to pepsin degradation up to 2 h, but it was susceptible to digestion using pancreatin. Many positive attributes were presented for Mo-CBP3. However, this protein showed high sequence homology with allergens and resistance to pepsin digestion that indicates that further hypothesis-based testing on its potential allergenicity must be done. Additionally, animal toxicity evaluations (e.g. acute and repeated dose oral exposure assays) must be performed to meet the mandatory requirements of several regulatory agencies. Finally, the approach adopted here exemplified the importance of performing an early risk assessment of candidate proteins for use in plant transformation programs.

Electrocardiogram, heart movement and heart rate in the awake gecko (Hemidactylus mabouia).

The electrocardiogram (ECG) is the simplest and most effective non-invasive method to assess the electrical activity of the heart and to obtain information on the heart rate (HR) and rhythm. Because information on the HR of very small reptiles (body mass <10 g) is still scarce in the literature, in the present work we describe a procedure for recording the ECG in non-anesthetized geckos (Hemidactylus mabouia, Moreau de Jonnès, 1818) under different conditions, namely manual restraint (MR), spontaneous tonic immobility (TI), and in the non-restrained condition (NR). In the gecko ECG, the P, QRS and T waves were clearly distinguishable. The HR was 2.83 ± 0.02 Hz under MR, which was significantly greater (p < 0.001) than the HR under the TI (1.65 ± 0.09 Hz) and NR (1.60 ± 0.10 Hz) conditions. Spontaneously beating isolated gecko hearts contracted at 0.84 ± 0.03 Hz. The in vitro beating rate was affected in a concentration-dependent fashion by adrenoceptor stimulation with noradrenaline, as well as by the muscarinic cholinergic agonist carbachol, which produced significant positive and negative chronotropic effects, respectively (p < 0.001). To our knowledge, this is the first report on the ECG morphology and HR values in geckos, particularly under TI. The methodology and instrumentation developed here are useful for non-invasive in vivo physiological and pharmacological studies in small reptiles without the need of physical restraint or anesthesia.

Análise microscópica e ultraestrutural das glândulas salivares mandibulares de Procyon cancrivorus / Microscopy and ultrastructural analysis of mandibular salivary glands of Procyon cancrivorus

Devido a importância ecológica dos Procyon cancrivorus, o objetivo deste trabalho foi caracterizar morfologicamente e ultra estruturalmente a glândula salivar mandibular desta espécie. Foram utilizadas 10 pares (direita e esquerda) de glândulas salivares mandibulares de cinco animais adultos. As glândulas salivares mandibulares foram dissecadas e mensuradas com paquímetro de precisão e posteriormente processadas por técnica rotineira de histologia e coradas por HE (hematoxilina e eosina) e Picrossírius. Fragmentos das glândulas foram processados para análise em microscopia eletrônica de varredura (MEV). As glândulas salivares mandibulares direitas e esquerdas de P. cancrivorus apresentaram-se lobuladas, formato ovalado, e posicionadas entre a fossa atlantis e o osso basihyoideum do crânio. Microscopicamente, estas glândulas estavam revestidas por uma cápsula de tecido conjuntivo denso não modelado, a qual adentra a glândula através de septos, dividindo-a em lóbulos. Nos septos de tecido conjuntivo estão presentes vasos sanguíneos e nervos, além de ductos interlobulares excretores. Dentro dos lóbulos das glândulas salivares mandibulares, são encontrados ductos do tipo estriado e intercalar, além de predomínio de ácinos do tipo mucoso. Os resultados permitem concluir que as glândulas salivares mandibulares dos Procyon cancrivorus, seguem o padrão estrutural descrito em outras espécies de mamíferos. No entanto os tipos de ácinos podem variar entre as espécies, sendo, portanto, necessários futuros estudos histoquímicos e de biologia celular para desvendar a importância dessa variação para a espécie aqui estudada, comparando com seu hábito alimentar.

Due to the ecological importance of Procyon cancrivorus, the aim of this study was to characterize morphologically and ultrastructurally the mandibular salivary gland of this specie. We used 10 pairs (right and left) of mandibular salivary glands from five adult animals. The mandibular salivary glands were dissected and measured by precision calipers and later processed by histological routine technique, stained by HE (hematoxylin and eosin) and Picrossirius. Fragments of the glands were processed for analysis by scanning electron microscopy (SEM). The mandibular salivary gland, right and left from P. cancrivorus presents lobulated and oval-shape, and it was positioned between the fossa atlantis and basihyoideum ossis of skull. Microscopically, these glands are lined by a capsule formed by dense irregular connective tissue, which enters into the gland by septa, dividing it into lobules. Into the septa of connective tissue were present blood vessels and nerves, and interlobular excretory ducts. Inside the lobes of the mandibular salivary glands, striatum and intermediate-type ducts are found, and the predominance of mucous acini was observed. The results indicate that the mandibular salivary glands of Procyon cancrivorus, follows the structural pattern described in other mammalian species. However the types of acini may vary among species, and thus further histochemical and cell biology studies are required in order to uncover the significance of this variation for the specie here studied, according with their feeding habit.

Morfologia da glândula mamária do quati (Nasua nasua) / Morphology of mammary gland of the coati (Nasua nasua)

Nasua nasua é um animal onívoro, encontrado em todo Brasil. A reprodução da espécie ocorre somente uma vez ao ano, na primavera. As fêmeas são matriarcas e amamentam suas crias até os 5 meses de idade, vivem com seus filhotes em bandos de até 30 indivíduos. Para descrição morfológica da glândula mamaria do Nasua nasua foram utilizados seis animais provenientes do Criatório Cientifico (Cecrimpas), Unifeob. Autorizado pelo Ibama (Proc.02027.002322/98-99). Para análise macroscópica um animal foi injetado com látex neoprene, sendo a artéria femoral injetada com látex de cor vermelha e a veia jugular de cor azul. Os demais animais foram fixados em solução aquosa a 10% de formaldeído. Para análise microscópica, fragmentos glandulares foram coletados e submetidos ao processo rotineiro, embebido em parafina e corados com Hematoxilina e Eosina, Picrossírius e Azul de Toluidina. Macroscopicamente foram evidenciados três pares de glândulas mamárias, sendo dois pares posicionados na região abdominal e um par na região inguinal. Microscopicamente, notou-se epitélio de revestimento externo das papilas mamárias, epitélio pavimentoso estratificado queratinizado, o qual seguia por toda glândula de forma irregular. Na entrada do óstio, o epitélio da epiderme era modificado ocorrendo uma transição de epitélio pavimentoso estratificado para um epitélio cúbico no ducto papilar. O parênquima glandular era caracteristicamente túbulo alveolar com células secretoras, evidenciado principalmente no animal lactente. Os resultados macroscópicos e microscópicos assemelham-se aos já descritos nas cadelas (Canis familiaris) e os do Procyon cancrivorus pertencente à mesma família do quati, Família Procyonidae.

Coati is an omnivorous animal, found throughout of Brazil. The reproduction of this species occurs only once a year during spring. Females are matriarchs and nurse their young until 5 months old, living with their young in groups of up to 30. For morphological description of the mammary gland Nasua nasua six animals were used coming from the hatchery Scientific (Cecrimpas), Unifeob. Authorized by IBAMA (Proc. 02027.002322/98-99). For the macroscopic analysis one animal was injected with neoprene latex, was injected into the femoral artery with red latex and the jugular vein with blue colored one. The others animals were fixed in a 10% aqueous solution of formaldehyde. For microscopic analysis, glandular fragments were collected and submitted to routine process, embedded in paraffin and stained with hematoxylin and eosin, toluidine blue and picrossirius. Macroscopically were evidenced three pairs of mammary glands, two pairs positioned in the abdominal region and one pair in the inguinal region. Microscopically, was found a simple epithelium lining external mammary papillae, keratinized stratified epithelium, which continued throughout the gland. At the entrance of the papillae ostium the epithelium of the epidermis was modified occurring a transition from stratified epithelium to cubical epithelium into the papillary duct. The glandular parenchyma was characteristically alveolar with secretory cells observed predominantly in the lactating female. The macroscopic and microscopic results are similar to those already described in bitches (Cannis familiaris) and Procyon cancrivorus belonging to the same family of coati, the family Procyonidae.

A morphological analysis of the transition between the embryonic primitive intestine and yolk sac in bovine embryos and fetuses.

The yolk sac (YS) is the main source of embryonic nutrition during the period when the placenta has not yet formed. It is also responsible for hematopoiesis because the blood cells develop from it as part of the primitive embryonic circulation. The objective of this study was to characterize the transitional area between the YS and primitive gut using the techniques of light microscopy, transmission electron microscopy, and immunohistochemistry to detect populations of pluripotent cells by labeling with Oct4 antibody. In all investigated embryos, serial sections were made to permit the identification of this small, restricted area. We identified the YS connection with the primitive intestine and found that it is composed of many blood islands, which correspond to the vessels covered by vitelline and mesenchymal cells. We identified large numbers of hemangioblasts inside the vessels. The mesenchymal layer was thin and composed of elongated cells, and the vitelline endodermal membrane was composed of large, mono- or binucleated cells. The epithelium of the primitive intestine comprised stratified columnar cells and undifferentiated mesenchymal cells. The transitional area between the YS and the primitive intestine was very thin and composed of cells with irregular shapes, which formed a delicate lumen containing hemangioblasts. In the mesenchyme of the transitional area, there were a considerable number of small vessels containing hemangioblasts. Using Oct4 as a primary antibody, we identified positive cells in the metanephros, primordial gonad, and hepatic parenchyma as well as in YS cells, suggesting that these regions contain populations of pluripotent cells.

Identificação do nicho de progenitores mesenquimais no fígado de embriões e fetos caninos: uma fonte de células-tronco para terapia celular / Identification of mesenchymal progenitor niches from liver of embryo and fetuses of canines: a source of stem cells for cell therapy

As células-tronco (CT) derivadas dos tecidos fetais (TF) foram as mais recentes descobertas entre as CT, e ultimamente tem demonstrado amplo potencial terapêutico, dentre os TF o fígado fetal (FF) apresenta grande potencial terapêutico. Este órgão durante o período fetal em mamíferos é um nicho hematopoético transitório, sendo o principal órgão responsável pela hematopoese no feto, além de contribuir com a formação do nicho definitivo na medula óssea adulta, portanto pode ser considerado um nicho de células-tronco mesenquimais (CTM) e progenitores. No entanto, pouco se sabe sobre a localização destas células no FF, desta forma o presente estudo visa identificar o nicho de CTM e progenitores em FF de cães, a fim de contribuir com as técnicas de isolamento e extração celular. Em conjunto foi realizada a verificação da expressão do fator de transcrição Oct-3/4 e da proteína delta polimerase do DNA (PCNA). Para a análise foram utilizados cinco embriões e 11 fetos caninos com idades gestacionais variando de 25-60 dias. Os resultados elucidaram a partir de 25 dias de gestação o FF apresentou-se volumoso e composto por todas as estruturas típicas, dentre elas a tríade portal, ductos biliares e ramos das artérias hepáticas. Com 30 dias de gestação foram identificados os primeiros sitos de progenitores mesenquimais (PM) enquanto que aos 60 dias os nichos estavam completamente formados com localização semelhante ao fígado adulto (FA). No entanto, células imunopositivas para Oct-3/4 não foram identificadas; sendo assim, destacamos que o FF é uma fonte de PM, apresentando-se como uma alternativa para a utilização terapêutica, bem como para os estudos da biologia do desenvolvimento das CTM e progenitores.(AU)

Stem cells (SC) derived from fetal tissues (FT) were the most recent discoveries among the SC, and lately had demonstrated broad therapeutic potential; the FT from the fetal liver (FL) presents great therapeutic potential. This organ during the fetal period in mammals acts as a transient hematopoietic niche, being the main organ responsible for hematopoiesis in the fetus, and contribute to the formation of permanent niche in the adult bone marrow, thus can be considered a niche for mesenchymal stem cells (MSC) and parents. However, little is known about the location of these cells in FF, so the present study aims to identify the niche of mesenchymal progenitors in FF and dogs in order to contribute to the techniques of cell isolation and extraction. Together was performed to verify the expression of the transcription factor Oct-3/4 of the protein and DNA polymerase delta (PCNA). For the analysis of five embryos were used and 11 canine fetuses with gestational ages ranging from 25-60 days. The results elucidated from 25 days of gestation showed the FF is bulky and composed of all the typical structures, among them the portal triad, bile ducts and hepatic artery branches. With 30 days of gestation were identified the first requirements of mesenchymal progenitors (MP) at 60 days while the niches were completely formed with location similar to adult liver (AL). However, cells immunoreactives for Oct-3/4 were not identified, therefore, point out that the FL is a source of PM, presenting as an alternative for therapeutic purposes as well as for studying the developmental biology of MSC and parents.(AU)

Macroscopic and microscopic analysis of the tongue of the common opossum (Didelphis marsupialis).

We performed a macroscopic and microscopic study of the tongues of common opossums, Didelphis marsupialis, from South America. We studied two males and two females. We collected morphometric data on the tongue with precision calipers. For the light microscopy and scanning electron microscopy analyses, we fixed tissue fragments in 10% formaldehyde and 2.5% glutaraldehyde, respectively. The opossum tongues averaged 5.87 ± 0.20 cm in length, 3.27 ± 0.15 cm in width at the lingual body, and 3.82 ± 0.15 cm in width at the root. The mean thickness of the lingual body was 1.8 ± 0.1 cm, and the thickness of the root was 3.82 ± 0.15 cm. Sharp filiform papillae were scattered across the entire tongue; conical filiform papillae occurred on the lingual body and tongue tip; fungiform papillae were scattered among the filiform papillae on the lingual body and tongue tip; and there were three vallate papillae at the root of the tongue. We found two strands of papillary projections in the tongue root. Despite the low variability observed in the lingual papillae, the morphological data obtained in this study may be related to the opossum's diverse food habits and the extensive geographic distribution of the species throughout America.

A brief report on some health aspects of rats fed with crescent levels of recombinant chagasin, a potential plant defense protein.

Chagasin may be considered a potential plant-incorporated protectant (PIP) protein due to its deleterious effects on insect pests. However, extensive safety studies with PIP's are necessary before introducing them into the target plant. Thus, a short-term feeding trial in rats with high doses of r-chagasin was conducted to provide evidences about its safety. Three test diets containing casein + r-chagasin (0.25, 0.5 and 1% of total protein) were offered to rats (10 days). The test diets did not show adverse effects upon the development, organ weight, hematological parameters and serum protein profiles of rats, providing preliminary information on the safety of r-chagasin.

Morfologia macro e microscópica das papilas linguais do quati (Nasua nasua) / Gross and microscopic morphology of the coati tongue (Nasua nasua)

O quati (Nasua nasua) é um animal que pertence à Família Procyonidae. Foram utilizados três animais ortotanasiados, de ambos os sexos, provenientes do Criatório Científico de Animais Silvestres, Centro Universitário Fundação de Ensino Octávio Bastos (Cecrimpas, Unifeob) autorizado pelo IBAMA (Proc.02027.003731/04-76). Para a análise macroscópica, as línguas foram retiradas, analisadas e foto-documentadas. Para análise microscópica, as línguas foram processadas rotineiramente pela técnica de microscopia eletrônica de varredura e inclusão em Paraplast; pela técnica de microscopia de luz os fragmentos foram cortados em micrótomo, com espessura média de 5mm e corados em HE e Picrosírius com fundo de hematoxilina. Os resultados macroscópicos e microscópicos mostram que a língua do quati apresenta papilas filiformes, fungiformes, valadas e cônicas sendo estas distribuídas nas regiões rostralis, medialis e caudalis. Histologicamente, a língua do quati é revestida por um epitélio pavimentoso estratificado queratinizado apresentando camada basal, espinhosa, granulosa e córnea com fibras de músculos estriados esqueléticos longitudinais e transversais e diversas glândulas. De acordo com os resultados pode-se concluir que a língua do quati possui características macroscópicas e microscópicas semelhantes aos canídeos, tendo como diferença o número de papilas valadas e o grau de queratinização.

The coati (Nasua nasua) is an animal that belongs to the Procyonidae family. For this study we used three orthotanasized animals of both sexes, obtained from the Scientific Center for Wild Animal Breeding, University Center Octavio Bastos Educational Foundation (Cecrimpas, Unifeob), authorized by IBAMA (Proc.02027.003731/04-76). For microscopic analysis, tongues were processed routinely through electron microscopy scanning and inclusion in Paraplast; for light microscopy, the fragments were cut by microtome with an average thickness of 5mm and stained with H&E and Picrosirius counterstained with hematoxylin. The macroscopic and microscopic results show that the tongue of coati has filiform, fungiform, vallate and conical papillae distributed in the rostralis, medialis and caudalis regions. Histologically, the tongue is lined by a keratinized stratified squamous epithelium with spinous, granulous and keratin basal layer, with striated longitudinal and transverse skeletal muscles fibers, and several glands. We can conclude that coati´s tongue has macroscopic and microscopic features similar to the ones of Canidae, with differences in the number of vallate papillae and degree of keratinization.

Study of the antiproliferative potential of seed extracts from Northeastern Brazilian plants

This study assessed the antiproliferative and cytotoxic potential against tumor lines of ethanolic seed extracts of 21 plant species belonging to different families from Northeastern Brazil. In addition, some underlying mechanisms involved in this cytotoxicity were also investigated. Among the 21 extracts tested, the MTT assay after 72 h of incubation demonstrated that only the ethanolic extract obtained from Myracrodruon urundeuva seeds (EEMUS), which has steroids, alkaloids and phenols, showed in vitro cytotoxic activity against human cancer cells, being 2-fold more active on leukemia HL-60 line [IC50 value of 12.5 (9.5-16.7) μg/mL] than on glioblastoma SF-295 [IC50 of 25.1 (17.3-36.3) μg/mL] and Sarcoma 180 cells [IC50 of 38.1 (33.5-43.4) μg/mL]. After 72h exposure, flow cytometric and morphological analyses of HL-60-treated cells showed that EEMUS caused decrease in cell number, volume and viability as well as internucleosomal DNA fragmentation in a dose-dependent way, suggesting that the EEMUS triggers apoptotic pathways of cell death.

Este estudo avaliou o potencial antiproliferativo e citotóxico contra linhagens de células tumorais de extratos etanólicos de sementes de vinte e uma espécies vegetais pertencentes a diferentes famílias do Nordeste brasileiro. Além disso, alguns mecanismos subjacentes envolvidos nesta citotoxidade também foram investigados. Dentre os 21 extratos testados pelo ensaio do MTT após 72 h de incubação, apenas o extrato etanólico obtido a partir de sementes de Myracrodruon urundeuva (EEMUS), o qual apresentou traços de esteróides, alcalóides e fenóis em sua composição, demonstrou atividade citotóxica in vitro contra células tumorais humanas, sendo 2 vezes mais ativo sobre a linhagem leucêmica HL-60 [IC50 valor de 12,5 (9,5-16,7) μg/mL] do que sobre células de glioblastoma SF-295 [IC50 de 25,1 (17,3-36,3) μg/mL] e de sarcoma 180 [IC50 de 38,1 (33,5-43,4) μg/mL]. Após 72 h de exposição, as análises morfológicas e por citometria de fluxo de células HL-60 tratadas com EEMUS mostraram diminuição no número de células, seu volume e viabilidade, assim como fragmentação internucleosomal do DNA de forma dose-dependente, sugerindo que a ação antiproliferativa de EEMUS pode ser ativada por vias apoptóticas.