Atividades Biológicas De Xilana De Sabugo De Milho

The corn cob is an agricultural by-product still little used, this in part due to the low knowledge of the biotechnological potential of their molecules. Xylan from corn cobs (XSM) is a polysaccharide present in greater quantity in the structure of plant and its biotechnology potential is little known. This study aimed to the extraction, chemical characterization and evaluation of biological activities of xylan from corn cobs. To this end, corncobs were cleaned, cut, dried and crushed, resulting in flour. This was subjected to a methodology that combines the use of alkaline conditions with waves of ultrasound. After methanol precipitation, centrifugation and drying was obtained a yield of 40% (g/g flour). Chemical analysis indicated a high percentage of polysaccharides in the sample (60%) and low contamination by protein (0.4%) and phenolic compounds (> 0.01%). Analysis of monosaccharide composition indicated the presence of xylose:glucose:arabinose:galactose:mannose:glucuronic acid in a molar ratio 50:20:15:10:2.5:2.5. The presence of xylan in the sample was confirmed by nuclear magnetic resonance (¹H and ¹³C) and infrared spectroscopy (IR). Tests were conducted to evaluate the antioxidant potential of XSM. This showed a total antioxidant capacity of 48.45 EAA/g sample. However, did not show scavenging activity of superoxide and hydroxyl radical and also reducing power. But, showing a high capacity chelating iron ions with 70% with about 2 mg/mL. The ability to XSM to influence cell proliferation in culture was also evaluated. This polymer did not influence the proliferation of normal fibroblast cells (3T3), however, decreased the rate of proliferation of tumor cells (HeLa) in a dose-dependent, reaching an inhibition of about 50% with a concentration around 2 mg/mL. Analyzing proteins related to cell death, by immunoblotting, XSM increases the amount of Bax, Bcl-2 decrease, increase cytochrome c and AIF, and reduce pro-caspase-3, indicating the induction of cell death induced apoptosis dependent and independent of caspase. XSM did not show anticoagulant activity in the PT test. However, the test of activated partial thromboplastin time (aPTT), XSM increased clotting time at about 5 times with 600 μg of sample compared with the negative control. The presence of sulfate on the XSM was discarded by agarose gel electrophoresis and IR. After carboxyl-reduction of XSM the anticoagulant activity decreased dramatically. The data of this study demonstrate that XSM has potential as antioxidant, antiproliferative and anticoagulant compound. Future studies to characterize these activities of XSM will help to increase knowledge about this molecule extracted from corn and allow their use in functional foods, pharmaceuticals and chemical industries.

Indução de morte celular em células de adenocarcinoma cervical humano (HeLa) pela lectina da esponja Cinachyrella apion (CaL)

Cancer is a term used to represent a set of more than 100 diseases, including malignant tumors from different locations. The malignancies are the second leading cause of death in the population, representing approximately 17% of deaths of known cause. Strategies that induce differentiation have had limited success in the treatment of established cancers. In this work, a lectin purified from the marine sponge Cinachyrella apion (CaL) was evaluated due to its hemolytic, cytotoxic and antiproliferative properties, besides the ability to induce cell death via apoptosis in tumor cells. The antiproliferative activity of CaL was tested against cell lines, with the highest inhibition of tumor growth for HeLa, reducing cell growth at a dose dependent manner, with a concentration of 10 μg/mL. The hemolytic activity and toxicity against peripheral blood cells were tested using the concentration of IC50 for both trials and twice the IC50 for analysis in flow cytometry, indicating that CaL is not toxic to these cells. To assess the mechanism of cell death caused by CaL in HeLa cells, we performed flow cytometry and western blotting. The results showed the lectin probably induces cell death by apoptosis activation by pro-apoptotic protein Bax, promoting mitochondrial membrane permeabilization, cell cycle arrest in S phase, with accumulation of cells of approximately 57% in this phase, and acting as both dependent and/or independent of caspases pathway. These results suggest that CaL has the potential to be used as drug treatment against cancer.

Avaliação do efeito do composto tipo heparina isolado do caranguejo Chaceon fenneri na hemostasia e na morte celular

Heparin is a pharmaceutical animal widely used in medicine due to its potent anticoagulant effect. Furthermore, it has the ability to inhibit the proliferation, invasion and adhesion of cancer cells to vascular endothelium. However, its clinical applicability can be compromised by side effects such as bleeding. Thus, the search for natural compounds with low bleeding risk and possible therapeutic applicability has been targeted by several research groups. From this perspective, this study aims to evaluate the hemorrhagic and anticoagulant activities and citotoxic effect for different tumor cell lines (HeLa, B16-F10, HepG2, HS-5,) and fibroblast cells (3T3) of the Heparin-like from the crab Chaceon fenneri (HEP-like). The HEP-like was purified after proteolysis, ion-exchange chromatography, fractionation with acetone and characterized by electrophoresis (agarose gel) and enzymatic degradation. Hep-like showed eletroforetic behavior similar to mammalian heparin, and high trisulfated /Nacetylated disaccharides ratio. In addition, HEP-like presented low in vitro anticoagulant activity using aPTT and a minor hemorrhagic effect when compared to mammalian heparin. Furthermore, the HEP-like showed significant cytotoxic effect (p<0.001) on HeLa, HepG2 and B16-F10 tumor cells with IC50 values of 1000 ug/mL, after incubation for 72 hours. To assess the influence of heparin-like on the cell cycle in HeLa cells, analysis was performed by flow cytometry. The results of this analysis showed that HEP-like influence on the cell cycle increasing S phase and decreasing phase G2. Thus, these properties of HEP-like make these compounds potential therapeutic agents

Avaliação das atividades antioxidantes e citotóxicas de extratos ricos em polissacarídeos extraídos das hastes de mandacaru (Cereus Jamacaru de Candolle, Cactaceae)

The use of medicinal plants to cure and treat various diseases is a common practice in the world and in Brazil. In several regions of the Brazil´s Northeast, the cactus Cereus jamacaru, known as mandacaru, is used popularly as a treatment to many diseases, including those related to heart respiratory diseases, gastric ulcers, scurvy, and kidney diseases. However, there is a scarcity in the scientific literature that proves scientifically the popular application of this cactus. Like other plants, Cereus jamacaru synthesizes several potentially bioactive molecules, like as polysaccharides. In this work, three polysaccharides-rich aqueous extracts, MCA80, MPM and MCP60, were obtained from this plant and analyzed chemically, as well as their cytotoxic and antioxidant potential. The data showed that all extracts consist mainly of polysaccharides (89.42 to 95.76%), but also protein (> 2%) and phenolic (3 to 8.87%) contaminants were detected. All extracts are rich in galactose, glucose and mannose. In addition, glucuronic acid was found in MCA80 and MCP60. The extracts showed total antioxidant capacity ranged from 55.21 to 68.13 of ascorbic acid equivalents (AAE). Besides, they exhibited reducer power and cupric chelation in a dose-dependent manner. None of the extracts inhibited the MTT reduction in the presence of prostate tumor cells (PC-3). However, MCP60 was the most effective extract by preventing the reduction of MTT by about 80% in the presence of cells 786. Nuclear fragmentation tests showed that this extract induces cell death. The data indicated that mandacaru synthesizes bioactive polysaccharides with potential as antioxidant and antitumor agents. For future studies, it is intended to purify and characterize these polysaccharides and its antioxidant and antitumor mechanisms

Produção e caracterização de quitooligossacarídeos produzidos pelo fungo Metarhizium anisopliae e avaliação da citotoxicidade em células tumorais

Chitosan is a natural polymer, biodegradable, nontoxic, high molecular weight derived from marine animals, insects and microorganisms. Oligomers of glucosamine (GlcN) and N-acetylglucosamine (GlcNAc) have interesting biological activities, including antitumor effects, antimicrobial activity, antioxidant and others. The alternative proposed by this work was to study the viability of producing chitooligosaccharides using a crude enzymes extract produced by the fungus Metarhizium anisopliae. Hydrolysis of chitosan was carried out at different times, from 10 to 60 minutes to produce chitooligosaccharides with detection and quantification performed by High Performace Liquid Chromatography (HPLC). The evaluation of cytotoxicity of chitosan oligomers was carried out in tumor cells (HepG2 and HeLa) and non-tumor (3T3). The cells were treated for 72 hours with the oligomers and cell viability investigated using the method of MTT. The production of chitosan oligomers was higher for 10 minutes of hydrolysis, with pentamers concentration of 0.15 mg/mL, but the hexamers, the molecules showing greater interest in biological properties, were observed only with 30 minutes of hydrolysis with a concentration of 0.004 mg/mL. A study to evaluate the biological activities of COS including cytotoxicity in tumor and normal cells and various tests in vitro antioxidant activity of pure chitosan oligomers and the mixture of oligomers produced by the crude enzyme was performed. Moreover, the compound with the highest cytotoxicity among the oligomers was pure glucosamine, with IC50 values of 0.30; 0.49; 0.44 mg/mL for HepG2 cells, HeLa and 3T3, respectively. Superoxide anion scavenging was the mainly antioxidant activity showed by the COS and oligomers. This activity was also depending on the oligomer composition in the chitosan hydrolysates. The oligomers produced by hydrolysis for 20 minutes was analyzed for the ability to inhibit tumor cells showing inhibition of proliferation only in HeLa cells, did not show any effect in HepG2 cells and fibroblast cells (3T3)

Avaliação da atividade antioxidante e antiproliferativa do extrato aquoso de frutas tropicais

The antioxidant activity of aqueous extracts of five edible tropical fruits (Spondias lutea, Hancornia speciosa, Spondias purpurea, Manilkara zapota and Averrhoa carambola) was investigated using different methods. The amount of phenolic compounds was determined by the Folin-Ciocalteu reagent. The M. zapota had Total Antioxidant Capacity (TAC) higher than the other fruits. Extracts showed neither reducing power nor iron chelation (between 0.01 and 2.0 mg/mL). H. speciosa exhibited the highest superoxide scavenging activity (80%, 0.5 mg/mL). However, at high concentrations (8.0 mg/mL) only A. carambola, S. purpurea and S. lutea scavenging 100% of radicals formed. M. zapota and S. purpurea had higher phenolic compound levels and greater OH radical scavenging activity (92 %, 2.0 mg/mL). Antiproliferative activity was assessed with 3T3 fibroblasts and cervical tumor cells (HeLa). The most potent extract was S. purpurea (0.5 mg/mL), which inhibited HeLa cell proliferation by 52%. The most fruits showed antioxidant and antiproliferative properties, characterizing them as functional foods.

Avaliação in vitro da eficácia de produtos homeopáticos contendo momordica charantia através de bioensaios

Homeopathic medicines have been used for over two hundred years without the examination of their effects on in vivo and in vitro assays, due to the peculiarity of homeopathic preparations, the high dilution, which creates a challenge for the use of usual analytical techniques of quality control of medicine.Although there is scarcity of literature and variety of experiments, recently there have been some studies with few in vitro assays which have shown positive responses when evaluating the mechanism of action of homeopathic medicines which are able to act on a specific system.The present study aims to evaluate the efficacy of homeopathic products containing Momordica charantia through bioassays.Homeopathic products were tested by the MTT to assess cytotoxicity in RAW 264.7 (macrophage-like cells) and in tumor cells HeLa (human cervical adenocarcinoma cells), CHO K1 (Chinese hamster ovary cells), PANC-1 (human pancreas cancer cells) and PC-3 (human prostate cancer cells), dosage of inflammatory mediators NO, TNF-α and IL-6 released by RAW 264.7 cells, analysis of the death process and cell cycle changes of PC-3 by flow cytometry. The data demonstrate that homeopathic products of Momordica charantia did not show cytotoxicity to RAW 264.7, increased the production of inflammatory mediators by RAW 264.7 synergistically with LPS, showed cytotoxicity to PC-3 with change in its cell cycle inhibiting its proliferation, being the 30CH the most potent sample. Correlation studies were conducted in order to evaluate the possible in vitro applicable models to the quality control of homeopathic products with Momordica charantia. The data showed that the best applicable models in assessing the quality are the MTT to assess cytotoxicity in RAW 264.7 and PC-3 in 24 hours for Momordica charantia fruit products and dosage of NO production by RAW 264.7 with and without LPS

Inhibitory Effect of Gallic Acid and Its Esters on 2,2 '-Azobis(2-amidinopropane)hydrochloride (AAPH)-Induced Hemolysis and Depletion of Intracellular Glutathione in Erythrocytes

The protective effect of gallic acid and its esters, methyl, propyl, and lauryl gallate, against 2,2'-azobis(2-amidinopropane)hydrochloride (AAPH)-induced hemolysis and depletion of intracellular glutathione (GSH) in erythrocytes was studied. The inhibition of hemolysis was dose-dependent, and the esters were significantly more effective than gallic acid. Gallic acid and its esters were compared with regard to their reactivity to free radicals, using the DPPH and AAPH/pyranine free-cell assays, and no significant difference was obtained. Gallic acid and its esters not only failed to inhibit the depletion of intracellular GSH in erythrocytes induced by AAPH but exacerbated it. Similarly, the oxidation of GSH by AAPH or horseradish peroxidase/H(2)O(2) in cell-free systems was exacerbated by gallic acid or gallates. This property could be involved in the recent findings on proapoptotic and pro-oxidant activities of gallates in tumor cells. We provide evidence that lipophilicity and not only radical scavenger potency is an important factor regarding the efficiency of antihemolytic substances.

Synthesis, characterization, X-ray structure and in vitro anti mycobacterial and antitumoral activities of Ru(II) phosphine/diimine complexes containing the "SpymMe(2)" ligand, SpymMe(2)=4,6-dimethyl-2-mercaptopyrimidine

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Antitumor and immunomodulatory effects of the naphthoquinone 5-methoxy-3,4-dehydroxanthomegnin

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

2-Bromo-1,4-naphthoquinone: a potentially improved substitute of menadione in Apatone therapy

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Estudo da imunoexpressão dos transportadores de glicose 1 e 3 em carcinomas epidermóides de lábio inferior e sua relação com parâmetros clínico-patológicos

The Epidermoid Carcinoma (EC) is the most common lesions located in the region of the head and neck and, despite advances in treatment modalities, the prognosis is still poor. The malignant cells show an increase in glucose uptake, process mediated by glucose transporters (GLUTs). Increased expression of GLUT 1 and GLUT 3 is related to the aggressive behavior of this lesion. The aim of this study was to evaluate, through immunohistochemistry, the expression of GLUTs 1 and 3 in EC of the lower lip. The sample consisted of 40 cases of EC of the lower lip, of which 20 had regional lymph node metastasis and the remaining 20 with absence of metastasis. The percentages of immunostained cells in front of tumor invasion and in the center of tumor were evaluated. These results were related to the presence and absence of lymph node metastasis, TNM classification and histological grading. The percentage of cytoplasmic/membranous expression of GLUT 1 ranged from 77.35% to 100%, while for GLUT 3 this value ranged from 0.79% to 100%. As for nuclear staining for GLUT 1, this percentage ranged from 0 to 0.42%, however. GLUT 3 showed only one case with nuclear staining. Despite the significant expression of tumor cells related to the proteins studied, we observed no statistically significant relationship between the variables and the antibodies analyzed, regardless of the region evaluated. However, there was a moderate positive correlation between cytoplasmic/membranous immunoexpressions of GLUT 1 in invasion front and in the tumor center (r = 0.679, p <0.001). Similarly, moderate positive correlation was found between the nuclear immunoexpressions of GLUT 1 in the invasion front and in the tumor center (r = 0.547, p <0.001). For GLUT 3, was also observed a moderate statistically significant positive correlation between cytoplasmic/membranous expression in tumor invasion front and in tumor center (r = 0.589, p <0.001). We also observed that the immunoreactivity for GLUT 1 was higher than GLUT 3 expression in invasion front (p <0.001) and tumor center (p <0.001). From these results, this study suggests that tumor hypoxia is a remarkable characteristic of the EC of the lower lip and GLUT 1 may be primarily responsible for glucose uptake into the interior of the malignant cells

Estudo imuno-histoquímico da expressão da glut-1 e mensuração do índice angiogênico (CD34) em adenomas pleomórficos, carcinomas adenóides císticos e carcinomas mucoepidermóides de glândulas salivares

The expression of glucose transporter protein 1 (GLUT-1), as well the angiogenesis has been associated to clinical behavior and aggressiveness in tumors of various origin. It is believed that the expression of this protein denotes metabolic demand of the tumor cells and, thus its influence upon the formation of new blood vessels. Pleomorphic adenoma (PA) and the adenoid cystic carcinoma (ACC) and mucoepidermoid carcinoma (MEC) represent, respectively, the most commom benign and malignant tumors of salivary glands. The aim of this study was to analyze and compare the immunohistochemical expression of GLUT-1 and its correlation with angiogenesis in cases of PAs, ACCs and MECs considering their histological grades. The sample consisted of 20 PAs, 20 ACCs and 10 MECs. The cases were analyzed and classified according to their histological grades. The expression of GLUT-1 was evaluated in the parenchyma lesions, establishing the percentage of immunopositive cells, according to the following scores: 0 (no cell immunomarked), 1 (up to 25% of tumor cells immunostained), 2 (25 - 50% of tumor cells immunostained) and 3 (more than 50% of tumor cells immunostained). The angiogenic index was analyzed by counting the microvessels immunostained by anti-CD34 antibody, in 5 fields (200X). The analysis of the expression of GLUT-1 in tumor parenchyma showed statistically significant differences between benign and malignant groups (p = 0.022). The average number of microvessels in PAs was 40.4, 21.2 in ACCs and 66.5 in MECs, with significant differences between groups (p <0.001). When compared to the expression of GLUT-1 and angiogenic index as a whole, there was no significant correlation between the number of microvessels and the expression of GLUT-1 (r = 0.211, p = 0.141). In conclusion, the results of this study suggest not only that differences in biological behavior between PAs, ACCs and MECs may be associated to the expression of GLUT-1, but also that benign and malignant salivary gland present differences in the average number of microvessels, with higher levels considered more aggressive tumors. Furthermore, the number of newly formed microvessels can be independent of the metabolic demand of the tumor cells

Estudo da expressão imuno-histoquímica das MMPs -2, -7, -9 e -26 e TIMPs -1 e -2 em adenomas pleomórficos e carcinomas adenóides císticos de glândulas salivares menores

The balance between the expression of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) has been related to various physiological and pathological processes, including salivary gland morphogenesis and tumor invasion and metastasis processes. Pleomorphic adenoma (PA) and adenoid cystic carcinoma (ACC) respectively represent benign and malignant neoplasias of salivary glands. Although they share the same cell origin, they present distinct biological behavior. The aim of this study was to compare the immunohistochemical expression of MMPs -2, -7, -9 and -26, and of TIMPs -1 and -2, in cases of PA and ACC of minor salivary glands. Twenty cases of PA and twenty cases of ACC were assessed according to the presence, intensity and location of MMPs and TIMPs in the tumor parenchyma. Most of the PAs and ACCs presented a high expression of MMP -2, -7, -9 and -26 and of TIMP -1 and -2, predominantly located in tumor cells. There was no significant difference in the expression of MMPs -2 (p=0.359), -7 (p=0.081) and -26 (p=0.553), as well as of TIMPs -1 (p=0.657) and -2 (p=0.248), between the parenchyma of PAs and ACCs. However, MMP-9 showed a significant difference of expression between the two tumors, with the ACC showing more intense marking for this gelatinase (p=0.041). The strong expression of MMP-9 observed in the parenchyma suggests that this gelatinase may play an important role in the biological behavior of these tumors. On the other hand, although there was no significant difference between the marking of MMP -2, 7 and 26 in the studied tumors, the data, when analyzed as a whole, suggest that these proteases may take part in the process of tissue remodeling in both tumors, but do not present a direct relation with the pattern of aggressiveness of ACC. Nonetheless, matrilisins may indirectly influence the behavior of this tumor due to their capacity of activating MMP-9, strongly expressed in the parenchyma of ACC

Expressão imuno-histoquímica das integrinas α2ß1, α3ß, e α5ß1, em carcinoma epidermóide de lábio inferior e língua

The unpredictable biologic behavior of the oral squamous cells carcinoma has determined extensive research on the evolution of such tumor. Due to the existing relation between the outer cell matrix and the tumor cells, the integrins have been used as markers in the predictive study of the cell behavior. This study aims to analyze immunohistochemically the expression of the integrin α2β1, α3β1, and α5β1 connections for the collagen, the laminin and the fibronectin respectively in 15 cases of squamous cells carcinoma from the lower lip and 15 from the tongue, with different scores of malignance grading. A predominantly diffuse, cytoplasm and granular immunological marking was observed in the majority of the analyzed cases. According to the marking intensity, integrin α2β1 appeared positive in 80% of the lip and in 93,3% of the tongue cases. The immunological reactivity of integrin α3β1 was classified as positive in 60% of both the tongue and lip cases. For this integrin, 20% and 33.3% of the tongue and lip cases, respectively, were negative. In relation to integrin α5β1 the intensity was classified as positive in 53,3% of the cases and strongly positive in 46,7% of those located in the lip. In the tongue carcinomas, the intensity was positive in 46,7% of the cases and strongly positive in 53,3%. The statistic analysis did not show any significant differences or correlation of expression between these integrins nor between the anatomical sites or between different scores of malignancy grading. The expressive immunological marking of the integrins, α2β1, α3β1, and α5β1 in the studied cases of squamous cell carcinomas leads us to think of a great participation of these proteins in oral carcinogenesis; however, our results do not allow us to correlate its expression as an indicator of variations in the biological behavior of this neoplasia