Marcos Paulo Machado Thomé

Acridine orange (AO) is a cell permeable green fluorophore that can be protonated and trapped in Acidic Vesicular Organelles (AVOs). Its metachromatic shift to red fluorescence is concentration-dependent and, therefore, AO fluoresces red in AVOs, such as autolysosomes. This makes AO staining a quick, accessible and reliable method to assess the volume of AVOs, which increases upon autophagy induction. Here we describe a ratiometric analysis of autophagy using AO, considering the red-to-green fluorescence intensity ratio (R/GFIR) to quantify flow cytometry and fluorescence microscopy data of AO-stained cells. This method measured with accuracy the increase in autophagy induced by starvation or rapamycin and the blockage by bafilomycin A1 or the knockdown of Beclin1 or ATG7. Results obtained with AO, considering R/GFIR, correlated with LC3-I to LC3-II conversion, SQSTM1 degradation and GFP-LC3 puncta formation, thus validating this assay to be used as an initial and quantitative method for evaluating the late step of autophagy in individual cells, complementing other methods.

Temozolomide (TMZ) is the most widely used drug to treat glioblastoma (GBM), which is the most common and aggressive primary tumor of the Central Nervous System and one of the hardest challenges in oncotherapy. TMZ is an alkylating agent that induces autophagy, apoptosis and senescence in GBM cells. However, therapy with TMZ increases survival after diagnosis only from 12 to 14.4 months, making the development of combined therapies to treat GBM fundamental. One candidate for GBM therapy is Resveratrol (Rsv), which has additive toxicity with TMZ in several glioma cells in vitro and in vivo. However, the mechanism of Rsv and TMZ additive toxicity, which is the aim of the present work, is not clear, especially concerning cell cycle dynamics and long term effects.Glioma cell lines were treated with Rsv and TMZ, alone or in combinations, and the induction and the role of autophagy, apoptosis, cell cycle dynamics, protein expression and phosphorylation status were measured. We further evaluated the long term senescence induction and clonogenic capacity.As expected, temozolomide caused a G2 cell cycle arrest and extensive DNA damage response. Rsv did not reduced this response, even increasing pATM, pChk2 and gammaH2Ax levels, but abrogated the temozolomide-induced G2 arrest, increasing levels of cyclin B and pRb(S807/811) and reducing levels of pWee1(S642) and pCdk1(Y15). This suggests a cellular state of forced passage through G2 checkpoint despite large DNA damage, a scenario that may produce mitotic catastrophe. Indeed, the proportion of cells with high nuclear irregularity increased from 6 to 26% in 48 h after cotreatment. At a long term, a reduction in clonogenic capacity was observed, accompanied by a large induction of senescence.The presence of Rsv forces cells treated with TMZ through mitosis leading to mitotic catastrophe and senescence, reducing the clonogenic capacity of glioma cells and increasing the chronic effects of temozolomide.

Autophagy and senescence have been described as central features of cell biology, but the interplay between these mechanisms remains obscure. Using a therapeutically relevant model of DNA damage-induced senescence in human glioma cells, we demonstrated that acute treatment with temozolomide induces DNA damage, a transitory activation of PRKAA/AMPK-ULK1 and MAPK14/p38 and the sustained inhibition of AKT-MTOR. This produced a transient induction of autophagy, which was followed by senescence. However, at the single cell level, this coordinated transition was not observed, and autophagy and senescence were triggered in a very heterogeneous manner. Indeed, at a population level, autophagy was highly negatively correlated with senescence markers, while in single cells this correlation did not exist. The inhibition of autophagy triggered apoptosis and decreased senescence, while its activation increased temozolomide-induced senescence, showing that DNA damage-induced autophagy acts by suppressing apoptosis.

Hepatocellular carcinoma is the most prevalent primary liver tumor and is among the top ten cancer that affect the world population. Its development is related, in most cases, to the existence of chronic liver injury, such as in cirrhosis. The knowledge about the correlation between chronic inflammation and cancer has driven new researches with anti-inflammatory agents that have potential for the development of antitumor drugs. Gallic acid is a phenolic acid found in many natural products and have shown anti-inflammatory, anti-tumor, anti-mutagenic and antioxidant actions. The purpose of this study was to investigate the effect of gallic acid on acute and chronic cell proliferation and inflammatory parameters of hepatocellular carcinoma cells (HepG2), as well as to investigate the mechanisms involved. Results showed that the gallic acid decreased the proliferation of HepG2 cells in a dose-dependent manner (Trypan blue exclusion assay), without causing necrosis (LDH assay). We observed a significant increase in the percentage of small and regular nuclei (Nuclear Morphometric Analysis assay), a significant induction of apoptosis by Annexin V-FITC and PI assay and no interference with the cell cycle using the FITC BrdU Flow Kit. We observed a significant reduction in the levels of IL-8 and increased levels of IL-10 and IL-12 (Cytometric Bead Array Human Inflammation Assay). Furthermore, gallic acid caused no cancer cells regrowth at a long term (Cumulative Population Doubling assay). According to these results, gallic acid showed a strong potential as an anti-tumor agent in hepatocellular carcinoma cells.

Autophagy is a catabolic process that is largely regulated by extracellular and intracellular signaling pathways that are central to cellular metabolism and growth. Mounting evidence has shown that ion channels and transporters are important for basal autophagy functioning and influence autophagy to handle stressful situations. Besides its role in cell proliferation and apoptosis, intracellular Ca²⁺ is widely recognized as a key regulator of autophagy, acting through the modulation of pathways such as the mechanistic target of rapamycin complex 1, calcium/calmodulin-dependent protein kinase kinase 2, and protein kinase C. Proper spatiotemporal Ca²⁺ availability, coupled with a controlled ionic flow among the extracellular milieu, storage compartments, and the cytosol, is critical in determining the role played by Ca²⁺ on autophagy and on cell fate. The crosstalk between Ca²⁺ and autophagy has a central role in cellular homeostasis and survival during several physiologic and pathologic conditions. Here we review the main findings concerning the mechanisms and roles of Ca²⁺-modulated autophagy, focusing on human disorders ranging from cancer to neurologic diseases and immunity. By identifying mechanisms, players, and pathways that either induce or suppress autophagy, new promising approaches for preventing and treating human disorders emerge, including those based on the modulation of Ca²⁺-mediated autophagy.

Glioblastoma multiforme is the most aggressive type of primary brain tumor associated with few therapeutic opportunities and poor prognosis. In this study, we evaluated the efficacy of combining temozolomide (TMZ) with suberoylanilide hydroxamic acid (SAHA) - a specific histone deacetylases inhibitor - in glioma models in vitro and in vivo. In glioma cell lines, combined TMZ/SAHA promoted more cytotoxicity, G2/M arrest and apoptosis than either drugs alone. G2/M arrest was detected as soon as 24 h post drug exposure and preceded apoptosis, which occurred from 72 h treatment. TMZ and SAHA, alone or combined, also stimulated autophagy as evaluated by means of acridine orange staining and immunodetection of LC3I-II conversion and p62/SQSTM1 degradation. Time-course of autophagy accompanied G2/M arrest and preceded apoptosis, and blockage of late steps of autophagy with chloroquine (CQ) augmented SAHA/TMZ toxicity leading to apoptosis. In orthotopic gliomas in vivo, combined SAHA/TMZ showed better antitumor efficacy than either drugs alone, and adding CQ to the regimen improved antiglioma effects of SAHA and TMZ monotherapies without further benefit on combined SAHA/TMZ. In summary, the herein presented data suggest that autophagy acts as a protective response that impairs efficacy of SAHA and TMZ. Inhibiting autophagy termination with CQ may offer means to improve antitumor effects of SAHA and TMZ in gliomas and possibly other cancers.

Autophagy is a cellular bulk degradation process used as an alternative source of energy and metabolites and implicated in various diseases. Inefficient autophagy in nutrient-deprived cancer cells would be beneficial for cancer therapy making its modulation valuable as a therapeutic strategy for cancer treatment, especially in combination with chemotherapy. Dipyridamole (DIP) is a vasodilator and antithrombotic drug. Its major effects involve the block of nucleoside uptake and phosphodiestesase inhibition, leading to increased levels of intracellular cAMP. Here we report that DIP increases autophagic markers due to autophagic flux blockage, resembling autophagosome maturation and/or closure impairment. Treatment with DIP results in an increased number of autophagosomes and autolysosomes and impairs degradation of SQSTM1/p62. As blockage of autophagic flux decreases the recycling of cellular components, DIP reduced the intracellular ATP levels in cancer cells. Autophagic flux blockage was neither through inhibition of lysosome function nor blockage of nucleoside uptake, but could be prevented by treatment with a PKA inhibitor, suggesting that autophagic flux failure mediated by DIP results from increased intracellular levels of cAMP. Treatment with DIP presented antiproliferative effects in vitro alone and in combination with chemotherapy drugs. Collectively, these data demonstrate that DIP can impair autophagic degradation, by preventing the normal autophagosome maturation, and might be useful in combination anticancer therapy.

Vincristine (VCR) is a classical chemotherapeutic that has been revisited to treat refractory solid tumors producing encouraging results. VCR binds to tubulin and decreases the rate of microtubule dynamics, thus triggering many cellular responses and behaviors. However, the dynamics of these responses and fates are uncharacterized. This study combined systems biology approaches with acute and long-term in vitro experiments to predict key pathways and mechanisms associated with cell fates during and after VCR treatment. Glioblastoma (GBM) cells were treated with clinically relevant doses of VCR, and interconnected cell fates were explored. A correlation matrix based on experimental cell analysis reported strong negative correlations between cell number, nuclear irregularities, senescence, or apoptosis, depending on the cells' genetic makeup and treatment regimen. P53 would be essential in all analyzed processes according to topological network analysis. Furthermore, despite the high acute sensitivity, both cell lines re-growth in the long term after a single VCR treatment, especially in those populations with high levels of autophagy. These multiple responses may also be triggered in patients' exposed tumors, which should be considered to allow the rational design of VCR protocols, including modulators of the cell fates and pathways mentioned above.

Epstein-Barr virus (EBV) is a widely distributed gamma-herpesvirus that has been associated with various cancers mainly from lymphocytic and epithelial origin. Although EBV-mediated oncogenesis has been associated with viral oncogenes expressed during latency, a growing set of evidence suggested that antiviral treatments directed against EBV lytic phase may contribute to prevent some forms of cancers, including EBV-positive Post-Transplant Lymphoproliferative Diseases. It is shown here that dipyridamole (DIP), a safe drug with favorable and broad pharmacological properties, inhibits EBV reactivation from B-cell lines. DIP repressed immediate early and early genes expression mostly through its ability to inhibit nucleoside uptake. Considering its wide clinical use, DIP repurposing could shortly be evaluated, alone or in combination with other antivirals, to treat EBV-related diseases where lytic replication plays a deleterious role.

Alkylating agents damage DNA and proteins and are widely used in cancer chemotherapy. While cellular responses to alkylation-induced DNA damage have been explored, knowledge of how alkylation affects global cellular stress responses is sparse. Here, we examined the effects of the alkylating agent methylmethane sulfonate (MMS) on gene expression in mouse liver, using mice deficient in alkyladenine DNA glycosylase (Aag), the enzyme that initiates the repair of alkylated DNA bases. MMS induced a robust transcriptional response in wild-type liver that included markers of the endoplasmic reticulum (ER) stress/unfolded protein response (UPR) known to be controlled by XBP1, a key UPR effector. Importantly, this response is significantly reduced in the Aag knockout. To investigate how AAG affects alkylation-induced UPR, the expression of UPR markers after MMS treatment was interrogated in human glioblastoma cells expressing different AAG levels. Alkylation induced the UPR in cells expressing AAG; conversely, AAG knockdown compromised UPR induction and led to a defect in XBP1 activation. To verify the requirements for the DNA repair activity of AAG in this response, AAG knockdown cells were complemented with wild-type Aag or with an Aag variant producing a glycosylase-deficient AAG protein. As expected, the glycosylase-defective Aag does not fully protect AAG knockdown cells against MMS-induced cytotoxicity. Remarkably, however, alkylation-induced XBP1 activation is fully complemented by the catalytically inactive AAG enzyme. This work establishes that, besides its enzymatic activity, AAG has noncanonical functions in alkylation-induced UPR that contribute to cellular responses to alkylation.

Caveolin-1 (Cav-1) is an integral membrane protein present in all organelles, responsible for regulating and integrating multiple signals as a platform. Mitochondria are extremely adaptable to external cues in chronic liver diseases, and expression of Cav-1 may affect mitochondrial flexibility in hepatic stellate cells (HSCs) activation. We previously demonstrated that exogenous expression of Cav-1 was sufficient to increase some classical markers of activation in HSCs. Here, we aimed to evaluate the influence of exogenous expression and knockdown of Cav-1 on regulating the mitochondrial plasticity, metabolism, endoplasmic reticulum (ER)-mitochondria distance, and lysosomal activity in HSCs. To characterize the mitochondrial, lysosomal morphology, and ER-mitochondria distance, we perform transmission electron microscope analysis. We accessed mitochondria and lysosomal networks and functions through a confocal microscope and flow cytometry. The expression of mitochondrial machinery fusion/fission genes was examined by real-time polymerase chain reaction. Total and mitochondrial cholesterol content was measured using Amplex Red. To define energy metabolism, we used the Oroboros system in the cells. We report that GRX cells with exogenous expression or knockdown of Cav-1 changed mitochondrial morphometric parameters, OXPHOS metabolism, ER-mitochondria distance, lysosomal activity, and may change the activation state of HSC. This study highlights that Cav-1 may modulate mitochondrial function and structural reorganization in HSC activation, being a potential candidate marker for chronic liver diseases and a molecular target for therapeutic intervention.

Glioblastoma (GBM) is the most aggressive and lethal among the primary brain tumors, with a low survival rate and resistance to radio and chemotherapy. The P2Y12 is an adenosine diphosphate (ADP) purinergic chemoreceptor, found mainly in platelets. In cancer cells, its activation has been described to induce proliferation and metastasis. Bearing in mind the need to find new treatments for GBM, this study aimed to investigate the role of the P2Y12R in the proliferation and migration of GBM cells, as well as to evaluate the expression of this receptor in patients’ data obtained from the TCGA data bank. Here, we used the P2Y12R antagonist, ticagrelor, which belongs to the antiplatelet agent’s class. The different GBM cells (cell line and patient-derived cells) were treated with ticagrelor, with the agonist, ADP, or both, and the effects on cell proliferation, colony formation, ADP hydrolysis, cell cycle and death, migration, and cell adhesion were analyzed. The results showed that ticagrelor decreased the viability and the proliferation of GBM cells. P2Y12R antagonism also reduced colony formation and migration potentials, with alterations on the expression of metalloproteinases, and induced autophagy in GBM cells. Changes were observed at the cell cycle level, and only the U251 cell line showed a significant reduction in the ADP hydrolysis profile. TCGA data analysis showed a higher expression of P2Y12R in gliomas samples when compared to the other tumors. These data demonstrate the importance of the P2Y12 receptor in gliomas development and reinforce its potential as a pharmacological target for glioma treatment.

Cryptococcus neoformans and Cryptococcus gattii are the etiological agents of cryptococcosis, a high mortality disease. The development of such disease depends on the interaction of fungal cells with macrophages, in which they can reside and replicate. In order to dissect the molecular mechanisms by which cryptococcal cells modulate the activity of macrophages, a genome-scale comparative analysis of transcriptional changes in macrophages exposed to Cryptococcus spp. was conducted. Altered expression of nearly 40 genes was detected in macrophages exposed to cryptococcal cells. The major processes were associated with the mTOR pathway, whose associated genes exhibited decreased expression in macrophages incubated with cryptococcal cells. Phosphorylation of p70S6K and GSK-3β was also decreased in macrophages incubated with fungal cells. In this way, Cryptococci presence could drive the modulation of mTOR pathway in macrophages possibly to increase the survival of the pathogen.

Aves sao bioindicadores utilizados para a deteccao de alteracoes ambientais, e o estudo da biodiversidade destes organismos-chave torna-se essencial para tal papel, porem pouco se sabe das especies que ocorrem no Noroeste Fluminense, um ambiente com alto indice de degradacao. Sendo assim, o presente trabalho apresenta um inventario da avifauna ocorrente em areas degradadas localizadas no Noroeste do Estado do Rio de Janeiro. O levantamento ocorreu durante doze meses. Foi realizado em duas areas previamente delimitadas da zona rural pertencente ao Municipio de Natividade, RJ, totalizando 154 horas de observacoes em cada area e uma lista de 76 especies, com as maiores riquezas pertencente as familias Tyrannidae e Thraupidae. Dos habitos alimentares prevaleceram o insetivoro, frugivoro e onivoro.

As infeccoes parasitarias ainda apresentam-se em elevados indices, principalmente nos paises subdesenvolvidos, assim como naqueles em desenvolvimento. Neste contexto, as criancas constituem o grupo mais atingido, devido a alguns fatores como: imunidade, higiene pessoal e outros. Sendo assim, o presente estudo objetivou a investigacao dos indices parasitarios nos alunos da Escola Municipal Antonio Ferreira da Fonseca, localidade rural do municipio de Italva-RJ. O estudo foi realizado atraves dos resultados de exames parasitologicos realizados no Laboratorio da Faculdade Redentor em Itaperuna-RJ. Foram utilizados os metodos de Hoffman e de Willis. Foram examinadas 38 amostras, e destas verificou-se uma prevalencia de 71,05% de positividade, sendo a Entamoeba histolytica o protozoario mais frequente (42,1%), seguido pela Giardia lamblia (21,05%) e Entamoeba coli (5,26%). Concluiu-se que o indice parasitario nos escolares e elevado, e que a prevalencia de infeccao foi pelo protozoario Entamoeba histolytica.

Cyphocharax gilbert is an abundant species of fish in lagoons and rivers of northern Rio de Janeiro State, been important to the low-income populations of Campos dos Goytacazes city and adjacent regions. The present study shows the prevalence and the intensity of infestation of Riggia paranensis on C. gilbert from de Cima lagoon and Feia lagoon and from Itabapoana River, tests if the parasite affects the body size of infested fish, and evaluates if the parasitism affect the fish market activities. The results show that the prevalence of infestation was higher in Itabapoana River (between 49% and 64%), lower in the de Cima lagoon (between 2.6% and 2.8%) and intermediate in Feia lagoon (3.3% and 3.7%). Most of host fish presented only one parasite. Fish with two parasites were observed at Itabapoana River in 10% of the infested fish. The infested females were significantly bigger (P ≤ 0.05) than the no infested ones. Only in the Itabapoana River the consumers reject the infested fish. The high frequency of R. paranensis in C. gilbert population from the middle Itabapoana River might be a secondary impact of the exotic fish species introduction.

Devido os ciclistas necessitarem de um alto aporte de oxigenio para a realizacao de uma competicao eficaz o presente estudo objetivou avaliar se os indices de hematocrito e eritropoetina aumentam com a pratica de exercicio fisico como o ciclismo, em relacao a pessoas sedentarias. O estudo foi submetido e aprovado pelo comite de etica em pesquisa da Faculdade Redentor em Itaperuna. Os participantes da pesquisa apresentaram faixa etaria variando entre 23 a 54 anos, assinaram um termo de consentimento livre e esclarecido, preencheram um questionario e em seguida foi realizado a puncao venosa e retirada de 5mL de sangue antes e imediatamente apos a competicao. Os resultados indicam que nao houve diferenca estatistica dos resultados de hematocrito entre sedentarios (42,33/±2,72) e ciclistas pre (43,33/±2,29) e entre este ultimo e ciclistas pos (43,00/±1,91) para p ≤ 0,05. Os resultados referentes a eritropoetina tambem nao se mostraram significativos entre o grupo sedentario (7,28/±2,49) e ciclistas pre (9,44/±4,37)  e entre este ultimo comparado aos ciclistas pos (8,95/±3,73). Entretanto, a serie branca mostrou-se alterada apos a competicao. Logo, conclui-se que o tempo de coleta nao possibilitou averiguar diferencas significativas das variaveis entre os grupos estudados.

Most rural areas in Brazil do not receive treated water, and the population of these areas uses springs or wells for their daily consumption without control potability. Thus, the water becomes a vector of transmission of diseases, caused mostly by enteric pathogens origin. Therefore, the aim of this study was to verify the microbiological quality of water from two springs in the rural municipality of Varre-Sai, RJ. Samples were collected for seven months, and analyzed by the Multiple Tube Method for detection of total and thermotolerant coliforms. Contamination was found in four samples of point 01, maximum 1600 NMP/100mL, while the point 02 showed contamination in only two samples with a maximum of 23 NMP/100mL. The presence of protective structures in point 02 and their absence in point 01 may have contributed to these differences. The construction of structures protect of the springs associated with the preservation of natural vegetation can positively influence water quality.

Objetivou-se verificar a fauna ectoparasitaria de Helostoma temminkii (beijador) e Carassius Auratus (kinguio) cultivados em uma piscicultura no municipio de Muriae- MG. Para tanto, foram utilizados os indices de prevalencia e intensidade (minima, maxima e media) para cada parasita encontrados nos hospedeiros analisados. Ao todo foram coletados 36 peixes de ambas as especies. Dos quais 41,66% exemplares dos H. temminkii apresentaram sinais clinicos e 22,21% eram sadios, e da especie C. Auratus, 19,44% individuos apresentaram sinais clinicos e 16,66% sadios. Todos os peixes analisados estavam parasitados por uma ou mais especies de parasitas. A maior prevalencia foi para Epistylis sp presente no tegumento, considerando os exemplares de ambas as especies. Quanto aos parasitas obrigatorios as monogeneas obtiveram maior prevalencia enquanto o crustaceo Argulus sp. apresentou menor prevalencia

A vegetacao de mata ciliar constitui uma das principais protecoes dos recursos hidricos, contribuindo tanto para a manutencao da qualidade da agua, quanto para o quantitativo desse recurso. O presente estudo teve por objetivo descrever o grau de desenvolvimento de um reflorestamento as margens do ribeirao de Varre-Sai, no municipio de Varre-Sai. Os dados foram obtidos atraves de observacao da cobertura vegetal e amostragem do quantitativo de serapilheira e analise quimica de amostras de solo. O presente estudo possibilitou detectar que a vegetacao apresenta especies arboreas com porte de estagio intermediarios de sucessao ecologica. Foi ainda detectado presenca de serapilheira o que viabiliza uma melhor fertilidade e retencao de agua no solo, fato tambem associado ao maior teor nutricional encontrado no solo quando comparado com amostras de areas adjacentes com cobertura de pastagens. Atraves da analise dos dados obtidos pode-se afirmar que o reflorestamento esta em pleno desenvolvimento e que a vegetacao restabelecida esta desempenhando sua funcao ecologica de protecao e viabilidade do recurso hidrico que margeia.

As enteroparasitoses constituem-se um grave problema de saude publica. Sendo assim, o presente estudo teve por objetivo a investigacao dos indices parasitarios em alunos da Escola Municipalizada Deputado Salim Simao em Santo Antonio de Padua – RJ. Das 54 amostras analisadas, verificou-se uma prevalencia de 61,1% de positividade, sendo a Giardia lamblia o protozoario mais frequente (78,8%). Desta forma, faz-se necessario uma melhoria nos programas de controle, na conscientizacao dos pais sobre o assunto, bem como no saneamento basico, proporcionando uma melhoria na qualidade de vida da populacao.

Os ictioparasitas, em situacao de desequilibrio ambiental podem trazer serios danos as populacoes de seus hospedeiros, acarretando a reducao de estoques pesqueiros naturais e de cultivo. Portanto, este trabalho teve como objetivo verificar a fauna parasitaria do tegumento da especie Cyphocharax gilbert obtidos pelas praticas de captura utilizadas por pescadores em dois riachos da porcao media da sub-bacia do rio Muriae. Ao todo, foram capturados 45 hospedeiros nos dois pontos de coletas, sendo 29 individuos no Riacho do Limoeiro e 16 individuos no Riacho da Paixao. Nos hospedeiros, foram encontrados diferentes tipos de parasitas pertencentes ao filo Protozoa (3 generos, 3 especies), classe Monogenoedea (larva), classe Crustacea (2 generos, 2 especies). Foram identificados para o Riacho do Limoero Mixobolus sp, Epystilis sp,  larva de Monogenea, Trichodina sp. , Riggia paranensis, e Lernea sp. No Riacho da Paixao foram identificados larva de Monogenea e Trichodina sp. A variacao de diversidade de parasitas entre os ambientes pode estar relacionada as diferencas naturais entre os locais estudadoss e a existencia de duas populacoes de hospedeiros diferentes, uma para cada riacho.

Os lagos urbanos desempenham funcao estetica e de equilibrio da umidade nas cidades, promovendo o bem estar. Frequentemente os lagos sao povoados por especies exoticas de peixes, que por sua vez trazem consigo parasitas que podem levar disturbios as especies naturais. Entao o objetivo deste estudo foi verificar a fauna parasitaria das brânquias de Oreocrhomis niloticus em um lago urbano no Municipio de Itaperuna – RJ. Para tanto realizou -se duas coletas (Fevereiro e Maio) totalizando 18 exemplares de O. niloticus . No mes de Fevereiro, apenas Cichlidogyrus tilapiae foi encontrado parasitando as brânquias de O. niloticus apresentando 100% de prevalencia. Na coleta realizada em Maio foram encontrados os monogenoides Cichlidogyrus sclerosus (88,88%) e Cichlidogyrus tilapiae (44,44%) . Sugere-se que, por se tratar de um lago relativamente raso, O. niloticus apresenta adensamento populacional o que favorece o aumento da ocorrencia de parasitas da mesma forma tal qual e encontrada em pisciculturas.

As tecnicas de nucleacao tornam-se uma acao vital para a recuperacao de areas degradadas, pois propiciam reconstituir um ambiente o mais proximo possivel da sua condicao original. Este estudo teve por objetivo analisar a capacidade de uma tecnica nucleadora para recuperacao ecologica de uma area de recarga hidrica em topo de morro com 1,4 ha. O projeto de restauracao da area de recarga foi implantado em dezembro de 2011, em uma propriedade rural, localizada a 2,5 km do centro urbano de Raposo 7o distrito de Itaperuna no estado do rio de Janeiro. A propriedade esta inserida em uma paisagem cuja matriz e a pastagem degradada. Foi utilizada a tecnica de nucleacao “nucleos de Anderson, que consistem no plantio de especies pioneiras e secundarias de forma consorciada. As avaliacoes para monitoramento da restauracao foram realizadas no mes de maio, julho e outubro de 2012, com dados de alturas e diâmetros de caules. Foi observado que todas as especies inseridas tiveram crescimento perceptivel, mesmo em periodo de estiagem, o que e um indicio de sucesso no processo de recuperacao. Apesar do presente estudo ter transcorrido em curto periodo de tempo, os dados demonstram uma perspectiva otimista em relacao ao restabelecimento da vegetacao no local. As especies secundarias Angico ( Anadenanthera macrocarpaI ) e Inga ( Inga marginata ) se mostraram propicias a integracao da pratica de reflorestamento, apresentando boa implementacao na area estudada. As tecnicas de nucleacao pela sua essencia de simular o processo de sucessao ecologica natural, ainda podem favorecer o aumento da biodiversidade, contribuindo de forma expressiva para o equilibrio ecologico local.

Neste trabalho objetivou-se analisar qual o melhor meio alternativo para o desenvolvimento de orquideas da especie Cattleya guttata Lindl.. Para esse fim foram utilizados tres meios nutritivos polpa de tomate, 10 gL -1 (T), polpa de mamao, 10 gL -1 (M),  e polpa de mamao mais tomate, 10 gL -1 , durante um periodo de 90 dias, analisando as variaveis altura da parte aerea, numero de raizes, comprimento da maior raiz, peso da massa fresca. Apos esse periodo observou-se que o meio nutritivo de mamao se mostrou mais eficaz nas varaveis altura da parte aerea, comprimento da maior raiz e peso da massa fresca, o meio de mamao + tomate obteve os melhores resultados na variavel numero de raizes. O meio nutritivo que mais se mostrou eficaz para o desenvolvimento da Cattley guttata Lindl. foi o de mamao, mas por apresentarem bons resultados nas variaveis estudadas, a utilizacao de meios alternativos para o crescimento e desenvolvimento de orquideas se mostra viavel para o desenvolvimento da plântula in vitro .

A presenca da tuberculose em sistemas prisionais e descrito como uma ameaca a saude publica. O presente estudo objetivou a determinar a prevalencia da Mycobacterium tuberculosis no sistema prisional Diomedes Vinhosa Muniz em Itaperuna, Rio de Janeiro. O estudo foi submetido e aprovado pelo comite de etica em pesquisa da Faculdade Redentor em Itaperuna e autorizado pela Secretaria de Estado de Administracao Penitenciaria (SEAP) do estado do Rio de Janeiro. Participaram da pesquisa 10% do total de detentos condenados, estes assinaram um termo de consentimento livre e esclarecido, preencheram um questionario socio economico e epidemiologico em seguida foi coletado o material biologico para realizacao da baciloscopia. Os resultados indicaram que 69% dos detentos analisados nao completaram o ensino fundamental, 30% e originario de comunidades faveladas, 55% possuem uma renda mensal de ate 2 salarios minimos, 79% consumiam tabaco, 64% relataram que tiveram tosse por mais de 3 semanas, 65% declaram a maconha como a droga ilicita preferencial e 51% usavam cocaina, 3% declararam se portador do HIV, 42% tiveram contato com individuos com a tuberculose e 3% dos detentos haviam a presenca da Mycobacterium tuberculosis . Logo, conclui-se que no sistema prisional Diomedes Vinhosa Muniz houve uma baixa prevalencia da M.ycobacterium tuberculosis.

Autophagy and senescence have been described as central features of cell biology, but the interplay between these mechanisms remains obscure. Using a therapeutically relevant model of DNA damage-induced senescence in human glioma cells, we demonstrated that acute treatment with temozolomide induces DNA damage, a transitory activation of PRKAA/AMPK-ULK1 and MAPK14/p38 and the sustained inhibition of AKT-MTOR. This produced a transient induction of autophagy, which was followed by senescence. However, at the single cell level, this coordinated transition was not observed, and autophagy and senescence were triggered in a very heterogeneous manner. Indeed, at a population level, autophagy was highly negatively correlated with senescence markers, while in single cells this correlation did not exist. The inhibition of autophagy triggered apoptosis and decreased senescence, while its activation increased temozolomide-induced senescence, showing that DNA damage-induced autophagy acts by suppressing apoptosis.

Autophagy and senescence have been described as central features of cell biology, but the interplay between these mechanisms remains obscure. Using a therapeutically relevant model of DNA damage-induced senescence in human glioma cells, we demonstrated that acute treatment with temozolomide induces DNA damage, a transitory activation of PRKAA/AMPK-ULK1 and MAPK14/p38 and the sustained inhibition of AKT-MTOR. This produced a transient induction of autophagy, which was followed by senescence. However, at the single cell level, this coordinated transition was not observed, and autophagy and senescence were triggered in a very heterogeneous manner. Indeed, at a population level, autophagy was highly negatively correlated with senescence markers, while in single cells this correlation did not exist. The inhibition of autophagy triggered apoptosis and decreased senescence, while its activation increased temozolomide-induced senescence, showing that DNA damage-induced autophagy acts by suppressing apoptosis.

Autophagy and senescence have been described as central features of cell biology, but the interplay between these mechanisms remains obscure. Using a therapeutically relevant model of DNA damage-induced senescence in human glioma cells, we demonstrated that acute treatment with temozolomide induces DNA damage, a transitory activation of PRKAA/AMPK-ULK1 and MAPK14/p38 and the sustained inhibition of AKT-MTOR. This produced a transient induction of autophagy, which was followed by senescence. However, at the single cell level, this coordinated transition was not observed, and autophagy and senescence were triggered in a very heterogeneous manner. Indeed, at a population level, autophagy was highly negatively correlated with senescence markers, while in single cells this correlation did not exist. The inhibition of autophagy triggered apoptosis and decreased senescence, while its activation increased temozolomide-induced senescence, showing that DNA damage-induced autophagy acts by suppressing apoptosis.

Autophagy and senescence have been described as central features of cell biology, but the interplay between these mechanisms remains obscure. Using a therapeutically relevant model of DNA damage-induced senescence in human glioma cells, we demonstrated that acute treatment with temozolomide induces DNA damage, a transitory activation of PRKAA/AMPK-ULK1 and MAPK14/p38 and the sustained inhibition of AKT-MTOR. This produced a transient induction of autophagy, which was followed by senescence. However, at the single cell level, this coordinated transition was not observed, and autophagy and senescence were triggered in a very heterogeneous manner. Indeed, at a population level, autophagy was highly negatively correlated with senescence markers, while in single cells this correlation did not exist. The inhibition of autophagy triggered apoptosis and decreased senescence, while its activation increased temozolomide-induced senescence, showing that DNA damage-induced autophagy acts by suppressing apoptosis.

Autophagy and senescence have been described as central features of cell biology, but the interplay between these mechanisms remains obscure. Using a therapeutically relevant model of DNA damage-induced senescence in human glioma cells, we demonstrated that acute treatment with temozolomide induces DNA damage, a transitory activation of PRKAA/AMPK-ULK1 and MAPK14/p38 and the sustained inhibition of AKT-MTOR. This produced a transient induction of autophagy, which was followed by senescence. However, at the single cell level, this coordinated transition was not observed, and autophagy and senescence were triggered in a very heterogeneous manner. Indeed, at a population level, autophagy was highly negatively correlated with senescence markers, while in single cells this correlation did not exist. The inhibition of autophagy triggered apoptosis and decreased senescence, while its activation increased temozolomide-induced senescence, showing that DNA damage-induced autophagy acts by suppressing apoptosis.

Autophagy and senescence have been described as central features of cell biology, but the interplay between these mechanisms remains obscure. Using a therapeutically relevant model of DNA damage-induced senescence in human glioma cells, we demonstrated that acute treatment with temozolomide induces DNA damage, a transitory activation of PRKAA/AMPK-ULK1 and MAPK14/p38 and the sustained inhibition of AKT-MTOR. This produced a transient induction of autophagy, which was followed by senescence. However, at the single cell level, this coordinated transition was not observed, and autophagy and senescence were triggered in a very heterogeneous manner. Indeed, at a population level, autophagy was highly negatively correlated with senescence markers, while in single cells this correlation did not exist. The inhibition of autophagy triggered apoptosis and decreased senescence, while its activation increased temozolomide-induced senescence, showing that DNA damage-induced autophagy acts by suppressing apoptosis.

Autophagy and senescence have been described as central features of cell biology, but the interplay between these mechanisms remains obscure. Using a therapeutically relevant model of DNA damage-induced senescence in human glioma cells, we demonstrated that acute treatment with temozolomide induces DNA damage, a transitory activation of PRKAA/AMPK-ULK1 and MAPK14/p38 and the sustained inhibition of AKT-MTOR. This produced a transient induction of autophagy, which was followed by senescence. However, at the single cell level, this coordinated transition was not observed, and autophagy and senescence were triggered in a very heterogeneous manner. Indeed, at a population level, autophagy was highly negatively correlated with senescence markers, while in single cells this correlation did not exist. The inhibition of autophagy triggered apoptosis and decreased senescence, while its activation increased temozolomide-induced senescence, showing that DNA damage-induced autophagy acts by suppressing apoptosis.

Autophagy and senescence have been described as central features of cell biology, but the interplay between these mechanisms remains obscure. Using a therapeutically relevant model of DNA damage-induced senescence in human glioma cells, we demonstrated that acute treatment with temozolomide induces DNA damage, a transitory activation of PRKAA/AMPK-ULK1 and MAPK14/p38 and the sustained inhibition of AKT-MTOR. This produced a transient induction of autophagy, which was followed by senescence. However, at the single cell level, this coordinated transition was not observed, and autophagy and senescence were triggered in a very heterogeneous manner. Indeed, at a population level, autophagy was highly negatively correlated with senescence markers, while in single cells this correlation did not exist. The inhibition of autophagy triggered apoptosis and decreased senescence, while its activation increased temozolomide-induced senescence, showing that DNA damage-induced autophagy acts by suppressing apoptosis.

Esse estudo reporta a presenca de Henneguya sp. no sangue de Astyanax fasciatus de ambiente natural, em um corrego da sub-bacia do rio Muriae, pertencente a bacia do rio do Paraiba do Sul de Itaperuna, estado do Rio de Janeiro, Brasil. Oito coletas foram realizadas em outubro de 2009 e setembro de 2010, com 45 dias de intervalo, em cada coleta foram capturados dez especimes de Astyanax fasciatus dando um total de 80 lâminas de sangue. Esse estudo relata a presenca de Henneguya sp. em duas especimes coletadas em novembro de 2009. Essa baixa incidencia de parasitas ocorreu porque o genero Henneguya ja foi registrado em varios orgaos de especies do genero Astyanax . Embora os filamentos branquiais sejam de locais com maior tropismo e alta prevalencia, nao foram encontrados registros na literatura de parasitas com esse genero no sangue de peixes.

A hepatite B esta entre as doencas infecciosas que mais causam morbidade e mortalidade no mundo, tornando-se uma grande preocupacao entre os trabalhadores de diversas areas. Entre os profissionais da saude e estetica, a hepatite B configura-se como uma doenca ocupacional. Portanto, o objetivo do presente trabalho foi realizar um levantamento da contaminacao pelo virus da hepatite B, bem como fornecer informacoes a respeito dessa doenca para manicures do municipio de Itaperuna, RJ. Os resultados obtidos sugerem negatividade de casos com a presenca do HBsAg, porem, nao se pode deixar de ressaltar que 78% das entrevistadas nao sabem de forma completa os meios de transmissao da hepatite B e apenas 2% recebeu o esquema completo de vacinacao do virus,  mostrando que ainda existe falta de conhecimento acerca da doenca e prevencao tanto ocupacional como pessoal, sendo assim, importante o estudo desse grupo que tem se destacado tanto nos ultimos anos, podendo desta forma, informa-las sobre os riscos e as maneiras como elas podem estar se prevenindo.

A utilizacao de plantas medicinais e orientada por conhecimentos acumulados mediante a relacao direta com o meio ambiente. O objetivo deste estudo e descrever o conhecimento local da populacao do distrito de Catune, municipio de Tombos (MG) em relacao as plantas utilizadas na medicina tradicional e identificar se tal conhecimento e repassado para as novas geracoes. Foram realizadas 40 entrevistas com moradores locais, dos quais 11 (38%) apresentaram idade superior a 60 anos e vinte e nove entrevistados (72%) afirmaram utilizar recursos vegetais como fitoterapicos. O conhecimento local do uso de plantas medicinais nao e utilizado por geracoes mais jovens.

A utilização de plantas medicinais é orientada por conhecimentos acumulados mediante a relação direta com o meio ambiente. O objetivo deste estudo é descrever o conhecimento local da população do distrito de Catuné, município de Tombos (MG) em relação às plantas utilizadas na medicina tradicional e identificar se tal conhecimento é repassado para as novas gerações. Foram realizadas 40 entrevistas com moradores locais, dos quais 11 (38%) apresentaram idade superior a 60 anos e vinte e nove entrevistados (72%) afirmaram utilizar recursos vegetais como fitoterápicos. O conhecimento local do uso de plantas medicinais não é utilizado por gerações mais jovens.

tanques na irrigação de hortaliças criando um

Objetivou-se com o presente estudo analisar a diversidade e as condições viárias e fitossanitárias da arborização urbana no município de Aperibé – RJ. Foram analisadas as ruas sede e dos bairros que estão ao seu redor através de visitas semanais, onde foram feitas anotações em planilha e as árvores encontradas foram fotografadas e analisadas individualmente. Observou-se que apesar da diversidade encontrada, que há um desequilíbrio no que diz respeito à distribuição de uma mesma espécie arbórea. Das 30 famílias encontradas a Fabaceae foi a mais abundante, sendo que as espécies nativas foram suprimidas pelas exóticas e todas as árvores pesquisadas eram frutíferas. A Licania tomentosa Benth., a Mangifera indica L. e o Ficus benjamina L. foram as que mais apresentaram problemas fitossanitários. 98,8% das árvores estavam saudáveis e 94% não apresentaram problemas na raiz, mas muitas encontravam-se desprovidas de recuo e área livre ao seu redor, e os conflitos das árvores com a rede elétrica representou metade dos problemas viários. O inventário permitiu analisar as espécies arbóreas e forneceu um diagnóstico atualizado da arborização urbana do município, o que futuramente servirá como base para a manutenção e manejo da área por órgãos responsáveis.

TUNDISI, 2014).

Abstract Cryptococcus neoformans and Cryptococcus gattii are the etiological agents of cryptococcosis, a high mortality disease. The development of such disease depends on the interaction of fungal cells with macrophages, in which they can reside and replicate. In order to dissect the molecular mechanisms by which cryptococcal cells modulate the activity of macrophages, a genome-scale comparative analysis of transcriptional changes in macrophages exposed to Cryptococcus spp. was conducted. Altered expression of nearly 40 genes was detected in macrophages exposed to cryptococcal cells. The major processes were associated with the mTOR pathway, whose associated genes exhibited decreased expression in macrophages incubated with cryptococcal cells. Phosphorylation of p70S6K and GSK-3β was also decreased in macrophages incubated with fungal cells. In this way, Cryptococci presence could drive the modulation of mTOR pathway in macrophages possibly to increase the survival of the pathogen.

O uso de plantas no combate a doencas e tao antigo quanto a propria humanidade e, muitas vezes, e o unico recurso de muitas comunidades. Portanto, objetivou-se com este trabalho realizar o levantamento das plantas medicinais utilizadas pelos moradores do distrito de Agua Santa de Minas, Tombos-MG, verificando suas indicacoes terapeuticas. A coleta de dados foi realizada nos meses de maio e junho com aplicacao de questionarios semiestruturados. Para tal estudo, foram entrevistados trinta moradores do distrito, onde 87,67% eram do sexo feminino. Foi verificado que 76,66% fazem uso de plantas medicinais e foram levantadas 33 especies medicinais, sendo que 10 sao nativas e 23 sao exoticas. O Boldo (Plectranthus barbatus Andr) com 14,28% e a Hortela (Mentha spicata L.) com 8,33% foram as especies com maior numero de citacao. Em relacao as familias: Lamiaceae, Asteraceae, Apiaceae e Malvaceae foram as mais representativas. A forma de preparo mais utilizada foi o cha com 97,06% de citacoes, sendo que a folha e a parte mais selecionada para o preparo com 82,80%. Foram relacionadas plantas para 26 usos medicinais, a maioria foi para o tratamento da gripe (17,46%), colica menstrual (7,14%) e problemas relacionados ao estomago (7,14).

O uso de plantas no combate a doencas e tao antigo quanto a propria humanidade e, muitas vezes, e o unico recurso de muitas comunidades. Portanto, objetivou-se com este trabalho realizar o levantamento das plantas medicinais utilizadas pelos moradores do distrito de Agua Santa de Minas, Tombos-MG, verificando suas indicacoes terapeuticas. A coleta de dados foi realizada nos meses de maio e junho com aplicacao de questionarios semiestruturados. Para tal estudo, foram entrevistados trinta moradores do distrito, onde 87,67% eram do sexo feminino. Foi verificado que 76,66% fazem uso de plantas medicinais e foram levantadas 33 especies medicinais, sendo que 10 sao nativas e 23 sao exoticas. O Boldo (Plectranthus barbatus Andr) com 14,28% e a Hortela (Mentha spicata L.) com 8,33% foram as especies com maior numero de citacao. Em relacao as familias: Lamiaceae, Asteraceae, Apiaceae e Malvaceae foram as mais representativas. A forma de preparo mais utilizada foi o cha com 97,06% de citacoes, sendo que a folha e a parte mais selecionada para o preparo com 82,80%. Foram relacionadas plantas para 26 usos medicinais, a maioria foi para o tratamento da gripe (17,46%), colica menstrual (7,14%) e problemas relacionados ao estomago (7,14).

As areas umidas vem sofrendo alteracoes graduais ocasionadas pela agropecuaria convencional que substitui sua conformacao original por pastagens modificando as comunidades de aves que possuem importantes funcoes na manutencao do ecossistema. Portanto, foi escolhida uma paisagem modificada que inclui distintas classificacoes de areas umidas afim de averiguar a ocorrencia de aves aquaticas e a utilizacao do espaco pelas mesmas. No local foram constatadas especies consideradas incomuns e um casal da Garca real (Pilherodius pileatus), considerada Provavelmente Ameacada de Extincao para o Estado do Rio de Janeiro utilizando-se de um resquicio de mata riparia no entorno de um corrego. Isto permitiu constatar que esta area apesar de bastante alterada pela criacao extensiva de gado, e de grande importância para a conservacao da avifauna na regiao com um dos mais baixos indices de biodiversidade avicola do Estado.

Os riachos de altitude no Brasil tem um excelente potencial para a implantacao de usinas Geradoras de Energia (3MGwatts / h). No entanto, pouco se sabe sobre a ictiofauna nessas areas por varias razoes. Portanto, objetivou-se caracterizar a ictiofauna do riacho quilombo no periodo de seca, afluente do rio Paraiba do Sul. Os pontos de coleta foram distribuidos em Montante e Jusante, do projeto de implantacao. Ao todo, 47 exemplares, distribuidos em 11 especies, sendo Poecilia reticulata (exotica) a mais abundante. A maior riqueza foi atribuida a Jusante e observou-se quedas verticais na calha do corpo hidrico que poderiam constituir uma barreira natural a migracao de peixes a jusante. Quase todas as especies eram de pequeno porte, a excecao de Rhamdia quelen. A vegetacao nativa do entorno do riacho foi substituida por pastagens, suprimindo os microhabitats do corpo hidrico, o que contribuiu para a numero reduzido de peixes capturados. Por isso, recomenda-se a recuperacao da vegetacao nas margens do riacho antes e concomitante a possivel implantacao do empreendimento.

Abstract Alkylating agents damage DNA and proteins and are widely used in cancer chemotherapy. While the cellular responses to alkylation-induced DNA damage have been explored, knowledge of how alkylation damage affects global cellular stress responses is still sparse. Here, we examined the effects of the alkylating agent methylmethane sulfonate (MMS) on gene expression in mouse liver taking advantage of mice deficient in alkyladenine DNA glycosylase (Aag), the enzyme that initiates the repair of alkylated DNA bases. MMS induced a robust transcriptional response in wild-type liver that included markers of the endoplasmic reticulum (ER) stress/unfolded protein response (UPR) known to be controlled by the transcription factor XBP1, a key UPR effector. Importantly, this response is significantly reduced in the Aag knockout. To investigate a potential role for AAG in alkylation-induced UPR, the expression of UPR markers after MMS treatment was interrogated in human glioblastoma cell lines expressing different AAG levels. Alkylation induced the UPR in cells expressing AAG; conversely, AAG knock-down compromised UPR induction and led to a defect in XBP1 activation plus a decrease in the expression of the ER chaperone BiP. To verify that the DNA repair activity of AAG is required for this response, AAG knockdown cells were complemented with wild-type Aag or with a mutant version of the Aag gene producing a glycosylase-deficient AAG protein. As expected, the glycosylase-defective mutant Aag does not fully protect AAG knockdown cells against MMS-induced cytotoxicity. Remarkably, however, alkylation-induced XBP1 activation is fully complemented by the catalytically inactive AAG enzyme. This work establishes that, in addition to its enzymatic activity, AAG has non-canonical functions in alkylation-induced UPR that contribute to the overall cellular response to alkylation. Significance Statement Stress response pathways, such as the DNA damage response (DDR) and the UPR, are critical in both the etiology and treatment of cancer and other chronic diseases. Knowledge of an interplay between ER stress and genome damage repair is emerging, but evidence linking defective DNA repair and impaired ER stress response is lacking. Here, we show that AAG is necessary for UPR activation in response to alkylating agents. AAG-deficient mice and human cancer cells are impaired in alkylation-induced UPR. Strikingly, this defect can be complemented by an AAG variant defective in glycosylase activity. Our studies suggest AAG has non-canonical functions and identify AAG as a point of convergence for stress response pathways. This knowledge could be explored to improve cancer treatment.

As condicoes sanitarias e o manejo na incubacao, larvicultura e alevinagem do tambaqui Colossoma macropomum (Cuvier, 1818) (Osteichthyes: Serrasalmidae) no Estado do Amazonas / Marcos Paulo Machado Thome