Antioxidant effect of endothelin-1 receptor antagonist protects the rat kidney against chronic injury induced by hypertension and hyperglycemia / Efeito antioxidante de antagonista dos receptores de endotelina-1 protege ratos contra lesão renal crônica induzida por hipertensão e hiperglicemia

ABSTRACT Hypertension and Diabetes mellitus are the two main causes of chronic kidney disease that culminate in the final stage of kidney disease. Since these two risk factors are common and can overlap, new approaches to prevent or treat them are needed. Macitentan (MAC) is a new non-selective antagonist of the endothelin-1 (ET-1) receptor. This study aimed to evaluate the effect of chronic blockade of ET-1 receptor with MAC on the alteration of renal function observed in hypertensive and hyperglycemic animals. Genetically hypertensive rats were divided into control hypertensive (HT-CTL) group, hypertensive and hyperglycemic (HT+DIAB) group, and hypertensive and hyperglycemic group that received 25 mg/kg macitentan (HT-DIAB+MAC25) via gavage for 60 days. Kidney function and parameters associated with oxidative and nitrosative stress were evaluated. Immunohistochemistry for neutrophil gelatinase-associated lipocalin (NGAL), ET-1, and catalase in the renal cortex was performed. The HT+DIAB group showed a decrease in kidney function and an increase in NGAL expression in the renal cortex, as well as an increase in oxidative stress. MAC treatment was associated with attenuated ET-1 and NGAL production and increases in antioxidant defense (catalase expression) and nitric oxide production. In addition, MAC prevented an increase in oxidant injury (as measured by urinary hydroperoxide and lipid peroxidation), thus improving renal function. Our results suggest that the antioxidant effect of the ET-1 receptor antagonist MAC is involved in the improvement of kidney function observed in hypertensive and hyperglycemic rats.

RESUMO Hipertensão e Diabetes Mellitus figuram como as duas principais causas de doença renal crônica que culmina em doença renal terminal. Uma vez que os dois fatores de risco são comuns e podem se sobrepor, novas abordagens preventivas e terapêuticas se fazem necessárias. O macitentan (MAC) é um novo antagonista não-seletivo dos receptores da endotelina-1 (ET-1). O presente estudo teve como objetivo avaliar os efeitos do bloqueio crônico dos receptores da ET-1 com MAC sobre a alteração da função renal em animais hipertensos e hiperglicêmicos. Ratos geneticamente hipertensos foram divididos em grupos com animais hipertensos de controle (HT-CTL), hipertensos e hiperglicêmicos (HT+DIAB) e hipertensos e hiperglicêmicos tratados com 25 mg/kg de macitentan (HT-DIAB+MAC25) via gavagem por 60 dias. Foram avaliados função renal e parâmetros associados ao estresse oxidativo e nitrosativo. Exames de imunoistoquímica foram realizados para lipocalina associada à gelatinase neutrofílica (NGAL), ET-1 e catalase no córtex renal. O grupo HT+DIAB exibiu diminuição da função renal e aumento na expressão de NGAL no córtex renal, bem como estresse oxidativo aumentado. O tratamento com MAC foi associado a atenuação da produção de ET-1 e NGAL e maior ativação das defesas antioxidantes (expressão de catalase) e elevação da produção de óxido nítrico. Além disso, o MAC evitou exacerbação da lesão oxidante (medida por hidroperóxidos urinários e peroxidação lipídica), melhorando assim a função renal. Nossos resultados sugerem que o efeito antioxidante do antagonista dos receptores da ET-1 MAC esteja imbricado no aprimoramento da função renal observada em ratos hipertensos e hiperglicêmicos.

Genotype and expression of the enhanced gree fluorescent protein in LEW-Tg (EGFP) F455.5/Rrrc rats / Genótipo e expressão da proteína fluorescente verde melhorada em ratos da linhagem LEW-Tg (EGFP) F455.5/Rrrc

The Green fluorescent protein (GFP) was first described after being extracted from Aequorea victoria in 1987; Since then, GFP and its derivatives have been widely used in several experiments as cell and protein marker. In the present study it was verified the genotype of the offspring from crosses between heterozygote Lewis LEW-Tg (EGFP) F455.5/Rrrc rats and analyzed the expression of the enhanced green fluorescent protein (EGFP) in different cell types and genotypes. The genotype of the offspring was assessed by PCR and analysis of EGFP expression in different cells and genotypes, including mesenchymal stem cells (MSC) derived from adipose tissue and calvarial osteoblast cells. Expression of EGFP was verified by flow cytometry, fluorescence microscopy, and immunostaining. Through these methods, it was identified the genotypes of the offspring and determined the levels of expression of EGFP in two cell types. A difference in expression between the (EGFP +/+) and (EGFP +/-) genotypes was also observed in addition to the presence of autofluorescence. Further studies on the natural fluorescence of cells with the (EGFP +/-) genotype and that induced by presence of the EGFP are necessary.

A proteína fluorescente verde (GFP) foi descrita pela primeira vez após ter sido extraída de Aequorea victoria em 1987. Desde então, a GFP e seus derivados têm sido amplamente utilizados em várias experiências como marcador celular e de proteínas. O objetivo do presente estudo foi o de verificar o genótipo dos descendentes de cruzamentos entre ratos Lewis LEW-Tg (EGFP) F455.5/Rrrc heterozigotos e de analisar a expressão da proteína fluorescente verde melhorada (EGFP) em diferentes tipos celulares e genótipos. O genótipo da descendência foi avaliado por PCR e pela análise da expressão da EGFP em diferentes células e genótipos, incluindo-se as células-tronco mesenquimais (MSC) derivadas de tecido adiposo e de osteoblastos de calvária. A expressão da EGFP foi verificada por citometria de fluxo, microscopia de fluorescência e imunocoloração. Foram, identificados os genótipos da descendência e determinados os níveis de expressão de EGFP em dois tipos de células. Foi também constatada uma diferença de expressão entre os genótipos (EGFP +/+) e (EGFP +/-) além da presença de autofluorescência. Mais estudos são necessários para esclarecer a fluorescência natural de células com o genótipo (EGFP +/-) e aquela induzida pela presença da EGFP.

Cultivo de células estromales de rata. Influencia del suero fetal bovino / Stromal rat cell culture. Influence of fetal calf serum

Las células madre estromales humanas y de roedores cultivadas pueden ser inducidas a diferenciarse en neuronas, enfatizando su utilidad potencial en la terapia celular neurorrestaurativa. Los sistemas de cultivo para la expansión de estas células describen el uso de diferentes proporciones de suero fetal, lo que motivó a estudiar qué concentración de suero fetal bovino era capaz de garantizar un adecuado rendimiento celular. Las células de la médula ósea de rata se cultivaron en medio a-MEM suplementado con 10 y 20 por cientode suero fetal bovino y se subcultivaron hasta 3 veces. La viabilidad celular de los cultivos primarios y los subcultivos estuvo por encima del 98 por ciento en ambos experimentos. Los cultivos primarios demoraron 17,4 días en confluir y los subcultivos 7,7 días. La concentración de suero fetal al 20 por ciento no aumentó significativamente la velocidad de multiplicación celular; no obstante, se obtuvo un mayor número de células estromales. El sistema de expansión in vitro podría utilizarse en estudios futuros para la expansión de las células estromales humanas, lo que sienta mejores bases para su aplicación clínica

Cultured human and rodents stromal stem cells can be induced to differentiate into neurons, emphasizing its potential use in neurorestorative cell therapy. Cropping systems for the expansion of these cells describe the use of different ratios of fetal serum, which led to study what concentration of fetal calf serum was able to ensure an adequate cell yield. Cells from rat bone marrow were cultured in medium supplemented with a-MEM 10 and 20 percent fetal bovine serum and subcultured up to 3 times. Cell viability of primary cultures and subcultures was above 98 percent in both experiments. Primary cultures converge delayed in 17.4 days and 7.7 days subcultures. The concentration of 20 percent fetal calf serum did not significantly increase the speed of cell division, however, we obtained a greater number of stromal cells. The expansion in vitro system could be used in future studies for the expansion of human stromal cells, which feels better basis for clinical application

Influence of low frequency electric and magnetic fields exposure on albino rat placentation following maternal exposure during early and late periods of embryogenesis

Background: with the phenomenal growth in the use of extremely low frequency electromagnetic fields EMFs, there has been a surge of interest concerning the possible harmful biological effects of electromagnetic fields EMFs with a frequency of 50-60 Hz on the human body

Aim of the work: The aim of the present study is to evaluate the histological changes in the placenta after exposure to low frequency electric and magnetic fields during two different critical periods of embryogenesis

Methods: twenty-four pregnant albino rats were randomly divided into three equal groups: control, early exposed and late exposed. Both exposed groups were exposed to 50 Hz magnetic field of intensity 1.5G and a strong electric field of intensity 10KV/m. The EMFs exposure started from gestational day 6 through day 10 in the early exposed group and from gestational day 11 through day 15 in the late exposed group. Twenty day full term placenta were then collected and examined histological through light microscopy using hematoxylin and Eosin stains

Results: The present study indicated that 50Hz EMFs exposure the low frequency EMFs exposure in the early and late period affect the placentation of albino rats. Also EMFs exposure in the early period cause changes in rat placenta more than that occurred in the late exposure except the deciduas was more affected in late exposure

Conclusions: So it is suggested that exposure to appliances that producing EMFs must be limited especially during pregnancy

Population structure and genetic variability of mainland and insular populations of the Neotropical water rat, Nectomys squamipes (Rodentia, Sigmodontinae)

Seven microsatellite loci were used to investigate the genetic variability and structure of six mainland and two island populations of the Neotropical water rat Nectomys squamipes, a South American semi-aquatic rodent species with a wide distribution. High levels of variability were found within mainland populations while island populations were less variable but the more differentiated in respect to allele number and frequency. The time of biological divergence between mainland and island populations coincided with geological data. A significant geographic structure was found in mainland populations (q = 0.099; r = 0.086) although the degree of differentiation was relatively low in respect to the distance between surveyed localities (24 to 740 km). Genetic and geographic distances were not positively correlated as previously found with random amplified polymorphic DNA (RAPD) markers. Significant but low genetic differentiation in the mainland and lack of isolation by distance can be explained by large population size and/or recent population expansion. Additionally, the agreement between the age of geologic events (sea level fluctuations) and divergence times for insular populations points to a good reference for molecular clock calibration to associate recent environmental changes and the distribution pattern of small mammals in the Brazilian Atlantic Forest

Nine karyomorphs for spiny rats of the genus Proechimys (Echimyidae, Rodentia) from North and Central Brazil

Spiny rats of the genus Proechimys are morphologically diverse, widely distributed and have diploid numbers ranging from 2n = 14-16 to 2n = 62. In this paper we present cytogenetical data and brief comments on morphological and biogeographical issues related to spiny rats. In our sample of 42 spiny rats collected from 12 Brazilian Amazonian tropical rainforest and the Cerrado (Brazilian savanna) sites we detected nine karyological entities: four different karyomorphs with 2n = 30, three with 2n = 28, one with 2n = 15 and one with 2n = 44. Based on qualitative morphological characters these karyomorphs can be allocated to five species within the goeldii, guyannensis and longicaudatus species groups

In vitro translation of RNA to lactase during postnatal development of rat intestine.

mRNA levels encoding lactase were detected by Northern blot analysis using two different probes in developing rat intestine. Probe I and probe II corresponding to second half of prolactase gene showed a 6.8 kb mRNA transcript in 7, 14, 21 and 30 day old rat intestine. There was no change in quantity of lactase mRNA detected using probe II, but hybridization with probe I showed a progressive decrease in mRNA transcript encoding lactase with age. At day 7 and 14 of postnatal development, the lactase mRNA was quite high, but it reduced upon weaning. The in vitro translation products of RNA detected by Western blot analysis using brush border lactase antibodies showed several isoforms of lactase antigen with molecular weight ranging from 100-220 kDa. Analysed at days 7 and 30 of postnatal development, lactase isoforms of molecular weight 130 kDa and 220 kDa were similar to those found in purified brush border membranes. The translation of RNA to 220 kDa lactase protein was high in 7 and 14 day old pups, but it was markedly reduced in 30 day old animals. These findings support the contention that translation of mRNA to lactase is impaired in weaned animals, which may also be responsible for the maturational decline in lactase activity in adult rat intestine.

Plasmodium yoelii: identification of a gene encoding a putative ADP-ribosylation factor-1 GTPase-activating protein, PyAG1

The PyAG1 gene, identified by the screening of a Plasmodium yoelii genomic DNA library with a rhoptry-specific Mab, encodes a protein with a zinc finger structure immediately followed by the consensus sequence of the Arf GAP catalytic site. The serum of mice immunized with the recombinant protein recognized specifically the rhoptries of the late infected erythrocytic stages. Blast analysis using the Genbank database gave the highest scores with four proteins presenting an Arf1 GAP activity. If presenting also this activity, the PyAG1 protein could be involved in the regulation of the secreted protein vesicular transport and, consequently, in the rhoptry biogenesis.

Marcaçäo fenotípica para cor e padräo de pelagem em Rattus norvegicus - I / Phenotypic marking for pattern and coat color Rattus norvegicus - I

A cor e padräo de pelagem em Rattus norvegicus säo fenótipos de mais fácil identificaçäo prática deste animal. A seleçäo de matrizes genotipicamente definidas para estes fenótipos foi obtida a partir de ratos Wistar e Long Evans, submetidos a cruzamentos experimentais, seguidos de cruzamentos-teste. Tais matrizes foram utilizadas para iniciar uma série de cruzamentos endogâmicos (cinco geraçöes consecutivas; incrossing = 67.2 por cento), com a finalidade de serem posteriormente comparadas entre si e a um grupo controle, inicialmente quanto aos efeitos da consangüinidade. A análise dos resultados permitiu concluir que tais efeitos levaram a uma reduçäo da fertilidade nas linhagens selecionadas, o que näo se verificou no grupo controle, mesmo submetido a endogamia.