Joana Santos

meis1 regulates cyclin D1 and c-myc expression, and controls the proliferation of the multipotent cells in the early developing zebrafish eye

During eye development, retinal progenitors are drawn from a multipotent, proliferative cell population. In Drosophila the maintenance of this cell population requires the function of the TALE-homeodomain transcription factor Hth, although its mechanisms of action are still unknown. Here we investigate whether members of the Meis gene family, the vertebrate homologs of hth, are also involved in early stages of eye development in the zebrafish. We show that meis1 is initially expressed throughout the eye primordium. Later, meis1 becomes repressed as neurogenesis is initiated, and its expression is confined to the ciliary margin, where the retinal stem population resides. Knocking down meis1 function through morpholino injection causes a delay in the G1-to-S phase transition of the eye cells, and results in severely reduced eyes. This role in cell cycle control is mediated by meis1 regulating cyclin D1 and c-myc transcription. The forced maintenance of meis1 expression in cell clones is incompatible with the normal differentiation of the meis1-expressing cells, which in turn tend to reside in undifferentiated regions of the retinal neuroepithelium, such as the ciliary margin. Together, these results implicate meis1 as a positive cell cycle regulator in early retinal cells, and provide evidence of an evolutionary conserved function for Hth/Meis genes in the maintenance of the proliferative, multipotent cell state during early eye development.

Sequential determination of fat- and water-soluble vitamins in green leafy vegetables during storage

The simultaneous analysis of fat- and water-soluble vitamins from foods is a difficult task considering the wide range of chemical structures involved. In this work, a new procedure based on a sequential extraction and analysis of both types of vitamins is presented. The procedure couples several simple extraction steps to LC-MS/MS and LC-DAD in order to quantify the free vitamins contents in fresh-cut vegetables before and after a 10-days storage period. The developed method allows the correct quantification of vitamins C, B(1), B(2), B(3), B(5), B(6), B(9), E and provitamin A in ready-to-eat green leafy vegetable products including green lettuce, ruby red lettuce, watercress, swiss chard, lambs lettuce, spearmint, spinach, wild rocket, pea leaves, mizuna, garden cress and red mustard. Using this optimized methodology, low LOQs were attained for the analyzed vitamins in less than 100 min, including extraction and vitamin analysis using 2 optimized procedures; good repeatability and linearity was achieved for all vitamins studied, while recoveries ranged from 83% to 105%. The most abundant free vitamins found in leafy vegetable products were vitamin C, provitamin A and vitamin E. The richest sample on vitamin C and provitamin A was pea leaves (154 mg/g fresh weight and 14.4 mg/100g fresh weight, respectively), whereas lambs lettuce was the vegetable with the highest content on vitamin E (3.1 mg/100 g fresh weight). Generally, some losses of vitamins were detected after storage, although the behavior of each vitamin varied strongly among samples.

Phenolic profile evolution of different ready-to-eat baby-leaf vegetables during storage.

Ready-to-eat baby-leaf vegetables market has been growing and offering to consumers convenient, healthy and appealing products, which may contain interesting bioactive compounds. In this work, the composition and the evolution of the phenolic compounds from different baby-leaf vegetables during refrigerated storage was studied. The phenolic compounds were extracted using pressurized liquid extraction (PLE) and the phenolic profile of each sample was analyzed and quantified by using LC-MS and LC-DAD methods, respectively, at the beginning and at the end of a 10-day storage period. The baby-leaf vegetables studied included green lettuce, ruby red lettuce, swiss chard, spinach, pea shoots, watercress, garden cress, mizuna, red mustard, wild rocket and spearmint samples and a total of 203 phenolic compounds were tentatively identified and quantified. The main naturally phenolic compounds identified correspond to glycosylated flavonoids, with exception of green lettuce and spearmint leaves which had a higher content of hydroxycinnamic acids. Quantification of the main compounds showed a 10-fold higher content of total phenolic content of ruby red lettuce (483mgg(-1)) in relation to the other samples, being the lowest values found in the garden cress (12.8mgg(-1)) and wild rocket leaves (8.1mgg(-1)). The total phenolic content only showed a significant change (p<0.05) after storage in the green lettuce (+17.5%), mizuna (+7.8%), red mustard (-23.7%) and spearmint (-13.8%) leaves. Within the different classes of phenolic compounds monitored, the flavonols showed more stable contents than the hydroxycinnamic and hydroxybenzoic acids, although the behavior of each compound varied strongly among samples.

Yellow eel (Anguilla anguilla) development in NW Portuguese estuaries with different contamination levels.

The aims of the present study were to compare the health status of yellow eels (Anguilla anguilla) developing in three estuaries of the NW Portuguese coast with different levels of pollution and their physiological responses to combined effects of environmental variation and pollution. For this, a field study was performed using a multi-parameter approach, including eels condition indexes and biomarkers, water quality variables and other environmental factors. Sixteen biological parameters were assessed, namely: hepatosomatic index (LSI), Fulton’s condition index (K), lipid peroxidation (LPO), total glutathione (TG), reduced glutathione (GSH), oxidised glutathione (GSSG), GSH/GSSG, and the activity of the enzymes acetylcholinesterase (AChE), lactate dehydrogenase (LDH), sodium-potassium ATPase (Na+/K+-ATPase), ethoxyresorufin O-deethylase (EROD), glutathione S-transferases (GST), catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GR). Ten environmental factors were also measured in water: temperature, salinity, pH, phosphates, nitrates, nitrites, ammonium, silica, phenol and hardness. Globally, the biomarkers indicate exposure and toxic effects of pollutants on eels living in contaminated estuaries. The relationships between biological and environmental variables were assessed through redundancy analysis. K and LSI indexes, AChE and Na+/K+-ATPase, total glutathione levels and the antioxidant enzymes CAT, GR, and SOD where the factors most discriminating reference (Minho River estuary) from contaminated estuaries (Lima and Douro Rivers estuaries). Moreover, the most striking outcomes of pollutants exposure on biological responses were observed during winter, probably due to a joint effect of cold weather and pollution stress. Altogether, the results indicate that the development of eels in the polluted estuaries of Lima and Douro rivers is interfering with physiological functions determinant for their survival and performance. This may increase the mortality rates during the continental life-phase of the species and decrease the percentage of animals able to successfully complete their oceanic migration and, thus, reduce the contribution of each generation to the next one.

Comparative study about the effects of pollution on glass and yellow eels (Anguilla anguilla) from the estuaries of Minho, Lima and Douro Rivers (NW Portugal).

The health status of eels (Anguilla anguilla) developing in three estuaries of the NW Portuguese coast with different types and levels of pollution was compared in relation to morphometric parameters, Fulton condition index (F index) and several biomarkers. Relatively to the reference population, glass eels from the Lima estuary had lower weight and length, cholinesterase (ChE) and lactate dehydrogenase (LDH) inhibition, and lower levels of some anti-oxidant parameters, while yellow eels also showed a decreased F index, and increased Na(+)/K(+)-ATPase and lipid peroxidation (LPO) levels. Relatively to the reference population, glass eels from the Douro estuary had increased Na(+)/K(+)-ATPase and glutathione-S-transferase activities and LDH inhibition, while yellow eels also had ChE inhibition and increased LPO. Overall, these results indicate that eels from polluted estuaries showed a poor health status than those from a reference estuary, and that adverse effects become more pronounced after spending several years in polluted estuaries.

Multi-elemental analysis of ready-to-eat “baby leaf” vegetables using microwave digestion and high-resolution continuum source atomic absorption spectrometry

The mineral content (phosphorous (P), potassium (K), sodium (Na), calcium (Ca), magnesium (Mg), iron (Fe), manganese (Mn), zinc (Zn) and copper (Cu)) of eight ready-to-eat baby leaf vegetables was determined. The samples were subjected to microwave-assisted digestion and the minerals were quantified by High-Resolution Continuum Source Atomic Absorption Spectrometry (HR-CS-AAS) with flame and electrothermal atomisation. The methods were optimised and validated producing low LOQs, good repeatability and linearity, and recoveries, ranging from 91% to 110% for the minerals analysed. Phosphorous was determined by a standard colorimetric method. The accuracy of the method was checked by analysing a certified reference material; results were in agreement with the quantified value. The samples had a high content of potassium and calcium, but the principal mineral was iron. The mineral content was stable during storage and baby leaf vegetables could represent a good source of minerals in a balanced diet. A linear discriminant analysis was performed to compare the mineral profile obtained and showed, as expected, that the mineral content was similar between samples from the same family. The Linear Discriminant Analysis was able to discriminate different samples based on their mineral profile.

Mycobacterium gordonae urinary infection in a renal transplant recipient

Abstract: The authors present a case of urinary infection by a non‐tuberculous mycobacteria (NTM) species, Mycobacterium gordonae, in a renal transplant recipient. A 29‐year‐old female patient had persistent sterile pyuria after her second kidney transplant. An NTM, M. gordonae, was isolated, and the patient was started on antituberculous treatment, with resolution of leukocyturia. Ureteral stenosis with hydronephrosis and deterioration of allograft function was diagnosed later on and, despite the introduction of intraureteral catheter and resolution of hydronephrosis, there was no recovery of baseline renal function. She ultimately resumed dialysis after a severe pyelonephritis. The authors discuss the problems of establishing diagnosis of infection (versus colonization) by NTM and highlight the difficulty of treating these infections, especially because of the possible interaction with immunosuppressant agents, facilitating anti‐allograft immune response.

Evaluating the impact of sprouting conditions on the glucosinolate content of Brassica oleracea sprouts.

The glucosinolates content of brassica plants is a distinctive characteristic, representing a healthy advantage as many of these compounds are associated to antioxidant and anti-carcinogenic properties. Brassica sprouts are still an underutilized source of these bioactive compounds. In this work, four varieties of brassica sprouts (red cabbage, broccoli, Galega kale and Penca cabbage), including two local varieties from the North of Portugal, were grown to evaluate the glucosinolate profile and myrosinase activity during the sprouting. Also the influence of light/darkness exposure during sprouting on the glucosinolate content was assessed. Glucosinolate content and myrosinase activity of the sprouts was evaluated by HPLC methods. All sprouts revealed a higher content of aliphatic glucosinolates than of indole glucosinolates, contrary to the profile described for most of brassica mature plants. Galega kale sprouts had the highest glucosinolate content, mainly sinigrin and glucoiberin, which are recognized for their beneficial health effects. Penca cabbage sprouts were particularly richer in glucoraphanin, who was also one of the major compounds in broccoli sprouts. Red cabbage showed a higher content of progoitrin. Regarding myrosinase activity, Galega kale sprouts showed the highest values, revealing that the use of light/dark cycles and a sprouting phase of 7-9 days could be beneficial to preserve the glucosinolate content of this variety.

Cerebral coccidioidomycosis after renal transplantation in a non‐endemic area

C. Carvalho, I. Ferreira, S. Gaião, S. Guimarães, R. Costa, J. Santos, S. Sampaio, M. Bustorff, G. Oliveira, M. Pestana. Cerebral coccidioidomycosis after renal transplantation in a non‐endemic area.
Transpl Infect Dis 2010: 12: 151–154. All rights reserved

Phylogeny of the teashirt-related zinc finger (tshz) gene family and analysis of the developmental expression of tshz2 and tshz3b in the zebrafish.

The tshz genes comprise a family of evolutionarily conserved transcription factors. However, despite the major role played by Drosophila tsh during the development of the fruit fly, the expression and function of other tshz genes have been analyzed in a very limited set of organisms and, therefore, our current knowledge of these genes is still fragmentary. In this study, we perform detailed phylogenetic analyses of the tshz genes, identify the members of this gene family in zebrafish and describe the developmental expressions of two of them, tshz2 and tshz3b, and compare them with meis1, meis2.1, meis2.2, pax6a, and pax6b expression patterns. The expression patterns of these genes define a complex set of coexpression domains in the developing zebrafish brain where their gene products have the potential to interact. Developmental Dynamics 239:1010–1018, 2010.