Aromatase expression in cultured fetal sheep astrocytes after nitrosative/oxidative damage.

Aromatase, the enzyme converting androgens into estrogens, is involved in many brain processes such as neural differentiation and plasticity or the prevention of cell death. We have previously observed an increase in aromatase immunoreactivity in sheep neurons exposed in vitro to the oxidant 3-nitro-L: -tyrosine. However, little is known regarding the way that sheep astrocytes cope with nitrosative stress, a condition occurring in sheep in the pathogenesis of neurodegenerative disorders such as scrapie and Maedi-Visna. Our aim has been to evaluate the effects of 3-nitro-L-tyrosine on astrocyte primary cultures from 90-day-old fetal sheep brain. Living cells were observed and characterized by immunofluorescence with a GFAP antibody, which indicated that the majority of the cells were astrocytes. A viability assay was performed on both untreated and treated cells. Reverse transcription with the polymerase chain reaction was undertaken to monitor time- and dose-dependent variations in aromatase gene expression. Stressed astrocytes showed signs of deterioration, were reduced in number, and appeared round with few short processes; the cell death rate was ∼30%. Aromatase expression was detected starting from a 24-h exposure to 1 mM 3-nitro-L-tyrosine and reached the highest levels at 72 h. Thus, oxidative damage probably results in the local production of neuroprotective estradiol by reactive astrocytes via the aromatization of testosterone.

Microcystins Presence in Mussels (M. galloprovincialis) and Water of Two Productive Mediterranean's Lagoons (Sardinia, Italy).

Microcystins (MCs) are hepatotoxins harmful for animal and human health. The most toxic type between them is MC-LR whose presence has been investigated in different reservoirs all around the world. In this work microcystins were monitored in spring and summer in water and mussels (Mytilus galloprovincialis) of two Sardinia lagoons: Cabras and Calich lagoons. A Solid Phase Extraction method was developed to clean and concentrate samples before the Enzyme Linked Immunosorbent Assay (ELISA) and the following Mass Spectrometry detection. MCs presence was detected using the screening ELISA test in both lagoons. MCs peak was revealed in July for water and mussels belonging to Cabras lagoon (0.75 ± 0.07 ng/L in water and 0.12 ± 0.04 ng/g ww in mussels). In water of Calich lagoon there was a constant trend in the concentration of MCs during the considered months, while there was a MCs peak in July (0.6 ± 0.5 ng/g ww) in mussels. The following LC-MS/MS analysis did not reveal MC-LR presence in all analyzed samples. These results can be useful to enrich knowledge on public health and consumer's safeguard.

Reorganization of actin cytoskeleton in L929 cells infected with Coxiella burnetii strains isolated from placenta and foetal brain of sheep (Sardinia, Italy).

Coxiella burnetii, the etiological agent of Q Fever, is a zoonotic pathogen distributed worldwide. It has been reported that virulent strains of C. burnetii are poorly internalized by monocytes compared to avirulent variants. Virulence is also associated to the formation of pseudopodal extensions and transient reorganization of filamentous actin. In this article, we investigated the ability of 2 Coxiella strains isolated from ovine aborted samples to induce reorganization of the actin cytoskeleton in mouse fibroblast cells. Cells were exposed for 24 and 48 hours to ovine placenta and foetal brain tissue homogenates and then analysed by polymerase chain reaction (PCR) in order to detect Coxiella infection. The formation of pseudopodal extensions, the polarized distribution of F­actin, and the involvement of C. burnetii strain in cytoskeleton reorganization have been assessed using a laser scanning confocal fluorescence microscope. Results indicate that similarly to the virulent reference strain, strains of C. burnetii isolated from foetal brain induced morphological changes - modification in F­actin distribution and in the localization of bacteria. By contrast, C. burnetii strain isolated from ovine placenta did not induce any significant change in L929 cell morphology. In conclusion, both C. burnetii strains isolated from ovine placenta and foetal brain were pathogenic causing ovine abortion, but in vitro the C. burnetii strain isolated from brain only was able to induce F­actin reorganization in L929 infected cells.