Although several lines of evidence have indicated that menopause is associated with increased susceptibility to neurological disorders, the mechanisms involved in this phenomenon remain to be elucidated. Because neuroinflammation is a common feature of a number of brain diseases, we hypothesized that the cessation of ovarian functions and the consequent decrease in estrogen receptor (ER)-mediated antiinflammatory activity may represent a trigger for postmenopausal brain dysfunctions. The aim of the present study was to investigate the effects of aging and surgical menopause on the activity of ER in neuroinflammation. The present study shows that ER genes are expressed in the hippocampus, but ER transcriptional activity decreases significantly beginning at 12 months of age in intact and ovariectomized mice. With ovariectomy, we observe an age-dependent accumulation of mRNA encoding inflammatory mediators (e.g. TNFα, IL1β, and macrophage inflammatory protein-2) and changes in the morphology of astroglia and microglia. In addition, we show that aging itself is coupled with an exaggerated response to acute inflammatory stimuli with a major accumulation of TNFα, IL1β, macrophage inflammatory protein-2, and macrophage chemoattractant protein-1 mRNA in response to lipopolysaccharide administration. The response to acute inflammatory stimuli appears to be differentially modulated by the duration of hormone deprivation in 12-month-old mice. Taken together, the present results show that aging is associated with decreased ER activity, despite continuous ER synthesis, and that age-dependent neuroinflammation is strongly influenced by hormone deprivation.

By the use of in vivo imaging, we investigated the dynamics of estrogen receptor (ER) activity in intact, ovariectomized, and hormone-replaced estrogen response element-luciferase reporter mice. The study revealed the existence of a long-paced, noncircadian oscillation of ER transcriptional activity. Among the ER-expressing organs, this oscillation was asynchronous and its amplitude and period were tissue dependent. Ovariectomy affected the amplitude but did not suppress ER oscillations, suggesting the presence of tissue endogenous oscillators. Long-term administration of raloxifene, bazedoxifene, combined estrogens alone or with basedoxifene to ovariectomized estrogen response element-luciferase mice showed that each treatment induced a distinct spatiotemporal profile of ER activity, demonstrating that the phasing of ER activity among tissues may be regulated by the chemical nature and the concentration of circulating estrogen. This points to the possibility of a hierarchical organization of the tissue-specific pacemakers. Conceivably, the rhythm of ER transcriptional activity translates locally into the activation of specific gene networks enabling ER to significantly change its physiological activity according to circulating estrogens. In reproductive and nonreproductive organs this hierarchical regulation may provide ER with the signaling plasticity necessary to drive the complex metabolic changes occurring at each female reproductive status. We propose that the tissue-specific oscillatory activity here described is an important component of ER signaling necessary for the full hormone action including the beneficial effects reported for nonreproductive organs. Thus, this mechanism needs to be taken in due consideration to develop novel, more efficacious, and safer hormone replacement therapies.

The k-casein fraction plays an important role in the formation, stabilisation and aggregation on casein micelles and thus affects technological and nutritional properties of milk. In this study, exon 4 of k-casein (CSN3) gene was sequenced and analysed in Girgentana goat breed. Analyses of the obtained sequences showed the presence of A, B, D, and G known alleles and two new genetic variants, named D' and N. The new D' allele differs from D in one transition, G284→A284, which did not cause amino acid change. The new N allele differs from A in five single nucleotide polymorphisms (SNPs): T245/C245, G284/A284, G309/A309, G471/A471 and T591/C591, while it differs from C in one transition, i.e. T583→C583. Comparing the amino acid sequences of N and A alleles, the first two SNPs caused no amino acid change, whereas the other SNPs produced changes (Val65/Ile65, Val119/Ile119, and Ser159/Pro159, respectively). Comparison of N allele with C revealed the amino acid change Val156→Ala156. The most frequent allele was A (0.480) followed by B (0.363), D (0.112), and N (0.034). The D' and G alleles were identified only in two animals and in heterozygous conditions with a very low frequency (0.005). The most common genotype was AB (39.5%) followed by AA (19.5%), AD (12.7%), and BB (11.7%). Homozygous D'D', GG, and NN individuals were not found. Further analysis will be performed in order to establish associations among genotypes and quantitative and qualitative milk traits.

The aim of this work was to study β-defensin 1 (SBD1) and β-defensin 2 (SBD2) genes in Valle del Belice dairy sheep in order to identify polymorphisms that can be utilized as markers of the analyzed genes, and search for the functional effects and roles of the identified polymorphisms (variation of the amino acid sequence of the protein and stability of mRNA molecule). The study was conducted on 300 randomly selected animals belonging to four flocks. A total of seven SNPs were identified, two in SBD1 and five in SBD2. The two SNPs identified in SBD2 coding region, at position 1659 and position 1667, were non-synonymous, leading to amino acid changes in the protein product. Nevertheless, the functional effects predicted by the two SNPs demonstrated that amino acid substitutions may not have effect on β-defensin 2 protein function. Moreover, we demonstrated that SBD2 mutant sequence shows changes in mRNA secondary structure. These results suggest that identified SNPs could play a role in the modulation of the immune response.

The knowledge of milk proteome has been greatly enhanced by technological advances in the proteomics field as the use of the two-dimensional differential in-gel electrophoresis, a gel-based approach which allowed the analysis of proteins from complex mixtures and the comparing of several protein samples in the same experiment. The aim of this study was to characterise the whole milk proteomic profile in Girgentana dairy goat breed by two-dimensional differential in-gel elecrophoresis. The obtained representative 2D whole milk proteomic map showed a general picture of the protein distributions over the pH 3-10 NL including about 100 spots, most of them organised like a spot train. Among differentially abundant spots in the three experimental conditions, milk fat globule EGF factor 8 protein, b-lactoglobulin, b-casein and serum albumin were successfully identified. The three-dyes system employed in two-dimensional differential in-gel electrophoresis (2D-DIGE) analysis allowed us to obtain a global representation of the Girgentana whole milk proteome. These preliminary results could be used to generate a milk reference proteomics map for the Girgentana goat breed.