Relationships among GnRH, substance P, prostaglandins, sex steroids and aromatase activity in the brain of the male lizard Podarcis sicula sicula during reproduction.

The release of PGF2 alpha and PGE2, progesterone, androgens and oestradiol in vitro, and the aromatase activity in the brain of the male lizard Podarcis sicula sicula during three different phases of the reproductive period were evaluated. In addition, the effects of salmon GnRH, substance P, salmon GnRH antagonist, substance P antagonist, PGF2 alpha, PGE2 and acetylsalicylic acid on the release of prostaglandins and sex steroids and on aromatase activity in the brain were evaluated during the same three phases. PGF2 alpha, oestradiol and aromatase activity were higher during the refractory phase, androgens during the fighting phase, and progesterone during the mating phase, while PGE2 was lower during the refractory phase. Treatment with salmon GnRH increased PGF2 alpha, oestradiol and aromatase activity, but decreased the amount of androgens released. Substance P decreased PGF2 alpha, oestradiol and aromatase activity, but increased the amount of androgens released. PGF2 alpha increased oestradiol and aromatase activity, but decreased the amount of androgens released. Acetylsalicylic acid decreased PGF2 alpha, oestradiol and aromatase activity, but increased the amount of androgens released. These data suggest that salmon GnRH and substance P have different roles in reproductive processes, with opposite mechanisms, in the central nervous system of this male lizard: salmon GnRH seems to be involved in regulating the refractory phase, while substance P plays a role in regulating the fighting phase

Tyrosine Phosphorylation of Eps15 Is Required for Ligand-Regulated, but Not Constitutive, Endocytosis

Membrane receptors are internalized either constitutively or upon ligand engagement. Whereas there is evidence for differential regulation of the two processes, little is known about the molecular machinery involved. Previous studies have shown that an unidentified kinase substrate is required for endocytosis of the epidermal growth factor receptor (EGFR), the prototypical ligand-inducible receptor, but not of the transferrin receptor (TfR), the prototypical constitutively internalized receptor. Eps15, an endocytic protein that is tyrosine phosphorylated by EGFR, is a candidate for such a function. Here, we show that tyrosine phosphorylation of Eps15 is necessary for internalization of the EGFR, but not of the TfR. We mapped Tyr 850 as the major in vivo tyrosine phosphorylation site of Eps15. A phosphorylation-negative mutant of Eps15 acted as a dominant negative on the internalization of the EGFR, but not of the TfR. A phosphopeptide, corresponding to the phosphorylated sequence of Eps15, inhibited EGFR endocytosis, suggesting that phosphotyrosine in Eps15 serves as a docking site for a phosphotyrosine binding protein. Thus, tyrosine phosphorylation of Eps15 represents the first molecular determinant, other than those contained in the receptors themselves, which is involved in the differential regulation of constitutive vs. regulated endocytosis

Protective role of carbonic anhydrases III and VII in cellular defense mechanisms upon redox unbalance

Under oxidative stress conditions, several constitutive cellular defense systems are activated, which involve both enzymatic systems and molecules with antioxidant properties such as glutathione and vitamins. In addition, proteins containing reactive sulfhydryl groups may eventually undergo reversible redox modifications whose products act as protective shields able to avoid further permanent molecular oxidative damage either in stressful conditions or under pathological circumstances. After the recovery of normal redox conditions, the reduced state of protein sulfhydryl groups is restored. In this context, carbonic anhydrases (CAs) III and VII, which are human metalloenzymes catalyzing the reversible hydration of carbon dioxide to bicarbonate and proton, have been identified to play an antioxidant role in cells where oxidative damage occurs. Both proteins are mainly localized in tissues characterized by a high rate of oxygen consumption, and contain on their molecular surface two reactive cysteine residues eventually undergoing S-glutathionylation. Here, we will provide an overview on the molecular and functional features of these proteins highlighting their implications into molecular processes occurring during oxidative stress conditions

Insights into the role of reactive sulfhydryl groups of Carbonic Anhydrase III and VII during oxidative damage

Carbonic anhydrases (CAs) III and VII are two cytosolic isoforms of the α-CA family which catalyze the physiological reaction of carbon dioxide hydration to bicarbonate and proton. Despite these two enzymes share a 49% sequence identity and present a very similar three-dimensional structure, they show profound differences when comparing the specific activity for CO2 hydration reaction, with CA VII being much more active than CA III. Recently, CA III and CA VII have been proposed to play a new role as scavenger enzymes in cells where oxidative damage occurs. Here, we will examine functional and structural features of these two isoforms giving insights into their newly proposed protective role against oxidative stress

Empathy and perceived burden in caregivers of patients with schizophrenia spectrum disorders

Background Caregivers of patients load different kinds of burdens, including emotional distress. Aims of this study were to evaluate both burden and empathy of caregivers who assist patients with schizophrenia spectrum disorders. Methods We selected a sample of 60 caregivers (34 women and 26 men), who assisted patients with schizophrenia spectrum disorders treated in our local Community Mental Health Center for a 1-year minimum period. We administered two scales to our sample, Zarit Burden Interview (ZBI) and Balanced Emotional Empathy Scale (BEES), and collected data of caregivers and their assisted patients in a 3-month period. Data were statistically analyzed. Results We reported a mean ZBI score of 49.68 (±15.03 SD) and a mean BEES score of 14.35 (±9.05 SD), indicating the perception of moderate-severe burden and low level of empathy, respectively. The analysis of internal consistency confirmed the good reliability of both ZBI (Cronbach’s alpha = 0.90) and BEES (Cronbach’s alpha = 0.77). The correlation between the two scales was not statistically significant at Spearman test. At our multiple linear regression, many variables of both caregiver and patient showed a significant correlation with the ZBI score. In particular, not living with the assisted patient and female gender of caregiver potentially decreased the burden, whereas clinical severity of assisted patient and two caregiver conditions, middle school education and spouse relationship with patient, could worsen the burden. We highlighted two positive statistically significant correlations between the total score of BEES and caregiver characteristics: being spouse and not living with assisted patient. Conclusions Our study highlights that the caregiver burden of patients with severe psychiatric disorders is high and is associated with low emotional empathy experienced by caregivers, probably due to a defensive psychological mechanism. The conditions of spouse and cohabitation can concomitantly increase both empathy and burden in caregivers

The Eps15 Homology (Eh) Domain-Based Interaction between Eps15 and Hrb Connects the Molecular Machinery of Endocytosis to That of Nucleocytosolic Transport

The Eps15 homology (EH) module is a protein–protein interaction domain that establishes a network of connections involved in various aspects of endocytosis and sorting. The finding that EH-containing proteins bind to Hrb (a cellular cofactor of the Rev protein) and to the related protein Hrbl raised the possibility that the EH network might also influence the so-called Rev export pathway, which mediates nucleocytoplasmic transfer of proteins and RNAs. In this study, we demonstrate that Eps15 and Eps15R, two EH-containing proteins, synergize with Hrb and Hrbl to enhance the function of Rev in the export pathway. In addition, the EH-mediated association between Eps15 and Hrb is required for the synergistic effect. The interaction between Eps15 and Hrb occurs in the cytoplasm, thus pointing to an unexpected site of action of Hrb, and to a possible role of the Eps15–Hrb complex in regulating the stability of Rev

Involvement of PAR-4 in Cannabinoid-Dependent Sensitization of Osteosarcoma Cells to TRAIL-Induced Apoptosis

The synthetic cannabinoid WIN 55,212-2 is a potent cannabinoid receptor agonist with anticancer potential. Experiments were performed to determine the effects of WIN on proliferation, cell cycle distribution, and programmed cell death in human osteosarcoma MG63 and Saos-2 cells. Results show that WIN induced G2/M cell cycle arrest, which was associated with the induction of the main markers of ER stress (GRP78, CHOP and TRB3). In treated cells we also observed the conversion of the cytosolic form of the autophagosome marker LC3-I into LC3-II (the lipidated form located on the autophagosome membrane) and the enhanced incorporation of monodansylcadaverine and acridine orange, two markers of the autophagic compartments such as autolysosomes. WIN also induced morphological effects in MG63 cells consisting in an increase in cell size and a marked cytoplasmic vacuolization. However, WIN effects were not associated with a canonical apoptotic pathway, as demonstrated by the absence of specific features, and only the addition of TRAIL to WIN-treated cells led to apoptotic death probably mediated by up-regulation of the tumor suppressor factor PAR-4, whose levels increased after WIN treatment, and by the translocation of GRP78 on cell surface

Tubulin nitration in human gliomas

Immunohistochem. and biochem. investigations showed that significant protein nitration occurs in human gliomas, esp. in grade IV glioblastomas at the level of astrocytes and oligodendrocytes and neurons. Enhanced alpha-tubulin immunoreactivity was co-present in the same elements in the glioblastomas. Proteomic methodologies were employed to identify a nitrated protein band at 55 kDa as alpha-tubulin. Peptide mass fingerprinting procedures demonstrated that tubulin is nitrated at Tyr224 in grade IV tumor samples but is unmodified in grade I samples and in non-cancerous brain tissue. These results provide the first characterization of endogenously nitrated tubulin from human tumor samples

Corneal incision width after lens implantation: Comparing delivery systems: Corneal incision width and injector systems

Abstract Purpose To compare corneal incision width after phacoemulsification and intraocular lens implantation (IOL) using different delivery systems. Methods One hundred and seventeen patients with cataract and no other anterior segment pathological features or previous eye surgery underwent cataract surgery with IOL implantation through a 2.2 mm incision. Three foldable IOL were implanted with their recommended delivery systems: Acrysof© SN60WF with Monarch© III/cartridge D (Group A, 38 patients); Tecnis© ZCB00 with Unfolder Platinum/cartridge easy load (Group B, 38 patients); Acrysof© SN60WF with Ultrasert™ preloaded system (Group C, 42 patients). Incision width was measured before and after phacoemulsification and IOL implantation. Results Before and after phacoemulsification incision width was, respectively, 2.21 ± 0.02 mm and 2.34 ± 0.08 mm in group A; 2.20 ± 0.02 mm and 2.31 ± 0.06 mm in group B; 2.20 ± 0.02 mm and 2.30 ± 0.07 mm in group C. Incision width was not significantly enlarged after phacoemulsification. Before and after IOL implantation incision width was, respectively, 2.34 ± 0.07 mm and 2.47 ± 0.07 mm in group A; 2.32 ± 0.06 mm and 2.45 ± 0.08 mm in group B; 2.30 ± 0.07 mm and 2.39 ± 0.07 mm in group C. Incision widths in group C were significantly different to groups A and B. No relationship was found between incision sizes and phacoemulsification time, ultrasound energy and IOL powers. Conclusion In cataract surgery Ultrasert™ enlarges the corneal incision less than other delivery systems

Numb Is an Endocytic Protein

Numb is a protein that in Drosophila determines cell fate as a result of its asymmetric partitioning at mitosis. The function of Numb has been linked to its ability to bind and to biologically antagonize Notch, a membrane receptor that also specifies cell fate. The biochemical mechanisms underlying the action of Numb, however, are still largely unknown. The wide pattern of expression of Numb suggests a general function in cellular homeostasis that could be additional to, or part of, its action in fate determination. Such a function could be endocytosis, as suggested by the interaction of Numb with Eps15, a component of the endocytic machinery. Here, we demonstrate that Numb is an endocytic protein. We found that Numb localizes to endocytic organelles and is cotrafficked with internalizing receptors. Moreover, it associates with the appendage domain of α adaptin, a subunit of AP2, a major component of clathrin-coated pits. Finally, fragments of Numb act as dominant negatives on both constitutive and ligand-regulated receptor-mediated internalization, suggesting a general role for Numb in the endocytic process