Impaired phospholipases A2 production by stimulated macrophages from patients with acute respiratory distress syndrome

The aim of this study was to investigate whether early phase of acute respiratory distress syndrome (ARDS) is associated with changes in immune response, either systemic or localized to the lung. ARDS and control mechanically ventilated patients, as well as healthy volunteers were studied. Alveolar macrophages (AMΦ) and blood monocytes (BM) were treated ex vivo with lipopolysaccharide (LPS), interferon-γ (IFNγ), and surfactant. Phospholipase A₂ (PLA₂) activity and TLR4 expression were evaluated as markers of cell response. AMΦ from ARDS patients did not respond upon treatment with either LPS or IFN-γ by inducing PLA₂ production. On the contrary, upon stimulation, in control patients the intracellular PLA₂, (mainly cPLA₂) levels were increased, but secretion of PLA₂ (mainly sPLA₂-IIA) was observed only after treatment with LPS. Surfactant suppressed PLA₂ production in cells from both groups of patients. Increased relative changes of total PLA₂ activity and an upregulation of TLR4 expression upon stimulation was observed in BM from primary ARDS, control patients and healthy volunteers. In BM from secondary ARDS patients, however, no PLA₂ induction was observed, with a concomitant down-regulation of TLR4 expression. Cytosolic PLA₂, its activated form, p-cPLA_2, and sPLA₂-IIA were the predominant PLA₂ types within the cells, while extracellularly only sPLA₂-IIA was identified. These results support the concept of down-regulated innate immunity in early ARDS that is compartmentalized in primary and systemic in secondary ARDS. PLA₂ isoforms could serve as markers of the immunity status in ARDS. Finally, our data highlight the role of surfactant in controlling inflammation.     

E-cadherin in oral SCC: an analysis of the confusing literature and new insights related to its immunohistochemical expression

E-cadherin plays a crucial structural role in cell-cell contacts in epithelial tissues, and a functional role in signaling pathways that regulate cell proliferation, differentiation, and survival. Reduced immunoexpression of E-cadherin adhesions is largely considered as being equivalent to defective functionality and malignancy, and has been used as a prognostic parameter. A critical analysis of studies on E-cadherin immunoexpression in oral carcinomas revealed a wide range of both technical and interpretational aspects. This paper highlights biological characteristics of E-cadherin with respect to its expression in normal and neoplastic epithelial cells and to its interrelations with the tumor microenvironment that can have an impact on immunohistochemical results and their application in the clinical setting.     

The Importance of a Proper Aetiological Diagnosis in the Management of Patients with Invasive Mycoses: A Case Report of a Brain Abscess by <Emphasis Type="Italic">Scedosporium apiospermum</Emphasis>

Scedosporium apiospermum is a saprobic fungus responsible for many different clinical manifestations. Although it affects mostly immunocompromised patients, pulmonary and disseminated scedosporiosis have also been reported in immunocompetent subjects. It often causes subcutaneous mycetoma, despite its preferential tropism to CNS. The authors describe a fatal case of a S. apiospermum brain abscess in a 58-year-old female. She was affected by chronic liver disease and idiopathic pulmonary fibrosis and had been treated with corticosteroid therapy for a long time. She recovered in a neurosurgery unit, wherein TC scan and cerebral MRI revealed an expansive left temporo-parietal process with vasogenic oedema. A stereotactic puncture of the lesion was carried out, and pus of brain abscess was evacuated. Empirical antifungal therapy was initiated with liposomal amphotericine B based on the clinical suspicion of Zygomycetes infection; after 3 days, posaconazole was added. The correct aetiological diagnosis arrived too late and the patient was treated with no specific therapy. This fatal case confirms the necessity of having a fast and correct aetiological diagnosis to improve the patient’s outcome.     

The distribution of genetic variation in Norway spruce (Picea abies Karst.) populations in the western Alps

Aim  In order to look for a possible centre of survival for the Norway spruce ( Picea abies Karst.) in the south-western Alps, six natural populations of this area were investigated by means of genetic markers in order to assess the degree and the distribution of genetic diversity within the species.
Location  Western and South-western Alps.
Methods  Populations were genotyped using seven simple sequence repeat (SSR) markers. Basic population genetics parameters were estimated and the amount of genetic differentiation calculated.
Results  A large amount of variability was found (0.59 <  H _e < 0.67); genetic differentiation as measured by F _ST was 0.05, close to other similar studies; no isolation by distance was detected by a Mantel test. Analysis of molecular variance confirmed a high degree of variability within populations and a low degree of variability among populations. Finally, the number of populations from which those observed could have arisen was estimated by Bayesian analysis.
Main conclusions  The results presented here suggest that the present populations derive their genetic make-up from three inferred clusters. The possible existence in this area of a relict/refuge population during the last glaciation is discussed.     

Mass spectrometric identification of the trypsin cleavage pathway in lysyl-proline containing oligotuftsin peptides.

Trypsin cleaves specifically peptide bonds at the C-terminal side of lysine and arginine residues, except for -Arg-Pro- and -Lys-Pro- bonds which are normally resistant to proteolysis. Here we report evidence for a -Lys-Pro- tryptic cleavage in modified oligotuftsin derivatives, Ac-[TKPKG]4-NH2) (1), using high-resolution mass spectrometry and HPLC as primary methods for analysis of proteolytic reactions. The proteolytic susceptibility of -Lys-Pro- bonds was strongly dependent on flanking residues, and the flexibility of the peptide backbone might be a prerequisite for this unusual cleavage. While -Lys-Gly- bonds in 1 were rapidly cleaved, the modification of these Lys residues by the attachment of a ss-amyloid(4-10) epitope to yield -Lys(X)-Gly derivatives prevented cleavage of this bond, and provided trypsin cleavage of -Lys-Pro- bonds, the pathway of this degradation being independent on the type of Lys-N(epsilon)-side chains (acetyl group, amino acid, peptide). Substitution of the Lys residues by Ala at the P'2 positions decreased the tryptic cleavage, while replacement of the bulky side chain of Thr at the P2 positions strongly increased the cleavage of -Lys-Pro- bonds. Circular dichroism (CD) data of the modified oligotuftsin derivatives are in accord with enhanced flexibility of the peptide backbone, as a prerequisite for increased susceptibility to cleavage of -Lys-Pro- bonds. These results obtained of oligotuftsin derivatives might have implications for the proteolytic degradation of target peptides that require specific conformational preconditions.     

The Depolarizing Action of GABA Controls Early Network Activity in the Developing Hippocampus

Early in postnatal life γ-aminobutyric acid (GABA), the primary inhibitory transmitter in adults, excites targeted neurons by an outwardly directed flux of chloride which results from the unbalance between the cation–chloride cotransporters NKCC1 and KCC2, involved in chloride uptake and extrusion, respectively. This effect contributes to generate synchronized network activity or giant depolarizing potentials (GDPs) in the developing hippocampus. Here, we review some recent data concerning the mechanisms by which GDPs are generated and their functional role in enhancing synaptic efficacy at poorly developed GABAergic and glutamatergic synapses. In adulthood, reshaping neuronal circuits due to changes in chloride homeostasis and to the shift of GABA from hyperpolarizing to depolarizing, has been implicated in several neurological disorders, including epilepsy. Evidence has been recently provided that in chronically nerve growth factor-deprived mice expressing a progressive age-dependent neurodegenerative pathology resembling that observed in patients with Alzheimer’s disease, the reduced expression of mRNA encoding for the Kcc2 gene and the depolarizing action of GABA lead to the reorganization of the neuronal hippocampal network. This may represent a novel mechanism by which GABAergic signaling counterbalances the loss of synaptic activity in neurodegenerative diseases.     

3D tumour models: novel in vitro approaches to cancer studies

3D in vitro models have been used in cancer research as a compromise between 2-dimensional cultures of isolated cancer cells and the manufactured complexity of xenografts of human cancers in immunocompromised animal hosts. 3D models can be tailored to be biomimetic and accurately recapitulate the native in vivo scenario in which they are found. These 3D in vitro models provide an important alternative to both complex in vivo whole organism approaches, and 2D culture with its spatial limitations. Approaches to create more biomimetic 3D models of cancer include, but are not limited to, (i) providing the appropriate matrix components in a 3D configuration found in vivo, (ii) co-culturing cancer cells, endothelial cells and other associated cells in a spatially relevant manner, (iii) monitoring and controlling hypoxia- to mimic levels found in native tumours and (iv) monitoring the release of angiogenic factors by cancer cells in response to hypoxia. This article aims to overview current 3D in vitro models of cancer and review strategies employed by researchers to tackle these aspects with special reference to recent promising developments, as well as the current limitations of 2D cultures and in vivo models. 3D in vitro models provide an important alternative to both complex in vivo whole organism approaches, and 2D culture with its spatial limitations. Here we review current strategies in the field of modelling cancer, with special reference to advances in complex 3D in vitro models.     

Nandrolone and stanozolol induce leydig cell tumor proliferation through an estrogen‐dependent mechanism involving IGF‐I system

Several substances such as anabolic androgenic steroids (AAS), peptide hormones like insulin‐like growth factor‐I (IGF‐I), aromatase inhibitors and estrogen antagonists are offered via the Internet, and are assumed without considering the potential deleterious effects that can be caused by their administration. In this study we aimed to determine if nandrolone and stanozolol, two commonly used AAS, could have an effect on Leydig cell tumor proliferation and if their effects could be potentiated by the concomitant use of IGF‐I. Using a rat Leydig tumor cell line, R2C cells, as experimental model we found that nandrolone and stanozolol caused a dose‐dependent induction of aromatase expression and estradiol (E2) production. When used in combination with IGF‐I they were more effective than single molecules in inducing aromatase expression. AAS exhibited estrogenic activity and induced rapid estrogen receptor (ER)‐dependent pathways involving IGF1R, AKT, and ERK1/2 phosphorylation. Inhibitors for these kinases decreased AAS‐dependent aromatase expression. Up‐regulated aromatase levels and related E2 production increased cell proliferation as a consequence of increased cyclin E expression. The observation that ER antagonist ICI182,780 was also able to significantly reduce ASS‐ and AAS + IGF‐induced cell proliferation, confirmed a role for estrogens in AAS‐dependent proliferative effects. Taken together these data clearly indicate that the use of high doses of AAS, as it occurs in doping practice, enhances Leydig cell proliferation, increasing the risk of tumor development. This risk is higher when AAS are used in association with IGF‐I. To our knowledge this is the first report directly associating AAS and testicular cancer. J. Cell. Physiol. 227: 2079–2088, 2012. © 2011 Wiley Periodicals, Inc.