Macrophage polarisation affects their regulation of trophoblast behaviour.

INTRODUCTION: During the first trimester of human pregnancy, fetally-derived extravillous trophoblast (EVT) invade into the uterine decidua and remodel the uterine spiral arteries to ensure that sufficient blood reaches the maternal-fetal interface. Decidual macrophages have been implicated in the regulation of decidual remodelling, and aberrant activation of these immune cells is associated with pre-eclampsia. METHODS: The monocytic cell line THP-1 was activated to induce a classically- or alternatively-activated macrophage phenotype and the conditioned media was used to treat the EVT cell line SGHPL-4 in order to determine the effect of macrophage polarisation on trophoblast behaviour in-vitro. SGHPL-4 cell functions were assessed using time-lapse microscopy, endothelial-like tube formation assays, and western blot. RESULTS: The polarisation state of the THP-1 cells was found to differentially alter the behaviour of trophoblast cells in-vitro with pro-inflammatory classically-activated macrophage conditioned media significantly inhibiting trophoblast motility, impeding trophoblast tube formation, and inducing trophoblast expression of cleaved caspase 3, when compared to anti-inflammatory alternatively-activated macrophage conditioned media. DISCUSSION: Macrophages can regulate trophoblast functions that are critical during decidual remodelling in early pregnancy. Importantly, there is differential regulation of trophoblast function in response to the polarisation state of these cells. Our studies indicate that the balance between a pro- and anti-inflammatory environment is important in regulating the cellular interactions at the maternal-fetal interface and that disturbances in this balance likely contribute to pregnancy disorders associated with poor trophoblast invasion and vessel remodelling.

Impact of p16 status on pro- and anti-angiogenesis factors in head and neck cancers.

BACKGROUND: Head and neck cancers (HNC) are aggressive tumours. Overexpression of p16 in HNC correlates with human papilloma virus (HPV)-associated HNC that carry a better prognosis than HPV-negative tumours. Angiogenesis is an important factor in tumour progression. Our aim was to dissect the impact of p16 expression on angiogenesis factors in HNC. METHODS: Eighteen newly diagnosed HNC patients and controls were analysed. Eleven pro- and anti-angiogenesis factors were quantified using multiplex ELISA in HNC patients and controls. Angiogenesis factors were analysed in tumour tissue using immunohistochemistry. RESULTS: Circulating levels of endostatin (anti-angiogenesis factor) were higher in the HNC group compared with healthy donors. Interestingly, the pro-angiogenesis factors angiopoietin-1 and vascular endothelial growth factor (VEGF) were significantly higher in patients with p16-negative compared with p16-positive HNC. Moreover, the major source of VEGF in p16-positive HNC tissue was tumour stromal cells. In contrast, both tumour cells and stromal cells expressed VEGF in p16-negative tissue. CONCLUSIONS: We show that p16-negative tumours associate with increased circulating levels of pro-angiogenic VEGF and angiopoietin-1. Tissue expression of VEGF differs between p16-positive and p16-negative tumours. These findings may explain differences in the biological behaviour of p16-positive and p16-negative HNC. Better understanding of mechanisms by which the p16 status influences tumour angiogenesis may guide the development of targeted therapies.

The role of T and B cells in atherosclerosis: potential clinical implications.

The chronic inflammation process that characterises atherosclerosis involves both the innate and adaptive arms of the immune system. Several lines of evidence have recently highlighted pivotal roles for T and B lymphocytes - cells that belong to the adaptive immune system - in the development and progression of atherosclerosis. In this review, we summarise the current knowledge on the roles of adaptive immune responses in atherosclerosis and present our views on how a better understanding of these immune mechanisms could shape future therapies to slow down or even prevent this disease.

Fragmentation properties of three-membered heterocyclic molecules by partial ion yield spectroscopy: C(2)H(4)O and C(2)H(4)S.

We investigated the photofragmentation properties of two three-membered ring heterocyclic molecules, C(2)H(4)O and C(2)H(4)S, by total and partial ion yield spectroscopy. Positive and negative ions have been collected as a function of photon energy around the C 1s and O 1s ionization thresholds in C(2)H(4)O, and around the S 2p and C 1s thresholds in C(2)H(4)S. We underline similarities and differences between these two analogous systems. We present a new assignment of the spectral features around the C K-edge and the sulfur L(2,3) edges in C(2)H(4)S. In both systems, we observe high fragmentation efficiency leading to positive and negative ions when exciting these molecules at resonances involving core-to-Rydberg transitions. The system, with one electron in an orbital far from the ionic core, relaxes preferentially by spectator Auger decay, and the resulting singly charged ion with two valence holes and one electron in an outer diffuse orbital can remain in excited states more susceptible to dissociation. A state-selective fragmentation pattern is analyzed in C(2)H(4)S which leads to direct production of S(2+) following the decay of virtual-orbital excitations to final states above the double-ionization threshold.

Molecular-frame angular distributions of resonant CO:C(1s) Auger electrons.

The molecular-frame angular distributions of resonantly excited CO:C(1s) --> pi* Auger electrons were determined using angle-resolved electron-ion coincidence spectroscopy in combination with a novel transformation procedure. Our new methodology yields full three-dimensional electron angular distributions with high energy resolution from the measurement of electrons at only two angles. The experimentally determined distributions are well reproduced by calculations performed in a simple one-center approximation, allowing an unambiguous identification of several overlapping Auger lines.

Inner-shell photodetachment thresholds: unexpected long-range validity of the Wigner law.

Threshold behavior in inner-shell photodetachment is studied for the first time, specifically with 2-, 3-, or 4-electron emission from He- and S-. The threshold shapes are surprisingly consistent with the Wigner threshold law in all cases, despite large PCI effects observed in He-. In S-, the s-wave law is observed to agree with the data over an unprecedented range, more than an order of magnitude greater than predictions, and allows for the observation of the d-wave component. The measurements also demonstrate a means for obtaining precise core-excitation energies of free atoms.

UV irradiation inhibits ABC transporters via generation of ADP-ribose by concerted action of poly(ADP-ribose) polymerase-1 and glycohydrolase.

ATP-binding cassette (ABC) transporters are involved in the transport of multiple substrates across cellular membranes, including metabolites, proteins, and drugs. Employing a functional fluorochrome export assay, we found that UVB irradiation strongly inhibits the activity of ABC transporters. Specific inhibitors of poly(ADP-ribose) polymerase-1 (PARP-1) restored the function of ABC transporters in UVB-irradiated cells, and PARP-1-deficient cells did not undergo UVB-induced membrane transport inhibition. These data suggest that PARP-1 activation is necessary for ABC transporter functional downregulation. The hydrolysis of poly(ADP-ribose) by poly(ADP-ribose) glycohydrolase (PARG) was also required, since specific PARG inhibitors, which limit the production of ADP-ribose molecules, restored the function of ABC transporters. Furthermore, ADP-ribose molecules potently inhibited the activity of the ABC transporter P-glycoprotein. Hence, poly(ADP-ribose) metabolism appears to play a novel role in the regulation of ABC transporters.

Corpse disposal after apoptosis.

The termination of the apoptotic program occurs in most cases via recognition and clearance by phagocytes. Engulfed cells do not simply disappear from the midst of living tissues. Constituents of the corpse indeed survive the intracellular processing and are recycled to the membrane of the phagocyte. The presentation of yielded antigens to T cells is a central event in the induction and the maintenance of peripheral tolerance. Conversely, errors in this pathway contribute to the pathogenesis of systemic and organ specific autoimmune diseases. Here we discuss the available information on the events that follow active engulfment of dying cells, with attention to the events involved in vitro and in vivo in apoptotic cell processing. The outcome of the processing is the cross-priming or the functional inactivation of T cells that specifically recognise antigens contained in the cell corpse.

Apoptosis of the teratocarcinoma cell line Tera-1 leads to the cleavage of HERV-K10gag proteins by caspases and/or granzyme B.

Redistribution, post-translational modifications and coclustering with viral antigens contribute to the immunogenicity of apoptotic cell-derived autoantigens. Almost all known targets of the humoral autoimmune response in systemic lupus erythematosus (SLE) are cleaved by caspases or granzyme B during apoptosis. Antibodies against retroviral proteins can frequently be detected in the sera of SLE patients without overt retroviral infections. These antibodies may represent cross-reactive antibodies or may have been induced by proteins encoded by endogenous retroviral sequences. We used Tera-1 cells that abundantly express a group-specific antigen of human endogenous retroviruses, HERV-K10gag polyprotein, to investigate its processing during apoptosis. Tera-1 cells induced to undergo apoptosis showed an altered HERV-K10gag processing compared with viable cells. In addition, granzyme B was able to cleave HERV-K10gag isolated from viable Tera-1 cells. Similar to nuclear autoantigens, endogenous retroviral proteins are cleaved during the execution phase of apoptosis. These post-translational modifications may result in the generation of T-cell neoepitopes or a changed epitope hierarchy of retroviral proteins. Therefore, immunogenicity of retroviral antigens in SLE patients may result from a similar mechanism as described for nuclear autoantigens.

5,6-carboxyfluorescein diacetate succinimidyl ester-labeled apoptotic and necrotic as well as detergent-treated cells can be traced in composite cell samples.

Detection of dividing cells by staining with 5,6-carboxyfluorescein diacetate succinimidyl ester (CFSE) has been widely used in flow-cytometric protocols. We analyzed the fate of CFSE in cells undergoing apoptotic or necrotic cell death, respectively. Peripheral blood mononuclear cells (PBMC) were stained with CFSE. Apoptosis was induced by UVB irradiation and necrosis by incubation at 56 degrees C for 30 min. In some experiments, labeled cells were permeabilized with detergent and CFSE association with nuclei was assessed. We observed that (i) CFSE remains stably detectable in apoptotic and necrotic cells; (ii) CFSE remains stably associated with the nuclei of cells even after their lysis by detergent; (iii) CFSE labeling does not interfere with the induction of cell death; and (iv) CFSE is not transferred from stained dying cells to unstained neighboring counterparts. We conclude that, in addition to tracking viable cells, CFSE can be used to trace dying cells in composite samples. We demonstrated that CFSE labeling does not influence the induction and the execution of apoptosis or necrosis.

Activity of Na+/K+-ATPase and of Ca++-ATPase under the action of adenosine triphosphate in experimental myocardial hypertrophy.

This paper presents the results of our research on the mechanisms involved in the modifications occurring in the activity of ionic pumps (Na+/K+-ATPase and Ca++-ATPase) at the level of the sarcolemma, the sarcoplasmic reticulum and mitochondrial membrane of the myocardial cell in experimental myocardial hypertrophy induced by isoproterenol. We also studied the effects of concomitant administration of adenosine triphosphate. Thus, we found the activity of the sarcolemmal Ca++-ATPase intensely increased under the action of isoproterenol, while Ca++-ATPase activity in the sarcoplasmic reticulum decreased significantly. At the same time, the Na+/K+-ATPase activity decreased both in the sarcolemma and in the sarcoplasmic reticulum. Administration of adenosine triphosphate induces opposite effects, of lower intensity, upon the activity of the two ATPase, that tend to offset the the effects of isoproterenol. This was found in the group of rats given concomitantly both isoproterenol and adenosine triphosphate. A better understanding of the processes involved in those modifications of membrane ATPase activity allows us to consider their different behaviour to isoproterenol and adenosine triphosphate as the expression of intrinsic mechanisms by means of which the myocardial cell intervenes in maintaining a physiological concentration of calcium ions (Ca++) that is necessary both in order to avoid a failure of the contractile apparatus and in order to preserve the mitochondria role of ATP supplier. Our results demonstrate the antiadrenergic action of adenosine triphosphate that attenuates isoproterenol effects on cardiac myocytes.

Research on some metabolic and enzymatic modifications in experimental hypertensive myocardium.

This paper presents our research on metabolic and enzymatic changes in the experimental, Isoproterenol-induced (ISO) hypertensive myocardium of rats. We analyze the effects produced by the simultaneous administration of adenosinetriphosphate (fosfobion) (FOS) and Isoproterenol on the changes in the body weight/heart weight ratio, and on the biochemical changes of cardiac metabolism. We studied the following parameters in the myocardium tissue and blood: plasmatic and tissular creatinin-phosphokinase, Na+K+ ATP-ase in the sarcolemma and the sarcoplasmic reticulum, Ca+2 ATP-ase in the mitochondrial membrane, sarcoplasmic reticulum and sarcolemma, as well as plasmatic and tissular lactate. Our data show an increase of heart weight to 939 mg, compared to 752 mg in the control group, while the heart weight/body weight ratio (mg/g), which was 3.8 in the control group, increased to 5.8 in the group to which Isoproterenol (ISO) was administered, and to 5.2 when fosfobion was associated. Investigation of myocardial metabolism has also shown the fact that under the influence of Isoproterenol, plasma creatinin-phosphokinase rises by 20%, while the association of fosfobion reduces it, in the myocardium tissue, down to 73%, in comparison with the values in the control group. Significant changes were found in the myocardium lactate that decreased by 26% under ISO influence, in comparison with normal values, and that decreased by 90% when FOS and ISO were administered together. This study produces arguments about metabolism-induced cardiac changes under the action of ISO and also contributes to the identification of ways that lead to cardiac hypertrophy. The experiment also demonstrates that the ATP-ases responsible for ion transportation across the membrane are actively involved in myocardium hypertrophy. The disturbances occurring in the investigated enzymatic systems are closed related with the myocardial metabolic ones. Fosfobion does not prevent the appearance and development of Isoproterenol-induced myocardial hypertrophy, but diminishes the increase of myocardial lactate produced by this synthesised catecholamine. At the same time, fosfobion significantly decreases the activity of Ca+2 ATP-ase in the plasmalemma and increases the activity of the Na+ - K+ ATP-ase both in the plasmalemma and in the sarcoplasmic reticulum, indirectly favouring the mechanical processes of cardiac myocytes relaxation. The study of the enzymatic activity of Na+K+ and Ca+2 ATP-ases in our experimental conditions contributes to a better understanding of the mechanisms that produce myocardial and coronary disturbances in myocardial hypertrophy.

Zinc and copper in plasma and erythrocytes of Down's syndrome children.

The concentration of zinc and copper in the plasma and the erythrocytes of 24 children with Down's syndrome was measured and compared with the values in a control group of normal children. Zinc and copper were determined in this biologic material by flame atomic absorption spectrophotometry. A significant (p less than 0.001) decrease of the plasma zinc content well as an increase of copper (p less than 0.024) and zinc (p less than 0.001) in the erythrocytes of Down's syndrome patients were found. The possible mechanisms of these changes are discussed.

Investigation of thermic transition and renaturation of high polymerized DNA extracted from thyroid nodule.

The study was carried out on 16 cases of mixed thyroid hypertrophy in which total or partial thyroidectomy was performed. Fragments collected intraoperatively from the thyroid nodule were used for extraction of high polymerized DNA. At the same time with quantitative evaluation of DNA, RNA and proteins were also assayed. The thermic transition mean temperature of the DNA extracted from the thyroid nodule is compared to other standards (DNA-HP-standard, calf thymus DNA, normal leukocytic DNA) and the thermic transition curves are presented. Hyperchromicity after thermic denaturation and renaturation is analysed and expressed in per cent values.