Identification of proteins bearing beta1-6 branched N-glycans in human melanoma cell lines from different progression stages by tandem mass spectrometry analysis

The common structural alterations in the cell-surface glycoproteins concern the highly elevated expression of tri- and tetra-antennary beta1-6-N-acetylglucosamine (beta1-6 GlcNAc) bearing N-glycans, which are recognised by Phaseolus vulgaris agglutinin (PHA-L). In this report we identified proteins bearing beta1-6 GlcNAc branched N-glycans in three human melanoma cell lines: WM35--from the primary tumour site, as well as WM239 and WM9 from different metastatic sites: the skin and the lymph node, respectively, by tandem mass spectrometry (MS/MS) on PHA-L agarose bound material, followed by immunochemical identification. Our results show that melanoma cell lines differ from each other in the number of N-glycoproteins bearing beta1-6 GlcNAc branched oligosaccharides. Among identified proteins the largest group consists of integrin subunits. In addition, L1-CAM, Mac-2 binding protein, melanoma cell adhesion molecule, intercellular adhesion molecule, melanoma associated antigen, tumour rejection antigen-1, melanoma-associated chondroitin sulfate proteoglycan 4 and lysosome-associated membrane protein (LAMP-1) were found. It was indicated that WM35 cell line showed the lowest number of proteins possessing beta1-6 GlcNAc branched N-glycans in comparison to metastatic WM9 and WM239 cell lines. Our data suggest that changes in the number of proteins being a substrate for GlcNAc-TV are better correlated with melanoma development and progression than with expression of cell adhesion molecules.     

Ion conductance pathways in potato tuber (Solanum tuberosum) inner mitochondrial membrane

Single-ion channel activities were measured after reconstitution of potato tuber mitochondrial inner membranes into planar lipid bilayers. In addition to the recently described large-conductance Ca(2+)-activated potassium channel activity (Koszela-Piotrowska et al., 2009), the following mitochondrial ion conductance pathways were recorded: (i) an ATP-regulated potassium channel (mitoK(ATP) channel) activity with a conductance of 164+/-8pS, (ii) a large-conductance Ca(2+)-insensitive iberiotoxin-sensitive potassium channel activity with a conductance of 312 pS+/-23, and (iii) a chloride 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS)-inhibited channel activity with a conductance of 117 pS+/-4. In isolated non-phosphorylating potato tuber mitochondria, individual and combined potassium channel activities caused significant (up to 14mV) but not collapsing K(+)-influx-induced membrane potential depolarisation. Under phosphorylating conditions, the coupling parameters were unchanged in the presence of high K(+) level, indicating that plant K(+) channels function as energy-dissipating systems that are not able to divert energy from oxidative phosphorylation. A potato tuber K(+) channel that is ATP-, 5-hydroxydecanonic acid-, glybenclamide-inhibited and diazoxide-stimulated caused low cation flux, modestly decreasing membrane potential (up to a few mV) and increasing respiration in non-phosphorylating mitochondria. Immunological analysis with antibodies raised against the mammalian plasma membrane ATP-regulated K(+) channel identified a pore-forming subunit of the Kir-like family in potato tuber mitochondrial inner membrane. These results suggest that a mitoK(ATP) channel similar to that of mammalian mitochondria is present in potato tuber mitochondria.     

Dietary Intake and Serum and Hair Concentrations of Minerals and their Relationship with Serum Lipids and Glucose Levels in Hypertensive and Obese Patients with Insulin Resistance

Inadequate minerals intake, as well as disruption of some metabolic processes in which microelements are cofactors, are suggested to lead to the development of hypertension. The role of minerals in the pathogenesis of hypertension still remains to be explained. In the present study, we sought to determine associations between serum and hair mineral concentrations and serum lipids and glucose levels. Forty obese hypertensive subjects with insulin resistance and 40 healthy volunteers were recruited in the study. Blood pressure, BMI, and insulin resistance were recorded in all subjects. Levels of lipids, glucose, sodium and potassium, iron, copper, zinc, magnesium, and calcium were assessed in serum. Iron, copper, zinc, magnesium, and calcium were assessed in hair. Dietary intake of the analyzed minerals was estimated. We found distinctly higher concentrations of serum iron and serum and hair calcium as well as markedly lower levels of hair zinc in the hypertensive subjects. The study group manifested also significantly lower daily intake of calcium, magnesium, and iron. We observed a relationship between the concentrations of iron, zinc, and copper in serum and hair and high and low range of cholesterol, triglycerides, and glucose serum levels in the studied patients. Moreover, this study demonstrated significant correlation between serum and hair concentrations of selected minerals and their dietary intake and levels of serum lipids and glucose and blood pressure in the study and the control groups. The obtained results seem to indicate the association between lipid and glucose metabolism and iron, copper, zinc, and calcium concentrations in blood and hair of hypertensive and obese patients with insulin resistance.     

Synthesis and anticonvulsant activity of new N-Mannich bases derived from 5-cyclopropyl-5-phenyl- and 5-cyclopropyl-5-(4-chlorophenyl)-imidazolidine-2,4-diones

Synthesis, physicochemical and anticonvulsant properties of new N-Mannich bases derived from 5-cyclopropyl-5-phenyl- and 5-cyclopropyl-5-(4-chlorophenyl)-imidazolidine-2,4-diones have been described. Initial anticonvulsant screening was performed using intraperitoneal (ip.) maximal electroshock (MES) and subcutaneous pentylenetetrazole (scPTZ) seizure tests. The neurotoxicity was determined applying the rotarod test. The in vivo results in mice showed that all compounds were effective especially in the MES screen. The quantitative evaluation after oral administration in rats showed that the most active was 5-cyclopropyl-5-phenyl-imidazolidine-2,4-dione (1) with ED(50) values of 5.76 mg/kg (MES) and 57.31 mg/kg (scPTZ). This molecule was more potent than phenytoin and ethosuximide which were used as reference antiepileptic drugs. Additionally compound 1 with ED(50) of 26.06 mg/kg in psychomotor seizure test (6-Hz) in mice showed comparable activity to new generation anticonvulsant - levetiracetam.     

Xanthophyll cycle--a mechanism protecting plants against oxidative stress

Six different xanthophyll cycles have been described in photosynthetic organisms. All of them protect the photosynthetic apparatus from photodamage caused by light-induced oxidative stress. Overexcitation conditions lead, in the chloroplast, to the over-reduction of the NADP pool and production of superoxide, which can subsequently be metabolized to hydrogen peroxide or a hydroxyl radical, other reactive oxygen species (ROS). On the other hand, overexcitation of photosystems leads to an increased lifetime of the chlorophyll excited state, increasing the probability of chlorophyll triplet formation which reacts with triplet oxygen forming single oxygen, another ROS. The products of the light-dependent phase of xanthophyll cycles play an important role in the protection against oxidative stress generated not only by an excess of light but also by other ROS-generating factors such as drought, chilling, heat, senescence, or salinity stress. Four, mainly hypothetical, mechanisms explaining the protective role of xanthophyll cycles in oxidative stress are presented. One of them is the direct quenching of overexcitation by products of the light phase of xanthophyll cycles and three others are based on the indirect participation of xanthophyll cycle carotenoids in the process of photoprotection. They include: (1) indirect quenching of overexcitation by aggregation-dependent light-harvesting complexes (LHCII) quenching; (2) light-driven mechanisms in LHCII; and (3) a model based on charge transfer quenching between Chl a and Zx. Moreover, results of the studies on the antioxidant properties of xanthophyll cycle pigments in model systems are also presented.     

Lead as an inductor of some morphological and functional changes in synaptosomes from rat brain

Summary 1. The effect of lead (in vivo) on the uptake of GABA, dopamine, and histidine as a precursor of histamine in synaptosomes obtained from chronically lead-treated rats was studied.2. Lead decreased the uptake of GABA, increased the uptake of dopamine, and did not change the uptake of histidine. These effects were independent of calcium concentration.3. Lead administration to the rat changed the morphology of the synaptosomes, as manifested in the decreased number of synaptic vesicles and disturbed mitochondrial structure.4. The results suggest the existence of several mechanisms of lead toxicity on uptake, related to individual neurotransmitters, which are not necessarily connected with a Pb²⁺/Ca²⁺ interaction.     

Ghrelin ovarian cell expression in patients with polycystic ovary syndrome: an immunohistochemical evaluation.

INTRODUCTION: The influence of ghrelin on different organs has been studied recently, e.g. in the regulation of pituitary hormone release, regulation of energy homeostasis, glucose metabolism and insulin secretion, cell proliferation, and reproductive function. However, the etiology of polycystic ovary syndrome has not been fully explained. The aim of our study was to estimate the presence of ghrelin in polycystic ovaries cells and evaluation of the relationship between ghrelin occurrence and cells proliferation. METHODS: In the present work we have compared ten polycystic ovaries with ovaries without pathology as the control group. We used immunohistochemical method to detect ghrelin. The cells proliferation was evaluated by Ki 67 proliferation index. RESULTS: Ghrelin immunostaining was demonstrated in cytoplasm of ovarian secondary interstitial cells and in atretic corpus luteum. The cell nuclei were ghrelin positive in granulosa, theca layers of follicular cyst in both groups as well as in luteal cells of young corpus luteum in healthy ovaries. Ki 67 immunostaining was observed in granulosa and theca layers of follicular cyst in polycystic and healthy ovaries. CONCLUSIONS: It is possible that local ghrelin expression plays an important role in the direct control of ovarian development and function and ghrelin may participate in patomechanism of PCOS.     

Fibroblast-like synoviocytes from rheumatoid arthritis patients express functional IL-15 receptor complex: endogenous IL-15 in autocrine fashion enhances cell proliferation and expression of Bcl-x(L) and Bcl-2

The hallmarks of rheumatoid arthritis (RA) are leukocytic infiltration of the synovium and expansiveness of fibroblast-like synoviocytes (FLS). The abnormal proliferation of FLS and their resistance to apoptosis is mediated, at least in part, by present in RA joints proinflammatory cytokines and growth factors. Because IL-15 exerts properties of antiapoptotic and growth factors, and is produced by RA FLS, we hypothesized that IL-15 participates in RA FLS activation. To test this hypothesis, we first examined whether RA FLS express chains required for high affinity functional IL-15R. Indeed, RA FLS express IL-15Ralpha at mRNA and protein levels. Moreover, we confirmed the presence of IL-2Rbeta and common gamma-chains. Interestingly, TNF-alpha or IL-1beta triggered significant elevation of IL-15Ralpha chain at mRNA and protein levels. Next, we investigated the effects of exogenous or endogenous IL-15 on Bcl-2 and Bcl-x(L) expression, FLS proliferation, and apoptosis. Exogenous IL-15 enhanced RA FLS proliferation and increased the level of mRNA-encoding Bcl-x(L). To test the role of endogenous IL-15 in the activation of RA FLS, an IL-15 mutant/Fcgamma2a protein exerting properties of specific antagonist to the IL-15Ralpha chain was used. We found that blocking IL-15 biological activities using this protein substantially reduced endogenous expression of Bcl-2 and Bcl-x(L), and RA FLS proliferation that was reflected by increased apoptosis. Thus, we have demonstrated that a distinctive phenotype of RA FLS, i.e., persistent activation, proliferation, and resistance to apoptosis, is related to the autocrine activation of IL-15Rs by FLS-derived IL-15.     

Structure of trichomonads as revealed by scanning electron microscopy.

A scanning electron microscope (SEM) study of Hypotrichomonas acosta (Moskowitz), Trichomonas vaginalis Donné, Pentatrichomonas hominis (Davaine), and Tritrichomonas foetus (Riedmüller) provided new information about the structure of the periflagellar canal; emergence of the flagella from the cell body; structure of the undulating membrane; and position, shape, and size of the pelta. Of special interest were the spatial relationships of the attached part of the recurrent flagellum and the accessory filament in Hypotrichomonas and in the members of Trichomonadinae, i.e. Trichomonas and Pentatrichomonas.