Microbubbles as a scattering contrast agent for grating-based x-ray dark-field imaging.

In clinically established-absorption-based-biomedical x-ray imaging, contrast agents with high atomic numbers (e.g. iodine) are commonly used for contrast enhancement. The development of novel x-ray contrast modalities such as phase contrast and dark-field contrast opens up the possible use of alternative contrast media in x-ray imaging. We investigate using ultrasound contrast agents, which unlike iodine-based contrast agents can also be administered to patients with renal impairment and thyroid dysfunction, for application with a recently developed novel x-ray dark-field imaging modality. To produce contrast from these microbubble-based contrast agents, our method exploits ultra-small-angle coherent x-ray scattering. Such scattering dark-field x-ray images can be obtained with a grating-based x-ray imaging setup, together with refraction-based differential phase-contrast and the conventional attenuation contrast images. In this work we specifically show that ultrasound contrast agents based on microbubbles can be used to produce strongly enhanced dark-field contrast, with superior contrast-to-noise ratio compared to the attenuation signal. We also demonstrate that this method works well with an x-ray tube-based setup and that the relative contrast gain even increases when the pixel size is increased from tenths of microns to clinically compatible detector resolutions about up to a millimetre.

Limbal stem cells transplantation in the reconstruction of the ocular surface: 6 years experience.

PURPOSE: To analyze the graft survival rate and stability of the corneal surface in patients who underwent limbal stem cell transplantation. Three surgical techniques were performed based on the origin of the ocular surface lesion: conjunctival limbal autograft (CLAU), living-related conjunctival limbal autograft (lr-CLAL), and keratolimbal allograft (KLAL) transplantations. METHODS: Nonrandomized consecutive comparative case series study. Eighty-four patients (90 eyes; 31 women and 53 men; age range: 11-78 years) were included in the study. Mean follow-up was 31.2 months (range: 6-72 months). Patients were divided into three groups: CLAU, lr-CLAL, and KLAL, comprising 21, 26, and 43 eyes, respectively. Graft survival rate and clinical success of the stem cell transplantation was confirmed by impression cytology. The Kaplan Meier survival curve and generalized Peto tests were used for the analyses. RESULTS: Graft survival rate and the regularity of the corneal surface differed significantly between the allo- and autografts. The 3-year and 6-year graft survival rates were 76.1% and 61.9%, respectively, for the autologous transplantation group, and 59.4% and 46.3%, respectively, for the allogeneic transplantation group. Corneal surface restoration correlated with positive staining for corneal epithelial cells in impression cytology. CONCLUSIONS: Significantly better long-term outcomes were achieved with autotransplantation of the limbus compared with allogeneic limbal grafts from living-related and cadaveric donors.

Platelet phospholipase A2 activity in patients with Alzheimer's disease, vascular dementia and ischemic stroke.

Phospholipase A(2) (E.C. 3.1.1.4, PLA(2)) plays an essential role in metabolism of membrane phospholipids, it is related to inflammatory reactions, secretion of amyloid precursor protein and activation of NMDA receptor after ischemia. In the present study we investigated PLA(2) activity in platelets from 37 Alzheimer's disease (AD) patients, 32 vascular dementia (VaD) patients and 32 individuals with ischemic stroke as compared to 27 healthy elderly controls. PLA(2) activity was determined using radiometric assay. Mean platelet PLA(2) activity was increased in individuals with Alzheimer's disease (p < 0.001). In VaD group the enzyme activity was between the values in AD and controls, these differences being significant from both groups. In the group of patients with ischemic stroke mean PLA(2) activity was higher either 48 h after the stroke or 7 days later (in both cases p < 0.001). The results may be particularly interesting in light of the fact, that inhibitors of PLA(2) activity are known.

Effects of lipids and antioxidants on PCB-mediated dysfunction of vascular endothelial cells (EC).

Our findings suggest that exposure to specific environmental contaminants can trigger diseases of the vasculature, e.g., cardiovascular disease. In addition, high-fat diets may potentiate and diets high in antioxidant nutrients may protect against PCB-mediated endothelial cell dysfunction. Our data give an insight into the potential use of vitamin E and related antioxidants to limit PCB-mediated cell injury. These studies are significant for providing new insights into potential nutrition interventions in diseases that can be induced by the toxicity of PCBs and other halogenated compounds.

4-Hydroxynonenal induces oxidative stress and death of cultured spinal cord neurons.

Primary spinal cord trauma can trigger a cascade of secondary processes leading to delayed and amplified injury to spinal cord neurons. Release of fatty acids, in particular arachidonic acid, from cell membranes is believed to contribute significantly to these events. Mechanisms of fatty acid-induced injury to spinal cord neurons may include lipid peroxidation. One of the major biologically active products of arachidonic acid peroxidation is 4-hydroxynonenal (HNE). The levels of HNE-protein conjugates in cultured spinal cord neurons increased in a dose-dependent manner after a 24-h exposure to arachidonic acid. To study cellular effects of HNE, spinal cord neurons were treated with different doses of HNE, and cellular oxidative stress, intracellular calcium, and cell viability were determined. A 3-h exposure to 10 microM HNE caused approximately 80% increase in oxidative stress and 30% elevation of intracellular calcium. Exposure of spinal cord neurons to HNE caused a dramatic loss of cellular viability, indicated by a dose-dependent decrease in MTS [3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-s ulfophenyl)- 2H-tetrazolium, inner salt] conversion. The cytotoxic effect of HNE was diminished by pretreating neurons with ebselen or N-acetylcysteine. These data support the hypothesis that formation of HNE may be responsible, at least in part, for the cytotoxic effects of membrane-released arachidonic acid to spinal cord neurons.

SYMTERM--program for modelling chemical processes in non-isothermal conditions

The program SYMTERM allows to perform the numerical simulations of chemical processes which take place in the thermal analysis apparatus, considering two important factors: the exchange of heat between a sample and an environment, and the influence of heat evolved in the reaction on its kinetics. The program makes it possible to obtain kinetics profiles (DTA, TG, DTG or DSC curves) at various heating rates and permits to determine both the activation energy Eact and the reaction order of a single rate-limited step reaction with using DTA (DSC) and TG data simultaneously.

Nicotine attenuates arachidonic acid-induced neurotoxicity in cultured spinal cord neurons.

Arachidonic acid release from cellular membranes due to spinal cord trauma may be one of the principal destructive events that can lead to progressive injury to spinal cord tissue. Exposure to arachidonic acid can compromise neuronal survival and viability. Because nicotine is known to be a neuroprotective agent, we propose that it can prevent arachidonic acid-induced neurotoxicity. To study this hypothesis, effects of nicotine on mitochondrial function, cellular energy content and apoptotic cell death were measured in cultured spinal cord neurons treated with arachidonic acid. Nicotine attenuated arachidonic acid-induced compromised cell viability and cellular ATP levels in spinal cord neurons. Nicotine exerted these protective effects when used at the concentration of 10 microM and only after a 2-h pre-treatment before a co-exposure to arachidonic acid. Antagonists of nicotinic receptors, such as alpha-bungarotoxin or mecamylamine, only partially reversed these neuroprotective effects of nicotine. In addition, nicotine prevented arachidonic acid-induced activation of caspase-3 activity and apoptotic cell death. These results indicate that nicotine pre-treatment can exert a protective effect against arachidonic acid-induced injury to spinal cord neurons.

Nicotine attenuates arachidonic acid-induced overexpression of nitric oxide synthase in cultured spinal cord neurons.

Primary spinal cord trauma can initiate a cascade of pathophysiologic events which markedly contribute to the expansion and amplification of the primary insult. The detailed mechanisms of these secondary neurochemical reactions are largely unknown; however, they involve membrane lipid derangements with the release of free fatty acids, in particular, arachidonic acid (AA). AA can induce several injury effects on spinal cord neurons. We hypothesize that upregulation of nitric oxide synthase (NOS) is among the most important mechanisms of arachidonic-acid-induced neuronal dysfunction and that nicotine can attenuate this effect. To study these hypotheses, spinal cord neurons were exposed to AA and/or nicotine, and several markers of neuronal nitric oxide synthase (nNOS) metabolism were measured. In addition, cotreatments with either inhibitors of nicotinic receptors or inhibitors of specific NOS isoforms were employed. Treatment with AA markedly increased activity of nNOS, as well as mRNA and protein levels of this enzyme. Changes in nNOS expression were accompanied by an increase in cellular cGMP and medium nitrite levels. Pretreatment with nicotine decreased AA-induced overexpression of nNOS and elevation of nitrite levels. In addition, it appeared that these nicotine effects could be partially modulated both by the alpha7 nicotinic receptors or by nonreceptor mechanisms. Alternatively, the observed changes could also be mediated by an alternate nicotinic receptor mechanism which is not blocked by alpha-bungarotoxin or mecamylamine. Results of the present study indicate that exposure to AA can lead to induction of nNOS in cultured spinal cord neurons. In addition, nicotine can exert a neuroprotective effect by attenuation of AA-induced upregulation of nNOS metabolism. These data may have therapeutic implications for the treatment of acute spinal cord trauma.

Zinc nutrition and apoptosis of vascular endothelial cells: implications in atherosclerosis.

Little is known about the requirements and function of zinc in maintaining endothelial cell integrity, especially during stressful conditions, such as the inflammatory response in cardiovascular disease. There is evidence that zinc requirements of the vascular endothelium are increased during inflammatory conditions such as atherosclerosis, where apoptotic cell death is also prevalent. Apoptosis is a morphologically distinct mechanism of programmed cell death which involves the activation of a cell-intrinsic suicide program, and there is evidence that factors such as inflammatory cytokines (e.g., tumor necrosis factor [TNF]) and pure or oxidized lipids are necessary to induce the cell death pathway. Because of its constant exposure to blood components, including prooxidants, diet-derived fats, and their derivatives, the endothelium is very susceptible to oxidative stress and to apoptotic injury mediated by blood lipid components, prooxidants, and cytokines. Thus, it is likely that the cellular lipid environment, primarily polyunsaturated fatty acids, can potentiate the overall endothelial cell injury by increasing cellular oxidative stress and cytokine release in proximity to the endothelium, which then could further induce apoptosis and disrupt endothelial barrier function. Our data suggest that zinc deficiency exacerbates the detrimental effects of specific fatty acids (e.g., linoleic acid) and inflammatory cytokines, such as TNF, on vascular endothelial functions. We propose that a major mechanism of zinc protection against disruption of endothelial cell integrity during inflammatory conditions, is by the ability of zinc to inhibit the pathways of signal transduction leading to apoptosis and especially mechanisms that lead to upregulation of caspase genes.

Arachidonic acid-induced oxidative injury to cultured spinal cord neurons.

Spinal cord trauma can cause a marked release of free fatty acids, in particular, arachidonic acid (AA), from cell membranes. Free fatty acids, and AA by itself, may lead to secondary damage to spinal cord neurons. To study this hypothesis, cultured spinal cord neurons were exposed to increasing concentrations of AA (0.01-10 microM). AA-induced injury to spinal cord neurons was assessed by measurements of cellular oxidative stress, intracellular calcium levels, activation of nuclear factor-KB (NF-kappaB), and cell viability. AA treatment increased intracellular calcium concentrations and decreased cell viability. Oxidative stress increased significantly in neurons exposed to 1 and 10 microM AA. In addition, AA treatment activated NF-kappaB and decreased levels of the inhibitory subunit, IKB. It is interesting that manganese superoxide dismutase protein levels and levels of intracellular total glutathione increased in neurons exposed to this fatty acid for 24 h, consistent with a compensatory response to increased oxidative stress. These results strongly support the hypothesis that free fatty acids contribute to the tissue injury observed following spinal cord trauma.

Effect of neuroleptics on phospholipase A2 activity in the brain of rats.

The effect of neuroleptics on phospholipase A2 (PLA2) activity in rat brain plasma membranes was studied. Chlorpromazine (10 mg/kg), fluphenazine (5 mg/kg), thioridazine (5 mg/kg), trifluoperazine (5 mg/kg), haloperidol (2 mg/kg), and sulpiride (100 mg/kg) were administered to rats intraperitoneally as a single dose or long-term treatment (4 weeks). The PLA2 activity was determined 24, 48, and 72 h after the last injection of a drug. The enzyme activity was decreased after a single or 4-week administration of chlorpromazine, trifluoperazine, haloperidol, and sulpiride. Fluphenazine and thioridazine caused an increase of PLA2 activity in rat brain both after a single dose and long-term administration. For the first time it was shown that neuroleptics cannot only inhibit but also increase, PLA2 activity. Elucidation of this fact requires further studies.

Effect of antidepressants on ATP content, 3H-valine incorporation and cell morphometry of astrocytes cultured from rat brain.

ATP is a potential marker of cell vialibility and growth. The content of ATP and 3H-valine incorporation into proteins were measured and the morphometry was performed after antidepressant treatment of astrocytes cultured in vitro with or without dibutyryl 3'5'-cyclic adenosine monophosphate (dB-cAMP). Antidepressants were added into the culture medium (for 24 h) at a final concentration of 10(-4)M (imipramine, amitriptyline, clomipramine, doxepine, mianserin) or 10(-5)M (maprotiline). It was shown that all antidepressants except maprotiline and imipramine increased ATP level and decreased 3H-valine incorporation into astrocytes. All drugs except clomipramine and maprotiline, diminished cell area and perimeter of astrocytes. The addition of dB-cAMP to cultures caused an increase of astrocyte form factor. It can be concluded that antidepressants have a significant effect on energy metabolism and differentiation of astrocytes cultured in vitro.

BmCF1, a Bombyx mori RXR-type receptor related to the Drosophila ultraspiracle.

A PCR approach has been used to obtain an ovarian cDNA clone from the silkmoth Bombyx mori, encoding a 50 kDa protein (BmCF1) that belongs to the RXR subfamily of nuclear hormone receptors and is most similar to the CF1/USP protein (DmCF1) encoded by the ultraspiracle gene of Drosophila. The similarity is high in the DNA-binding and moderately so in the ligand-binding domains, although not in the N-terminal, putatively activator A/B domain. Protein sequence comparisons with the available RXR sequences indicate that although insect USP-like sequences are more related to each other than to vertebrate RXRs, their inter se similarities are lower than in the case of the vertebrate RXRs. Two distinct BmCF1-homologous transcripts are observed consistently, and are indicative either of alternative splicing or of the existence of a second RXR gene in the moth. The transcripts are widely distributed, suggesting functions at multiple developmental stages, as in the case of Drosophila ultraspiracle.

Aging modulates calcium-dependent phosphatidylinositol degradation by cerebral cortex synaptic plasma membrane phospholipases.

The synaptic plasma membrane (SPM) and cytosol fractions from cerebral cortex of adult (4-mo-old) and aged (27-mo-old) rats were used as a source of phospholipase A2 (PLA2) and phospholipase C (PLC). The activity of PLC acting on [3H-inositol]phosphatidylinositol ([3H]PtdIns) was investigated in the presence of endogenous and 2 mM Ca2+. Arachidonic acid (AA) release was studied in the same conditions, using 1-stearoyl-[2-14C]arachidonyl-sn-glycerophosphoinositol ([14C]PtdIns) as a substrate. In the presence of endogenous Ca2+ (i.e., no added Ca2+) SPM-bound PLC and PLA2 or diacylglycerol (DAG) lipase of aged brain exert significantly higher activity in degradation of PtdIns as compared to their activities in adult brain. Moreover, these enzymes of aged brain are less or not further activated by 2 mM Ca2+, contrary to the enzymes isolated from adult brain. The activity of cytosolic enzymes involved in degradation [3H]PtdIns and [14C]PtdIns and their regulation by Ca2+ ions are not significantly changed in senescent cerebral cortex as compared to the adult. The intracellular calcium concentration ([Ca2+]i), measured with fura-2, is lower in aged brain compared to adult brain, which may suggest the modification in Ca2+ ion redistribution in aged brain and probably its higher concentration in membranes. These results indicate that aging modifies significantly the activity of membrane-bound, Ca(2+)-dependent phospholipase(s) degrading PtdIns, which may be connected with alteration of Ca2+ ion redistribution and may influence the formation and accumulation of very potent lipid messengers as diacylglycerol, lysophospholipid, and arachidonic acid, known to be involved in neurotransmission processes.

Behavioral effects of long-term exposure to magnetic fields in rats.

Male rats and pregnant and nonpregnant female rats of the Wistar strain were sham-exposed or exposed to static (0.49 T) or to extremely low frequency (50 Hz) magnetic fields (0.018 T) 2 h per day for 20 consecutive days. Measures of irritability, exploratory activity, and locomotion were made in that order before and after the 4th, 10th, and 17th 2-h exposures. A reliable decrease in the irritability of rats after repeated exposure to a static or undulating field was found. No significant effects of treatment conditions on open-field behavior and locomotor activity were observed. Pregnancy had no influence on the behavioral end points. These results indicate that irritability of rats may be used as a simple behavioral indicant of mammalian sensitivity to magnetic fields.

[Gangliosides in neurological pharmacotherapy]. / Gangliozydy w farmakoterapii neurologicznej.

Gangliosides take part in synaptic transmission, neuronal metabolism and development of nervous tissue. They cooperate with nerve growth factor (NGF) and have positive influence on regeneration of the nervous system impairments. There exist many behavioural and biochemical evidences of gangliosides participation in the regeneration of experimentally injured animal nervous system. The therapeutic effectivity of gangliosides in clinical practice is encouraging. Commercial preparates of gangliosides (Cronassial, Sygen) have been successfully used in the therapy of chronic neuropathies, strokes and subarachnoidal haemorrhages. Among the adverse reactions to these drugs are: local irritation, anxiety and possible detrimental effect in immunological system. Ganglioside preparations need further clinical examinations.