CSF proteome analysis in clinically isolated syndrome (CIS): candidate markers for conversion to definite multiple sclerosis.

Cerebrospinal fluid (CSF) is a promising source of biomarkers in clinically isolated syndrome (CIS), which frequently presents as a first episode of multiple sclerosis (MS). Using the two-dimensional difference in gel electrophoresis (2-D DIGE), we compared CSF samples from patients with CIS that remained CIS (CIS-CIS, n=8) over a follow-up time of 2 years and from patients with CIS that developed definite MS of the relapsing-remitting subtype (CIS-RRMS, n=8) over the same period. Protein spots that showed significant differences between patients and controls were selected for further analysis by MALDI-TOF mass spectrometry. For validation of identified spots ELISA experiments were performed. We identified one protein that was upregulated in CIS-RRMS (serin peptidase inhibitor) and eight proteins (alpha-1-B-glycoprotein, Fetuin-A, apolipoprotein A4, haptoglobin, human Zinc-alpha-2-glycoprotein (ZAG), Retinol-binding protein, superoxid dismutase 1, transferrin) that were down-regulated in CIS-RRMS vs. CIS-CIS. For Fetuin-A, our findings could be confirmed by ELISA. The pathophysiological role as well as clinical relevance of these candidate proteins in CIS remains to be further clarified by future studies.

R56865 is antifibrillatory in reperfused ischemic guinea-pig hearts, even when given only during reperfusion.

R56865 was previously characterized as an inhibitor of Na and Ca overload that has beneficial effects during ischemia and reperfusion. An isolated guinea-pig heart preparation was subjected to 60 minutes of ischemia and 60 minutes of reperfusion. R56865 was given before ischemia and with the onset of reperfusion, applying different dosing schedules, including an initial loading dose. R56865 below 0.1 mumol/l had no cardiodepressant effects in normoxic hearts and at 0.1 mumol/l reduced left ventricular pressure marginally. The onset of ischemic contracture was delayed only at this concentration. R56865 given before ischemia potently inhibits delayed sustained fibrillation occurring during reperfusion in the concentration range between 0.01 mumol/l and 0.1 mumol/l. Analysis of cellular Na+, K+, and Ca2+ concentrations revealed that R56865 substantially improves the ionic homeostasis of myocardial cells. Most importantly, the compound also reduced the incidence of delayed sustained fibrillation when given at the onset of reperfusion. R56865 was most effective when fast equilibration of drug with tissue was achieved by giving an initial loading dose. In particular, the cellular Na+ and Ca2+ contents were improved using this dosing scheme. The results are compatible with the classification of R56865 as an inhibitor of Na+ and Ca2+ overload.

Effect of Na+ reduction and monensin on ion content and contractile response in normoxic and ischaemic reperfused rat hearts.

The possibility was explored whether the functional properties of Na+/Ca2+ exchange are altered after ischaemia, thereby contributing to the elevated intracellular (i) Ca2+ levels in ischaemic reperfused hearts. The intracellular Na+, K+ and Ca2+ contents in rat Langendorff heart preparations were determined by atomic absorption spectrometry under normoxic conditions, after ischaemia (30 min) and after ischaemia (30 min) plus reperfusion (30 min). In addition, the influence of modulating the Na+ gradient (Na+o/Na+i) across the sarcolemma was studied with respect to cardiac contractility and intracellular ion content. This was done by either decreasing extracellular (o) Na+ or by increasing Na+i with monensin, both in normoxic and reperfused hearts. Both Na+o reduction and monensin led to an increase in contractility and coronary flow, an effect which was nearly abolished in reperfused hearts. Under normoxic conditions the intracellular ion contents amounted to Na+ = 12.4 +/- 0.4, K+ = 99.0 +/- 3.1 and Ca2+ = 0.64 +/- 0.02 mmol/kg cell (means +/- SEM, n = 7). In normoxic hearts, lowering Na+o reduced and monensin increased Na+i, thereby both leading to a decrease in Na+ gradient; no effect on total Ca2+i content was observed. Na+i increased twofold after ischaemia as compared to the normoxic situation, an effect which was aggravated (4 fold increase) in reperfused hearts. The opposite effects were observed for K+i with a 25% decrease after ischaemia and a 40% decrease in reperfused hearts. Only after ischaemia plus reperfusion was Ca2+i increased (6 fold).(ABSTRACT TRUNCATED AT 250 WORDS)

Ouabain-induced changes of calcium and potassium in slices of hippocampus of the rat: comparison to hypoxia and effect of R 56865.

Simple and reliable in vitro models of cerebral ischaemia are important for the identification of antiischaemic/antihypoxic compounds. Alterations of the concentrations of potassium and calcium were recorded in slices of hippocampus of the rat. The slices were subjected to hypoxia in the presence and absence of intoxication with glucose or ouabain (1 mmol/l). Normoxic slices of hippocampus showed an extracellular space of 57% and a tissue concentration of potassium of 45 mmol/kg wet wt. A cellular concentration of potassium of 92 mmol/kg was calculated. Hypoxia, in the presence of glucose, only slightly reduced tissue concentrations of potassium and did not influence concentrations of calcium. Omission of glucose during hypoxia led to tissue concentrations of potassium below 10 mmol/kg, within 10-30 min of hypoxia. Concentrations of calcium only increased from 3.3 to 3.5 mmol/kg after 30 min of hypoxia, without glucose. Intoxication with ouabain is proposed as alternative experimental model of ionic movements, associated with cerebral ischaemia/hypoxia. Tissue concentrations of potassium fell rapidly to values below 10 mmol/kg, within 5 min and concentrations of calcium rose to 5.2 mmol/kg, within 30 min of intoxication with ouabain. In quantitative terms, the model for cerebral ischaemia with intoxication with ouabain is suggested to be superior to the model based on hypoxia without glucose. To verify intoxication with ouabain as an experimental model for ischaemic/hypoxic insults, the effect of an investigational drug with antiischaemic/hypoxic properties (R 56865) was evaluated in the model. The drug R 56865 produced dose-dependent attenuation of the fall in tissue concentrations of potassium, between 3 x 10(-7) and 5 x 10(-6) mol/l.(ABSTRACT TRUNCATED AT 250 WORDS)