Assessment of cerebral perfusion in post-traumatic brain injury patients with the use of ICG-bolus tracking method.

The aim of this study was to verify the usefulness of the time-resolved optical method utilizing diffusely reflected photons and fluorescence signals combined with intravenous injection of indocyanine green (ICG) in the assessment of brain perfusion in post-traumatic brain injury patients. The distributions of times of flight (DTOFs) of diffusely reflected photons were acquired together with the distributions of times of arrival (DTAs) of fluorescence photons. The data analysis methodology was based on the observation of delays between the signals of statistical moments (number of photons, mean time of flight and variance) of DTOFs and DTAs related to the inflow of ICG to the extra- and intracerebral tissue compartments. Eleven patients with brain hematoma, 15 patients with brain edema and a group of 9 healthy subjects were included in this study. Statistically significant differences between parameters obtained in healthy subjects and patients with brain hematoma and brain edema were observed. The best optical parameter to differentiate patients and control group was variance of the DTOFs or DTAs. Results of the study suggest that time-resolved optical monitoring of inflow of the ICG seems to be a promising tool for detecting cerebral perfusion insufficiencies in critically ill patients.

Effectiveness of water treatment for the removal of Cryptosporidium and Giardia spp.

Cryptosporidium and Giardia are intestinal parasites of humans and of many other species of animals. Water constitutes an important route of transmission for human infections in both developed and developing countries. In Poland, contamination of water sources with oocysts/cysts is not routinely monitored and scientific research in this field is scarce. Our aim was to compare the contamination of surface and treated water and thus the success of water treatment processes. Water samples (n=94) of between 30 l (surface water) to over 1000 l for tap water, were taken in the period of 2008-2009 using specially constructed equipment with cartridge filtration (Filta-Max; IDEXX, USA). Immunofluorescent assay, and nested polymerase chain reaction were used for the detection of parasites. Cryptosporidium oocysts were found in 85% of surface water and in 59% of raw (intake) water samples. Oocysts were also detected in treated water (16%) but were absent in samples of swimming pool water. The highest mean number of Cryptosporidium oocysts [geometric mean (GM)=61/10 l] was found in samples of rinsing water. Giardia cysts were observed in 61% of surface water samples, in 6% of raw water and in 19% of treated water, with the highest number of cysts noted in rinsing water samples (GM=70 cysts/10 l). Our study highlights the frequent occurrence of parasites in surface waters in Poland and the effectiveness of water treatment for the removal of parasites from drinking water.

NMR-Based Metabolomics of Rat Hippocampus, Serum, and Urine in Two Models of Autism.

Autism spectrum disorders (ASDs) are increasingly diagnosed as developmental disabilities of unclear etiology related to genetic, epigenetic, or environmental factors. The diagnosis of ASD in children is based on the recognition of typical behavioral symptoms, while no reliable biomarkers are available. Rats in whom ASD-like symptoms are due to maternal administration of the teratogenic drugs valproate or thalidomide on critical day 11 of pregnancy are widely used models in autism research. The present studies, aimed at detecting changes in the levels of hydrophilic and hydrophobic metabolites, were carried out on 1-month-old rats belonging to the abovementioned two ASD models and on a control group. Analysis of both hydrophilic and hydrophobic metabolite levels gives a broader view of possible mechanisms involved in the pathogenesis of autism. Hippocampal proton magnetic resonance (MRS) spectroscopy and ex vivo nuclear magnetic resonance (NMR) analysis of serum and urine samples were used. The results were analyzed using advanced statistical tests. Both the results of our present MRS studies of the hippocampus and of the NMR studies of body fluids in both ASD models, particularly from the THAL model, appeared to be consistent with previously published NMR results of hippocampal homogenates and data from the literature on autistic children. We detected symptoms of disturbances in neurotransmitter metabolism, energy deficit, and oxidative stress, as well as intestinal malfunction, which shed light on the pathogenesis of ASD and could be used for diagnostic purposes. These results confirm the usefulness of the noninvasive techniques used in ASD studies.

Hippocampal Metabolite Profiles in Two Rat Models of Autism: NMR-Based Metabolomics Studies.

Autism spectrum disorders (ASDs) are increasingly being diagnosed. Hypotheses link ASD to genetic, epigenetic, or environmental factors. The role of oxidative stress and the imbalance between excitatory and inhibitory neurotransmission in the pathogenesis of ASD has been suggested. Rats in which ASD symptoms are induced by valproate (VPA) or thalidomide (THAL) application in utero are useful models in ASD studies. Our study investigated whether rats in ASD models show changes in metabolite levels in the brain consistent with the hypothetical pathomechanisms of ASD. Female rats were fed one dose of 800 mg/kg VPA or 500 mg/kg THAL orally on the 11th day of gestation, and 1-month offspring were used for the experiments. Metabolic profiles from proton nuclear magnetic resonance spectroscopy of hydrophilic and hydrophobic extracts of rat hippocampi were subjected to OPLS-DA statistical analysis. Large differences between both models in the content of several metabolites in the rat hippocampus were noticed. The following metabolic pathways were identified as being disturbed in both ASD models: steroid hormone biosynthesis; fatty acid biosynthesis; the synthesis and degradation of ketone bodies; glycerophospholipid metabolism; cholesterol metabolism; purine metabolism; arginine and proline metabolism; valine, leucine, and isoleucine biosynthesis and degradation. These results indicate disorders of energy metabolism, altered structure of cell membranes, changes in neurotransmission, and the induction of oxidative stress in the hippocampus. Our data, consistent with hypotheses of ASD pathomechanisms, may be useful in future ASD studies, especially for the interpretation of the results of metabolomics analysis of body fluids in rat ASD models.

Altered Cerebrospinal Fluid Concentrations of Hydrophobic and Hydrophilic Compounds in Early Stages of Multiple Sclerosis-Metabolic Profile Analyses.

The lack of a single predictive or diagnostic test in multiple sclerosis (MS) remains a major obstacle in the patient's care. The aim of this study was to investigate metabolic profiles, especially lipids in cerebrospinal fluid (CSF) using 1H-NMR spectroscopy and metabolomics analysis to discriminate MS patient group from the control ones. In this study, 19 MS patients and 19 controls, without neurological problems, patients were enrolled. To obtain the CSF metabolic profiles, NMR spectroscopy was used. Hydrophilic and hydrophobic compounds were analyzed using univariate and multivariate supervised analysis orthogonal partial least square discriminant analysis (OPLS-DA). Targeted OPLS-DA analysis of 32 hydrophilic and 17 hydrophobic compounds obtained 9 hydrophilic metabolites and 8 lipid functional groups which had the highest contribution to patient's group separation. Lower concentrations of CSF hydrophilic and hydrophobic compounds were observed in MS patients as compared to control group. Acetone, choline, urea, 1,3-dimethylurate, creatinine, isoleucine, myo-inositol, leucine, and 3-OH butyrate; saturated and monounsaturated acyl groups of ω-9, ω-7, ω-6, ω-3, and fatty acid, triglycerides, 1,3-DG, 1-MG, and unassigned component signal at 3.33 ppm were the most important signal compounds in group separation. Analysis of metabolic profile of raw CSF and their lipid extract shows decreased levels of many compounds and led to the conclusion that MS patients could have a disturbance in many metabolic pathways perhaps leading to the decreased level of acetyl-CoA and/or inflammation. CSF metabolic profile analyses could be used as a fingerprint for early MS diagnosis.

High-resolution 1H nuclear magnetic resonance spectroscopy analysis of bile samples obtained from a patient after orthotopic liver transplantation: new perspectives.

The liver is involved in many synthetic and metabolic functions. It takes part in the regulation of circulation, thermoregulation, and digestion. All liver functions are disturbed during harvesting, preservation, and transplantation. The assessment of graft function is still a matter of concern. Many methods including analysis of hepatic enzymes, ketone body ratio, and level of drug metabolites do not give complete and reliable information on graft function. Liver biopsy is still the best diagnostic tool, however, it is invasive, carrying the risk of complications. High-resolution proton spectroscopy of nuclear magnetic resonance was used for analysis of the bile sample obtained from a patient after orthotopic liver transplantation.

Bastadin 12 and ryanodine reveal similarities between thapsigargin- and tetrabromobisphenol A-induced intracellular Ca(2+) release in cultured cerebellar granule cells.

Tetrabromobisphenol A (TBBPA) is a commonly used brominated flame retardant with recognized neuro- and cytotoxic properties that are presumably mediated by intracellular Ca(2+) release. Other studies have demonstrated that ryanodine is able to inhibit Ca(2+) efflux from skeletal sarcoplasmic reticulum (SR) membranes in response to the known Ca(2+) releaser thapsigargin, provided that the macrocyclic brominated tyrosine derivative bastadin 5 is also present. Similar effects supporting the role of ryanodine receptors in thapsigargin-evoked Ca(2+) release have been observed in primary cultures of rat cerebellar granule cells (CGCs). Here, we used CGCs and the fluorescent intracellular Ca(2+) probe fluo-3 to test the following hypotheses: (1) TBBPA shares Ca(2+) releasing properties with thapsigargin, and (2) synthetic bastadin 12 can replace bastadin 5 as a pharmacological tool to identify these similarities. The results demonstrated that either 200 nM thapsigargin or 30 µM bastadin 12 alone induced an increase in the intracellular Ca(2+) level in CGCs, whereas 2.5 and 10 µM bastadin 12 had no effect on the basal Ca(2+) concentration. The thapsigargin-induced Ca(2+) release was partially reduced by co-administration of either 2.5 µM bastadin 12 or 200 µM ryanodine, and the release of Ca(2+) was nearly completely attenuated by these compounds when they were given together. TBBPA (5, 10 and 25 µM) administration caused a concentration-dependent increase in CGC Ca(2+) levels. Administration of 2.5 µM bastadin 12 with 200 µM ryanodine blocked the increase in intracellular Ca(2+) evoked by 10 µM TBBPA, although these compounds were ineffective when applied separately. These results indicate that bastadin 12 may replace bastadin 5 when testing the ability of ryanodine to inhibit Ca(2+) release from the intracellular stores of cultured neurons, and our findings support the hypothesis that TBBPA and thapsigargin induce intracellular Ca(2+) release through a common mechanism.

Low molecular weight thiols reduce thimerosal neurotoxicity in vitro: modulation by proteins.

Thimerosal (TH), an ethylmercury complex of thiosalicylic acid has been used as preservative in vaccines. In vitro neurotoxicity of TH at high nM concentrations has been reported. Although a number of toxicological experiments demonstrated high affinity of mercury to thiol groups of the extracellular amino acids and proteins that may decrease concentration of free TH in the organism, less is known about the role of interactions between proteins and amino acids in protection against TH neurotoxicity. In the present study we examined whether the presence of serum proteins and of l-cysteine (Cys), d,l-homocysteine (Hcy), N-acetyl cysteine (NAC), l-methionine (Met) and glutathione (GSH) in the incubation medium affects the TH-induced changes in the viability, the intracellular levels of calcium and zinc and mitochondrial membrane potential in primary cultures of rat cerebellar granule cells. The cells were exposed to 500 nM TH for 48 h or to 15-25 µM TH for 10 min. Our results demonstrated a decrease in the cells viability evoked by TH, which could be prevented partially by serum proteins, albumin or in a dose-dependent manner by 60, 120 or 600 µM Cys, Hcy, NAC and GSH, but not by Met. This neuroprotection was less pronounced in the presence of proteins. Incubation of neurons with TH also induced the rise in the intracellular calcium and zinc concentration and decrease in mitochondrial membrane potential, and these effects were abolished by all the sulfur containing compounds studied and administered at 600 µM concentration, except Met. The loss of the ethylmercury moiety from TH as a result of interaction with thiols studied was monitored by (1)H NMR spectroscopy. This extracellular process may be responsible for the neuroprotection seen in the cerebellar cell cultures, but also provides a molecular pathway for redistribution of TH-derived toxic ethylmercury in the organism. In conclusion, these results confirmed that proteins and sulfur-containing amino acids applied separately reduce TH neurotoxicity, while their combination modulates in more complex way neuronal survival in the presence of TH.