Cannabinoid receptor Type 1 densities reflect social organization in Microtus.

Across many species, endocannabinoids play an important role in regulating social play, reward, and anxiety. These processes are mediated through at least two distinct cannabinoid receptors (CB), CB1 and CB2. CB1 expression is found in appreciable densities across regions of the brain that integrate memory with socio-spatial information; many of these regions have been directly linked to the neurobiology of pair bonding in monogamous species. Using receptor autoradiography, we provide the first distributional map of CB1 within the brains of closely related monogamous prairie voles and promiscuous meadow voles, and compare receptor densities across sexes and species in limbic regions. We observe CB1-specific signal using [3H] CP-55,940 and [3H] SR141716A, though the latter exhibited a lower signal to noise ratio. We confirmed the presence of CB2 in prairie vole spleen tissue using [3H] CP-55,940. However, we found no evidence of CB2 in the brain using either [3H] CP-55,940 or [3H] A-836,339. The overall distribution of putative CB1 in the brain was similar across vole species and followed the pattern of CB1 expression observed in other species-high intensity binding within the telencephalon, moderate binding within the diencephalon, and mild binding within the mesencephalon and metencephalon (aside from the cerebellar cortex). However, we found profound differences in CB1 densities across species, with prairie voles having higher CB1 binding in regions implicated in social attachment and spatial memory (e.g., periaqueductal gray, hippocampus). These findings suggest that CB1 densities, but not distribution, correlate with the social systems of vole species.

Accumulation of mineral oil saturated hydrocarbons (MOSH) in female Fischer 344 rats: Comparison with human data and consequences for risk assessment.

Female Fischer 344 rats were orally exposed to a mixture of mineral oil saturated hydrocarbons (MOSH) of broad molecular mass range at doses of 40, 400 and 4000mg/kg feed. Amounts and compositions of the MOSH were analyzed in liver, spleen, adipose tissue and the carcass after exposure during 30, 60, 90 and 120d as well as after 90d exposure followed by 30d depuration. At 40mg/kg in the feed, after 30d of exposure, 10.9% of the ingested MOSH were recovered from the animal body; after 90d plus 30d depuration it was 3.9%. In liver and spleen, the maximum retention in terms of molecular mass (simulated distillation) was at n-C29; in adipose tissue and carcass it was at n-C15/16. The differentiation between MOSH below and above n-C25 (Class I versus Class II and III oils), used for present regulation, is not supported by the present data on accumulation; structural characteristics seem more pertinent than molecular mass. Concentrations in the tissues increased far less than proportionally with the dose, rendering linear extrapolation to low doses questionable. No steady state was reached after 120d. In fact, comparing with the concentrations in human tissues at the estimated exposure, extrapolation from animal experiments seems to grossly underestimate human internal exposure. Comprehensive two-dimensional gas chromatography (GCxGC) was used to characterize the MOSH residues in the tissues with the aim of identifying the most strongly accumulated types. In the liver and spleen, the highly branched hydrocarbons dominated, whereas in the adipose tissue it was the n-alkanes and species with main n-alkyl moieties. Strong MOSH accumulation is not of concern per se, but the safety at the high concentrations in human tissues needs to be re-evaluated, possibly taking into account also end points other than granuloma formation.

In vivo biodistribution and toxicity assessment of triplet-triplet annihilation-based upconversion nanocapsules.

Triplet-triplet annihilation (TTA)-based upconversion nanocapsules (UCNCs) have great potential in biological and medical applications. However, there are numerous unresolved issues with respect to the safety of these novel nanomaterials. In this work, for the first time, we studied the in vivo biodistribution of UCNCs which were synthesized by co-loading platinum (II)-tetraphenyl-tetrabenzoporphyrin (PtTPBP) and boron dipyrromethene derivative (BDP) into bovine serum albumin (BSA)-stabilized soybean oil droplets, and systematically assessed the potential toxicity of UCNCs both in vitro and in vivo. The results showed that UCNCs had no significant influence on the proliferation or the migration of HeLa cells even when the dosage was increased to 12 mg/mL. The biodistribution results demonstrated that UCNCs mainly accumulated in the mononuclear phagocyte system (MPS) including the liver and spleen after intravenous injection of the nanocapsules. When mice were intravenously injected with 1200 mg/kg of the UCNCs over a period of 60 days, no noticeable toxicity was observed under these treatment conditions as shown by body weight results, histological analyses, hematological analyses and blood biochemical examinations. This research inspires further studies on UCNCs for biomedical applications.

Ultrapure laser-synthesized Si-based nanomaterials for biomedical applications: in vivo assessment of safety and biodistribution.

Si/SiOx nanoparticles (NPs) produced by laser ablation in deionized water or aqueous biocompatible solutions present a novel extremely promising object for biomedical applications, but the interaction of these NPs with biological systems has not yet been systematically examined. Here, we present the first comprehensive study of biodistribution, biodegradability and toxicity of laser-synthesized Si-SiOx nanoparticles using a small animal model. Despite a relatively high dose of Si-NPs (20 mg/kg) administered intravenously in mice, all controlled parameters (serum, enzymatic, histological etc.) were found to be within safe limits 3 h, 24 h, 48 h and 7 days after the administration. We also determined that the nanoparticles are rapidly sequestered by the liver and spleen, then further biodegraded and directly eliminated in urine without any toxicity effects. Finally, we found that intracellular accumulation of Si-NPs does not induce any oxidative stress damage. Our results evidence a huge potential in using these safe and biodegradable NPs in biomedical applications, in particular as vectors, contrast agents and sensitizers in cancer therapy and diagnostics (theranostics).

Non-invasive assessment of tissue iron overload.

In recent years, there has been increasing interest in non-invasive iron measurement, especially of the liver and heart, in patients with iron overload. Serum ferritin still remains an essential monitoring parameter in intervals between liver iron measurements; however, confounding factors such as inflammation, chelation treatment changes and the specific disease have to be taken into account. Liver iron measurements can now routinely be performed in clinical applications either by quantitative magnetic resonance imaging (MRI) using the transverse magnetic relaxation rate R(2) or R(2)* (1/T(2)*) or by biomagnetic liver susceptometry. For iron measurements in the heart, the single-breathhold multi-echo MRI-R(2)* method has become a standard modality and is now applied in clinical settings beyond research studies. In other tissues like the pancreas, pituitary, and brain, different MRI methods are employed, but their clinical benefit has yet to be proven.

Assessment of the relationship between cerebral and splanchnic oxygen saturations measured by near-infrared spectroscopy and direct measurements of systemic haemodynamic variables and oxygen transport after the Norwood procedure.

OBJECTIVES: To evaluate the clinical utility of near-infrared spectroscopic (NIRS) monitoring of cerebral (ScO2) and splanchnic (SsO2) oxygen saturations for estimation of systemic oxygen transport after the Norwood procedure. METHODS: ScO2 and SsO2 were measured with NIRS cerebral and thoracolumbar probes (in humans). Respiratory mass spectrometry was used to measure systemic oxygen consumption (O2). Arterial (SaO2), superior vena caval (SvO2) and pulmonary venous oxygen saturations were measured at 2 to 4 h intervals to derive pulmonary (Qp) and systemic blood flow (Qs), systemic oxygen delivery (DO2) and oxygen extraction ratio (ERO2). Mixed linear regression was used to test correlations. A study of 7 pigs after cardiopulmonary bypass (study 1) was followed by a study of 11 children after the Norwood procedure (study 2). RESULTS: Study 1. ScO2 moderately correlated with SvO2, mean arterial pressure, Qs, DO2 and ERO2 (slope 0.30, 0.64. 2.30, 0.017 and -32.5, p < 0.0001) but not with SaO2, arterial oxygen pressure (PaO2), haemoglobin and O2. Study 2. ScO2 correlated well with SvO2, SaO2, PaO2 and mean arterial pressure (slope 0.43, 0.61, 0.99 and 0.52, p < 0.0001) but not with haemoglobin (slope 0.24, p > 0.05). ScO2 correlated weakly with O2 (slope -0.07, p = 0.05) and moderately with Qs, DO2 and ERO2 (slope 3.2, 0.03, -33.2, p < 0.0001). SsO2 showed similar but weaker correlations. CONCLUSIONS: ScO2 and SsO2 may reflect the influence of haemodynamic variables and oxygen transport after the Norwood procedure. However, the interpretation of NIRS data, in terms of both absolute values and trends, is difficult to rely on clinically.

An assessment of extraction and assay techniques for quantification of calpain and calpastatin from small tissue samples.

Our objective was to evaluate whether small (biopsy-sized) samples could be used to measure calpain and calpastatin activities in skeletal muscle. The accuracy of different separation and assay methods for the quantification of calpains and calpastatin from small (1.0 and 0.2 g) skeletal muscle samples was tested. In Exp. 1, the LM was removed from six lambs, and a 50-g subsample was processed using the reference method (DEAE-Sephacel chromatography and casein assay). Subsamples (1.0 and 0.2 g) also were processed using the two-step separation (1 mL DEAE-Sephacel and bulk elution using 200 and 400 mM NaCl) and heated calpastatin methods; in both cases, fractions were assayed with Bodipy-labeled and [14C]-labeled casein microassays. Finally, casein zymography was used to separate and quantify the calpain proteases from 1.0-and 0.2-g samples. The values obtained after processing the 50-g sample using the reference method were judged most accurate, and the alternative approaches were compared with these. For each extraction and assay approach, we considered: 1) the effect of the sample size on the mean activity; 2) increased or decreased variation of data; and 3) the correlation relative to the reference method. Where possible, we compared the ratio of calpain to calpastatin activities determined using the alternative approaches with the ratios found using the reference method. These methodologies were further investigated in Exp. 2, where single homogenates from different tissues (heart, spleen, lung, and muscle) were assayed using the alternative approaches. Experiment 1 established that most of the approaches suffered from poor correlations and/or unacceptable variation. By using a large, homogenous sample in Exp. 2, however, we determined that this error was not due to the methodologies themselves. Therefore, the unacceptable variation found in Exp. 1 resulted from the small sample size, and we recommend that large tissue samples (e.g., 50 g) should be used for calpain and calpastatin activity measurements in skeletal muscle instead of small tissue biopsies (e.g., 0.2 and 1.0 g).

Assessment of a systematic expression profiling approach in ENU-induced mouse mutant lines.

Comparative genomewide expression profiling is a powerful tool in the effort to annotate the mouse genome with biological function. The systematic analysis of RNA expression data of mouse lines from the Munich ENU mutagenesis screen might support the understanding of the molecular biology of such mutants and provide new insights into mammalian gene function. In a direct comparison of DNA microarray experiments of individual versus pooled RNA samples of organs from ENU-induced mouse mutants, we provide evidence that individual RNA samples may outperform pools in some aspects. Genes with high biological variability in their expression levels (noisy genes) are identified as false positives in pooled samples. Evidence suggests that highly stringent housing conditions and standardized procedures for the isolation of organs significantly reduce biological variability in gene expression profiling experiments. Data on wild-type individuals demonstrate the positive effect of controlling variables such as social status, food intake before organ sampling, and stress with regard to reproducibility of gene expression patterns. Analyses of several organs from various ENU-induced mutant lines in general show low numbers of differentially expressed genes. We demonstrate the feasibility to detect transcriptionally affected organs employing RNA expression profiling as a tool for molecular phenotyping.

Assessment of iron stores in children with transfusion siderosis by biomagnetic liver susceptometry.

To investigate the applicability of noninvasive Superconducting Quantum Interference Device (SQUID) biomagnetic liver susceptometry and its limitations in thalassemic children, 23 patients with beta-thalassemia major and other iron loading anemias (age: 4-16 years) and 16 age-related normal children were studied. Liver iron concentrations ranged from 600 to 11,000 microg/g(liver) for thalassemic patients and from 60 to 340 microg/g(liver) for normal patients. Measuring the respective organ volumes by sonography, liver and spleen iron stores, accounting for 80% of total body iron stores, were estimated. Nonliver contributions from the lung or intestine to the measured SQUID signals in the small-sized patients were not observed. Moreover, livers in thalassemia were found to be enlarged by 18% per 1,000 microg/g (r = 0.75, P < 10(-3)). Serum ferritin values correlate significantly with iron stores (r = 0.64, P < 10(-3)), but predict iron stores only within large error intervals of 4,000 microg/g(liver). Analyzing the experimental data from biomagnetometry and from related transfusion and chelation treatment data within the framework of a two-compartment model, we were able to derive additional information on total body iron elimination and chelation therapy efficacy. The exponential decline of iron stores allows forecast of steady-state conditions of the final iron load for a particular transfusion and chelation therapy regimen.

Magnetic resonance imaging and assessment of liver iron content in genetic hemochromatosis.

Computed tomography (CT) scanning is not highly sensitive in the assessment of liver iron content and magnetic resonance imaging (MRI) appears to be more efficient. The aim of this study was to determine the effectiveness of MRI in the evaluation of liver iron content using a standard spin-echo technique. The study included 23 patients with genetic hemochromatosis and 24 non-iron-overloaded patients as controls. A comparison was made of: (a) MRI signal intensity of liver, spleen, paravertebral muscles and subcutaneous adipose tissue using two different spin-echo sequences (SE 500/28; SE 2000/28,56); (b) liver attenuation determined by a single energy CT scan; and (c) a biochemical determination of hepatic iron. There was a significant decrease in liver signal intensity in the genetic hemochromatosis group (256 +/- 201, mean +/- S.D.) compared with the control group (801 +/- 413, p less than 0.001), but there was no correlation with liver iron concentration. However, such a correlation was found and was even more highly significant than in CT when the ratio between the liver and another organ was taken into account. For a lower limit of liver/spleen ratio calculated at 0.46 (mean 2 S.D. in the control group), the specificity (0.96) of MRI was satisfactory, but the sensitivity (0.78) remained insufficient (MRI being unable to detect an iron overload of up to 125 mumol/g). Hopefully, these results might be improved in the near future by using more sensitive sequences such as gradient echo sequences.