Altered energy status of primary cerebellar granule neuronal cultures from rats exposed to lead in the pre- and neonatal period.

This paper examines the effect of pre- and neonatal exposure of rats to lead (0.1% lead acetate in drinking water, resulting in rat offspring whole blood lead concentration (Pb-B) 4µg/dL) on the energy status of neuronal mitochondria by measuring changes in ATP, ADP, AMP, adenosine, TAN concentration, adenylate energy charge value (AEC) and mitochondrial membrane potential in primary cerebellar granule neurons (CGC) in dissociated cultures. Fluorescence studies were performed to imaging and evaluate mitochondria mass, mitochondrial membrane potential, intracellular and mitochondrial reactive oxygen species (ROS) production. The Na(+)/K(+) ATPase activity in intact CGC was measured spectrophotometrically. Our data shows that pre- and neonatal exposure of rats to Pb, even below the threshold of whole blood Pb value considered safe for people, affects the energy status of cultured primary cerebellar granule neurons through a decrease in ATP and TAN concentrations and AEC value, inhibition of Na(+)/K(+) ATPase, and increase in intracellular and mitochondrial ROS concentration. These observations suggest that even these low levels of Pb are likely to induce important alterations in neuronal function that could play a role in neurodegeneration.

Blood arachidonic acid and HDL cholesterol influence the phagocytic abilities of human monocytes/macrophages.

AIMS: Many immunomodulators may intensify the process of phagocytosis in monocytes. Among them are the fatty acids contained in cellular membrane phospholipids. But in the available literature there are no reports on how blood fatty acids and lipoproteins can modulate the phagocytic abilities of cells. Therefore, the aim of this study was the evaluation of the phagocytic activity of monocytes isolated from the blood of healthy human subjects with defined fatty acids and lipoprotein content. METHODS: Cells obtained from 24 donors were used for measuring phagocytic activity and for the quantification of serum fatty acids, total cholesterol, TG, HDL, and LDL, respectively. Phagocytosis was determined using a PHAGOTEST kit, fatty acids using a GC chromatograph, and lipids using test kits. RESULTS/CONCLUSIONS: The analysis shows an influence of serum HDL concentrations on the process of phagocytosis in the isolated cells and suggests that the concentration of arachidonic acid in blood is a factor that determines the phagocytic ability of monocytes. Moreover, the concentration of conjugated linoleic acid trans-10 cis-12 has considerable influence on phagocytosis.

Fluoride as a pro-inflammatory factor and inhibitor of ATP bioavailability in differentiated human THP1 monocytic cells.

Chronic exposure of humans to fluorine compounds in the air, water and food may be atherogenic via the activation of oxidative stress and increased ROS production. The most important factor that promotes the formation of ROS seems to be the oxidoreduction of electron carriers in the critical points of the respiratory chain, which depends, among other things, on the cellular demand for ATP. This paper examines the effect of fluorides in concentrations determined in human serum on the intracellular synthesis of ROS, the activity of the respiratory chain enzymes and the synthesis of ATP via oxidative and substrate-level phosphorylation. The incubation of macrophages in fluoride solutions significantly decreased the amount of synthesized cellular ATP and increased formation of ROS and apoptosis in a dose-dependent pattern. The addition of respiratory chain inhibitors resulted in a significant decrease in the synthesized ROS. Sodium fluoride probably promotes oxidative stress in macrophages, which is manifested by a strong increase in ROS synthesis and a decrease in ATP. We suppose that fluoride may destabilize the action of respiratory chain. Our results indicate that the respiratory chain is the main site of ROS synthesis. One cannot exclude the stimulating role of fluorine compounds on the formation of ROS that is independent of the respiratory chain.

Short-term storage of human haematopoietic cells. Influence of air and deoxyribonuclease I.

The aim of this study was to optimize strategy for ex vivo short-term storage of human cord blood and bone marrow haematopoietic cells. We report that the presence of air in the vials (1/2 of their volume), in which hematopoietic cells are stored, improves the survival of clonogenic progenitors. We observed that presence of air prevented a rapid decrease in the pH of storage medium. Similarly, a beneficial effect on cell survival and recovery also had an addition of Deoxyribonuclease I (DNase I). We observed that DNase I efficiently prevented cell clumping, and moreover, did not affect the clonogenecity of the haematopoietic progenitors. Therefore containers, in which haematopoietic cells are stored, should contain enough air (source of oxygen) and the storage medium itself should be supplemented with DNase I.

In vivo and in vitro studies on the toxicity of Hoechst 33342 (Ho342). Implications for employing Ho342 for the isolation of haematopoietic stem cells.

The fluorescence dye Hoechst 33342 (Ho342) is employed for isolating early haematopoietic cells and the aim of this study was to evaluate the in vivo and in vitro toxicity of this compound. First, by employing a murine model we studied the influence of this dye on the morphology of the different organs of animals that have been injected intravenously with increasing doses of Ho342. Accordingly, we found that Ho342 at relatively low doses (0,3 M) caused morphological changes in the spleen and lungs and at higher doses (1,5 & 6 M) damaged also the liver. In contrast, kidneys appear to be relatively resistant to this dye. Next, since Ho342 is employed for isolating early haematopoietic cells by FACS, we have been looking for potential toxicity of this dye against normal human haematopoietic progenitors. Accordingly, CD34+ cells isolated from cord blood (CB) samples were exposed to increasing doses of Ho342 (0-50 microM). We found that the low concentration of Ho342 (10 microM) recommended for isolating HSC significantly inhibited the clonogenecity of human erythroid progenitors (BFU-E). The higher doses of Ho342 have also been toxic against normal human myeloid progenitors (CFU-GM). This study shows that Ho342 could potentially damage human cells. This fact should be considered whenever Ho342 has to be employed for isolating living cells.