Histochemical changes in neonatal liver caused by vaginal instillation of magnetic nanoparticles in pregnant mice

Drug delivery through the vagina is a novel and effective approach for treating embryonic diseases. Magnetic nanoparticles (MNPs) currently are used as drug delivery systems. The safety of MNPs for use with embryonic tissues remains unclear. We used pregnant mice to investigate the possible toxicity of MNPs toward neonatal liver at three embryonic ages using histochemical and immunohistochemical techniques. MNPs were instilled through the vaginas of pregnant mice at days 12 (E12), 15 (E15) and 17 (E17) after fertilization. We found MNPs in the neonatal liver parenchyma after delivery of the pups on day 20. We observed that MNPs caused mild apoptosis of hepatocytes, cytoplasmic vacuolation and lymphocytic infiltration in the neonatal liver after treatment at E15 compared to instillation at E12 and E17. We observed also that MNPs increased the production of caspase proteins and tumor necrosis factor receptor 2 proteins, which are indicators of apoptosis, in the neonatal liver after instillation of MNPs at E15 compared to instillation at E12 and E17. MNPs also increased the number of collagen fibers and the amounts of connective tissue growth factors in the neonatal liver parenchyma after instillation at E15 compared to instillation at E12 and E17. The general carbohydrates in the neonatal liver were decreased in a time-dependent manner after instillation at E17, E15 and E12 owing to the presence of MNPs in the parenchyma. Overall, we determined that MNPs were mildly toxic to neonatal liver.     

Chronic stress and its effects on adrenal cortex apoptosis in pregnant rats

The model of chronic intermittent stress by immobilization during pregnancy may produce alterations in the mechanisms that maintain adrenal gland homeostasis. In earlier investigations using this model, significant variations in plasma prolactin and corticosterone levels, and adrenal gland weights were observed. We hypothesized that chronic stress causes changes in apoptosis in the adrenal glands of pregnant rats. We identified and quantified apoptotic cells in the adrenal cortex and examined their ultrastructural characteristics using transmission electron microscopy. Adrenal glands of pregnant rats at gestation days 12, 17 and 21 were studied for control and experimental (stressed) rats. Immunolabelling techniques, stereological analysis and image quantification of adrenal gland sections were combined to determine differences in apoptosis in the different cell populations of the adrenal cortex. The apoptotic index of the experimental rats showed a significant reduction at gestation day 17, while at days 12 and 21 there were no differences from controls. Moreover, the apoptotic index of the reticular zones in control and experimental animals showed a significant increase compared to the glomerular and fascicular zones at the three gestation times studied. Chronic stress by immobilization reduced the caspase-dependent apoptotic index at gestation day 17, which may be related to variations in plasma concentrations of estrogens and prolactin.     

Identifying the neural circuitry of alcohol craving and relapse vulnerability

With no further intervention, relapse rates in detoxified alcoholics are high and usually exceed 80% of all detoxified patients. It has been suggested that stress and exposure to priming doses of alcohol and to alcohol-associated stimuli (cues) contribute to the relapse risk after detoxification. This article focuses on neuronal correlates of cue responses in detoxified alcoholics. Current brain imaging studies indicate that dysfunction of dopaminergic, glutamatergic and opioidergic neurotransmission in the brain reward system (ventral striatum including the nucleus accumbens) can be associated with alcohol craving and functional brain activation in neuronal systems that process attentional relevant stimuli, reward expectancy and experience. Increased functional brain activation elicited by such alcohol-associated cues predicted an increased relapse risk, whereas high brain activity elicited by affectively positive stimuli may represent a protective factor and was correlated with a decreased prospective relapse risk. These findings are discussed with respect to psychotherapeutic and pharmacological treatment options.     

Direct exchange of vitamin B12 is demonstrated by modelling the growth dynamics of algal–bacterial cocultures

The growth dynamics of populations of interacting species in the aquatic environment is of great importance, both for understanding natural ecosystems and in efforts to cultivate these organisms for industrial purposes. Here we consider a simple two-species system wherein the bacterium Mesorhizobium loti supplies vitamin B₁₂ (cobalamin) to the freshwater green alga Lobomonas rostrata, which requires this organic micronutrient for growth. In return, the bacterium receives photosynthate from the alga. Mathematical models are developed that describe minimally the interdependence between the two organisms, and that fit the experimental observations of the consortium. These models enable us to distinguish between different mechanisms of nutrient exchange between the organisms, and provide strong evidence that, rather than undergoing simple lysis and release of nutrients into the medium, M. loti regulates the levels of cobalamin it produces, resulting in a true mutualism with L. rostrata. Over half of all microalgae are dependent on an exogenous source of cobalamin for growth, and this vitamin is synthesised only by bacteria; it is very likely that similar symbiotic interactions underpin algal productivity more generally.     

Selective permeabilization of lipid membranes by photodynamic action via formation of hydrophobic defects or pre-pores

To gain insight into mechanisms of photodynamic modification of biological membranes, we studied an impact of visible light in combination with a photosensitizer on translocation of various substances across artificial (vesicular and planar) bilayer lipid membranes (BLMs). Along with induction of carboxyfluorescein leakage from liposomes, pronounced stimulation of lipid flip-flop between the two monolayers was found after photosensitization, both processes being prevented by the singlet oxygen quencher sodium azide. On the contrary, no enhancement of potassium chloride efflux from liposomes was detected by conductometry under these conditions. Illumination of planar BLMs in the presence of a photosensitizer led to a marked increase in membrane permeability to amphiphilic 2-n-octylmalonic acid, but practically no change in the permeability to ammonia, which agreed with selective character of the photosensitized leakage of fluorescent dyes from liposomes (Pashkovskaya et al., Langmuir, 2010). Thus, the effect on transbilayer movement of molecules elicited by the photodynamic treatment substantially depended on the kind of translocated species, in particular, on their lipophilicity. Based on similarity with results of previous electroporation studies, we hypothesized about photodynamic induction of "pre-pores" or "hydrophobic defects" permeable to amphiphilic compounds and less permeable to hydrophilic substances and inorganic ions.     

In vivo fluxes in the ammonium-assimilatory pathways in corynebacterium glutamicum studied by 15N nuclear magnetic resonance

Glutamate dehydrogenase (GDH) and glutamine synthetase (GS)-glutamine 2-oxoglutarate-aminotransferase (GOGAT) represent the two main pathways of ammonium assimilation in Corynebacterium glutamicum. In this study, the ammonium assimilating fluxes in vivo in the wild-type ATCC 13032 strain and its GDH mutant were quantitated in continuous cultures. To do this, the incorporation of 15N label from [15N]ammonium in glutamate and glutamine was monitored with a time resolution of about 10 min with in vivo 15N nuclear magnetic resonance (NMR) used in combination with a recently developed high-cell-density membrane-cyclone NMR bioreactor system. The data were used to tune a standard differential equation model of ammonium assimilation that comprised ammonia transmembrane diffusion, GDH, GS, GOGAT, and glutamine amidotransferases, as well as the anabolic incorporation of glutamate and glutamine into biomass. The results provided a detailed picture of the fluxes involved in ammonium assimilation in the two different C. glutamicum strains in vivo. In both strains, transmembrane equilibration of 100 mM [15N]ammonium took less than 2 min. In the wild type, an unexpectedly high fraction of 28% of the NH4+ was assimilated via the GS reaction in glutamine, while 72% were assimilated by the reversible GDH reaction via glutamate. GOGAT was inactive. The analysis identified glutamine as an important nitrogen donor in amidotransferase reactions. The experimentally determined amount of 28% of nitrogen assimilated via glutamine is close to a theoretical 21% calculated from the high peptidoglycan content of C. glutamicum. In the GDH mutant, glutamate was exclusively synthesized over the GS/GOGAT pathway. Its level was threefold reduced compared to the wild type.     

Adjustment of juvenile tuatara to a cooler,southern climate: operative temperatures,emergence behaviour and growth rate

Translocations for conservation often involve species limited to relict distributions. However, uncertainty can exist regarding the ability of source individuals to acclimatise following a shift to a distant location. We investigated the ability of captive-reared juvenile tuatara (Sphenodon punctatus) of Cook Strait stock (41°S) to adjust to outdoor, predator-protected pens within Orokonui Ecosanctuary (45 °S). We examined potential basking and within burrow temperatures, the influence of temperature on emergence, and growth rates in comparison with other locations. Tuatara at Orokonui reached their preferred temperature when basking over summer, and burrows provided protection from freezing over winter. Emergence was temperature-dependent and essentially ceased during winter. Growth rates of Orokonui-held juveniles were within the range for four other captive-rearing facilities and faster than for wild juveniles from a Cook Strait population. As all Orokonui-held juveniles have survived and grown we conclude that the climate at this southern location is suitable to consider a free-release.     

Rate of progression of CT-quantified emphysema in male current and ex-smokers: a follow-up study

Background

Little is known about the factors associated with CT-quantified emphysema progression in heavy smokers. The objective of this study was to investigate the effect of length of smoking cessation and clinical / demographical factors on the rate of emphysema progression and FEV₁-decline in male heavy smokers.

Methods

3,670 male smokers with mean (SD) 40.8 (17.9) packyears underwent chest CT scans and pulmonary function tests at baseline and after 1 and 3 years follow-up. Smoking status (quitted ≥5, ≥1-<5, <1 years or current smoker) was noted. Rate of progression of emphysema and FEV₁-decline after follow-up were assessed by analysis of variance adjusting for age, height, baseline pulmonary function and emphysema severity, packyears, years in study and respiratory symptoms. The quitted ≥5 group was used as reference.

Results

Median (Q1-Q3) emphysema severity,<-950 HU, was 8.8 (5.1 – 14.1) and mean (SD) FEV₁ was 3.4 (0.73) L or 98.5 (18.5) % of predicted. The group quitted ‘>5 years’ showed significantly lower rates of progression of emphysema compared to current smokers, 1.07% and 1.12% per year, respectively (p<0.001). Current smokers had a yearly FEV₁-decline of 69 ml, while subjects quit smoking >5 years had a yearly decline of 57.5 ml (p<0.001).

Conclusion

Quit smoking >5 years significantly slows the rate of emphysema progression and lung function decline.

Trial registration

Registered at http://www.trialregister.nl with trial number ISRCTN63545820.     

Production of reactive oxygen species in mitochondria of HeLa cells under oxidative stress

Mitochondria can be a source of reactive oxygen species (ROS) and a target of oxidative damage during oxidative stress. In this connection, the effect of photodynamic treatment (PDT) with Mitotracker Red (MR) as a mitochondria-targeted photosensitizer has been studied in HeLa cells. It is shown that MR produces both singlet oxygen and superoxide anion upon photoactivation and causes photoinactivation of gramicidin channels in a model system (planar lipid bilayer). Mitochondria-targeted antioxidant (MitoQ) inhibits this effect. In living cells, MR-mediated PDT initiates a delayed ("dark") accumulation of ROS, which is accelerated by inhibitors of the respiratory chain (piericidin, rotenone and myxothiazol) and inhibited by MitoQ and diphenyleneiodonium (an inhibitor of flavin enzymes), indicating that flavin of Complex I is involved in the ROS production. PDT causes necrosis that is prevented by MitoQ. Treatment of the cell with hydrogen peroxide causes accumulation of ROS, and the effects of inhibitors and MitoQ are similar to that described for the PDT model. Apoptosis caused by H2O2 is augmented by the inhibitors of respiration and suppressed by MitoQ. It is concluded that the initial segments of the respiratory chain can be an important source of ROS, which are targeted to mitochondria, determining the fate of the cell subjected to oxidative stress.