Antioxidant properties of MitoTEMPOL and its hydroxylamine

Piperidine nitroxides such as TEMPOL have been widely used as antioxidants in vitro and in vivo. MitoTEMPOL is a mitochondria-targeted derivative of TEMPOL designed to protect mitochondria from the oxidative damage that they accumulate, but once there is rapidly reduced to its hydroxylamine, MitoTEMPOL-H. As little is known about the antioxidant efficacy of hydroxylamines, this study has assessed the antioxidant activity of both MitoTEMPOL and MitoTEMPOL-H. The hydroxylamine was more effective at preventing lipid-peroxidation than MitoTEMPOL and decreased oxidative damage to mitochondrial DNA caused by menadione. In contrast to MitoTEMPOL, MitoTEMPOL-H has no superoxide dismutase activity and its antioxidant actions are likely to be mediated by hydrogen atom donation. Therefore, even though MitoTEMPOL is rapidly reduced to MitoTEMPOL-H in cells, it remains an effective antioxidant. Furthermore, as TEMPOL is also reduced to a hydroxylamine in vivo, many of its antioxidant effects may also be mediated by its hydroxylamine

Patterns of urinary cortisol levels during ontogeny appear population specific rather than species specific in wild chimpanzees and bonobos

Compared with most mammals, postnatal development in great apes is protracted, presenting both an extended period of phenotypic plasticity to environmental conditions and the potential for sustained mother-offspring and/or sibling conflict over resources. Comparisons of cortisol levels during ontogeny can reveal physiological plasticity to species or population specific socioecological factors and in turn how these factors might ameliorate or exaggerate mother-offspring and sibling conflict. Here, we examine developmental patterns of cortisol levels in two wild chimpanzee populations (Budongo and Taï), with two and three communities each, and one wild bonobo population (LuiKotale), with two communities. Both species have similar juvenile life histories. Nonetheless, we predicted that key differences in socioecological factors, such as feeding competition, would lead to interspecific variation in mother-offspring and sibling conflict and thus variation in ontogenetic cortisol patterns. We measured urinary cortisol levels in 1394 samples collected from 37 bonobos and 100 chimpanzees aged up to 12 years. The significant differences in age-related variation in cortisol levels appeared population specific rather than species specific. Both bonobos and Taï chimpanzees had comparatively stable and gradually increasing cortisol levels throughout development; Budongo chimpanzees experienced declining cortisol levels before increases in later ontogeny. These age-related population differences in cortisol patterns were not explained by mother-offspring or sibling conflict specifically; instead, the comparatively stable cortisol patterns of bonobos and Taï chimpanzees likely reflect a consistency in experience of competition and the social environment compared with Budongo chimpanzees, where mothers may adopt more variable strategies related to infanticide risk and resource availability. The clear population-level differences within chimpanzees highlight potential intraspecific flexibility in developmental processes in apes, suggesting the flexibility and diversity in rearing strategies seen in humans may have a deep evolutionary history.Publisher PDFPeer reviewe

Domain formation in DODAB–cholesterol mixed systems monitored via nile red anisotropy

The effect of the cholesterol (Ch) on liposomes composed of the cationic lipid dioctadecyldimethylammonium bromide (DODAB) was assessed by studying both the steady-state and time-resolved fluorescence anisotropy of the dye Nile Red. The information obtained combined with analysis of the steady-state emission and luorescence lifetime of Nile Red (NR) for different cholesterol concentrations (5–50%) elucidated the presence of “condensed complexes” and cholesterol-rich domains in these mixed systems. The steady-state fluorescence spectra were decomposed into the sum of two lognormal emissions, emanating from two different states, and the effect of temperature on the anisotropy decay of Nile Red for different cholesterol concentrations was observed. At room temperature, the time-resolved anisotropy decays are indicative of NR being relatively immobile (manifest by a high r∞ value). At higher temperature, rotational times ca. 1 ns were obtained throughout and a trend in increasing hindrance was seen with increase of Ch content

Protective effect of EDTA preadministration on renal ischemia

BACKGROUND: Chelation therapy with sodium edetate (EDTA) improved renal function and slowed the progression of renal insufficiency in patients subjected to lead intoxication. This study was performed to identify the underlying mechanism of the ability of EDTA treatment to protect kidneys from damage. METHODS: The effects of EDTA administration were studied in a rat model of acute renal failure induced by 60 minutes ischemia followed or not by 60 minutes reperfusion. Renal ischemic damage was evaluated by histological studies and by functional studies, namely serum creatinine and blood urea nitrogen levels. Treatment with EDTA was performed 30 minutes before the induction of ischemia. Polymorphonuclear cell (PMN) adhesion capability, plasmatic nitric oxide (NO) levels and endothelial NO synthase (eNOS) renal expression were studied as well as the EDTA protection from the TNFα-induced vascular leakage in the kidneys. Data was compared by two-way analysis of variance followed by a post hoc test. RESULTS: EDTA administration resulted in the preservation of both functional and histological parameters of rat kidneys. PMN obtained from peripheral blood of EDTA-treated ischemized rats, displayed a significant reduction in the expression of the adhesion molecule Mac-1 with respect to controls. NO was significantly increased by EDTA administration and eNOS expression was higher and more diffuse in kidneys of rats treated with EDTA than in the controls. Finally, EDTA administration was able to prevent in vivo the TNFα-induced vascular leakage in the kidneys. CONCLUSION: This data provides evidence that EDTA treatment is able to protect rat kidneys from ischemic damage possibly through the stimulation of NO production

Fabrication Principles and Their Contribution to the Superior In Vivo Therapeutic Efficacy of Nano-Liposomes Remote Loaded with Glucocorticoids

We report here the design, development and performance of a novel formulation of liposome- encapsulated glucocorticoids (GCs). A highly efficient (>90%) and stable GC encapsulation was obtained based on a transmembrane calcium acetate gradient driving the active accumulation of an amphipathic weak acid GC pro-drug into the intraliposome aqueous compartment, where it forms a GC-calcium precipitate. We demonstrate fabrication principles that derive from the physicochemical properties of the GC and the liposomal lipids, which play a crucial role in GC release rate and kinetics. These principles allow fabrication of formulations that exhibit either a fast, second-order (t1/2 ∼1 h), or a slow, zero-order release rate (t1/2 ∼ 50 h) kinetics. A high therapeutic efficacy was found in murine models of experimental autoimmune encephalomyelitis (EAE) and hematological malignancies

Celastrol targets mitochondrial respiratory chain complex I to induce reactive oxygen species-dependent cytotoxicity in tumor cells

<p>Abstract</p> <p>Background</p> <p>Celastrol is an active ingredient of the traditional Chinese medicinal plant <it>Tripterygium Wilfordii</it>, which exhibits significant antitumor activity in different cancer models <it>in vitro </it>and <it>in vivo</it>; however, the lack of information on the target and mechanism of action of this compound have impeded its clinical application. In this study, we sought to determine the mode of action of celastrol by focusing on the processes that mediate its anticancer activity.</p> <p>Methods</p> <p>The downregulation of heat shock protein 90 (HSP90) client proteins, phosphorylation of c-Jun NH2-terminal kinase (JNK), and cleavage of PARP, caspase 9 and caspase 3 were detected by western blotting. The accumulation of reactive oxygen species (ROS) was analyzed by flow cytometry and fluorescence microscopy. Cell cycle progression, mitochondrial membrane potential (MMP) and apoptosis were determined by flow cytometry. Absorption spectroscopy was used to determine the activity of mitochondrial respiratory chain (MRC) complexes.</p> <p>Results</p> <p>Celastrol induced ROS accumulation, G2-M phase blockage, apoptosis and necrosis in H1299 and HepG2 cells in a dose-dependent manner. N-acetylcysteine (NAC), an antioxidative agent, inhibited celastrol-induced ROS accumulation and cytotoxicity. JNK phosphorylation induced by celastrol was suppressed by NAC and JNK inhibitor SP600125 (SP). Moreover, SP significantly inhibited celastrol-induced loss of MMP, cleavage of PARP, caspase 9 and caspase 3, mitochondrial translocation of Bad, cytoplasmic release of cytochrome c, and cell death. However, SP did not inhibit celastrol-induced ROS accumulation. Celastrol downregulated HSP90 client proteins but did not disrupt the interaction between HSP90 and cdc37. NAC completely inhibited celastrol-induced decrease of HSP90 client proteins, catalase and thioredoxin. The activity of MRC complex I was completely inhibited in H1299 cells treated with 6 μM celastrol in the absence and presence of NAC. Moreover, the inhibition of MRC complex I activity preceded ROS accumulation in H1299 cells after celastrol treatment.</p> <p>Conclusion</p> <p>We identified ROS as the key intermediate for celastrol-induced cytotoxicity. JNK was activated by celastrol-induced ROS accumulation and then initiated mitochondrial-mediated apoptosis. Celastrol induced the downregulation of HSP90 client proteins through ROS accumulation and facilitated ROS accumulation by inhibiting MRC complex I activity. These results identify a novel target for celastrol-induced anticancer activity and define its mode of action.</p

Shared community effects and the non-genetic maternal environment shape cortisol levels in wild chimpanzees

Mechanisms of inheritance remain poorly defined for many fitness-mediating traits, especially in long-lived animals with protracted development. Using 6,123 urinary samples from 170 wild chimpanzees, we examined the contributions of genetics, non-genetic maternal effects, and shared community effects on variation in cortisol levels, an established predictor of survival in long-lived primates. Despite evidence for consistent individual variation in cortisol levels across years, between-group effects were more influential and made an overwhelming contribution to variation in this trait. Focusing on within-group variation, non-genetic maternal effects accounted for 8% of the individual differences in average cortisol levels, significantly more than that attributable to genetic factors, which was indistinguishable from zero. These maternal effects are consistent with a primary role of a shared environment in shaping physiology. For chimpanzees, and perhaps other species with long life histories, community and maternal effects appear more relevant than genetic inheritance in shaping key physiological traits.Additional co-authors: Klaus Zuberbuehler, Linda Vigilant, Tobias Deschner, Roman M. Wittig & Catherine Crockfor

Mesenchymal Stromal Cells Improve Salivary Function and Reduce Lymphocytic Infiltrates in Mice with Sjögren's-Like Disease

Non-obese diabetic (NOD) mice develop Sjögren's-like disease (SS-like) with loss of saliva flow and increased lymphocytic infiltrates in salivary glands (SGs). There are recent reports using multipotent mesenchymal stromal cells (MSCs) as a therapeutic strategy for autoimmune diseases due to their anti-inflammatory and immunomodulatory capabilities. This paper proposed a combined immuno- and cell-based therapy consisting of: A) an injection of complete Freund's adjuvant (CFA) to eradicate autoreactive T lymphocytes, and B) transplantations of MSCs to reselect lymphocytes. The objective of this was to test the effectiveness of CD45(-)/TER119(-) cells (MSCs) in re-establishing salivary function and in reducing the number of lymphocytic infiltrates (foci) in SGs. The second objective was to study if the mechanisms underlying a decrease in inflammation (focus score) was due to CFA, MSCs, or CFA+MSCs combined.Donor MSCs were isolated from bones of male transgenic eGFP mice. Eight week-old female NOD mice received one of the following treatments: insulin, CFA, MSC, or CFA+MSC (combined therapy). Mice were followed for 14 weeks post-therapy. CD45(-)/TER119(-) cells demonstrated characteristics of MSCs as they were positive for Sca-1, CD106, CD105, CD73, CD29, CD44, negative for CD45, TER119, CD11b, had high number of CFU-F, and differentiated into osteocytes, chondrocytes and adipocytes. Both MSC and MSC+CFA groups prevented loss of saliva flow and reduced lymphocytic infiltrations in SGs. Moreover, the influx of T and B cells decreased in all foci in MSC and MSC+CFA groups, while the frequency of Foxp3(+) (T(reg)) cell was increased. MSC-therapy alone reduced inflammation (TNF-α, TGF-β), but the combination of MSC+CFA reduced inflammation and increased the regenerative potential of SGs (FGF-2, EGF).The combined use of MSC+CFA was effective in both preventing saliva secretion loss and reducing lymphocytic influx in salivary glands

Two-Photon Fluorescence Microscopy Imaging of Cellular Oxidative Stress Using Profluorescent Nitroxides

A range of varying chromophore nitroxide free radicals and their nonradical methoxyamine analogues were synthesized and their linear photophysical properties examined. The presence of the proximate free radical masks the chromophore’s usual fluorescence emission, and these species are described as profluorescent. Two nitroxides incorporating anthracene and fluorescein chromophores (compounds 7 and 19, respectively) exhibited two-photon absorption (2PA) cross sections of approximately 400 G.M. when excited at wavelengths greater than 800 nm. Both of these profluorescent nitroxides demonstrated low cytotoxicity toward Chinese hamster ovary (CHO) cells. Imaging colocalization experiments with the commercially available CellROX Deep Red oxidative stress monitor demonstrated good cellular uptake of the nitroxide probes. Sensitivity of the nitroxide probes to H2O2-induced damage was also demonstrated by both one- and two-photon fluorescence microscopy. These profluorescent nitroxide probes are potentially powerful tools for imaging oxidative stress in biological systems, and they essentially “light up” in the presence of certain species generated from oxidative stress. The high ratio of the fluorescence quantum yield between the profluorescent nitroxide species and their nonradical adducts provides the sensitivity required for measuring a range of cellular redox environments. Furthermore, their reasonable 2PA cross sections provide for the option of using two-photon fluorescence microscopy, which circumvents commonly encountered disadvantages associated with one-photon imaging such as photobleaching and poor tissue penetration