Alpha-tomatine induces apoptosis and inhibits nuclear factor-kappa B activation on human prostatic adenocarcinoma PC-3 cells

Background

Alpha-tomatine (α-tomatine) is the major saponin in tomato (Lycopersicon esculentum). This study investigates the chemopreventive potential of α-tomatine on androgen-independent human prostatic adenocarcinoma PC-3 cells.

Methodology/Principal Findings

Treatment of highly aggressive human prostate cancer PC-3 cells with α-tomatine resulted in a concentration-dependent inhibition of cell growth with a half-maximal efficient concentration (EC₅₀) value of 1.67±0.3 µM. It is also less cytotoxic to normal human liver WRL-68 cells and normal human prostate RWPE-1 cells. Assessment of real-time growth kinetics by cell impedance-based Real-Time Cell Analyzer (RTCA) showed that α-tomatine exhibited its cytotoxic effects against PC-3 cells as early as an hour after treatment. The inhibitory effect of α-tomatine on PC-3 cancer cell growth was mainly due to induction of apoptosis as evidenced by positive Annexin V staining and decreased in mitochondrial membrane potential but increased in nuclear condensation, polarization of F-actin, cell membrane permeability and cytochrome c expressions. Results also showed that α-tomatine induced activation of caspase-3, -8 and -9, suggesting that both intrinsic and extrinsic apoptosis pathways are involved. Furthermore, nuclear factor-kappa B (NF-κB) nuclear translocation was inhibited, which in turn resulted in significant decreased in NF-κB/p50 and NF-κB/p65 in the nuclear fraction of the treated cells compared to the control untreated cells. These results provide further insights into the molecular mechanism of the anti-proliferative actions of α-tomatine.

Conclusion/Significance

α-tomatine induces apoptosis and inhibits NF-κB activation on prostate cancer cells. These results suggest that α-tomatine may be beneficial for protection against prostate cancer development and progression.     

Heterozygous <Emphasis Type="BoldItalic">tx</Emphasis> mice have an increased sensitivity to copper loading: Implications for Wilson’s disease carriers

Wilson’s disease carriers constitute 1% of the human population. It is unknown whether Wilson’s disease carriers are at increased susceptibility to copper overload when exposed to chronically high levels of ingested copper. This study investigated the effect of chronic excess copper in drinking water on the heterozygous form of the Wilson's disease mouse model – the toxic milk (tx) mouse. Mice were provided with drinking water containing 300 mg/l copper for 4–7, 8–11, 12–15 or 16–20 months. At the completion of the study liver, spleen, kidney and brain tissue were analyzed by atomic absorption spectroscopy to determine copper concentration. Plasma ceruloplasmin oxidase activity and liver histology were also assessed. Chronic copper loading resulted in significantly increased liver copper in both tx heterozygous and tx homozygous mice, while wild type mice were resistant to the effects of copper loading. Copper loading effects were greatest in tx homozygous mice, with increased extrahepatic copper deposition in spleen and kidney – an effect absent in heterozygote and wild type mice. Although liver histology in homozygous mice was markedly abnormal, no histological differences were noted between heterozygous and wild type mice with copper loading. Tx heterozygous mice have a reduced ability to excrete excess copper, indicating that half of the normal liver Atp7b copper transporter activity is insufficient to deal with large copper intakes. Our results suggest that Wilson’s disease carriers in the human population may be at increased risk of copper loading if chronically exposed to elevated copper in food or drinking water.     

The electron transport systems of Fasciola hepatica mitochondria.

The electron transport systems of Fasciola hepatica mitochondria were investigated spectrophotometrically at room temperature and at −196°. The mitochondria were found to contain substrate reducible a-, b- and c-type cytochromes. All of the cytochrome components of the classical mammalian type of respiratory chain were present, although the concentration of cytochromes aa₃ was low. In addition to the mammalian type of respiratory chain, the Fasciola mitochondria contained a substrate reducible b-type cytochrome component (557 nm) which included a CO reactive o-type cytochrome. The results suggest that F. hepatica mitochondria contain a branched electron transport system including a mammalian type of chain and involving two terminal oxidases and at least two b-type cytochromes.     

Specific receptor sites for 1-O-alkyl-2-O-acetyl-sn-glycero-3-phosphocholine (platelet activating factor) on rabbit platelet and guinea pig smooth muscle membranes

By using tritiated 1-O-alkyl-2-O-acetyl-sn-glycero-3-phosphocholine (3H-PAF), we have directly identified its specific binding sites on rabbit platelet plasma membranes. The equilibrium dissociation constant for 3H-PAF is 1.36 (+/- 0.05) X 10(-9) M at 0 degrees C. The number of binding sites is 1.61 (+/- 0.34) X 10(12)/mg of membrane, which corresponds to approximately 150-300 receptors/platelet (depending on membrane vesicle orientation). Binding of 3H-PAF to rabbit platelet plasma membrane is rapid (t1/2 less than 5 min at 0 degrees C) and reversible. For a series of PAF analogues, their affinity for the receptor sites parallels with their relative potency to induce platelet aggregation. PAF can cause contraction of smooth muscle of heart, parenchymal strip, trachea, and ileum. Specific PAF receptor binding was demonstrated with purified plasma membrane from several smooth muscles and from polymorphonuclear leukocytes but not from presumably PAF nonresponsive cells such as erythrocytes and alveolar macrophages. It is likely that the interaction of PAF with these binding sites initiates the specific responses of platelets, polymorphonuclear leukocytes, and smooth muscles.     

Paracrystalline arrays in mitochondria following ageing of mitochondriain situ

Paracrystalline arrays of helical configuration were observed in the mitochondrial intracrystal spaces following prolonged ageing of mitochondriain situ. The occurrence of these mitochondria with the paracrystalline arrays, average diameter of about 70 Å, appeared to increase following an increase in the time of ageingin situ.The exact function of the mitochondria containing the paracrystalline arrays is unknown. These mitochondria could not possibly be responsible for the overall decline in the State 3 respiration, respiratory control index and the ADP/O ratio observed with intact mitochondria isolated after prolonged ageingin situ [21].     

Dynamic anchoring of PKA is essential during oocyte maturation

In the final stages of ovarian follicular development, the mouse oocyte remains arrested in the first meiotic prophase, and cAMP-stimulated PKA plays an essential role in this arrest. After the LH surge, a decrease in cAMP and PKA activity in the oocyte initiates an irreversible maturation process that culminates in a second arrest at metaphase II prior to fertilization. A-kinase anchoring proteins (AKAPs) mediate the intracellular localization of PKA and control the specificity and kinetics of substrate phosphorylation. Several AKAPs have been identified in oocytes including one at 140 kDa that we now identify as a product of the Akap1 gene. We show that PKA interaction with AKAPs is essential for two sequential steps in the maturation process: the initial maintenance of meiotic arrest and the subsequent irreversible progression to the polar body extruded stage. A peptide inhibitor (HT31) that disrupts AKAP/PKA interactions stimulates oocyte maturation in the continued presence of high cAMP. However, during the early minutes of maturation, type II PKA moves from cytoplasmic sites to the mitochondria, where it associates with AKAP1, and this is shown to be essential for maturation to continue irreversibly.     

Increased levels of (class switched) memory B cells in peripheral blood of current smokers

There is increasing evidence that a specific immune response contributes to the pathogenesis of COPD. B-cell follicles are present in lung tissue and increased anti-elastin titers have been found in plasma of COPD patients. Additionally, regulatory T cells (Tregs) have been implicated in its pathogenesis as they control immunological reactions. We hypothesize that the specific immune response in COPD is smoke induced, either by a direct effect of smoking or as a result of smoke-induced lung tissue destruction (i.e. formation of neo-epitopes or auto antigens). Furthermore, we propose that Tregs are involved in the suppression of this smoke-induced specific immune response.The presence of B cells, memory B cells and Tregs was assessed by flow cytometry in peripheral blood of 20 COPD patients and 29 healthy individuals and related to their current smoking status.COPD patients had lower (memory) B-cell percentages and higher Treg percentages in peripheral blood than healthy individuals, with a significant negative correlation between these cells. Interestingly, current smokers had higher percentages of (class-switched) memory B cells than ex-smokers and never smokers, irrespective of COPD.This increase in (class-switched) memory B cells in current smokers is intriguing and suggests that smoke-induced neo-antigens may be constantly induced in the lung. The negative correlation between B cells and Tregs in blood is in line with previously published observations that Tregs can suppress B cells. Future studies focusing on the presence of these (class switched) memory B cells in the lung, their antigen specificity and their interaction with Tregs are necessary to further elucidate the specific B-cell response in COPD.     

<Emphasis Type="Italic">In vitro</Emphasis> and <Emphasis Type="Italic">ex vivo</Emphasis> effect of hyaluronic acid on erythrocyte flow properties

Background  

Hyaluronic acid (HA) is present in many tissues; its presence in serum may be related to certain inflammatory conditions, tissue damage, sepsis, liver malfunction and some malignancies. In the present work, our goal was to investigate the significance of hyaluronic acid effect on erythrocyte flow properties. Therefore we performed in vitro experiments incubating red blood cells (RBCs) with several HA concentrations. Afterwards, in order to corroborate the pathophysiological significance of the results obtained, we replicated the in vitro experiment with ex vivo RBCs from diagnosed rheumatoid arthritis (RA) patients, a serum HA-increasing pathology.     

Assessment of the relative contribution of asexual propagation in a population of the coral-excavating sponge Cliona delitrix from the Bahamas

Cliona delitrix is a very destructive coral-excavating sponge in Caribbean coral reef systems, particularly for Montastraea species. Little is known about how these excavating sponges propagate across coral reefs. In this study a hypothesis was tested that coral breakage caused by the bioeroding activity facilitates the asexual propagation of this sponge and in turn favors the spread of the most aggressive sponge genotypes. An allozyme analysis, involving 12 loci systems of 52 sponge individuals from a total of 13 Montastraea heads, found that no two sponges possessed identical multi-locus genotypes. Contrary to the pattern expected for fragmenting species, the incidence of clonality and asexual propagation at the population level was minimal. The lack of correlation between genetic and physical distances for the studied sponges also suggests that population maintenance appears to derive from larval dispersal, with a spatial range of dispersal larger than the average distance between the coral heads (10–10² m).