Prokaryotic lifestyles in deep sea habitats

Gradients of physicochemical factors influence the growth and survival of life in deep-sea environments. Insights into the characteristics of deep marine prokaryotes has greatly benefited from recent progress in whole genome and metagenome sequence analyses. Here we review the current state-of-the-art of deep-sea microbial genomics. Ongoing and future genome-enabled studies will allow for a better understanding of deep-sea evolution, physiology, biochemistry, community structure and nutrient cycling. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. An erratum to this article can be found at     

Physicochemical and biological study of selected hydrophobic polyethylenimine-based polycationic liposomes and their complexes with DNA

Non-viral gene therapy is based on the development of efficient and safe gene carrier systems able to transfer DNA into cells. Polyethylenimine (PEI), the most promising non-viral vector, with its high cationic-charge-density potential is able (1) to compact DNA in complexes (polyplexes) smaller than those formed by liposomes (lipoplexes) and (2) to destabilize the endosomal membrane by a 'proton sponge' effect. Several PEI's hydrophobic modifications were reported in the last several years but in some cases a reduced transfection efficiency was observed. The mechanism underlying this phenomenon is not well understood so far. In order to extensively investigate these mechanisms, we reported a physicochemical and biological study of selected hydrophobic PEI's derivatives grafted with chains of different length and percentages of substitution able to form vesicles (polycationic liposomes) and to bind DNA. Their properties were studied by means of dynamic light scattering, freeze-fracture microscopy, potentiometric titrations, gel retardation assays, polyanion exchange reactions, toxicity assays, in vitro transfections, and fluorescence microscopy. Our results indicate that even if polyplexes are able to pass through the cellular membrane, the stability of PEI's hydrophobic polyplexes likely explain their different transfection efficiency in vitro.     

Evidence for a role of glycosphingolipids in CXCR4-dependent cell migration

Chemotaxis induction is a major effect evoked by stimulation of the chemokine receptor CXCR4 with its sole ligand CXCL12. We now report that treatment of CHP-100 human neuroepithelioma cells with the glucosylceramide synthase (GCS) inhibitor DL-threo-1-phenyl-2-hexadecanoylamino-3-pyrrolidino-1-propanol inhibits CXCR4-dependent chemotaxis. We provide evidence that the phenomenon is not due to unspecific effects of the inhibitor employed and that inhibition of GCS neither affects total or plasmamembrane CXCR4 expression, nor CXCL12-induced Ca(2+) mobilization. The effects of the GCS inhibitor on impairment of CXCL12-induced cell migration temporally correlated with a pronounced downregulation of neutral glycosphingolipids, particularly glucosylceramide, and with a delayed and more moderate downregulation of gangliosides; moreover, exogenously administered glycosphingolipids allowed resumption of CXCR4-dependent chemotaxis. Altogether our results provide evidence, for the first time, for a role glycosphingolipids in sustaining CXCL12-induced cell migration.     

Statin therapy lowers muscle sympathetic nerve activity and oxidative stress in patients with heart failure

Despite standard drug therapy, sympathetic nerve activity (SNA) remains high in heart failure (HF) patients making the sympathetic nervous system a primary drug target in the treatment of HF. Studies in rabbits with pacing-induced HF have demonstrated that statins reduce resting SNA, in part, due to reductions in reactive oxygen species (ROS). Whether these findings can be extended to the clinical setting of human HF remains unclear. We first performed a study in seven statin-na?ve HF patients (56 ± 2 yr; ejection fraction: 31 ± 4%) to determine if 1 mo of simvastatin (40 mg/day) reduces muscle SNA (MSNA). Next, to control for possible placebo effects and determine the effect of simvastatin on ROS, a double-blinded, placebo-controlled crossover design study was performed in six additional HF patients (51 ± 3 yr; ejection fraction: 22 ± 4%), and MSNA, ROS, and superoxide were measured. We tested the hypothesis that statin therapy decreases resting MSNA in HF patients and this would be associated with reductions in ROS. In study 1, simvastatin reduced resting MSNA (75 ± 5 baseline vs. 65 ± 5 statin bursts/100 heartbeats; P < 0.05). Likewise, in study 2, simvastatin also decreased resting MSNA (59 ± 5 placebo vs. 45 ± 6 statin bursts/100 heartbeats; P < 0.05). In addition, statin therapy significantly reduced total ROS and superoxide. As expected, cholesterol was reduced after simvastatin. Collectively, these findings indicate that short-term statin therapy concomitantly reduces resting MSNA and total ROS and superoxide in HF patients. Thus, in addition to lowering cholesterol, statins may also be beneficial in reducing sympathetic overactivity and oxidative stress in HF patients.     

Interaction of a solitary larval endoparasitoid, Microplitis mediator, with its host, Mamestra brassicae: host acceptance and host suitability

Abstract:  Microplitis mediator (Haliday) (Hym., Braconidae) is an important parasitoid of early instar larvae of the European cabbage moth, Mamestra brassicae L. (Lep., Noctuidae). In the laboratory, we examined attack responses of female M. mediator to the first three larval instars of M. brassicae . Females were presented with M. brassicae larvae either one individual at a time in a no-choice experiment, or three individuals, one from each instar, simultaneously in a choice experiment. Whether or not there was choice, naïve female parasitoids attacked a high proportion of larvae and did not discriminate among instars. In the no-choice experiment, attacked larvae were reared, and parasitoid cocoons were produced from about 76% of larvae attacked as first and second instars, but from only 19% of larvae attacked as third instars. Dissections of attacked larvae from the choice experiment showed that about 79% of attacks on first and second instars resulted in oviposition compared with only 49% for third instars. When given choice, frequency and number of attacks on first instar larvae increased with increasing parasitoid experience. Our results suggest that first and second instar larvae of M. brassicae are suitable hosts for M. mediator , but that third instar larvae are suboptimal both because oviposition attempts were frequently unsuccessful and because immature parasitoids failed to complete development. Nevertheless, naïve attacking parasitoids exhibited minimal discrimination among instars, although experienced parasitoids most frequently attacked first instar larvae. The host selection behaviour of M. mediator is discussed in the context of optimal foraging theory and implications for biological control.     

Histological process and dynamics of germ cell degeneration in pejerrey Odontesthes bonariensis larvae and juveniles during exposure to warm water

Elevated temperature causes degeneration and disappearance of the germ cells in the males of scrotal mammals. It was recently shown that heat-induced germ cell degeneration occurs also in fish but, unlike in mammals, it occurs not only in males but also in females. The purpose of this study was to clarify the histological process and dynamics of heat-induced germ cell disappearance in pejerrey Odontesthes bonariensis larvae and juveniles. Monosex and mixed-sex fish produced by thermal manipulation of sex (temperature-dependent sex determination) were subjected to 29 degrees C for periods between 1 and 12 weeks, and used to analyze, by histological methods, the changes in gonadal size and the number of normal and degenerating germ cells. Groups exposed to 29 degrees C for 8-12 weeks were subsequently transferred to 24 degrees C to verify if any gonadal damage would be permanent. Germ cell degeneration, histologically characterized by nuclear pyknosis or eosinophilia and cytoplasmic eosinophilia, was observed with increasing frequency at higher temperatures (29>24> 17 degrees C) and more in males than in females. Clear degenerative changes in the germinal epithelium usually began within one week of exposure to 29 degrees C and appeared clearer in females than in males. Complete loss of germ cells was observed only in individuals exposed for periods of 8-12 weeks to 29 degrees C but no treatment produced 100% sterile fish. Germ cells that remained in the gonads after exposure to 29 degrees C retained the capacity to rapidly recolonize germ cell-depleted areas, suggesting that the associated somatic cells in the gonads are little or not affected by this temperature.     

Effects of crude extracts of Agaricus blazei on DNA damage and on rat liver carcinogenesis induced by diethylnitrosamine

The effects of crude extracts of the mushroom Agaricus blazei Murrill (Agaricaceae) on both DNA damage and placental form glutathione S-transferase (GST-P)-positive liver foci induced by diethylnitrosamine (DEN) were investigated. Six groups of adult male Wistar rats were used. For two weeks, animals of groups 3 to 6 were treated with three aqueous solutions of A. blazei (mean dry weight of solids being 1.2, 5.6, 11.5 and 11.5 mg/ml, respectively). After this period, groups 2 to 5 were given a single ip injection 200 mg/kg DEN and groups 1 and 6 were treated with 0.9% NaCl. All animals were subjected to 70% partial hepatectomy at week five and sacrificed 4, 24 and 48 h or 8 weeks after DEN or 0.9% NaCl treatments (10th week after the beginning of the experiment). The alkaline comet assay and GST-P-positive liver foci development were used to evaluate the influence of the mushroom extracts on liver cell DNA damage and on the initiation of liver carcinogenesis, respectively. Previous treatment with the highest concentration of A. blazei (11.5 mg/ml) significantly reduced DNA damage, indicating a protective effect against DEN-induced liver cytotoxicity/genotoxicity. However, the same dose of mushroom extract significantly increased the number of GST-P-positive liver foci.     

Characterization of apoptosis signal transduction pathways in HL-5 cardiomyocytes exposed to ischemia/reperfusion oxidative stress model

During ischemia/reperfusion (I/R), cardiomyocytes are exposed to sudden lack of nutrients and successively to radical oxygen species (ROS). In the present study, we used the HL-5 cardiac atrial myocyte cell line exposed to serum/glucose depletion added or not in H(2)O(2) to mimic ROS during ischemia, then replaced in their standard culture medium to simulate reperfusion. We investigated the effects of serum/glucose depletion combined or not to ROS exposure on AKT and MAP kinases activation to address the role of each event with respect to apoptosis. We demonstrate that serum/glucose depletion per se did not induce apoptosis when compared to ROS exposure. In particular, ROS recruited p38MAPK and JNK pathways. SB202190 preventing p38MAPK activity, partially protected HL-5 from apoptosis while blocking JNK, thanks to JNKI, further enhanced apoptosis. Blocking phosphatidylinositol (PI) 3-kinase with LY294002 or ERKs with U0126 was without consequence on apoptosis. Finally, BCL-2 and BCL-X(L/S) expression levels were analyzed in cells exposed to 1 h ischemia followed by 12-h reperfusion in the presence or not of SB202190; BCL-2, but not BCL-X(L/S), expression was decreased in ROS treated cells but SB202190 failed to restore BCL-2 level. Our data suggest that p38MAPK activation primarily mediates ROS-induced apoptosis while concomitant JNK activation would represent a scavenger pathway for cells trying to escape apoptosis.     

Mandibuloacral dysplasia is caused by a mutation in LMNA-encoding lamin A/C

Mandibuloacral dysplasia (MAD) is a rare autosomal recessive disorder, characterized by postnatal growth retardation, craniofacial anomalies, skeletal malformations, and mottled cutaneous pigmentation. The LMNA gene encoding two nuclear envelope proteins (lamins A and C [lamin A/C]) maps to chromosome 1q21 and has been associated with five distinct pathologies, including Dunnigan-type familial partial lipodystrophy, a condition that is characterized by subcutaneous fat loss and is invariably associated with insulin resistance and diabetes. Since patients with MAD frequently have partial lipodystrophy and insulin resistance, we hypothesized that the disease may be caused by mutations in the LMNA gene. We analyzed five consanguineous Italian families and demonstrated linkage of MAD to chromosome 1q21, by use of homozygosity mapping. We then sequenced the LMNA gene and identified a homozygous missense mutation (R527H) that was shared by all affected patients. Patient skin fibroblasts showed nuclei that presented abnormal lamin A/C distribution and a dysmorphic envelope, thus demonstrating the pathogenic effect of the R527H LMNA mutation.     

Synthesis and evaluation of the cardiovascular effects of two, membrane impermeant, macromolecular complexes of dextran-testosterone

The incidence of cardiovascular disease is greater in men than in premenopausal women. Testosterone has been considered a significant risk factor for cardiovascular disease, but testosterone's mechanism of action and its cellular site of action are still not clear. However, it is likely that non-genomic extracellular effects of the hormone are involved. With the aim of providing further information about this phenomenon, two membrane impermeant, macromolecular complexes of testosterone were synthesized and their cardiovascular effects were evaluated. We covalently bound testosterone (through carbon 3 or C-17 functional groups) to dextran (2 MDa) and evaluated its effects on isolated and perfused rat hearts (Langerdorff model). Our results showed that the macromolecular complexes increased vascular resistance similarly to free testosterone and blocked adenosine-induced vasodilatation. These effects were exerted rapidly and possibly through a non-genomic mechanism. Blockade of C-3 or C-17 functional groups by binding to macromolecular dextran induced no qualitative and/or quantitative changes in testosterone-induced effects.