Respiration-dependent proton translocation and the mechanism of protonmotive force generation in Nitrobacter winogradskyi

Abstract Proton translocation associated with electron flow to oxygen has been observed with cells of Nitrobacter winogradskyi in the presence of either potassium ferrocyanide or isoascorbate plus N , N , N ', N ' tetramethyl- p -phenylenediamine. The data are consistent with a proton pumping function for the terminal oxidase, cytochrome aa ₃, in this organism as the mechanism for generating a protonmotive force. The failure of previous work with Nitrobacter [4] to detect proton translocation linked to oxidation of nitrite, the physiological substrate, is discussed.     

Preliminary Findings of Fecal Gonadal Hormone Concentrations in Six Captive Sea Otters (Enhydra lutris) after Deslorelin Implantation

The sea otter (Enhydra lutris) is a popular exhibit animal in many zoos and aquariums worldwide. Captive sea otters from these populations are owned by the United States Fish and Wildlife Service (USFWS). The USFWS has requested that these sea otters be prevented from breeding in order to save captive space for wild rescued animals. Sea otters are often housed in mixed sex groups, therefore a chemical contraceptive method or surgical removal of gonads must be used to prevent potential pregnancy. The contraceptive, Suprelorin® or deslorelin, has been used in many different species to effectively suppress reproduction but duration of effect may vary not only between species but also individuals. Here, we report the effects of one to several consecutive deslorelin implants on gonadal reproductive hormones found in fecal samples from six captive sea otters (two males and four females) compared to two control otters (one male and one female) housed at three zoological institutions. We documented the longitudinal hormone signatures of many stages of the contraceptive cycle including pretreatment (PT), stimulatory phase (S), effective contraception (EC), and hormone reversal (HR) that was characterized by a return to normal hormone levels. Deslorelin was found to be an effective contraceptive in sea otters and was found to be reversible documented by a live birth following treatment, however the duration of suppression in females was much longer than expected with a 6‐month and a 1‐year implant lasting between 3 and 4 years in females. Zoo Biol. 32:307–315, 2013. © 2012 Wiley Periodicals, Inc.     

Tumor attenuation by 2(6-hydroxynaphthyl)-beta-D-xylopyranoside requires priming of heparan sulfate and nuclear targeting of the products

We have previously reported that the heparan sulfate-priming glycoside 2-(6-hydroxynaphthyl)-beta-D-xylopyranoside selectively inhibits growth of transformed or tumor-derived cells. To investigate the specificity of this xyloside various analogs were synthesized and tested in vitro. Selective growth inhibition was dependent on the presence of a free 6-hydroxyl in the aglycon. Because cells deficient in heparan sulfate synthesis were insensitive to the xyloside, we conclude that priming of heparan sulfate synthesis was required for growth inhibition. In growth-inhibited cells, heparan sulfate chains primed by the active xyloside were degraded to products that contained anhydromannose and appeared in the nuclei. Hence the degradation products were generated by nitric oxide-dependent cleavage. Accordingly, nitric oxide depletion reduced nuclear localization of the degradation products and counteracted the growth-inhibitory effect of the xyloside. We propose that 2-(6-hydroxynaphthyl)-beta-D-xylopyranoside entered cells and primed synthesis of heparan sulfate chains that were subsequently degraded by nitric oxide into products that accumulated in the nucleus. In vivo experiments demonstrated that the xyloside administered subcutaneously, perorally, or intraperitoneally was adsorbed and made available to tumor cells located subcutaneously. Treatment with the xyloside reduced the average tumor load by 70-97% in SCID mice. The present xyloside may serve as a lead compound for the development of novel antitumor strategies.     

Heparan sulfate proteoglycan as a plasma membrane carrier

The plasma membrane defines the border of living cells and provides a barrier to extracellular components. Advances in molecular biology have resulted in the development of novel therapeutic strategies (e.g. gene therapy and cellular protein delivery) which rely on the entry of charged macromolecules into the intracellular compartment. Recent reports demonstrate an intriguing role for heparan sulfate proteoglycans in cellular internalization of viruses, basic peptides and polycation-nucleic-acid complexes and the possibility that they have important implications for gene transfer and protein delivery to mammalian cells. This review focuses on heparan sulfate proteoglycan as a plasma membrane carrier.     

No association between Helicobacter pylori genotypes and antibiotic resistance phenotypes within families

Background. Triple therapy combining a proton pump inhibitor with two antibiotics, e.g. clarythromycin (CLR), metronidazole (MTZ) or amoxicillin (AMX), represents the standard in Helicobacter pylori eradication regimens. Resistance to antimicrobial agents, particularly MTZ (up to 56% in Western countries) and CLR (up to 15% in southern Europe), is frequently observed and may be associated with treatment failure [ 1 ]. Recently, several studies indicated that individual H. pylori colonies from a single anatomic site may not always yield identical genotypes, or the identical patterns of susceptibility to antibiotics [ 2 - 5 ]. Representative for every single patient we analyzed 27 H. pylori antrum isolates for susceptibility to antimicrobial agents in order to test whether identical H. pylori genotypes exhibit a similar pattern of susceptibility to antibiotics. Methods. PCR, RELP, PFGE, antibiotic susceptibility testing. Results. H. pylori genotype and antibiotic susceptibility pattern in families do not segregrate. Conclusion. Molecular typing of H. pylori from family members does not predict antibiotic susceptibility pattern.     

The hypothalamic neuropeptide oxytocin is required for formation of the neurovascular interface of the pituitary

The hypothalamo-neurohypophyseal system (HNS) is?the neurovascular structure through which the hypothalamic neuropeptides oxytocin and arginine-vasopressin exit the brain into the bloodstream, where they go on to affect peripheral physiology. Here, we investigate the molecular cues that regulate the neurovascular contact between hypothalamic axons and neurohypophyseal capillaries of the zebrafish. We developed a transgenic system in which both hypothalamic axons and neurohypophyseal vasculature can be analyzed in?vivo. We identified the cellular organization of the zebrafish HNS as well as the dynamic processes that contribute to formation of the HNS neurovascular interface. We show that formation of this interface is regulated during development by local release of oxytocin, which affects endothelial morphogenesis. This cell communication process is essential for the establishment of a tight axovasal interface between the neurons and blood vessels of the HNS. We present a unique example of axons affecting endothelial morphogenesis through secretion of a neuropeptide.     

DIGE proteome analysis reveals suitability of ischemic cardiac in vitro model for studying cellular response to acute ischemia and regeneration

Proteomic analysis of myocardial tissue from patient population is suited to yield insights into cellular and molecular mechanisms taking place in cardiovascular diseases. However, it has been limited by small sized biopsies and complicated by high variances between patients. Therefore, there is a high demand for suitable model systems with the capability to simulate ischemic and cardiotoxic effects in vitro, under defined conditions. In this context, we established an in vitro ischemia/reperfusion cardiac disease model based on the contractile HL-1 cell line. To identify pathways involved in the cellular alterations induced by ischemia and thereby defining disease-specific biomarkers and potential target structures for new drug candidates we used fluorescence 2D-difference gel electrophoresis. By comparing spot density changes in ischemic and reperfusion samples we detected several protein spots that were differentially abundant. Using MALDI-TOF/TOF-MS and ESI-MS the proteins were identified and subsequently grouped by functionality. Most prominent were changes in apoptosis signalling, cell structure and energy-metabolism. Alterations were confirmed by analysis of human biopsies from patients with ischemic cardiomyopathy.With the establishment of our in vitro disease model for ischemia injury target identification via proteomic research becomes independent from rare human material and will create new possibilities in cardiac research.     

Nanotubes, exosomes, and nucleic acid–binding peptides provide novel mechanisms of intercellular communication in eukaryotic cells: implications in health and disease

The prevailing view that eukaryotic cells are restrained from intercellular exchange of genetic information has been challenged by recent reports on nanotubes, exosomes, apoptotic bodies, and nucleic acid–binding peptides that provide novel pathways for cell–cell communication, with implications in health and disease.     

Complex cell rearrangements during intersegmental vessel sprouting and vessel fusion in the zebrafish embryo

The formation of intersegmental blood vessels (ISVs) in the zebrafish embryo serves as a paradigm to study angiogenesis in vivo. ISV formation is thought to occur in discrete steps. First, endothelial cells of the dorsal aorta migrate out and align along the dorsoventral axis. The dorsal-most cell, also called tip cell, then joins with its anterior and posterior neighbours, thus establishing a simple vascular network. The vascular lumen is then established via formation of vacuoles, which eventually fuse with those of adjacent endothelial cells to generate a seamless tube with an intracellular lumen. To investigate the cellular architecture and the development of ISVs in detail, we have analysed the arrangement of endothelial cell junctions and have performed single cell live imaging. In contrast to previous reports, we find that endothelial cells are not arranged in a linear head-to-tail configuration but overlap extensively and form a multicellular tube, which contains an extracellular lumen. Our studies demonstrate that a number of cellular behaviours, such as cell divisions, cell rearrangements and dynamic alterations in cell-cell contacts, have to be considered when studying the morphological and molecular processes involved in ISV and endothelial lumen formation in vivo.     

Microcavity array (MCA)-based biosensor chip for functional drug screening of 3D tissue models

Multicellular tumour spheroids that mimic a native cellular environment are widely used as model systems for drug testing. To study drug effects on three-dimensional cultures in real-time we designed and fabricated a novel type of sensor chip for fast, non-destructive impedance spectroscopy and extracellular recording. Precultured spheroids are trapped between four gold electrodes. Fifteen individual 100microm deep square microcavities with sizes from 200 to 400microm allow an optimised positioning during the measurement. Although apoptosis was induced in human melanoma spheroids by Camptothecin (CTT), treated cultures did not show disintegration but displayed increased impedance magnitudes compared to controls after 8h resulting from an altered morphology of the outer cells. Contractions in cardiomyocyte spheroids were monitored when the innovative chip was used for recording of extracellular potentials. The silicon-based electrode array is used as an acute test system for the monitoring of any kind of 3D cell cultures. Since no adherence of cells or labelling is necessary the multifunctional sensor chip provides a basis for improved drug development by high content screenings with reduced costs and assay times. Additional improvements for parallel testing of different substances on one chip are presented.