A role for antizyme inhibitor in cell proliferation

Acknowledgments We thank Ewa Dahlberg and Lena Thiman for expert technical help with cell culturing and HPLC, respectively. The authors acknowledge financial support from the Portuguese Foundation for Science and Technology—SFRH/BD/46364/2008 (PhD fellowship to T. M. S.), the Gunnar Nilssons Cancer Foundation, the Mrs Berta Kamprad Foundation and the Per-Eric and Ulla Schyberg Foundation. Heather Wallace was supported by a visiting scholarship from Lund University. We are grateful for the generous supply of AzI antibody and pcDNA3.1-AzI from Dr. Senya Matsufuji and Dr. Yasuku Murakami (Tokyo, Japan), and Dr. Leif Andersson (Helsinki, Finland), respectively.Peer reviewedPublisher PD

Biocompatibility of a polymer based on Off-Stoichiometry Thiol-Enes + Epoxy (OSTE+) for neural implants.

The flexibility of implantable neural probes has increased during the last 10 years, starting with stiff materials such as silicone to more flexible materials like polyimide. We have developed a novel polymer based on Off-Stoichiometry Thiol-Enes + Epoxy (OSTE+, consisting of a thiol, two allyls, an epoxy resin and two initiators), which is up to 100 times more flexible than polyimide. Since a flexible neural probe should be more biocompatible than a stiff probe, an OSTE+ probe should be more biocompatible than one composed of a more rigid material. We have investigated the toxicity of OSTE+ as well as of OSTE+ that had been incubated in water for a week (OSTE+H2O) using MTT assays with mouse L929 fibroblasts. We found that OSTE+ showed cytotoxicity, but OSTE+H2O did not. Extracts were analyzed using LC-MS and GC-MS in order to identify leaked chemicals

Cells and Holograms – Holograms and Digital Holographic Microscopy as a Tool to Study the Morphology of Living Cells

We present a method to study the morphology of living, dividing and dying cells using DHM. DHM is a non-invasive, non-destructive and non-phototoxic method which allows the user to perform both qualitative and quantitative measurements of living cells over time. We show here our results on cell division and cell death in single cells. The morphological analyses performed here show changes caused by cell death and cell division, and indicate the possibilities to discriminate between different types of cell death. Cells dying in an apoptosis-like manner display different cell area and cell thickness profiles over time compared to cells dying in a necrosis-like manner, although their volume profiles are very similar. Dividing cells show a characteristic dip in the volume profile, which makes them easily distinguishable. Also, several previous studies show the versatile abilities of DHM. Different cell types have been studied and the morphology has been used to determine cell functionality as well as changes in morphology related to the environment. Cell morphology parameters can be very useful when following the effects of different treatments, the process of differentiation as well as cell growth and cell death. Cell morphology studied by DHM can be useful in toxicology, stem cell and cancer research

Digital Holography and Cell Studies

Digital holography microscopy (DHM) has developed into a broad field, and one of all the interesting applications is to study cells without staining, labeling or in any other way affecting them. Both fixed and living, dying or dead cells can be studied. The first DHM images showing living cells were published in 2004 and 2005 (Carl et al. 2004, Marquet et al. 2005), making this field of research rather new. Digital holography makes it possible to easily measure cell properties that previously have been very difficult to study, such as cell thickness and volume (Marquet et al. 2005, Mölder et al. 2008). Two of the major advantages of DHM is the 3-D imaging possibility and measurements over time. Digital holography has ben used to study several types of cells, such as nerve cells, red blood cells, stem cells and cancer cells (Emery et al. 2007, Kemper et al. 2006, Langehanenberg et al. 2009) . It has also been applied for studies of cell proliferation, cell movement, sub-cellular structures and cell morphology (Kemper et al. 2009, Yu et al. 2009). Both 2-D and 3-D cell movement can be determined ( Langehanenberg et al. 2009). Even cell viability status can be determined using DHM. Interestingly, it is possible to study both single cells and entire populations simultaneously, allowing for very nuanced studies. Older, well known techniques often require some degree of cell disturbance such as the fluorescent antibody labeling required for fluorescense or confocal microscopy studies. In this paper we will present some of the studies made possible by DHM. We will compare DHM with previously used techniques and discuss the benefits and drawbacks of digital holography cell measurements

COMPRENDO: Focus and approach

Tens of thousands of man-made chemicals are in regular use and discharged into the environment. Many of them are known to interfere with the hormonal systems in humans and wildlife. Given the complexity of endocrine systems, there are many ways in which endocrine-disrupting chemicals (EDCs) can affect the body’s signaling system, and this makes unraveling the mechanisms of action of these chemicals difficult. A major concern is that some of these EDCs appear to be biologically active at extremely low concentrations. There is growing evidence to indicate that the guiding principle of traditional toxicology that “the dose makes the poison” may not always be the case because some EDCs do not induce the classical dose–response relationships. The European Union project COMPRENDO (Comparative Research on Endocrine Disrupters—Phylogenetic Approach and Common Principles focussing on Androgenic/Antiandrogenic Compounds) therefore aims to develop an understanding of potential health problems posed by androgenic and antiandrogenic compounds (AACs) to wildlife and humans by focusing on the commonalities and differences in responses to AACs across the animal kingdom (from invertebrates to vertebrates)

Polyamine dependence of normal cell-cycle progression

The driving force of the cell cycle is the activities of cyclin-dependent kinases (CDKs). Key steps in the regulation of the cell cycle therefore must impinge upon the activities of the CDKs. CDKs exert their functions when bound to cyclins that are expressed cyclically during the cell cycle. Polyamine biosynthesis varies bicyclically during the cell cycle with peaks in enzyme activities at the G(1)/S and S/G(2) transitions. The enzyme activities are regulated at transcriptional, translational and post-translational levels. When cells are seeded in the presence of drugs that interfere with polyamine biosynthesis, cell cycle progression is affected within one cell cycle after seeding. The cell cycle phase that is most sensitive to polyamine biosynthesis inhibition is the S phase, while effects on the G(1) and G(2)/M phases occur at later time points. The elongation step of DNA replication is negatively affected when polyamine pools are not allowed to increase normally during cell proliferation. Cyclin A is expressed during the S phase and cyclin A/CDK2 is important for a normal rate of DNA elongation. Cyclin A expression is lowered in cells treated with polyamine biosynthesis inhibitors. Thus, polyamines may affect S phase progression by participating in the regulation of cyclin A expression

Polyamine-Dependent Early Cellular Signals and Cell Proliferation

When a resting cell is awakened to enter the cell cycle by mitogenic stimulation, a complex molecular machinery involving positive and negative regulation of cell-cycle progression is initiated. Part of the machinery is quite well-known. It involves receptors of various kinds, protein interactions, phosphorylation reactions, and molecular cascades carrying the signal to enter the cell cycle from the cell surface to the nucleus. In the nucleus, transcriptional activation results in an increased involvement of genes in the process. In the cytoplasm, translational and posttranslational mechanisms also have a part in the process. In this complex partly known molecular machinery, there are also molecules that not yet have been assigned a specific role; however, there are enough data to state that their role in normal cell-cycle progression is ubiquitous. The level of the polyamines—putrescine, spermidine, and spermine—are low in nonproliferating cells, but their levels increase early after mitogenic stimulation and these increases are necessary for normal rates of cell-cycle progression (1,2). In this chapter, the focus will be on the role of polyamines in early signaling processes that are required for normal cell-cycle progression

Cells and polyamines do it cyclically

Cell-cycle progression is a one-way journey where the cell grows in size to be able to divide into two equally sized daughter cells. The cell cycle is divided into distinct consecutive phases defined as G(1) (first gap), S (synthesis), G(2) (second gap) and M (mitosis). A non-proliferating cell, which has retained the ability to enter the cell cycle when it receives appropriate signals, is in G(0) phase, and cycling cells that do not receive proper signals leave the cell cycle from G(1) into G(0). One of the major events of the cell cycle is the duplication of DNA during S-phase. A group of molecules that are important for proper cell-cycle progression is the polyamines. Polyamine biosynthesis occurs cyclically during the cell cycle with peaks in activity in conjunction with the G(1)/S transition and at the end of S-phase and during G(2)-phase. The negative regulator of polyamine biosynthesis, antizyme, shows an inverse activity compared with the polyamine biosynthetic activity. The levels of the polyamines, putrescine, spermidine and spermine, double during the cell cycle and show a certain degree of cyclic variation in accordance with the biosynthetic activity. When cells in G(0)/G(1) -phase are seeded in the presence of compounds that prevent the cell-cycle-related increases in the polyamine pools, the S-phase of the first cell cycle is prolonged, whereas the other phases are initially unaffected. The results point to an important role for polyamines with regard to the ability of the cell to attain optimal rates of DNA replication