Search

Building a blastema from the stump is a key step of salamander limb regeneration. Stump-derived cells temporarily suspend their identity as they contribute to the blastema by a process generally referred to as dedifferentiation. Here, we provide evidence for a mechanism that involves an active inhibition of protein syn-thesis during blastema formation and growth. Relieving this inhibition results in a higher number of cycling cells and enhances the pace of limb regeneration. By small RNA profiling and fate mapping of skeletal muscle progeny as a cellular model for dedifferentiation, we find that the downregulation of miR-10b-5p is critical for rebooting the translation machinery. miR-10b-5p targets ribosomal mRNAs, and its artificial upregulation causes decreased blastema cell proliferation, reduction in transcripts that encode ribosomal subunits, dimin-ished nascent protein synthesis, and retardation of limb regeneration. Taken together, our data identify a link between miRNA regulation, ribosome biogenesis, and protein synthesis during newt limb regeneration.

One of the greatest enigmas of modern biology is how the geometry of muscular and skeletal structures are created and how their development is controlled during growth and regeneration. Scaling and shaping of vertebrate muscles and skeletal elements has always been enigmatic and required an advanced technical level in order to analyse the cell distribution in 3D. In this work, synchrotron X-ray computed microtomography (µCT) and chemical contrasting has been exploited for a quantitative analysis of the 3D-cell distribution in tissues of a developing salamander (Pleurodeles waltl) limb - a key model organism for vertebrate regeneration studies. We mapped the limb muscles, their size and shape as well as the number and density of cells within the extracellular matrix of the developing cartilage. By using tomographic approach, we explored the polarity of the cells in 3D, in relation to the structure of developing joints. We found that the polarity of chondrocytes correlates with the planes in joint surfaces and also changes along the length of the cartilaginous elements. Our approach generates data for the precise computer simulations of muscle-skeletal regeneration using cell dynamics models, which is necessary for the understanding how anisotropic growth results in the precise shapes of skeletal structures.

Salamanders have been hailed as champions of regeneration, exhibiting a remarkable ability to regrow tissues, organs and even whole body parts, e.g. their limbs. As such, salamanders have provided key insights into the mechanisms by which cells, tissues and organs sense and regenerate missing or damaged parts. In this Primer, we cover the evolutionary context in which salamanders emerged. We outline the varieties of mechanisms deployed during salamander regeneration, and discuss how these mechanisms are currently being explored and how they have advanced our understanding of animal regeneration. We also present arguments about why it is important to study closely related species in regeneration research.

Regeneration and tissue repair are widely spread in the animal kingdom, and actually present in many phyla, although with surprisingly different efficiencies within a given phylum [1]. Moreover, even though most regenerative processes depend on blastema formation, i.e. proliferation of mesenchymal cells in the vicinity of the amputation plane, the ways to achieve this process are multiple (Fig. 1). In fact, the questions linked to the phenomena of regeneration and tissue repair are numerous, and each forms in itself one piece of this amazing puzzle for biologists to reconstitute. Some of those are listed here: Where is the boundary between regeneration and tissue repair? How much does tissue repair impinge on regeneration? What are the mechanisms that control cell proliferation in cases where a blastema is formed and where it does not? Are these mechanisms conserved across evolution? Where are the differences between the mechanisms that control cell proliferation in blastemas and those that control cell proliferation in oncogenic tumors? Are the regenerative processes leading to the replacement of amputated structures close to embryonic/fetal developmental processes, or do they rather represent distinct adult developmental processes ? What are the memory mechanisms that allow the reactivation of an embryonic/fetal/larval developmental program? What are the criteria that distinguish regeneration in closely related species such as in the neotenic axolotl or in the postmetamorphic newt? Is it possible to consider appendage regeneration in a larval-stage animal as a short memory process, i.e. memory of a recently achieved developmental process, whereas that occurring in adult animals would involve a long memory process? At the other extremity of the life cycle, what are the links between the regeneration potential and the ageing process in a given species? These questions are fundamental biological questions, and their understanding will open new avenues to regenerative medicine. An important prerequisite, however, is systematic comparative functional analyses of the various regenerative modes at hand in vertebrates as well as invertebrates. In this issue we propose to approach some of these questions through a variety of angles and model systems.

The overall bauplan of the tetrapod brain is highly conserved, yet significant variations exist among species in terms of brain size, structural composition and cellular diversity. Understanding processes underlying neural and behavioral development in a wide range of species is important both from an evolutionary developmental perspective as well as for the identification of cell sources with post-developmental neurogenic potential. Here we characterize germinal processes in the brain of Notophthalmus viridescens and Pleurodeles waltl during both development and adulthood. Using a combination of cell tracking tools, including clonal analyses in new transgenic salamander lines we examine the origin of neural stem and progenitor cells found in the adult brain, determine regional variability in cell cycle length of progenitor cells, and show spatio-temporally orchestrated neurogenesis. We analyze how maturation of different brain regions and neuronal subpopulations are linked to the acquisition of complex behaviors, and how these behaviors are altered upon chemical ablation of dopamine neurons. Our data analyzed from an evolutionary perspective reveal both common and species-specific processes in tetrapod brain formation and function.

Three new secondary metabolites kermesoside (1), cocciferoside (2) and (-)-8-chlorocatechin (3), were isolated from the the stems with barks of Quercus coccifera along with five known phenolic compounds, 3-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-propan-1-one (4) and 3-hydroxy-1-(4-hydroxy-3,5-dimethoxyphenyl)-propan-1-one (5), trans-resveratrol-3-O-β-glucopyranoside (6) lyoniresinol-9-O-β-xylopyranoside (7), lyoniresinol-9-O-β-glucopyranoside (8). The structure elucidation of the isolated compounds was performed by spectroscopic methods (UV, 1D- and 2D- NMR and HR-MS).

Ecdysteroids, molting hormones of insects, can exert several mild, non-hormonal bioactivities in mammals, including humans. In a previous study, we have found a significant effect of certain derivatives on the ABCB1 transporter mediated multi-drug resistance of a transfected murine leukemia cell line. In this paper, we present a structure-activity relationship study focused on the apolar dioxolane derivatives of 20-hydroxyecdysone. Semi-synthesis and bioactivity of a total of 32 ecdysteroids, including 20 new compounds, is presented, supplemented with their complete 1H- and 13C-NMR signal assignment. publishersversion published

Degeneration of dopamine neurons in the midbrain causes symptoms of the movement disorder, Parkinson disease. Dopamine neurons are generated from proliferating progenitor cells localized in the embryonic ventral midbrain. However, it remains unclear for how long cells with dopamine progenitor character are retained and if there is any potential for reactivation of such cells after cessation of normal dopamine neurogenesis. We show here that cells expressing Lmx1a and other progenitor markers remain in the midbrain aqueductal zone beyond the major dopamine neurogenic period. These cells express dopamine receptors, are located in regions heavily innervated by midbrain dopamine fibres and their proliferation can be stimulated by antagonizing dopamine receptors, ultimately leading to increased neurogenesis in vivo. Furthermore, treatment with dopamine receptor antagonists enhances neurogenesis in vitro, both from embryonic midbrain progenitors as well as from embryonic stem cells. Altogether our results indicate a potential for reactivation of resident midbrain cells with dopamine progenitor potential beyond the normal period of dopamine neurogenesis.

Organisms need to adapt to the ecological constraints in their habitat. How specific processes reflect such adaptations are difficult to model experimentally. We tested whether environmental shifts in oxygen tension lead to events in the adult newt brain that share features with processes occurring during neuronal regeneration under normoxia. By experimental simulation of varying oxygen concentrations, we show that hypoxia followed by re-oxygenation lead to neuronal death and hallmarks of an injury response, including activation of neural stem cells ultimately leading to neurogenesis. Neural stem cells accumulate reactive oxygen species (ROS) during re-oxygenation and inhibition of ROS biosynthesis counteracts their proliferation as well as neurogenesis. Importantly, regeneration of dopamine neurons under normoxia also depends on ROS-production. These data demonstrate a role for ROS-production in neurogenesis in newts and suggest that this role may have been recruited to the capacity to replace lost neurons in the brain of an adult vertebrate. DOI: http://dx.doi.org/10.7554/eLife.08422.001, eLife digest During the winter, red-spotted newts remain active in water that is covered by ice. The oxygen levels under the ice tend to drop and so the newts adjust their metabolism to cope with these conditions. However, when oxygen levels return to normal, this may result in the newts producing larger amounts of chemically reactive molecules called reactive oxygen species (ROS). These molecules form naturally as a by-product of oxygen metabolism, but in high quantities they can damage cells and tissues. It has been proposed that red-spotted newts and other animals that experience periods of low oxygen may have evolved processes to repair such damage. Unlike us, red-spotted newts are able to replace nerve cells in the brain that have died or been injured. This regeneration is fuelled by stem cells called ependymoglia cells, which divide to produce new nerve cells. Here, Hameed et al. investigated whether the return of oxygen to normal levels after a period of low oxygen can damage nerve cells in the newts, and whether this is followed by regeneration. The experiments show that nerve cells in the newt brain do indeed die when oxygen levels return to normal. Also, the brain activates an injury response that triggers the ependymoglia cells to divide. During this process, the ependymoglia cells accumulate ROS and their ability to divide is impaired if the production of ROS is blocked. The replacement of injured brain cells in normal oxygen conditions also depends on increased ROS levels. Together, Hameed et al.'s findings demonstrate a key role for ROS production in controlling the regeneration of damaged nerve cells in the red-spotted newt. A future challenge is to identify the genes that control the survival and activation of ependymoglia cells in response to increased ROS levels in the brain. DOI: http://dx.doi.org/10.7554/eLife.08422.002

In contrast to mammals, salamanders can regenerate complex structures after injury, including entire limbs. A central question is whether the generation of progenitor cells during limb regeneration and mammalian tissue repair occur via separate or overlapping mechanisms. Limb regeneration depends on the formation of a blastema, from which the new appendage develops. Dedifferentiation of stump tissues, such as skeletal muscle, precedes blastema formation, but it was not known whether dedifferentiation involves stem cell activation. We describe a multipotent Pax7+ satellite cell population located within the skeletal muscle of the salamander limb. We demonstrate that skeletal muscle dedifferentiation involves satellite cell activation and that these cells can contribute to new limb tissues. Activation of salamander satellite cells occurs in an analogous manner to how the mammalian myofiber mobilizes stem cells during skeletal muscle tissue repair. Thus, limb regeneration and mammalian tissue repair share common cellular and molecular programs. Our findings also identify satellite cells as potential targets in promoting mammalian blastema formation.

Based on the reaction of Zn(II), thiocyanate and 2-[2-(5-dimethylamino-thiophen-2-yl)-vinyl]-1,3,3-trimethyl-3H-indolium bromide (DTVTI), a donor-active solvent-assisted liquid-phase microextraction procedure followed by spectrophotometric determination of zinc at 570 nm was developed. The optimum experimental conditions were investigated and found to be as follows: concentration of NH4SCN 0.02 mol L-1, concentration of DTVTI 4 × 10-5 mol L-1. Various extraction solvents were studied alone as well as in mixtures with different improvers, and a mixture of toluene as the extraction solvent and tributylphosphate as the donor-active solvent in a 4:1 v/v ratio was selected. The calibration plot was linear up to 2.62 mg L-1 of zinc with limit of detection 0.09 mg L-1. The developed procedure was applied for zinc determination in dietary supplements. O presente trabalho descreve o uso de microextração em fase líquida com um solvente doador-ativo para a determinação espectrofotométrica de zinco, em 570 nm, usando a reação com tiocianato e com o reagente brometo de 2-[2-(5-dimetilamino-2-tiofenil)-vinil]-1,3,3-trimetil-3H-indolium (DTVTI). As condições experimentais ótimas foram estabelecidas usando NH4SCN 0,02 mol L-1 e DTVTI 4 × 10-5 mol L-1. Estudaram-se ainda vários solventes extratores, separados e em misturas, com diferentes promotores, sendo selecionada a mistura de tolueno como solvente de extração e tributilfosfato como solvente doador-ativo na razão de 4:1 v/v. O sistema mostrou resposta linear até a concentração de 2,62 mg L-1 de zinco com um limite de detecção de 0,09 mg L-1. O método desenvolvido foi aplicado na determinação de zinco em suplementos dietéticos.

Two new and one known ecdysteroids were identified in the methanolic extract of the roots ofSerratula wolffii. The new compounds isolated were ponasterone A-22-apioside (1) and 3-epi-shidasterone (3), together with the known 3-epi-22-deoxy-20-hydroxyecdysone (2). The structures of compounds1–3were determined by extensive spectroscopic techniques, including one- and two-dimensional NMR methods.

The catalytic enantioselective epoxidation of the isoflavones 1a–f has been performed by the Mn(III)salen complexes (R,R)-3 and (S,S)-3 as catalysts and dimethyldioxirane as the oxygen-atom source to afford optically active isoflavone epoxides 2a–f. The absolute configuration of the nonracemic epoxides 2 have been determined by X-ray diffraction analysis. Our present results constitute the first examples of the preparation of optically active isoflavone epoxides.

We study the notion of H-dimension and the formally stronger k-variable property, as considered by Gabbay, Immerman and Kozen. We exhibit a class of flows of time that has H-dimension 3, and admits a finite expressively complete set of one-dimensional temporal connectives, but does not have the k-variable property for any finite k.