Immunohistochemical characterisation of the adult Nothobranchius furzeri intestine.

Nothobranchius furzeri is emerging as an exciting vertebrate organism in the field of biomedicine, developmental biology and ecotoxicology research. Its short generation time, compressed lifespan and accelerated ageing make it a versatile model for longitudinal studies with high traceability. Although in recent years the use of this model has increased enormously, there is still little information on the anatomy, morphology and histology of its main organs. In this paper, we present a description of the digestive system of N. furzeri, with emphasis on the intestine. We note that the general architecture of the intestinal tissue is shared with other vertebrates, and includes a folding mucosa, an outer muscle layer and a myenteric plexus. By immunohistochemical analysis, we reveal that the mucosa harbours the same type of epithelial cells observed in mammals, including enterocytes, goblet cells and enteroendocrine cells, and that the myenteric neurons express neurotransmitters common to other species, such as serotonin, substance P and tyrosine hydroxylase. In addition, we detect the presence of a proliferative compartment at the base of the intestinal folds. The description of the normal intestinal morphology provided here constitutes a baseline information to contrast with tissue alterations in future lines of research assessing pathologies, ageing-related diseases or damage caused by toxic agents.

Modeling familial and sporadic Parkinson's disease in small fishes.

The establishment of animal models for Parkinson's disease (PD) has been challenging. Nevertheless, once established, they will serve as valuable tools for elucidating the causes and pathogenesis of PD, as well as for developing new strategies for its treatment. Following the recent discovery of a series of PD causative genes in familial cases, teleost fishes, including zebrafish and medaka, have often been used to establish genetic PD models because of their ease of breeding and gene manipulation, as well as the high conservation of gene orthologs. Some of the fish lines can recapitulate PD phenotypes, which are often more pronounced than those in rodent genetic models. In addition, a new experimental teleost fish, turquoise killifish, can be used as a sporadic PD model, because it spontaneously manifests age-dependent PD phenotypes. Several PD fish models have already made significant contributions to the discovery of novel PD pathological features, such as cytosolic leakage of mitochondrial DNA and pathogenic phosphorylation in α-synuclein. Therefore, utilizing various PD fish models with distinct degenerative phenotypes will be an effective strategy for identifying emerging facets of PD pathogenesis and therapeutic modalities.

RNAseq Analysis of Brain Aging in Wild Specimens of Short-Lived Turquoise Killifish: Commonalities and Differences With Aging Under Laboratory Conditions.

A vast body of studies is available that describe age-dependent gene expression in relation to aging in a number of different model species. These data were obtained from animals kept in conditions with reduced environmental challenges, abundant food, and deprivation of natural sensory stimulation. Here, we compared wild- and captive aging in the short-lived turquoise killifish (Nothobranchius furzeri). These fish inhabit temporary ponds in the African savannah. When the ponds are flooded, eggs hatch synchronously, enabling a precise timing of their individual and population age. We collected the brains of wild fish of different ages and quantified the global age-dependent regulation of transcripts using RNAseq. A major difference between captive and wild populations is that wild populations had unlimited access to food and hence grew to larger sizes and reached asymptotic size more rapidly, enabling the analysis of age-dependent gene expression without the confounding effect of adult brain growth. We found that the majority of differentially expressed genes show the same direction of regulation in wild and captive populations. However, a number of genes were regulated in opposite direction. Genes downregulated in the wild and upregulated in captivity were enriched for terms related to neuronal communication. Genes upregulated in the wild and downregulated in captive conditions were enriched in terms related to DNA replication. Finally, the rate of age-dependent gene regulation was higher in wild animals, suggesting a phenomenon of accelerated aging.

Identification of Isthmin1 in the small annual fish, Nothobranchius guentheri, as a novel biomarker of aging and its potential rejuvenation activity.

Isthmin 1 (Ism1) has been shown to play roles in multiple biological processes including morphogenesis, hematopoiesis, antiviral immune response and suppression of tumor growth. However, it remains unknown if it plays any role in aging process. Here we showed for the first time that Ism1 was a new age-related biomarker, which decreased with age in fish, mice and humans. Interestingly, Ism1 was also useful to measure the "rejuvenated" age of fish Nothobranchius guentheri reversed by salidroside treatment and temperature reduction, providing additional evidence that Ism1 was an aging biomarker. In addition, we clearly showed that dietary intake of recombinant Ism1 had little effects on the body length and weight of aging N. guentheri, but it retarded the onset of age-related biomarkers and prolonged both the maximum and median lifespan of the fish. We also showed that Ism1 exerted its rejuvenation activity via the enhancement of antioxidant system. Collectively, our results indicate that Ism1 is not only is a novel biomarker of aging but also a potential rejuvenation factor capable of reversing aging of N. guentheri.

The sources of sex differences in aging in annual fishes.

Intersexual differences in life span (age at death) and aging (increase in mortality risk associated with functional deterioration) are widespread among animals, from nematodes to humans. Males often live shorter than females, but there is substantial unexplained variation among species and populations. Despite extensive research, it is poorly understood how life span differences between the sexes are modulated by an interplay among genetic, environmental and social factors. The goal of our study was to test how sex differences in life span and ageing are modulated by social and environmental factors, and by intrinsic differences between males and females. To disentangle the complex basis of sex differences in life span and aging, we combined comparative data from sex ratios in 367 natural populations of four species of African annual killifish with experimental results on sex differences in life span and aging from eight laboratory populations tested in treatments that varied social and environmental conditions. In the wild, females consistently outlived males. In captivity, sex-specific mortality depended on social conditions. In social-housed experimental groups, male-biased mortality persisted in two aggressive species, but ceased in two placid species. When social and physical contacts were prevented by housing all fish individually, male-biased mortality ceased in all four species. This outcome held across benign and challenging environmental conditions. Fitting demographic survival models revealed that increased baseline mortality was primarily responsible for a shorter male life span in social-housing conditions. The timing and rate of aging were not different between the sexes. No marker of functional aging we recorded in our study (lipofuscin accumulation, proliferative changes in kidney and liver) differed between males and females, despite their previously confirmed association with functional aging in Nothobranchius killifish. We show that sex differences in life span and aging in killifish are driven by a combination of social and environmental conditions, rather than differential functional aging. They are primarily linked to sexual selection but precipitated through multiple processes (predation, social interference). This demonstrates how sex-specific mortality varies among species even within an ecologically and evolutionary discrete lineage and explains how external factors mediate this difference.

Nothobranchius furzeri, the Turquoise Killifish: A Model of Age-Related Osteoporosis?

INTRODUCTION: Osteoporosis is a frequent age-related disease, which affects millions of people worldwide. Despite significant progress in the treatment of the disease, a high number of patients still are underdiagnosed and undertreated. Therefore, novel animal models for the investigation of the disease are necessary. Nothobranchius furzeri is the shortest-lived vertebrate (with a lifespan of 3-7 months) that can be kept in captivity. Although it is an established model for aging research, studies on bone are lacking. The aim of this study was therefore to characterize N. furzeri as a potential model for age-related osteoporosis. MATERIALS AND METHODS: Bone properties of aging N. furzeri were investigated in male and female fish of the Gona Re Zhou strain, which were between 8 and 20 weeks old. Micro-computed tomography (Scanco Medical µCT35) was performed to determine the bone properties of the vertebral bodies. Bone structure and remodeling were investigated by different histological staining techniques and histomorphometry. The chemical composition of fish vertebrae and intervertebral discs was analyzed by Raman microspectroscopy. RESULTS: Osteoblasts, mono- and multinucleated osteoclasts but no osteocytes could be observed in the vertebral area of N. furzeri. Histomorphometric evaluations revealed a significant decrease of the number of osteoblasts/bone perimeter and for osteoid volume/bone volume (BV) a trend toward a decrease in old male N. furzeri. Comparing male and female fish, males showed higher BV densities and cortical thickness. The relative values of the bone volume density of 20-week-old male N. furzeri were significantly lower than 10-week-old ones. The mineral to matrix ratio increased with age in male and female fish. In the intervertebral discs, proteoglycans in relation to the organic matrix were significantly lower in older female fish. CONCLUSION: Our finding of a lack of osteocytes is in agreement with the fact that N. furzeri belongs to the evolutionarily advanced teleost fish. Furthermore, not only age-specific but also sex-specific differences were visible in the bone properties of N. furzeri, which can be taken into consideration for the study of gender aspects of age-related musculoskeletal diseases.

Multifaceted Nothobranchius.

Annual killifish of the genus Nothobranchius are seeing a rapid increase in scientific interest over the years. A variety of aspects surrounding the egg-laying Cyprinodontiformes is being extensively studied, including their aging. Inhabiting drying water bodies of Africa rarely allows survival through more than one rainy season for the Nothobranchius populations. Therefore, there is no lifespan-related bias in natural selection, which has ultimately led to the decreased efficiency of DNA repair system. Aging of the Nothobranchius species is studied both under normal conditions and under the influence of potential geroprotectors, as well as genetic modifications. Most biogerontological studies are conducted using the species Nothobranchius furzeri (GRZ isolate), which has a lifespan of 3 to 7 months. However, the list of model species of Nothobranchius is considerably wider, and the range of advanced research areas with their participation extends far beyond gerontology. This review summarizes the most interesting and promising topics developing in the studies of the fish of Nothobranchius genus. Both classical studies related to lifespan control and rather new ones are discussed, including mechanisms of diapause, challenges of systematics and phylogeny, evolution of sex determination mechanisms, changes in chromosome count, occurrence of multiple repeated DNA sequences in the genome, cognitive and behavioral features and social stratification, as well as methodological difficulties in working with Nothobranchius.

The Effect of Meclofenoxate on the Transcriptome of Aging Brain of Nothobranchius guentheri Annual Killifish.

Annual fish of the genus Nothobranchius are promising models for aging research. Nothobranchius reproduces typical aspects of vertebrate aging, including hallmarks of brain aging. Meclofenoxate (MF) is a well-known compound that can enhance cognitive performance. The drug is prescribed for asthenic conditions, trauma, and vascular diseases of the brain. It is believed that MF is able to delay age-dependent changes in the human brain. However, until now, there has been no study of the MF effect on the brain transcriptome. In the present work, we performed an RNA-Seq study of brain tissues from aged Nothobranchius guentheri, which were almost lifetime administered with MF, as well as young and aged control fish. As expected, in response to MF, we revealed significant overexpression of neuron-specific genes including genes involved in synaptic activity and plasticity, neurotransmitter secretion, and neuron projection. The effect was more pronounced in female fish. In this aspect, MF alleviated age-dependent decreased expression of genes involved in neuronal activity. In both treated and untreated animals, we observed strong aging-associated overexpression of immune and inflammatory response genes. MF treatment did not prevent this effect, and moreover, some of these genes tended to be slightly upregulated under MF treatment. Additionally, we noticed upregulation of some genes associated with aging and cellular senescence, including isoforms of putative vascular cell adhesion molecule 1 (VCAM1), protein O-GlcNAcase (OGA), protein kinase C alpha type (KPCA), prolow-density lipoprotein receptor-related protein 1 (LRP1). Noteworthy, MF treatment was also associated with the elevated transcription of transposons, which are highly abundant in the N. guentheri genome. In conclusion, MF compensates for the age-dependent downregulation of neuronal activity genes, but its effect on aging brain transcriptome still cannot be considered unambiguously positive.

Neuronal Phenotype of col4a1 and col25a1: An Intriguing Hypothesis in Vertebrates Brain Aging.

Collagens are the most abundant proteins in vertebrates and constitute the major components of the extracellular matrix. Collagens play an important and multifaceted role in the development and functioning of the nervous system and undergo structural remodeling and quantitative modifications during aging. Here, we investigated the age-dependent regulation of col4a1 and col25a1 in the brain of the short-lived vertebrate Nothobranchius furzeri, a powerful model organism for aging research due to its natural fast-aging process and further characterized typical hallmarks of brain aging in this species. We showed that col4a1 and col25a1 are relatively well conserved during vertebrate evolution, and their expression significantly increases in the brain of N. furzeri upon aging. Noteworthy, we report that both col4a1 and col25a1 are expressed in cells with a neuronal phenotype, unlike what has already been documented in mammalian brain, in which only col25a1 is considered a neuronal marker, whereas col4a1 seems to be expressed only in endothelial cells. Overall, our findings encourage further investigation on the role of col4a1 and col25a1 in the biology of the vertebrate brain as well as the onset of aging and neurodegenerative diseases.

Genomic fingerprints of palaeogeographic history: The tempo and mode of rift tectonics across tropical Africa has shaped the diversification of the killifish genus Nothobranchius (Teleostei: Cyprinodontiformes).

This paper reports a phylogeny of the African killifishes (Genus Nothobranchius, Order Cyprinodontiformes) informed by five genetic markers (three nuclear, two mitochondrial) of 80 taxa (seven undescribed and 73 of the 92 recognized species). These short-lived annual fishes occupy seasonally wet habitats in central and eastern Africa, and their distribution coincides largely with the East African Rift System (EARS). The fossil dates of sister clades used to constrain a chronometric tree of all sampled Nothobranchius recovered the origin of the genus at ~13.27 Mya. It was followed by the radiations of six principal clades through the Neogene. An ancestral area estimation tested competing biogeographical hypotheses to constrain the ancestral origin of the genus to the Nilo-Sudan Ecoregion, which seeded a mid-Miocene dispersal event into the Coastal ecoregion, followed closely (~10 Mya) by dispersals southward across the Mozambique coastal plain into the Limpopo Ecoregion. Extending westwards across the Tanzanian plateau, a pulse of radiations through the Pliocene were associated with dispersals and fragmentation of wetlands across the Kalahari and Uganda Ecoregions. We interpret this congruence of drainage rearrangements with dispersals and cladogenic events of Nothobranchius to reflect congruent responses to recurrent uplift and rifting. The coevolution of these freshwater fishes and wetlands is attributed to ultimate control by tectonics, as the EARS extended southwards during the Neogene. Geobiological consilience of the combined evidence supports a tectonic hypothesis for the evolution of Nothobranchius.

Reproductive senescence in a short-lived fish.

Reproductive senescence is an age-associated decline in reproductive performance, which often arises as a trade-off between current and future reproduction. Given that mortality is inevitable, increased allocation into current reproduction is favoured despite costs paid later in life. This assumption is violated in organisms with post-maturity growth whose reproductive output increases long after maturity. While reproductive senescence is frequently studied in animals with determinate growth at maturity, such as insects or mammals, we have very limited understanding of reproductive senescence in organisms with an extensive post-maturity growth period. The fact that many post-maturity growers experience strong adult mortality leads to conflicting expectations for reproductive senescence. The aim of this study was to investigate how co-occurrence of rapid life history and post-maturity growth mould reproductive senescence in a short-lived killifish, Nothobranchius furzeri, using longitudinal data on laboratory and wild-type populations. We followed the individual fecundity, fertility and fertilization of 132 singly housed fish from the perspectives of chronological and biological age. At the onset of senescence, the sex-specific contribution to decrease in fertilization capacity was investigated. Allocation trade-offs were estimated through the association between reproductive parameters and life span, and between early-life and late-life fecundity. We demonstrate that female fecundity increased steadily after maturity and reproductive senescence occurred long after the growth asymptote. The prime age for fecundity coincided with 50% female survival and consequent decline in fecundity implies an association with somatic deterioration. Reproductive senescence in fertilization rate was stronger in females than in males. Females with high early fecundity experienced a long life span and high late-life fecundity, discounting the role of allocation trade-offs in reproductive senescence. The present study reports a clear case of reproductive senescence in a fish with a long post-maturation growth period, unusually rapid development and short life span. The onset of reproductive senescence was postponed compared to animals that cease growing at sexual maturity. Fish and other animals with post-maturity growth have long been considered insusceptible to ageing but this conclusion may be related to the previous lack of longitudinal data rather than to the absence of reproductive senescence in such organisms.

Mass spectrometric characterization of cyclic dinucleotides (CDNs) in vivo.

Cyclic dinucleotides (CDNs) are key secondary messenger molecules produced by cyclic dinucleotide synthases that trigger various cellular signaling cascades from bacteria to vertebrates. In mammals, cyclic GMP-AMP synthase (cGAS) has been shown to bind to intracellular DNA and catalyze the production of the dinucleotide 2'3' cGAMP, which signals downstream effectors to regulate immune function, interferon signaling, and the antiviral response. Despite the importance of CDNs, sensitive and accurate methods to measure their levels in vivo are lacking. Here, we report a novel LC-MS/MS method to quantify CDNs in vivo. We characterized the mass spectrometric behavior of four different biologically relevant CDNs (c-di-AMP, c-di-GMP, 3'3' cGAMP, 2'3' cGAMP) and provided a means of visually representing fragmentation resulting from collision-induced dissociation at different energies using collision energy breakdown graphs. We then validated the method and quantified CDNs in two in vivo systems, the bacteria Escherichia coli OP50 and the killifish Nothobranchius furzeri. We found that optimization of LC-MS/MS parameters is crucial to sensitivity and accuracy. These technical advances should help illuminate physiological and pathological roles of these CDNs in in vivo settings. Graphical abstract.

Localization of Neurotrophin Specific Trk Receptors in Mechanosensory Systems of Killifish (Nothobranchius guentheri).

Neurotrophins (NTs) and their signal-transducing Trk receptors play a crucial role in the development and maintenance of specific neuronal subpopulations in nervous and sensory systems. NTs are supposed to regulate two sensory systems in fish, the inner ear and the lateral line system (LLS). The latter is one of the major mechanosensory systems in fish. Considering that annual fishes of the genus Nothobranchius, with their short life expectancy, have become a suitable model for aging studies and that the occurrence and distribution of neurotrophin Trk receptors have never been investigated in the inner ear and LLS of killifish (Nothobranchius guentheri), our study aimed to investigate the localization of neurotrophin-specific Trk receptors in mechanosensory systems of N. guentheri. For histological and immunohistochemical analysis, adult specimens of N. guentheri were processed using antibodies against Trk receptors and S100 protein. An intense immunoreaction for TrkA and TrkC was found in the sensory cells of the inner ear as well as in the hair cells of LLS. Moreover, also the neurons localized in the acoustic ganglia displayed a specific immunoreaction for all Trk receptors (TrkA, B, and C) analyzed. Taken together, our results demonstrate, for the first time, that neurotrophins and their specific receptors could play a pivotal role in the biology of the sensory cells of the inner ear and LLS of N. guentheri and might also be involved in the hair cells regeneration process in normal and aged conditions.

The turquoise killifish: a genetically tractable model for the study of aging.

Lifespan is a remarkably diverse trait in nature, ranging from just hours in adult mayflies to hundreds of years in the Greenland shark and quahog clam. Great disparities in lifespan are often observed even among somewhat closely related species; for example, in the laboratory, wild-derived strains of the common house mouse have a maximum observed lifespan of approximately 6 years, while a similarly sized rodent, the naked mole rat, can live for over 30 years. Comparative biology of aging across the tree of life provides a tremendous opportunity for understanding the molecular and genetic basis underlying lifespan and aging. However, a lack of molecular and laboratory tools has limited the ability of researchers to take full advantage of the incredible diversity of aging phenotypes in nature. Recent developments in genomic technology have made it increasingly possible to study non-canonical model organisms for aging. One promising new genetic model organism amenable to a range of experimental interventions is the turquoise killifish (Nothobranchius furzeri). This fish species has a naturally short lifespan and undergoes a wide range of aging-related transformations. These fish have a fully sequenced genome and transcriptome, and killifish embryos are accessible to transgenesis and genome editing. Furthermore, different killifish species and populations show striking differences in lifespan, providing the opportunity for comparative analysis of aging. This Review introduces the natural life history of the turquoise killifish, its emerging applicability as an aging model system, the genetic tools that have been developed to study aging for this species and a summary of recent studies facilitated by these new tools.

Spontaneous adenocarcinoma of the gas gland in Nothobranchius fishes.

Nothobranchius fishes (Cyprinodontiformes), known for their genetically encoded extremely compressed lifespan, are considered an excellent vertebrate model for the research of aging. Unlike the rapid accumulation of data concerning their biology, ecology and genome, knowledge of their age-related diseases, including tumours, is still very limited. This Note reports spontaneous neoplastic lesions in the swim bladder gas glands of Nothobranchius furzeri, N. kadleci and N. orthonotus. Based on light and transmission electron microscopy, the neoplastic proliferation of gas gland cells was classified as adenocarcinoma. There was a concurrent proliferation of haemopoietic cells in the kidney interstitium in all individuals diagnosed with this type of primary neoplasia.

Egg size does not universally predict embryonic resources and hatchling size across annual killifish species.

Egg size has a crucial impact on the reproductive success of a mother and the performance of her offspring. It is therefore reasonable to employ egg size as a proxy for egg content when studying variation in offspring performance. Here, we tested species differences in allometries of several egg content parameters with egg area. We measured individual eggs in five species of annual killifish (Cyprinodontiformes), a group of fish where egg banks permit population survival over dry season. Apart from comparing allometric scaling exponents, amounts and compositions of egg components across the different species, we assessed the explanatory power of egg area for egg wet and dry weight and for hatchling size. We found notable species-specific allometries between egg area and the other egg parameters (egg dry weight and water content, elemental composition and triglyceride content). Across species, egg area predicted egg wet weight with highest power. Within species, coefficients of determination were largest in A. elongatus, a large piscivorous species with large eggs. Our study shows that systematically using egg area as a proxy of egg content between different species can ignore relevant species-specific differences and mask within-species variability in egg content.

What have we learned on aging from omics studies?

Aging is a complex process. Transcriptomic studies of the last decade have identified genes and pathways that are regulated during aging in multiple species and organs. Yet, since a manifold of pathways are regulated and the amplitude of regulation is often small, reproducibility across studies is moderate and disentangling cause-consequence relationships has proven challenging. Here, we review a number of consistent findings in the light of more recent, longitudinal studies and of studies combining transcriptomics and proteomics that identified deregulation of protein biosynthetic pathways as an early event and likely driver of aging.

Dispersion/reaggregation in early development of annual killifishes: Phylogenetic distribution and evolutionary significance of a unique feature.

Annual killifishes are members of the Aplocheiloidea and live in ephemeral habitats that desiccate regularly during the dry season and refill during the rainy season. Populations of these fishes survive the dry season by producing drought-resistant diapausing eggs that are buried in the substrate. When the pool refills during the rainy season the juveniles hatch, grow rapidly and reproduce until the pool desiccates again during the next dry season. The association with such unpredictable habitats has led to the evolution to a variety of developmental adaptations such as a dispersed/reaggregation phase of the deep blastomeres, three possible diapause stages, extreme tolerance to high salinity and anoxia, an efficient DNA repair system and an extremely short life span. Here, we review the course of the dispersed/reaggregation phase, its evolution and phylogenetic distribution and diversity within the Aplocheiloidea. The phenomenon of blastomere dispersion/reaggregation in these fishes was first described in the 1960s and 70s. Blastomeres of most teleost fishes segregate into three groups that give rise to the enveloping cell layer, the yolk syncytial layer and the deep blastomeres that will form the embryo itself. When epiboly commences, the deep blastomeres form a more or less coherent cell sheet with a so called embryonic shield at it marginal zone marking the area where gastrulation takes place. In annual killifishes, the deep blastomeres segregate when epiboly starts and disperse when epiboly commences. After epiboly has been completed, the deep blastomeres are randomly distributed and migrate all over the enveloping cell layer. After several days they start to reaggregate and form the actual embryo that starts gastrulation. The evolutionary origin and mechanism behind this peculiar developmental pathway have puzzled developmental biologists for almost 50 years. However, several of these annual killifishes (Nothobranchius furzeri, Austrofundulus limnaeus, Austrolebias charrua and Austrolebias bellottii) have become model organisms in studies on developmental physiology, aging and stress tolerance. This has led to the establishment of modern genetic techniques such as transgenesis and cell fate mapping that are now used to tackle questions about the origin and mechanisms behind the dispersal/reaggregation phase.

Live fast, diversify non-adaptively: evolutionary diversification of exceptionally short-lived annual killifishes.

BACKGROUND: Adaptive radiations are triggered by ecological opportunity - the access to novel niche domains with abundant available resources that facilitate the formation of new ecologically divergent species. Therefore, as new species saturate niche space, clades experience a diversity-dependent slowdown of diversification over time. At the other extreme of the radiation continuum, non-adaptively radiating lineages undergo diversification with minimal niche differentiation when 'spatial opportunity' (i.e. areas with suitable 'ancestral' ecological conditions) is available. Traditionally, most research has focused on adaptive radiations, while empirical studies on non-adaptive radiations remain lagging behind. A prolific clade of African fish with extremely short lifespan (Nothobranchius killifish), show the key evolutionary features of a candidate non-adaptive radiation - primarily allopatric species with minimal niche and phenotypic divergence. Here, we test the hypothesis that Nothobranchius killifish have non-adaptively diversified. We employ phylogenetic modelling to investigate the tempo and mode of macroevolutionary diversification of these organisms. RESULTS: Nothobranchius diversification has proceeded with minor niche differentiation and minimal morphological disparity among allopatric species. Additionally, we failed to identify evidence for a role of body size or biogeography in influencing diversification rates. Diversification has been homogeneous within this genus, with the only hotspot of species-richness not resulting from rapid diversification. However, species in sympatry show higher disparity, which may have been caused by character displacement among coexisting species. CONCLUSIONS: Nothobranchius killifish have proliferated following the tempo and mode of a non-adaptive radiation. Our study confirms that this exceptionally short-lived group have diversified with minimal divergent niche adaptation, while one group of coexisting species seems to have facilitated spatial overlap among these taxa via the evolution of ecological character displacement.

Administration of rGDF11 retards the aging process in male mice via action of anti-oxidant system.

One of the most studied and widely accepted conjectures of aging process is the oxidative stress theory. Current studies have generated disputes on the effects of GDF11 and GDF8, a closely related member of GDF11, on rejuvenation and anti-aging properties. In this study, we first demonstrated that when recombinant GDF8 (rGDF8) and GDF11 (rGDF11) of the fish Nothobranchius guentheri were injected into 20-month-old male mice, their serum GDF8 and GDF11 levels were clearly increased. We also showed that injection of rGDF8 and rGDF11 had little influences on the body weight and serological parameters of the mice, indicating their general condition and physiology were not affected. Based on these findings, we started to test the effects of administration of piscine rGDF11 and rGDF8 on the aging process of male mice and to explore the underlying mechanisms. It was found that rGDF11 was able to reduce the levels of AGEs, protein oxidation and lipid peroxidation, and to slow down the accumulation of age-related histological markers, while rGDF8 was not. Moreover, rGDF11 significantly prevented the decrease in CAT, GPX and SOD activities, but rGDF8 did not. Collectively, these results suggest that it is GDF11 but not GDF8 that can exert rejuvenation and anti-aging activities via the action of antioxidant system. It is also the first report that shows the activity of GDF11 is not species-specific, implicating potential usefulness of piscine GDF11 in prolonging the lifespan of the elderly.