Saikosaponin-d,a novel SERCA inhibitor,induces autophagic cell death in apoptosis-defective cells

Autophagy is an important cellular process that controls cells in a normal homeostatic state by recycling nutrients to maintain cellular energy levels for cell survival via the turnover of proteins and damaged organelles. However, persistent activation of autophagy can lead to excessive depletion of cellular organelles and essential proteins, leading to caspase-independent autophagic cell death. As such, inducing cell death through this autophagic mechanism could be an alternative approach to the treatment of cancers. Recently, we have identified a novel autophagic inducer, saikosaponin-d (Ssd), from a medicinal plant that induces autophagy in various types of cancer cells through the formation of autophagosomes as measured by GFP-LC3 puncta formation. By computational virtual docking analysis, biochemical assays and advanced live-cell imaging techniques, Ssd was shown to increase cytosolic calcium level via direct inhibition of sarcoplasmic/endoplasmic reticulum Ca²⁺ ATPase pump, leading to autophagy induction through the activation of the Ca²⁺/calmodulin-dependent kinase kinase–AMP-activated protein kinase–mammalian target of rapamycin pathway. In addition, Ssd treatment causes the disruption of calcium homeostasis, which induces endoplasmic reticulum stress as well as the unfolded protein responses pathway. Ssd also proved to be a potent cytotoxic agent in apoptosis-defective or apoptosis-resistant mouse embryonic fibroblast cells, which either lack caspases 3, 7 or 8 or had the Bax-Bak double knockout. These results provide a detailed understanding of the mechanism of action of Ssd, as a novel autophagic inducer, which has the potential of being developed into an anti-cancer agent for targeting apoptosis-resistant cancer cells.     

Recent experience modulates forebrain gene-expression in response to mate-choice cues in European starlings

Mate-choice decisions can be experience dependent, but we know little about how the brain processes stimuli that release such decisions. Female European starlings (Sturnus vulgaris) prefer males with long-bout songs over males with short-bout songs, and show higher expression of the immediate early gene (IEG) ZENK in the auditory forebrain when exposed to long-bout songs than when exposed to short-bout songs. We exposed female starlings to a short-day photoperiod for one of three durations and then, on an increased photophase, exposed them to one week of long-bout or short-bout song experience. We then examined their IEG response to novel long-bout versus novel short-bout songs by quantifying ZENK protein in two song-processing areas: the caudo-medial hyperstriatum ventrale and the caudo-medial neostriatum. ZENK expression in both areas increased with tenure on short-day photoperiods, suggesting that short days sensitize females to song. The ZENK response bias toward long-bout songs was greater in females with long-bout experience than in females with short-bout experience, indicating that the forebrain response bias toward a preferred trait depends on recent experience with that category of trait. This surprising level of neuroplasticity is immediately relevant to the natural history and fitness of the organism, and may underlie a mechanism for optimizing mate-choice criteria amidst locally variable distributions of secondary sexual characteristics.     

The role of prolactin in the regulation of clutch size and onset of incubation behavior in the American kestrel

In most bird species, the timing of incubation onset may influence the degree of hatching asynchrony, which, together with variation in clutch size, affects reproductive success. In some domesticated species that usually show no hatching asynchrony, plasma prolactin concentrations in females rise with the onset of incubation and the end of laying, and this rise enhances incubation behavior and may terminate laying. To investigate whether a rise in prolactin during laying is involved in the regulation of clutch size and incubation onset in a species with hatching asynchrony, we measured plasma concentrations of immunoreactive prolactin (ir-prolactin) in laying American kestrels, Falco sparverius, and quantified clutch size and incubation behavior. In a separate study, we administered one of three concentrations of ovine prolactin (o-prolactin) via osmotic pumps implanted in females when egg 2 of a clutch was laid. ir-Prolactin concentrations during laying were higher in small than in large clutches and increased in parallel with the development of incubation behavior. o-Prolactin treatment enhanced incubation behavior, but did not affect clutch size, possibly because the manipulation was performed after clutch size had already been determined. Consistent with studies on domesticated species that show synchronous hatching, our results indicate that rising prolactin during laying enhances the expression of incubation behavior in a species that shows hatching asynchrony. Further studies are necessary to determine whether the relationship between prolactin and clutch size in the American kestrel is one of causation or of mere association.     

Removing the confound of time in investigating the regulation of serial behaviours: testosterone, prolactin and the transition from sexual to parental activity in male American kestrels

Understanding the relationships among variables can be difficult if the variables covary with time. Using a simple, commonly used statistical procedure to control for the passage of time, we demonstrate that some hormonal changes traditionally assumed to be causally related to the transition from sexual to parental behaviour can be explained by seasonal changes in the hormones and the fact that parental behaviour follows sexual behaviour in the course of breeding. We quantified immunoreactive prolactin (ir-prolactin) and testosterone concentrations in laboratory-housed and free-living male American kestrels, Falco sparverius, over the course of the breeding season and during sexual and parental phases of reproduction. We found that ir-prolactin increased while testosterone decreased with the transition from sexual to parental behaviour. ir-Prolactin increased with date (i.e. seasonally), whereas testosterone peaked when the frequency of clutch initiation was greatest and then decreased with date thereafter. Controlling for the effects of date eliminated the change in ir-prolactin associated with the transition from sexual to parental behaviour, revealing that prolactin variation associated with this behavioural transition in male kestrels is due, in large part, to seasonal changes in hormone concentrations and the serial nature of these behavioural states. Controlling for the effects of date further revealed that the testosterone difference between sexual and parental males decreases seasonally. It is possible that other phenomena traditionally attributed to environmental, behavioural, social, or other variables also may be explained by the association between such variables and the passage of time.     

Independent effects of song quality and experience with photostimulation on expression of the immediate,early gene ZENK (EGR‐1) in the auditory telencephalon of female European starlings

Age influences behavioral decisions such as reproductive timing and effort. In photoperiodic species, such age effects may be mediated, in part, by the individual's age‐accrued experience with photostimulation. In female European starlings (Sturnus vulgaris) that do not differ in age, experimental manipulation of photostimulation experience (photoexperience) affects hypothalamic, pituitary, and gonadal activity associated with reproductive development. Does photoexperience also affect activity in forebrain regions involved in processing a social cue, the song of males, which can influence mate choice and reproductive timing in females? Female starlings prefer long songs over short songs in a mate‐choice context, and, like that in other songbird species, their auditory telencephalon plays a major role in processing these signals. We manipulated the photoexperience of female starlings, photostimulated them, briefly exposed them to either long or short songs, and quantified the expression of the immediate‐early gene ZENK (EGR‐1) in the caudomedial nidopallium as a measure of activity in the auditory telencephalon. Using an information theoretic approach, we found higher ZENK immunoreactivity in females with prior photostimulation experience than in females experiencing photostimulation for the first time. We also found that long songs elicited greater ZENK immunoreactivity than short songs did. We did not find an effect of the interaction between photoexperience and song length, suggesting that photoexperience does not affect forebrain ZENK‐responsiveness to song quality. Thus, photoexperience affects activity in an area of the forebrain that processes social signals, an effect that we hypothesize mediates, in part, the effects of age on reproductive decisions in photoperiodic songbirds. © 2009 Wiley Periodicals, Inc. Develop Neurobiol 2009     

Cytoplasmic filaments and cellular wound healing in amoeba proteus

The flexibility and self-healing properties of animal cell surface membranes are well known. These properties have been best exploited in various micrurgical studies on living cells (2, 3), especially in amoebae (7, 20). During nuclear transplantation in amoebae, the hole in the membrane through which a nucleus passes can have a diameter of 20-30 μm, and yet such holes are quickly sealed, although some cytoplasm usually escapes during the transfer. While enucleating amoebae in previous studies, we found that if a very small portion of a nucleus was pushed through the membrane and exposed to the external medium, the amoeba expelled such a nucleus on its own accord. When this happened, a new membrane appeared to form around the embedded portion of the nucleus and no visible loss of cytoplasm occurred during nuclear extrusion. In the present study, we examined amoebae that were at different stages of expelling partially exposed nuclei, to follow the sequence of events during the apparent new membrane formation. Unexpectedly, we found that a new membrane is not formed around the nucleus from inside but a hole is sealed primarily by a constriction of the existing membrane, and that cytoplasmic filaments are responsible for the prevention of the loss of cytoplasm.     

Which population level environmental factors are associated with asthma,rhinoconjunctivitis and eczema? Review of the ecological analyses of ISAAC Phase One

The International Study of Asthma and Allergies in Childhood (ISAAC) Phase One showed large worldwide variations in the prevalence of symptoms of asthma, rhinoconjunctivitis and eczema, up to 10 to 20 fold between countries. Ecological analyses were undertaken with ISAAC Phase One data to explore factors that may have contributed to these variations, and are summarised and reviewed here.In ISAAC Phase One the prevalence of symptoms in the past 12 months of asthma, rhinoconjunctivitis and eczema were estimated from studies in 463,801 children aged 13 - 14 years in 155 centres in 56 countries, and in 257,800 children aged 6-7 years in 91 centres in 38 countries. Ecological analyses were undertaken between symptom prevalence and the following: Gross National Product per capita (GNP), food intake, immunisation rates, tuberculosis notifications, climatic factors, tobacco consumption, pollen, antibiotic sales, paracetamol sales, and outdoor air pollution.Symptom prevalence of all three conditions was positively associated with GNP, trans fatty acids, paracetamol, and women smoking, and inversely associated with food of plant origin, pollen, immunisations, tuberculosis notifications, air pollution, and men smoking. The magnitude of these associations was small, but consistent in direction between conditions. There were mixed associations of climate and antibiotic sales with symptom prevalence.The potential causality of these associations warrant further investigation. Factors which prevent the development of these conditions, or where there is an absence of a positive correlation at a population level may be as important from the policy viewpoint as a focus on the positive risk factors. Interventions based on small associations may have the potential for a large public health benefit.     

Prior experience with photostimulation enhances photo-induced reproductive development in female European starlings: a possible basis for the age-related increase in avian reproductive performance

Reproductive performance in female birds improves with age, and this is generally attributed to experiences obtained during breeding. In temperate-zone species, experience with photostimulation during the first breeding year may prime the hypothalamo-pituitary-gonadal axis to respond to photic cues more rapidly or robustly in subsequent years. To test this idea, we captured 32 photorefractory juvenile (hence naive to photostimulation) female European starlings (Sturnus vulgaris) and held half of them (naive group) on a photoperiod of 8L:16D for 32 wk and the other half (experienced group) on 8L:16D for 12 wk, 16L:8D for 12 wk, and then 8L:16D for 8 wk. When we subsequently transferred all birds to 16L:8D, the increase in body mass, which may presage egg laying in the wild, was more robust in experienced than in naive females. Experienced females also showed a more robust elevation in plasma concentrations of the yolk-precursor protein vitellogenin, although naive females showed an initial rapid but transient rise in vitellogenin that we attribute to their extended exposure to short-day photoperiods prior to photostimulation. Finally, the photo-induced increase in diameter of the largest ovarian follicle, in plasma concentrations of luteinizing hormone, and in the number of septo-preoptic fibers relative to the number of cell bodies immunoreactive to GnRH was greater in experienced than in naive females. Thus, prior experience with photostimulation enhances some initial phases of photo-induced reproductive development and may explain, in part, why reproductive performance improves with age in temperate-zone birds.     

Yolk androgens reduce offspring survival

Females may favour some offspring over others by differential deposition of yolk hormones. In American kestrels (Falco sparverius), we found that yolks of eggs laid late in the sequence of a clutch had more testosterone (T) and androstenedione (A4) than yolks of first-laid eggs. To investigate the effects of these yolk androgens on nestling 'fitness', we injected both T and A4 into the yolks of first-laid eggs and compared their hatching time, nestling growth and nestling survival with those of first-laid eggs in which we injected vehicle as a control. Compared to controls, injection of T and A4 at a dose intended to increase their levels to those of later-laid eggs delayed hatching and reduced nestling growth and survival rates. Yolk androgen treatment of egg 1 had no effect on survival of siblings hatching from subsequently laid eggs. The adverse actions of yolk androgen treatment in the kestrel are in contrast to the favourable actions of yolk T treatment found previously in canaries (Serinus canaria). Additional studies are necessary in order to determine whether the deposition of yolk androgens is an adaptive form of parental favouritism or an adverse by-product of endocrine processes during egg formation. Despite its adaptive significance, such 'transgenerational' effects of steroid hormones may have helped to evolutionarily shape the hormonal mechanisms regulating reproduction.