Correlates and Consequences of Dominance in a Social Rodent

In harem‐polygynous societies, body condition is often correlated with dominance rank. However, the consequences of dominance are less clear. High‐ranking males do not inevitably have the highest reproductive success, especially in systems where females mate with multiple males. In such societies, we expect male reproductive success to be more highly skewed than female reproductive success, but reproductive skew in females can still arise from rankings established within matrilineal societies. Dominance can also impact life‐history decisions by influencing dispersal patterns in yearlings. To better understand the function of dominance in harem‐polygynous societies, we studied the causes and consequences of dominance in yellow‐bellied marmots (Marmota flaviventris), a social rodent with skewed male reproductive success and female reproductive suppression. Specifically, we examined body condition as a predictor and the probability of breeding, number of offspring, and dispersal as outcomes of dominance. Additionally, we looked at variation in dominance between males and females and adults and yearlings, because marmots can engage in distinct interactions depending on the type of individuals involved. We found that marmots in better body condition have higher dominance rank than those in poorer condition. In addition, adults are dominant over yearlings. Within yearlings, dominance does not influence dispersal, but those in better body condition are less likely to disperse. Within all adults, individuals in better condition produce more offspring per year. Within adult males, more dominant males have greater reproductive success. Despite previous evidence of reproductive suppression in females, we found no effects of dominance rank on female reproductive success in the current study. The function of dominance in female marmots remains enigmatic.     

Is the combination of topsoil replacement and inoculation with plant material an effective tool for the restoration of threatened sandy grassland?

Question: Is it possible to restore dry calcareous inland sand ecosystems with their characteristic plant community structure within a 4‐yr period by means of combined abiotic–biotic techniques (topsoil replacement, inoculation with raked/mown plant material from target areas)? Location: Upper Rhine valley, Germany. Methods: Two 4‐year experiments were carried out on former arable land, each in the proximity of a reference area bearing a similar complex of threatened sandy grasslands (experiment 1: fine‐scale; experiment 2: landscape scale). In both experiments we used nutrient‐poor deep‐sand substrate (abiotic approach), raked/mown inoculation material from target areas and grazing as management tool (biotic and management approach). The vegetation of the restoration and donor areas was sampled once a year and analysed by non‐metric multidimensional scaling (NMDS) ordination and target‐species ratios. Mixed linear models were calculated to determine effects of grazing (experiment 1) and year (both experiments). Results: NMDS revealed a continuous development of the restored sites towards the corresponding donor sites. Similarly, target‐species ratios of the restored sites tended towards the ratios of the donor sites. To date, grazing effects have mainly been structural: reduction of a carpet‐forming pleurocarpous moss species and of litter. In addition, cover of target species in relation to total plant cover was significantly enhanced by grazing in the last two study years. Conclusions: The combination of nutrient‐poor substrate, inoculation with raked/mown plant material and grazing proved to be a very effective restoration method for dry base‐rich sand ecosystems. After 4 yr the restored plant communities serve as well‐developed parts of a habitat network.     

The stabilisation of purified, reconstituted P-glycoprotein by freeze drying with disaccharides

The drug efflux pump P-glycoprotein (P-gp) (ABCB1) confers multidrug resistance, a major cause of failure in the chemotherapy of tumours, exacerbated by a shortage of potent and selective inhibitors. A high throughput assay using purified P-gp to screen and characterise potential inhibitors would greatly accelerate their development. However, long-term stability of purified reconstituted ABCB1 can only be reliably achieved with storage at −80 °C. For example, at 20 °C, the activity of ABCB1 was abrogated with a half-life of <1 day. The aim of this investigation was to stabilise purified, reconstituted ABCB1 to enable storage at higher temperatures and thereby enable design of a high throughput assay system. The ABCB1 purification procedure was optimised to allow successful freeze drying by substitution of glycerol with the disaccharides trehalose or maltose. Addition of disaccharides resulted in ATPase activity being retained immediately following lyophilisation with no significant difference between the two disaccharides. However, during storage trehalose preserved ATPase activity for several months regardless of the temperature (e.g. 60% retention at 150 days), whereas ATPase activity in maltose purified P-gp was affected by both storage time and temperature. The data provide an effective mechanism for the production of resilient purified, reconstituted ABCB1.     

The enemy release hypothesis as a hierarchy of hypotheses

In ecology, a hypothesis is usually not discarded if a few studies reject it, as long as there are other studies supporting it. How to assess the usefulness of ecological hypotheses is therefore not straightforward. Using the enemy release hypothesis as an example, we show how creating a hierarchy of hypotheses (HoH) can help reviewing and evaluating evidence for and against an ecological hypothesis. In a HoH, a broad, overarching hypothesis branches into more specific and better testable sub‐hypotheses. The enemy release hypothesis is a major hypothesis in invasion ecology and posits that the absence of enemies in the exotic range of an alien species is a cause of its invasion success. Based on a systematic review of empirical tests of this hypothesis, we divided it into sub‐hypotheses, differentiating among 1) indicators for enemy release, 2) types of comparisons, and 3) types of enemies. We identified 176 empirical tests and weighted each test according to the number of alien species studied and the research method (experimental vs observational, field vs enclosure vs laboratory). For the broadly formulated enemy release hypothesis, we found nearly as much supporting (36%) as questioning evidence (43%). At the sub‐hypotheses level, however, we found that some sub‐hypotheses are strongly supported by empirical evidence, whereas others receive hardly any support. These differences are further emphasized for some types of habitat and focal taxonomic groups. Our findings suggest that several specific formulations (i.e. sub‐hypotheses) of the broad enemy release hypothesis are useful, whereas other formulations should be viewed more critically. In general, the approach outlined here can help evaluate major ecological hypotheses and their specific sub‐hypotheses. Our study also highlights the need for a scientific debate on how much supporting evidence is sufficient to consider an ecological hypothesis to be useful.     

Alginate Microspheres Containing Temperature Sensitive Liposomes (TSL) for MR-Guided Embolization and Triggered Release of Doxorubicin

Objective

The objective of this study was to develop and characterize alginate microspheres suitable for embolization with on-demand triggered doxorubicin (DOX) release and whereby the microspheres as well as the drug releasing process can be visualized in vivo using MRI.

Methods and Findings

For this purpose, barium crosslinked alginate microspheres were loaded with temperature sensitive liposomes (TSL/TSL-Ba-ms), which release their payload upon mild hyperthermia. These TSL contained DOX and [Gd(HPDO3A)(H₂O)], a T₁ MRI contrast agent, for real time visualization of the release. Empty alginate microspheres crosslinked with holmium ions (T₂* MRI contrast agent, Ho-ms) were mixed with TSL-Ba-ms to allow microsphere visualization. TSL-Ba-ms and Ho-ms were prepared with a homemade spray device and sized by sieving. Encapsulation of TSL in barium crosslinked microspheres changed the triggered release properties only slightly: 95% of the loaded DOX was released from free TSL vs. 86% release for TSL-Ba-ms within 30 seconds in 50% FBS at 42°C. TSL-Ba-ms (76 ± 41 μm) and Ho-ms (64 ± 29 μm) had a comparable size, which most likely will result in a similar in vivo tissue distribution after an i.v. co-injection and therefore Ho-ms can be used as tracer for the TSL-Ba-ms. MR imaging of a TSL-Ba-ms and Ho-ms mixture (ratio 95:5) before and after hyperthermia allowed in vitro and in vivo visualization of microsphere deposition (T₂*-weighted images) as well as temperature-triggered release (T₁-weighted images). The [Gd(HPDO3A)(H₂O)] release and clusters of microspheres containing holmium ions were visualized in a VX₂ tumor model in a rabbit using MRI.

Conclusions

In conclusion, these TSL-Ba-ms and Ho-ms are promising systems for real-time, MR-guided embolization and triggered release of drugs in vivo.     

Sustained release of an anti-glaucoma drug: demonstration of efficacy of a liposomal formulation in the rabbit eye

Topical medication remains the first line treatment of glaucoma; however, sustained ocular drug delivery via topical administration is difficult to achieve. Most drugs have poor penetration due to the multiple physiological barriers of the eye and are rapidly cleared if applied topically. Currently, daily topical administration for lowering the intra-ocular pressure (IOP), has many limitations, such as poor patient compliance and ocular allergy from repeated drug administration. Poor compliance leads to suboptimal control of IOP and disease progression with eventual blindness. The delivery of drugs in a sustained manner could provide the patient with a more attractive alternative by providing optimal therapeutic dosing, with minimal local toxicity and inconvenience. To investigate this, we incorporated latanoprost into LUVs (large unilamellar vesicles) derived from the liposome of DPPC (di-palmitoyl-phosphatidyl-choline) by the film hydration technique. Relatively high amounts of drug could be incorporated into this vesicle, and the drug resides predominantly in the bilayer. Vesicle stability monitored by size measurement and DSC (differential scanning calorimetry) analysis showed that formulations with a drug/lipid mole ratio of about 10% have good physical stability during storage and release. This formulation demonstrated sustained release of latanoprost in vitro, and then tested for efficacy in 23 rabbits. Subconjunctival injection and topical eye drop administration of the latanoprost/liposomal formulation were compared with conventional daily administration of latanoprost eye drops. The IOP lowering effect with a single subconjunctival injection was shown to be sustained for up to 50 days, and the extent of IOP lowering was comparable to daily eye drop administration. Toxicity and localized inflammation were not observed in any treatment groups. We believe that this is the first demonstration, in vivo, of sustained delivery to the anterior segment of the eye that is safe and efficacious for 50 days.     

Conditioned pain modulation is associated with common polymorphisms in the serotonin transporter gene

Background

Variation in the serotonin transporter (5-HTT) gene (SLC6A4) has been shown to influence a wide range of affective processes. Low 5-HTT gene-expression has also been suggested to increase the risk of chronic pain. Conditioned pain modulation (CPM) - i.e. ‘pain inhibits pain’ - is impaired in chronic pain states and, reciprocally, aberrations of CPM may predict the development of chronic pain. Therefore we hypothesized that a common variation in the SLC6A4 is associated with inter-individual variation in CPM. Forty-five healthy subjects recruited on the basis of tri-allelic 5-HTTLPR genotype, with inferred high or low 5-HTT-expression, were included in a double-blind study. A submaximal-effort tourniquet test was used to provide a standardized degree of conditioning ischemic pain. Individualized noxious heat and pressure pain thresholds (PPTs) were used as subjective test-modalities and the nociceptive flexion reflex (NFR) was used to provide an objective neurophysiological window into spinal processing.

Results

The low, as compared to the high, 5-HTT-expressing group exhibited significantly reduced CPM-mediated pain inhibition for PPTs (p = 0.02) and heat-pain (p = 0.02). The CPM-mediated inhibition of the NFR, gauged by increases in NFR-threshold, did not differ significantly between groups (p = 0.75). Inhibition of PPTs and heat-pain were correlated (Spearman’s rho = 0.35, p = 0.02), whereas the NFR-threshold increase was not significantly correlated with degree of inhibition of these subjectively reported modalities.

Conclusions

Our results demonstrate the involvement of the tri-allelic 5-HTTLPR genotype in explaining clinically relevant inter-individual differences in pain perception and regulation. Our results also illustrate that shifts in NFR-thresholds do not necessarily correlate to the modulation of experienced pain. We discuss various possible mechanisms underlying these findings and suggest a role of regulation of 5-HT receptors along the neuraxis as a function of differential 5-HTT-expression.     

By Regulating Mitochondrial Ca2+-Uptake UCP2 Modulates Intracellular Ca2+

Introduction

The possible role of UCP2 in modulating mitochondrial Ca²⁺-uptake (mCa²⁺-uptake) via the mitochondrial calcium uniporter (MCU) is highly controversial.

Methods

Thus, we analyzed mCa²⁺-uptake in isolated cardiac mitochondria, MCU single-channel activity in cardiac mitoplasts, dual Ca²⁺-transients from mitochondrial ((Ca²⁺)m) and intracellular compartment ((Ca²⁺)c) in the whole-cell configuration in cardiomyocytes of wild-type (WT) and UCP2^-/- mice.

Results

Isolated mitochondria showed a Ru360 sensitive mCa²⁺-uptake, which was significantly decreased in UCP2^-/- (229.4±30.8 FU vs. 146.3±23.4 FU, P<0.05). Single-channel registrations confirmed a Ru360 sensitive voltage-gated Ca²⁺-channel in mitoplasts, i.e. mCa1, showing a reduced single-channel activity in UCP2^-/- (Po,total: 0.34±0.05% vs. 0.07±0.01%, P<0.05). In UCP2^-/- cardiomyocytes (Ca²⁺)m was decreased (0.050±0.009 FU vs. 0.021±0.005 FU, P<0.05) while (Ca²⁺)c was unchanged (0.032±0.002 FU vs. 0.028±0.004 FU, P>0.05) and transsarcolemmal Ca²⁺-influx was inhibited suggesting a possible compensatory mechanism. Additionally, we observed an inhibitory effect of ATP on mCa²⁺-uptake in WT mitoplasts and (Ca²⁺)m of cardiomyocytes leading to an increase of (Ca²⁺)c while no ATP dependent effect was observed in UCP2^-/-.

Conclusion

Our results indicate regulatory effects of UCP2 on mCa²⁺-uptake. Furthermore, we propose, that previously described inhibitory effects on MCU by ATP may be mediated via UCP2 resulting in changes of excitation contraction coupling.     

Role of nonstructural proteins 3A and 3B in host range and pathogenicity of foot-and-mouth disease virus

The genome of foot-and-mouth disease virus (FMDV) differs from that of other picornaviruses in that it encodes a larger 3A protein (>50% longer than poliovirus 3A), as well as three copies of protein 3B (also known as VPg). Previous studies have shown that a deletion of amino acids 93 to 102 of the 153-codon 3A protein is associated with an inability of a Taiwanese strain of FMDV (O/TAW/97) to cause disease in bovines. Recently, an Asian virus with a second 3A deletion (amino acids 133 to 143) has also been detected (N. J. Knowles et al., J. Virol. 75:1551-1556, 2001). Genetically engineered viruses harboring the amino acids 93 to 102 or 133 to 143 grew well in porcine cells but replicated poorly in bovine cells, whereas a genetically engineered derivative of the O/TAW/97 virus expressing a full-length 3A (strain A12) grew well in both cell types. Interestingly, a virus with a deletion spanning amino acid 93 to 144 also grew well in porcine cells and caused disease in swine. Further, genetically engineered viruses containing only a single copy of VPg were readily recovered with the full-length 3A, the deleted 3A (amino acids 93 to 102), or the "super" deleted forms of 3A (missing amino acids 93 to 144). All of the single-VPg viruses were attenuated in porcine cells and replicated poorly in bovine cells. The single-VPg viruses produced a mild disease in swine, indicating that the VPg copy number is an important determinant of host range and virulence. The association of VPg copy number with increased virulence in vivo may help to explain why all naturally occurring FMDVs have retained three copies of VPg.