Tissue response and wound healing after placement of two types of bioengineered grafts containing vital cells in submucosal maxillary pouches: an experimental pilot study in rabbits

PURPOSE: This pilot study evaluated the wound healing and tissue response after placement of two different skin substitutes in subgingival mucosal pouches in rabbits. MATERIALS AND METHODS: Four rabbits were selected to receive a commercially available skin substitute consisting of a collagen matrix with fibroblasts and an epithelial layer (test membrane 1) and a prototype device consisting of a collagen matrix with fibroblasts only (test membrane 2). In each rabbit, two horizontal incisions were made in the buccal alveolar mucosa of the maxilla bilaterally to create submucosal pouches. Three pouches in each animal were filled with either the test 1 or test 2 membranes, and one pouch was left without a membrane (sham-operated control). All rabbits were sacrificed after a healing period of 4 weeks, and histologic samples were prepared and examined. RESULTS: After a healing period of 1 month, both tested membranes were still visible in the sections. Test membrane 1 was still bilayered, contained inflammatory cells in its center, and was encapsulated by a thick fibrous tissue. Numerous ectopic calcifications were evident in the collagenous part of the membrane and in association with some basal epithelial cells. Test membrane 2 was also encapsulated in fibrous tissue, with inflammatory cells present only between the fibrous encapsulation and the remnants of the membrane. For test membrane 2, no calcifications were visible. CONCLUSIONS: Test membrane 1 seemed to be more resistant to degradation, but there was also a more pronounced inflammatory reaction in comparison to test membrane 2, especially in the vicinity of the keratinocytes. The significance of the ectopic calcifications, along with that of the resorption or degradation processes of both tested membranes, must be evaluated in future experimental studies, with different time points after implantation examine

No evidence for large differences in genomic methylation between wild and hatchery steelhead (Oncorhynchus mykiss)

When salmonid fish that have been raised in hatcheries spawn in the wild, they often produce fewer surviving adult offspring than wild fish. Recent data from steelhead (Oncorhynchus mykiss) in the Hood River (Oregon, USA) show that even one or two generations of hatchery culture can result in dramatic declines in fitness. Although intense domestication selection could cause such declines, it is worth considering alternative explanations. One possibility is heritable epigenetic changes induced by the hatchery environment. Here, we show, using methylation-sensitive amplified fragment length polymorphism, that hatchery and wild adult steelhead from the Hood River do not appear to differ substantially in overall levels of genomic methylation. Thus, although altered methylation of specific DNA sites or other epigenetic processes could still be important, the hatchery environment does not appear to cause a global hypo- or hypermethylation of the genome or create a large number of sites that are differentially methylated.

A novel steroidal antiandrogen targeting wild type and mutant androgen receptors

Prostate cancer (PCa) progression is enhanced by androgen and treatment with antiandrogens represents an alternative to castration. While patients initially respond favorably to androgen ablation therapy, most experience a relapse of the disease within 1-2 years by expressing androgen receptor (AR) mutants. Such mutations, indeed, promote unfavorable agonistic behavior from classical antagonists. Here, we have synthesized and screened 37 novel compounds derived from dihydrotestosterone (DHT), cyanolutamide and hydroxyflutamide. These derivatives were tested for their potential antagonistic activity using a luciferase reporter gene assay and binding properties were determined for wild type (WT) and mutant ARs (T877A, W741C, W741L, H874Y). In the absence and presence of antiandrogens, androgen dependent cellular proliferation and prostate specific antigen (PSA) expression were assayed in the prostate cancer cell line LNCaP by crystal violet, real time PCR and by Western blots. Also, cellular proliferation and PSA expression were assayed in 22Rv1. A novel compound RB346, derived from DHT, was found to be an antagonist for all tested AR forms, preventing DHT induced proliferation and PSA expression in LNCaP and 22Rv1 cells. RB346 displayed no agonistic activity, in contrast to the non-steroidal antiandrogen bicalutamide (Casodex) with unfavorable agonistic activity for W741L-AR. Additionally, RB346 has a slightly higher binding affinity for WT-AR, T877A-AR and H874Y-AR than bicalutamide. Thus, RB346 is the first potent steroidal antiandrogen with efficacy for WT and various AR mutants.

Outer skull landmark-based coordinates for measurement of cerebral blood flow and intracranial pressure in rabbits

Despite the increased use of intracranial neuromonitoring during experimental subarachnoid hemorrhage (SAH), coordinates for probe placement in rabbits are lacking. This study evaluates the safety and reliability of using outer skull landmarks to identify locations for placement of cerebral blood flow (CBF) and intraparenchymal intracranial pressure (ICP) probes. Experimental SAH was performed in 17 rabbits using an extracranial-intracranial shunt model. ICP probes were placed in the frontal lobe and compared to measurements recorded from the olfactory bulb. CBF probes were placed in various locations in the frontal cortex anterior to the coronary suture. Insertion depth, relation to the ventricular system, and ideal placement location were determined by post-mortem examination. ICP recordings at the time of SAH from the frontal lobe did not differ significantly from those obtained from the right olfactory bulb. Ideal coordinates for intraparenchymal CBF probes in the left and right frontal lobe were found to be located 4.6±0.9 and 4.5±1.2 anterior to the bregma, 4.7±0.7mm and 4.7±0.5mm parasagittal, and at depths of 4±0.5mm and 3.9±0.5mm, respectively. The results demonstrate that the presented coordinates based on skull landmarks allow reliable placement of intraparenchymal ICP and CBF probes in rabbit brains without the use of a stereotactic frame.

Improved microsurgical creation of venous pouch arterial bifurcation aneurysms in rabbits

The choice of the experimental aneurysm model is essential for valid embolization-device evaluations. So far, the use of the rabbit venous pouch arterial bifurcation aneurysm model has been limited by demanding microsurgery, low aneurysm patency rates, and high mortality. This study aimed to facilitate microsurgery and to reduce mortality by optimized peri-/postoperative management.

Wild immunology: converging on the real world

Recently, the Centre for Immunity, Infection and Evolution sponsored a one-day symposium entitled "Wild Immunology." The CIIE is a new Wellcome Trust-funded initiative with the remit to connect evolutionary biology and ecology with research in immunology and infectious diseases in order to gain an interdisciplinary perspective on challenges to global health. The central question of the symposium was, "Why should we try to understand infection and immunity in wild systems?" Specifically, how does the immune response operate in the wild and how do multiple coinfections and commensalism affect immune responses and host health in these wild systems? The symposium brought together a broad program of speakers, ranging from laboratory immunologists to infectious disease ecologists, working on wild birds, unmanaged animals, wild and laboratory rodents, and on questions ranging from the dynamics of coinfection to how commensal bacteria affect the development of the immune system. The meeting on wild immunology, organized by Amy Pedersen, Simon Babayan, and Rick Maizels, was held at the University of Edinburgh on 30 June 2011.

Seropositivity and Risk Factors Associated with Toxoplasma gondii Infection in Wild Birds from Spain

Toxoplasma gondii is a zoonotic intracellular protozoan parasite of worldwide distribution that infects many species of warm-blooded animals, including birds. To date, there is scant information about the seropositivity of T. gondii and the risk factors associated with T. gondii infection in wild bird populations. In the present study, T. gondii infection was evaluated on sera obtained from 1079 wild birds belonging to 56 species (including Falconiformes (n = 610), Strigiformes (n = 260), Ciconiiformes (n = 156), Gruiformes (n = 21), and other orders (n = 32), from different areas of Spain. Antibodies to T. gondii (modified agglutination test, MAT titer ≥1:25) were found in 282 (26.1%, IC95%:23.5–28.7) of the 1079 birds. This study constitute the first extensive survey in wild birds species in Spain and reports for the first time T. gondii antibodies in the griffon vulture (Gyps fulvus), short-toed snake-eagle (Circaetus gallicus), Bonelli's eagle (Aquila fasciata), golden eagle (Aquila chrysaetos), bearded vulture (Gypaetus barbatus), osprey (Pandion haliaetus), Montagu's harrier (Circus pygargus), Western marsh-harrier (Circus aeruginosus), peregrine falcon (Falco peregrinus), long-eared owl (Asio otus), common scops owl (Otus scops), Eurasian spoonbill (Platalea leucorodia), white stork (Ciconia ciconia), grey heron (Ardea cinerea), common moorhen (Gallinula chloropus); in the International Union for Conservation of Nature (IUCN) “vulnerable” Spanish imperial eagle (Aquila adalberti), lesser kestrel (Falco naumanni) and great bustard (Otis tarda); and in the IUCN “near threatened” red kite (Milvus milvus). The highest seropositivity by species was observed in the Eurasian eagle owl (Bubo bubo) (68.1%, 98 of 144). The main risk factors associated with T. gondii seropositivity in wild birds were age and diet, with the highest exposure in older animals and in carnivorous wild birds. The results showed that T. gondii infection is widespread and can be at a high level in many wild birds in Spain, most likely related to their feeding behaviour.

Can wild ungulate carcasses provide enough biomass to maintain avian scavenger populations? An empirical assessment using a bio-inspired computational model

Background The reduction in the amount of food available for European avian scavengers as a consequence of restrictive public health policies is a concern for managers and conservationists. Since 2002, the application of several sanitary regulations has limited the availability of feeding resources provided by domestic carcasses, but theoretical studies assessing whether the availability of food resources provided by wild ungulates are enough to cover energetic requirements are lacking. Methodology/Findings We assessed food provided by a wild ungulate population in two areas of NE Spain inhabited by three vulture species and developed a P System computational model to assess the effects of the carrion resources provided on their population dynamics. We compared the real population trend with to a hypothetical scenario in which only food provided by wild ungulates was available. Simulation testing of the model suggests that wild ungulates constitute an important food resource in the Pyrenees and the vulture population inhabiting this area could grow if only the food provided by wild ungulates would be available. On the contrary, in the Pre-Pyrenees there is insufficient food to cover the energy requirements of avian scavenger guilds, declining sharply if biomass from domestic animals would not be available. Conclusions/Significance Our results suggest that public health legislation can modify scavenger population trends if a large number of domestic ungulate carcasses disappear from the mountains. In this case, food provided by wild ungulates could be not enough and supplementary feeding could be necessary if other alternative food resources are not available (i.e. the reintroduction of wild ungulates), preferably in European Mediterranean scenarios sharing similar and socio-economic conditions where there are low densities of wild ungulates. Managers should anticipate the conservation actions required by assessing food availability and the possible scenarios in order to make the most suitable decisions.

Spatial and temporal variation in population genetic structure of wild Nile tilapia (Oreochromis niloticus) across Africa

Background: Reconstructing the evolutionary history of a species is challenging. It often depends not only on the past biogeographic and climatic events but also the contemporary and ecological factors, such as current connectivity and habitat heterogeneity. In fact, these factors might interact with each other and shape the current species distribution. However, to what extent the current population genetic structure reflects the past and the contemporary factors is largely unknown. Here we investigated spatio-temporal genetic structures of Nile tilapia (Oreochromis niloticus) populations, across their natural distribution in Africa. While its large biogeographic distribution can cause genetic differentiation at the paleo-biogeographic scales, its restricted dispersal capacity might induce a strong genetic structure at micro-geographic scales. Results: Using nine microsatellite loci and 350 samples from ten natural populations, we found the highest genetic differentiation among the three ichthyofaunal provinces and regions (Ethiopian, Nilotic and Sudano-Sahelian) (R(ST) = 0.38 - 0.69). This result suggests the predominant effect of paleo-geographic events at macro-geographic scale. In addition, intermediate divergences were found between rivers and lakes within the regions, presumably reflecting relatively recent interruptions of gene flow between hydrographic basins (R(ST) = 0.24 - 0.32). The lowest differentiations were observed among connected populations within a basin (R(ST) = 0.015 in the Volta basin). Comparison of temporal sample series revealed subtle changes in the gene pools in a few generations (F = 0 - 0.053). The estimated effective population sizes were 23 - 143 and the estimated migration rate was moderate (m similar to 0.094 - 0.097) in the Volta populations. Conclusions: This study revealed clear hierarchical patterns of the population genetic structuring of O. niloticus in Africa. The effects of paleo-geographic and climatic events were predominant at macro-geographic scale, and the significant effect of geographic connectivity was detected at micro-geographic scale. The estimated effective population size, the moderate level of dispersal and the rapid temporal change in genetic composition might reflect a potential effect of life history strategy on population dynamics. This hypothesis deserves further investigation. The dynamic pattern revealed at micro-geographic and temporal scales appears important from a genetic resource management as well as from a biodiversity conservation point of view.

A novel isolation method of Brucella species and molecular tracking of Brucella suis biovar 2 in domestic and wild animals

Brucella suis biovar 2 is the most common aetiological agent of porcine brucellosis in Europe. B. suis biovar 2 is considered to have low zoonotic potential, but is a causative agent of reproductive losses in pigs, and it is thus economically important. The multilocus variable-number of tandem repeats genotyping analysis of 16 loci (MLVA-16) has proven to be highly discriminatory and is the most suitable assay for simultaneously identifying B. suis and tracking infections. The aim of this study was to investigate the relatedness between isolates of B. suis biovar 2 obtained during a brucellosis outbreak in domestic pigs and isolates from wild boars and hares collected from proximal or remote geographical areas by MLVA-16. A cluster analysis of the MLVA-16 data revealed that most of the isolates obtained from Switzerland clustered together, with the exception of one isolate. The outbreak isolates constituted a unique subcluster (with a genetic similarity >93.8%) distinct from that of the isolates obtained from wild animals, suggesting that direct transmission of the bacterium from wild boars to domestic pigs did not occur in this outbreak. To obtain a representative number of isolates for MLVA-16, alternative methods of Brucella spp. isolation from tissue samples were compared with conventional direct cultivation on a Brucella-selective agar. We observed an enhanced sensitivity when mechanical homogenisation was followed by host cell lysis prior to cultivation on the Brucella-selective agar. This work demonstrates that MLVA-16 is an excellent tool for both monitoring brucellosis and investigating outbreaks. Additionally, we present efficient alternatives for the isolation of Brucella spp.