Individual-based simulations of the War of Attrition

The War of Attrition model of John Maynard Smith predicts a single, mixed evolutionarily stable strategy (ESS) for animal contests which are settled by conventional displays with no assessment of the opponent's fighting ability. We test the predictions of the model by simulating the evolution of strategies in a finite population of animals under various assumptions on how possible strategies are coded and mutated. While our simulations for the most part confirm the predictions of the model, we also discovered some significant deviations from the theoretically predicted ESS. Specifically, we found that if inheritance of strategies is somewhat imprecise, then a population can evolve that achieves on average a higher payoff than a population at the theoretically predicted ESS. Moreover, if the ESS is realized as a polymorphism of fixed persistence times, then for small populations, sufficiently stringent statistical tests will reject the hypothesis that these times are distributed as theoretically predicted.     

Effects of adrenocorticotropin stimulation on cortisol dynamics of pregnant gilts and their fetuses: implications for prenatal stress studies

The effect of adrenocorticotropic hormone (ACTH) administration on plasma cortisol concentrations was determined in pregnant gilts and their fetuses. In a first experiment, 100 IU ACTH (Synacthen Depot) was administered intramuscularly to the gilts every second day from Days 49 to 75 of gestation. ACTH injections were carried out at 08:00 h and, thereafter, 10 blood samples were taken within the following 8h via jugular catheters. Blood samples were analysed for plasma cortisol concentrations, and results were compared with values from animals which were treated with physiological saline and untreated animals (blood sampling only). The values for plasma cortisol concentrations increased until 3h after ACTH applications to a mean maximum level of 276.5+/-17.2 nmol/l in the whole 4-week stimulation period. Plasma cortisol levels did not return to pre-treatment values within the 8 h post-injection. No differences in cortisol levels were found between the physiological saline and untreated control, and no habituation of the adrenocortical response to ACTH was found during the 4-week stimulation period. In a second experiment, 100 IU ACTH were administered to pregnant gilts at gestation Day 65. After 3 h, fetuses were recovered under general anaesthesia and blood samples were taken from the umbilical vein, artery, and, after decapitation, from periphery. Application of ACTH to the sows significantly increased their plasma cortisol concentrations (P<0.001), and also increased plasma cortisol concentrations in peripheral blood samples from the fetuses (P=0.09) and in the umbilical vein (P<0.001) and artery (P<0.01), respectively. Plasma ACTH concentrations did not differ in fetuses from ACTH-treated or control sows. The results show that in gilts the adrenocortical response to an exogenous application of Synacthen Depot is consistent over time during mid-gestation. Furthermore, cortisol but not ACTH levels were increased in fetuses from ACTH-treated sows, indicating that maternal cortisol can cross the placenta during mid-gestation. The stimulation of maternal cortisol release through exogenous ACTH with subsequent elevation of fetal cortisol levels is, therefore, a useful approach for studying effects of elevated maternal glucocorticoids in prenatal stress studies in pigs.     

Polymorphonuclear granulocytes induce antibody-dependent apoptosis in human breast cancer cells

Recent studies in HER-2/neu-targeted immunotherapy demonstrated that polymorphonuclear neutrophils (PMN) mediated Ab-dependent cellular cytotoxicity against HER-2/neu-positive breast cancer cell lines. However, the mechanism of cell death remained unclear. We used several assays to analyze the induction of apoptosis in the breast cancer cell line SK-BR-3 via PMN-dependent Ab-dependent cellular cytotoxicity. In the presence of the HER-2/neu Ab 520C9 and PMN from healthy donors, apoptosis occurred as detected by annexin V binding and disappearance of euploid SK-BR-3 nuclei, which can be differentiated from PMN nuclei by their increased DNA contents. Apoptosis induction was observed with E:T cell ratios as low as 10:1. Laser scanning fluorescence microscopy of TUNEL tumor cells or staining for cleaved cytokeratin-18 further confirmed apoptosis of the SK-BR-3 breast cancer cells. Killing via 520C9 was dependent on the interaction with FcR on PMN, because 1) F(ab')(2) fragments of 520C9 mediated no cytotoxicity, 2) target cell death was influenced by a biallelic polymorphism of FcgammaRIIa on the effector cells, and 3) a bispecific Ab against HER-2/neu and the IgA receptor (FcalphaRI) expressed on effector cells significantly induced apoptosis. Thus, PMN induce Ab-dependent apoptosis against human breast cancer cells targeted with HER-2/neu-directed mAbs or FcR directed bispecific Abs.     

Protein kinase C-delta is a negative regulator of antigen-induced mast cell degranulation

Regulation of mast cell degranulation is dependent on the subtle interplay of cellular signaling proteins. The Src homology 2 (SH2) domain-containing inositol-5'-phosphatase (SHIP), which acts as the gatekeeper of degranulation, binds via both its SH2 domain and its phosphorylated NPXY motifs to the adapter protein Shc via the latter's phosphorylated tyrosines and phosphotyrosine-binding domain, respectively. This theoretically leaves Shc's SH2 domain available to bind proteins, which might be part of the SHIP/Shc complex. In a search for such proteins, protein kinase C-delta (PKC-delta) was found to coprecipitate in mast cells with Shc and to interact with Shc's SH2 domain following antigen or pervanadate stimulation. Phosphorylation of PKC-delta's Y(332), most likely by Lyn, was found to be responsible for PKC-delta's binding to Shc's SH2 domain. Using PKC-delta(-/-) bone marrow-derived mast cells (BMMCs), we found that the antigen-induced tyrosine phosphorylation of Shc was similar to that in wild-type (WT) BMMCs while that of SHIP was significantly increased. Moreover, increased translocation of PKC-delta to the membrane, as well as phosphorylation at T505, was observed in SHIP(-/-) BMMCs, demonstrating that while PKC-delta regulates SHIP phosphorylation, SHIP regulates PKC-delta localization and activation. Interestingly, stimulation of PKC-delta(-/-) BMMCs with suboptimal doses of antigen yielded a more sustained calcium mobilization and a significantly higher level of degranulation than that of WT cells. Altogether, our data suggest that PKC-delta is a negative regulator of antigen-induced mast cell degranulation.     

Morphological phenotyping of enteric neurons using neurofilament immunohistochemistry renders chemical phenotyping more precise in porcine ileum

The aim of the study was: (1) to test the suitability of neurofilament (NF) immunohistochemistry for representing the shapes of morphologically defined neuron types in the pig ileum myenteric plexus, (2) to estimate the proportions of these neuron types as related to the whole myenteric neuron population and (3) to demonstrate the usefulness of a refined morphological classification of enteric neurons on the paradigm of calcitonin gene-related peptide (CGRP)-immunoreactive neurons. So far, immunoreactivity for this peptide was supposed to be present in the pig enteric nervous system only in type II neurons. Ileal whole mounts of two pigs were stained with the cuprolinic blue (CB) method and, thereafter, incubated with an antibody pool against NF proteins (70, 160 and 210 kDa), visualised with a fluorochrome-tagged secondary antibody. The structural representation of morphologically defined myenteric neuron types typical for pig ileum (Stach I, II, IV, V and VI) was equivalent to their silver impregnated image, as demonstrated in previous studies. Counts of CB-stained neurons revealed between 2,526 and 2,662 neurons per square centimetre in one pig and between 2,027 and 2,763 in the other. As related to these total neuron numbers, the proportions of type I neurons were 1.7% and 1.5%, of type II neurons 7.2% and 7.9%, of type IV neurons 1.9% and 2.4%, of type V neurons 1.1% and 1.5%, and of type VI neurons 1.3% each. These values are generally comparable with those estimated earlier on silver impregnated material. Double labelling for NF and CGRP indicated that CGRP-immunoreactive smooth contoured neurons with long processes could be subdivided into two distinct morphological neuron types, i.e. type II and type V. We conclude that NF immunohistochemistry is an appropriate tool for representation of morphologically defined enteric neuron types in the pig. Combination of this technique with immunohistochemistry for neuroactive substances may be useful for making both morphological and chemical classification schemes mutually more precise.     

Three-dimensional histomorphometric analysis of distraction osteogenesis using an implanted device for mandibular lengthening in sheep

The aim of this study was to lengthen the sheep mandible with a fully buried device and to quantitatively analyze the tissue regenerate in the distraction gap by means of two-dimensional and three-dimensional histomorphometry. A custom-made device for continuous distraction was used in five adult sheep and fixed with three bicortical screws on either side of an osteotomy, anterior to the premolar region of the mandible. A cable-connected power and control unit was implanted in the neck region. After a 5-day latency period, distraction was activated every 2 hours and advanced at a rate of 1.01 mm per day. The distraction period was planned for 14 days, but because of stability problems and cable breakage, the actual distraction period ranged from 2 to 17 days, resulting in gap distances from 1.7 to 17.1 mm (mean, 0.95 mm/day). Osteogenesis was followed by radiographic imaging, and after a 6-week consolidation period, the harvested mandibles were serially sectioned for histologic and two-dimensional histomorphometric analysis, with three-dimensional reconstruction. Histologic examination of the specimens demonstrated predominantly membranous bone formation with remodeling bridging the distraction gap mainly in the periosteal region of the lingual side. In addition, cartilaginous areas and chondral bone formation were observed where the bridging appeared incomplete. Because of device fixation on the buccal side of the mandible, the preservation of the lingual periosteum seemed to play the major role for sufficient bone repair in the distraction gap. Cartilage within the distraction gap suggests fixation instability in this animal model.