Establishment and characterization of continuous hematopoietic progenitors-derived pig normal mast cell lines

Mast cells (MCs) are tissue resident, hematopoietic stem cells-derived elements, distributed throughout the body. They are the pivotal mediating cells of allergic reactions. In addition, in mice, MCs play a critical role in the defense against several pathogens, such as bacteria, parasites and viruses. Whereas the biology of rodent and human MCs has been extensively studied using in vitro derived populations, the role of MCs in pigs has not yet been evaluated, given the very low availability of pure porcine MCs populations. In the present report, we describe an original method to obtain continuous factor-dependent normal pig MCs (PMC) lines from fetal hematopoietic progenitors. These Stem Cell Factor (SCF) and Interleukin-3- (IL-3)-dependent PMC lines retain their capacity to growth after conventional freezing methods and exhibit most of the morphological and biochemical properties of normal, although immature, MCs, including metachromatic granules containing sulfated polysaccharides, the expression of c-kit and high-affinity IgE receptors (FcepsilonRI), and the ability to store histamine that is released upon cross-linking of FcepsilonRI. In vitro derived PMC lines might thus be valuable tools to further investigate the reactivity of these elements towards several parasites frequently encountered in pig, such as, but not limited to, Ascaris suum, Trichinella spiralis or Trichuris suis, or towards antigens derived from these pathogens.     

Long-term in vitro culture of wheat grains

Wheat (Triticum aestivum, Triticum durum) grains were excised immediately following fertilization and cultured until maturity. A rachis fragment attached to the grain was required to ensure an increase in grain size for the first 10 days following fertilization. A ¹⁴C-labeling study revealed that 8-day-old grains accumulated more dry matter into the ethanol-insoluble fraction when grown on agar rather than when immersed in liquid medium. Light enhanced the absorption of sucrose from the medium only in the latter case. In agar-based culture, when no contact was made between the grain surface and the medium, peeling off the outer pericarp layers increased sugar absorption, leading to a threefold increase in the amount of accumulated dry matter in the ethanol-insoluble fraction. Culturing of wheat grains with attached rachis fragment and peeled pericarp is recommended for maximum in vitro growth.     

Formation of the boundary between the midbrain and the hindbrain: involvement of Otx2 and Gbx2 genes

We have studied the neuromeric organisation of the mesencephalic-metencephalic (mes-met) territory of the avian neural tube using chick/quail transplantation experiments and analysing the expression of various regulatory genes in chimeric and normal embryos. Homotopic grafts demonstrate the presence of an interneuromeric boundary separating the mesencephalic and cerebellar territories (the mes-met or midbrain/hindbrain boundary). This boundary is characterised from HH10 onwards by the confrontation of the Otx2-Wnt1 and Gbx2-Fgf8 expressing domains, while En2 and Pax2 genes are expressed at both sides of the mes-met boundary. The evolution of the position of the Otx2/Gbx2 boundary with respect to the vesicles and constriction observed within the mes-met domain between stages HH10 and HH20, allows us to redefine the fate map of this region and to propose a new nomenclature for HH10. Transplantation between the prosencephalic neuroepithelium and the mes-met domain shows the possibility of inducing a mes-met phenotype within the two caudal-most prosomeres, preceded by its characteristic genetic cascade. The induction selectively takes place along the boundary between the graft (Otx2 positive) and the host cerebellar territory (expressing high levels of Gbx2); this includes the induction inside the graft of a new Otx2/Gbx2 boundary. Conversely, no induction is ever observed when the graft is confronted to the host Otx2 expressing domain. Although Fgf8 may be involved in the inductive events, our data strongly suggest that confrontation between Otx2 and Gbx2 is essential as an organiser of the mes-met domain.     

Preliminary investigations on somatic embryogenesis from leaf discs of red oak (Quercus rubra L.)

Somatic embryos were obtained from leaf discs of juvenile red oak plants. Basal inductive nutrient medium was a modified Murashige and Skoog solution enriched with 500 mg L^–1 casein hydrolysate, 100 mg L^–1 polyvinylpyrrolidone, 5.4 M naphthaleneacetic acid and 0.09 M benzyladenine. Embryogenesis was obtained only from leaf discs in the presence of light and increased when the adaxial surface of the explants (with midrib or main veins present) was in contact with the medium. Large variation was observed in all experiments. Recurrent embryogenesis was observed at the base of embryo clusters with callus present; conversely, embryogenic potential was rapidly lost by subculturing full calli. Maturation, germination and development of isolated somatic embryos were obtained. However, the vast majority of embryos did not have viable apical bud meristems and on only a few occasions were shoots produced.Abbreviations BA N⁶-benzyladenine - CH casein hydrolysate - 2iP isopentenyladenine - NAA naphthaleneacetic acid - 2.4-D 2.4-dichlorophenoxyacetic acid - GA gibberellic acid - PVP polyvinylpyrrolidone     

Effect of dietary energy level in finishing phase on performance,carcass and meat quality in immunocastrates and barrows in comparison with gilts and entire male pigs

Immunocastration, a technique consisting of two vaccinations against gonadotropin-releasing hormone (GnRH), can be used as alternative to surgical castration of piglets. It reduces boar taint and allows higher economic and ecological efficiency compared to barrows. The feeding strategy of immunocastrates, however, can still be improved. After second vaccination, when immunisation becomes fully effective, feed intake of immunocastrates increases sharply. This study aimed to investigate whether energy intake of immunocastrates after second vaccination could be reduced by lowering the dietary energy level of the finishing phase, without negatively affecting animal performance and quality of pork production. We hypothesised that immunocastrates already reach their limits in voluntary feed intake after second vaccination, and therefore would not be able to compensate the lower dietary energy level, in contrast to barrows. Therefore, this study aimed to assess the effect of high-energy diet (HE, net energy (NE) = 10.2 MJ/kg) compared to low-energy diet (LE, NE = 8.8 MJ/kg) in barrows and immunocastrates and as a reference, gilts and entire male pigs on a standard high-energy diet were included. CP and standardised ileal digestible amino acid levels were similar in both diets. For each treatment, eight pen replicates of six pigs per pen were evaluated on performance, carcass quality, meat and fat quality, digestibility, economic and ecological sustainability, behaviour and effectiveness of immune response. No difference in feed intake of immunocastrates between LE and HE could be demonstrated. As a result, daily energy intake of immunocastrates was higher on HE compared to LE, which resulted in a higher daily gain on HE. Feed conversion ratio (FCR) of immunocastrates on HE did not differ significantly with FCR of entire males. Barrows did not show higher average daily gain on HE compared to LE. Nitrogen efficiency was better in HE compared to LE, without negative effects on digestibility, carcass quality, economic parameters, behaviour or immune response. Small positive effects on the palatability of the meat of immunocastrates on HE were observed, although consumers did not prefer one of both feeds. Immunocastration was successful in reducing sexual and aggressive behaviour as well as in lowering the prevalence of boar taint from 15% in EM to 0% in immunocastrates. However, in two out of 96 immunocastrates (one on HE and one on LE), the immunocastration was not fully effective. In conclusion, this study did not show advantages of feeding immunocastrates or barrows a low-energy diet.     

Effect of nicotine on in vivo secretion of melanocorticotropic hormones in the rat

Corticosterone, ACTH, β-endorphin and α-MSH were measured in rat plasma by radioimmunoassay before and 2,5,15,30 minutes after an intraperitoneal injection of nicotine (500 μg/Kg b.w.). Nicotine induced an increase of plasma corticosterone (p < 0.05 at t + 15 min), ACTH and β-endorphin (p < 0.01 at t + 5 min) and a decrease of α-MSH (p < 0.005 at t + 15 min). Dose response experiments showed an increase of corticosterone, ACTH, β-endorphin 15 min after 250 μg/Kg b.w. nicotine I.P., no effect being observed after injection of 100 μg/Kg b.w. The decrease of α-MSH was observed 15 min after 100, 250 or 500 μg/Kg b.w. nicotine I.P. Our results suggest that the increase of corticosterone is mediated through ACTH release.     

Effect of seven hours intermittent suckling and flavour recognition on piglet performance

A low feed intake during the first days after weaning is predisposing for weaning diarrhoea and weight loss. In this experiment we tried to increase the feed intake of the piglets after weaning by stimulating the solid feed intake during the last two weeks before weaning by separating them from the sows for 7 h/d. In addition, the effect of flavour recognition and the interaction of flavour recognition with intermittent suckling were tested. In two consecutive weaning rounds, sows were divided over two compartments with 7 to 10 sows each. They were assigned to one of four treatments in a two factorial design: control housing/control feed (n = 7); control housing/ flavoured feed (n = 8); intermittent suckling/control feed (n = 7); intermittent suckling/ flavoured feed (n = 9). After weaning, for each round 3 pens of 6 pigs were selected per treatment group. All these piglets received the same feed with the same flavour at the same concentration. Contrary to the expectations, intermittent suckling decreased the solid feed intake during the last two weeks before weaning. Flavour addition to the creep feed did not increase feed intake or other performance parameters before and after weaning, nor did it interact with intermittent suckling. Although after intermittent suckling pigs ate less creep feed before weaning, they did not perform worse after weaning. However, to increase feed intake after weaning, longer periods of separation might be necessary.     

A mutant of Azospirillum brasilense Sp7 impaired in flocculation with a modified colonization pattern and superior nitrogen fixation in association with wheat.

We report here significant phenotypic and genetic differences between Azospirillum brasilense Sp7 and spontaneous mutant Sp7-S and their related properties in association with wheat. In contrast to the wild-type strain of Sp7, colonies of Sp7-S stained weakly with Congo red when grown on agar media containing the dye and did not flocculate in the presence of fructose and nitrate. Scanning and transmission electron micrographs showed clearly that the Sp7-S strain lacked surface materials present as a thick layer on the surface of the wild-type Sp7 strain. Different patterns of colonization on wheat roots between Sp7 and Sp7-S, revealed by in situ studies using nifA-lacZ as a reporter gene, were related to a large increase in nitrogenase activity (acetylene reduction) with Sp7-S in association with normal and 2,4-dichlorophenoxyacetic acid-treated wheat for assays conducted under conditions in which the nitrogenase activity of free-living Azospirillum organisms was inhibited by an excess of oxygen. Randomly amplified polymorphic DNA analysis indicated the close genetic relationship of Sp7-S to several other sources of Sp7, by comparison to other recognized strains of A. brasilense. Genetic complementation of Sp7-S was achieved with a 9.4-kb fragment of DNA cloned from wild-type Sp7, restoring Congo red staining and flocculation.     

Imbalanced distribution of Plasmodium falciparum MSP-1 genotypes related to sickle-cell trait.

BACKGROUND: The sickle-cell trait protects against severe Plasmodium falciparum malaria and reduces susceptibility to mild malaria but does not prevent infection. The exact mechanism of this protection remains unclear. We have hypothesized that AS individuals are protected by virtue of being less susceptible to a subset of parasite strains; thus we compared some genetic characteristics of parasites infecting AS and AA subjects. MATERIALS AND METHODS: Blood was collected from asymptomatic individuals living in two different regions of Africa. The polymorphic MSP-1 and MSP-2 loci were genotyped using a PCR-based methodology. Individual alleles were identified by size polymorphism, amplification using family-specific primers, and hybridization using family-specific probes. Multivariate logistic regression was used to analyze allele distribution. RESULTS: In Senegalese carriers, age and hemoglobin type influenced differently the distribution of the three MSP-1 families and had an impact on distinct individual alleles, whereas the distribution of MSP-2 alleles was marginally affected. There was no influence of other genetic traits, including the HLA Bw53 genotype, or factors such as place of residence within the village. In a cohort of Gabonese schoolchildren in which the influence of age was abrogated, a similar imbalance in the MSP-1 allelic distribution but not of MSP-2 allelic distribution by hemoglobin type was observed. CONCLUSIONS: The influence of the host's hemoglobin type on P. falciparum genotypes suggests that parasite fitness for a specific host is strain-dependent, which is consistent with our hypothesis that innate resistance might result from reduced fitness of some parasite strains for individuals with sickle-cell traits.