Network on veterinary medicines initiated by the European Federation For Pharmaceutical Sciences.

The European Federation for Pharmaceutical Sciences (EUFEPS) was founded 25 years ago by more than 20 national pharmaceutical societies and faculty members. As a pan-European organization, it brings together pharmaceutical societies as well as academic, industrial and regulatory scientists engaged in drug research and development, drug regulation and education of professionals working in these fields. EUFEPS represents pharmaceutical sciences in Europe and is recognized as such by both the European Commission and the European Medicines Agency. EUFEPS cooperates with the European Federation of Pharmaceutical Industries and other European organizations and maintains global connections with agencies such as the US Food and Drug Administration and the American Association of Pharmaceutical Scientists. EUFEPS has established specified networks forming the basis of its activities. The creation of a Network on Veterinary Medicines is prompted by the manifold problems resulting from the use of veterinary drugs and its inherent interconnections with human medicine, environmental and public health. A long-term goal of this initiative was to expand the spectrum of available therapeutics for use in animals, including the development of innovative delivery systems.

Comparison of milk oligosaccharides pattern in colostrum of different horse breeds.

Colostrum oligosaccharides are known to exhibit prebiotic and immunomodulatory properties. Oligosaccharide composition is species-specific, and equine colostrum has been reported to contain unique oligosaccharides. Therefore, equine oligosaccharides (EMOS) from colostrum from different horse breeds were analyzed by CE-LIF, CE-MS(n), HILIC-MS(n), and exoglycosidase degradation. Sixteen EMOS were characterized and quantified, of which half were neutral and half were acidic. EMOS showed about 63% structural overlap with human milk oligosaccharides, known for their bioactivity. Seven EMOS were not reported before in equine oligosaccharides literature: neutral Gal(ß1-4)HexNAc, Gal(ß1-4)Hex-Hex, ß4'-galactosyllactose, and lactose-N-hexaose, as well as acidic 6'-Sialyl-Hex-Ac-HexNAc, sialyllacto-N-tetraose-a, and disialylacto-N-tetraose (isomer not further specified). In all colostrum samples, the average oligosaccharide concentration ranged from 2.12 to 4.63 g/L; with ß 6'and 3'- galactosyllactose, 3'-sialyllactose, and disialyllactose as the most abundant of all oligosaccharides (27-59, 16-37, 1-8, and 1-6%, respectively). Differences in presence and in abundance of specific EMOS were evident not only between the four breeds but also within the breed.

Equine colostral carbohydrates reduce lipopolysaccharide-induced inflammatory responses in equine peripheral blood mononuclear cells.

REASONS FOR PERFORMING STUDY: Increasing evidence suggests that reactions to lipopolysaccharide (LPS), particularly in the gut, can be partly or completely mitigated by colostrum- and milk-derived oligosaccharides. Confirmation of this hypothesis could lead to the development of new therapeutic concepts. OBJECTIVES: To demonstrate the influence of equine colostral carbohydrates on the inflammatory response in an in vitro model with equine peripheral blood mononuclear cells (PBMCs). METHODS: Carbohydrates were extracted from mare colostrum, and then evaluated for their influence on LPS-induced inflammatory responses in PBMCs isolated from the same mares, mRNA expression of tumour necrosis factor-alpha, interleukin-6 and interleukin-10 was measured as well as the protein levels of tumour necrosis factor-alpha (TNF-alpha) and interleukin-10 (IL-10). RESULTS: Equine colostral carbohydrates significantly reduced LPS-induced TNF-alpha protein at both times measured and significantly reduced LPS-induced TNF-alpha, IL-6 and IL-10 mRNA expression by PBMCs. Moreover, cell viability significantly increased in the presence of high concentrations of colostral carbohydrates. CONCLUSIONS: Carbohydrates derived from equine colostrum reduce LPS-induced inflammatory responses of equine PBMCs. POTENTIAL RELEVANCE: Colostrum and milk-derived carbohydrates are promising candidates for new concepts in preventive and regenerative medicine.

Genotyping of Toll-like receptor 4, myeloid differentiation factor 2 and CD-14 in the horse: an investigation into the influence of genetic polymorphisms on the LPS induced TNF-alpha response in equine whole blood.

The inter- and intra-species differences in the response to lipopolysaccharides (LPS) are well recognised in mammalian species. It has been hypothesized that these differences can be attributed to genetic polymorphisms in the components involved in LPS signal transduction. These components include the cluster of differentiation factor 14 (CD-14), a membrane bound protein on the surface of mononuclear cells that recognises LPS and a receptor complex consisting of Toll-like receptor-4 (TLR-4) and myeloid differentiation factor-2 (MD-2). Sequencing of these three proteins in humans and mice revealed that all three are susceptible to polymorphic alterations, influencing the response to LPS. Previous experiments in the horse showed large inter-individual variations in the response to LPS. With the aim to assess this inter-individual variation, we performed a whole blood assay in 10 healthy horses as a functional assay to study the responsiveness to LPS. In 3 out of the 10 horses, LPS-induced TNF-alpha production was significantly lower compared to the overall mean. Subsequently the entire cDNA sequence encoding for the TLR-4, MD-2 and CD-14 protein was documented for each horse. Although mutations were observed in the sequence of TLR-4, these could not be related to an altered response to LPS in the concentration used in this study, as determined in the whole blood assay. Despite the various mutations found in the TLR-4 receptor protein, no alterations could be found in either the MD-2 or CD-14 gene, which are obviously more conserved structures.

Purification and N-terminal sequencing of style glycoproteins associated with self-incompatibility in Petunia hybrida.

We report isolation and N-terminal amino acid sequencing of three style glycoproteins, which segregate with three S (self-incompatibility) alleles of Petunia hybrida. The S-glycoproteins were expressed mainly in the upper part of the pistil and showed an increasing concentration during flower development. The glycoproteins were purified by a combination of ConA-Sepharose and cation exchange fast protein liquid chromatography. The amount of S-glycoproteins recovered from style extracts varied from 0.5 to 1.6 micrograms per style, which was 40-60% of the amount recovered by a simplified analytical method. N-terminal amino acid sequences of S1-, S2- and S3-glycoprotein showed homology within the fifteen amino terminal residues. These amino acid sequences were compared with the previously published sequences of S-glycoproteins from Nicotiana alata and Lycopersicon peruvianum.

Evidence for the existence of a universal crown-gall tumor initiation enhancer.

Tumor initiation of different dicotyledonous plant species inoculated with Agrobacterium tumefaciens B6 has been studied in vivo and in vitro. The tumor formation in weakly susceptible plants can be strongly enhanced by exogenously applied active extract fractions derived from highly susceptible Helianthus cotyledons. It is found that highly susceptible plants (Kalanchoë, Lycopersicon and Pinto beans) contain an active tumor initiation enhancer which is clearly similar to the compound(s) found in Helianthus cotyledons. No activity can be detected in extracts derived from weakly susceptible plants (Coleus, Phaseolus) or in those obtained from crown-gall tumor tissues.

Influence of a crown-gall tumor initiation enhancer on bacterial attachment to the host plant cell wall.

The competitive activities of different plant cell walls upon Agrobacterium tumefaciens attachment have been studied in vitro by means of two crown-gall tumor initiation assays. The low or high susceptibility of different plant species is independent of their capacity to cause bacterial cells to adhere to specific sites on the plant cell walls. However, the attachment properties of cell wall fragments derived from Helianthus cotyledons seem to be age-dependent. It is found that a tumor initiation enhancer, present in extract fractions derived from highly susceptible plants and closely related with the competence for tumor formation, does not influence bacterial adherence. The two steps, attachment and the step by which the tumor initiation enhancer is involved, clearly differ in the processes leading to the transformation of a normal cell into a tumor cell.

Effects of abscisic acid, cytokinins, and light on isoflavonoid phytoalexin accumulation inPhaseolus vulgaris L.

Cotyledons ofPhaseolus vulgaris L. contain small amounts of phaseollin and kievitone. Isolating the cotyledons from the plant does not alter phaseollin levels. Kievitone levels, however, although not affected in light-incubated cotyledons, increased rapidly in dark-incubated cotyledons. Abscisic acid (ABA) at 10(-4) M stimulated the accumulation of phaseollin in excised cotyledons in both light and darkness, whereas benzylaminopurine (BAP) increased these levels only in the light. The kievitone level was influenced by ABA and BAP only in dark-incubated cotyledons, i.e., inhibited at 10(-4) M. When excised cotyledons were treated with mercuric chloride, both phaseollin and kievitone accumulated rapidly in both light and darkness. The effect of ABA on these cotyledons was similar to that on non-treated cotyledons. The results demonstrate that the synthesis of the two phytoalexins is regulated by separate mechanisms and indicate that the phytoalexin composition is dependent on the physiological condition of the cotyledons. ABA and BAP may play a role in the resistance response of the plant.