Angiogenesis is the process that drives blood vessel development in growing tissues in response to the local production of angiogenic factors. With the present research the authors have studied vascular endothelial growth factor (VEGF) production in ovarian follicles as a potential mechanism of ovarian activity regulation. Prepubertal gilts were treated with 1250 IU equine chorionic gonadotropin (eCG) followed 60 h later by 750 IU of human chorionic gonadotropin (hCG) in order to induce follicle growth and ovulation. Ovaries were collected at different times of the treatment and single follicles were isolated and classified according to their diameter as small (<4 mm), medium (4-5 mm), or large (>5 mm). VEGF levels were measured in follicular fluid by enzyme immunoassay, and VEGF mRNA content was evaluated in isolated theca and granulosa compartments. Equine chorionic gonadotropin stimulated a prompt follicular growth and induced a parallel evident rise in VEGF levels in follicular fluid of medium and large follicles. Analysis of VEGF mRNA levels confirmed the stimulatory effect of eCG, showing that it is confined to granulosa cells, whereas theca cells maintained their VEGF steady state mRNA. Administration of hCG 60 h after eCG caused a dramatic drop in follicular fluid VEGF that reached undetectable levels in 36 h. A parallel reduction in VEGF mRNA expression was recorded in granulosa cells. The stimulating effect of eCG was also confirmed by in vitro experiments, provided that follicles in toto were used, whereas isolated follicle cells did not respond to this hormonal stimulation. Consistent with the observation in vivo, granulosa cells in culture reacted to hCG with a clear block of VEGF production. These results demonstrate that while follicles of untreated animals produce stable and low levels of the angiogenic factor, VEGF markedly rose in medium and large follicles after eCG administration. The increasing levels, essentially attributable to granulosa cells, are likely to be involved in blood vessel development in the wall of growing follicles, and may play a local key role in gonadotropin-induced follicle development. When ovulation approaches, under the effect of hCG, the production of VEGF is switched off, probably creating the safest conditions for the rupture of the follicle wall while theca cells maintained unaltered angiogenic activity, which is probably required for corpus luteum development.

A large number of hDAF transgenic pigs to be used for xenotransplantation research were generated by using sperm-mediated gene transfer (SMGT). The efficiency of transgenesis obtained with SMGT was much greater than with any other method. In the experiments reported, up to 80% of pigs had the transgene integrated into the genome. Most of the pigs carrying the hDAF gene transcribed it in a stable manner (64%). The great majority of pigs that transcribed the gene expressed the protein (83%). The hDAF gene was transmitted to progeny. Expression was stable and found in caveolae as it is in human cells. The expressed gene was functional based on in vitro experiments performed on peripheral blood mononuclear cells. These results show that our SMGT approach to transgenesis provides an efficient procedure for studies involving large animal models.SMGT ͉ hyperacute rejection ͉ swine

BackgroundThe similarities between swine and humans in physiological and genomic patterns, and the great correlation in size and anatomy, make pigs extremely useful in preclinical studies. New-born piglets can represent a model for congenital and genetic diseases in new-born children. It is known that piglets may have significant differences in clinicopathological results compared to adult pigs. Therefore, adult laboratory reference intervals cannot be applied to piglets. The aim of this study was to compare haematological and chemical variables in piglets of two ages and determinate age-related reference intervals for commercial hybrid young pigs.Blood samples were collected under general anaesthesia from 130 animals divided into five- (P5) and 30- (P30) day-old piglets. Only P30 animals were treated with parenteral iron after birth. Samples were analysed using automated haematology (ADVIA 2120) and chemistry analysers, and age-related reference intervals were calculated.ResultsSignificant higher values of RBC, Hb and HCT were observed in P30 animals when compared to P5, with an opposite trend for MCV. These results were associated with a reduction of the RBC regeneration process and the thrombopoietic response. The TSAT and TIBC were significantly higher in P30 compared to P5; however, piglets remained iron deficient compared to adult reference intervals reported previously.ConclusionsIn conclusion, this paper emphasises the high variability occurring in clinicopathological variables between new-born and 30-day-old pigs, and between piglets and adult pigs. This study provides valuable reference data for piglets at precise ages and could be used in the future as historical control improving the Reduction in animal experiments, as suggested by the 3Rs principle.Electronic supplementary materialThe online version of this article (doi:10.1186/s12917-017-0946-2) contains supplementary material, which is available to authorized users.

Endotoxic shock, one of the most prominent causes of mortality in intensive care units, is characterized by pulmonary hypertension, systemic hypotension, heart failure, widespread endothelial activation/injury, and clotting culminating in disseminated intravascular coagulation and multi-organ system failure. In the last few years, studies in rodents have shown that administration of low concentrations of carbon monoxide (CO) exerts potent therapeutic effects in a variety of diseases/disorders. In this study, we have administered CO (one our pretreatment at 250 ppm) in a clinically relevant, well-characterized model of LPS-induced acute lung injury in pigs. Pretreatment only with inhaled CO significantly ameliorated several of the acute pathological changes induced by endotoxic shock. In terms of lung physiology, CO pretreatment corrected the LPS-induced changes in resistance and compliance and improved the derangement in pulmonary gas exchange. In terms of coagulation and inflammation, CO reduced the development of disseminated intravascular coagulation and completely suppressed serum levels of the proinflammatory IL-1beta in response to LPS, while augmenting the anti-inflammatory cytokine IL-10. Moreover, the effects of CO blunted the deterioration of kidney and liver function, suggesting a beneficial effect in terms of end organ damage associated with endotoxic shock. Lastly, CO pretreatment prevents LPS-induced ICAM expression on lung endothelium and inhibits leukocyte marginalization on lung parenchyma.