Early Castration in Horses Does Not Impact Osteoarticular Metabolism.

The castration of stallions is traditionally performed after puberty, at around the age of 2 years old. No studies have focused on the effects of early castration on osteoarticular metabolism. Thus, we aimed to compare early castration (3 days after birth) with traditional castration (18 months of age) in horses. Testosterone and estradiol levels were monitored from birth to 33 months in both groups. We quantified the levels of biomarkers of cartilage and bone anabolism (CPII and N-MID) and catabolism (CTX-I and CTX-II), as well as of osteoarthritis (HA and COMP) and inflammation (IL-6 and PGE2). We observed a lack of parallelism between testosterone and estradiol synthesis after birth and during puberty in both groups. The extra-gonadal synthesis of steroids was observed around the 28-month mark, regardless of the castration age. We found the expression of estrogen receptor (ESR1) in cartilage and bone, whereas androgen receptor (AR) expression appeared to be restricted to bone. Nevertheless, with respect to osteoarticular metabolism, steroid hormone deprivation resulting from early castration had no discernable impact on the levels of biomarkers related to bone and cartilage metabolism, nor on those associated with OA and inflammation. Consequently, our research demonstrated that early castration does not disrupt bone and cartilage homeostasis.

Interactions between ß-endorphin and kisspeptin neurons of the ewe arcuate nucleus are modulated by photoperiod.

Opioid peptides are well-known modulators of the central control of reproduction. Among them, dynorphin coexpressed in kisspeptin (KP) neurons of the arcuate nucleus (ARC) has been thoroughly studied for its autocrine effect on KP release through κ opioid receptors. Other studies have suggested a role for ß-endorphin (BEND), a peptide cleaved from the pro-opiomelanocortin precursor, on food intake and central control of reproduction. Similar to KP, BEND content in the ARC of sheep is modulated by day length and BEND modulates food intake in a dose-dependent manner. Because KP levels in the ARC vary with photoperiodic and metabolic status, a photoperiod-driven influence of BEND neurons on neighboring KP neurons is plausible. The present study aimed to investigate a possible modulatory action of BEND on KP neurons located in the ovine ARC. Using confocal microscopy, numerous KP appositions on BEND neurons were found but there was no photoperiodic variation of the number of these interactions in ovariectomized, estradiol-replaced ewes. By contrast, BEND terminals on KP neurons were twice as numerous under short days, in ewes having an activated gonadotropic axis, compared to anestrus ewes under long days. Injection of 5 µg BEND into the third ventricle of short-day ewes induced a significant and specific increase of activated KP neurons (16% vs. 9% in controls), whereas the percentage of overall activated (c-Fos positive) neurons, was similar between both groups. These data suggest a photoperiod-dependent influence of BEND on KP neurons of the ARC, which may influence gonadotropin-releasing hormone pulsatile secretion and inform KP neurons about metabolic status.

Early castration in foals: Consequences on physical and behavioural development.

BACKGROUND: The impact of very early castration of foals has not yet been studied despite the many positive effects observed in dogs and cats. OBJECTIVES: The objective of the study was to compare castration at 3 days and 18 months and assess their subsequent morphological and behavioural development. STUDY DESIGN: This was a randomised, blinded clinical study. METHODS: Twenty-two Welsh ponies underwent either early (3-day old, EC group, n = 11) or traditional (18-month old, TC group, n = 11) castration. Animals were followed up to 3 years of age. All ponies were castrated using a primary closure technique under general anaesthesia. Weight and morphometric measurements were monitored monthly from birth until 8 months of age in both groups. Then, measurements were taken every 3 months until 2 years of age and then every 6 months until 3 years of age. Temperament tests were performed on all animals when they were 1- and 3-years old. RESULTS: No differences were observed between the EC and TC groups in terms of physical development from birth until 40 months of age or in terms of temperament and behaviour at either 1 or 3 years of age. MAIN LIMITATIONS: The study included only one breed (Welsh ponies) and only 22 animals that were castrated before 2 years of age, precluding comparison with castration performed at older ages. CONCLUSIONS: We demonstrate that early castration at 3 days does not interfere with morphological or behavioural development.

Chronic low BPS exposure through diet impairs in vitro embryo production parameters according to metabolic status in the ewe.

Bisphenol A (BPA), an endocrine disruptor, has been replaced by structural analogues including bisphenol S (BPS). BPA and BPS exhibited similar effects regarding reproductive functions. Moreover, metabolic status and lipid metabolism are related to female fertility and could worsen BPS effects. The objective was to determine BPS in vivo effects on folliculogenesis and embryo production after chronic exposure through diet, and the influence of metabolic status in adult ewes. Sixty primiparous 2.5 year-old ewes, undergoing a restricted or well fed diet, were exposed to BPS (0, 4 or 50 µg/kg/day) for at least three months. After hormonal oestrus synchronisation and ovarian stimulation, ewes were subjected to ovum pick-up (OPU) procedures to collect immature oocytes, that underwent in vitro maturation, fertilisation and embryo production. Body weight, body condition score and plasma glucose were higher in well-fed compared to restricted ewes, while plasma NEFA was lower during the 4-5 months after the beginning of the diets. Plasma progesterone levels increased on day 5 before OPU session in well-fed compared to restricted ewes. No effect of BPS dose was observed on follicle population, plasma AMH levels and embryo production numbers and rates. However, a significant diet x BPS dose interaction was reported for cleaved embryos, > 4-cell embryos, blastocyst and early blastocyst numbers, and plasma triiodothyronine levels. Our study showed that a contrasted diet did not affect follicle population nor embryo production in adult ewes but could affect the quality and progesterone secretion of the corpus luteum. Chronic low BPS exposure had no effect on follicular population and oocyte competence. Nevertheless, the significant diet x dose interactions observed on embryo production suggest that BPS effect is modulated by metabolic status. Further studies are required to assess the risk of BPS exposure for public reproductive health.

Investigation of equine testis contribution to vitamin D bioactivation.

Although vitamin D acts in various biological processes, it plays a critical role in the maintenance of bone health, and regulates calcium homeostasis. In humans and rodents, the main tissues involved in vitamin D metabolism are the liver and the kidneys, however it has been shown that the testis has strongly participated in its bioactivation. Indeed, in these different species, enzymes metabolizing vitamin D (CYP27A1, CYP27B1 and CYP2R1) have been demonstrated in this tissue. Moreover, men with hypogonadism have shown a decrease in circulating levels of vitamin D. In equine species, the castration of males is a regular practice to reduce the behavior of stallions deemed too aggressive. Castration is carried out at various ages: in foals during their growth or in adulthood once they have reached their optimum size. Although horses exhibit atypical vitamin D metabolism with low circulating levels of vitamin D, it was suggested that testis may contribute to its activation as has been described in rodents and humans; castration could therefore be likely to affect its metabolism. In this study, blood levels of bioactive form of vitamin D (1 α,25[OH] 2 vitamin D 3 ) were measured before and after castration at different ages: 1 wk, after puberty (2 yr) and at adulthood (6 yr). The gene expression of enzymes involved in vitamin D metabolism has been sought in the testis of different experimental groups. No change in bioactive vitamin D3 levels was observed after castration regardless of the age at the time of surgery. The exceptional status of equine species is confirmed with a low or a lack of testis contribution to vitamin D metabolism, regardless of testicular development. This is demonstrated by a low or a lack of signal from enzymes involved in vitamin D bioactivation. Therefore, horses constitute a unique model in comparative endocrinology.

Establishment of a Newborn Lamb Gut-Loop Model to Evaluate New Methods of Enteric Disease Control and Reduce Experimental Animal Use.

Enteric infectious diseases are not all well controlled, which leads to animal suffering and sometimes death in the most severe cases, in addition to economic losses for farmers. Typical symptoms of enteric infections include watery diarrhea, stomach cramps or pain, dehydration, nausea, vomiting, fever and weight loss. Evaluation of new control methods against enteric infections requires the use of many animals. We aimed to develop a new method for an initial in vivo screen of promising compounds against neonatal diseases such as cryptosporidiosis while limiting experimental animal use. We therefore adapted an in vivo method of multiple consecutive but independent intestinal loops to newborn lambs delivered by cesarean section, in which endotoxin responsiveness is retained. This new method allowed for the screening of natural yeast fractions for their ability to stimulate immune responses and to limit early Cryptosporidium parvum development. This model may also be used to investigate host-pathogen interactions and immune responses in a neonatal controlled environment.

Neonatal Suckling, Oxytocin, and Early Infant Attachment to the Mother.

The neuropeptide oxytocin (OT) promotes maternal care and social affiliation in adults but its importance in infant attachment still remains unknown. True animal models of infant attachment are extremely rare, and the sheep (in complement to non-human primates) is one of the few that provides the opportunity to investigate its neuroendocrinological basis. In the lamb, access to the udder has strong rewarding properties for the establishment of a preferential relationship with the mother. Therefore, the present study explored the possible involvement of OT through its release during close social contacts with the mother. The first experiment revealed that lambs having free access to the udder from birth onward developed, by 12 h of age, a clear preference for their mothers over another maternal ewe. Delaying access to the udder for six, four or even only 2 h starting at birth, by covering the ewe's udder, resulted in the lack of such a preference without affecting general activity. These effects persisted in most cases at 24 h but by 72 h of age a bond with the mother was clearly expressed. Experiment two showed that social interactions with the mother were followed by a release of OT in the plasma when lambs had the possibility to suckle. Non-nutritive interactions were without effects. Preliminary data on two subjects suggested that OT might also increase in the cerebrospinal fluid after suckling. Finally, in the third experiment, oral administration of a non-peptide OT receptor antagonist (L-368-899, Merck) over the first 4 h after birth led to decreased exploration of the mother's body compared to lambs receiving saline, and impaired the expression of a preference for the mother at 24 h. The effects were no longer observed at 48 h. Our findings demonstrate that both delayed access to the mother's udder and OT receptor antagonist alter the onset of mother preference in newborn lambs. This suggests that central OT facilitates the development of filial attachment through its release during suckling.

Prenatal programming by testosterone of follicular theca cell functions in ovary.

In mammalian ovaries, the theca layers of growing follicles are critical for maintaining their structural integrity and supporting androgen synthesis. Through combining the postnatal monitoring of ovaries by abdominal magnetic resonance imaging, endocrine profiling, hormonal analysis of the follicular fluid of growing follicles, and transcriptomic analysis of follicular theca cells, we provide evidence that the exposure of ovine fetuses to testosterone excess activates postnatal follicular growth and strongly affects the functions of follicular theca in adulthood. Prenatal exposure to testosterone impaired androgen synthesis in the small antral follicles of adults and affected the expression in their theca cells of a wide array of genes encoding extracellular matrix components, their membrane receptors, and signaling pathways. Most expression changes were uncorrelated with the concentrations of gonadotropins, steroids, and anti-Müllerian hormone in the recent hormonal environment of theca cells, suggesting that these changes rather result from the long-term developmental effects of testosterone on theca cell precursors in fetal ovaries. Disruptions of the extracellular matrix structure and signaling in the follicular theca and ovarian cortex can explain the acceleration of follicle growth through altering the stiffness of ovarian tissue. We propose that these mechanisms participate in the etiology of the polycystic ovarian syndrome, a major reproductive pathology in woman.

Pineal-dependent increase of hypothalamic neurogenesis contributes to the timing of seasonal reproduction in sheep.

To survive in temperate latitudes, species rely on the photoperiod to synchronize their physiological functions, including reproduction, with the predictable changes in the environment. In sheep, exposure to decreasing day length reactivates the hypothalamo-pituitary-gonadal axis, while during increasing day length, animals enter a period of sexual rest. Neural stem cells have been detected in the sheep hypothalamus and hypothalamic neurogenesis was found to respond to the photoperiod. However, the physiological relevance of this seasonal adult neurogenesis is still unexplored. This longitudinal study, therefore aimed to thoroughly characterize photoperiod-stimulated neurogenesis and to investigate whether the hypothalamic adult born-cells were involved in the seasonal timing of reproduction. Results showed that time course of cell proliferation reached a peak in the middle of the period of sexual activity, corresponding to decreasing day length period. This enhancement was suppressed when animals were deprived of seasonal time cues by pinealectomy, suggesting a role of melatonin in the seasonal regulation of cell proliferation. Furthermore, when the mitotic blocker cytosine-b-D-arabinofuranoside was administered centrally, the timing of seasonal reproduction was affected. Overall, our findings link the cyclic increase in hypothalamic neurogenesis to seasonal reproduction and suggest that photoperiod-regulated hypothalamic neurogenesis plays a substantial role in seasonal reproductive physiology.

The impact of thyroid hormone in seasonal breeding has a restricted transcriptional signature.

Thyroid hormone (TH) directs seasonal breeding through reciprocal regulation of TH deiodinase (Dio2/Dio3) gene expression in tanycytes in the ependymal zone of the medio-basal hypothalamus (MBH). Thyrotropin secretion by the pars tuberalis (PT) is a major photoperiod-dependent upstream regulator of Dio2/Dio3 gene expression. Long days enhance thyrotropin production, which increases Dio2 expression and suppresses Dio3 expression, thereby heightening TH signaling in the MBH. Short days appear to exert the converse effect. Here, we combined endocrine profiling and transcriptomics to understand how photoperiod and TH control the ovine reproductive status through effects on hypothalamic function. Almost 3000 genes showed altered hypothalamic expression between the breeding- and non-breeding seasons, showing gene ontology enrichment for cell signaling, epigenetics and neural plasticity. In contrast, acute switching from a short (SP) to a long photoperiod (LP) affected the expression of a much smaller core of 134 LP-responsive genes, including a canonical group previously linked to photoperiodic synchronization. Reproductive switch-off at the end of the winter breeding season was completely blocked by thyroidectomy (THX), despite a very modest effect on the hypothalamic transcriptome. Only 49 genes displayed altered expression between intact and THX ewes, including less than 10% of the LP-induced gene set. Neuroanatomical mapping showed that many LP-induced genes were expressed in the PT, independently of the TH status. In contrast, TH-sensitive seasonal genes were principally expressed in the ependymal zone. These data highlight the distinctions between seasonal remodeling effects, which appear to be largely independent of TH, and TH-dependent localised effects which are permissive for transition to the non-breeding state.

The Two Populations of Kisspeptin Neurons Are Involved in the Ram-Induced LH Pulsatile Secretion and LH Surge in Anestrous Ewes.

Exposure to a ram during spring stimulates luteinizing hormone (LH) secretion and can induce ovulation in sexually quiescent ewes ("ram effect"). Kisspeptin (Kiss) present in the arcuate nucleus (ARC) and the preoptic area (POA) is a potent stimulators of LH secretion. Our aim was to investigate whether Kiss neurons mediate the increase in LH secretion during the ram effect. With double immunofluorescent detection, we identified Kiss neurons (Kiss IR) activated (Fos IR) by exposure to a ram for 2 hours (M2) or 12 hours (M12) or to ewes for 2 hours (C). The density of cells Kiss + Fos IR and the proportion of Kiss IR cells that were also Fos IR cells were higher in M2 and M12 than in C in ARC (P < 0.002) and POA (P < 0.02). In ARC, these parameters were also higher in M12 than in M2 (P < 0.02 and P < 0.05). Kiss antagonist (P234 10-6M) administered by retrodialysis in POA for 3 hours at the time of introduction of the ram reduced the amplitude of the male-induced increase in LH concentration compared with solvent (P < 0.02). In ARC, P234 had a more limited effect (P < 0.038 1 hour after P234) but pulse frequency increased less than after solvent (P = 0.07). In contrast, Kiss antagonist (P271 10-4M) infused in ARC but not POA 6 to 18 hours after introduction of the ram prevented the LH surge in the ewe (0/6 vs 4/5 and 4/6 in C). These results suggest that both populations of Kiss neurons are involved in the ram-induced pulsatile LH secretion and in the LH surge.

Effect of a long-chain n-3 polyunsaturated fatty acid-enriched diet on adipose tissue lipid profiles and gene expression in Holstein dairy cows.

The objective of this study was to determine whether fish oil supplement has an effect on adipose tissue lipid profiles and gene expression in postpartum dairy cows. Holstein cows were supplemented with either long-chain n-3 polyunsaturated fatty acid (PUFA; protected fish oil) or control PUFA (n-6; toasted soybeans) for 2mo after calving (n=23 per diet). These cows showed no difference in milk production or metabolic parameters, but exhibited a tendency toward a decrease in early embryo mortality rate after artificial insemination. We hypothesized that, in addition to this effect, modifications in adipose tissue (AT) gene expression and lipid profiles would occur in response to diet. Subcutaneous AT samples were thus collected from the dewlaps of n-3 and n-6 dairy cows at 1mo antepartum, and 1wk, 2mo, and 5mo postpartum for the analysis of lipids and gene expression. Lipid profiles were obtained by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry in both positive and negative modes. We found 37 lipid species in the 200 to 1,200 m/z range, which differed between the n-3 and control groups, suggesting that the n-3 supplement affected the lipid composition through the enrichment of lipids integrating long-chain PUFA from fish oil sources: eicosapentaenoic and docosahexaenoic acid. Moreover, a decrease in triacylglycerolipids was observed in AT of n-3 supplemented cows. The expression of 44 genes involved in fatty acid metabolism and the adipokine system was assessed by real-time reverse-transcription PCR. Hierarchical clustering, according to either postpartum stage or diet, enabled us to group genes exhibiting similar kinetic properties during lactation or by those that varied in similar ways after n-3 supplementation, respectively. Among the genes exhibiting a dietary effect, FABP4, LIPE, CD36, and PLIN1 were overexpressed in n-3 AT samples compared with the control, suggesting an increase in lipolysis due to n-3 supplementation, which was reflected on lipolytic activity at the protein level (i.e., protein expression of fatty acid binding protein 4, phosphorylated perilipin 1, and phosphorylated hormone-sensitive lipase). This increase in lipolysis is relevant to the decrease in triglycerides observed in these samples. Gene expression analyses between n-3 and control AT samples also suggested that the n-3 diet could modulate the secretory functions of AT, possibly by affecting adipokine expression; however, this has to be confirmed at the protein level.

Influence of transvaginal ultrasound-guided follicular punctures in the mare on heart rate, respiratory rate, facial expression changes, and salivary cortisol as pain scoring.

Transvaginal ultrasound-guided follicular punctures are widely used in the mare for diagnosis, research, and commercial applications. The objective of our study was to determine their influence on pain, stress, and well-being in the mare, by evaluating heart rate, breath rate, facial expression changes, and salivary cortisol before, during, and after puncture. For this experiment, 21 pony mares were used. Transvaginal ultrasound-guided aspirations were performed on 11 mares. After injections for sedation, analgesia, and antispasmodia, the follicles from both ovaries were aspirated with a needle introduced through the vagina wall into the ovary. In the control group, 10 mares underwent similar treatments and injections, but no follicular aspiration. Along the session, heart rate and breath rate were evaluated by a trained veterinarian, ears position, eyelid closure, and contraction of facial muscles were evaluated, and salivary samples were taken for evaluation of cortisol concentration. A significant relaxation was observed after sedative injection in the punctured and control mares, according to ear position, eyelid closure, and contraction of facial muscles, but no difference between punctured and control animals was recorded. No significant modification of salivary cortisol concentration during puncture and no difference between punctured and control mares at any time were observed. No significant modification of the breath rate was observed along the procedure for the punctured and the control mares. Heart rate increased significantly but transiently when the needle was introduced in the ovary and was significantly higher at that time for the punctured mares than that for control mares. None of the other investigated parameters were affected at that time, suggesting discomfort is minimal and transient. Improving analgesia, e.g., through a multimodal approach, during that possibly more sensitive step could be recommended. The evaluation of facial expression changes and heart rate is easy-to-use and accurate tools to evaluate pain and well-being of the mare.

The mycotoxins deoxynivalenol and nivalenol show in vivo synergism on jejunum enterocytes apoptosis.

The mycotoxins deoxynivalenol (DON) and nivalenol (NIV), worldwide cereal contaminants, raise concerns for human and animal gut health, following exposure through contaminated food and feed. The aim of this work was to analyze the effects of DON and NIV, alone or associated, on the intestinal pig mucosa. Jejunal loops were used for testing DON and NIV individually and in combination (1:1) after a single exposure, for 24 h. For repeated exposure, piglets received a natural contaminated feed, with DON or with DON + NIV for 28 days. Histological investigations, proliferation and apoptosis assessments were conducted. Both experiments were concordant for the total-cell proliferation decreased at the villus tips after DON or DON + NIV at 10 µM acutely, or repeatedly, by 30-35% and 20-25%, respectively. In loops model, apoptotic enterocytes at villus tips increased dose-dependently after DON, NIV alone or DON + NIV in combination. The combination in loops at 10 µM showed higher effects on proliferation and apoptosis than DON alone, and synergism was shown for villus apoptotic enterocyte. These results are to be considered for NIV consumer risk assessment. Our results demonstrate the in vivo disruption of the intestinal balance proliferation/apoptosis explaining, at least partly, the disruption of intestinal barrier by these mycotoxins.

Effect of a long chain n-3 PUFA-enriched diet on production and reproduction variables in Holstein dairy cows.

The objective of this study was to determine the effect of a rumen-protected fish oil supplement on the production and reproduction variables in postpartum dairy cows. Holstein cows (n=46) were given a basal total mixed diet plus one PUFA supplement: n-3 (n-3; protected fish oil; 1% dry matter intake (DMI); n=23) or control (n-6; toasted soybeans; 1.8% DMI; n=23), in a switchback design over two consecutive lactations. Supplements were added to the diet between calving and 2 months after calving to assess the effect on growth and maturation of ovarian follicles from which ovulation occurred around the day of insemination. Body weight (BW), milk yield (MY) and composition, dry matter intake (DMI), energy balance (EB), subcutaneous fat thickness, plasma fatty acid composition, plasma nonesterified fatty acids (NEFA), glucose and urea concentrations, follicular activity, embryo mortalities and fertility (conception rate after first AI, AI1) were assessed. BW, MY, DMI, plasma NEFA, glucose and urea were unaffected by the diet. There was a trend of an increased number of large follicles (diameter≥10mm) with the n-3 dietary supplementation (P=0.06) and a decrease in infertility or early embryo mortality rate 21 days after AI, 13.5% in the n-3 compared with 38.8% in the n-6 group (P=0.09), with no effect on the conception rate at 35d or 90d after AI1. These data suggest that the effect seen on ovarian variables is not associated with an effect on production and metabolic variables and is specific to n-3 PUFA supplementation. Further studies are necessary to determine whether DHA or EPA enhances fertility in lactating dairy cattle.

Nivalenol has a greater impact than deoxynivalenol on pig jejunum mucosa in vitro on explants and in vivo on intestinal loops.

The mycotoxins deoxynivalenol (DON) and nivalenol (NIV), worldwide cereal contaminants, raise concerns for animal and human gut health, following contaminated food or feed ingestion. The impact of DON and NIV on intestinal mucosa was investigated after acute exposure, in vitro and in vivo. The histological changes induced by DON and NIV were analyzed after four-hour exposure on pig jejunum explants and loops, two alternative models. On explants, dose-dependent increases in the histological changes were induced by DON and NIV, with a two-fold increase in lesion severity at 10 µM NIV. On loops, NIV had a greater impact on the mucosa than DON. The overall proliferative cells showed 30% and 13% decrease after NIV and DON exposure, respectively, and NIV increased the proliferative index of crypt enterocytes. NIV also increased apoptosis at the top of villi and reduced by almost half the proliferative/apoptotic cell ratio. Lamina propria cells (mainly immune cells) were more sensitive than enterocytes (epithelial cells) to apoptosis induced by NIV. Our results demonstrate a greater impact of NIV than DON on the intestinal mucosa, both in vitro and in vivo, and highlight the need of a specific hazard characterization for NIV risk assessment.

Resistin in dairy cows: plasma concentrations during early lactation, expression and potential role in adipose tissue.

Resistin is an adipokine that has been implicated in energy metabolism regulation in rodents but has been little studied in dairy cows. We determined plasma resistin concentrations in early lactation in dairy cows and investigated the levels of resistin mRNA and protein in adipose tissue and the phosphorylation of several components of insulin signaling pathways one week post partum (1 WPP) and at five months of gestation (5 MG). We detected resistin in mature bovine adipocytes and investigated the effect of recombinant bovine resistin on lipolysis in bovine adipose tissue explants. ELISA showed that plasma resistin concentration was low before calving, subsequently increasing and reaching a peak at 1 WPP, decreasing steadily thereafter to reach pre-calving levels at 6 WPP. Plasma resistin concentration was significantly positively correlated with plasma non esterified fatty acid (NEFA) levels and negatively with milk yield, dry matter intake and energy balance between WPP1 to WPP22. We showed, by quantitative RT-PCR and western blotting, that resistin mRNA and protein levels in adipose tissue were higher at WPP1 than at 5 MG. The level of phosphorylation of several early and downstream insulin signaling components (IRß, IRS-1, IRS-2, Akt, MAPK ERK1/2, P70S6K and S6) in adipose tissue was also lower at 1 WPP than at 5 MG. Finally, we showed that recombinant bovine resistin increased the release of glycerol and mRNA levels for ATGL (adipose triglyceride lipase) and HSL (hormone-sensitive lipase) in adipose tissue explants. Overall, resistin levels were high in the plasma and adipose tissue and were positively correlated with NEFA levels after calving. Resistin is expressed in bovine mature adipocytes and promotes lipid mobilization in adipose explants in vitro.

Expression of adipokine and lipid metabolism genes in adipose tissue of dairy cows differing in a female fertility quantitative trait locus.

We have previously characterized 2 haplotypes (Fertil+ and Fertil-) of Holstein dairy cows differing in 1 female fertility quantitative trait locus (QTL) located on chromosome 3 (QTL-Fert-F-BTA3) between positions 9.8 and 13.5 cM. This QTL is composed of 124 genes, some of them being involved in metabolism or reproduction. Primiparous Fertil+ and Fertil- cows exhibited 69 and 39% pregnancy rate at first service, respectively. A difference in plasma nonesterified fatty acid concentrations observed between both haplotypes might indicate a difference in adipose tissue mobilization. We compared adipose tissue gene expression in Fertil+ and Fertil- cows during their second lactation, at 2 physiological stages, implying either intense lipid mobilization (1 wk postpartum) or fat storage (5 mo of gestation). We investigated by reverse-transcription quantitative PCR the mRNA gene expression of 5 positional candidate genes located in the QTL-Fert-F-BTA3, as well as 18 other functional candidate genes encoding proteins involved in lipid metabolism and several adipokines. Among them, genes involved in either lipolysis or lipogenesis were chosen as controls because they were previously described in dairy cow adipose tissue. A hierarchical clustering was performed to group genes according to their expression pattern, allowing 2 clusters to be determined. Cluster 1 was composed of genes that were overexpressed during mobilization (ADIPOQ, ADIPOR2, LIPE, FABP4, PLIN1, RARRES, LEPR, and CPT1A) and cluster 2 of genes overexpressed during reconstitution of body reserves (ACACA, FASN, and SCD). Genes belonging to cluster 1 (LIPE, FABP4, PLIN1, and CPT1A) are known to be involved in lipolysis and fatty acid oxidation, and genes belonging to cluster 2 (ACACA, FASN, and SCD) are known to be involved in fatty acid synthesis. The expression of 5 genes from cluster 1 was correlated to plasma nonesterified fatty acid levels and thus to mobilization of body reserves in dairy cows (ADIPOQ, ADIPOR2, LIPE, PLIN1, and FABP4). During the mobilization stage, none of the positional candidate genes belonging to QTL-Fert-F-BTA3 (ADAR, MTX1, SHC1, SPTA1, and PAQR6) showed a difference in expression between the 2 haplotypes. Interestingly, ADIPOQ and ADIPOR2 were the only genes showing a significant mRNA overexpression in Fertil- cows at the mobilization stage. Further studies focusing on plasma adiponectin level and adipokine actions on the ovary are needed to investigate its potential role in dairy cow fertility.

An experimental study to evaluate the Fluobeam 800 imaging system for fluorescence-guided lymphatic imaging and sentinel node biopsy.

BACKGROUND: Visualization of the lymphatic system is challenging. Lymphatic imaging is a crucial diagnostic tool for benign and malignant lymphatic pathologies. Fluorescence-guided imaging allows selective lymphatic mapping and sentinel lymph node (SLN) identification. There are a few fluorescence systems, but some drawbacks remain due to technical and ergonomic aspects. The aim of this study was to evaluate the feasibility of the new Fluobeam 800 imaging system. METHODS: After approval by the ethics committee, the system was evaluated for lymphography and SLN biopsy in an animal model. Five pigs each with 4 lymph node (LN) stations (n = 20 LN stations) were subjected to lymphatic imaging using indocyanine green (ICG). Additionally, the use of ICG was compared with ICG adsorbed to human serum albumin (ICG-HSA). Lymphatic vessels and SLN identification rates were measured. RESULTS: After injection, a clear fluorescence signal of the lymphatic vessels was visualized leading to the LN station. Overall, ICG fluorescence imaging identified a mean of 2.0 lymphatic vessels and 1.1 (range = 1-2) SLN in 20 of 20 LN stations. Reverse lymphography was feasible. A clinical difference in resolution was not detected between use of ICG-HSA and ICG. CONCLUSION: This is the first study analyzing the feasibility of the Fluobeam 800 imaging system allowing transcutaneous real-time imaging. It enables detection of the SLN by fluorescence retention with increased detection depth and resolution. After fixation to the ceiling, the ergonomics advanced for simultaneous field navigation and dissection. The new system can be applied for lymphatic imaging for lympatico-reconstructive surgery and SLN biopsy.

Deoxynivalenol as a new factor in the persistence of intestinal inflammatory diseases: an emerging hypothesis through possible modulation of Th17-mediated response.

BACKGROUND/AIMS: Deoxynivalenol (DON) is a mycotoxin produced by Fusarium species which is commonly found in temperate regions worldwide as a natural contaminant of cereals. It is of great concern not only in terms of economic losses but also in terms of animal and public health. The digestive tract is the first and main target of this food contaminant and it represents a major site of immune tolerance. A finely tuned cross-talk between the innate and the adaptive immune systems ensures the homeostatic equilibrium between the mucosal immune system and commensal microorganisms. The aim of this study was to analyze the impact of DON on the intestinal immune response. METHODOLOGY: Non-transformed intestinal porcine epithelial cells IPEC-1 and porcine jejunal explants were used to investigate the effect of DON on the intestinal immune response and the modulation of naive T cells differentiation. Transcriptomic proteomic and flow cytometry analysis were performed. RESULTS: DON induced a pro-inflammatory response with a significant increase of expression of mRNA encoding for IL-8, IL-1α and IL-1ß, TNF-α in all used models. Additionally, DON significantly induced the expression of genes involved in the differentiation of Th17 cells (STAT3, IL-17A, IL-6, IL-1ß) at the expenses of the pathway of regulatory T cells (Treg) (FoxP3, RALDH1). DON also induced genes related to the pathogenic Th17 cells subset such as IL-23A, IL-22 and IL-21 and not genes related to the regulatory Th17 cells (rTh17) such as TGF-ß and IL-10. CONCLUSION: DON triggered multiple immune modulatory effects which could be associated with an increased susceptibility to intestinal inflammatory diseases.