Effects of Different Oral Doses of Sodium Chloride on the Basal Acid-Base and Mineral Status of Exercising Horses Fed Low Amounts of Hay

The provision of NaCl, according to current recommendations, to horses in moderate work has been shown to induce immediate postprandial acidosis. The present study aimed to clarify whether this NaCl induced acidosis i) persists beyond the immediate postprandial period, and ii) is still present after a 2 week adaptation period. Six adult warmblood mares in moderate work received daily 1.00 kg hay per 100 kg body weight (bwt) only together with 0.64 kg unprocessed cereal grains/100 kg bwt. d as fed basis. Using a 3x3 Latin Square, either 0 (NaCl-0), 50 (NaCl-50) or 100 (NaCl-100) g NaCl/d were fed together with the concentrates in two equal doses for 3 weeks. During the final week, a mineral digestibility trial was undertaken. The middle sodium and chloride intake (NaCl-50) at least met the most common recommendations for moderate work. Morning (7: 00 AM) urine and venous blood samples were collected on days 0, 1-4, 8, and 15, and analysed for pH, acid-base status, creatinine and electrolyte concentrations. Fractional electrolyte clearances (FC) were determined. Mean apparent sodium digestibility ranged between 60-62% whereas chloride digestibility was consistently above 94%. Supplementing 100 g but not 50 g of NaCl resulted in significant reduction of blood pH and base excess as well as urinary pH and urine acid excretion. Both 50 g and 100 g NaCl supplementation caused a significant reduction in base and net acid-base excretion, urine density and potassium concentration, but increased urine sodium concentration and the FC of sodium and chloride (P < 0.05). This suggests that a high proportion of the recommended salt doses is excreted renally. The above effects of NaCl supplementation persisted over the 2 week measurement period. Results suggest that feeding 100 g NaCl to moderately exercising horses results in mild metabolic acidosis, whereas feeding 50 g according to current recommendations resulted in compensated acidosis

Amniotic fluid INSL3 measured during the critical time window in human pregnancy relates to cryptorchidism, hypospadias and phthalate load: a large case-control study

The period of the first to second trimester transition in human pregnancy represents a sensitive window for fetal organogenesis, particularly in regard to the development of the male reproductive system. This is a time of relative analytical inaccessibility. We have used a large national biobank of amniotic fluid samples collected at routine amniocentesis to determine the impacts of exogenous endocrine disruptor load on specific fetal biomarkers at this critical time. While adrenal and testicular steroids are highly correlated, they are also mostly positively influenced by increasing phthalate load, represented by the metabolites 7cx-MMeHP and 5cx-MEPP, by PFOS exposure, and by smoking, suggesting an adrenal stress response. In contrast, the testis specific biomarkers INSL3 and androstenedione are negatively impacted by the phthalate endocrine disruptors. Using a case-control design, we show that cryptorchidism and hypospadias are both significantly associated with increased amniotic concentration of INSL3 during gestational weeks 13 to 16, and some, though not all steroid biomarkers. Cases are also linked to a specifically increased variance in the Leydig cell biomarker INSL3 compared to controls, an effect exacerbated by maternal smoking. No influence of phthalate metabolites or PFOS was evident on the distribution of cases and controls. Considering that several animal and human studies have shown a negative impact of phthalate load on fetal and cord blood INSL3, respectively, the present results suggest that such endocrine disruptors may rather be altering the relative dynamics of testicular development and consequent hormone production, leading to a desynchronization of tissue organization during fetal development. Being born small for gestational age appears not to impact on the testicular biomarker INSL3 in second trimester amniotic fluid

The male fetal biomarker INSL3 reveals substantial hormone exchange between fetuses in early pig gestation

The peptide hormone INSL3 is uniquely produced by the fetal testis to promote the transabdominal phase of testicular descent. Because it is fetal sex specific, and is present in only very low amounts in the maternal circulation, INSL3 acts as an ideal biomarker with which to monitor the movement of fetal hormones within the pregnant uterus of a polytocous species, the pig. INSL3 production by the fetal testis begins at around GD30. At GD45 of the ca.114 day gestation, a time at which testicular descent is promoted, INSL3 evidently moves from male to female allantoic compartments, presumably impacting also on the female fetal circulation. At later time-points (GD63, GD92) there is less inter-fetal transfer, although there still appears to be significant INSL3, presumably of male origin, in the plasma of female fetuses. This study thus provides evidence for substantial transfer of a peptide hormone between fetuses, and probably also across the placenta, emphasizing the vulnerability of the fetus to extrinsic hormonal influences within the uterus

Relaxin-like peptides in male reproduction: a human perspective

The relaxin family of peptide hormones and their cognate GPCRs are becoming physiologically well-characterized in the cardiovascular system and particularly in female reproductive processes. Much less is known about the physiology and pharmacology of these peptides in male reproduction, particularly as regards humans. H2-relaxin is involved in prostate function and growth, while insulin-like peptide 3 (INSL3) is a major product of the testicular Leydig cells and, in the adult, appears to modulate steroidogenesis and germ cell survival. In the fetus, INSL3 is a key hormone expressed shortly after sex determination and is responsible for the first transabdominal phase of testicular descent. Importantly, INSL3 is becoming a very useful constitutive biomarker reflecting both fetal and post-natal development. Nothing is known about roles for INSL4 in male reproduction and only very little about relaxin-3, which is mostly considered as a brain peptide, or INSL5. The former is expressed at very low levels in the testes, but has no known physiology there, whereas the INSL5 knockout mouse does exhibit a testicular phenotype with mild effects on spermatogenesis,probably due to a disruption of glucose homeostasis. INSL6 is a major product of male germ cells, although it is relatively unexplored with regard to its physiology or pharmacology, except that in mice disruption of the INSL6 gene leads to a disruption of spermatogenesis. Clinically, relaxin analogues may be useful in the control of prostate cancer, and both relaxin and INSL3 have been considered as sperm adjuvants for in vitro fertilization

Studies on oocyte quality of different follicle populations in mares

1 Einleitung und Zielstellung 1 2 Literaturübersicht 3 2.1 Physiologie der Fortpflanzung bei Stuten 3 2.2 Follikulogenese 6 2.3 Funktionelle Follikelentwicklung 9 2.4 Eizellreifung 14 2.5 Bedeutung der Mitochondrien in Oozyten während der Maturation 22 2.6 Bedeutung der Kumuluszellen für die Prozesse der Eizellreifung 24 2.7 Parameter zur Bestimmung der Eizellqualität 26 2.8 In-vivo-Oozytengewinnung bei Stuten 33 3 Material und Methode 37 4\. Ergebnisse 54 4.1 Kumulus-Oozyten-Komplex-Gewinnung mittels transvaginaler ultraschallgeleiteter Follikelpunktion 54 4.2. Steroidkonzentrationen in den Follikelflüssigkeiten der unterschiedlichen Follikelpopulationen 56 4.3 Aktivität der Glukose-6-Phosphat-Dehydrogenase der gewonnenen Oozyten 58 4.4 Untersuchungen zur Chromatinkonfiguration der Oozyten 63 4.5 Untersuchungen zur mitochondrialen Aktivität in den gewonnenen Oozyten 70 4.6 Untersuchungen zur mitochondrialen Aggregation der gewonnenen Oozyten 76 5 Diskussion 82 6 Zusammenfassung 106 7 Summary 108 8 Zitierte Literatur 110 9 Publikationen 124 10 Danksagung 125 11 Selbstständigkeitsversicherung 126Ziel des tierexperimentellen Designs war es Eizellen in vivo zu gewinnen, die verschiedenen, definierten physiologischen Entwicklungsstadien von Follikelpopulationen zugeordnet werden konnten. Durch die parallele Bestimmung mehrerer Parameter der Eizellqualität an jeder einzelnen Eizelle, sollten Zusammenhänge in vivo zwischen chromosomalen und zytoplasmatischen Reifungsprozessen in den Oozyten untersucht werden. Insbesondere sollte dabei die follikuläre Herkunft als Einflussfaktor auf die Eigenschaften von Oozyten näher beschrieben werden. Die Oozytengewinnung erfolgte durch wiederholte transvaginale ultraschallgeleitete Follikelpunktionen, die in 120 Follikelpunktionssitzungen an 14 Mecklenburger Warmblutstuten während einer Zuchtsaison vorgenommen wurden. Die Eizellgewinnung erfolgte zunächst bei rossigen Stuten, denen 24 Stunden zuvor ein hCG-Präparat appliziert worden war. Die Follikelaspirate wurden getrennt nach präovulatorischen Follikeln und subordinanten Follikeln gesammelt. Nach einer Ablation aller sichtbaren Follikel, wurden die Stuten einer zweiten Follikelaspiration unterzogen, noch bevor sich in der neu herangewachsenen Follikelpopulation ein dominanter Follikel entwickelt hatte. Zur genaueren Charakterisierung der verschiedenen Follikelgruppen erfolgten stichprobenartig Analysen der Östradiol-17β- und Progesteronkonzentrationen in den Follikelflüssigkeiten. Die Kumulus-Oozyten- Komplexe (KOK) wurden nach der Gewinnung unter einem Stereomikroskop morphologisch beurteilt und mit Brillant-Cresyl-Blau (BCB) unter Kulturbedingungen zur Bestimmung der Aktivität der Glukose-6-Phosphat- Dehydrogenase (G-6-PDH) inkubiert. Anschließend wurden die denudierten Oozyten mit dem Fluoreszenzfarbstoff MitoTracker Orange CMTM Ros zur Bestimmung der mitochondrialen Aktivität und Aggregation vital inkubiert und nach einer Fixierung in Paraformaldehyd mit Hoechst 33258 zur Bestimmung der Chromatinkonfiguration parallel gefärbt. Die Östradiol-17β-Konzentrationen waren mit 1911,4±184,5 ng/ml in präovulatorischen Follikeln, mit 885,6±120,6 ng/ml in Follikeln der wachsenden und mit 54,4±155,9 ng/ml in Follikeln der subordinanten Follikelgruppe zwischen den Follikelgruppen signifikant unterschiedlich (p<0,05). Kompakte KOK wurden signifikant häufiger aus wachsenden Follikelgruppen (p<0,05) und Oozyten, die nur die Corona radiata aufwiesen tendenziell häufiger aus den subordinanten Follikelgruppen (p=0,06) gewonnen. Insgesamt wurde mittels der BCB-Färbung in einem Drittel der Oozyten eine hohe G-6-PDH-Aktivität festgestellt. Es wurden keine signifikanten Einflüsse der Follikelgruppen, der KOK-Morphologie, des mitochondrialen Aggregationsmusters oder der mitochondrialen Aktivität auf die Verteilung von Oozyten mit unterschiedlicher G-6-PDH-Aktivität beobachtet. Bei der Untersuchung der Chromatinkonfigurationen konnte gezeigt werden, dass wachsende Follikelpopulationen signifikant höhere Anteile Oozyten im fibrillären Diplotänstadium enthalten, während Oozyten aus subordinanten Follikelpopulationen gehäuft ein kondensiertes Diplotän aufwiesen (p<0,05). Oozyten, deren Kern sich im fibrillären Diplotänstadium befand, wiesen einen signifikant größeren Anteil Oozyten mit einer niedrigen G-6-PDH-Aktivität und Oozyten mit pyknotischen Chromatin einen signifikant größeren Anteil Oozyten mit einer hohen G-6-PDH-Aktivität auf (p<0,05). Oozyten, deren Chromatin pyknotisch war, fanden sich nur in subordinanten Follikelgruppen. In Oozyten aus wachsenden Follikelgruppen wurden signifikant höhere Werte (Fluoreszenzintensität/Oozyte) der mitochondrialen Aktivität gemessen als in Oozyten aus subordinanten Follikelgruppen (p<0,05). Oozyten, die nur eine Corona radiata aufwiesen und Oozyten der expandierten KOK zeigten ebenfalls signifikant höhere Werte der mitochondrialen Aktivität als Oozyten mit kompaktem Kumulus (p<0,05). Oozyten mit grob granulierten Mitochondrien zeigten eine signifikant höhere mitochondriale Aktivität als Oozyten mit fein granulierten Mitochondrien (p<0,0001). Insgesamt zeigten 28,1% der untersuchten Oozyten ein grobgranuliertes mitochondriales Aggregationsmuster und 71,8% ein fein granuliertes Aggregationsmuster. Ein signifikanter Einfluss der follikulären Herkunft oder der Chromatinkonfiguration der Oozyten auf ihr mitochondriales Aggregationsmuster wurde nicht beobachtet. Die Untersuchungsergebnisse zeigten deutlich, dass in subordinanten Follikelgruppen hauptsächlich atretische und in den wachsenden Follikelgruppen zum größten Teil vitale Follikel enthalten waren. Aufgrund der Ergebnisse kann angenommen werden, dass es bereits in unreifen Follikeln zu einer Entkopplung zwischen der chromosomalen und zytoplasmatischen Reifung in Oozyten kommen kann. Besonders in stark atretischen Follikeln könnte dieser Prozess bereits irreversibel fortgeschritten sein. Oozyten mit der gleichen Chromatinkonfiguration oder Kumulusmorphologie sollten daher bei unterschiedlicher follikulärer Herkunft hinsichtlich ihrer Entwicklungskompetenz in vitro nicht als gleichwertig betrachtet werden. Da die follikuläre Herkunft chromosomale und zytoplasmatische Eigenschaften von equinen Oozyten signifikant beeinflusst, kann das Wissen über die follikuläre Herkunft einer Oozyte als weiterer Parameter zur Abschätzung der Oozytenqualität dienen. Die erzielten Ergebnisse können in Zukunft dazu beitragen In-vitro-Systeme der Eizellreifung zu optimieren und die Oozytengewinnung auf eine möglichst hohe Anzahl entwicklungskompetenter Oozyten zu orientieren.The aim of this study was to recover equine oocytes in vivo, which could be assigned to follicle populations in different physiologic developmental stages. By a parallel determination of several parameters of oocyte quality for every single oocyte, links between chromosomal and cytoplasmic maturation processes in vivo should be investigated. Especially the impact of the different follicular origins on the characteristics of oocytes should be described in detail. Oocyte recovery was done by repeated transvaginal ultrasound guided follicle aspiration in 14 Mecklenburger Warmblood mares, which underwent 120 follicle aspiration sessions during one breeding season. Follicle aspiration sessions were performed first during the heat of the mares, 24 hours after they had received hCG. The aspirates of the preovulatory follicles and the subordinate follicle populations were collected separately. After an ablation of all visible follicles of the mare, a second follicle aspiration session was done just before a dominant follicle had developed in the newly grown progressive follicle population. To characterise the follicle populations more closely random samples of follicle fluid from the different follicle populations were taken simultaneously and analysed for their oestradiol and progesterone content. Immediately after recovery the cumulus- oocytes-complexes (COCs) were divided in groups depending on their cumulus morphology under a stereomicroscope and incubated under culture conditions with brilliant cresyl blue (BCB), to evaluate the activity of glucose-6-phosphate dehydrogenase (G-6-PDH) enzyme. The denuded oocytes were then incubated with the Mito Tracker Orange CMTM Ros vital stain to measure the mitochondrial activity and aggregation. Following a fixation with paraformaldehyde the oocytes were also subjected to Hoechst 33258 stain to detect the chromatin configuration. The oestradiol concentrations showed a level of 1911.7 ± 185.5 ng/ml in preovulatory follicles, 885.6 ± 120.6 ng/ml in growing and 54.4 ± 155.9 ng/ml in subordinate follicle populations indicating significant differences between the follicular fluid samples of the different follicle populations (p < 0.05). Significantly more oocytes with compact cumuli were found in the progressive follicle population (p < 0.05), whereas the subordinate follicle population tended to contain more oocytes, which were only covered by corona radiata cells (p = 0.06). By the measurement with BCB stain, altogether one third of the recovered oocytes showed a high G-6-PDH activity. No significant impacts of the follicle population of origin, the cumulus morphology, the mitochondrial aggregation patterns or the mitochondrial activity of the oocytes were observed on the distribution patterns of oocytes with different G-6-PDH activity. The evaluation of the chromatin configuration showed that follicles of growing follicle populations contained significantly higher proportions of oocytes with a fibrillar diplotene, while subordinante follicle populations had higher proportions of oocytes with a condensed diplotene (p < 0.05). While oocytes with the chromatin configuration of a fibrillar diplotene showed a significantly higher amount of oocytes with a low G-6-PDH activity, oocytes with pycnotic chromatin had a significantly higher number of oocytes with a high G-6-PDH activity (p < 0.05). Oocytes with a pycnotic chromatin were found in subordinate follicle populations only. The mitochondrial activity (fluorescence intensity/oocyte) was significantly increased in oocytes of growing follicle populations as compared to oocytes from subordinate follicle populations (p < 0.05). Higher levels of mitochondrial activity were found also in oocytes with an expanded cumulus and in oocytes with only corona radiata cells than in the oocytes with a compact cumulus (p < 0.05). The mitochondrial activity was significantly higher in oocytes which showed mitochondria in granulated aggregation pattern than in oocytes with fine mitochondrial aggregation pattern (P<0,0001). In total, 28.1% of the oocytes showed a granulated mitochondrial aggregation pattern and 71.8% a fine aggregation pattern of the mitochondria but no significant impact of the follicle population of origin or the chromatin configuration were found. The results of this study show clearly that subordinate follicle populations consisted mainly of atretic follicles while the growing follicle population consisted of a greater number of viable follicles. Based on the results it could be supposed that already in immature follicles chromosomal and cytoplasmic maturation processes of the enclosed oocytes can be teared apart. Especially in follicles with progressed atresia this deviation process could have already reached an irreversible stage. Therefore, oocytes with the same chromatin configuration or the same cumulus morphology but with a different follicular origin should not be taken as having the same developmental competence. Because of the significant impact of the follicular origin on chromosomal and cytoplasmic characteristics of equine oocytes, the knowledge about the follicular origin of an oocyte can be used as a parameter to assess the quality of an oocyte. The results can contribute to optimize in vitro oocyte maturation systems and should be helpful to design oocyte-recovery treatments in vivo, which results in highest numbers of developmental competent oocytes

Markierungen XVIII

Premi 200

Ultrasonographic Monitoring of Fetal Growth and Fetal Weight Calculation in Sows during Gestation

Ultrasound examinations offer the possibility to monitor fetal growth and estimate fetal weight, but reference data for such techniques in pigs are rare. The aim of this study was therefore to identify suitable anatomical fetal structures for monitoring physiological growth dynamics by ultrasound examinations and to estimate fetal weight using appropriate mathematical models. For this purpose, 198 fetuses of 15 primiparous Landrace sows were examined by ultrasound on days 36, 50, 64, 79 and 92 of gestation in live sows and in utero after slaughter. Biparietal distance (BPD), rostro-occipital distance (ROD), corpus vitreum diameter, heart length, abdominal circumference (AC) and transverse and sagittal abdominal diameter were determined by ultrasound in utero, and the fetuses were subsequently ex uteri determined and weighed. Reference curves for the continuous increase in fetal parameters over the pregnancy were established. Weight estimation could be performed with linear models at a known stage of pregnancy using one or a combination of parameters. Cubic equations were developed to describe the relationships between body measurements and weight over the course of gestation. BPD, ROD and AC have been shown to be the most suitable parameters for fetal weight estimation, but in live sows, only the fetal head parameters could be easily and reliably determined. These techniques could initially be of interest for research into fetal growth, but future application in veterinary practice is also conceivable

Data on piglet bodyweight_body temperature_catheter functionality_plasma xylose concentration_after jugular vein catheter surgery

&lt;p&gt;Data on piglet bodyweight and body temperature before and after jugular vein&nbsp;catheter surgery in suckling piglets at the age of 10 to 15 days compared to control piglets without catheter surgery.&lt;/p&gt; &lt;p&gt;Data on plasma xylose concentration in samples from jugular vein catheter and catheter functionality in suckling piglets..&lt;/p&gt; &lt;p&gt;Supplementary material including surgical materials and pictures_De Leonardis et al. animal open space&lt;/p&gt

Data on piglet bodyweight_body temperature_catheter functionality_plasma xylose concentration_after jugular vein catheter surgery

Data on piglet bodyweight and body temperature before and after jugular vein catheter surgery in suckling piglets at the age of 10 to 15 days compared to control piglets without catheter surgery. Data on plasma xylose concentration in samples from jugular vein catheter and catheter functionality in suckling piglets.. Supplementary material including surgical materials and pictures_De Leonardis et al. animal open spac