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Felids generally follow a poly-estrous reproductive strategy. Eurasian lynx (Lynx lynx) display a different pattern of reproductive cyclicity where physiologically persistent corpora lutea (CLs) induce a mono-estrous condition which results in highly seasonal reproduction. The present study was based around a sono-morphological and endocrine study of captive Eurasian lynx, and a control-study on free-ranging lynx. We verified that CLs persist after pregnancy and pseudo-pregnancy for at least a two-year period. We could show that lynx are able to enter estrus in the following year, while CLs from the previous years persisted in structure and only temporarily reduced their function for the period of estrus onset or birth, which is unique among felids. The almost constant luteal progesterone secretion (average of 5 ng/ml serum) seems to prevent folliculogenesis outside the breeding season and has converted a poly-estrous general felid cycle into a mono-estrous cycle specific for lynx. The hormonal regulation mechanism which causes lynx to have the longest CL lifespan amongst mammals remains unclear. The described non-felid like ovarian physiology appears to be a remarkably non-plastic system. The lynx's reproductive ability to adapt to environmental and anthropogenic changes needs further investigation.

Die vorliegende Arbeit beschäftigt sich mit den physiologischen Veränderungen porciner Spermatozoen, die durch einen metabolischen Einbau von Fettsäuren in Spermatozoenlipide hervorgerufen werden. Ziel dieser Arbeit war die Untersuchung der metabolischen Aufnahme von Fettsäuren in die Spermatozoenlipide und die Bewertung des physiologischen Zustandes porciner Spermatozoen mit Hinblick auf die Niedrigtemperaturlagerung. Alle in den porcinen Spermatozoen vorkommenden Lipide wurden mittels GC und MALDI-TOF-MS analysiert. Hauptvertreter der polaren Lipidklassen sind Glycerophospholipide (GPC, GPE). Der Hauptvertreter der neutralen Lipidklassen ist Diacylglycerol (DAG). Die metabolische Aufnahme von Fettsäuren in die Lipide wurde durch die Supplementierung des Flüssigkonservierungsmediums mit [14C]-Octadecadiensäure radiochemisch untersucht. Anhand dieser Experimente wurde gezeigt, dass die Temperatur und die Inkubationsdauer wichtige Faktoren für die metabolische Aufnahme dieser Radiochemikalie in die Spermatozoenlipide sind. Die zugesetzten Fettsäuren werden sowohl in die neutralen (DAG) als auch in die polaren Lipide (diacyl-GPC) der Spermatozoen eingebaut. Nach Supplementierung mit 13C-markierter Octadecadiensäure wurden die Lipide mittels MALDI- und Q-TOF-MS als DAG (18:2/18:2), GPC (16:0/18:2) und GPC (18:2/18:2) charakterisiert. Die gleichen Ergebnisse wurden auch für die in den Spermatozoenlipiden vorkommenden Hexadecen-, Octadecen-, und Octadecatriensäure erhalten. Bei der Untersuchung des physiologischen Zustandes von Spermatozoen wurde gezeigt, dass insbesondere Supplementierungsvarianten mit endogen vorkommenden Fettsäuren zu einer besseren Spermatozoenvitalität und Motilität bei Niedrigtemperaturlagerung führten. Gleichzeitig wurde eine Verminderung des Auftretens von akrosomalen Schäden festgestellt. Damit stellt eine Supplementierung der Spermatozoen mit ausgewählten Fettsäuren eine effektive Maßnahme zur Lagerung von Spermatozoen bei 4 bis 6°C dar., This study examines the metabolic incorporation of selected fatty acids into the lipids of porcine spermatozoa and evaluates the physiological state of spermatozoa subsequent to low temperature storage supplementation with selected free fatty acids. The aim was to understand the role of fatty acids in relation to the (cryo-)preservation of spermatozoa and successful reproduction in more detail. All lipids present in porcine spermatozoa were analysed using gas chromatography (GC) and mass spectrometry (MALDI-TOF-MS). The main representatives of the polar lipid classes are glycerophospholipids (in particular GPC and GPE). The main representatives of the neutral lipid classes are diacylglycerols (DAG). Metabolic incorporation of fatty acids into lipids was radiochemically monitored using [14C]-octadecadienoic acid in the supplied spermatozoa-preservation medium. Temperature and incubation time were shown to be particularly important determinants. The added fatty acids were incorporated into both the spermatozoas’ neutral (DAG) and polar lipids (diacyl-GPC). The affected lipids were characterised by means of MALDI- and Q-TOF-MS subsequent to the supplementation of uniformly 13C-labelled octadecadienoic acid. DAG (18:2/18:2), GPC (16:0/18:2) and GPC (18:2/18:2) could be identified and a de-novo biosynthesis of DAG (18:2/18:2) could be proven. The same results were obtained when spermatozoa were supplemented with hexadecenoic, octadecenoic and octadecatrienoic acids. Finally, it was shown that the physiological state of the spermatozoa, especially those supplemented with endogeneously present fatty acids, led to an enhanced vitality and motility in spermatozoa subsequent to low temperature storage. It was also observed that acrosomal damage was reduced and that hexadecenoic acid significantly stabilised all the vitality parameters. In conclusion, supplementing spermatozoa with selected fatty acids is an effective solution for the storage of spermatozoa at 4 to 6°C.

Assisted reproductive technology (ART) is considered an important tool in the conservation of endangered species, but often the most limiting factor of ART is the availability of mature oocytes. The aim of the present study was to investigate the feasibility of preserving female germ cells from ovaries of female lions (Panthera leo). Good quality cumulus-oocyte complexes (COCs) were isolated and subjected to in vitro maturation (IVM). In addition, ovarian cortex was obtained and cut into pieces for culture and cryopreservation by slow freezing. The survival of ovarian follicles was assessed by histology. Frozen-thawed samples of ovarian cortex samples were xenotransplanted under the skin of ovariectomized immunodeficient mouse for 28 days. Overall, 178 intact COCs were obtained from 13 lions, but only 28.1% were matured in vitro indicating insufficient IVM conditions. In contrast, almost all follicles within the ovarian cortex survived culture when the original sample was from a young healthy lion collected immediately after euthanasia. Within the xenotransplants, the number of primordial follicles decreased after 28 days by 20%, but the relation between primordial and growing follicles changed in favour of follicular growth. Female gamete rescue from valuable felids may be performed by slow freeze cryopreservation of ovarian cortex. Although the IVM protocol for lions is not yet optimized, mature oocytes may be obtained after long-term xenotransplantation and IVM and could potentially represent one way of salvage of endangered felid species in the future.

Female cancer patients are offered 'banking' of gametes before starting fertility-threatening cancer therapy. Transplants of fresh and frozen ovarian tissue between healthy fertile and infertile women have demonstrated the utility of the tissue banked for restoration of endocrine and fertility function. Additional methods, like follicle culture and isolated follicle transplantation, are in development. Specialist reproductive medicine scientists and clinicians with complementary expertise in ovarian tissue culture and transplantation presented relevant published literature in their field of expertise and also unpublished promising data for discussion. As the major aims were to identify the current gaps prohibiting advancement, to share technical experience and to orient new research, contributors were allowed to provide their opinioned expert views on future research. Normal healthy children have been born in cancer survivors after orthotopic transplantation of their cryopreserved ovarian tissue. Longevity of the graft might be optimized by using new vitrification techniques and by promoting rapid revascularization of the graft. For the in vitro culture of follicles, a successive battery of culture methods including the use of defined media, growth factors and three-dimensional extracellular matrix support might overcome growth arrest of the follicles. Molecular methods and immunoassay can evaluate stage of maturation and guide adequate differentiation. Large animals, including non-human primates, are essential working models. Experiments on ovarian tissue from non-human primate models and from consenting fertile and infertile patients benefit from a multidisciplinary approach. The new discipline of oncofertility requires professionalization, multidisciplinarity and mobilization of funding for basic and translational research.

The Iberian lynx is considered the most endangered felid species. Therefore, an ex situ conservation program was initiated to protect this species from extinction. Additional knowledge on lynx reproduction biology and reliable methods for reproductive monitoring are important for developing a captive breeding program. The aim of this study in lynx was to implement transrectal ultrasonography to visualize ovarian structures (follicles, corpora lutea) and to assess ovarian activity in addition to analysis of serum progesterone and oestradiol. Because of limited access to Iberian lynxes, the less-endangered Eurasian lynx and bobcat were also studied in this comparative study. Recent endocrinological studies based on faecal and urinary progesterone and oestrogen metabolites revealed that steroid profiles in both these species were alike and did not follow the typical pattern of other felids. Pregnancy diagnosis was not possible, since progesterone concentrations did not differ between pregnant and pseudopregnant animals. Progesterone was also detected after parturition as well as after weaning until the onset of a new oestrous cycle. In the present study, the presence of corpora lutea during the non-breeding season was confirmed by ultrasonography and by elevated serum levels of progesterone averaging 3.56 ± 1.3 ng/ml in Eurasian and 6.1 ± 0.26 ng/ml in Iberian lynx, respectively. The ultrasonographical findings on the ovarian structures suggest strongly that corpora lutea developed after ovulation stay active until November and regress before the onset of the next oestrus.