Different patterns of metabolic cryo-damage in domestic cat (Felis catus) and cheetah (Acinonyx jubatus) spermatozoa.

Felid spermatozoa are sensitive to cryopreservation-induced damage, but functional losses can be mitigated by post-thaw swim-up or density gradient processing methods that selectively recover motile or structurally-normal spermatozoa, respectively. Despite the importance of sperm energy production to achieving fertilization, there is little knowledge about the influence of cryopreservation or post-thaw processing on felid sperm metabolism. We conducted a comparative study of domestic cat and cheetah sperm metabolism after cryopreservation and post-thaw processing.

Oxidative phosphorylation is essential for felid sperm function, but is substantially lower in cheetah (Acinonyx jubatus) compared to domestic cat (Felis catus) ejaculate.

Compared with the normospermic domestic cat, sperm metabolic function is compromised in the teratospermic cat and cheetah, but the pathway(s) involved in this deficiency are unknown. Glycolysis is essential for sperm motility, yet it appears to function normally in spermatozoa of either species regardless of structural morphology. We conducted a comparative study to further understand the mechanisms of energy production in felid spermatozoa, with the hypothesis that oxidative phosphorylation is required for normal sperm function and is impaired in teratospermic ejaculates.

Glycolytic enzyme activity is essential for domestic cat (Felis catus) and cheetah (Acinonyx jubatus) sperm motility and viability in a sugar-free medium.

We have previously reported a lack of glucose uptake in domestic cat and cheetah spermatozoa, despite observing that these cells produce lactate at rates that correlate positively with sperm function. To elucidate the role of glycolysis in felid sperm energy production, we conducted a comparative study in the domestic cat and cheetah, with the hypothesis that sperm motility and viability are maintained in both species in the absence of glycolytic metabolism and are fueled by endogenous substrates. Washed ejaculates were incubated in chemically defined medium in the presence/absence of glucose and pyruvate.

Evidence for compromised metabolic function and limited glucose uptake in spermatozoa from the teratospermic domestic cat (Felis catus) and cheetah (Acinonyx jubatus).

Cheetahs and certain other felids consistently ejaculate high proportions (≥ 60%) of malformed spermatozoa, a condition known as teratospermia, which is prevalent in humans. Even seemingly normal spermatozoa from domestic cat teratospermic ejaculates have reduced fertilizing capacity. To understand the role of sperm metabolism in this phenomenon, we conducted a comparative study in the normospermic domestic cat versus the teratospermic cat and cheetah with the general hypothesis that sperm metabolic function is impaired in males producing predominantly pleiomorphic spermatozoa.

Zona-free embryo culture: is it a viable option to improve pregnancy rates?

Sporadic reports published during the previous decade have documented pregnancies achieved with transfer of zona-free human embryos. Although the overall efficiency seems to be good and some authors have suggested systematic application for special infertility problems, there have been only a few attempts to compare the benefits of zona-free embryo culture and transfer with the traditional approach using zona-intact embryos. So far, the majority of instances in which zona-free culture has been applied have occurred accidentally.

Long-distance transportation of primate embryos developing in culture: a preliminary study.

Non-human primate embryos are invaluable for conducting research relevant to human infertility and stem cells, but their availability is restricted. In this preliminary study, rhesus monkey embryos were produced by IVF at the Caribbean Primate Research Centre and shipped in tubes of gassed culture medium within a battery-powered transport incubator by overnight courier to Wayne State University in Michigan. Upon arrival, the embryos were incubated in fresh culture medium to evaluate further development.

Incidence of chromosomal mosaicism in morphologically normal nonhuman primate preimplantation embryos.

Objective: To establish the exact rates of chromosomal mosaicism in morphologically normal rhesus macaque embryos by determining the chromosomal complement of all blastomeres.

Design: Retrospective rhesus monkey IVF study.

Setting: Academic laboratory and primate research center.

Effects of in vitro maturation and age on oocyte quality in the rhesus macaque Macaca mulatta.

Objective: To evaluate oocyte quality in a primate model.

Design: Analysis of oocyte karyotype by chromosome spreading and oocyte spindles by confocal microscopy.

Setting: Research laboratory, Caribbean Primate Research Center.

Rhesus macaque embryos derived from MI oocytes maturing after retrieval display high rates of chromosomal anomalies.

Background: Rhesus macaque and human preimplantation embryos display similar rates of chromosomal abnormalities. The aim of this study was to determine whether embryos developing from MI oocytes that mature post-retrieval display more chromosomal anomalies than those embryos that are generated from oocytes that are at MII at the time of retrieval.

Methods: Rhesus macaque oocytes were obtained after hormonal ovarian stimulation.

Chromosomal instability in rhesus macaque preimplantation embryos.

Objective: To establish a relevant animal model to systematically investigate chromosomal instability in human oocytes and preimplantation embryos.

Design: Prospective rhesus monkey IVF study.

Setting: Academic laboratory, Oregon National Primate Research Center and Caribbean Primate Research Center.

Mitochondria in stem cells.

The current status of knowledge about mitochondrial properties in mouse, monkey and human embryonic, adult and precursor stem cells is discussed. Topics include mitochondrial localization patterns, oxygen consumption and ATP content in cells as they relate to the maintenance of stem cell properties and subsequent differentiation of stem cells into specific cell types. The significance of the perinuclear arrangement of mitochondria, which may be a characteristic feature of stem cells, as well as the expression of mitochondrial DNA regulatory proteins and mutations in the mitochondrial stem cell genome is also discussed.

The mitochondrial contribution to stem cell biology.

The distribution and functions of mitochondria in stem cells have not been examined, yet the contributions of these organelles to stem cell viability and differentiation must be vitally important in view of their critical roles in all other cell types. A key role for mitochondria in stem cells is indicated by reports that they translocate in the oocyte during fertilisation to cluster around the pronuclei and can remain in a perinuclear pattern during embryo development. This clustering appears to be essential for normal embryonic development.

Differentiation-related changes in mitochondrial properties as indicators of stem cell competence.

Several methods may be used to assess stem cell competence, including the expression of cell surface markers and telomerase activity. We hypothesized that mitochondrial characteristics might be an additional and reliable way to verify stem cell competence. In a multipotent, adult monkey stromal stem cell line, previously shown to differentiate into adipocytes, chondrocytes, and osteocytes, we found that several mitochondrial properties change with increasing passage number in culture.

Challenges of primate embryonic stem cell research.

Embryonic stem (ES) cells hold great promise for treating degenerative diseases, including diabetes, Parkinson's, Alzheimer's, neural degeneration, and cardiomyopathies. This research is controversial to some because producing ES cells requires destroying embryos, which generally means human embryos. However, some of the surplus human embryos available from in vitro fertilization (IVF) clinics may have a high rate of genetic errors and therefore would be unsuitable for ES cell research.

The role of animal studies in supporting human assisted reproductive technology.

Although average success rates of human IVF have increased progressively during the past two decades, the efficiency of this technique, based on each embryo produced or transferred, is still low. High success rates are usually achieved by transferring several embryos to the patient, which is often associated with multiple pregnancies. The quality of in vitro produced embryos is a major area that needs attention.

Duration and temperature of culture medium equilibration affect frequency of blastocyst development.

Hamster 2-cell embryos were cultured in 50 microl drops of chemically defined medium (HECM-9) under oil in 60 mm Petri dishes. In the first experiment, the dishes were equilibrated with 5% O(2) /10% CO(2) /85% N(2) for 2 h either within sealed plastic bags or exposed directly to the same gas mixture in a tissue culture incubator. After culture of embryos for 48 h, there was no difference in development to the blastocyst stage.

Oxygen concentration and preimplantation development.

The recent article by Karagenc et al. once again shows that atmospheric oxygen is detrimental to embryo development in vitro. This study demonstrates that zygotes can readily develop into blastocysts under ambient oxygen, but in spite of their morphologically normal appearance, the viability of many of these embryos is compromised, with the most pronounced effect on ICM development.

Human ovarian tissue cultures: extracellular matrix composition, coating density and tissue dimensions.

Human ovarian tissue can be successfully cryopreserved for fertility preservation. Optimal use of this approach requires the development of reliable restoration methods, including in-vitro culture of follicles. A culture system has been established, but improvement of the basic handling and techniques is necessary.

17Beta-estradiol and progesterone improve in-vitro cytoplasmic maturation of oocytes from unstimulated prepubertal and adult rhesus monkeys.

Background: Effects of 17beta-estradiol and progesterone on rhesus monkey oocyte maturation in vitro were evaluated by embryo development subsequent to IVF.

Methods And Results: In experiment 1, immature cumulus-oocyte complexes collected from unstimulated adult females during the non-breeding season were matured in modified medium CMRL-1066 containing various combinations of gonadotrophins (FSH + LH), estradiol and/or progesterone. Formation of morulae and blastocysts was greatest in oocytes matured in medium containing estradiol and/or progesterone, with or without gonadotrophins (morula 38-46%, blastocyst 14-20%) than in control oocytes matured without estradiol or progesterone (morula 14%, blastocyst 0%).

Blastocyst development after intergeneric nuclear transfer of mountain bongo antelope somatic cells into bovine oocytes.

Intergeneric embryos were constructed by nuclear transfer using Mountain Bongo antelope somatic cells fused with enucleated bovine oocytes and their subsequent development in vitro was investigated. After two to six passages, starved or non-starved skin fibroblast cells were used as donor nuclei. In vitro matured bovine oocytes were enucleated by squeezing the first polar body and surrounding cytoplasm through a slit in the zona pellucida.

Early history of in vitro fertilization.

Although in vitro fertilization (IVF) is used widely for a variety of purposes, it is often not appreciated how this technology was developed. A large number of experiments beginning in 1878 contributed to the first successful reports of IVF over 75 years later. The discovery of sperm capacitation in 1951 was central to the development of IVF technology, and it was rapidly followed by the first convincing reports of IVF in several species.

Features associated with reproductive ageing in female rhesus monkeys.

Background: The specific aims were to determine the effects of maternal age on the meiotic and developmental competence of oocytes and incidence of chromosomal anomalies in oocytes from a population of fertile rhesus monkeys.

Methods: Monkeys were divided into two age groups (4-15 and 16-26 years of age) and underwent ovarian stimulation for collection of oocytes.

Results: In the older, compared with younger, monkeys, serum basal concentrations of FSH were elevated (P < 0.

Biochemical and biological properties of urinary follicle-stimulating hormone (FSH) from the rhesus monkey (Macaca mulatta).

Follicle-stimulating hormone (FSH) is routinely used for the induction of superovulation in women. Homologous gonadotropin preparations that could be used for reproductive studies in macaques would have valuable research applications. Accordingly, we set out to characterize the physical and biological characteristics of urinary FSH (UFSH) in the ovariectomized rhesus monkey.

RECENT PROGRESS IN THE STUDY OF EARLY EVENTS IN MAMMALIAN FERTILIZATION.

During the past 25 years, great advances have been made in understanding the physiology, morphology and biochemistry of fertilization in invertebrate animal species. In contrast to this situation, there is a paucity of knowledge pertaining to mammalian fertilization. Major areas in which information is lacking are the nature of changes undergone by spermatozoa in preparation for fertilization, and the mechanisms involved in sperm penetration of the egg investments.