A methodology is described for analyzing single human ova for 8 or 9 different metabolic enzymes, or 4 or 5 enzymes plus as many metabolites. This overcomes an obstacle to the study of human ovum metabolism: the severe limitation of usable material. Results obtained with this methodology, applied to discarded specimens from an in vitro fertilization program, indicate that in spite of imperfections these ova can provide a valid picture of the metabolic characteristics of normal human ova. Data are presented for 17 enzymes from 8 metabolic pathways in human and mouse ova. Relative to size, 10 of the enzymes were substantially higher in human than mouse ova. Most dramatically so were 2 enzymes of fatty acid metabolism (10-fold and 15-fold), hexokinase (9-fold), and aspartate aminotransferase (19-fold). This suggests that major species differences in metabolism are present. The validity of the human data, in spite of restriction to discarded material, is supported by (1) consistency of results among most of the ova, 2] concordance between average levels with those of rare specimens that were discarded because sperm were not available, and (3) the presence of adenosine triphosphate (ATP) concentrations similar to those of normal mouse ova. Surprisingly, both human and mouse ova contain phosphocreatine at levels nearly equal of those of ATP.

No abstract

With the advent of new techniques of human in vitro fertilization (IVF), identifying parameters of oocyte quality to allow selection of those most likely to fertilize becomes crucial. Morphology of oocytes, which correlates positively with biological performance, is the currently utilized classification criterion. However, biological links between form and function are tenuous, and underlying mechanisms remain elusive. We investigated whether biochemical activation is quantitatively associated with the stages of maturation in ova obtained from patients undergoing gynecologic surgery during unstimulated cycles and women undergoing IVF after exogenous gonadotropin stimulation. Changes in selected enzymes from protein, lipid, and carbohydrate metabolism (hexokinase, phosphoglucomutase, glycogen synthetase, uridine diphosphoglucose pyrophosphorylase, glucose-6-phosphate dehydrogenase, cytosolic thiolase, beta-hydroxyacyl-CoA dehydrogenase, alanine aminotransferase, and aspartate aminotransferase) were determined simultaneously, in individual oocytes, utilizing a highly sensitive biochemical methodology. Several enzyme activities paralleled maturation grade and were higher in stimulated oocytes after correction for grade. These biochemical findings quantify metabolic and functional changes that increase as ova mature, possibly contributing to their reproductive performance.

Swiss white mice were superovulated, mated, and sacrificed to recover two‐cell embryos that were cultured in Ham's F‐10 supplemented with 15% fetal serum. In 16 experiments, media enriched with fetal bovine serum (FBS) supported blastocyst development from 80% ± 19% (mean ± S.D.) of two‐cell embryos. Culture media + FBS was the positive control when 74 batches of heat‐inactivated human fetal cord serum (hFCS) were tested. Statistical analyses indicated two distinct populations: 49 hFCS promoted blastocyst formation and 25 hFCS grew fewer blastocysts. In five studies, 35/47 two‐cell embryos recovered from mice oviducts in media + FBS and immediately incubated formed blastocysts (75% ± 10%). In six comparison studies where the recovered embryos stood at room temperature for 30 minutes before incubation, only 18/57 (29% ± 21%) became blastocysts. When the colony was housed for 1 week in rooms with Shell No Pest Strips as treatment for mites, only 11/125 two‐cell embryos became blastocysts (9%). In contrast, animals housed in quarters decontaminated with chlorine bleach had reduced breeding efficiency and produced fewer two‐cell embryos. We conclude that (1) Ham's F‐10 + FBS is an excellent positive control to test new batches of hFCS; (2) hFCS that supports blastocyst formation from ≥75% of two‐cell embryos is adequate for human use; (3) pesticide treatment of breeding colonies and cooling of murine embryos during harvest both impaired in vitro blastocyst development; and (4) chlorine bleach cleansing of animal quarters reduced the number of successful matings.