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Biomedical subjects

K Hardy

Publications and source records attributed to K Hardy.

At least 73 records · Page 4Linked to original sources
PubMed1996

Metabolism and cell allocation during parthenogenetic preimplantation mouse development.

Diploid parthenogenetic postimplantation mouse embryos, containing two maternal genomes, are characterized by poor development of extraembryonic membranes derived from the trophectoderm and primitive endoderm of the blastocyst. This is thought to be caused by a deficiency of expression of paternally derived imprinted genes. Here we have compared the inner cell mass, from which the primitive endoderm and fetal lineages are derived, and the trophectoderm, which forms a major component of the placenta, in parthenogenetic and fertilized preimplantation embryos. We have also studied the metabolism from the 1-cell to the blastocyst stage. Cell numbers were reduced in the ICM and TE of parthenogenetic blastocysts compared to fertilized blastocysts. This was thought to be due to the increased levels of cell death observed in these lineages. Pyruvate and glucose uptake by parthenogenetic embryos was similar to that by fertilized embryos throughout preimplantation development. However, at the expanded blastocyst stage glucose uptake by parthenogenetic embryos was significantly higher than by fertilized embryos. The implications of the actions of imprinted genes and of X-inactivation is discussed.

Animals↗
PubMed1996

Expression of intercellular junctions during preimplantation development of the human embryo.

A total of 74 human embryos were stained with gap junction protein specific anti-peptide antibodies an antibodies to the desmosomal protein desmoplakin to reveal the expression pattern of intercellular junctions during preimplantation development. Prior to implantation, the human embryo expresses predominantly connexin (Cx43)-containing gap junctions. Gap junctions were first detected in apposing cell membranes at the 4-cell stage and became increasingly organized as development proceeded. In normal blastocysts, trophectoderm (TE) cells were linked by dense arrays of gap junctions while inner cell mass (ICM) cells were linked by small, punctate gap junctions. Gap junctions containing Cx32 or Cx26 were observed occasionally in the TE of late blastocysts. Desmosomes appeared between outer cells prior to cavitation and were retained in the TE, but not in the ICM. Levels of gap junction protein expression were variable in morphologically normal embryos at the same stage, suggesting that a normal appearance may not be a reliable indicator of future viability. Morphologically normal embryos often possessed multinucleate, apoptotic and decompacting cells. They could show either extensive, disorganized over-expression or reduced expression of gap junction protein. The results fit the view that only embryos destined to survive display an organized pattern of intercellular junctions.

Blastocyst↗
PubMed1995

Delaying transfer to the third day post-insemination, to select non-arrested embryos, increases development to the fetal heart stage.

The purpose of this study was to determine whether delaying embryo transfer by 24 h, until day 3 post-insemination, allowed improved selection of non-arrested embryos for transfer. We have retrospectively analysed pregnancy rates in a large series of patients who had embryo transfer either on day 2 or on day 3 post-insemination over a 27 month period. From January 1990 to March 1992, 567 patients received embryo transfer on day 2, and 661 patients had transfer on day 3 post-insemination, but these transfers were not contemporary. Pregnancy rates were slightly higher in patients who had embryo transfer on day 3 (37%) than in those patients who had their embryos transferred on day 2 (35%), but this difference was not significant. The implantation rate, as measured by the proportion of embryos developing to the fetal heart stage, was significantly higher following transfer on day 3 (23%) than after transfer on day 2 (19%) (P < 0.05), suggesting that selection of viable embryos is improved on day 3. Furthermore, of the embryos which gave rise to a fetal sac, significantly fewer miscarried before the fetal heart stage (P < 0.05) following transfer on day 3 (6%) than after transfer on day 2 (12%). Delaying transfer until day 3 provides a further 24 h to observe embryo development. During this period 16% of embryos arrested or became developmentally retarded; thus waiting until day 3 allowed these embryos to be identified and avoided for consideration for transfer. Embryo transfer may be safely delayed until day 3, and this may help in selecting embryos most likely to implant and develop after transfer.

Adult↗
PubMed1995

Normal development and metabolic activity of preimplantation embryos in vitro from patients with polycystic ovaries.

Polycystic ovary syndrome (PCOS) is closely associated with high miscarriage rates and, following in-vitro fertilization (IVF), with decreased fertilization rates, suggesting that oocytes and embryos are of poor quality. In this prospective study, we examined the development, metabolic activity and blastocyst cell number of embryos following IVF from 51 patients with either anovulatory PCOS, ovulatory PCOS or tubal disease. The number of oocytes retrieved and the fertilization rates were similar for patients with PCOS and tubal disease. Following embryo transfer, 46% of the patients with PCOS and 36% of patients with tubal disease became pregnant. A similar proportion of surplus embryos from patients with PCOS and tubal disease developed to the blastocyst stage (38% and 43% respectively). Patients with anovulatory PCOS had embryos with less fragmentation which cleaved faster, cavitated earlier and had more cells at the blastocyst stage than embryos from patients with tubal disease. While the profile of glucose uptake and lactate production was similar for all groups throughout preimplantation development, patients with tubal disease who underwent ovulation induction using the 'titrated' regimen optimized for PCOS patients resulted in embryos with reduced pyruvate uptake, in addition to low blastocyst cell numbers. This study demonstrates that with an optimized ovulation induction regimen, embryos from PCOS patients are of good quality and developmental potential.

Adult↗
PubMed1995

Bilateral endobronchial non-Hodgkin's lymphoma.

Pulmonary involvement with non-Hodgkin's lymphoma, while well recognized, is less common than with Hodgkin's lymphoma. Endobronchial non-Hodgkin's lymphoma is even rarer and usually occurs in the presence of disseminated disease. We present a case of bilateral endobronchial non-Hodgkin's lymphoma.

Aged↗
PubMed1994

Head and neck tumors after energetic proton irradiation in rats.

This is a two-year progress report on a life span dose-response study of brain tumor risk at moderate to high doses of energetic protons. It was initiated because a joint NASA/USAF life span study of rhesus monkeys that were irradiated with 55-MeV protons (average surface dose, 3.5 Gy) indicated that the incidence of brain tumors per unit surface absorbed dose was over 19 times that of the human tinea capitis patients whose heads were exposed to 100 kv x-rays. Examination of those rats that died in the two-year interval after irradiation of the head revealed a linear dose-response for total head and neck tumor incidence in the dose range of 0-8.5 Gy. The exposed rats had a greater incidence of pituitary chromophobe adenomas, epithelial and mesothelial cell tumors than the unexposed controls but the excessive occurrence of malignant gliomas that was observed in the monkeys was absent in the rats. The estimated dose required to double the number of all types of head and neck tumors was 5.2 Gy. The highest dose, 18 Gy, resulted in high mortality due to obstructive squamous metaplasia at less than 50 weeks, prompting a new study of the relative biological effectiveness of high energy protons in producing this lesion.

Adenoma↗
PubMed1994

Results of time-resolved radiation exposure measurements made during U.S. Shuttle missions with a tissue equivalent proportional counter.

Time-resolved radiation exposure measurements inside the crew compartment have been made during recent Shuttle missions with the USAF Radiation Monitoring Equipment-III (RME-III), a portable four-channel tissue equivalent proportional counter. Results from the first six missions are presented and discussed. The missions had orbital inclinations ranging from 28.5 degrees to 57 degrees, and altitudes from 200-600 km. Dose equivalent rates ranged from 40-5300 micro Sv/dy. The RME-III measurements are in good agreement with other dosimetry measurements made aboard the vehicle. Measurements indicate that medium- and high-LET particles contribute less than 2% of the particle fluence for all missions, but up to 50% of the dose equivalent, depending on the spacecraft's altitude and orbital inclination. Iso-dose rate contours have been developed from measurements made during the STS-28 mission. The drift rate of the South Atlantic Anomaly (SAA) is estimated to be 0.49 degrees W/yr and 0.12 degrees N/yr. The calculated trapped proton and Galactic Cosmic Radiation (GCR) dose for the STS-28 mission were significantly lower than the measured values.

Cosmic Radiation↗
PubMed1994

Radiation dosimetry measurements during U.S. Space Shuttle missions with the RME-III.

Time-resolved radiation dosimetry measurements inside the crew compartment have been made during recent Shuttle missions with the U.S. Air Force Radiation Monitoring Equipment-III (RME-III), a portable battery-powered four-channel tissue equivalent proportional counter. Results from the first six missions are presented and discussed. Half of the missions had orbital inclinations of 28.5 degrees with the remainder at inclinations of 57 degrees or greater; altitudes ranged from 300 to 600 km. The determined dose equivalent rates ranged from 70 to 5300 microSv/day. The RME-III measurements are in good agreement with other dosimetry measurements made aboard the vehicles. Measurements indicate that medium- and high-LET particles contribute less than 2% of the particle fluence for all missions, but up to 50% of the dose equivalent, depending on the spacecraft's altitude and orbital inclination. Isocontours of fluence, dose and dose equivalent rate have been developed from measurements made during the STS-28 mission. The drift rate of the South Atlantic Anomaly is estimated to be 0.49 degrees W/yr and 0.12 degrees N/yr. The calculated trapped proton and GCR dose for the STS-28 mission was significantly lower than the measured values.

Atlantic Ocean↗
PubMed1993

Early human embryo metabolism.

Non-invasive microanalytical methods have been devised to study the energy metabolism of single human preimplantation embryos. Pyruvate, which is added routinely to all media used to culture human embryos, is consumed throughout the preimplantation period, with glucose assuming an increasing role at embryo compaction and blastocyst formation. All of the glucose consumed may be accounted for by the appearance of lactate in the incubation medium. The enzyme hexokinase may be involved in regulating this aerobic glycolysis. There is considerable indirect evidence for the utilisation of endogenous as opposed to exogenous energy substrates, the most likely candidate being protein. Information on early human embryo metabolism is likely to find application in a number of areas: these include the improvement of techniques for assisted human conception, notably in the selection of embryos for transfer following In Vitro Fertilisation; the diagnosis of genetic defects at the preimplantation stage; increased understanding of the causes of implantation failure and miscarriage, and the development of novel post-coital contraceptives.

Adenosine Triphosphate↗
PubMed1993

Cell allocation in twin half mouse embryos bisected at the 8-cell stage: implications for preimplantation diagnosis.

Previous work in the mouse and human has shown that removal of cells from 2- and 4-cell embryos disproportionately reduces the inner cell mass (ICM), from which the fetus is derived, suggesting that preimplantation biopsy at these stages would be inadvisable. Here we have examined allocation in twin half mouse embryos produced by bisection at the 8-cell stage. Half embryos and intact zona-free control embryos were differentially labeled at the time of cavitation using polynucleotide-specific fluorochromes, and the numbers of cells in the ICM and trophectoderm (TE) counted. Adding the numbers of cells in the twin halves together, the numbers of ICM cells are not significantly different from those in intact controls. In contrast, numbers of TE cells were significantly higher (P < 0.01), producing a small increase in the total cell number of half embryos. There is no concomitant decrease in ICM cell number. We propose that as the surface area of a half embryo is greater than half an intact control, TE cells in half embryos divide faster in order to reach an optimal packing density similar to that in intact control embryos. Furthermore, we suggest that preimplantation biopsy only be performed at the 8-cell stage or later.

Animals↗
PubMed1993

Selection criteria for human embryo transfer: a comparison of pyruvate uptake and morphology.

PURPOSE: Pyruvate uptake is higher in human embryos developing to the blastocyst stage than those arresting at cleavage stages. To investigate whether pyruvate uptake provides an improved criterion for selecting embryos for transfer, we have measured uptakes by individual embryos noninvasively over 24-hr periods between the first day (day 1) postinsemination and embryo transfer on day 2 to 3 and correlated the levels with implantation and pregnancy outcome. RESULTS: The mean uptake was significantly lower for embryos that implanted than for those which failed to implant: 22.9 +/- 1.0 and 27.1 +/- 0.6 pmol/embryo/hr, respectively on day 2, and 22.4 +/- 1.5 and 26.9 +/- 0.8 pmol/embryo/hr, respectively, on day 3, but the wide range of uptakes by individual embryos was overlapping. CONCLUSION: We conclude that pyruvate uptake as the sole criterion for embryo selection cannot predict which embryos will implant after transfer. Assessment of embryos using morphological and developmental criteria, therefore, remains the most consistent, though inefficient, indicator of pregnancy potential.

Adult↗
PubMed1993

Pharmacokinetics and biological activity of kinetensin in conscious sheep.

Kinetensin is a nonapeptide, originally isolated from pepsin-treated plasma, that shares some sequence homology with the C-terminal end of neurotensin. The present study was designed to determine, by infusing kinetensin to conscious sheep, the pharmacokinetics and a neurotensin-like biological activity (pancreatic polypeptide response) of kinetensin. Kinetensin was rapidly metabolized, approximately 200-fold more rapidly than neurotensin. The majority of the metabolism occurred in the circulation as demonstrated both in vivo and in vitro. The lung and gut cleared kinetensin also. Inhibition of converting enzyme, present in highest concentration in the lung, abolished lung clearance but was without effect on kinetensin metabolism by the gut or in the general circulation. Arterial infusion of kinetensin which achieved high blood kinetensin levels at the pancreas did not increase plasma pancreatic polypeptide. We conclude that the extremely rapid degradation of exogenous kinetensin, together with the lack of biological activity, makes it unlikely that kinetensin plays a role as a circulating regulatory peptide. Nevertheless, since the putative kinetensin substrate circulates at microM concentrations, it is feasible that kinetensin is generated and metabolized at the target organ.

Animals↗
PubMed1993

Mammalian bombesin as a hormone in ovine pregnancy: ontogeny, origin, and molecular forms.

Mammalian gastrin-releasing peptides (GRP) are present in female reproductive tissues and stimulate uterine contraction and DNA synthesis in the endometrium. We set out to establish whether the GRP were likely to play a role in fetal development by measuring the fetal and maternal plasma concentrations in chronically cannulated fetal sheep from 115 days gestation to term (145 days) and for 18 days after parturition. Placental fluids and fetal urine were also obtained. In a separate series of animals, uterine, placental, and fetal gut and lung tissues were collected. Samples were extracted in acid and assayed by radioimmunoassay, and molecular forms were characterized by high-performance liquid chromatography. GRP were present in the decidua (> 10 pmol/g), as well as placental, uterine, lung, and gastrointestinal tissues (all < 6 pmol/g). Fetal and maternal plasma GRP were elevated compared with nonpregnant ewes, falling sharply after parturition. Placental fluids and urine also contained GRP (80-410 fmol/ml). The main molecular form in all tissues and fluids examined coeluted with porcine GRP-(18-27). GRP immunoreactivity was primarily localized to epithelial cells of the decidua. These data suggest that plasma GRP is probably derived from the decidua and may play a role as a circulating hormone in ovine fetal and uterine development.

Animals↗
PubMed1993

Binucleate blastomeres in preimplantation human embryos in vitro: failure of cytokinesis during early cleavage.

The nuclei of disaggregated blastomeres from two hundred preimplantation human embryos were examined between days 2 and 4 after insemination in vitro by vital labelling with a polynucleotide-specific fluorochrome. Although the majority of blastomeres had a single nucleus, binucleate blastomeres containing two nuclei of equal size were common and other blastomeres had fragmented nuclei or were anucleate. Seventeen per cent of normally fertilized embryos at two- to four-cell stage had at least one binucleate blastomere, and this increased to 65% at the nine- to 16-cell stage when individual embryos had between one and six binucleate blastomeres. The proportion of binucleate blastomeres in normally fertilized embryos increased from 5 to 10% over this period, whereas in abnormally fertilized, polyspermic or parthenogenetic, embryos the proportion was significantly higher during early cleavage stages but decreased at the nine- to 16-cell stage when the majority of these embryos arrest (25 and 6%, respectively). The incidence of anucleate blastomeres in normally fertilized embryos was also high, especially in those of poor morphology. In contrast, blastomeres with fragmented nuclei were relatively uncommon and the incidence was variable among classes and stages of development. Estimates of the volume of binucleate blastomeres based on measurement of their diameters and comparison with mononucleate blastomeres at various cleavage stages indicated that these blastomeres arise from a failure of cytokinesis between the second and fourth cleavage divisions. On this basis, assignment of binucleate blastomeres to particular cleavage stages in normally fertilized day 4 embryos suggests that at least some of these blastomeres arising during early cleavage persist without further cell division for up to 48 h. At the cellular level, therefore, blastomeres with either binucleate or abnormal nuclei contribute to cleavage stage arrest in vitro.

Blastomeres↗
PubMed1993

Activity of enzymes of energy metabolism in single human preimplantation embryos.

A method was developed to measure the activities of enzymes in extracts from single human preimplantation embryos. The method permits the analysis of two enzymes plus appropriate controls in an extract from a single embryo, and was used to investigate the control of energy metabolism during the development of human embryos from the two-cell to the blastocyst stage. Hexokinase (HK), 6-phosphofructokinase (PFK), pyruvate kinase (PK), fructose-1,6-diphosphate aldolase (ALD), glucose phosphate isomerase (GPI), lactate dehydrogenase (LDH), glucose-6-phosphate dehydrogenase (G6PDH) and 2-oxoglutarate dehydrogenase (ODH) were all detectable, whereas glycogen phosphorylase (GP) was not. The enzyme activities of ODH, PFK, LDH, PK, GPI and G6PDH, averaged over all stages of development from the two-cell to blastocyst stage (days 2-6 after insemination), were 3.5, 6.6, 15, 69, 73 and 87 times greater than HK, respectively. The activity of ALD was very similar to that of HK. The activities of ALD, GPI, PFK, PK and LDH showed no significant variation with stage of development, although the activity of GPI fell significantly from the four-eight cell to the eight-sixteen cell stage (P < 0.05). HK activity decreased from the two-eight cell to the eight-sixteen cell (P < 0.05), and increased significantly from the eight-sixteen cell to the blastocyst stage (P < 0.01). The overall relationship between hexokinase activity and stage approached significance (P = 0.059, one-way analysis of variance). The activity of G6PDH decreased significantly with development (P < 0.001, one way analysis of variance).(ABSTRACT TRUNCATED AT 250 WORDS)

Blastocyst↗