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Biomedical subjects

K L Martin

Publications and source records attributed to K L Martin.

At least 19 recordsLinked to original sources
PubMed2005

The myo-inositol-1-phosphate synthase gene is essential in Trypanosoma brucei.

The de novo synthesis of myo-inositol occurs via a two-step process: first, glucose 6-phosphate is converted into inositol 1-phosphate by an INO1 (myo-inositol-1-phosphate synthase; EC 5.5.1.4); then, it is dephosphorylated by an inositol monophosphatase. The myo-inositol can then be incorporated into PI (phosphatidylinositol), which is utilized in a variety of cellular functions, including the biosynthesis of GPI (glycosylphosphatidylinositol) anchors. A putative INO1 was identified in the Trypanosoma brucei genome database and, by recombinant expression in Escherichia coli, was shown to be a catalytically active INO1. To investigate the importance of INO1, we created a conditional knockout, which, under non-permissive conditions, showed that INO1 is an essential gene in bloodstream form T. brucei and that the de novo synthesized myo-inositol is used for the formation of PI and GPI anchors.

Animals↗
PubMed2001

Structure of epiglucan, a highly side-chain/branched (1 --> 3;1 --> 6)-beta-glucan from the micro fungus Epicoccum nigrum Ehrenb. ex Schlecht.

The extracellular fungal polysaccharide, epiglucan, synthesised by Epicoccum nigrum is a side-chain/branched (1 --> 3;1 --> 6)-D-beta-glucan. Methylation analysis, 13C DEPT NMR and specific enzymic digestion data show slight variation in branching frequency among the epiglucans from the three strains examined. The (1 --> 3)-beta-linked backbone has (1 --> 6)-beta-linked branches at frequencies greater than the homologous glucans, scleroglucan and schizophyllan, from Sclerotium spp. and Schizophyllum commune, respectively. The structural analyses do not allow a distinction to be made between structures I and II. [structures: see text] Epiglucan displays non-Newtonian shear thinning rheological properties, typical of these glucans.

Ascomycota↗
PubMed2000

The rate and anisotropy of impulse propagation in the postnatal terminal crest are correlated with remodeling of Cx43 gap junction pattern.

BACKGROUND: Disruptions to intermyocyte coupling have been implicated in arrhythmogenesis and development of conduction disturbances. At present, understanding of the relationship between the microscopic organization of intercellular coupling and the macroscopic spread of impulse in the normal and diseased heart is largely confined to theoretical analyses. METHODS AND RESULTS: The abundance and arrangement of gap junctions, as well as conduction properties, were assessed in terminal crest preparations isolated from the atria of neonate, weanling, and adult rabbits. We report that the connexin composition of terminal crest was uncomplicated, with Cx43 being the most prominent isoform detectable by Western blotting and immunostaining. Terminal crest myocytes showed little change in total Cx43-gap junction per cell during postnatal growth as assessed by stereology. However, marked non-uniformities emerged in the sarcolemmal distribution of Cx43-gap junctions. Cx43-gap junction area at myocyte termini increased 3.5-fold from birth to adulthood. Correlated with this change in Cx43, impulse propagation velocity parallel to the myofiber axis, as assessed by multi-site optical mapping using voltage-sensitive dye (di-4-ANEPPS), increased 2.4-fold. Conversely, the amount of Cx43-gap junctions on myocyte sides, and the conduction velocity transverse to the myofiber axis, remained relatively invariant during maturation. Hence, the increasing electrical anisotropy of maturing terminal crest was wholly accounted for by increases in conductance velocity along the bundle. This increase in longitudinal conduction velocity was correlated with changes in the sarcolemmal pattern, but not the overall density, of Cx43-gap junctions. CONCLUSIONS: This study provides the first correlative structure/function analysis of the relationship between the macroscopic conduction of impulse and the microscopic cellular organization of gap junctions in a differentiating cardiac bundle. Confirmation is provided for theoretical predictions which emphasize the importance of the cell-to-cell geometry of coupling in determining the spread and pattern of myocardial activation.

Analysis of Variance↗
PubMed1999

Role of developmental factors in the switch from pyruvate to glucose as the major exogenous energy substrate in the preimplantation mouse embryo.

Preimplantation mouse embryos, cultured in vitro and those freshly flushed from the reproductive tract, exhibit a switch in energy substrate preference, from pyruvate during the early preimplantation stages, to glucose at the blastocyst stage. Although the biochemical basis of this phenomenon is quite well characterized, its timing and possible association with developmental factors have not been considered. We have therefore examined the role of five developmental factors in determining the timing of the switch, namely: (1) embryo age (in hours post hCG); (2) developmental stage; (3) cytokinesis; (4) cell number; and (5) activation of the embryonic genome. One-cell embryos, which develop more slowly than 2-cell embryos in vitro, were used to investigate the role of embryo age and developmental stage. Cytochalasin D, which inhibits cytokinesis and delays the timing of compaction and cavitation, was used to investigate the role of cell division and developmental stage. Finally, transcription of the embryonic genome was examined with the inhibitor, alpha-amanitin. Pyruvate and glucose consumption by single embryos were measured using a noninvasive ultramicrofluorometric technique. The results showed that the timing of the switch in energy substrate preference is precisely regulated in the mouse preimplantation embryo. Activation of the embryonic genome is a prerequisite for the switch and its timing is closely associated with developmental stage, specifically compaction and/or cavitation. Cell number, cytokinesis and embryo age appeared to be unrelated to the timing of the switch. These conclusions may well be extrapolated to other species, since an increase in net glucose uptake, if not always at the expense of pyruvate, is a feature of preimplantation embryo metabolism in all mammals studied.

Amanitins↗
PubMed1998

Heparin-binding epidermal growth factor significantly improves human blastocyst development and hatching in serum-free medium.

The purpose of this study was to determine the effect of heparin-binding epidermal growth factor (HB-EGF) on human embryo development in vitro from days 2 to 14 post-insemination. Embryos were cultured in a complex serum-free medium (CSFM3) in the absence and presence of 1 nM and 100 nM HB-EGF. Development to the blastocyst stage of A-C-grade embryos (A grade = highest quality) was improved in the presence of 1 nM HB-EGF from 40.7% to 65.4% and significantly increased to 71.0% in the presence of 100 nM HB-EGF (P < 0.05). Moreover, the percentage of blastocysts hatching was improved in the presence of 1 nM HB-EGF from 45.5% to 70.5% and almost doubled to 81.8% (P < 0.05) in the presence of 100 nM HB-EGF. HB-EGF promoted the development of high-grade (classed as BG1) and medium-grade (BG2) blastocysts. There was no difference in blastocyst quality between the control and HB-EGF-treated embryos as assessed by blastocyst cell number and consumption of the major energy substrates, pyruvate and glucose, measured on day 6 of culture. Further development was assessed by culturing the blastocysts on growth factor-reduced Matrigel (GFR-Matrigel). Adherence and outgrowth were observed, with these embryos producing significantly more human chorionic gonadotrophin over days 7-14 compared with those cultured on plastic (47.8 +/- 8.0 mU versus 23.0 +/- 8.6 mU). The addition of recombinant human growth factors to clinical in-vitro fertilization medium may be useful in promoting embryo development with a view to carrying out blastocyst transfers.

Blastomeres↗
PubMed1998

The use of recombinant growth factors to promote human embryo development in serum-free medium.

Research into human implantation requires embryo culture systems that yield high numbers of good quality blastocysts. One approach is to use co-culture but the presence of feeder cells and serum may confound analysis of paracrine or autocrine factors involving blastocyst implantation. An alternative approach is to produce a defined serum-free culture medium supplemented with growth factors. We have shown that the addition of Leukaemia Inhibitory Factor increased blastocyst formation from 18.4-43.6% but none of these embryos developed beyond day 7 or hatched, suggesting that additional factors are required. Development to the blastocyst stage was significantly increased to 71.0% in the presence of 100 nM heparin-binding epidermal growth factor (HB-EGF) and 81.0% of these embryos went on to hatch. No difference in blastocyst quality between the control and HB-EGF-treated embryos was found. These experiments clearly demonstrate the potential of this system to generate blastocysts in vitro. Further investigation of the normality of these blastocysts must be carried out before they are used clinically, since it has been demonstrated in other species that apparent improvements in culture conditions may be detrimental to pregnancy outcome.

Culture Media, Serum-Free↗
PubMed1998

Metabolism of pyruvate by the early human embryo.

Pyruvate is added to all media used for human in vitro fertilization and embryo culture, but its function(s) in the early embryo is unknown. We tested the possibility that pyruvate can act as an oxidizable energy source by measuring the consumption of pyruvate and oxygen by Day 2 and Day 3 human embryos, using microfluorometric techniques. Oxygen consumption (19.6 pmol/embryo per hour) could account for the oxidation of only 56% of the pyruvate consumed (13.9 pmol/embryo per hour). Oxygen was also consumed in the absence of exogenous substrates. Lactate appeared in the incubation medium with pyruvate (0.47 mM) as sole exogenous substrate at a rate of 12.1 pmol/embryo per hour, at a similar rate (10.85 pmol/embryo per hour) in the presence of 1 mM glucose and 0.47 mM pyruvate, and at 2.25 pmol/embryo per hour in the absence of exogenous substrates, suggesting that a high proportion of the pyruvate taken up by early human embryos is converted to lactate. Pyruvate uptake in the presence of UK5099, a pyruvate transport inhibitor, was reduced to 10% of control values, consistent with the presence of the monocarboxylate carrier in the human embryo plasma membrane.

Acrylates↗
PubMed1997

Programs and services to prevent pregnancy, childbearing, and poor birth outcomes among adolescents in rural areas of the southeastern United States.

PURPOSE: To illustrate how rural adolescents' needs for pregnancy prevention and improved birth outcomes are currently being addressed, and to suggest strategies for future programs. METHODS: Local and state-level informants knowledgeable about services to adolescents in the Southeastern United States were identified. Semistructured interviews were used to determine the program start date and time frame, funding sources, target population, participating counties, implementing agency or organization, specific program services, and status of program activities. These programs were categorized by the type of services offered and the population targeted. RESULTS: The most common adolescent services in the rural Southeast attempt either to improve life options of youth, reduce sexual activity, or provide prenatal and postnatal care. Unlike urban areas where there are a variety of family planning providers, in the rural Southeast, health departments are the primary source of family planning for adolescents. There are no abortion providers in most rural areas of the Southeast. The majority of rural programs that include adolescents among the population served are developed for all women rather than specifically for adolescents. Programs specific to rural adolescents are described. CONCLUSIONS: The majority of programs in the rural Southeast address only selected adolescent health issues. Successful interventions require locally supported, multipronged, intensive approaches with consistent messages targeted to high-risk populations. Evaluation tools are needed to determine the effectiveness of each component of prevention programs.

Adolescent↗
PubMed1996

Effects of metabolic inhibitors on mouse preimplantation embryo development and the energy metabolism of isolated inner cell masses.

The effects of two metabolic inhibitors, methyl palmoxirate (MP) and amino-oxyacetate (AOA), on mouse preimplantation embryo development and cell number, and inner cell mass. (ICM) cell metabolism have been examined. Two-cell embryos were cultured in media supplemented with either MP, which inhibits fatty acid oxidation, or AOA, which inhibits the transamination of glutamate into alpha-ketoglutarate. Embryos were scored for development daily. On day 5, expanded blastocysts were differentially labeled with fluorochromes to visualize TE and ICM cell nuclei, or the ICMs isolated by immunosurgery and their energy metabolism determined using microfluorometric methods. Embryos exposed to the two inhibitors developed into fully expanded blastocysts, although cell numbers of both the TE and ICM cells were significantly reduced compared to controls. The uptake of glucose in the presence of 1 mM MP or AOA did not differ from the controls, but less glucose was accountable for by lactate production. MP significantly reduced lactate production. In the presence of 4 mM AOA, the amount of glucose oxidized and the amount of lactate formed by ICMs were significantly reduced. The results indicate that the fuels used by isolated mouse ICMs vary in response to substrate availability and that fatty acids may be a potential energy source.

Aminooxyacetic Acid↗
PubMed1996

Glucose utilization during gonadotropin-induced meiotic maturation in cumulus cell-enclosed mouse oocytes.

Earlier work from this laboratory has determined that glucose plays an important role in the mechanisms regulating meiotic maturation in mammalian oocytes. In the current study, we have further explored the role of glucose in hormone-induced germinal vesicle breakdown (GVB) in an effort to better understand how glucose utilization and metabolism relate to the control of meiotic maturation in mouse cumulus cell-enclosed oocytes (CEO). When CEO were cultured in medium containing 4 mM hypoxanthine (to maintain meiotic arrest), 5.5 mM glucose, and 0.23 mM pyruvate, follicle-stimulating hormone (FSH) stimulated lactate accumulation in a time-dependent manner. Addition of 2-deoxyglucose (2-DG) to the medium at various times after the initiation of culture resulted in rapid termination of lactate production and suppression of FSH-induced GVB scored after 18 hr of culture, the effectiveness diminishing the longer the delay before addition of 2-DG. By 8 hr, addition of 2-DG was without effect on GVB. Similar effects were seen when FSH-treated CEO were washed free of glucose. In a 2-DG dose-response experiment, gonadotropin-induced lactate production was prevented, but this inhibition did not necessarily prevent GVB. The activities of six metabolic enzymes were measured in extracts of freshly isolated complexes, and in order of increasing activity were: hexokinase, 6-phosphogluconate dehydrogenase, glucose-6-phosphate dehydrogenase, phosphofructokinase, lactate dehydrogenase, and pyruvate kinase. Of the six enzymes examined, only hexokinase activity was increased in CEO exposed to FSH. CEO were cultured in microdrops in the presence or absence of FSH, and aliquots from the same microdrop were assayed for glucose, lactate, and pyruvate. In response to FSH, utilization of glucose in microdrop cultures by CEO was markedly increased and was accompanied by comparable lactate production and limited pyruvate production. Cycloheximide and alpha-amanitin both blocked FSH-induced oocyte maturation, but only cycloheximide prevented the increase in hexokinase activity and glucose consumption. These data suggest that hexokinase is an important rate-limiting enzyme for glucose utilization that is under translational control and participates in the mechanisms controlling the reinitiation of meiosis. However, stimulation of glycolytic activity does not appear to be a necessary concomitant for meiotic induction.

Animals↗
PubMed1996

Changes in atrioventricular conduction properties with refractory-period modulation.

Dofetilide, clofilium, and risotilide, three drugs known to prolong cardiac action potentials and refractory periods, were studied by using a perfused isolated rabbit heart preparation with intermittent premature pacing and bipolar surface electrograms. The rate-related effects of these drugs on atrioventricular (AV) conduction were tested by pacing at a long (400 ms) and a short (250 ms) basic cycle length (BCL). All three drugs increased refractory periods in a concentration-dependent manner in most segments of the AV axis. The maximal atrio-His (AH) conduction interval (AHmax) and delta AH (AHmax - AHmin) produced by premature pacing was decreased by the highest concentration of each drug at the 400-ms BCL, whereas only clofilium reduced AHmax and delta AH at the 250-ms BCL. Changes in delta AH correlated best with changes in the atrial functional refractory period. The His-Purkinje system conduction interval (HV), represented by delta HV, was unaffected by any drug at either BCL. These results show that if atrial or nodal refractory periods are increased sufficiently, AHmax but not AHmin was decreased at the 400-ms BCL. Because dofetilide and risotilide did not affect AHmax at the 250-ms BCL, these drugs may be less effective at preventing AV nodal reentrant tachycardias than a drug such as clofilium that displays less rate dependency.

Animals↗
PubMed1996

Calcium decreases and parathyroid hormone increases in serum of periparturient mares.

Changes in serum concentrations of Ca and parathyroid hormone (PTH) may develop in periparturient mares, may be influenced by dietary Ca, and may be associated with changes in Ca concentration of mammary secretion. Milk and blood samples were taken from eight mares on Farm A and eight on Farm B for 10 d before parturition and from four mares on each farm for 5 d postpartum. Milk Ca was measured by two commercial tests. Serum samples were analyzed for PTH and total Ca in 16 mares and for ionized Ca in six (Farm A). Parturition was induced in eight mares on Farm A and four on Farm B; no significant difference in serum Ca or PTH was found between mares with induced and spontaneous foaling. Mean serum total Ca decreased from 12.5 mg/dL to a nadir of 11 mg/dL on d 2 postpartum, and mean PTH increased from 46 pg/mL to a peak of 186 pg/mL on d 2 postpartum. Serum total Ca concentrations were lower and serum PTH concentrations were higher in Farm A mares than in Farm B mares, probably reflecting dietary Ca (.35% of DM on Farm A and .55% on Farm B). The serum PTH peak and Ca nadir occurred on d 2 postpartum, 1 d later than reported previously in dairy cows. Milk Ca concentration increased progressively from 7 d before parturition; this increase preceded, so was not dependent on, prepartal changes in serum Ca and PTH.

Animals↗
PubMed1995

Role of glucose in mouse preimplantation embryo development.

Mouse preimplantation embryos consume pyruvate preferentially during the early developmental stages, before glucose becomes the predominant energy substrate in the blastocyst. To investigate the importance of the switch to glucose utilization at the later developmental stages, mouse embryos from F1 hybrid mice (CBA/Ca x C57BL/6) were cultured from the one- and two-cell stages (22 and 46 h post hCG, respectively) for 5 days in a modified medium, M16, containing 0.33 mM pyruvate and 5 or 23 mM D + L-lactate, in the presence and absence of 1 mM glucose (M16 + G and M16 - G, respectively). Nutrient uptakes were also determined over this time. Some embryos cultured in M16 - G were transferred to M16 + G at 94 or 118 h post hCG. Embryos cultured from the two-cell stage in M16 + G exhibited the characteristic fall in pyruvate consumption between the morula and the blastocyst stage; those cultured from the two-cell stage in M16 - G compensated for the lack of glucose by consuming increasing amounts of pyruvate, from 2.78 pmol/embryo/h at 58 h post hCG to 5.21 pmol/embryo/h at 154 h post hCG. However, the percentage of embryos developing to the blastocyst stage, the hatching rate, and blastocyst cell numbers (50.6 +/- 2.5 [28] vs. 105 +/- 3.8 [37]) were all lower in this group. When exposed to glucose at 94 or 118 h post hCG, embryos cultured from the two-cell stage in M16 - G readily consumed glucose in preference to pyruvate, although the characteristic fall in pyruvate consumption was not observed. One-cell embryos cultured continuously in M16 - G were only able to develop to the morula stage, after which time they degenerated. In these embryos pyruvate was readily consumed between 22 and 94 h post hCG, before falling from 2.77 pmol/embryo/h at 83 h post hCG to 0.045 pmol/embryo/h at 130 h post hCG. Transfer of these embryos to M16 + G at 94 and 118 h post hCG did not support development to the hatching blastocyst stage.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Scanning electron microscopy and molecular modeling of inhibition of calcium oxalate monohydrate crystal growth by citrate and phosphocitrate.

Binding of citrate and phosphocitrate to calcium oxalate monohydrate crystals has been studied using scanning electron microscopy (SEM) and molecular modeling. Phosphocitrate structure has been resolved using low temperature X-ray analysis and ab initio computational methods. The (-1 0 1) crystal surface of calcium oxalate monohydrate is involved in binding of citrate and phosphocitrate, as shown by SEM and molecular modeling. Citrate and phosphocitrate conformations and binding energies to (-1 0 1) faces have been obtained and compared to binding to another set of calcium-rich planes (0 1 0). Difference in inhibitory properties of these compounds has been attributed to better coordination of functional groups of phosphocitrate with calcium ions in (-1 0 1). Relevance of this study to design of new calcium oxalate monohydrate inhibitors is discussed.

Calcium Oxalate↗
PubMed1995

Glucose utilization by components of the mouse conceptus during early embryogenesis.

Mouse conceptuses were collected from female mice between day 6.5 and day 9.5 of pregnancy, dissected into their component parts and incubated for 2.5 h at 37 degrees C in droplets of Hepes-buffered medium containing 1 or 5 mmol glucose l-1 supplemented with 0.33 mmol pyruvate l-1 plus either 1 or 5 mmol (DL) lactate l-1 under oil. Glucose disappearance and lactate appearance were measured enzymatically at the end of incubation. High glucose concentration doubled utilization of this substrate per microgram of embryonic protein, but the change in lactate concentration had no effect on glucose turnover. Over the whole period of development studied, tissue from the ectoplacental cone exhibited the lowest rate of glucose turnover of all tissues isolated. At day 8.5, there was little difference between yolk sac and embryonic tissues, but by day 9.5, the yolk sac had a higher rate of utilization of glucose than did embryonic tissues. By this time, the embryonic tissues had started to show some metabolic differentiation, with head and visceral tissue exhibiting 20% higher turnover of glucose than that of body tissue. Overall, the rate of glucose utilization fell as development progressed and the estimate of the relative rate of glucose utilization on day 9.5 was half the value on day 6.5.

Animals↗
PubMed1994

Comparison of pyruvate uptake by embryos derived from conception and non-conception natural cycles.

The uptake of pyruvate by human embryos derived from natural cycles in the first 24 h following fertilization was examined. Since only one egg was obtained and therefore only one embryo transferred to the woman, it was possible to relate pyruvate consumption by a particular embryo to the outcome of that cycle (pregnancy or no pregnancy). The results showed that embryos have a wide range of pyruvate uptake values (2-53 pmol/embryo/h) but that this variation was reduced significantly to an intermediate range of values in those embryos that were able to implant (10-30 pmol/embryo/h). An association was found between embryo morphology and pyruvate consumption. Morphologically good embryos were more likely to implant if they demonstrated an intermediate pyruvate uptake. However, poor embryos did not implant even if they had a pyruvate uptake of 10-30 pmol/embryo/h. No relationship was found between the type of infertility and pyruvate consumption of individual embryos. It is suggested that the ability of an embryo to implant is multifactorial and that both morphology and pyruvate uptake may be factors.

Embryo, Mammalian↗
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↗