5'-Nucleotidase of human placental trophoblastic microvilli possesses cobalt-stimulated FAD pyrophosphatase activity

An enzyme with FAD pyrophosphatase activity was extracted from human placental syncytiotrophoblast microvilli and purified to near-homogeneity. The enzyme has been identified as 5'-nucleotidase by several criteria. Throughout purification, parallel increases in the specific activities of FAD pyrophosphatase and AMP phosphatase were observed. The enzyme was a glycoprotein with a subunit molecular weight of 74,000. EDTA treatment resulted in a marked decline in both activities, and restoration of FAD pyrophosphatase activity but not 5'-nucleotidase activity was accomplished by the addition of Co2+ or, to a lesser extent, Mn2+. The substrate specificity of the 5'-nucleotidase activity that we observed agreed closely with the results of others. The pyrophosphatase activity was relatively specific for FAD. ADP, ATP, NAD(H), and FMN were not hydrolyzed, and ADP strongly inhibited both activities. For FAD pyrophosphatase activity, a Km of 1.2 x 10(-5) M and a Vmax of 1.1 mumol/min/mg protein were determined in assays performed in the presence of Co2+. In the absence of added Co2+, the Vmax declined but the Km was unchanged. For 5'-nucleotidase (AMP as substrate) the Km was 4.1 x 10(-5) M and the Vmax 109 mumol/min/mg protein. Hydrolysis of FMN to riboflavin was observed in partially purified detergent extracts of microvilli that contained alkaline phosphatase activity and lacked FAD pyrophosphatase and 5'-nucleotidase activity. The presence of both FAD pyrophosphatase and FMN phosphatase activities in syncytiotrophoblast microvilli supports the view that the placental uptake of vitamin B2 involves the hydrolysis of FAD and FMN to riboflavin which is then absorbed, a sequence postulated for intestinal absorption and liver uptake.     

Developmental expression of bovine adrenocortical steroid hydroxylases. Regulation of P-450(17 alpha) expression leads to episodic fetal cortisol production

The developmental expression of adrenocortical steroid hydroxylases was studied in bovine fetuses from 40 to 280 days gestational age. The expression of P-450(17 alpha) is first detected at a gestational age of 50 days and reaches a maximum at 60-70 days. The expression of P-450(17 alpha) then declines and is nondetectable at a gestational age of 100 days. P-450(17 alpha) is not expressed again until about 240 days, i.e. shortly before birth (approximately 280 days). P-450scc, P-450c21, P-450(11 beta) and adrenodoxin were present in fetal adrenals throughout gestation. This "on-off-on" pattern of P-450(17 alpha) expression during fetal development was associated with a corresponding episodic production of cortisol. Immunoreactive corticotropin (ACTH) levels in fetal plasma were elevated in small fetuses (corresponding to less than or equal to 100 days) and in near-term fetuses (corresponding to greater than 250 days) compared with those in mid-gestation fetuses. In primary culture, adrenal cells from mid-gestation fetuses contained no detectable P-450(17 alpha) but rapidly responded to ACTH with an increase in P-450(17 alpha) protein and mRNA. The tissue specificity of the developmental patterns is emphasized by the fact that both P-450(17 alpha) and P-450scc were detectable throughout the development of the fetal testes, whereas only P-450scc was detectable in fetal bovine ovary prior to 200 days. Thus, in fetal bovine adrenal it appears that ACTH is the major regulatory factor effecting the intermittent presence of P-450(17 alpha), whereas the presence of the other steroid hydroxylases is either regulated by additional factors or shows a much different sensitivity to ACTH.     

Emotional and behavioral reactions to facially deformed patients before and after craniofacial surgery

The present experiment investigated whether observers' emotional and behavioral reactions to facially deformed patients could be substantially improved by surgical procedures conducted by well-trained specialists in an experienced multidisciplinary team. Also investigated was the hypothesis that emotional states mediate the effects of physical attractiveness and facial deformity on social interaction. Twenty patients between the ages of 3 months and 17 years were randomly selected from over 2000 patients' files of Kenneth E. Salyer of Dallas, Texas. Patient diagnoses included facial clefts, hypertelorism, Treacher Collins syndrome, and craniofacial dysostoses (Crouzon's and Apert's syndromes). Rigorously standardized photographs of patients taken before and after surgery were shown to 22 "naive" raters ranging in age from 18 to 54 years. Raters were asked to predict their emotional and behavioral responses to the patients. These ratings indicated that observers' behavioral reactions to facially deformed children and adolescents would be more positive following craniofacial surgery. Similarly, the ratings indicated that observers' emotional reactions to these patients would be more positive following surgery. The results are discussed in terms of current sociopsychologic theoretical models for the effects of attractiveness on social interaction. A new model is presented that implicates induced emotional states as a mediating process in explaining the effects of attractiveness and facial deformity on the quality of social interactions. Limitations of the current investigation and directions for future research are also discussed.     

Direct effects of oestradiol-17 beta and prostaglandin E-2 in protecting pig corpora lutea from a luteolytic dose of prostaglandin F-2 alpha

Implants containing vehicle or oestradiol-17 beta (10 mg) were placed into pairs of corpora lutea (CL) with and without prostaglandin F-2 alpha (PGF-2 alpha) (100 micrograms) on Day 11 and CL were collected on Day 19, in cyclic gilts (Exp. 1). The results demonstrated that CL implanted with PGF-2 alpha with or without oestradiol-17 beta had a markedly lower (P less than 0.01) weight (mg) and progesterone concentration (ng/mg) than CL with vehicle-or oestradiol-17 beta-implanted or unimplanted CL, which were similar (149 and 7.2 vs. 304 and 49.6, respectively). In Exp. 2, CL implanted with vehicle, oestradiol-17 beta or PGE-2 remained fully functional until Day 19, whereas CL implanted with oestradiol-17 beta +/- PGF-2 alpha and PGE-2 + PGF-2 alpha exhibited lower (P less than 0.05) weight and progesterone concentrations; CL implanted with PGE-2 + PGF-2 alpha were heavier (P less than 0.05) and tended (P less than 0.10) to have greater progesterone concentrations than CL implanted with oestradiol-17 beta + PGF-2 alpha. In Exp. 3, a dose-dependent (P less than 0.05) effect of PGE-2 on preventing regression induced by PGF-2 alpha was observed on Day 19. These data demonstrate a direct effect of PGE-2, but not of oestradiol-17 beta in protecting the CL against luteolysis induced by PGF-2 alpha.     

Physiological and enzymatic activity of pepper seeds (Capsicum annuum) during priming

Pepper ( Capsicum annuum L. cv. Keystone Resistance Giant 3) seeds were monitored during priming to determine if seed treatments which accelerate the rate of germination could be correlated with specific physiological changes within the seeds. Pepper seeds primed with −0.90 and −1.35 MPa NaCl solutions at 23°C for 18 days did not completely equilibrate with the osmotic potential of the priming solution. Seed respiratory rates indicated that priming extends the lag phase of germination following imbibition. Soluble protein levels increased 115% in primed seeds, and the uptake and incorporation of [¹⁴C(U)] labelled amino acids into the acid insoluble fraction increased throughout the priming treatments. Alcohol dehydrogenase (EC 1.1.1.1, anaerobic metabolism), glucose-6-phosphate dehydrogenase (EC 1.1.1.49) and 6-phosphogluconate dehydrogenase (EC 1.1.1.44, pentose phosphate pathway) activities remained stable throughout the priming treatment, but were higher after 6 days. than the water-imbibed controls. Aldolase (EC 4.2.1.1. glycolysis) and isocitrate lyase (EC 4.1.3.1, glyoxylate cycle) activities increased with imbibition and were 61 and 56% (respectively) higher in primed seeds as compared to the water-imbibed controls after 12 days. Treatment with the −0.90 MPa NaCl solution was more effective than the −1.35 MPa solution in improving the rate of germination, yet there were no significant differences between the protein concentrations or enzyme activities of the two priming treatments. However, the incorporation of labelled amino acids into pepper seeds was significantly higher in the −0.90 MPa priming treatment.     

Expectations of assisted conception for infertility.

OBJECTIVE--To provide reliable prognostic information for couples seeking assisted conception. DESIGN--Analysis of four years'' practice (1988-91). SETTING--Private university service linked with NHS reproductive medicine services. PATIENTS--804 couples with various causes of subfertility, median duration five years, median age of women 34 years. INTERVENTIONS--1280 completed cycles: 950 in vitro fertilisation, 144 gamete intrafallopian transfer, and 186 intrauterine insemination and superovulation. MAIN OUTCOME MEASURES--Pregnancy and birth rates per cycle and cumulative pregnancy and take home baby rates per couple. RESULTS--In women under 40 years and men with normal sperm, whatever the cause of infertility, results with in vitro fertilisation improved steadily reaching a pregnancy rate per cycle of 30% (95% confidence interval 26% to 35%) during 1990-1 and birth rate per cycle of 29% (23% to 35%) in 1990. Pregnancy and birth rates for gamete intrafallopian transfer were 36% (28% to 44%) and 26% (17% to 37%) and for intrauterine insemination 18% (12% to 24%) and 16% (10% to 22%). After six cycles cumulative probability of pregnancy was 82% and cumulative take home baby rate 70%. Considering only in vitro fertilisation and gamete intrafallopian transfer after four cycles the pregnancy rate was 78% (66% to 91%). CONCLUSIONS--Conception is less likely in women over 40 and men with sperm dysfunction. For other couples the prognosis for a live birth is at least as good as for fertile couples if they persist with treatment.     

Fluid dynamics and microscale chemical movement in the chemosensory appendages of the lobster, Homarus americanus

Every chemosensory structure has a boundary layer surroundingit through which chemical signals must pass before contactingreceptor cells. Fluid motion in this boundary layer is slowand odor movement is mainly by diffusion. The boundary layerstructure depends upon external fluid velocities and the morphologyof the appendage. High-speed (10–200 Hz) electrochemicalrecordings from microchemical electrodes were used to quantifychemical transport in the microscale environment of three morphologicallydifferent chemosensory appendages of the lobster, Homarus americanus:lateral antennule, medial antennule and walking legs. Controlledpulses of the odor tracer (dopamine) were delivered to the threeappendages at three different flow speeds (0, 3, 6 cm/s). Theamplitudes of the pulses increased with increasing flow speed,indicating that boundary layer thickness decreased with increasingflow speed. Larger pulse amplitudes were measured in the walkinglegs than in the lateral or medial antennules at all flow speeds.In addition, larger amplitudes were recorded in the medial antennulethan the lateral antennule. Changes in pulse amplitude withincreasing flow speed were larger than changes in pulse duration.These results demonstrate that pulse amplitude is affected morethan pulse duration by boundary layer thickness and that themorphology of the receptor strucure helps determine the structureof signals arriving at receptor cells. This may explain whyanimals have adopted sampling strategies that reduce boundarylayer thickness.     

A fluorescent sterol probe study of cholesterol/phospholipid membranes

The behavior of dehydroergosterol in -α-dimyristoylphosphatidylcholine (DMPC) unsonicated multilamellar liposomes was characterized by absorption spectroscopy and fluorescence measurements. Dehydroergosterol exhibited a lowered absorption coefficient in multilamellar liposomes whiel the steady-state fluorescence anisotropy of dehydroergosterol in these membranes decreased significantly with increasing dehydroergosterol concentration, suggesting membrane sterol-sterol interactions. The comparative steady-state anisotropy of 0.9 mole percent dehydroergosterol in multilamellar liposomes was lower than in small unilamellar vesicles suggesting different sterol environments for dehydroergosterol. Dehydroergosterol fluorescence lifetime was relatively independent of membrane sterol content and yielded similar values in sonicated and unsonicated model membranes. In multilamellar liposomes containing 5 mole percent cholesterol, the gel-to-liqui crystalline phase transition of DMPC detected by 0.9 mole percent dehydroergosterol was significantly broadened when compared to the phase transition detected by dehydroergosterol in the absence of membrane cholesterol (Smutzer, G. et al. (1986) Biochim. Biophys. Acta 862, 361–371). In multilamellar liposomes containing 10 mole percent cholesterol, the major fluorescence lifetime of dehydroergosterol did not detect the gel-to-liquid crystalline phase transition of DMPC. Time-correlated fluorescence anisotropy decays of dehydroergosterol in DMPC multilamellar liposomes in the absence and presence of 5 mole percent cholesterol exhibited a single rotational correlation time near one nanosecond that was relatively independent of temperature and low concentrations of membrane cholesterol. The limiting anisotropy of 0.9 mole percent dehydroergosterol decreased above the gel-to-liquid crystalline phase transition in membranes without cholesterol and was not significantly affected by the phase transition in membranes containing 5 mole percent cholesterol. These results suggested hindered rotational diffusion of dehydroergosterol in multilamellar liposomes. Lifetime and time-correlated fluorescence measurements of 0.9 mole percent dehydroergosterol in multilamellar liposomes further suggested this fluorophore was detecting physical properties of the bulk membrane phospholipids in membranes devoid of cholesterol and was detecting sterol-rich regions in membranes of low sterol concentration.     

Changes in Leydig cell ultrastructure and function during pubertal development in the boar

Changes in the ultrastructure of Leydig cells during pubertal development in the boar (40 to 250 days of age) were assessed using quantitative morphometric procedures, and the results were compared to the in vitro steroid-producing capacity and gonadotropin sensitivity of testicular tissue obtained from the same boars. Volume of individual Leydig cells declined through 100 days of age, increased rapidly to a peak at 130-160 days (i.e., puberty), and then declined to intermediate levels by 220-250 days of age. The pattern of change in the number of intracellular organelles per Leydig cell was very similar to the change that occurred in Leydig cell volume. Changes in the total intracellular volume occupied by each type of organelle were highly correlated with changes in Leydig cell volume (r = 0.40-0.99, p less than 0.01), and this was particularly true for the nucleus (r = 0.63), mitochondria (r = 0.88), smooth endoplasmic reticulum (SER; r = 0.97), and total cytoplasm (r = 0.99) of the boar Leydig cell. In vitro production of testosterone and estradiol, expressed per Leydig cell, also peaked at 130-160 days, and was highly correlated to average Leydig cell volume, volume of SER, and number and total volume of mitochondria (r = 0.63-0.84; p less than 0.01). Observations in the present study indicated that onset of puberty in boars coincides with a dramatic increase in average Leydig cell size and SER volume per Leydig cell, accompanied by an increase in number of other intracellular organelles, including mitochondria, lysosomes, and lipid droplets, and a peak in the steroid-producing capacity per Leydig cell. A decline in Leydig cell size, intracellular organelles, and sensitivity to gonadotropin stimulation occurred postpubertally.