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Sample records for quantitative human chorionic

  1. Quantitative human chorionic gonadotropin analysis. I. Comparison of an immunoradiometric assay and a radioimmunoassay

    SciTech Connect

    Shapiro, A.I.; Wu, T.F.; Ballon, S.C.; Lamb, E.J.

    1984-01-01

    An immunoradiometric assay (IRMA) for the quantitative analysis of human chorionic gonadotropin (hCG) was evaluated for specificity, sensitivity, accuracy and precision. The results were compared with those of the conventional radioimmunoassay (RIA) used in our laboratory. The IRMA is a solid-phase, double-antibody immunoassay that sandwiches the intact hCG molecule between the two antibodies. It has specificity, accuracy, and precision which are similar to those of the RIA. The RIA is based upon the assumptions that the antigenicity of the tracer is not altered by the iodination process and that the antibody reacts equally with all of the antigens, including the radiolabeled antigen. The IRMA does not use radiolabeled antigens and thus is free of the assumptions made in the conventional RIA. The IRMA may be more accurate at the lower limits of the assay because it does not require logarithmic transformations. Since the IRMA does not measure the free beta-subunit of hCG, it cannot be endorsed as the sole technique to quantitate hCG in patients with gestational trophoblastic neoplasia until the significance of the free beta-subunit in these patients is determined.

  2. Human chorionic gonadotrophin and sport.

    PubMed Central

    Kicman, A T; Brooks, R V; Cowan, D A

    1991-01-01

    Human chorionic gonadotrophin (hCG) is a glycoprotein hormone which is produced in large amounts during pregnancy and also by certain types of tumour. The biological action of hCG is identical to that of luteinizing hormone, although the former has a much longer plasma half-life. Some male athletes use pharmaceutical preparations of hCG to stimulate testosterone production before competition and/or to prevent testicular shutdown and atrophy during and after prolonged courses of androgen administration. Testosterone administration can be detected by measuring the ratio of concentrations of testosterone to epitestosterone (T/E). An athlete is often considered to have failed a drug test if the urinary T/E ratio is greater than 6. In contrast, hCG administration stimulates the endogenous production of both testosterone and epitestosterone without increasing the urinary T/E ratio above normal values. Although the administration of hCG was banned by the International Olympic Committee (IOC) in 1987, no definitive test for hCG has been approved by the IOC. Currently, the only way of measuring small concentrations of hCG is by immunoassay, and this does not have a discriminating power as great as gas-liquid chromatography with mass-spectrometry which is necessary to satisfy IOC requirements. Extraction procedures and chromatographic steps could be introduced before using a selected immunoassay for hCG to meet these requirements. PMID:1751893

  3. Human chorionic gonadotropin measurements in parathyroid carcinoma

    PubMed Central

    Rubin, Mishaela R; Bilezikian, John P; Birken, Steven; Silverberg, Shonni J

    2010-01-01

    Objective Preoperatively, it is difficult to differentiate between parathyroid cancer (PtCa) and severe primary hyperparathyroidism (PHPT) due to a benign tumor. Human chorionic gonadotropin (hCG) is a tumor marker in trophoblastic and nontrophoblastic cancers and hyperglycosylated hCG is increased in hCG-secreting malignancies. We investigated whether hCG can distinguish PtCa cancer from benign disease and add prognostic information. Design Observational study. Methods Measurement of urinary hCG (total and malignant isoforms) and serum malignant hCG in 8 subjects with PtCa and in 18 subjects with PHPT (measurement of urine in ten and serum in eight). Results Total urinary hCG was normal in the benign PHPT control subjects (range: 0–17 fmol/mg Cr; nl < 50). In the PtCa subjects, three had normal total urinary hCG levels and survived complication free for at least 2 years; three had persistently elevated total urinary hCG levels (range: 217–1986 fmol/mg Cr) and sustained hip fracture (n = 3) and died (n = 2) within 3 and 6 months respectively; two had a rise in total urinary hCG and had hip fracture (n = 1) and died (n = 2) within 4 and 10 months respectively. Elevated urinary hCG was of the malignant hyperglycosylated isoform. Serum malignant hyperglycosylated hCG values in all of the cancer patients exceeded the maximal serum malignant hCG level of the PHPT subjects with benign disease (3.77 pmol/l). Conclusion hCG, especially itshyperglycosylated isoform, might add diagnostic and prognostic information in PtCa. Further studies would help to elucidate the role of hCG as a potential tumor marker in this disease. PMID:18625691

  4. Biologically Active Chorionic Gonadotropin: Synthesis by the Human Fetus

    NASA Astrophysics Data System (ADS)

    McGregor, W. G.; Kuhn, R. W.; Jaffe, R. B.

    1983-04-01

    The kidney, and to a slight extent the liver, of human fetuses were found to synthesize and secrete the α subunit common to glycoprotein hormones. Fetal lung and muscle did not synthesize this protein. Since fetal kidney and liver were previously found to synthesize β chorionic gonadotropin, their ability to synthesize bioactive chorionic gonadotropin was also determined. The newly synthesized hormone bound to mouse Leydig cells and elicited a biological response: namely, the synthesis of testosterone. These results suggest that the human fetus may participate in metabolic homeostasis during its development.

  5. EFFECT OF BROMODICHLOROMETHANE ON HUMAN TROPHOBLAST CHORIONIC GONADOTROPHIN SECRETION

    EPA Science Inventory

    Effect of Bromodichloromethane on Human Trophoblast Chorionic Gonadotrophin Secretion

    Jiangang Chen1, Twanda L. Thirkill1, Peter N. Lohstroh1, Susan R. Bielmeier2, Michael G. Narotsky3, Deborah S. Best3, Randy A. Harrison3, Kala Natarajan1, Rex A. Pegram3, Gordon C. Dougla...

  6. Detection of human chorionic gonadotrophin misuse in sports.

    PubMed

    Delbeke, F T; Van Eenoo, P; De Backer, P

    1998-05-01

    The peptide hormone human chorionic gonadotrophin (hCG) was measured in the urine of 5663 male athletes collected for doping analysis in Flanders during the period 1993-1996. Using the Abbott IMx hCG procedure, the free and whole molecule associated beta-subunit of human chorionic gonadotrophin was determined. Statistical evaluation of the data results in a far outside value of 2.28 mIU/ml. The decision limit, i.e. the concentration at which a sportsman will be considered positive, is set at 5 mIU/ml to ensure with the greatest possible degree of certainty that no false positive results are reported. PMID:9657371

  7. Pattern of human chorionic gonadotropin binding in the polycystic ovary

    SciTech Connect

    Brawer, J.; Richard, M.; Farookhi, R. )

    1989-08-01

    The histologic evolution of polycystic ovaries in the estradiol valerate-treated rat coincides with the development of a unique plasma pattern of luteinizing hormone. To assess the role of luteinizing hormone in polycystic ovaries, it is necessary to evaluate the luteinizing hormone sensitivity of the specific tissues in the polycystic ovary. Therefore, we examined the pattern of luteinizing hormone binding sites in polycystic ovaries. Rats at 4 or 8 weeks after estradiol valerate treatment each received an intrajugular injection of iodine 125-labeled human chorionic gonadotropin. Some rats also received a 1000-fold excess of unlabeled human chorionic gonadotropin in the same injection. Ovaries were prepared for autoradiography. Dense accumulations of grains occurred over the theca of normal and atretic secondary follicles in all ovaries and over clusters of secondary interstitial cells. The iodine label was variable over the typically hypertrophied theca of precystic follicles. The theca of definitive cysts showed little or no label. These results indicate that cyst formation coincides with the loss of luteinizing hormone/human chorionic gonadotropin binding to the affected follicles.

  8. Human chorionic gonadotropin induces human macrophages to form intracytoplasmic vacuoles mimicking Hofbauer cells in human chorionic villi.

    PubMed

    Yamaguchi, Munekage; Ohba, Takashi; Tashiro, Hironori; Yamada, Gen; Katabuchi, Hidetaka

    2013-01-01

    The most characteristic morphological feature of macrophages in the stroma of placental villi, known as Hofbauer cells, is their highly vacuolated appearance. They also show positive immunostaining for human chorionic gonadotropin (hCG) and express messenger ribonucleic acid of the luteinizing hormone/chorionic gonadotropin receptor with a deletion of exon 9 (LH/CG-R Δ9). Maternal hCG enters fetal plasma through the mesenchyme of the placental villi and promotes male sexual differentiation in early pregnancy; therefore, excess hCG may induce aberrant genital differentiation and hCG must be adjusted at the fetomaternal interface. We hypothesized that hCG is regulated by Hofbauer cells and that their peculiar vacuoles are involved in a cell-specific function. To assess the morphological modification and expression of LH/CG-R Δ9 in human macrophages after hCG exposure, the present study examined phorbol 12-myristate 13-acetate (PMA)-treated THP-1 cells, a human monocyte-macrophage cell line. hCG induced transient vacuole formation in PMA-treated THP-1 cells, morphologically mimicking Hofbauer cells. Immunocytochemistry showed that PMA-treated THP-1 cells incorporated hCG but not luteinizing hormone or follicle-stimulating hormone. Western blotting analyses demonstrated that PMA-treated THP-1 cells expressed an immunoreactive 60-kDa protein, designated as endogenous LH/CG-R Δ9. hCG induced a transient reduction in the LH/CG-R Δ9, which was synchronous with the appearance of cytoplasmic vacuoles. In conclusion, human macrophages regulating hCG via cytoplasmic LH/CG-R Δ9 mimic the morphological characteristics of Hofbauer cells. Their vacuoles may be associated with their cell-specific function to protect the fetus from exposure to excess maternal hCG during pregnancy. PMID:23128164

  9. Luteinizing hormone/human chorionic gonadotropin receptors in breast cancer.

    PubMed

    Meduri, G; Charnaux, N; Loosfelt, H; Jolivet, A; Spyratos, F; Brailly, S; Milgrom, E

    1997-03-01

    Recent studies have suggested that human choriogonadotropin (hCG), in addition to its function in regulating steroidogenesis, may also play a role as a growth factor. Immunocytochemistry using two different monoclonal antibodies (LHR29 and LHR1055) raised against the human luteinizing hormone/human chorionic gonadotropin (LH/hCG) receptor allowed us to detect this receptor in breast cancer cell lines (T47D, MCF7, and ZR75) in individual cancer biopsies and in benign breast lesions. The receptor was also present in epithelial cells of normal human and sow breast. In the latter, its concentration increased after ovulation. The presence of LH/hCG receptor mRNA was confirmed by reverse transcription-PCR using primers extending over exons 2-4, 5-11, and 9-11. The proportion of LH/hCG-receptor positive cells and the intensity of the immunolabeling varied in individual biopsies, but there was no obvious correlation with the histological type of the cancer. These results are compatible with previous studies suggesting that during pregnancy, hCG is involved in the differentiation of breast glandular epithelium and that this hormone may play an inhibitory role in mammary carcinogenesis and in the growth of breast tumors. PMID:9041186

  10. Up-regulation of placental leptin by human chorionic gonadotropin.

    PubMed

    Maymó, Julieta L; Pérez Pérez, Antonio; Sánchez-Margalet, Víctor; Dueñas, José L; Calvo, Juan Carlos; Varone, Cecilia L

    2009-01-01

    Leptin, the 16,000 molecular weight protein product of the obese gene, was originally considered as an adipocyte-derived signaling molecule for the central control of metabolism. However, leptin has been suggested to be involved in other functions during pregnancy, particularly in placenta, in which it was found to be expressed. In the present work, we have found that recombinant human chorionic gonadotropin (hCG) added to BeWo choriocarcinoma cell line showed a stimulatory effect on endogenous leptin expression, when analyzed by Western blot. This effect was time and dose dependent. Maximal effect was achieved at hCG 100 IU/ml. Moreover, hCG treatment enhanced leptin promoter activity up to 12.9 times, evaluated by transient transfection with a plasmid construction containing different promoter regions and the reporter gene luciferase. This effect was dose dependent and evidenced with all the promoter regions analyzed, regardless of length. Similar results were obtained with placental explants, thus indicating physiological relevance. Because hCG signal transduction usually involves cAMP signaling, this pathway was analyzed. Contrarily, we found that dibutyryl cAMP counteracted hCG effect on leptin expression. Furthermore, cotransfection with the catalytic subunit of PKA and/or the transcription factor cAMP response element binding protein repressed leptin expression. Thereafter we determined that hCG effect could be partially blocked by pharmacologic inhibition of MAPK pathway with 50 microM PD98059 but not by the inhibition of the phosphatidylinositol 3-kinase pathway with 0.1 microm wortmannin. Moreover, hCG treatment promoted MAPK kinase and ERK1/ERK2 phosphorylation in placental cells. Finally, cotransfection with a dominant-negative mutant of MAPK blocked the hCG-mediated activation of leptin expression. In conclusion, we provide some evidence suggesting that hCG induces leptin expression in trophoblastic cells probably involving the MAPK signal transduction

  11. Metabolic and renal clearance rates of purified human chorionic gonadotropin.

    PubMed Central

    Wehmann, R E; Nisula, B C

    1981-01-01

    The metabolic clearance rate (MCR) and renal clearance rate (RCR) of human chorionic gonadotropin (hCG) were measured in healthy young men and women using techniques of continuous intravenous infusion and rapid intravenous injection of unlabeled, highly purified hCG. Seven subjects received 4 d of infusion at a rate of 0.2 microgram/min, followed by an additional 4 d of infusion at 0.8 microgram/min. Mean serum levels of hCG established at these rates of infusion were 61.1 +/- 3.3 and 237 +/- 16 ng/ml, respectively (mean +/- SEM). The MCR determined at the low infusion rate was not significantly different from that determined at the higher infusion rate (1.83 +/- 0.09 vs. 1.95 +/- 0.14 ml/min per m2). The mean MCR for all subjects was 1.88 +/- 0.08 ml/min per m2. The MCR was not significantly different between men amd women (2.04 +/- 0.13 vs. 1.76 +/- 0.07 ml/min per m2). The RCR also did not vary between low and high infusion rates (0.40 +/- 0.03 vs. 0.40 +/- 0.04 ml/min per m2). The mean RCR for all subjects was 0.40 +/- 0.02 ml/min per m2. There was no difference in RCR between men and women (0.42 +/- 0.05 vs. 0.39 +/- 0.03 ml/min per m2). Six subjects were given 1.0 mg of highly purified hCG by rapid intravenous injection. Initial serum levels of hCG were 300-400 ng/ml, and the subsequent disappearance curve was multiexponential over 8-10 d. The disappearance curve of hCG in each subject was fitted to a biexponential equation. The rapid component t1/2 was 5.97 +/- 0.63 h and the slow component t1/2 was 35.6 +/- 8.0 h. We conclude that the MCR of purified hCG in man is about 2 ml/min per m2 and the RCR is 0.4 ml/min per m2; these parameters are concentration independent and do not differ significantly between healthy young men and women. PMID:7251859

  12. Reproductive performance in sows treated with estradiol benzoate/human chorionic gonadotrophin combination at weaning.

    PubMed Central

    Friendship, R M; Bosu, W T; King, G J

    1982-01-01

    The effects of estradiol benzoate plus human chorionic gonadotrophin administered on the day of weaning on the reproductive performance in sows was studied. The treatment advanced the onset of estrus but the duration of estrus was slightly prolonged. Breeding on the post-weaning estrus was associated with poor pregnancy rate and smaller litters. The treatment did not cause permanent damage to the reproductive tract as the reproductive performance of the treated sows after subsequent breedings were unaffected. The possible causes of the poor fertility in the estradiol benzoate plus human chorionic gonadotrophin treated sows are discussed. PMID:6890868

  13. 21 CFR 862.1155 - Human chorionic gonadotropin (HCG) test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Human chorionic gonadotropin (HCG) test system. 862.1155 Section 862.1155 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  14. 21 CFR 862.1155 - Human chorionic gonadotropin (HCG) test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Human chorionic gonadotropin (HCG) test system. 862.1155 Section 862.1155 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  15. 21 CFR 862.1155 - Human chorionic gonadotropin (HCG) test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Human chorionic gonadotropin (HCG) test system. 862.1155 Section 862.1155 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical...

  16. EFFECT OF BROMODICHLOROMETHANE ON CHORIONIC GONADOTROPHIN SECRETION BY HUMAN PLACENTAL TROPHOBLAST CULTURES

    EPA Science Inventory

    EFFECT OF BROMODICHLOROMETHANE ON CHORIONIC GONADOTROPHIN SECRETION BY HUMAN PLACENTAL TROPHOBLAST CULTURES

    Jiangang Chen1, Gordon C. Douglas1?,Twanda L. Thirkill1?, Peter N. Lohstroh1, Susan R. Bielmeier2, Michael G. Narotsky3, Deborah S. Best3, Randy A. Harrison3, Kala ...

  17. Solid-phase competitive and sandwich-type erythro-immunoassays for human chorionic gonadotropin.

    PubMed

    Gupta, S K; Guesdon, J L; Avrameas, S; Talwar, G P

    1985-06-25

    A simple '1-step' competitive erythro-immunoassay for human chorionic gonadotropin (hCG) employing V-shaped well microtitration plates coated with monoclonal anti-beta-hCG antibody has been described. hCG of the test sample competes with the antigen-coupled sheep erythrocytes for binding to the antibody on the solid surface. The assay is able to detect up to 31.25 ng hCG/ml. A higher sensitivity enabling detection up to 0.25 ng hCG/ml is attained by the sandwich erythro-immunoassay using a chimera antibody prepared by coupling monoclonal anti-alpha-hCG antibody to an affinity-purified polyclonal antibody specific for sheep erythrocytes. This assay is amenable to the qualitative as well as quantitative use as described. The urinary components do not interfere in the assay. Results obtained by this assay on 47 human urine samples correlated well with the values obtained by '2-step' sandwich enzyme immunoassay and radioimmunoassay. PMID:2409175

  18. Ovarian stimulation with human chorionic gonadotropin and equine chorionic gonadotropin affects prostacyclin and its receptor expression in the porcine oviduct.

    PubMed

    Małysz-Cymborska, I; Andronowska, A

    2015-10-01

    Prostaglandins are well-known mediators of crucial events in the female reproductive tract, eg, early embryo development and implantation. Prostacyclin (PGI2) is the most synthesized prostaglandin in the human oviduct during the postovulatory period, indicating its important role in supporting and regulating the oviductal environment. The present study was undertaken to determine the influence of insemination and ovarian stimulation with human chorionic gonadotropin (hCG)/equine chorionic gonadotropin (eCG) on PGI2 synthesis in the porcine oviduct on day 3 post coitus. Mature gilts (n = 25) were assigned into 2 experiments. In experiment I, gilts were divided into cyclic (control; n = 5) and inseminated (control; n = 5) groups. In experiment II, there were 3 groups of animals: inseminated (n = 5), induced ovulation/inseminated (750 IU eCG, 500 IU hCG; n = 5), and superovulated/inseminated (1,500 IU eCG, 1,000 IU hCG; n = 5) gilts. Parts of oviducts (isthmus and ampulla) were collected 3 days after phosphate-buffered saline treatment (cyclic gilts of experiment I) or insemination (all other groups). Expression of messenger RNA for PGI2 synthase (PGIS) and its receptor (IP) was measured by real-time reverse transcription polymerase chain reaction (real-time RT PCR) and protein levels using Western blots. Concentrations of the PGI2 metabolite 6-keto PGF1α were evaluated by enzyme immunoassay and localization of PGIS and IP in the oviductal tissues using immunohistochemical staining. Insemination by itself increased PGIS protein levels in the oviductal isthmus (P < 0.05) and IP protein expression in the ampulla (P < 0.05). The concentration of 6-keto PGF1α increased significantly in the oviductal ampulla after insemination (P < 0.05). Induction of ovulation decreased IP protein levels in the oviductal ampulla (P < 0.05), whereas superovulation reduced IP levels in both parts of the oviduct (P < 0.01). Synthesis of 6-keto PGF1α was reduced by induction of ovulation

  19. Stimulation of Spermiation by Human Chorionic Gonadotropin and Carp Pituitary Extract in Grass Puffer, Takifugu niphobles

    PubMed Central

    Goo, In Bon; Park, In-Seok; Gil, Hyun Woo; Im, Jae Hyun

    2015-01-01

    Spermiation was stimulated in the mature grass puffer, Takifugu niphobles, with an injection of human chorionic gonadotropin (HCG) or carp pituitary extract (CPE). Spermatocrit and sperm density were reduced, but milt production was increased in both the HCG and CPE treatment groups relative to those in the control group (P <0.05). These results should be useful for increasing the fertilization efficiency in grass puffer breeding programs. PMID:26973977

  20. Gamma scintigraphy using Tc-99m labeled antibody to human chorionic gonadotropin

    SciTech Connect

    Morrison, R.T.; Lyster, D.M.; Alcorn, L.N.; Rhodes, B.A.; Breslow, K.; Burchiel, S.W.

    1984-01-01

    A case report is presented describing a 27-year-old woman with invasive trophoblastic hydatidiform mole metastatic to the lung. Gamma scintiscanning, using a polyclonal and monoclonal antibody specific to human chorionic gonadotropin, hCG, and labeled with Tc-99m, is described. The area of the primary lesion in the uterus was demonstrated with both antibodies tested without computer subtraction techniques; metastatic deposits in the lung were detected only with the aid of blood pool subtraction techniques.

  1. An unexpected reason for elevated human chorionic gonadotropin in a young woman. Cervical squamous carcinoma.

    PubMed

    Mustafa, Aynur; Bozdag, Zehra; Tepe, Neslihan B; Ozcan, Husiyen C

    2016-08-01

    Human chorionic gonadotropin has been used for decades, in addition to specific investigations, to detect pregnancy, trophoblastic tumors, as well as congenital defects. Rarely, it can be elevated in  non-trophoblastic tumors such as squamous cell cancers and germ cell tumors. A 33-year-old Asian Syrian female had irregular menses accompanied with feelings of heaviness in the vagina. In addition to routine investigations, we measured the serum beta human chorionic gonadotropin (ß-HCG) level (based on the patient's complaint of amenorrhea), which was 50.05 ml UI/ml. Cervical biopsy revealed a non-keratinized large cell squamous carcinoma. After excluding other causes, ß-hCG elevation was explained by the ectopic secretion of cancer cells line. Cervical biopsy was suggestive of large cell non-keratinizing squamous cell carcinoma and positive for human chorionic gonadotropin on immunohistochemistry. As a result, we manage the possibility of ectopic secretion of ß-HCG from non- trophoblastic disease. PMID:27464870

  2. Early pregnancy factor is an immunosuppressive contaminant of commercial preparations of human chorionic gonadotrophin.

    PubMed Central

    Rolfe, B E; Morton, H; Clarke, F M

    1983-01-01

    Early pregnancy factor (EPF) is a pregnancy associated substance detected in human serum and urine throughout the first and second trimesters of pregnancy. It has also been detected in several commercial preparations of human chorionic gonadotrophin (hCG). The various molecular weight forms of EPF which occur in human pregnancy serum, urine and commercial hCG preparations have been partially characterized and found to be similar to each other but distinct from hCG. Further evidence is presented which suggests that it is EPF rather than hCG which is responsible for the immunosuppressive activity of some crude hCG preparations. PMID:6831771

  3. Atypical Presentations of Molar Pregnancy: Diagnostic Roles of Imaging, β-Human Chorionic Gonadotropin Measurement, and p57 Immunostaining.

    PubMed

    Mohamed, Sara A; Al-Hendy, Ayman; Ghamande, Sharad; Chaffin, Joanna; Browne, Paul

    2016-03-01

    In modern practice , the diagnosis of molar pregnancy is made at an early gestational age. The opportunity to diagnose gestational trophoblastic disease (GTD) using sonography alone occurs less frequently. The classic appearance of a "snowstorm" in the endometrial cavity and bilateral theca lutein cysts still applies to the diagnosis of second-trimester GTD. The diagnosis of first-trimester GTD requires increased clinical suspicion. If the sonographic appearance of the pregnancy is atypical, GTD should be included in the differential diagnosis. Additional nonimaging criteria such as serial quantitative β-human chorionic gonadotropin levels, pathologic examination, and p57 (cyclin-dependent kinase inhibitor 1C protein) immunostaining can accurately confirm the diagnosis of GTD. PMID:26860483

  4. Regulation of human renin expression in chorion cell primary cultures

    SciTech Connect

    Duncan, K.G.; Haidar, M.A.; Baxter, J.D.; Reudelhuber, T.L. )

    1990-10-01

    The human renin gene is expressed in the kidney, placenta, and several other sites. The release of renin or its precursor, prorenin, can be affected by several regulatory agents. In this study, primary cultures of human placental cells were used to examine the regulation of prorenin release and renin mRNA levels and of the transfected human renin promoter linked to chloramphenicol acetyltransferase reporter sequences. Treatment of the cultures with a calcium ionophore alone, calcium ionophore plus forskolin (that activates adenylate cyclase), or forskolin plus a phorbol ester increased prorenin release and renin mRNA levels 1.3{endash} to 6{endash}fold, but several classes of steroids did not affect prorenin secretion or renin RNA levels. These results suggest that (i) the first 584 base pairs of the renin gene 5'{endash}flanking DNA do not contain functional glucocorticoid or estrogen response elements, (ii) placental prorenin release and renin mRNA are regulated by calcium ion and by the combinations of cAMP with either C kinase or calcium ion, and (iii) the first 100 base pairs of the human renin 5'{endash}flanking DNA direct accurate initiation of transcription and can be regulated by cAMP. Thus, some control of renin release in the placenta (and by inference in other tissues) occurs via transcriptional influences on its promoter.

  5. [Gestational trophoblastic neoplasia after spontaneous normalization of human chorionic gonadotropin in patient with partial hydatidiform mole].

    PubMed

    Matos, Michelle; Ferraz, Leda; Lopes, Patrícia de Fátima; Lozoya, Consuelo; Amim Junior, Joffre; Rezende-Filho, Jorge; Braga, Antonio

    2015-07-01

    We report here a case of gestational trophoblastic neoplasia after spontaneous normalization of human chorionic gonadotropin in a patient with a partial hydatidiform mole. This is the second occurrence of this event to be reported and the first one with proven immunohistochemical evidence. Besides showing the treatment for this pregnancy complication, this case report discusses the possibility of reducing the duration of post-molar follow-up, as well as strategies for early recognition of gestational trophoblastic neoplasia after spontaneous remission of molar pregnancy. PMID:26247255

  6. A link between high serum levels of human chorionic gonadotrophin and chorionic expression of its mature functional receptor (LHCGR) in Down's syndrome pregnancies

    PubMed Central

    Banerjee, Subhasis; Smallwood, Alan; Chambers, Anne E; Papageorghiou, Aris; Loosfelt, Hugues; Spencer, Kevin; Campbell, Stuart; Nicolaides, Kypros

    2005-01-01

    Human chorionic gonadotrophin (hCG) is released from placental trophoblasts and is involved in establishing pregnancy by maintaining progesterone secretion from the corpus luteum. Serum hCG is detected in the maternal circulation within the first 2–3 wks of gestation and peaks at the end of the first trimester before declining. In Down's syndrome (DS) pregnancies, serum hCG remains significantly high compared to gestation age-matched uncompromised pregnancies. It has been proposed that increased serum hCG levels could be due to transcriptional hyper-activation of the CGB (hCG beta) gene, or an increased half life of glycosylated hCG hormone, or both. Another possibility is that serum hCG levels remain high due to reduced availability of the hormone's cognate receptor, LHCGR, leading to lack of hormone utilization. We have tested this hypothesis by quantifying the expression of the hCG beta (CGB) RNA, LHCGR RNA and LHCGR proteins in chorionic villous samples. We demonstrate that chorionic expression of hCG beta (CGB) mRNA directly correlates with high serum hCG levels. The steady-state synthesis of LHCGR mRNA (exons 1–5) in DS pregnancies was significantly higher than that of controls, but the expression of full-length LHCGR mRNA (exons 1–11) in DS was comparable to that of uncompromised pregnancies. However, the synthesis of high molecular weight mature LHCGR proteins was significantly reduced in DS compared to uncompromised pregnancies, suggesting a lack of utilization of circulating hCG in DS pregnancies. PMID:15969756

  7. Comparative analysis of neural differentiation potential in human mesenchymal stem cells derived from chorion and adult bone marrow.

    PubMed

    Ziadlou, Reihane; Shahhoseini, Maryam; Safari, Fatemeh; Sayahpour, Forugh-Azam; Nemati, Shiva; Eslaminejad, Mohamadreza Baghaban

    2015-11-01

    The finding of a reliable and abundant source of stem cells for the replacement of missing neurons in nervous system diseases requires extensive characterization of neural-differentiation-associated markers in stem cells from various sources. Chorion-derived stem cells from the human placenta have recently been described as an abundant, ethically acceptable, and easily accessible source of cells that are not limited in the same way as bone marrow (BM) mesenchymal stem cells (MSCs). We have isolated and cultured chorion MSCs (C-MSCs) and compared their proliferative capacity, multipotency, and neural differentiation ability with BM-MSCs. C-MSCs showed a higher proliferative capacity compared with BM-MSCs. The expression and histone modification of Nestin, as a marker for neural stem/progenitor cells, was evaluated quantitatively between the two groups. The Nestin expression level in C-MSCs was significantly higher than that in BM-MSCs. Notably, modifications of lys9, lys4, and lys27 of histone H3 agreed with the remarkable higher expression of Nestin in C-MSCs than in BM-MSCs. Furthermore, after neural differentiation of MSCs upon retinoic acid induction, both immunocytochemical and flow cytometry analyses demonstrated that the expression of neural marker genes was significantly higher in neural-induced C-MSCs compared with BM-MSCs. Mature neuron marker genes were also expressed at a significantly higher level in C-MSCs than in BM-MSCs. Thus, C-MSCs have a greater potential than BM-MSCs for differentiation to neural cell lineages and can be regarded as a promising source of stem cells for the cell therapy of neurological disorders. PMID:26022335

  8. Protein folding in the endoplasmic reticulum: lessons from the human chorionic gonadotropin beta subunit.

    PubMed Central

    Ruddon, R. W.; Sherman, S. A.; Bedows, E.

    1996-01-01

    There have been few studies of protein folding in the endoplasmic reticulum of intact mammalian cells. In the one case where the in vivo and in vitro folding pathways of a mammalian secretory protein have been compared, the folding of the human chorionic gonadotropin beta subunit (hCG-beta), the order of formation of the detected folding intermediates is the same. The rate and efficiency with which multidomain proteins such as hCG-beta fold to native structure in intact cells is higher than in vitro, although intracellular rates of folding of the beta subunit can be approached in vitro in the presence of an optimal redox potential and protein disulfide isomerase. Understanding how proteins fold in vivo may provide a new way to diagnose and treat human illnesses that occur due to folding defects. PMID:8844836

  9. Secondary treatment failure without anti-human chorionic gonadotropin antibody in a patient with Kallmann syndrome.

    PubMed

    Kitahara, S; Yoshida, K; Ishizaka, K; Higashi, Y; Takagi, K; Oshima, H

    1998-07-01

    A 29-year-old man with Kallmann syndrome suddenly developed decreased semen volume, azoospermia, and facial hair loss after 11 years of successful human chorionic gonadotropin (hCG) and human menopausal gonadotropin (hMG) treatment. Anti-hCG antibody was not detected in the patient's serum. A high serum level of luteinizing hormone (LH) with nasal LH-releasing hormone analogue administration failed to increase serum testosterone to a sufficient level. Testosterone injection after cessation of hCG and hMG therapy was able to improve semen volume, but not azoospermia. Resumption of hCG and hMG therapy after 6 months cessation partially restored spermatogenesis. The secondary failure of hCG and hMG therapy suggests a decrease of testicular sensitivity to LH as well as hCG. PMID:9712456

  10. Expression of 15-Hydroxyprostaglandin Dehydrogenase in Human Chorion Is Associated with Peroxisome Proliferator-Activated Receptor Isoform Expression in Term Labor.

    PubMed

    He, Ping; Li, Yuan; Ding, Xiaoying; Sun, Qianqian; Huang, Ying; Gu, Hang; Ni, Xin

    2015-07-01

    Chorionic NAD-dependent 15-hydroxy prostaglandin dehydrogenase (PGDH) plays a pivotal role in controlling the amount of prostaglandins in the uterus. Peroxisome proliferator-activated receptors (PPARs) are implicated to be involved in parturition. In this study, we investigated whether PPARs are involved in control of PGDH expression in chorion. The chorionic tissues were collected from the following groups of the women with singleton pregnancy: term no labor (TNL), term labor (TL) and preterm labor (PTL). Chorionic trophoblasts were isolated and cultured in vitro. Immunocytochemistry analysis showed that PPARα, PPARβ, and PPARγ were localized to trophoblasts in chorion. The protein levels of PGDH, PPARβ, and PPARγ were localized to trophoblasts in chorion. The protein levels of PPARα, PPARβ, and PPARγ were reduced in TL tissues compared to that of TNL group. PPARα, PPARβ, and PPARγ expression correlated to PGDH in TNL tissues, whereas only PPARγ expression correlated to PGDH in TL chorion tissues. PGDH expression was decreased in PTL tissues compared with TL group, whereas the expression of PPARs was not significantly different between TL and PTL groups. The agonists of three PPARs dose-dependently stimulated PGDH activity, mRNA, and protein expression in cultured chorionic cells. PPARs did not affect the stability of PGDH mRNA but stimulated the transcriptional activity of HPGD gene. Our results suggest that PPARs play pivotal roles in maintenance of PGDH expression in chorion during human pregnancy. PMID:26093984

  11. Studies on stabilities of some human chorionic gonadotropin complexes with β-emitting radionuclides.

    PubMed

    Maiti, Moumita; Sen, Kamalika; Sen, Souvik; Lahiri, Susanta

    2011-02-01

    Human chorionic gonadotropin (hCG) is a peptide hormone, whose one of the structural subunits is identical to that of thyroid-stimulating hormone (TSH). As a consequence, the receptors of TSH also act as receptor for hCG hormone. Keeping in mind this interesting property of hCG we have studied the complex formation ability of various no-carrier-added β-emitting isotopes of (61)Cu (3.3 h), (62)Zn (9.2 h), (90)Nb (14.60 h) and (99)Mo (66.02 h) with hCG molecule. Stability of the hCG-M (M=metal ions) complexes was investigated by dialysis with respect to triple distilled water and ringer lactate solution, which has the same composition as extracellular fluid. PMID:21144760

  12. Mussel adhesive enhances the immobilization of human chorionic gonadotrophin to a solid support.

    PubMed

    Burzio, V A; Silva, T; Pardo, J; Burzio, L O

    1996-10-15

    Polystyrene microtiter plates coated with 0.30 microgram/ well of the adhesive polyphenolic protein purified from the mussel Aulacomya ater showed enhanced capacity to immobilize antigens such as human chorionic gonadotrophin (hCG). Uncoated and coated wells were activated with different amounts of hCG (from 2 to 500 ng), blocked with bovine serum albumin, and tested with anti-hCG monoclonal antibodies and antimouse IgG conjugated with peroxidase. The reading at 492 nm of the uncoated wells activated with 500 ng of hCG was similar to that obtained with coated wells but using 5 to 10 ng of antigen. The coating procedure also resulted in better sensitivity to detect low concentration of monoclonal antibodies and better signal-to-noise ratio. The capacity of the mussel coating to immobilize hCG, as well as the immunoreactivity of the attached antigen, remained stable for several months. PMID:8921186

  13. Evidence for, and Associated Risks with, the Human Chorionic Gonadotropin Supplemented Diet.

    PubMed

    Butler, Stephen A; Cole, Laurence A

    2016-11-01

    Trend diets can be commonplace amongst those who are trying to lose weight but in most cases there is some shred of evidence to suggest they might be of some benefit. Seldom is there a diet which is such a fad that it is not only completely unfounded but also potential harmful. The human chorionic gonadotropin or "hCG diet" is such a diet, which after half a century still has no evidence to support its efficacy; in fact all scientific publications subsequent to the original article counter these claims. In this short communication, we review the literature and present data on exactly what some of the hCG diet preparations actually contain and highlight that, based on current data, these may do more harm than good. It is worrying that more consideration is not given to the possible danger of administration of hCG preparations to individuals without an evidence-based rational. PMID:27010890

  14. Mechanism of Trypanosoma cruzi Placenta Invasion and Infection: The Use of Human Chorionic Villi Explants

    PubMed Central

    Fretes, Ricardo E.; Kemmerling, Ulrike

    2012-01-01

    Congenital Chagas disease, a neglected tropical disease, endemic in Latin America, is associated with premature labor and miscarriage. During vertical transmission the parasite Trypanosoma cruzi (T. cruzi) crosses the placental barrier. However, the exact mechanism of the placental infection remains unclear. We review the congenital transmission of T. cruzi, particularly the role of possible local placental factors that contribute to the vertical transmission of the parasite. Additionally, we analyze the different methods available for studying the congenital transmission of the parasite. In that context, the ex vivo infection with T. cruzi trypomastigotes of human placental chorionic villi constitutes an excellent tool for studying parasite infection strategies as well as possible local antiparasitic mechanisms. PMID:22701129

  15. Simultaneous photoelectrochemical and visualized immunoassay of β-human chorionic gonadotrophin.

    PubMed

    Zhang, Nan; Ruan, Yi-Fan; Ma, Zheng-Yuan; Zhao, Wei-Wei; Xu, Jing-Juan; Chen, Hong-Yuan

    2016-11-15

    Herein, on the basis of the alkaline phosphate (ALP) induced reaction, a simultaneous photoelectrochemical (PEC) and visualized immunoassay has been established for the detection of β-human chorionic gonadotrophin (β-HCG). Specifically, in the proposed system, ALP stimulated the oxidative hydrolyzing transformation of 5-bromo-4-chloro-3-indoyl phosphate (BCIP) to an indigo precipitation, generating an insulating layer that impeded the interfacial mass and electron transfer and thus the photocurrent production. Meanwhile, a visualized detection could be performed according to the change of color intensity. Upon proper experimental conditions, the protocol possessed a detection range of 0.5-1000IU/L with a detection limit of (0.20±0.011)IU/L toward β-HCG. With high sensitivity and specificity, this work presents the first general protocol for simultaneous PEC and visualized detection, which could be easily extended to addressing numerous other targets. PMID:27183279

  16. In vitro effects of chorionic gonadotropin hormone on human adipose development.

    PubMed

    Dos Santos, Esther; Dieudonné, Marie-Noëlle; Leneveu, Marie-Christine; Pecquery, René; Serazin, Valérie; Giudicelli, Yves

    2007-08-01

    It is well known that pregnancy is associated with fat weight gain. However, the mechanisms whereby fat mass accumulation is controlled during this period are poorly understood. Therefore, we attempted to determine whether human chorionic gonadotropin (HCG), in vitro, influences human adipose tissue development and/or metabolism. For the first time, HCG/LH receptor was characterized in human adipose cells. We also demonstrated that physiological concentrations of HCG, while unaltering both lipolysis and expression of two markers of lipogenesis (FAS and ADD1) in human mature adipocytes, stimulate human preadipocyte growth via the activation of a protein kinase A-independent mitogen-activated protein kinase/c-fos signaling pathway. HCG also moderately increases the preadipocyte differentiation capacity as reflected by enhanced glycerophosphate dehydrogenase activity and expression of key adipogenic transcriptional factors (C/enhancer-binding protein alpha and peroxisome proliferator-activated receptor gamma 2). Finally, HCG significantly stimulates the secretion of the pro-adipogenic factor, leptin, from human adipose tissue. Taken altogether, these data suggest that the pro-adipogenic effect of HCG in human preadipocytes contributes to explain why increased fat storage occurs during the first trimester of pregnancy. PMID:17641281

  17. Oxygen-Sensitive K+ Channels Modulate Human Chorionic Gonadotropin Secretion from Human Placental Trophoblast

    PubMed Central

    Díaz, Paula; Sibley, Colin P.; Greenwood, Susan L.

    2016-01-01

    Human chorionic gonadotropin (hCG) is a key autocrine/paracrine regulator of placental syncytiotrophoblast, the transport epithelium of the human placenta. Syncytiotrophoblast hCG secretion is modulated by the partial pressure of oxygen (pO2), reactive oxygen species (ROS) and potassium (K+) channels. Here we test the hypothesis that K+ channels mediate the effects of pO2 and ROS on hCG secretion. Placental villous explants from normal term pregnancies were cultured for 6 days at 6% (normoxia), 21% (hyperoxia) or 1% (hypoxia) pO2. On days 3–5, explants were treated with 5mM 4-aminopyridine (4-AP) or tetraethylammonium (TEA), blockers of pO2-sensitive voltage-gated K+ (KV) channels, or ROS (10–1000μM H2O2). hCG secretion and lactate dehydrogenase (LDH) release, a marker of necrosis, were determined daily. At day 6, hCG and LDH were measured in tissue lysate and 86Rb (K+) efflux assessed to estimate syncytiotrophoblast K+ permeability. hCG secretion and 86Rb efflux were significantly greater in explants maintained in 21% pO2 than normoxia. 4-AP/TEA inhibited hCG secretion to a greater extent at 21% than 6% and 1% pO2, and reduced 86Rb efflux at 21% but not 6% pO2. LDH release and tissue LDH/hCG were similar in 6%, 21% and 1% pO2 and unaffected by 4-AP/TEA. H2O2 stimulated 86Rb efflux and hCG secretion at normoxia but decreased 86Rb efflux, without affecting hCG secretion, at 21% pO2. 4-AP/TEA-sensitive K+ channels participate in pO2-sensitive hCG secretion from syncytiotrophoblast. ROS effects on both hCG secretion and 86Rb efflux are pO2-dependent but causal links between the two remain to be established. PMID:26863525

  18. Oxygen-Sensitive K+ Channels Modulate Human Chorionic Gonadotropin Secretion from Human Placental Trophoblast.

    PubMed

    Díaz, Paula; Sibley, Colin P; Greenwood, Susan L

    2016-01-01

    Human chorionic gonadotropin (hCG) is a key autocrine/paracrine regulator of placental syncytiotrophoblast, the transport epithelium of the human placenta. Syncytiotrophoblast hCG secretion is modulated by the partial pressure of oxygen (pO2), reactive oxygen species (ROS) and potassium (K+) channels. Here we test the hypothesis that K+ channels mediate the effects of pO2 and ROS on hCG secretion. Placental villous explants from normal term pregnancies were cultured for 6 days at 6% (normoxia), 21% (hyperoxia) or 1% (hypoxia) pO2. On days 3-5, explants were treated with 5mM 4-aminopyridine (4-AP) or tetraethylammonium (TEA), blockers of pO2-sensitive voltage-gated K+ (KV) channels, or ROS (10-1000μM H2O2). hCG secretion and lactate dehydrogenase (LDH) release, a marker of necrosis, were determined daily. At day 6, hCG and LDH were measured in tissue lysate and 86Rb (K+) efflux assessed to estimate syncytiotrophoblast K+ permeability. hCG secretion and 86Rb efflux were significantly greater in explants maintained in 21% pO2 than normoxia. 4-AP/TEA inhibited hCG secretion to a greater extent at 21% than 6% and 1% pO2, and reduced 86Rb efflux at 21% but not 6% pO2. LDH release and tissue LDH/hCG were similar in 6%, 21% and 1% pO2 and unaffected by 4-AP/TEA. H2O2 stimulated 86Rb efflux and hCG secretion at normoxia but decreased 86Rb efflux, without affecting hCG secretion, at 21% pO2. 4-AP/TEA-sensitive K+ channels participate in pO2-sensitive hCG secretion from syncytiotrophoblast. ROS effects on both hCG secretion and 86Rb efflux are pO2-dependent but causal links between the two remain to be established. PMID:26863525

  19. Equine Chorionic Gonadotropin and Human Chorionic Gonadotropin Stimulation Increase the Number of Luteinized Follicles and the Progesterone Level Compared with Cabergoline Stimulation in Anoestrus Bitches.

    PubMed

    Jurczak, A; Domosławska, A; Bukowska, B; Janowski, T

    2016-08-01

    In this study, ovarian morphologies and blood progesterone concentrations following oestrous induction in bitches were examined. Fifty-three clinically healthy anoestrus bitches received cabergoline at a daily dose of 5 μg/kg of body weight per os for 21 days (group I) or subcutaneous equine chorionic gonadotropin at a dose of 20 IU/kg of body weight for five consecutive days with an additional 500 IU s.c. per bitch of human chorionic gonadotropin on the last day of treatment (group II). Twenty bitches that spontaneously displayed oestrous signs were left untreated and served as controls (group III). The induced oestrous rates and ovulation rates in groups I and II were 60.0% vs 64.3% and 86.7% vs 83.3%, respectively. Morphological assessments of the ovarian structures after ovariohysterectomy revealed an increase in the number of luteinized follicles and cysts in group II compared with the two other groups (p < 0.001). In contrast, the numbers of corpora lutea and follicles were similar in all groups. In accordance with the above-mentioned alteration, the progesterone concentration in the gonadotropin group (II) was increased (p < 0.001) in the periovulatory period compared with the other two groups. During the entire sampling period, the progesterone profiles in the cabergoline (I) and control (III) groups were similar and typical of normally cycling bitches. In conclusion, gonadotropin treatment is associated with an increased progesterone level during the periovulatory period that probably originates from luteinized follicles, whereas cabergoline treatment induces cycles with both physiological progesterone concentrations and ovarian morphologies. PMID:27321641

  20. Human placenta and chorion: potential additional sources of hematopoietic stem cells for transplantation

    PubMed Central

    Bárcena, Alicia; Muench, Marcus O.; Kapidzic, Mirhan; Gormley, Matthew; Goldfien, Gabriel A.; Fisher, Susan J.

    2012-01-01

    Background Hematopoietic stem cell (HSC) transplantation is an essential element of medical therapy, leading to cures of previously incurable disease for hematological and non-hematological pathologies. Many patients do not find matched donors in a timely manner, which has driven efforts to find alternative pools of transplantable HSCs. The use of umbilical cord blood (UCB) as a source of transplantable HSCs began more than two decades ago. However, the use of UCB as a reliable source of HSCs for transplantation still faces crucial challenges: the number of HSCs present in a unit of UCB is usually sufficient for younger children but not for adults and the persistent delayed engraftment often seen can result in high rates of infection and mortality. Study Design and Methods We propose a new approach to a solution of these problems: a potential increase of the limited number of UCB–HSCs available by harvesting HSCs contained in the placenta and the fetal chorionic membrane available at birth. Results We investigated the presence of hematopoietic progenitors/HSC in human placenta and chorion at different gestational ages. The characterization of these cells was performed by flow cytometry and immunolocalization and their functional status was investigated by transplanting them into immunodeficient mice. Conclusion HSCs are present in extraembryonic tissues and could be banked in conjunction to the UCB-HSCs. This novel approach could have a large impact on the field of HSC banking and more crucially, on the outcome of patients undergoing this treatment by greatly improving the use of life-saving hematopoietic transplants. PMID:22074633

  1. A review of luteinising hormone and human chorionic gonadotropin when used in assisted reproductive technology.

    PubMed

    Ezcurra, Diego; Humaidan, Peter

    2014-01-01

    Gonadotropins extracted from the urine of post-menopausal women have traditionally been used to stimulate folliculogenesis in the treatment of infertility and in assisted reproductive technology (ART). Products, such as human menopausal gonadotropin (hMG), consist not only of a mixture of the hormones, follicle-stimulating hormone (FSH), luteinising hormone (LH) and human chorionic gonadotropin (hCG), but also other biologically active contaminants, such as growth factors, binding proteins and prion proteins. The actual amount of molecular LH in hMG preparations varies considerably due to the purification process, thus hCG, mimicking LH action, is added to standardise the product. However, unlike LH, hCG plays a different role during the natural human menstrual cycle. It is secreted by the embryo and placenta, and its main role is to support implantation and pregnancy. More recently, recombinant gonadotropins (r-hFSH and r-hLH) have become available for ART therapies. Recombinant LH contains only LH molecules. In the field of reproduction there has been controversy in recent years over whether r-hLH or hCG should be used for ART. This review examines the existing evidence for molecular and functional differences between LH and hCG and assesses the clinical implications of hCG-supplemented urinary therapy compared with recombinant therapies used for ART. PMID:25280580

  2. Evaluation of nicked human chorionic gonadotropin content in clinical specimens by a specific immunometric assay.

    PubMed

    Kovalevskaya, G; Birken, S; Kakuma, T; Schlatterer, J; O'Connor, J F

    1999-01-01

    We report the development and characterization of an IRMA for the direct measurement of nicked human chorionic gonadotropin (hCGn) in blood and urine. hCGn derived from a reference preparation of hCG used as an immunogen elicits monoclonal antibodies (mAbs) with enhanced recognition of human luteinizing hormone epitopes. The most specific assay for pregnancy hCGn is an IRMA composed of one mAb to choriocarcinoma-derived hCGn (C5) and a second mAb developed from immunization with normal-pregnancy hCGn. This assay was used to evaluate hCGn profiles in normal, in vitro fertilization, Down syndrome, and ectopic pregnancies. In all pregnancies, hCGn was usually present in much lower concentrations than the non-nicked hCG isoform. Our results suggest that some form of physical separation from the overwhelming quantities of non-nicked hCG present in clinical specimens will be required before accurate immunochemical estimations of hCGn can be made. PMID:9895340

  3. Why are We Waiting to Start Large Scale Clinical Testing of Human Chorionic Gonadotropin for the Treatment of Preterm Births?

    PubMed

    Rao, C V

    2016-07-01

    Preterm births are an expensive global health problem. Despite the basic science and clinical research advances to better understand and prevent preterm births, the rates are increasing. There are several therapeutic options. While some options such as progestins work for selected women, others such as magnesium sulfate can only be used for delaying births for 24 to 48 hours so that the patients can be treated with corticosteroids to promote fetal lung maturity. Based on the scientific and clinical evidence, we recommend testing human chorionic gonadotropin in a large multicenter, randomized, double-blind, and placebo-controlled clinical trials in women with active preterm labor and those with a previous history of preterm births. Human chorionic gonadotropin is not only inexpensive but also has not shown any side effects so far in the infants or in the mothers. PMID:26692543

  4. Human chorionic somatomammotropin in normal adolescent primiparous pregnancy. I. Effect of smoking.

    PubMed

    Moser, R J; Hollingsworth, D R; Carlson, J W; Lamotte, L

    1974-12-15

    Human chorionic somatomammotropin (HCS) levels were studied in normal smoking and nonsmoking primiparous adolescent pregnancies. 136 teenagers, aged 12-18 years, were divided into groups: nonsmokers, deep, and shallow inhalers, long, and short puffers, high, and low tar, and high, and low nicotin. Shallow inhaling and low nicotine exposure patients were found to have a later age of menarche than did nonsmokers (13.2 vs. 12.3 years, p=.03). The mean body weight of the mothers who smoked was slightly less (61 gm) than that of nonsmoking mothers. Except for long puffers, overall, smokers had significantly lower HCS values throughout pregnancy than noosmokers (p = .48 high tar-p = .002 low tar). However, in the third trimester those with the lowest smoking exposures had the lowest HCS values and the heavier smokers had slightly higher mean values than nonsmokers. These data suggest that HCS production may be more sensitive to low tar and nicotine exposure with possible tolerance or even stimulation occurring in larger doses. PMID:4432896

  5. Uterine choriocarcinoma accompanied by an extremely high human chorionic gonadotropin level and thyrotoxicosis.

    PubMed

    Hsieh, Tsung-Ying; Hsu, Keng-Fu; Kuo, Pao-Lin; Huang, Soon-Cen

    2008-04-01

    The conventional treatments given to a 24-year-old woman with metastatic uterine choriocarcinoma and clinical and biochemical thyrotoxicosis did not appear to have any effect, probably due to an extremely high serum human chorionic gonadotropin (hCG) level which was up to 11,910,000 mIU/mL, and were initially underscored in light of the 'high-dose hook effect'. To our knowledge, no extremely high hCG level in a uterine choriocarcinoma patient has been reported in the literature. Her decapacitating symptoms subsided after the first course of chemotherapy by etoposide, methotrexate, and actinomycin D-cyclophosphamide and vincristine (EMA-CO) regimen. The serum hCG level, which reflects the quantification of host tumor burden, returned to the reference range after the fifth course of chemotherapy and the thyroid function reached euthyroid status before the third course of chemotherapy; two final courses were administered after the hCG level became undetectable. Two years after remission of disease, the patient experienced a normal pregnancy, and a term baby girl was delivered vaginally. No recurrence of uterine choriocarcinoma has been noted for 7 years. PMID:18412797

  6. Antibody-free detection of human chorionic gonadotropin by use of liquid crystals.

    PubMed

    Ding, Xiaokang; Yang, Kun-Lin

    2013-11-19

    Human chorionic gonadotropin (hCG) is an important biomarker for the diagnosis of pregnancy and cancers. In this study, we report an antibody-free and label-free mechanism for detecting hCG. To replace enzyme-labeled antibodies, we use a short oligopeptide as an hCG receptor to bind hCG. The short oligopeptide sequence, (N-)PPLRINRHILTR(-C), is identified after 5 rounds of screening by use of a phage library. After binding, liquid crystal (LC) is used to transduce the binding event into optical signals. The captured hCG can disrupt a thin layer (~6 μm) of LC covered on the surface. Depending on the initial concentration of hCG, LC gives distinct optical signals visible to the naked eye. The limit of detection (LOD) for this method is approximately 1 IU/mL (2 nM) in both phosphate-buffered saline and urine samples, and only 0.6 μL of hCG solution is required. This means that as little as 45.5 pg of hCG can be detected by this method. Compared to other detection methods for hCG, this detection method does not require the use of antibody and is label-free. It has the potential to become a portable diagnostic kit for hCG. PMID:24147645

  7. Crystallization and characterization of human chorionic gonadotropin in chemically deglycosylated and enzymatically desialylated states

    SciTech Connect

    Lustbader, J.W.; Birken, S.; Pileggi, N.F.; Folks, M.A.G.; Pollak, S.; Cuff, M.E.; Yang, Wei; Hendrickson, W.A.; Canfield, R.E. )

    1989-11-28

    Crystals suitable for X-ray diffraction studies at moderate resolution have been grown from two forms of human chorionic gonadotropin (hCG): HF-treated hCG and neuraminidase-treated hCG. The enzymatically desialylated form of hCG produced crystals that diffract to 2.8 {angstrom} as compared to the HF-treated hCG crystals that diffract to 3.0 {angstrom}. Although it was assumed that the high and heterogeneous carbohydrate content of the glycoprotein hormones inhibited their crystallization, this report suggests that it is the negatively charged surface sugars and neither the total carbohydrate content nor its heterogeneity which interferes with crystal formation. Chemical deglycosylation resulted in significantly increased protein degradation during crystal growth. Such peptide bond cleavages were observed to a much lesser extent in the crystals grown from neuraminidase-digested hCG. Sequence analysis of the HF-treated hCG crystals suggested that up to 45% of the molecules within the crystal had an acid-labile peptide bond cleaved. In contrast, the neuraminidase-treated hCG exhibited less than 9% of this type of cleavage. The manner in which hCG was treated prior to crystallization was found to be a very important factor in the extent of peptide bound cleavages occurring during crystal growth. HF treatment of glycoproteins may render glycoproteins more susceptible to peptide bond cleavage during crystal growth.

  8. Effect of bromodichloromethane on chorionic gonadotrophin secretion by human placental trophoblast cultures.

    PubMed

    Chen, Jiangang; Douglas, Gordon C; Thirkill, Twanda L; Lohstroh, Peter N; Bielmeier, Susan R; Narotsky, Michael G; Best, Deborah S; Harrison, Randy A; Natarajan, Kala; Pegram, Rex A; Overstreet, James W; Lasley, Bill L

    2003-11-01

    Bromodichloromethane (BDCM) is a trihalomethane found in drinking water as a by-product of disinfection processes. BDCM is hepatotoxic and nephrotoxic in rodents and has been reported to cause strain-specific full-litter resorption in F344 rats during the luteinizing hormone-dependent phase of pregnancy. In humans, epidemiological studies suggest an association between exposure to BDCM in drinking water and increased risk of spontaneous abortion. To begin to address the mechanism(s) of BDCM-induced spontaneous abortion, we hypothesized that BDCM targets the placenta. Primary cultures of human term trophoblast cells were used as an in vitro model to test this hypothesis. Trophoblasts were allowed to differentiate into multinucleated syncytiotrophoblast-like colonies, after which they were incubated for 24 h with different concentrations of BDCM (20 nM to 2 mM). Culture media were collected and assayed for immunoreactive and bioactive chorionic gonadotropin (CG). Cultures exposed to BDCM showed a dose-dependent decrease in the secretion of immunoreactive CG as well as bioactive CG. The lowest effective BDCM concentration was 20 nM, approximately 35-times higher than the maximum concentration reported in human blood (0.57 nM). Trophoblast morphology and viability were similar in controls and cultures exposed to BDCM. We conclude that BDCM perturbs CG secretion by differentiated trophoblasts in vitro. This suggests that the placenta is a likely target of BDCM toxicity in the human and that this could be related to the adverse pregnancy outcomes associated with BDCM. PMID:12970577

  9. Luteinizing hormone/human chorionic gonadotrophin receptors in various epidermal structures.

    PubMed

    Venencie, P Y; Méduri, G; Pissard, S; Jolivet, A; Loosfelt, H; Milgrom, E; Misrahi, M

    1999-09-01

    Two different monoclonal antibodies recognizing different epitopes were used to study the localization of luteinizing hormone/human chorionic gonadotrophin (LH/hCG) receptors in human skin. Immunolabelling was observed only in the epidermis and derived structures but not in the dermis. The basal, spinal and granular layers were stained, whereas no receptors were detected in the non-nucleated horny cells. In the growing (anagen) hair, immunostaining was found in the inner root sheath below the level of the sebaceous glands and in the outer root sheath above this level. In the resting (telogen) hair, only the latter staining was observed. In the sebaceous glands, only the thin cells close to the walls of the ducts were immunolabelled. In the eccrine sweat glands, the external clear cells were stained in the secretory portion of the gland, whereas only the cells close to the lumen were labelled in the ducts. The distribution of LH/hCG receptors was compared with that of steroidogenic enzymes (side chain cleavage cytochrome P450, adrenodoxin, 3-beta-hydroxy-5-ene steroid dehydrogenase Delta5-Delta4 isomerase, 17-hydroxylase cytochrome P450 and cytochrome P450 aromatase). Only partial overlaps were observed. The presence of LH receptor mRNA in the skin was confirmed by reverse transcription-polymerase chain reaction. Monoclonal antibodies raised against the human follicle-stimulating hormone receptor failed to detect the latter in the epidermal structures and in the dermis. The role of LH and hCG in skin modifications occurring during pregnancy and after the menopause is unknown. These hormones may possibly act by regulating steroidogenic enzymes or by modulating cell growth and differentiation. PMID:10583046

  10. Human Chorionic Gonadotropin Partially Mediates Phthalate Association With Male and Female Anogenital Distance

    PubMed Central

    Lee, Myoung Keun; Naimi, Ashley I.; Barrett, Emily; Nguyen, Ruby H.; Sathyanarayana, Sheela; Zhao, Yaqi; Thiet, Mari-Paule; Redmon, J. Bruce; Swan, Shanna H.

    2015-01-01

    Context: Prenatal exposure to phthalates disrupts male sex development in rodents. In humans, the placental glycoprotein hormone human chorionic gonadotropin (hCG) is required for male development, and may be a target of phthalate exposure. Objective: This study aimed to test the hypothesis that phthalates disrupt placental hCG differentially in males and females with consequences for sexually dimorphic genital development. Design: The Infant Development and Environment Study (TIDES) is a prospective birth cohort. Pregnant women were enrolled from 2010–2012 at four university hospitals. Participants: Participants were TIDES subjects (n = 541) for whom genital and phthalate measurements were available and who underwent prenatal serum screening in the first or second trimester. Main Outcome Measures: Outcomes included hCG levels in maternal serum in the first and second trimesters and anogenital distance (AGD), which is the distance from the anus to the genitals in male and female neonates. Results: Higher first-trimester urinary mono-n-butyl phthalate (MnBP; P = .01), monobenzyl phthalate (MBzP; P = .03), and mono-carboxy-isooctyl phthalate (P < .01) were associated with higher first-trimester hCG in women carrying female fetuses, and lower hCG in women carrying males. First-trimester hCG was positively correlated with the AGD z score in female neonates, and inversely correlated in males (P = 0.01). We measured significant associations of MnBP (P < .01), MBzP (P = .02), and mono-2-ethylhexyl phthalate (MEHP; P < .01) with AGD, after adjusting for sex differences. Approximately 52% (MnBP) and 25% (MEHP) of this association in males, and 78% in females (MBzP), could be attributed to the phthalate association with hCG. Conclusions: First-trimester hCG levels, normalized by fetal sex, may reflect sexually dimorphic action of phthalates on placental function and on genital development. PMID:26200238

  11. Regenerative and reparative effects of human chorion-derived stem cell conditioned medium on photo-aged epidermal cells.

    PubMed

    Li, Qiankun; Chen, Yan; Ma, Kui; Zhao, Along; Zhang, Cuiping; Fu, Xiaobing

    2016-04-17

    Epidermal cells are an important regenerative source for skin wound healing. Aged epidermal cells have a low ability to renew themselves and repair skin injury. Ultraviolet (UV) radiation, particularly UVB, can cause photo-aging of the skin by suppressing the viability of human epidermal cells. A chorion-derived stem cell conditioned medium (CDSC-CNM) is thought to have regenerative properties. This study aimed to determine the regenerative effects of CDSC-CNM on UVB-induced photo-aged epidermal cells. Epidermal cells were passaged four times and irradiated with quantitative UVB, and non-irradiated cells served as a control group. Cells were then treated with different concentrations of CDSC-CNM. Compared to the non-irradiated group, the proliferation rates and migration rates of UVB-induced photo-aged epidermal cells significantly decreased (p < 0.05) with increasing intracellular radical oxygen species (ROS) generation and DNA damage. After treatment with CDSC-CNM, photo-aged epidermal cells significantly improved their viability, and their ROS generation and DNA damage decreased. The secretory factors in CDSC-CNM, including epidermal growth factor (EGF), transforming growth factor-β (TGF-β), interleukin (IL)-6, and IL-8 and the related signaling pathway protein levels, increased compared to the control medium (CM). The potential regenerative and reparative effects of CDSC-CNM indicate that it may be a candidate material for the treatment of prematurely aged skin. The functions of the secretory factors and the mechanisms of CDSC-CNM therapy deserve further attention. PMID:27097375

  12. Recognition of N-glycoforms in human chorionic gonadotropin by monoclonal antibodies and their interaction motifs.

    PubMed

    Li, Daoyuan; Zhang, Ping; Li, Fei; Chi, Lequan; Zhu, Deyu; Zhang, Qunye; Chi, Lianli

    2015-09-11

    The glycosylation of human chorionic gonadotropin (hCG) plays an important role in reproductive tumors. Detecting hCG N-glycosylation alteration may significantly improve the diagnostic accuracy and sensitivity of related cancers. However, developing an immunoassay directly against the N-linked oligosaccharides is unlikely because of the heterogeneity and low immunogenicity of carbohydrates. Here, we report a hydrogen/deuterium exchange and MS approach to investigate the effect of N-glycosylation on the binding of antibodies against different hCG glycoforms. Hyperglycosylated hCG was purified from the urine of invasive mole patients, and the structure of its N-linked oligosaccharides was confirmed to be more branched by MS. The binding kinetics of the anti-hCG antibodies MCA329 and MCA1024 against hCG and hyperglycosylated hCG were compared using biolayer interferometry. The binding affinity of MCA1024 changed significantly in response to the alteration of hCG N-linked oligosaccharides. Hydrogen/deuterium exchange-MS reveals that the peptide β65-83 of the hCG β subunit is the epitope for MCA1024. Site-specific N-glycosylation analysis suggests that N-linked oligosaccharides at Asn-13 and Asn-30 on the β subunit affect the binding affinity of MCA1024. These results prove that some antibodies are sensitive to the structural change of N-linked oligosaccharides, whereas others are not affected by N-glycosylation. It is promising to improve glycoprotein biomarker-based cancer diagnostics by developing combined immunoassays that can determine the level of protein and measure the degree of N-glycosylation simultaneously. PMID:26240146

  13. Human chorionic gonadotropin increases serum progesterone, number of corpora lutea and angiogenic factors in pregnant sheep.

    PubMed

    Coleson, Megan P T; Sanchez, Nicole S; Ashley, Amanda K; Ross, Timothy T; Ashley, Ryan L

    2015-07-01

    Early gestation is a critical period when implantation and placental vascularization are established, processes influenced by progesterone (P4). Although human chorionic gonadotropin (hCG) is not endogenously synthesized by livestock, it binds the LH receptor, stimulating P4 synthesis. We hypothesized treating pregnant ewes with hCG would increase serum P4, number of corpora lutea (CLs) and concepti, augment steroidogenic enzymes, and increase membrane P4 receptors (PAQRs) and angiogenic factors in reproductive tissues. The objective was to determine molecular alterations induced by hCG in pregnant sheep that may promote pregnancy. Ewes received either 600 IU of hCG or saline i.m. on day 4 post mating. Blood samples were collected daily from day 0 until tissue collection for serum P4 analysis. Reproductive tissues were collected on either day 13 or 25 of gestation and analyzed for PAQRs, CXCR4, proangiogenic factors and steroidogenic enzymes. Ewes receiving hCG had more CL and greater serum P4, which remained elevated. On day 25, StAR protein production decreased in CL from hCG-treated ewes while HSD3B1 was unchanged; further, expression of CXCR4 significantly increased and KDR tended to increase. PAQR7 and CXCR4 protein was increased in caruncle tissue from hCG-treated ewes. Maternal hCG exposure influenced fetal extraembryonic tissues, as VEGFA, VEGFB, FLT1, and ANGPT1 expression increased. Our results indicate hCG increases serum P4 due to augmented CL number per ewe. hCG treatment resulted in greater PAQR7 and CXCR4 in maternal endometrium and promoted expression of proangiogenic factors in fetal extraembryonic membranes. Supplementing livestock with hCG may boost P4 levels and improve reproductive efficiency. PMID:25861798

  14. CNS germinoma with elevated serum human chorionic gonadotropin level: Clinical characteristics and treatment outcome

    SciTech Connect

    Ogino, Hiroyuki . E-mail: ogino@med.nagoya-cu.ac.jp; Shibamoto, Yuta; Takanaka, Tsuyoshi; Suzuki, Kazunori; Ishihara, Shun-Ichi; Yamada, Tetsuya; Sugie, Chikao; Nomoto, Yoshihito; Mimura, Mikio

    2005-07-01

    Purpose: The prognostic significance of human chorionic gonadotropin (HCG) level in central nervous system germinoma remains controversial. The purpose of this study was to compare clinical characteristics and prognosis of germinoma patients with normal and high HCG titers in the serum. Methods and Materials: We undertook a multi-institutional retrospective analysis of 103 patients with central nervous system germinoma whose serum HCG and/or {beta}-HCG level had been measured before treatment between 1984 and 2002. All patients had been treated with radiation therapy either alone (n = 66) or in combination with chemotherapy (n = 37) with a median dose of 47.8 Gy. Results: HCG and/or {beta}-HCG level in the serum was high in 39% of all patients. The proportion of HCG-producing tumors was higher in the lesions at the basal ganglia than in the lesions at the other sites. No correlation was found between tumor size and HCG level, but there seemed to be a weak correlation between size and {beta}-HCG. The 5- and 10-year survival rates were 96% and 94%, respectively, in both patient groups with normal and high HCG (p = 0.99). The 5- and 10-year relapse-free survival rates were 87% and 82%, respectively, in patients with normal HCG level and were both 87% in patients with high HCG (p = 0.74). Also, no other patient-, tumor-, or treatment-related factors seemed to influence the prognosis of the patients. Conclusion: Serum HCG level does not seem to influence patient prognosis when treated with sufficient doses of radiation. Relationship between tumor size and site and HCG level should be investigated further.

  15. Choriocarcinoma-like human chorionic gonadotrophin (HCG) and HCG bioactivity during the first trimester of pregnancy.

    PubMed

    Mock, P; Kovalevskaya, G; O'Connor, J F; Campana, A

    2000-10-01

    The objective of this study was to evaluate the distribution of choriocarcinoma-like human chorionic gonadotrophin (HCG) isoforms during first trimester pregnancy and their relationship with in-vitro HCG bioactivity. This was done by means of a retrospective analysis of patients' sera with first trimester normal intrauterine and abnormal (ectopic) pregnancies. Serum samples were obtained from 38 women with an amenorrhoea of <10 weeks. From these, 19 had a normal intrauterine pregnancy (IUP) and 19 an ectopic pregnancy (EP). Total immunoreactive HCG (HCGi), free beta-HCGi and oestradiol were measured by enzyme immunoassays and bioactive HCG by the mouse Leydig cell bioassay. The alterations in HCG isoform content were measured by the combination of two immunometric assays, B152 for choriocarcinoma-like HCG and B109 for intact HCG detection and expressed as the B152/B109 ratio. Choriocarcinoma-like HCG isoforms ratio measured by B152 and B109 assays was significantly higher in the low subgroups of free beta-HCGi and gestational age (P = 0.0111 and 0.0036 respectively). Whereas bioactive to immunoreactive HCG ratios (b/i ratio) were significantly higher when free beta-HCGi concentrations were low (P = 0.0010), no correlation was found between the variation of bioactivity (b/i ratio) and the proportion of choriocarcinoma-like HCG isoforms (B159/B108). It is concluded that in first trimester pregnancies (i) the modulation of HCG in-vitro bioactivity is not related to the variation of choriocarcinoma-like HCG isoforms secretion and (ii) the amount of choriocarcinoma-like HCG isoforms secreted by the early trophoblast is predominant and may be the result of an early developmental regulation of glycosylation enzyme. PMID:11006201

  16. Identification of inadequate maternal progesterone concentrations in nulliparous dairy heifers and treatment with human chorionic gonadotrophin.

    PubMed

    King, S G; Dobson, H; Royal, M D; Christley, R M; Murray, R D; Routly, J E; Smith, R F; Mann, G E

    2013-11-01

    Plasma progesterone concentrations were estimated for 381 nulliparous dairy heifers from eight farms on days 4-6 after insemination. Heifers were synchronised using two treatments of prostaglandin F2α (PGF2α) 11 days apart. Alternate heifers were treated with 1500 IU human chorionic gonadotrophin (hCG) at the time of sampling, and the pregnancy status assessed ~35 days after insemination. Heifers with no corpus luteum (CL; n-30) at the second PGF2α had lower progesterone concentrations (P<0.001) and pregnancy rates (PR; P=0.001) compared with heifers with CL (n=351, defined as cyclic). The risk of no CL increased in heifers <15 months of age (OR=7.5, P<0.001) and with body condition score <2.5 (OR=4.5, P=0.001). Those with no CL were removed from further analysis. The 20 per cent cyclic control heifers with progesterone concentrations ≤2 ng/ml on days 4-6 had lower PRs compared with those with higher concentrations (42 per cent vs 64 per cent, OR=0.3, P=0.01). There was no overall effect of hCG on PR compared with controls (51 per cent vs 60 per cent, P=0.10). However, heifers with high progesterone on days 4-6 (>2 ng/ml) had a lower PR after treatment (51 per cent vs 64 per cent, OR=0.6, P=0.02). Heifers inseminated with sexed (n=18) compared with conventional (n=333) semen had a decreased PR (33 per cent vs 57 per cent, OR=0.2, P=0.01). This study demonstrates that higher plasma progesterone concentrations as early as days 4-6 after insemination have a positive effect on first insemination PR, but there was no beneficial effect of hCG treatment on these days in dairy heifers. PMID:24128945

  17. Human chorionic gonadotropin: Different glycoforms and biological activity depending on its source of production.

    PubMed

    Fournier, Thierry

    2016-06-01

    Human chorionic gonadotropin (hCG) is the first hormonal message from the placenta to the mother. It is detectable in maternal blood two days after implantation and behaves like a super LH agonist stimulating progesterone secretion by the corpus luteum. In addition to maintaining the production of progesterone until the placenta itself produces it, hCG also has a role in myometrial quiescence and local immune tolerance. Specific to humans, hCG is a complex glycoprotein composed of two highly glycosylated subunits. The α-subunit is identical to the pituitary gonadotropin hormones (LH, FSH, TSH), contains two N-glycosylation sites, and is encoded by a single gene (CGA). By contrast, the β-subunits are distinct for each hormones and confer both receptor and biological specificity, although LH and hCG bind to the same receptor (LH/CG-R). The hCG ß-subunit is encoded by a cluster of genes (CGB) and contains two sites of N-glycosylation and four sites of O-glycosylation. The hCG glycosylation state varies with the stage of pregnancy, its source of production and in the pathology. It is well established that hCG is mainly secreted into maternal blood, where it peaks at 8-10weeks of gestation (WG), by the syncytiotrophoblast (ST), which represents the endocrine tissue of the human placenta. The invasive extravillous trophoblast (iEVT) also secretes hCG, and in particular hyperglycosylated forms of hCG (hCG-H) also produced by choriocarcinoma cells. In maternal blood, hCG-H is elevated during early first trimester corresponding to the trophoblastic cell invasion process and then decreases. In addition to its endocrine role, hCG has autocrine and paracrine roles. It promotes formation of the ST and angiogenesis through LH/CG-R but has no effect on trophoblast invasion in vitro. By contrast, hCG-H stimulates trophoblast invasion and angiogenesis by interacting with the TGFß receptor in a LH/CG-R independent signalling pathway. hCG is largely used in antenatal screening

  18. New discoveries on the biology and detection of human chorionic gonadotropin

    PubMed Central

    Cole, Laurence A

    2009-01-01

    Human chorionic gonadotropin (hCG) is a glycoprotein hormone comprising 2 subunits, alpha and beta joined non covalently. While similar in structure to luteinizing hormone (LH), hCG exists in multiple hormonal and non-endocrine agents, rather than as a single molecule like LH and the other glycoprotein hormones. These are regular hCG, hyperglycosylated hCG and the free beta-subunit of hyperglycosylated hCG. For 88 years regular hCG has been known as a promoter of corpus luteal progesterone production, even though this function only explains 3 weeks of a full gestations production of regular hCG. Research in recent years has explained the full gestational production by demonstration of critical functions in trophoblast differentiation and in fetal nutrition through myometrial spiral artery angiogenesis. While regular hCG is made by fused villous syncytiotrophoblast cells, extravillous invasive cytotrophoblast cells make the variant hyperglycosylated hCG. This variant is an autocrine factor, acting on extravillous invasive cytotrophoblast cells to initiate and control invasion as occurs at implantation of pregnancy and the establishment of hemochorial placentation, and malignancy as occurs in invasive hydatidiform mole and choriocarcinoma. Hyperglycosylated hCG inhibits apoptosis in extravillous invasive cytotrophoblast cells promoting cell invasion, growth and malignancy. Other non-trophoblastic malignancies retro-differentiate and produce a hyperglycosylated free beta-subunit of hCG (hCG free beta). This has been shown to be an autocrine factor antagonizing apoptosis furthering cancer cell growth and malignancy. New applications have been demonstrated for total hCG measurements and detection of the 3 hCG variants in pregnancy detection, monitoring pregnancy outcome, determining risk for Down syndrome fetus, predicting preeclampsia, detecting pituitary hCG, detecting and managing gestational trophoblastic diseases, diagnosing quiescent gestational trophoblastic

  19. Separation of beta-human chorionic gonadotropin and immunoglobulin G by a miniaturized size exclusion chromatography column

    NASA Astrophysics Data System (ADS)

    Yang, Yongmo; Chae, Junseok

    2009-04-01

    This report describes a miniaturized size exclusion chromatography column that effectively preseparates raw samples for medical point-of-care testing (POCT) devices. The minicolumn is constructed of polydimethylsiloxane fabricated on a glass slide. The minicolumn separates 300 ng/ml of beta-human chorionic gonadotropin (β-hCG) from an immunoglobulin G (IgG)-rich solution (100 μg/ml) in 7.7 min, with 2.23 resolution and 0.018 mm plate height. The complete analyte discrimination shows potential for the sample preparation stage of POCT devices for cancer screening, prognosis, and monitoring.

  20. Value of human chorionic gonadotropin compared to CEA in discriminating benign from malignant effusions.

    PubMed

    Lamerz, R; Stoetzer, O J; Mezger, J; Brandt, A; Darsow, M; Wilmanns, W

    1999-01-01

    Human chorionic gonadotropin (HCG) is expressed in germ cell tumors and urothelial, breast, lung and colon cancers. The aim of the study was to investigate if the determination of HCG in comparison with CEA is able to discriminate between malignant and benign effusions. Effusion and partially serum samples of 61 patients with benign (g.i., heart/kidney isnuff.) and 116 patients with malignant diseases (g.i., gynec., lung, misc., CUP) were investigated. HCG was specifically determined by an IRMA using 2 monoclonal antibodies, CEA by a conventional double Ab RIA. Cytological staining was preformed using the Pappenheim-method on cytospin preparations. Significant differences (p < 0.001) were found for HCG between benign and malignant ascitic effusions with the best discrimination at 5 IU/l (ROC) and an overall sensitivity of 31.3% (spec. vs benign eff. 93.4%) increasing in subgroups from hematol. (5.8%) < misc. (31.3%) < gynec. (32.1%) < g.i. (36%) < lung (38.1%) to CUP (50%). CEA also showed significant differences between benign and malignant total and ascitic effusions, and weaker for the pleural subgroup (cutoff 9 ng/ml) with a total sensitivity of 44.6% (sp = 100%) increasing from misc. (30.8%) < lung (47.1%) < CUP (50%) < gynec. (60%) < g.i. (60.9%). Comparative cytology and TM determinations increased the positiverate of cytology (45.2%) to 58.3% for either cytology or HCG positive cases, or to 61.6% for either cytology or CEA positive cases. For the combined determination of cytologoy and HCG and CEA, the overall TM positive rate for 33 cytology-pos. cases was 78.8%, but in 40 cytology-negative cases 37.5% for TM positive cases. In conclusion HCG is useful in ascitic > pleural effusions with high specificity (90% at 5 IU/l) but low sensitivity of 31% increasing in g.i., lung and gynecologic cases, CEA a more general TM with higher sensitivity of 45% increasing in g.i., gynecologic and lung cases (sp. 100% at 9 ng/ml) both adding significantly to cytology

  1. Elevated serum progesterone/ MII oocyte ratio on the day of human chorionic gonadotropin administration can predict impaired endometrial receptivity

    PubMed Central

    Aflatoonian, Abbas; Davar, Robab; Hojjat, Farzaneh

    2014-01-01

    Background: Increased serum progesterone on the day of human chorionic gonadotropin administration may affect in vitro fertilization (IVF) outcome. Objective: The aim of this study was to evaluate whether progesterone elevation on the day of human chorionic gonadotropin administration is associated with poor IVF outcome. Materials and Methods: To determine the relationship between serum progesterone on the day of HCG and the outcome of IVF-embryo transfer treatment, 378 infertile patients undergoing IVF-embryo transfer at Yazd Research and Clinical Center for Infertility from October 2009 to March 2011 were prospectively studied. Results: In this study, absolute p-value and P/E2 ratio were not a good predictor outcome of in-vitro fertilization but progesterone per metaphase II were predictive of implantation rate and pregnancy rate with statistically significant results but had no effect on the fertilization rate. Conclusion: We suggest avoided the increased progesterone that the cause of advanced endometrial maturation and impaired endometrial receptivity. If the progesterone is greater than 0.32 per oocyte metaphase II, the embryo transfer can be canceled and freezing all embryos for future transfer must be considered, to increase acceptance of the endometrium and thus increase the success rate. PMID:25071852

  2. Small gene family encoding an eggshell (chorion) protein of the human parasite Schistosoma mansoni

    SciTech Connect

    Bobek, L.A.; Rekosh, D.M.; Lo Verde, P.T.

    1988-08-01

    The authors isolated six independent genomic clones encoding schistosome chorion or eggshell proteins from a Schistosoma mansoni genomic library. A linkage map of five of the clones spanning 35 kilobase pairs (kbp) of the S. mansoni genome was constructed. The region contained two eggshell protein genes closely linked, separated by 7.5 kbp of intergenic DNA. The two genes of the cluster were arranged in the same orientation, that is, they were transcribed from the same strand. The sixth clone probably represents a third copy of the eggshell gene that is not contained within the 35-kbp region. The 5- end of the mRNA transcribed from these genes was defined by primer extension directly off the RNA. The ATCAT cap site sequence was homologous to a silkmoth chorion PuTCATT cap site sequence, where Pu indicates any purine. DNA sequence analysis showed that there were no introns in these genes. The DNA sequences of the three genes were very homologous to each other and to a cDNA clone, pSMf61-46, differing only in three or four nucleotices. A multiple TATA box was located at positions -23 to -31, and a CAAAT sequence was located at -52 upstream of the eggshell transcription unit. Comparison of sequences in regions further upstream with silkmoth and Drosophila sequences revealed very short elements that were shared. One such element, TCACGT, recently shown to be an essential cis-regulatory element for silkmoth chorion gene promoter function, was found at a similar position in all three organisms.

  3. Modulation of rat testes lipid composition by hormones: Effect of PRL (prolactin) and hCG (human chorionic gonadotropin)

    SciTech Connect

    Sebokova, E.; Wierzbicki, A.; Clandinin, M.T. )

    1988-10-01

    The effect of prolactin (PRL) and human chorionic gonadotropin (hCG) administration for 7 days on the composition and function of rat testicular plasma membrane was investigated. Refractory state in Leydig cells desensitized by hCG decreased the binding capacity for {sup 125}I-labeled hCG and also luteinizing hormone (LH)-induced adenosine 3{prime},5{prime}-cyclic monophosphate (cAMP) and testosterone production. In testicular membranes of hCG-treated animals, a depletion of cholesterol and an increase in total phospholipid content was observed after gonadotropin injection, thereby decreasing the cholesterol-to-phospholipid ratio. Injection of high doses of PRL had no effect on the binding capacity or affinity of the LH-hCG receptor but decreased the response of Leydig cells to LH in terms of cAMP and testosterone synthesis. PRL also increased total and esterified cholesterol and decreased free cholesterol and membrane phospholipid content. The fatty acid composition of testicular lipids was significantly and selectively influenced by both hormonal treatments. These observations suggest that metabolism of cholesterol and long-chain polyunsaturated fatty acids in testicular tissue is affected by chorionic gonadotropin and PRL and may provide the mechanism for regulating steroidogenic functions.

  4. Secretion of Unconjugated Androgens and Estrogens by the Normal and Abnormal Human Testis before and after Human Chorionic Gonadotropin

    PubMed Central

    Weinstein, R. L.; Kelch, R. P.; Jenner, M. R.; Kaplan, S. L.; Grumbach, M. M.

    1974-01-01

    The secretion of androgens and estrogens by normal and abnormal testes was compared by determining the concentrations of dehydroepiandrosterone (DHEA), androstenedione (Δ4A), testosterone (T), estrone (E1), and 17β-estradiol (E2) in peripheral and spermatic venous plasma samples from 14 normal men and 5 men with unilateral testicular atrophy. Four normal men and one patient with unilateral atrophy of the testis were given human chorionic gonadotropin (HCG) before surgery. Plasma estrogens were determined by radioimmunoassay; plasma androgens were measured by the double-isotope dilution derivative technique. Peripheral concentrations of these steroids before and after HCG were similar in both the normal men and the patients with unilateral testicular atrophy. In normal men, the mean ±SE spermatic venous concentrations were DHEA, 73.1±11.7 ng/ml; Δ4A, 30.7±7.9 ng/ml; T, 751±114 ng/ml; E1, 306±55 pg/ml; and E2, 1298±216 pg/ml. Three of four subjects with unilateral testicular atrophy had greatly diminished spermatic venous levels of androgens and estrogens. HCG treatment increased the testicular secretion of DHEA and T fivefold, Δ4A threefold, E1 sixfold, and E2 eightfold in normal men. In the single subject with an atrophic testis who received HCG, the spermatic venous concentrations of androgens and estrogens were much less than in normal men similarly treated. We conclude that: (a) E1 is secreted by the human testis, but testicular secretion of E1 accounts for less than 5% of E1 production in normal men; (b) HCG stimulation produces increases in spermatic venous estrogens equal to or greater than the changes in androgens, including testosterone; and (c) strikingly decreased secretion of androgen and estrogen by unilateral atrophic human tests cannot be appreciated by analyses of peripheral steroid concentrations. PMID:4271572

  5. Overexpression of the β Subunit of Human Chorionic Gonadotropin Promotes the Transformation of Human Ovarian Epithelial Cells and Ovarian Tumorigenesis

    PubMed Central

    Guo, Xiaoqing; Liu, Guangzhi; Schauer, Isaiah G.; Yang, Gong; Mercado-Uribe, Imelda; Yang, Fan; Zhang, Shiwu; He, Yuanli; Liu, Jinsong

    2011-01-01

    Ovarian carcinoma is the most lethal gynecologic malignancy, however underlying molecular events remain elusive. Expression of human chorionic gonadotropin β subunit (β-hCG) is clinically significant for both trophoblastic and nontrophoblastic cancers; however, whether β-hCG facilitates ovarian epithelial cell tumorigenic potential remains uncharacterized. Immortalized nontumorigenic ovarian epithelial T29 and T80 cells stably overexpressing β-hCG were examined for alterations in cell cycle and apoptotic status by flow cytometry, expression of proteins regulating cell cycle and apoptosis by Western blot, proliferation status by MTT assay, anchorage-independent colony formation, and mouse tumor formation. Immunoreactivity for β-hCG was evaluated using mouse xenografts and on human normal ovarian, fallopian tube, endometrium, and ovarian carcinoma tissues. T29 and T80 cells overexpressing β-hCG demonstrated significantly increased proliferation, anchorage-independent colony formation, prosurvival Bcl-XL protein expression, G2-checkpoint progression, elevated cyclins E/D1 and Cdk 2/4/6, and decreased apoptosis. Collectively, these transformational alterations in phenotype facilitated increased xenograft tumorigenesis (P < 0.05). Furthermore, β-hCG immunoreactivity was elevated in malignant ovarian tumors, compared with normal epithelial expression in ovaries, fallopian tube, and endometrium (P < 0.001). Our data indicate that elevated β-hCG transforms ovarian surface epithelial cells, facilitating proliferation, cell cycle progression, and attenuated apoptosis to promote tumorigenesis. Our results further decipher the functional role and molecular mechanism of β-hCG in ovarian carcinoma. β-hCG may contribute to ovarian cancer etiology, which introduces a new therapeutic intervention target for ovarian cancer. PMID:21763678

  6. Relationship between blood and urine concentrations of intact human chorionic gonadotropin and its free subunits in early pregnancy

    SciTech Connect

    Norman, R.J.; Menabawey, M.; Lowings, C.; Buck, R.H.; Chard, T.

    1987-04-01

    Paired blood and urine samples were obtained from patients between the sixth and 14th weeks of normal pregnancy. The levels of intact human chorionic gonadotropin (hCG), and of the free alpha and beta subunits, were measured by specific radioimmunoassays. There was a close association between blood and urine levels of intact hCG and of the alpha subunit of hCG, but no relation between the levels of beta subunit in these sites. These findings suggest that the use of beta subunit assays may give discrepant results according to the fluid examined. By contrast, measurement of intact hCG appears to give similar results in blood and urine.

  7. Amniotic membrane is a potential regenerative option for chronic non-healing wounds: a report of five cases receiving dehydrated human amnion/chorion membrane allograft.

    PubMed

    Mrugala, Andrew; Sui, Audrey; Plummer, Malgorzata; Altman, Igor; Papineau, Elaine; Frandsen, Devn; Hill, Danielle; Ennis, William J

    2016-08-01

    A case series of five patients with a total of six chronic non-healing wounds (>30 day duration) were non-randomly selected to evaluate the performance, safety and handling properties of dehydrated human amnion/chorion membrane allograft, an amniotic membrane scaffolding product. The patients had lower extremity wounds that had previously failed standard of care within a university outpatient/inpatient wound healing programme. Five wounds treated with dehydrated amnion/chorion membrane allograft showed a mean 43% area reduction from baseline (51% median) at 3 weeks into treatment and completely healed with a 64-day median time to closure (SD ±27·6 days). One wound worsened at 3 weeks and was found to have a complete central vein obstruction that was treated with long-term mild compression but still eventually healed at 6 months. Removing this outlier, the four responding wounds had a 72% mean and 69% median change in area from baseline, at the 3 week point. All five patients received only one application of dehydrated human amnion/chorion membrane allograft, and there were no adverse events. The product was easy to use, administer and handle. In summary, dehydrated human amnion/chorion membrane allograft appears to be a safe, effective and easy to use therapy for chronic non-healing wounds. This study describes the details of these clinical cases and provides an overview of the current evidence on the use of amniotic tissue in clinical practice. PMID:25974156

  8. Simple way to determine nonspecific effects of plasma and serum components in radioreceptor assays and radioimmunoassays for human chorionic gonadotropin

    SciTech Connect

    Rao, C.V.; Hussa, R.O.

    1982-01-15

    A simple approach is validated for the determination of nonspecific effects of human plasma and serum in the radioreceptor assays and radioimmunoassays for human chorionic gonadotropin (hCG). The approach is based on the findings that, despite differences in the degree of inhibition of /sup 125/I-hCG binding to its receptors and antibodies by nonhormonal components in different dilutions of pool plasma and serum, the standard curves (plotted as the percentage of control for each serum or plasma dilution) are superimposable. The approach consists of (1) running a single standard curve with no pool plasma or serum, (2) including a set of ''correction'' tubes which contain pool plasma or serum diluted correspondingly to the dilution of the unknown samples in the assay, (3) dividing the counts per minute found in the correction tubes into the counts per minute of the unknown samples and multiplying by 100, and (4) using this value to obtain the amount of hCG in the unknown samples by comparison with the no pool plasma or serum standard curve.

  9. Simple way to determine nonspecific effects of plasma and serum components in radioreceptor assays and radioimmunoassays for human chorionic gonadotropin

    SciTech Connect

    Rao, C.V.; Hussa, R.O.

    1982-01-15

    A simple approach is validated for the determination of nonspecific effects of human plasma and serum in the radioreceptor assays and radioimmunoassays for human chorionic gonadotropin (hCG). The approach is based on the findings that, despite differences in the degree of inhibition of /sup 125/I-hCG binding to its receptors and antibodies by nonhormonal components in different dilutions of pool plasma and serum, the standard curves (plotted as the percentage of control for each serum or plasma dilution) are superimposable. The approach consists of (1) running a single standard curve with no pool plasma or serum, (2) including a set of correction tubes which contain pool plasma or serum diluted correspondingly to the dilution of the unknown samples in the assay, (3) dividing the counts per minute found in the correction tubes into the counts per minute of the unknown samples and multiplying by 100, and (4) using this value to obtain the amount of hCG in the unknown samples by comparison with the no pool plasma or serum standard curve.

  10. Effect of adding human chorionic gonadotropin to frozen thawed embryo transfer cycles with history of thin endometrium

    PubMed Central

    Davar, Robab; Miraj, Sepideh; Farid Mojtahedi, Maryam

    2016-01-01

    Background: Embryo implantation process is a complex phenomenon and depends on fetal and maternal factors interaction. Endometrial thickness is needed for successful implantation. Objective: We designed this study in order to assess adding human chorionic gonadotropin (HCG) to the conventional protocol in endometrial preparation in women with thin endometrium and a history of in vitro fertilization–embryo transfer (IVF-ET) failure. Materials and Methods: The non-randomized clinical trial study (quasi experimental design) was performed on 28 patients. Participants were women who were candidate for frozen-thawed (ET) and had two previous failed ET cycles because of thin endometrial. HCG was administrated (150 IU, intramuscular) from the 8th day of cycle and when endometrial thickness reached at least 7mm HCG was discontinued and frozen thawed ET was done. Results: Totally 28 patients were included. The mean ± SD age of participants was 30.39±4.7. The mean of endometrium thickness before and after HCG were 5.07±0.43 and 7.85±0.52, respectively p<0.001. Also, there were five clinically and chemically pregnant women. Conclusion: The findings of the study suggested that adding HCG to the conventional preparation method was an effective protocol and significantly improved endometrial thickness and pregnancy outcomes in women with previous embryo transfer failure because of thin endometrium. PMID:27141549

  11. Limitations in the process of transcription and translation inhibit recombinant human chorionic gonadotropin expression in CHO cells.

    PubMed

    Liu, Yang; Yi, Xiaoping; Zhuang, Yingping; Zhang, Siliang

    2015-06-20

    Human chorionic gonadotropin (hCG) is a glycoprotein hormone that exists as a heterodimer with a α subunit and β subunit assembled together with disulfide bridges. This hormone plays an important role in the detection of ovulation induction and in the treatment of certain diseases that cause female infertility. The effects of transcription, subunit expression, assembling and secretion on recombinant hCG expression in CHO cells were studied using stable high-producing and low-producing cell lines generated by the FLP-In™ system. The results indicated that the mRNA and polypeptide levels of the β subunit were always higher than those of the α subunit. Further study confirmed that the differences were caused by the transcription rate rather than by mRNA stability. In the high-producing cell lines, there was obvious transcription level limitation of the α subunit in contrast to the β subunit. In addition, there was obvious limitation of the synthetic steps from mRNA to polypeptide for both the α subunit and the β subunit, especially the β subunit. Significant limitations of the assembly and secretion levels were not observed in this research. This study presents a research methodology for double subunit protein expression and provides valuable evidence for the enhancement of recombinant hCG productivity. PMID:25529346

  12. Targeting vaccinia virus-expressed secretory beta subunit of human chorionic gonadotropin to the cell surface induces antibodies.

    PubMed Central

    Srinivasan, J; Singh, O; Chakrabarti, S; Talwar, G P

    1995-01-01

    We carried out experiments designed to study the effect of a protein's localization on its immunogenicity. A novel cell-surface protein was generated from a small, glycosylated secretory protein. The DNA sequence encoding the entire precursor of the human chorionic gonadotropin beta (beta hCG) subunit was fused in the correct reading frame to the DNA sequence encoding the transmembrane and cytoplasmic domains of vesicular stomatitis virus glycoprotein. This chimeric gene was introduced into the vaccinia virus genome to generate a recombinant virus. The recombinant virus, when used to infect animal cells, expressed a 135-amino-acid beta hCG subunit anchored in cellular membranes by the 48 carboxy-terminal amino acids of vesicular stomatitis virus glycoprotein. The immunogenicity of this recombinant virus with respect to its ability to generate anti-hCG antibodies was compared with that of a second recombinant vaccinia virus expressing the native secretory form of beta hCG. All animals immunized with the vaccinia virus expressing beta hCG on the cell surface elicited high titers of anti-hCG antibodies. Even after a single immunization with the recombinant vaccinia virus, the anti-hCG antibody titers persisted for a long period of time (more than 6 months). None of the animals immunized with vaccinia virus expressing the native secretory form of beta hCG showed any hCG-specific antibody response. PMID:7591154

  13. A controlled study of human chorionic gonadotrophin induced ovulation versus urinary luteinizing hormone surge for timing of intrauterine insemination.

    PubMed

    Martinez, A R; Bernadus, R E; Voorhorst, F J; Vermeiden, J P; Schoemaker, J

    1991-10-01

    Forty-eight patients in a programme of intrauterine insemination (IUI) were randomized in a cross-over study. All were stimulated with clomiphene citrate (CC) and inseminated either after follicular rupture induced by human chorionic gonadotrophin (HCG) or after a spontaneous urinary luteinizing hormone (LH) surge. The HCG was administered when follicles of 18-22 mm in diameter were observed on ultrasound and IUI was performed 37-40 h thereafter. The monitoring of a urinary LH peak was carried out using a rapid urinary LH test. IUI took place approximately 22 h after detection of the LH surge. Overall, the pregnancy rates were 9.3% (4/43) after HCG induced ovulation and 20.5% (9/44) after spontaneous ovulation (P = 0.12). Analysis of mid-cycle events showed that following sonographic criteria, the HCG injection was performed significantly earlier in the cycle compared with the spontaneous LH surge. In addition, the mean diameter of the preovulatory follicles was significantly smaller and insemination was substantially earlier in the HCG induced cycles. These findings suggest that a beneficial effect arises from allowing the natural process of final follicular maturation to occur. PMID:1752926

  14. Biosynthesis of a biologically active single peptide chain containing the human common alpha and chorionic gonadotropin beta subunits in tandem.

    PubMed Central

    Sugahara, T; Pixley, M R; Minami, S; Perlas, E; Ben-Menahem, D; Hsueh, A J; Boime, I

    1995-01-01

    One of the distinguishing features of the gonadotropin and thyrotropin hormone family is their heterodimeric structure, consisting of a common alpha subunit and a hormone-specific beta subunit. Subunit assembly is vital to the function of these hormones: The conformation of the heterodimer is essential for controlling secretion, hormone-specific posttranslational modifications, and signal transduction. To address whether alpha and beta subunits can be synthesized as one chain and also maintain biological activity, a chimera composed of the human chorionic gonadotropin (hCG) beta subunit genetically fused to the alpha subunit was constructed. The resulting polypeptide hCG molecule not only was efficiently secreted but also displayed an increased biological activity in vitro and in vivo. These data show that the alpha and hCG beta subunits encoded as a single chain retain a biologically active conformation similar to that seen in the heterodimer. This approach can be used to investigate structure-function relationships of the glycoprotein hormone family that were previously not tractable because of the absolute dependence on assembly for the biological response. Moreover, other bioactive multisubunit ligands can be engineered where the combination efficiency and specificity of heterodimers and homodimers are otherwise difficult to control. Images Fig. 2 Fig. 3 PMID:7892221

  15. Commercial radioimmunoassay for beta subunit of human chorionic gonadotropin: falsely positive determinations due to elevated serum luteinizing hormone

    SciTech Connect

    Fowler, J.E. Jr.; Platoff, G.E.; Kubrock, C.A.; Stuzman, R.E.

    1982-01-01

    Among 17 men who had received seemingly curative treatment for unilateral non-seminomatous germ cell tumors for the testis and who had consistently normal serum human chorionic gonadotropin (HCG) levels at a reference laboratory, 7 (41%) had at least one falsely positive commercial serum HCG determination. To investigate the cause of these falsely positive determinations the authors measured the cross reactivity of luteinizing hormone (LH) and follicle stimulating hormone (FSH) standards in the commercial HCG assay, and studied the relationships between commercial HCG levels and serum LH levels, serum FSH levels and gonadal status in men with and without normal gonadal function. The falsely positive HCG determinations appeared to be due to elevated serum LH levels and cross reactivity of LH in the commercial HCG assay because: 1) there was substantial cross reactivity of the LH standards in the commercial assay, 2) the serum LH was elevated in four of six men with solitary testes, 3) there was a striking correlation between elevated serum LH levels and falsely elevated commercial HCG levels in ten men with solitary or absent testes, and 4) there were no falsely positive HCG determinations in 13 normal men but there were falsely positive HCG determinations in seven of ten anorchid men.

  16. Predictive value of early serum beta-human chorionic gonadotrophin for the successful outcome in women undergoing in vitro fertilization

    PubMed Central

    Singh, Neeta; Goyal, Manu; Malhotra, Neena; Tiwari, Abanish; Badiger, Shreenivas

    2013-01-01

    AIMS: Pregnancies achieved by in vitro fertilization (IVF) are at increased risk of adverse outcome. The main objective of this study was to evaluate the predictive value of β-human chorionic gonadotrophin (β-HCG) and age of the patient for the successful outcome in IVF. MATERIALS AND METHODS: A retrospective study was done in 139 pregnancies after IVF at single IVF center from June 2007 to July 2012. The age of the patient and initial serum values of β-HCG on day 14 of embryo transfer were correlated with ongoing pregnancy (>12 weeks gestation). RESULTS: The β-HCG level on day 14 of more than 347 mIU/ml has a sensitivity of 72.2% and specificity of 73.6% in prediction of pregnancy beyond 12 weeks period of gestation. Positive likelihood ratio (LR) is 2.74 and negative LR is 0.37, (receiver operating characteristic area = 0.79). DISCUSSION: In IVF cycles, there is a lot of stress on the couples while the cycle is going on. There was a positive correlation between the higher values of early serum β-HCG levels and ongoing pregnancy. Hence, it can be used as an independent predictor of a successful outcome of IVF cycle. CONCLUSION: We concluded from our study that early serum β-HCG can be used as a predictor of a successful outcome in IVF. PMID:24672163

  17. Effects of preventing O-glycosylation on the secretion of human chorionic gonadotropin in Chinese hamster ovary cells

    SciTech Connect

    Matzuk, M.M.; Krieger, M.; Corless, C.L.; Boime, I.

    1987-09-01

    Human chorionic gonadotropin (hCG) is a member of a family of heterodimeric glycoprotein hormones that have a common ..cap alpha.. subunit but differ in their hormone-specific ..beta..-subunits. The ..beta.. subunit of hCG (hCG..beta..) is unique among the ..beta.. subunits in that it contains four mucin-like O-linked oligosaccharides attached to a carboxyl-terminal extension. To study the effects of O-glycosylation on the secretion and assembly of hCG, expression vectors containing either hCG..beta.. gene alone or together with the hCG..cap alpha.. gene were transfected into a mutant Chinese hamster ovary cell line, 1d1D, which exhibits a reversible defect in O-glycosylation. The results reveal that hCG..beta.. can be secreted normally in the absence of its O-linked oligosaccharides. hCG..beta.. devoid of O-linked carbohydrate can also combine efficiently with hCG..cap alpha.. and be secreted as an intact dimer. The authors conclude that in Chinese hamster ovary cells, the hCG..beta.. O-linked chains play no role in the assembly and secretion of hCG. The normal and O-linked oligosaccharide-deficient forms of hCG secreted by these cells should prove useful in examining the role of O-linked chains on the biological function of hCG.

  18. The Effect of Progestins on Tumor Necrosis Factor α-Induced Matrix Metalloproteinase 9 Activity and Gene Expression in Human Primary Amnion and Chorion Cells In Vitro

    PubMed Central

    Allen, Terrence K; Feng, Liping; Nazzal, Matthew; Grotegut, Chad A; Buhimschi, Irina A; Murtha, Amy P

    2015-01-01

    Background Current treatment modalities for preventing preterm premature rupture of membranes (PPROM) are limited, but progestins may play a role. Tumor necrosis factor α (TNFα) enhances matrix metalloproteinase 9 (MMP-9) gene expression and activity in fetal membranes, contributing to membrane weakening and rupture. We previously demonstrated that progestins attenuate TNFα-induced MMP-9 activity in a cytotrophoblast cell line. However, whether they have a similar effect in primary amnion and chorion cells of fetal membranes is unknown. In this study we evaluated the effect of progestins on basal and TNFα-induced MMP-9 activity and gene expression in primary chorion and amnion cells harvested from the fetal membranes of term non-laboring patients. Methods Primary amnion and chorion cells were isolated from fetal membranes obtained from term uncomplicated non-laboring patients following elective cesarean delivery (n=11). Confluent primary amnion and chorion cell cultures were both pretreated with vehicle (control), progesterone (P4), 17α-hydroxyprogesterone caproate (17P) or medroxyprogesterone acetate (MPA) at 10-6 M concentration for 6 h followed by stimulation with TNFα at 10 ng/mL for an additional 24 h. Cell cultures pretreated with the vehicle only served as the unstimulated control and the vehicle stimulated with TNFα served as the stimulated control. Both controls were assigned a value of 100 units. Cell culture medium was harvested for MMP-9 enzymatic activity quantification using gelatin zymography. Total RNA was extracted for quantifying MMP-9 gene expression using RT-qPCR. Basal MMP-9 activity and gene expression data were normalized to the unstimulated control. TNFα-stimulated MMP-9 activity and gene expression were normalized to the stimulated control. The primary outcome was the effect of progestins on TNFα-induced MMP-9 enzymatic activity in term human primary amnion and chorion cells in vitro. Secondary outcomes included the effect of

  19. A Point-of-Care Immunosensor for Human Chorionic Gonadotropin in Clinical Urine Samples Using a Cuneated Polysilicon Nanogap Lab-on-Chip

    PubMed Central

    Balakrishnan, S. R.; Hashim, U.; Gopinath, Subash C. B.; Poopalan, P.; Ramayya, H. R.; Iqbal Omar, M.; Haarindraprasad, R.; Veeradasan, P.

    2015-01-01

    Human chorionic gonadotropin (hCG), a glycoprotein hormone secreted from the placenta, is a key molecule that indicates pregnancy. Here, we have designed a cost-effective, label-free, in situ point-of-care (POC) immunosensor to estimate hCG using a cuneated 25 nm polysilicon nanogap electrode. A tiny chip with the dimensions of 20.5 × 12.5 mm was fabricated using conventional lithography and size expansion techniques. Furthermore, the sensing surface was functionalized by (3-aminopropyl)triethoxysilane and quantitatively measured the variations in hCG levels from clinically obtained human urine samples. The dielectric properties of the present sensor are shown with a capacitance above 40 nF for samples from pregnant women; it was lower with samples from non-pregnant women. Furthermore, it has been proven that our sensor has a wide linear range of detection, as a sensitivity of 835.88 μA mIU-1 ml-2 cm-2 was attained, and the detection limit was 0.28 mIU/ml (27.78 pg/ml). The dissociation constant Kd of the specific antigen binding to the anti-hCG was calculated as 2.23 ± 0.66 mIU, and the maximum number of binding sites per antigen was Bmax = 22.54 ± 1.46 mIU. The sensing system shown here, with a narrow nanogap, is suitable for high-throughput POC diagnosis, and a single injection can obtain triplicate data or parallel analyses of different targets. PMID:26368287

  20. A Point-of-Care Immunosensor for Human Chorionic Gonadotropin in Clinical Urine Samples Using a Cuneated Polysilicon Nanogap Lab-on-Chip.

    PubMed

    Balakrishnan, S R; Hashim, U; Gopinath, Subash C B; Poopalan, P; Ramayya, H R; Iqbal Omar, M; Haarindraprasad, R; Veeradasan, P

    2015-01-01

    Human chorionic gonadotropin (hCG), a glycoprotein hormone secreted from the placenta, is a key molecule that indicates pregnancy. Here, we have designed a cost-effective, label-free, in situ point-of-care (POC) immunosensor to estimate hCG using a cuneated 25 nm polysilicon nanogap electrode. A tiny chip with the dimensions of 20.5 × 12.5 mm was fabricated using conventional lithography and size expansion techniques. Furthermore, the sensing surface was functionalized by (3-aminopropyl)triethoxysilane and quantitatively measured the variations in hCG levels from clinically obtained human urine samples. The dielectric properties of the present sensor are shown with a capacitance above 40 nF for samples from pregnant women; it was lower with samples from non-pregnant women. Furthermore, it has been proven that our sensor has a wide linear range of detection, as a sensitivity of 835.88 μA mIU(-1) ml(-2) cm(-2) was attained, and the detection limit was 0.28 mIU/ml (27.78 pg/ml). The dissociation constant Kd of the specific antigen binding to the anti-hCG was calculated as 2.23 ± 0.66 mIU, and the maximum number of binding sites per antigen was Bmax = 22.54 ± 1.46 mIU. The sensing system shown here, with a narrow nanogap, is suitable for high-throughput POC diagnosis, and a single injection can obtain triplicate data or parallel analyses of different targets. PMID:26368287

  1. Comparison of recombinant human follicle stimulating hormone (rhFSH), human chorionic gonadotropin (HCG) and human menopausal gonadotropin (HMG) on semen parameters after varicocelectomy: a randomized clinical trial

    PubMed Central

    Amirzargar, Mohammad Ali; Yavangi, Mahnaz; Basiri, Abbass; Hosseini Moghaddam, Sayyed Mahdi; Babbolhavaeji, Hooshang; Amirzargar, Nasibeh; Amirzargar, Hossein; Moadabshoar, Leila

    2012-01-01

    Background: The most frequent physical finding in infertile men is varicocele, in which one of the mechanisms that can affect seminal parameters is oxidative stress. Objective: Our study aimed, for the first time, to compare the efficacy of recombinant human follicle-stimulating hormone (rhFSH), human chorionic gonadotropin (HCG) and human menopausal gonadotropin (HMG) on sperm parameters and fertility after varicocelectomy. Materials and Methods: 113 infertile men with varicocele were divided into four groups. Group A received HCG 5000 IU weekly, group B received HMG 75 IU three times a week, group C received rhFSH 75 IU three times a week and group D received no medical treatment after varicocelectomy. Results: After three months, in group A sperm morphology improved (p=0.007), causing a 32% pregnancy rate. In group B, sperm motility (p=0.023) and morphology (p=0.014) improved, causing a 57% pregnancy rate. In group C, all of the investigated semen parameters increased (p<0.05), causing a 62.5% pregnancy rate. Only rhFSH improved sperm concentrations to >20×106 mL (p=0.027). In group D, sperm morphology increased (p=0.038), but other parameters remained unchanged and no pregnancies occurred. Conclusion: It can be concluded that drugs can reduce induction time for spermatogenesis and fertility in comparison with varicocelectomy alone. For these purposes, rhFSH is more effective than other drugs. PMID:25246910

  2. Squelching of ETS2 transactivation by POU5F1 silences the human chorionic gonadotropin CGA subunit gene in human choriocarcinoma and embryonic stem cells.

    PubMed

    Gupta, Rangan; Ezashi, Toshihiko; Roberts, R Michael

    2012-05-01

    The subunit genes encoding human chorionic gonadotropin, CGA, and CGB, are up-regulated in human trophoblast. However, they are effectively silenced in choriocarcinoma cells by ectopically expressed POU domain class 5 transcription factor 1 (POU5F1). Here we show that POU5F1 represses activity of the CGA promoter through its interactions with ETS2, a transcription factor required for both placental development and human chorionic gonadotropin subunit gene expression, by forming a complex that precludes ETS2 from interacting with the CGA promoter. Mutation of a POU5F1 binding site proximal to the ETS2 binding site does not alter the ability of POU5F1 to act as a repressor but causes a drop in basal promoter activity due to overlap with the binding site for DLX3. DLX3 has only a modest ability to raise basal CGA promoter activity, but its coexpression with ETS2 can up-regulate it 100-fold or more. The two factors form a complex, and both must bind to the promoter for the combination to be transcriptionally effective, a synergy compromised by POU5F1. Similarly, in human embryonic stem cells, which express ETS2 but not CGA, ETS2 does not occupy its binding site on the CGA promoter but is found instead as a soluble complex with POU5F1. When human embryonic stem cells differentiate in response to bone morphogenetic protein-4 and concentrations of POU5F1 fall and hCG and DLX3 rise, ETS2 then occupies its binding site on the CGA promoter. Hence, a squelching mechanism underpins the transcriptional silencing of CGA by POU5F1 and could have general relevance to how pluripotency is maintained and how the trophoblast lineage emerges from pluripotent precursor cells. PMID:22446105

  3. Human chorionic gonadotropin (a luteinizing hormone homologue) decreases spatial memory and increases brain amyloid-β levels in female rats

    PubMed Central

    Berry, Anne S.; Tomidokoro, Yasushi; Ghiso, Jorge; Thornton, Jan

    2008-01-01

    Numerous studies have suggested that estradiol (E) improves spatial memory as female rats with E perform better than those without E. However there is an inverse relationship between E and luteinizing hormone (LH) levels and LH could play a role. We examined whether treatment with the LH homologue human chorionic gonadotropin (hCG), would impair spatial memory of adult E-treated female rats. In the Object Location Memory Task, ovariectomized (ovxed) rats treated with E and either a single high dose (400IU/kg) or a lower repeated dose of hCG (75 IU/kg hourly for 8 hours) showed spatial memory disruption compared to ovxed rats treated with estradiol alone. Impairment was attributed to memory disruption as performance improved with shortened delay between task exposure and testing. Tests on another spatial memory task, the Barnes maze, confirmed that hCG (400IU/kg) can impair memory: although E + veh treated animals made significantly fewer hole errors across time, E + hCG treated did not. In humans, high LH levels have been correlated with Alzheimer’s Disease (AD). Because brain amyloid-beta (Aβ) species have been implicated as a toxic factor thought to cause memory loss in AD, we analyzed whether hCG-treated animals had increased Aβ levels. Levels of Aβ from whole brains or hippocampi were assessed by Western Blot. hCG treatment to E-implanted females significantly increased soluble Aβ40 and Aβ42 levels. These results indicate that high levels of LH/hCG can impair spatial memory, and an increase in brain Aβ species may account for the memory impairment in hCG-treated rats. PMID:18413150

  4. 21 CFR 862.1155 - Human chorionic gonadotropin (HCG) test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... intended to measure HCG, a placental hormone, in plasma or urine. (2) Classification. Class II. (b) Human... persons with certain tumors or carcinomas) is intended to measure HCG, a placental hormone, in plasma...

  5. 21 CFR 862.1155 - Human chorionic gonadotropin (HCG) test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... intended to measure HCG, a placental hormone, in plasma or urine. (2) Classification. Class II. (b) Human... persons with certain tumors or carcinomas) is intended to measure HCG, a placental hormone, in plasma...

  6. Embryo survival from gossypol-fed heifers after transfer to lactating cows treated with human chorionic gonadotropin.

    PubMed

    Galvão, K N; Santos, J E P; Coscioni, A C; Juchem, S O; Chebel, R C; Sischo, W M; Villaseñor, M

    2006-06-01

    Objectives were to determine the effects of gossypol exposure during early embryo development on embryonic survival after transfer of frozen and thawed embryos to lactating dairy cows treated with human chorionic gonadotropin (hCG). Holstein cows (n = 269) were either treated or not treated with 3,300 IU of hCG on d 5 of the estrous cycle and received an embryo collected from heifers fed or not fed gossypol. Embryo donor heifers consumed either 0 or 12 g/d of free gossypol for 76 d prior to embryo collection, resulting in mean plasma gossypol concentrations of 0 and 7.38 microg/mL, respectively. Embryos were transferred on d 7 of the estrous cycle and pregnancy diagnosed 21 and 35 d later. Progesterone was analyzed in plasma collected on d 5 and 12 of the estrous cycle. Treatment with hCG increased the total luteal area on d 12 (818.0 vs. 461.1 mm2) because of increased number of corpora lutea (2.0 vs. 1.0) and increased area of the original corpora lutea (522.7 vs. 443.5 mm2). Plasma progesterone concentrations were similar between treatments on d 5, but increased by d 12 in hCG-treated cows (6.46 vs. 4.78 ng/ mL). Pregnancy rates on d 28 and 42 were not affected by hCG. However, after transfer into lactating cows, embryos collected from heifers not fed gossypol resulted in higher pregnancy rates at 28 d (33.3 vs. 23.1%) and 42 d (29.6 vs. 20.2%) of gestation compared with embryos collected from heifers fed gossypol. Our data suggest that the negative effects of gossypol on fertility are mediated by changes in embryo viability in spite of similar grade quality at transfer. PMID:16702269

  7. Second-trimester maternal serum alpha-fetoprotein, human chorionic gonadotropin, and unconjugated oestriol after early transvaginal multifetal pregnancy reduction.

    PubMed

    Groutz, A; Amit, A; Yaron, Y; Yovel, I; Wolman, I; Legum, C; Lessing, J B

    1996-08-01

    Maternal serum alpha-fetoprotein (MSAFP), human chorionic gonadotropin (hCG), and unconjugated oestriol (UE3) are used as second-trimester screening markers for the detection of various fetal abnormalities. Previous studies have suggested that second-trimester MSAFP is consistently elevated after late first-trimester transabdominal multifetal pregnancy reduction (MFPR). The present study was undertaken to evaluate the levels of all three markers after early transvaginal MFPR. Maternal serum was examined for MSAFP, hCG, and UE3 at 16-18 weeks' gestation in 28 patients who underwent transvaginal MFPR at approximately 10 weeks' gestation. The mean interval between the reduction procedure and the screening test was 7.2 +/- 0.9 weeks. The mean MSAFP value in 24 patients carrying viable twins was 2.49 +/- 0.99 multiples of the median (MOM). Two patients had elevated MSAFP values: one in association with omphalocoele and the other in relation to an adverse pregnancy outcome. All but two patients had normal hCG values (mean 1.98 +/- 1.26 MOM). Two cases with elevated hCG were associated with an adverse pregnancy outcome. Unconjugated oestriol values were within the normal range in all patients (mean 1.69 +/- 0.61 MOM). These results suggest that early transvaginal MFPR, at approximately 10 weeks' gestation, does not appear to influence second-trimester MSAFP, hCG, and UE3 levels. The values of these markers may therefore be interpreted by using the same criteria as those for the general obstetric population. PMID:8878282

  8. Determining an Optimal Cutoff of Serum β-Human Chorionic Gonadotropin for Assisting the Diagnosis of Intracranial Germinomas

    PubMed Central

    Zhang, Hui; Zhang, Peng; Fan, Jun; Qiu, Binghui; Pan, Jun; Zhang, Xi’an; Fang, Luxiong; Qi, Songtao

    2016-01-01

    Background Beta (β)-human chorionic gonadotropin (β-HCG) is used to confirm the diagnosis and plan treatment of intracranial germinomas. However, the cutoff values of serum β-HCG in diagnosis of intracranial germinomas reported in the literature are inconsistent. To establish an appropriate cutoff value of serum β-HCG for diagnosis of intracranial germinomas, we retrospectively reviewed the records of intracranial tumor patients who received serum β-HCG and α-fetoprotein (AFP) tests for diagnostic purposes at our hospital from 2005 to 2014. Methods A total of 93 intracranial germinomas and 289 intracranial non-germ cell tumors were included in this study. Receiver operating characteristic (ROC) analysis was used to evaluate the sensitivity and specificity of 3 cutoffs (0.1, 0.4, and 0.5 mIU/mL) for diagnosing intracranial germinomas. The serum β-HCG level of intracranial germinoma patients was further analyzed to investigate the effect of metastasis status and tumor location on serum β-HCG level. Results The area under the ROC curve was 0.81 (P < .001), suggesting β-HCG is an effective marker. Of the 3 cutoff values, 0.1 mIU/mL possessed a highest sensitivity (66.67%) and good specificity (91%). Although there was no β-HCG level difference between metastatic and non-metastatic intracranial germinoma patients, the diagnostic rate of metastatic neurohypophyseal germinomas was significantly higher than that of its non-metastatic counterpart (P < .05), implying that the location of the germinoma might need to be considered when β-HCG is used as a marker to predict metastasis. Conclusions Determining an optimal cutoff of serum β-HCG is helpful for assisting the diagnosis of intracranial germinoma. PMID:26771195

  9. Advancing or postponing the day of human chorionic gonadotropin does not matter for the outcome in assisted reproductive technology

    PubMed Central

    Pratap, Deepika; Kumar, Pratap; Adiga, Satish Kumar

    2014-01-01

    CONTEXT: The primary outcome is to remove the worry of getting immature oocytes with early administration of human chorionic gonadotropin (hCG). AIM: The aim was to find out the association between the day of hCG administration and follicular response in relation to the number and maturity of oocytes, and fertilization rate in assisted reproduction to avoid weekend oocyte recovery (OR). SETTINGS AND DESIGN: Retrospective study was carried out in the university infertility clinic. MATERIALS AND METHOD: Controlled ovarian hyperstimulation (COH) in 94 patients undergoing assisted reproductive technology (2010-2011) with recombinant follicle stimulating hormone and timely gonadotropin-releasing hormone antagonist were analyzed regarding day of hCG from day 8-11. Oocyte maturity and fertilization was analyzed and correlated with the day of hCG administration. STATISTICAL ANALYSIS: Kruskal–Wallis test. RESULTS: The average number of >18 mm follicles observed from day 8-11 of hCG administration was not statistically different. However, the OR rate (54.2%), number of mature oocytes (92.5%), and fertilization rate (78.5%) was maximum in the patients where hCG was administered on day 8 of COH. CONCLUSIONS: The day of hCG administration between 8 and 11 does not affect the OR rate significantly although the number of oocytes recovered on day 8 are marginally higher compared with day 9-11. Hence, it is possible to safely avoid weekend oocyte retrieval, by delaying or advancing hCG administration without compromising the outcome. PMID:25191023

  10. Synchronization of ovulation with human chorionic gonadotropin in lactating dairy cows with ovarian cysts during heat stress.

    PubMed

    Navanukraw, Chainarong; Khanthusaeng, Vilaivan; Kraisoon, Aree; Suwannarit, Duangkamon; Jarassaeng, Chaiwat; Aiumlamai, Suneerat

    2015-06-01

    A study was conducted during hot season to determine the effect of synchronization of ovulation with human chorionic gonadotropin (hCG) on fertility of lactating dairy cows with ovarian cysts. Non cyclic Holstein dairy cows (n = 80) were stratified by parity and diagnosed as having an ovarian cyst. The cows were further identified as follicular or luteal cysts according to the plasma progesterone (P4) concentration and the cystic image of ultrasonography. Cystic cows were randomly assigned to receive treatments (Ovsynch as the control or Ovsynch plus 3000 IU hCG). All cows were artificially inseminated at 16-18 h after the second gonadotropin releasing hormone injection. Cows supplemented with hCG had a greater number of corpus luteum (1.8 ± 0.2 and 0.8 ± 0.3; P < 0.05) and had greater P4 concentration on day 12 than those control cows (6.3 ± 0.3 and 3.9 ± 0.4 ng/ml; P < 0.05). Concentration of cortisol did not differ between groups of cystic cows. No significant differences were found in overall conception rates between the treatments; however, significantly greater conception rate (P = 0.03) was observed in cows with luteal cysts receiving Ovsynch plus hCG. This study highlights that administration of hCG following the Ovsynch-based timed artificial insemination (AI) is more effective than the control Ovsynch by which the hCG affects corpus luteum (CL) development, P4 concentration, and thus improves conception rate in dairy cows with luteal cysts. PMID:25863956

  11. The use of human amnion/chorion membrane in the clinical setting for lower extremity repair: a review.

    PubMed

    Zelen, Charles M; Snyder, Robert J; Serena, Thomas E; Li, William W

    2015-01-01

    Unsuccessful healing represents a significant medical dilemma for both patients and clinicians and create a financial burden on the health care system. Despite the efforts of physicians, many injuries fail to heal with standard care alone. For decades human amniotic membrane composed of both amnion and chorion has shown remarkable therapeutic potential, but only recent preservation methods have allowed its widespread use. Variations in configurations and the ability to micronize the material allow clinical uses that were previously not possible. Although there are limited data available regarding most amniotic membrane-based products, there is substantial preclinical and clinical evidence supporting the rationale and effectiveness of dHACM allograft as a treatment modality. The rapidly growing body of evidence suggests that the properties inherent in dHACM promote tissue regeneration and healing, recruiting patients' own stem cells into the wounded area. Randomized controlled trials evaluating dHACM now include more than 200 patients collectively and the results consistently show improved healing. Use of dHACM has been shown to be more clinically effective and cost-effective than other frequently used advanced wound care products. This cost-effectiveness results from dHACM showing higher healing rates and more rapid healing than other advanced wound care products. Cost-effectiveness is also enhanced through the availability of grafts of multiple sizes, which reduces wastage, and through ease of handling and storage for clinical use. Ongoing and future studies will further define and establish the value of amniotic membrane for chronic tissue repair and regeneration. PMID:25440424

  12. Premature progesterone rise at human chorionic gonadotropin triggering day has no correlation with intracytoplasmic sperm injection outcome

    PubMed Central

    Saharkhiz, Nasrin; Salehpour, Saghar; Tavasoli, Mahboobeh; Aghighi, Ali

    2015-01-01

    Background: Premature luteinization during in vitro fertilization was commonly happened before the introduction of GnRh analogues. High level of unwanted progesterone is associated with adverse pregnancy outcome and is thought to be induced by inappropriate LH elevation. Objective: To evaluate the progesterone level on the day of Human Chorionic Gonadotropin (HCG) triggering in GnRh agonist and antagonist protocols, and its correlation with clinical pregnancy rate and miscarriage rate. Materials and Methods: One hundred and seven women underwent intracytoplasmic sperm injection with long agonist protocol (n=46) or antagonist protocol (n=61). Blood sample was obtained from each patient for progesterone level measurement in HCG administration day, then patients were divided into two groups according to their serum progesterone levels on the HCG triggering day: progesterone level <1.2 ng/ml, and progesterone level ≥1.2 ng/ml. Clinical pregnancy and miscarriage rates were evaluated as main outcomes and biochemical pregnancy rate and implantation rate were considered as secondary outcomes. Results: The increased prevalence rate of premature progesterone (progesterone level ≥1.2 ng/ml) in total patients was 13.1% (14/107) and in long agonist protocol group and antagonist protocol group was 15.2% (7/46) and 11.5% (7/61) respectively. Premature progesterone rise had no significant correlation with clinical pregnancy rate in total patients (p=0.174), agonist protocol (p=0.545), and antagonist protocol (p=0.129). Also premature progesterone rise had no significant association with miscarriage rate in total patients (p=0.077), agonist protocol group (p=0.383) and antagonist protocol group (p=0.087). Conclusion: A significant rise in progesterone levels at the time of HCG triggering doesn’t lead to decrease in pregnancy rate and implantation rate and increase in miscarriage rate. PMID:25999996

  13. Energy status and HIF signalling in chorionic villi show no evidence of hypoxic stress during human early placental development.

    PubMed

    Cindrova-Davies, T; van Patot, M Tissot; Gardner, L; Jauniaux, E; Burton, G J; Charnock-Jones, D S

    2015-03-01

    Early human placental and embryonic development occurs in a physiologically low oxygen environment supported by histiotrophic secretions from endometrial glands. In this study, we compare the placental metabolomic profile in the first, second and third trimesters to determine whether the energy demands are adequately met in the first trimester. We investigated whether hypoxia-inducible factors, HIF-1α and/or HIF-2α, might regulate transcription during the first trimester. First and second trimester tissue was collected using a chorionic villus sampling-like (CVS) technique. Part of each villus sample was frozen immediately and the remainder cultured under 2 or 21% O2 ± 1 mM H2O2, and ±the p38 MAPK pathway inhibitor, PD169316. Levels of HIF-1α were assessed by western blotting and VEGFA, PlGF and GLUT3 transcripts were quantified by RT-PCR. Term samples were collected from normal elective Caesarean deliveries. There were no significant differences in concentrations of ADP, NAD(+), lactate, and glucose, and in the ATP/ADP ratio, across gestational age. Neither HIF-1α nor HIF-2α could be detected in time-zero CVS samples. However, culture under any condition (2 or 21% O2 ± 1 mM H2O2) increased HIF-1α and HIF-2α. HIF-1α and HIF-2α were additionally detected in specimens retrieved after curettage. HIF-1α stabilization was accompanied by significant increases in VEGFA and GLUT3 and a decrease in PlGF mRNAs. These effects were suppressed by PD169316. In conclusion, our data suggest that first trimester placental tissues are not energetically compromised, and that HIF-1α is unlikely to play an appreciable role in regulating transcriptional activity under steady-state conditions in vivo. However, the pathway may be activated by stress conditions. PMID:25391298

  14. Short-term human chorionic gonadotropin-induced testosterone rise does not modify leptin levels in eugonadal men.

    PubMed

    Panidis, D; Koliakos, G; Kourtis, A; Rousso, D; Mavromatidis, G; Triantos, A; Kalahanis, I

    2002-02-01

    The aim of this study was to monitor serum leptin concentrations after altering the levels of testosterone, by intramuscular administration of human chorionic gonadotropin (hCG), in eugonadal men. A 7-day monitoring of hCG, testosterone and leptin levels was performed after intramuscular administration of a dose of 5000 IU hCG in these men. Thirty fertile men aged 23-38 years were studied. In addition, 30 women aged 18-34 years with normal ovulatory cycles were studied, to verify reports of sexual dimorphism in serum leptin levels. These 60 individuals were divided into four groups, according to their sex and body mass index (BMI) values. In men, blood samples were collected at 09.00, after an overnight fast, for the determination of hCG, testosterone and leptin levels, and, immediately thereafter, a dose of 5000 IU hCG was administered intramuscularly. Further blood samples were collected at 24-h intervals for a period of 7 days for determination of the same hormones. In women, blood samples were collected only once, at 09.00, after an overnight fast between the 3rd and the 6th day of the menstrual cycle, for determination of serum estradiol and leptin levels. Our results showed that the mean value of leptin in thin men and women was significantly lower than that in obese men and women, respectively. The mean value of leptin in thin women was significantly higher than that in obese men. Serum leptin concentrations decreased significantly, 168 h after short-term hCG administration. There was a significant positive correlation between BMI values and serum leptin concentrations, in both men and women. Our results support the view that hCG administration in eugonadal men does not influence serum leptin levels. Moreover, a short-term increase of serum testosterone levels, after one dose of hCG, is not sufficient to affect and modify leptin secretion mechanisms in vivo. PMID:11915586

  15. Dehydrated Human Amnion/Chorion Membrane as Adjunctive Therapy in the Multidisciplinary Treatment of Pyoderma Gangrenosum: A Case Report.

    PubMed

    Snyder, Robert J; Ead, Joey; Glick, Brad; Cuffy, Cherlson

    2015-09-01

    Pyoderma gangrenosum (PG) is an uncommon chronic and progressive skin disorder that can lead to severe tissue necrosis, pathergy, horrendous pain, and disfigurement if not properly and promptly diagnosed and treated. Systemic treatment traditionally consists of long-term immunosuppression. Topical care of the painful wound often represents a clinical challenge. A 77-year-old woman with multiple comorbidities including venous insufficiency and diabetes mellitus was diagnosed through exclusion with refractory, painful PG. She was managed for 3 months by a multidisciplinary team comprised of an internist, 2 dermatologists, and a podiatric wound care specialist using immunosuppressive therapy, several local wound care modalities, and supportive bandages. During that time, severe wound pain continued unabated and the affected area changed from 3 separate wounds measuring 1.4 cm x 1.0 cm x .01 cm, 1.2 cm x 0.5 cm x 0.1 cm, and 0.6 cm x 0.5 cm x 0.1 cm to 1 wound measuring 8.0 cm x 10.3 cm x 0.1 cm. At that time, dehydrated human amnion/chorion membrane (dHACM) allograft, previously reported to facilitate healing venous leg and diabetic foot ulcers, was incorporated into the treatment plan. The patient reported wound pain decreased from 10 out of 10 to 5 out of 10 within hours following application. At the 4 day follow-up visit, she reported no pain; after 1 week, the wound decreased 6.4 cm x 9.4 cm x 0.1 cm in size and after 2 months (3 applications) the wound had reduced in area from 103 cm2 to 57.96 cm2 (reduced by more than half [56%]). In this patient, following the application of dHACM as an adjunct to immunosuppressive therapy, pain receded and wound healing commenced. Additional controlled studies are needed to ascertain the generalizability of this observation. PMID:26367481

  16. Insulin stimulates synthesis and release of human chorionic gonadotropin by choriocarcinoma cell lines

    SciTech Connect

    Ren, S.G.; Braunstein, G.D. )

    1991-03-01

    Recent studies have shown that insulin regulates placental lactogen, progesterone, and estrogen production from human trophoblast cells. This study was performed to examine whether insulin also regulates the production of hCG by this type of cell. After 24-36 h of preincubation, JEG-3 and JAR cells (2-3 x 10(5) cells/ml.well) or human term trophoblast cells (1 x 10(6) cells/ml.well) were exposed to the test hormone in serum-free Dulbecco's Modified Eagle's Medium for 24-96 h. Secretion of hCG from JEG-3 cells was stimulated by human insulin, human proinsulin, or porcine insulin in a dose-dependent manner, with lowest effective doses of 6.7, 96, and 53 mg/L, respectively. Time-course studies showed that hCG secretion peaked at 72-96 h with insulin exposure; in contrast, no decernable peak was seen without insulin in serum-free media. Exposure of JEG-3 cells for 24 h to 209 mg/liter insulin stimulated hCG synthesis, with 40 +/- 3% more immunoreactive intracellular hCG (P less than 0.05). Cells grown in the presence of insulin and (35S)methionine had 47 +/- 21% more labeled intracellular hCG and 56 +/- 13% more immunoprecipitable (35S)methionine-hCG secreted into the medium than the control cultures (P less than 0.05). During this time period, human placental lactogen release and total trichloroacetice acid-precipitable (35S)methionine protein were not increased. The insulin-induced stimulation of hCG synthesis was inhibited by cycloheximide. Additionally, insulin did not significantly affect total intracellular protein during 24-96 h of incubation. Insulin also increased hCG release from JAR cells, but not from human term trophoblast cells. A mouse monoclonal antibody to the IGF-I receptor inhibited the stimulation of insulin in JEG-3 cells.

  17. Analysis of cell-cell junctions in human amnion and chorionic plate affected by chorioamnionitis.

    PubMed

    Licini, Caterina; Tossetta, Giovanni; Avellini, Chiara; Ciarmela, Pasquapina; Lorenzi, Teresa; Toti, Paolo; Gesuita, Rosaria; Voltolini, Chiara; Petraglia, Felice; Castellucci, Mario; Marzioni, Daniela

    2016-07-01

    Chorioamnionitis is an acute inflammatory reaction associated with the premature rupture of the fetal membranes. It is caused mainly by invasion of bacteria from the vaginal tract that can penetrate the intact membranes and invade the amnion cavity and the decidua. Tight junctions (TJs) and adherent junctions (AJs) are intercellular junctions crucial for epithelia adhesion and permeability regulation in a wide variety of tissues and organs. Our aim is to investigate if TJ and AJ molecules are involved in human chorioamnionitis. We studied the protein expression (by immunohistochemistry and western blotting) and the mRNA levels (by RT-PCR) of some junction proteins such as Zonula Occludens-1 (ZO-1), occludin, VE-cadherin and β-catenin in fetal membranes from women with chorioamnionitis compared to those membranes derived from idiopathic pregnancies. Western blotting and immunohistochemical data established that occludin expression was decreased in amnion with chorioamnionitis compared to amnion from idiopathic pregnancies. Samples tested for ZO-1, VE-cadherin and β-catenin (proteins and mRNAs) showed no differences between idiopathic and pathological membranes. One of the most relevant results is the decrease of occludin in membranes with chorioamnionitis. Since we have previously demonstrated that some cytokines, particularly elevated in the chorioamnionitis, cause the disruption of TJs in placental villi, we suggest that the decrease of occludin in amnion may be the first change that leads to the rupture of the amniotic membrane in this pathology. PMID:26739007

  18. A unique case of growth hormone and human chorionic gonadotropin treatment in a 45,X male with Y: autosome translocation and literature review.

    PubMed

    Mareri, Arianna; Iezzi, MariaLaura; Salvatore, Alessia; Ligas, Claudio; D'Alessandro, Elvira

    2016-07-01

    Maleness associated with a 45,X karyotype is a rare condition in childhood. It is usually diagnosed in adult age because of infertility. We report a unique case of an unbalanced translocation t(Y;21) in a 14-year-old boy with 45,X karyotype referred because of short stature, thin habitus and puberty delay. Hormone analysis showed low serum levels of basal testosterone, insulin-like growth factor (IGF-I) and gonadotrophins. Diagnosis of GH deficiency and puberty delay were made. He was treated with human chorionic gonadotropin (hCG) and GH therapy, respectively, for 6 and 24 months. PMID:27054600

  19. Isolation and characterization of human placental trophoblast subpopulations from first-trimester chorionic villi.

    PubMed Central

    Aboagye-Mathiesen, G; Laugesen, J; Zdravkovic, M; Ebbesen, P

    1996-01-01

    A method for the simultaneous preparation of highly enriched human placental trophoblast populations (villous and extravillous) from first-trimester placental villi (5 to 12 weeks) by using sequential trypsinization, percoll gradient centrifugation, and negative selection with anti-CD9 immunomagnetic separation is described. The purification method resulted in the isolation of four distinct trophoblast populations identified on the basis of morphology and phenotyping: (i) mononuclear villous cytotrophoblast cells which, through differentiation, become committed to syncytium formation; (ii) an extravillous trophoblast population which appeared as a "crazy pavement" and, with subsequent subculturing, differentiated morphologically to mononuclear cells; (iii) an extravillous trophoblast fraction which fused to form multinucleated trophoblast giant cells; and (iv) floating intermediate extravillous trophoblast cells which fused together to form cell clumps and which further differentiated to a mononuclear anchoring intermediate extravillous trophoblast. Short-term cultures of the freshly isolated cell fractions consisted of heterogeneous trophoblasts at different differentiation stages as determined by their varied biochemical and morphological properties. All the isolated trophoblast populations expressed the cytokeratin intermediate filament and the epithelium-specific cell-cell adhesion molecule E-cadherin. The isolated villous trophoblasts in culture expressed integrins alpha 6 and beta 4 and reduced levels of beta 1 subunits, whereas the proliferating extravillous trophoblast cultures expressed alpha 1, alpha 3, and alpha 5 and high levels of beta 1 integrin subunits, vitronectin receptor (alpha V beta 3/beta 5), and major histocompatibility complex class 1 molecules. Furthermore, the isolated trophoblast populations secreted metalloproteases (such as type IV collagenases [mainly 72- and 92-kDa enzymes, i.e., gelatinases A and B]) and urokinase plasminogen

  20. Human chorionic gonadotropin and its relation to grade, stage and patient survival in ovarian cancer

    PubMed Central

    2012-01-01

    Background An influence of gonadotropins (hCG) on the development of ovarian cancer has been discussed. Therefore, we quantified serum hCG levels in patients with benign and malignant ovarian tumors and the hCG expression in ovarian cancer tissue in order to analyze its relation to grade, stage, gonadotropin receptor (LH-R, FSH-R) expression and survival in ovarian cancer patients. Methods Patients diagnosed and treated for ovarian tumors from 1990 to 2002 were included. Patient characteristics, histology including histological subtype, tumor stage, grading and follow-up data were available. Serum hCG concentration measurement was performed with ELISA technology, hCG tissue expression determined by immunohistochemistry. Results HCG-positive sera were found in 26.7% of patients with benign and 67% of patients with malignant ovarian tumors. In addition, significantly higher hCG serum concentrations were observed in patients with malignant compared to benign ovarian tumors (p = 0.000). Ovarian cancer tissue was positive for hCG expression in 68%. We identified significant differences in hCG tissue expression related to tumor grade (p = 0.022) but no differences with regard to the histological subtype. In addition, mucinous ovarian carcinomas showed a significantly increased hCG expression at FIGO stage III compared to stage I (p = 0.018). We also found a positive correlation of hCG expression to LH-R expression, but not to FSH-R expression. There was no significant correlation between tissue hCG expression and overall ovarian cancer patient survival, but subgroup analysis revealed an increased 5-year survival in LH-R positive/FSH-R negative and hCG positive tumors (hCG positive 75.0% vs. hCG negative 50.5%). Conclusions Serum human gonadotropin levels differ in patients with benign and malignant ovarian tumors. HCG is often expressed in ovarian cancer tissue with a certain variable relation to grade and stage. HCG expression correlates with LH-R expression in ovarian

  1. Chorionic villus sampling

    MedlinePlus

    Chorionic villus sampling (CVS) is a test some pregnant women have to screen their baby for genetic problems. ... CVS can be done through the cervix (transcervical) or through the belly (transabdominal). Miscarriage rates are slightly ...

  2. Ovarian and Adrenal Androgens and Their Link to High Human Chorionic Gonadotropin Levels: A Prospective Controlled Study

    PubMed Central

    Rodríguez-Gutiérrez, René; Villarreal-Pérez, Jesús Zacarías; Morales-Martinez, Felipe Arturo; Rodríguez-Guajardo, René; González-Saldivar, Gloria; Mancillas-Adame, Leonardo G.; Alvarez-Villalobos, Neri Alejandro; Lavalle-Gonzalez, Fernando Javier; González-González, José Gerardo

    2014-01-01

    Background. Although the association between human chorionic gonadotropin (hCG) and hyperandrogenism was identified more than 40 years ago, relevant questions remain unanswered. Design and Methods. We conducted a prospective, longitudinal, and controlled study in 23 women with a diagnosis of a complete hydatidiform mole (HM). Results. All participants completed the study. Before HM evacuation mean hCG was markedly higher in the cases than in the control group (P ≤ 0.001). Free testosterone (T) and dehydroepiandrosterone sulfate (DHEA-S) were found to be higher in the cases (2.78 ± 1.24 pg/mL and 231.50 ± 127.20 μ/dL) when compared to the control group (1.50 ± 0.75 pg/mL and 133.59 ± 60.69 μ/dL) (P = 0.0001 and 0.001), respectively. There was a strong correlation between hCG and free T/total T/DHEA-S concentrations (r = 0.78; P ≤ 0.001, r = 0.74;  P ≤ 0.001, and r = 0.71;  P ≤ 0.001), respectively. In the cases group 48 hours after HM evacuation, hCG levels were found to be significantly lower when compared to initial levels (P = 0.001) and free T and DHEA-S declined significantly (P = 0.0002 and 0.009). Conclusion. Before uterus evacuation, hCG, free T, and DHEA-S levels were significantly higher when compared with controls finding a strong correlation between hCG and free T/DHEA-S levels. Forty-eight hours after HM treatment hCG levels declined and the difference was lost. A novel finding of our study is that in cases, besides free T, DHEA-S was also found to be significantly higher and both the ovaries and adrenal glands appear to be the sites of this androgen overproduction. PMID:25505909

  3. Optimisation of an oviposition protocol employing human chorionic and pregnant mare serum gonadotropins in the Barred Frog Mixophyes fasciolatus (Myobatrachidae)

    PubMed Central

    2012-01-01

    Background Protocols for the hormonal induction of ovulation and oviposition are essential tools for managing threatened amphibians with assisted reproduction, but responses vary greatly between species and even broad taxon groups. Consequently, it is necessary to assess effectiveness of such protocols in representative species when new taxa become targets for induction. The threatened genus Mixophyes (family Myobatrachidae) has amongst the highest proportion of endangered species of all the Australian amphibians. This study developed and optimised the induction of oviposition in a non-threatened member of this taxon, the great barred frog (Mixophyes fasciolatus). Methods Gravid female M. fasciolatus were induced to oviposit on one or more occasions by administration of human chorionic gonadotropin (hCG) with or without priming with pregnant mare serum gonadotropin (PMSG). Treatments involved variations in hormone doses and combinations (administered via injection into the dorsal lymph sacs), and timing of administration. Pituitary homogenates from an unrelated bufonid species (Rhinella marina) were also examined with hCG. Results When injected alone, hCG (900 to 1400 IU) induced oviposition. However, priming with two time dependent doses of PMSG (50 IU, 25 IU) increased responses, with lower doses of hCG (200 IU). Priming increased response rates in females from around 30% (hCG alone) to more than 50% (p = 0.035), and up to 67%. Increasing the interval between the first PMSG dose and first hCG dose from 3 to 6 days also produced significant improvement (p<0.001). Heterologous pituitary extracts administered with hCG were no more effective than hCG alone (p = 0.628). Conclusions This study found that M. fasciolatus is amongst the few amphibian species (including Xenopus (Silurana) and some bufonids) that respond well to the induction of ovulation utilising mammalian gonadotropins (hCG). The optimal protocol for M. fasciolatus involved two priming doses of

  4. Human chorionic gonadotrophin in early gestation induces growth of estrogenic ovarian follicles and improves primiparous sow fertility during summer.

    PubMed

    Seyfang, Jemma; Langendijk, P; Chen, T Y; Bouwman, E; Kirkwood, R N

    2016-09-01

    Reduced summer farrowing rates may be due to inadequate corpora luteal (CL) support. Porcine CL become dependent on LH from 12 d of pregnancy and the embryonic estrogen signal for maternal recognition of pregnancy (MRP) is initiated at about 11-12 d after insemination. We hypothesised that injection of the LH analogue human chorionic gonadotropin (hCG) would induce growth of estrogenic follicles and, by mimicking the signal for MRP and stimulating progesterone secretion, increase primiparous sow fertility. In Experiment 1, during a 28 d lactation 53 mixed parity sows were full-fed either throughout lactation (n=16) or until 18 d and then feed restricted during the last 10 d of lactation (n=36). At 12 d after mating restrict-fed sows were injected with 1000IU hCG (n=17) or were not injected (n=19); the full-fed sows acted as non-treated positive controls. Transrectal ovarian ultrasound exams were performed on days 12, 16, 20, 24, and 28; blood samples were obtained on days 12, 14, and 15 for estradiol and progesterone assay. For Experiment 2, during the summer months primiparous sows received 1000IU hCG 12 d after mating (n=28) or were non-injected controls (n=27). Pregnancy status was determined at 28 d and sows allowed to go to term to determine farrowing rates and litter sizes. In Experiment 1, injection of hCG increased (P<0.001) follicle diameter and serum concentrations of estradiol (P<0.01) and progesterone (P<0.05). There were no effects of lactation feeding level on wean-estrus interval, farrowing rate or subsequent litter size. In Experiment 2, hCG injection was associated with a higher pregnancy rate (P<0.05) and farrowing rate (P<0.08). There was no effect on litter size. These data confirm that hCG stimulates growth of estrogenic follicles and CL function, and improves primiparous sow fertility during the summer months. PMID:27397793

  5. Expressions of candidate molecules in the human fallopian tube and chorionic villi of tubal pregnancy exposed to levonorgestrel emergency contraception

    PubMed Central

    2013-01-01

    Background Cases of ectopic pregnancy (EP) following levonorgestrel (LNG) emergency contraception (EC) failure were reported, however, the effects of LNG on tubal microenvironment or chorionic villi in EP have not yet been documented. Methods Fifty-five women with tubal pregnancy were divided into two groups according to whether LNG-EC was administrated during the cycle of conception. The serum concentrations of beta-hCG, E2 and P were measured. The mRNA and protein expressions of estrogen and progesterone receptors, leukemia inhibitory factor, vascular endothelial growth factor, inducible nitric oxide synthase, and endocannabinoid receptor - CB1 in the ectopic implantation site and chorionic villi were examined. Results Compared to those unexposed to LNG-EC, women with tubal pregnancy exposed to LNG-EC during the cycle of conception had no statistically significances in the serum concentrations of beta-hCG, E2 P, nor in the pathological types of tubal pregnancy or the expressions of ER-alpha, PR, LIF, VEGF, iNOS and CB1. Conclusions The expressions of candidate molecules in the fallopian tube and chorionic villi were not altered by exposure to LNG-EC. A routine therapy with no additional intervention might thus be applied to tubal pregnancy exposed to LNG-EC. PMID:23687977

  6. Magnetic particle-linked anti hCG β antibody for immunoassay of human chorionic gonadotropin (hCG), potential application to early pregnancy diagnosis.

    PubMed

    Kuo, Hsiao-Ting; Yeh, Jay Z; Jiang, Chi-Ming; Wu, Ming-Chang

    2012-07-31

    The objective of this study was to develop a magnetic particle-linked monoclonal antibody to hCG β for immunosorbent assay of human chorionic gonadotropin (hCG) with improved detection sensitivity. Monoclonal antibody against hCG β was found to be optimally cross-linked to the superparamagnetic particles (SPIO) using EDC and NHS as cross-linking reagents. This superparamagnetic particle-linked monoclonal antibody was able to concentrate hCG from a tested solution for further ELISA assay using horse radish peroxidase-labeled monoclonal antibody against hCG β. This hybrid technique had greatly decreased the detection limit to 0.1 mIU/mL, making an early detection of pregnancy possible. With an improved sensitivity and simple operation, the magnetic particle-linked anti hCG β antibody for immunoassay of human chorionic gonadotropin (hCG) has a great potential to supersede the traditional ELISA for pregnancy diagnosis. PMID:22542932

  7. First-trimester maternal serum human chorionic gonadotrophin as a marker for fetal chromosomal disorders. The Dutch Working Party on Prenatal Diagnosis.

    PubMed

    Van Lith, J M

    1992-06-01

    The Dutch Working Party on Prenatal Diagnosis has initiated a study on the possibilities of first-trimester screening for fetal chromosomal disorders. We report on maternal serum human chorionic gonadotrophin (MS-hCG) measurements in 1348 pregnancies with a chromosomally normal fetus and 53 pregnancies with a chromosomally abnormal fetus. The median MS-hCG concentration in 24 pregnancies with Down's syndrome was 1.19 multiples of the normal median (MoM). The MS-hCG distributions in normal and Down's syndrome pregnancies did not differ significantly (t-test: t = 1.945, p greater than 0.05). We also found no difference between normal pregnancies and pregnancies with other chromosomal disorders (six cases of trisomy 18, MoM = 0.80; four cases of sex chromosome abnormality, MoM = 1.01; 17 cases of chromosomal mosaicism in chorionic villi, MoM = 1.11). Selecting an upper limit at the 90th centile could detect 25 per cent of pregnancies with Down's syndrome. We conclude that, in the first trimester, MS-hCG as a screening factor for Down's syndrome is of minor value. However, MS-hCG could be a useful factor in a first-trimester screening programme based on a combination of markers. PMID:1387477

  8. Effect of treatment with human chorionic gonadotropin on day 5 after timed artificial insemination on fertility of lactating dairy cows.

    PubMed

    Nascimento, A B; Bender, R W; Souza, A H; Ayres, H; Araujo, R R; Guenther, J N; Sartori, R; Wiltbank, M C

    2013-05-01

    Reproductive management programs that synchronize ovulation can ovulate a smaller than normal follicle, potentially resulting in inadequate progesterone (P4) concentrations after artificial insemination (AI). Ovulation of the dominant follicle of the first follicular wave with human chorionic gonadotropin (hCG) treatment can produce an accessory corpus luteum and increase circulating P4 concentrations. This manuscript reports the results of 2 separate analyses that evaluated the effect of hCG treatment post-AI on fertility in lactating dairy cows. The first study used meta-analysis to combine the results from 10 different published studies that used hCG treatment on d 4 to 9 post-AI in lactating dairy cows. Overall, pregnancies per artificial insemination (P/AI) were increased 3.0% by hCG treatment post-AI [34% (752/2,213) vs. 37% (808/2,184); Control vs. hCG-treated, respectively]. The second study was a field research trial in which lactating Holstein cows (n=2,979) from 6 commercial dairy herds were stratified by parity and breeding number and then randomly assigned to one of 2 groups: control (no further treatment, n=1,519) or hCG [Chorulon i.m.: 2,000 IU (in 3 of the herds) or 3,300 IU (in 3 herds); n=1,460] on d 5 after a timed AI (ovulation synchronized with Ovsynch, Presynch-Ovsynch, or Double-Ovsynch). In a subset of cows, the hCG profile and P4 changes were determined. Treatment with hCG increased P4 (4.3 vs. 5.3 ng/mL on d 12). Pregnancies per AI were greater in cows treated with hCG (40.8%; 596/1,460) than control (37.3%; 566/1,519) cows. Interestingly, an interaction among treatment and parity was observed; primiparous cows had greater P/AI after hCG (49.7%; 266/535) than controls (39.5%; 215/544). In contrast, older cows receiving hCG (35.7%; 330/925) had similar P/AI to controls (36.0%; 351/975).Thus, targeted use of hCG on d 5 after TAI enhances fertility about 3.0% (based on meta-analysis) to 3.5% (based on our field trial). Surprisingly, this

  9. Quantitative photoacoustic elastography in humans

    NASA Astrophysics Data System (ADS)

    Hai, Pengfei; Zhou, Yong; Gong, Lei; Wang, Lihong V.

    2016-06-01

    We report quantitative photoacoustic elastography (QPAE) capable of measuring Young's modulus of biological tissue in vivo in humans. By combining conventional PAE with a stress sensor having known stress-strain behavior, QPAE can simultaneously measure strain and stress, from which Young's modulus is calculated. We first demonstrate the feasibility of QPAE in agar phantoms with different concentrations. The measured Young's modulus values fit well with both the empirical expectation based on the agar concentrations and those measured in an independent standard compression test. Next, QPAE was applied to quantify the Young's modulus of skeletal muscle in vivo in humans, showing a linear relationship between muscle stiffness and loading. The results demonstrated the capability of QPAE to assess the absolute elasticity of biological tissue noninvasively in vivo in humans, indicating its potential for tissue biomechanics studies and clinical applications.

  10. Human Chorionic Gonadotropin (hCG) Prevents the Transformed Phenotypes Induced by 17 β-estradiol in Human Breast Epithelial Cells

    PubMed Central

    Kocdor, Hilal; Kocdor, Mehmet A.; Russo, Jose; Snider, Kara E.; Vanegas, Johana E.; Russo, Irma H.; Fernandez, Sandra V.

    2009-01-01

    Human chorionic gonadotropin (hCG), a hormone produced during pregnancy, can elicit life-long refractoriness to carcinogenesis by differentiation of the breast epithelium. Human breast epithelial cells MCF-10F form tubules in collagen, mimicking the normal ductules, We have shown that 17 β-estradiol (E2) alter the ductulogenic pattern of these cells. The effect of the recombinant hCG (rhCG) in vitro was evaluated on the transformation of MCF-10F induced by E2. MCF-10F cells were treated with 70nM E2 alone or in combination with 50 IU/ml rhCG during 2 weeks, while the controls were treated with DMSO (the solvent in which E2 was dissolved) or rhCG alone. At the end of treatment, the cells were plated in type I collagen matrix (3D-cultures) for detecting 2 main phenotypes of cell transformation, namely the loss of ductulogenic capacity and the formation of solid masses. Although E2 significantly increased solid mass formation, this effect was prevented when MCF-10F cells were treated with E2 in combination with rhCG. Furthermore, E2 increased the main duct width (p<0.001), and caused a disruption of the luminal architecture, whereas rhCG increased the length of the tubules (p<0.001) and produced tertiary branching. In conclusion, rhCG was able to abrogate the transforming abilities of estradiol, and had the differentiating property by increasing the branching of the tubules formed by breast epithelial cells in collagen. These results further support our hypothesis, known as the terminal differentiation hypothesis of breast cancer prevention, that predicts that hCG treatment results in protection from tumorigenic changes by the loss of susceptible stem cells 1 through a differentiation to refractory stem cells 2 and increase differentiation of the mammary gland. PMID:19647089

  11. Effect of leptin on progesterone, human chorionic gonadotropin, and interleukin-6 secretion by human term trophoblast cells in culture.

    PubMed

    Cameo, Paula; Bischof, Paul; Calvo, Juan Carlos

    2003-02-01

    Leptin, the 16-kDa protein product of the obese gene, was originally seen as an adipocyte-derived signaling molecule. Recently, it has been suggested to be involved in some functions during pregnancy, particularly in the placenta. In the present study, we investigated the role of leptin in the secretion of hCG, progesterone, and interleukin-6 (IL-6) by human term trophoblast cells in culture. Placentae were obtained from cesarean sections following uncomplicated pregnancies and used immediately after delivery. Leptin, hCG, progesterone, and IL-6 were measured by ELISA, RIA, and immunoradiometric assay in the cultured media of trophoblast cells cultured for 48 and 96 h. Leptin mRNA expression in these cultures was determined by reverse transcription-polymerase chain reaction. Recombinant human leptin added to primary cultures of human term placental trophoblast cells showed a stimulatory effect on hCG and IL-6 secretion and an inhibitory effect on progesterone secretion. Primary cultures of term trophoblast cells expressed leptin mRNA. All these findings suggest a role for leptin in human placental endocrine function. PMID:12533410

  12. The Prenatal Environment in Twin Studies: A Review on Chorionicity.

    PubMed

    Marceau, Kristine; McMaster, Minni T B; Smith, Taylor F; Daams, Joost G; van Beijsterveldt, Catharina E M; Boomsma, Dorret I; Knopik, Valerie S

    2016-05-01

    A literature search was conducted to identify articles examining the association of chorionicity (e.g., whether twins share a single chorion and thus placenta or have separate chorions/placentas) and genetics, psychiatry/behavior, and neurological manifestations in humans twins and higher-order multiples. The main aim was to assess how frequently chorionicity has been examined in relation to heritability estimates, and to assess which phenotypes may be most sensitive to, or affected by, bias in heritability estimates because of chorionicity. Consistent with the theory that some chorionicity effects could lead to overestimation and others to underestimation of heritability, there were instances of each across the many phenotypes reviewed. However, firm conclusions should not be drawn since some of the outcomes were only examined in one or few studies and often sample sizes were small. While the evidence for bias due to chorionicity was mixed or null for many outcomes, results do, however, consistently suggest that heritability estimates are underestimated for measures of birth weight and early growth when chorionicity is not taken into account. PMID:26944881

  13. The tumor suppressor p53 induces expression of the pregnancy-supporting human chorionic gonadotropin (hCG) CGB7 gene

    PubMed Central

    Sohr, Sindy

    2011-01-01

    Successful pregnancy requires a functionally normal blastocyst encountering a receptive maternal endometrium. Interestingly, the cell cycle regulator and tumor suppressor p53 has been reported to support reproduction in mice by regulating the expression of the leukemia inhibitory factor gene in the maternal endometrium. However, in humans the hormonal system orchestrating successful pregnancy is considerably different from rodents. Particularly, the primatespecific dimeric glycoprotein hormone human chorionic gonadotropin (hCG) is essential for blastocyst implantation and maintenance of early human pregnancy. Here we provide evidence that p53 selectively induces expression of the hCGβ7 (CGB7) gene. None of the other CGB genes was found to be regulated by p53. We show that expression of the CGB7 gene is upregulated upon p53 induction in human HFF, HCT116 and DLD1 cells as well as in cell preparations enriched in human primary first-trimester trophoblasts. The increase in CGB7 levels upon doxorubicin treatment is lost after siRNA-directed knockdown of p53. Furthermore, we describe CGB7 as a direct transcriptional target gene of p53 by identifying a p53-responsive element in the CGB7 promoter using reporter assays, electrophoretic mobility shift assays and chromatin immunoprecipitations. With these results we provide a new link between p53 transcriptional activity and human reproduction. PMID:22032922

  14. Contribution of aquaporin 9 and multidrug resistance-associated protein 2 to differential sensitivity to arsenite between primary cultured chorion and amnion cells prepared from human fetal membranes

    SciTech Connect

    Yoshino, Yuta; Yuan, Bo; Kaise, Toshikazu; Takeichi, Makoto; Tanaka, Sachiko; Hirano, Toshihiko; Kroetz, Deanna L.; Toyoda, Hiroo

    2011-12-15

    Arsenic trioxide (arsenite, As{sup III}) has shown a remarkable clinical efficacy, whereas its side effects are still a serious concern. Therefore, it is critical to understand the effects of As{sup III} on human-derived normal cells for revealing the mechanisms underlying these side effects. We examined the effects of As{sup III} on primary cultured chorion (C) and amnion (A) cells prepared from human fetal membranes. A significant dose-dependent As{sup III}-mediated cytotoxicity was observed in the C-cells accompanied with an increase of lactate dehydrogenase (LDH) release. Higher concentrations of As{sup III} were required for the A-cells to show cytotoxicity and LDH release, suggesting that the C-cells were more sensitive to As{sup III} than the A-cells. The expression levels of aquaporin 9 (AQP9) were approximately 2 times higher in the C-cells than those in the A-cells. Both intracellular arsenic accumulation and its cytotoxicity in the C-cells were significantly abrogated by sorbitol, a competitive AQP9 inhibitor, in a dose-dependent manner. The protein expression levels of multidrug resistance-associated protein (MRP) 2 were downregulated by As{sup III} in the C-cells, but not in the A-cells. No significant differences in the expression levels of MRP1 were observed between C- and A-cells. The protein expression of P-glycoprotein (P-gp) was hardly detected in both cells, although a detectable amount of its mRNA was observed. Cyclosporine A, a broad-spectrum inhibitor for ABC transporters, and MK571, a MRP inhibitor, but not PGP-4008, a P-gp specific inhibitor, potently sensitized both cells to As{sup III}-mediated cytotoxicity. These results suggest that AQP9 and MRP2 are involved in controlling arsenic accumulation in these normal cells, which then contribute to differential sensitivity to As{sup III} cytotoxicity between these cells. -- Highlights: Black-Right-Pointing-Pointer Examination of effect of As{sup III} on primary cultured chorion (C) and amnion

  15. Intrauterine administration of recombinant human chorionic gonadotropin before embryo transfer on outcome of in vitro fertilization/ intracytoplasmic sperm injection: A randomized clinical trial

    PubMed Central

    Zarei, Afsoon; Parsanezhad, Mohammad Ebrahim; Younesi, Masoumeh; Alborzi, Saeed; Zolghadri, Jaleh; Samsami, Alamtaj; Amooee, Sedigheh; Aramesh, Shahintaj

    2014-01-01

    Background: The direct effect of hCG on the human endometrium was studied several times. Objective: The objectives of this study were to evaluate the effectiveness of intrauterine injection of recombinant human chorionic gonadotropin (rhCG) before embryo transfer (ET). Materials and Methods: In this randomized placebo-controlled clinical trial, a total number of 182 infertile patients undergoing their first in vitro fertilization/ intracytoplasmic sperm injection (IVF-ICSI) cycles were randomly assigned to receive 250μg intrauterine rhCG (n=84) or placebo (n=98) before ET. The implantation and pregnancy rates were compared between groups. Results: Patients who received intrauterine rhCG before ET had significantly higher implantation (36.9% vs. 22.4%; p=0.035), clinical pregnancy rates (34.5% vs. 20.4%; p=0.044) and ongoing pregnancy rate (32.1% vs. 18.4%; p=0.032) when compared to those who received placebo. The abortion (2.4% vs. 2.0%; p=0.929) and ectopic pregnancy rates (1.2% vs. 1.0%; p=0.976) were comparable between groups of rhCG and placebo, respectively. Conclusion: Intrauterine injection of 250μg of rhCG before ET significantly improves the implantation and pregnancy rates in IVF/ICSI cycles. Registration ID in IRCT: IRCT2012121711790N1 This article extracted from fellowship course thesis. (Masoumeh Younesi) PMID:24799855

  16. Liver and chorion cytochemistry.

    PubMed

    Roels, F; De Prest, B; De Pestel, G

    1995-01-01

    Microscopic visualization of peroxisomes in chorionic villus cytotrophoblast and in biopsy and autopsy samples of liver and kidney, the presence of enlarged liver macrophages containing lipid droplets insoluble in acetone and n-hexane as well as polarizing inclusions formed by stacks of trilamellar sheets are of diagnostic value in peroxisomal disorders. Methods are presented for evaluating these structures by light microscopy; trilamellar inclusions are only detected by electron microscopy. Macrophage features are preserved in archival paraffin blocks. In adrenal cortex, insoluble lipid, polarizing inclusions and trilamellar structures should be looked for. The stains are easily reproducible, and all reagents are commercially available. PMID:9053549

  17. The effect of human chorionic gonadotropin (HCG) in the treatment of obesity by means of the Simeons therapy: a criteria-based meta-analysis.

    PubMed Central

    Lijesen, G K; Theeuwen, I; Assendelft, W J; Van Der Wal, G

    1995-01-01

    1. A meta-analysis was conducted to assess if there is scientific ground for the use of human chorionic gonadotropin (HCG) as adjunctive therapy in the treatment of obesity. 2. Published papers relating to eight controlled and 16 uncontrolled trials that measured the effect of HCG in the treatment of obesity were traced by computer-aided search and citation tracking. 3. The trials were scored for the quality of the methods (based on four main categories: study population, interventions, measurement of effect, and data presentation and analysis) and the main conclusion of author(s) with regard to weight-loss, fat-redistribution, hunger, and feeling of well-being. 4. Methodological scores ranged from 16 to 73 points (maximum score 100), suggesting that most studies were of poor methodological quality. Of the 12 studies scoring 50 or more points, one reported that HCG was a useful adjunct. The studies scoring 50 or more points were all controlled. 5. We conclude that there is no scientific evidence that HCG is effective in the treatment of obesity; it does not bring about weight-loss of fat-redistribution, nor does it reduce hunger or induce a feeling of well-being. PMID:8527285

  18. Inhibition of in vitro human chorionic gonadotropin-stimulated testosterone production in testis and of ovulation in the rat by charcoal-treated rat testicular extract

    SciTech Connect

    de Bellabarba, G.A.; Bishop, W.; Rojas, F.J.

    1984-01-16

    Previously, the authors described the presence of a factor obtained from rat testis that was found to inhibit human chorionic gonadotropin (hCG) binding to gonadal receptors. In the present study, similarly prepared testicular extract was tested for its effects on in vitro hCG-stimulated testosterone production by isolated testis interstitial cells and for its effect on spontaneous ovulation in the rat. Incubation of interstitial cells with charcoal-treated extract significantly inhibited the steroidogenic response to hCG in a dose-related manner. This inhibition was also apparent after heating the extract for 10 min at 100/sup 0/C. A single i.p. injection of testicular extract inhibited spontaneous ovulation in the rat. This effect was also observed after heating the extract for 10 min at 100/sup 0/C. It is concluded that the aqueous testicular extract contains a factor able to antagonize the physiological events mediated by luteinizing hormone (LH)/hCG, and that this factor is consistent with the presence of an LH/hCG-binding inhibitory activity in rat testis.

  19. Early pregnancy human chorionic gonadotropin (hCG) isoforms measured by an immunometric assay for choriocarcinoma-like hCG.

    PubMed

    Kovalevskaya, G; Birken, S; Kakuma, T; O'Connor, J F

    1999-04-01

    Human chorionic gonadotropin (hCG) exhibits molecular heterogeneity in both its protein and carbohydrate moieties. This communication describes changes in hCG isoforms detected directly in clinical samples. These isoforms, quantified in blood or urine specimens, show a progression of change throughout normal pregnancy. Early pregnancy produces a type of hCG that resembles, in terms of immunoreactivity, a major form of hCG excreted in choriocarcinoma. The isoforms predominate for the first 5-6 weeks of gestation and then diminish, being replaced with the hCG isoforms which predominate throughout the remainder of pregnancy. The alteration in hCG isoform content occurs in both blood and urine. The progression of isoforms is best delineated by calculating the change in the ratio of the two forms, as many hCG assays either do not detect or fail to discriminate among these isoforms. An analogous pattern of hCG isoforms was observed in patients with in vitro fertilization pregnancies. hCG isolated from the pituitary displayed binding characteristics similar to those of the hCG derived from normal pregnancy urine. The early pregnancy hCG isoforms appear to have a differential expression in normal pregnancy as opposed to pregnancies which will not carry to term, suggesting that a determination of the relative balance of hCG isoforms may have diagnostic application in predicting pregnancy outcome. PMID:10194533

  20. Relationship between progesterone level on the day of human chorionic gonadotropin administration with outcomes of intra-cytoplasmic sperm injection in infertile couples

    PubMed Central

    Hajishafiha, Mahsomeh; Shahbazi, Zahra; Pakniyat, AbdolGhader; Oshnouei, Sima; Kiarang, Nazila

    2015-01-01

    Background: Gonadotropin-releasing hormone agonists or antagonists are used in assisted reproductive technique cycles as premature luteinizing hormone secretion inhibition. Studies have been reported different and contradictory results on the serum progesterone effect on intra-cytoplasmic sperm injection. Objective: The purpose of this study was to evaluate the effect of serum progesterone level on the day of Human chorionic gonadotropin (HCG) administration on the intra-cytoplasmic sperm injection (ICSI) outcome in infertile women. Materials and Methods: 249 infertile couples candidated for ICSI were enrolled in the study. Their serum progesterone level on the day of HCG administration was measured and according to serum level, patients were divided into four groups of less than 0.9, 0.9-1.4, 1.5-1.9, and ≥2 ng/mL. The four groups were compared with each other regarding fertility outcomes. Results: Pregnancy rate was not significantly different among the four groups (p>0.05). Also, there was no significant difference among the groups regarding frequency of abortion and ectopic pregnancy. Conclusion: Serum progesterone level on the day of HCG administration does not have any significant effect on pregnancy outcomes, including abortion, ectopic pregnancy, and pregnancy rate in patients undergoing ICSI treatment. PMID:26494986

  1. Antibodies against the beta-subunit of ovine luteinizing hormone can decrease the clearance of human chorionic gonadotropin in rhesus monkeys.

    PubMed

    Yamamoto, Y; Gunsalus, G L; Sundaram, K; Thau, R B

    1984-06-01

    Contraceptive vaccines based on active immunization against gonadotropic hormones are being investigated in humans and other primates. Immunization against the beta-subunit of ovine luteinizing hormone (oLH beta) reduces fertility in rhesus monkeys by inducing inadequate luteal phases and preventing corpus luteum rescue by rhesus chorionic gonadotropin (rhCG). These effects result from the cross-reactions of the oLH beta-antibodies with rhCG and rhLH. We used human CG (hCG), which also cross-reacts strongly with anti-oLH beta to examine how the circulating oLH beta-antibodies affect the metabolic clearance rates (MCR) of hCG in rhesus monkeys. 125I-hCG was injected into four nonimmunized and seven immunized monkeys and blood was collected at frequent intervals over 7 days. Total and immunoprecipitable radioactivity did not differ significantly, suggesting that the radioactivity in the plasma consisted almost entirely of 125I-hCG. This was confirmed by column chromatography. The MCR (mean +/- SE) was significantly lower (p less than 0.001) in six immunized monkeys (0.35 +/- 0.06 liters/day) as compared to controls (1.19 +/- 0.09 liters/day). The hCG disappearance curve in control monkeys was best described by a two-compartmental system (slow and fast) while an additional third (intermediate) compartment of distribution was typical for immunized animals. The half-lives of hCG for the two exponentials corresponding to the slow and fast components of distribution were not significantly different between the two groups. One immunized monkey had a MCR (1.44 liters/day) that was much greater than the MCR of the other six.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:6507705

  2. Luteal phase support in in-vitro fertilization using gonadotrophin releasing hormone analogue before ovarian stimulation: a prospective randomized study of human chorionic gonadotrophin versus intramuscular progesterone.

    PubMed

    Claman, P; Domingo, M; Leader, A

    1992-04-01

    This study was conducted to compare the endocrine milieu and pregnancy rates in an in-vitro fertilization and embryo transfer (IVF-ET) programme employing a gonadotrophin-releasing hormone agonist (GnRHa) and human menopausal gonadotrophin (HMG) when either human chorionic gonadotrophin (HCG) or progesterone were used for luteal phase support. A total of 121 IVF-ET treatment cycles were prospectively studied. All patients started leuprolide acetate in the midluteal phase and it was continued for at least 10 days. When oestradiol levels were less than 150 pmol/l, HMG was started. When at least three follicles were greater than or equal to 17 mm in diameter, HCG 5000 IU i.m. was given. Oocytes were retrieved using transvaginal ultrasound and embryos were transferred 48 h later. The patients' cycles were prospectively randomized to receive HCG (72 cycles) or progesterone (49 cycles) luteal support. The HCG group received 1500 IU i.m. on days 3, 6 and 9 after the initial trigger. The progesterone group received 12.5 mg i.m. q.d. starting from the day after the HCG trigger. The dose of progesterone was increased to 25 mg i.m. q.d. starting on the day of embryo transfer and continued for 17-21 days. If the patient became pregnant, this dose of progesterone was continued until fetal heart activity was visualized by ultrasound. Mean ages, number of eggs retrieved, embryos transferred, oestradiol levels on the day of the HCG trigger, oestradiol and progesterone at the time of embryo transfer were the same in both groups.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1522190

  3. A prospective, randomised comparative study of weekly versus biweekly application of dehydrated human amnion/chorion membrane allograft in the management of diabetic foot ulcers

    PubMed Central

    Zelen, Charles M; Serena, Thomas E; Snyder, Robert J

    2014-01-01

    The aim of this study is to determine if weekly application of dehydrated human amnion/chorion membrane allograft reduce time to heal more effectively than biweekly application for treatment of diabetic foot ulcers. This was an institutional review board-approved, registered, prospective, randomised, comparative, non-blinded, single-centre clinical trial. Patients with non-infected ulcers of ≥ 4 weeks duration were included for the study. They were randomised to receive weekly or biweekly application of allograft in addition to a non-adherent, moist dressing with compressive wrapping. All wounds were offloaded. The primary study outcome was mean time to healing. Overall, during the 12-week study period, 92·5% (37/40) ulcers completely healed. Mean time to complete healing was 4·1 ± 2·9 versus 2·4 ± 1·8 weeks (P = 0·039) in the biweekly versus weekly groups, respectively. Complete healing occurred in 50% versus 90% by 4 weeks in the biweekly and weekly groups, respectively (P = 0·014). Number of grafts applied to healed wounds was similar at 2·4 ± 1·5 and 2·3 ± 1·8 for biweekly versus weekly groups, respectively (P = 0·841). These results validate previous studies showing that the allograft is an effective treatment for diabetic ulcers and show that wounds treated with weekly application heal more rapidly than with biweekly application. More rapid healing may decrease clinical operational costs and prevent long-term medical complications. PMID:24618401

  4. Evaluation of dual trigger with gonadotropin-releasing hormone agonist and human chorionic gonadotropin in improving oocyte maturity rates: A prospective randomized study

    PubMed Central

    Mahajan, Nalini; Sharma, Shilpa; Arora, Puneet Rana; Gupta, Shalu; Rani, Kumkum; Naidu, Padmaja

    2016-01-01

    BACKGROUND: Mature oocytes are prerequisite for achieving the process of in vitro fertilization. Human chorionic gonadotropin (hCG) is the standard trigger used for stimulating ovulation but is associated with ovarian hyperstimulation syndrome (OHSS). Gonadotropin-releasing hormone agonist trigger achieves oocyte maturation and lowers the incidence of OHSS, but it has limitations of higher pregnancy loss rate and miscarriage rates. Coadministration of both hormones is found to improve the pregnancy rates and the number of mature oocytes retrieved. We aimed to assess if the dual trigger is better than the conventional hCG in triggering oocyte maturation. METHODOLOGY: The study included 76 female patients aged 24–43 years who were randomly divided into two groups with 38 patients in each arm. The study included patients with antimullerian hormone (AMH) <4 ng/ml, antral follicle counts (AFCs)/ovary <12. The study excluded high responders-AMH >4 ng/ml and AFC/ovary >12 to avoid OHSS risk with hCG trigger. RESULTS: The study showed statistically insignificant differences between dual group versus hCG group in terms of the number of oocytes retrieved (10.0 ± 5.6 vs. 8.7 ± 5.0; P = 0.2816), the number of mature oocytes recovered (8.4 ± 5.0 vs. 7.2 ± 4.0; P = 0.2588), fertilization rate (5.9 ± 4.2 vs. 5.6 ± 3.3; P = 0.7390), and the number of usable embryos on day 3 (4.0 ± 3.0 vs. 4.0 ± 2.4; P = 0.8991). CONCLUSION: The dual trigger is equivalent to hCG in triggering oocyte maturation. PMID:27382235

  5. The Higher Response of Vascular Endothelial Growth Factor and Angiotensin-II to Human Chorionic Gonadotropin in Women with Polycystic Ovary Syndrome

    PubMed Central

    Qu, Junwei; Che, Yena; Xu, Pei; Xia, Yanjie; Wu, Xiaoke; Wang, Yong

    2015-01-01

    Background This research investigated the response of vascular active factors, vascular endothelial growth factor (VEGF) and angiotensin-II (AT-II) to ovarian stimulation during 24 hours in patients with polycystic ovary syndrome (PCOS). Materials and Methods In this clinical trial study, 52 patients with PCOS and 8 control cases were stimulated with human chorionic gonadotropin (HCG) on the 4th to 7th day of the patients’ natural or induced menstrual cycles. We measured VEGF and AT-II by radioimmunoassay before the injection (0 hour) and 3, 8, 12, 18 and 24 hours after the stimulation. Results After ovarian stimulation, there was substantially higher level of VEGF in typical PCOS patients than the other three groups at the 3 hour time point (p<0.05), while there were no significant differences in VEGF at all the other time points among the four groups. As for AT-II, before and at all time points after the ovarian stimulation, it seemed that the AT-II levels in patients’ sera with different phenotypes of PCOS by the Rotterdam criteria were all higher than in the control group although the differences were not statistically significant. The level of AT-II in typical PCOS patients was also significantly higher than the other three groups at the 3 hour time point (p<0.05), while no significant differences at all the other time points among the four groups were observed. Conclusion The response to the stimulation varied among patients with different phenotypes of PCOS according to the Rotterdam criteria. Serum VEGF and AT-II were possible contributors to an increased risk of developing ovarian hyperstimulation syndrome (OHSS) in patients with typical PCOS during the early follicular phase (3 hours) after ovarian stimulation (Registration Number: NCT02265861). PMID:25780518

  6. A Positive Feedback Loop between Glial Cells Missing 1 and Human Chorionic Gonadotropin (hCG) Regulates Placental hCGβ Expression and Cell Differentiation

    PubMed Central

    Cheong, Mei-Leng; Wang, Liang-Jie; Chuang, Pei-Yun; Chang, Ching-Wen; Lee, Yun-Shien; Lo, Hsiao-Fan; Tsai, Ming-Song

    2015-01-01

    Human chorionic gonadotropin (hCG) is composed of a common α subunit and a placenta-specific β subunit. Importantly, hCG is highly expressed in the differentiated and multinucleated syncytiotrophoblast, which is formed via trophoblast cell fusion and stimulated by cyclic AMP (cAMP). Although the ubiquitous activating protein 2 (AP2) transcription factors TFAP2A and TFAP2C may regulate hCGβ expression, it remains unclear how cAMP stimulates placenta-specific hCGβ gene expression and trophoblastic differentiation. Here we demonstrated that the placental transcription factor glial cells missing 1 (GCM1) binds to a highly conserved promoter region in all six hCGβ paralogues by chromatin immunoprecipitation-on-chip (ChIP-chip) analyses. We further showed that cAMP stimulates GCM1 and the CBP coactivator to activate the hCGβ promoter through a GCM1-binding site (GBS1), which also constitutes a previously identified AP2 site. Given that TFAP2C may compete with GCM1 for GBS1, cAMP enhances the association between the hCGβ promoter and GCM1 but not TFAP2C. Indeed, the hCG-cAMP-protein kinase A (PKA) signaling pathway also stimulates Ser269 and Ser275 phosphorylation of GCM1, which recruits CBP to mediate GCM1 acetylation and stabilization. Consequently, hCG stimulates the expression of GCM1 target genes, including the fusogenic protein syncytin-1, to promote placental cell fusion. Our study reveals a positive feedback loop between GCM1 and hCG regulating placental hCGβ expression and cell differentiation. PMID:26503785

  7. A novel nanocatalytic SERS detection of trace human chorionic gonadotropin using labeled-free Vitoria blue 4R as molecular probe.

    PubMed

    Wen, Guiqing; Liang, Xiaojing; Liu, Qingye; Liang, Aihui; Jiang, Zhiliang

    2016-11-15

    In pH 7.4 Na2HPO4-NaH2PO4 buffer solution containing the peptide probes for human chorionic gonadotropin (hCG), silver nanoparticles (AgNPs) were aggregated to big AgNPs clusters that exhibited very weak catalytic effect on the gold nanoparticle reaction of H2O2-HAuCl4. When hCG was present in the peptide probe solution, the AgNPs did not aggregate and it had strong catalytic effect on the gold nanoparticle reaction with a strong resonance Rayleigh scattering (RRS) peak at 370nm and a strong surface enhanced Raman scattering (SERS) peak at 1615cm(-1) in the presence of molecular probe of Victoria blue 4R (VB4R). With the increase of the hCG concentration, the catalysis enhanced due to the nanocatalyst of AgNPs increasing, and the RRS intensity increased at 370nm. The increased RRS intensity was linear to the hCG concentration in 0.05-10ng/mL, with a linear regression equation of ΔI370nm=409.8C +294. And the SERS intensity at 1615cm(-1) increased linearly with the hCG concentration in the range of 0.05-20ng/mL, with a linear regression equation of ΔI1615cm-1=142C+134. Based on this, two new methods of nanocatalytic SERS and RRS were proposed for the determination of trace hCG. PMID:27208477

  8. Use of human chorionic gonadotropin in a male Pacific walrus (Odobenus rosmarus divergens) to induce rut and achieve a pregnancy in a nulliparous female.

    PubMed

    Muraco, Holley S; Coombs, Leah D; Procter, Dianna G; Turek, Paul J; Muraco, Michael J

    2012-01-01

    Walrus in US zoos have a very low reproductive rate of 11 births in 80 years, and little is known about Pacific walrus (Odobenus rosmarus divergens) reproductive biology. To address this, we initiated a program in which detailed biological data were recorded on captive walrus. As part of a 7-year study, 1 male and 1 female 16-year-old captive Pacific walrus were carefully monitored with weekly serum hormone analysis, daily glans penis smears for spermatozoa, and abdominal ultrasound for pregnancy. The female ovulated once annually from late December through mid-January and then exhibited 9 months of sustained elevated progesterone. This nonconceptive estrous cycle profile is consistent with reports from wild walrus females. In contrast, the male's seasonal rut routinely occurred in late February through May with a serum testosterone peak in March. This profile differed from the reported adult male cycle in wild walrus of November through March. During the period of the female's ovulation, the male had nadir testosterone levels and was consistently azoospermic. Likewise, during the male's spermatogenic rut in the spring, the female was anovulatory with elevated progesterone. On this basis, the male was treated for 14 weeks with human chorionic gonadotropin (hCG) in an attempt to increase testosterone levels in synchrony with the female's annual ovulation. The treatment successfully induced rut characterized by sustained elevated serum testosterone levels and production of spermatozoa. The male and female successfully bred, and the female became pregnant. Upon discontinuation of hCG treatment, the male resumed baseline testosterone levels. We theorize that the lack of synchronization of rut and ovulatory cycles is a primary reason for reproductive failure in these captive walrus. PMID:22207706

  9. Preliminary evidence for associations between second-trimester human chorionic gonadotropin and unconjugated oestriol levels with pregnancy outcome in Down syndrome pregnancies.

    PubMed

    Benn, P A

    1998-04-01

    Fifty-six cases of Down syndrome were identified in a population of women who had undergone maternal serum triple marker screening [alpha-fetoprotein (AFP), human chorionic gonadotropin (hCG), and unconjugated oestriol (uE3) analyses]. These affected pregnancies represented all known cases present in the population of 34,368 women screened. Using a 1:270 mid-trimester Down syndrome risk to define the screen-positive group, 42 affected pregnancies were screen-positive (medians: AFP = 0.79 MOM, hCG = 2.13 MOM, uE3 = 0.62 MOM, age 34.6 years) and 14 pregnancies were screen-negative (medians: AFP = 0.82 MOM, hCG = 1.57 MOM, uE3 = 0.92 MOM, age 24.2 years). Four affected pregnancies were associated with in utero death and each of these cases was associated with relatively extreme values of AFP, hCG, and uE3, including the three highest levels of hCG in the entire series of Down syndrome pregnancies. Twenty-nine (15 screen-positive and 14 screen-negative) affected pregnancies resulted in liveborns. Down syndrome pregnancies had a significantly shorter gestational term than controls, and Down syndrome babies were also lighter than controls, even after adjustment for sex and gestational age. In affected pregnancies, a low uE3 level appeared to be associated with a greater chance of a small-for-gestational age baby. No correlations could be demonstrated between AFP or hCG levels and gestational age-adjusted term weight. Based on this small series, it would appear that uE3 may be particularly useful in detecting those Down syndrome cases associated with small-for-gestational age fetuses. A very high hCG value may indicate a higher probability of fetal death. PMID:9602476

  10. A prospective, randomised comparative study of weekly versus biweekly application of dehydrated human amnion/chorion membrane allograft in the management of diabetic foot ulcers.

    PubMed

    Zelen, Charles M; Serena, Thomas E; Snyder, Robert J

    2014-04-01

    The aim of this study is to determine if weekly application of dehydrated human amnion/chorion membrane allograft reduce time to heal more effectively than biweekly application for treatment of diabetic foot ulcers. This was an institutional review board-approved, registered, prospective, randomised, comparative, non-blinded, single-centre clinical trial. Patients with non-infected ulcers of ≥ 4 weeks duration were included for the study. They were randomised to receive weekly or biweekly application of allograft in addition to a non-adherent, moist dressing with compressive wrapping. All wounds were offloaded. The primary study outcome was mean time to healing. Overall, during the 12-week study period, 92·5% (37/40) ulcers completely healed. Mean time to complete healing was 4·1 ± 2·9 versus 2·4 ± 1·8 weeks (P = 0·039) in the biweekly versus weekly groups, respectively. Complete healing occurred in 50% versus 90% by 4 weeks in the biweekly and weekly groups, respectively (P = 0·014). Number of grafts applied to healed wounds was similar at 2·4 ± 1·5 and 2·3 ± 1·8 for biweekly versus weekly groups, respectively (P = 0·841). These results validate previous studies showing that the allograft is an effective treatment for diabetic ulcers and show that wounds treated with weekly application heal more rapidly than with biweekly application. More rapid healing may decrease clinical operational costs and prevent long-term medical complications. PMID:24618401

  11. Determination of hyperglycosylated human chorionic gonadotropin produced by malignant gestational trophoblastic neoplasias and male germ cell tumors using a lectin-based immunoassay and surface plasmon resonance

    PubMed Central

    Kelly, Lisa S.; Birken, Steven; Puett, David

    2007-01-01

    The ability to reliably detect aberrant glycosylation of human chorionic gonadotropin (hCG) may have profound implications for the diagnosis and monitoring of malignant gestational trophoblastic neoplasia, germ cell tumors, other malignancies, and pregnancy complications. To become a clinically useful assay, however, this discrimination of glycoforms should be possible on minimally treated biological specimens. Towards this end, we have developed a lectin-based sandwich-type immunoassay to compare the glycosylation patterns of hCG among urine specimens from patients presenting with a normal pregnancy, invasive mole, choriocarcinoma, and male germ cell tumors using carbohydrate-free antibody fragments as capture reagents and a panel of eight lectins, five recognizing neutral sugars and three recognizing sialic acid. There was no significant difference in the binding of any of the lectins to hCG in the urine of women over the gestational range of 6 – 38 weeks. Three lectins, however, exhibited differential binding to urinary hCG derived from these normal pregnant controls and that from patients with malignant forms of gestational trophoblastic disease and male germ cell tumors. Galanthus nivalis agglutinin and Maackia amurensis lectin, which bind terminal mannose and α(2–3)sialic acid, respectively, preferentially bound pregnancy-derived hCG, whereas the lectin, wheat germ agglutinin, which binds sialic acid and β(1–4)N-acetylglucosamine, exhibited decreased binding to pregnancy-derived hCG compared to that from patients with male germ cell tumors and malignant gestational trophoblastic neoplasia. The differential binding observed with these three promising lectins is most encouraging and warrants further examination. The experimental paradigm also holds promise for the development of comparable assays for other glycosylated tumor markers. PMID:17081681

  12. Possible interactions between leptin, gonadotrophin-releasing hormone (GnRH-I and II) and human chorionic gonadotrophin (hCG).

    PubMed

    Islami, D; Bischof, P; Chardonnens, D

    2003-10-10

    Leptin is a metabolic signal to the reproductive axis, where it increases the plasma levels of luteinising hormone (LH) and follicle stimulating hormone (FSH). Since the placental regulation of human chorionic gonadotrophin (hCG) mimics that of the pituitary LH, we undertook this study to see if leptin could be involved in the secretion and synthesis of hCG in first-trimester trophoblast. We incubated cytotrophoblastic cells (CTB) with GnRH-I or GnRH-II, for 4 or 48 h and collected the media at different times thereafter. GnRH-II was more potent than GnRH-I when incubated for 4 h with CTB. Leptin secretion, as measured at 4 h, was significantly stimulated by GnRH-II. When measured at 24 h leptin values were also increased as compared to controls. Neither GnRH-I, nor GnRH-II had any effect on leptin secretion when incubated for 48 h with CTB. Leptin was also added to perifused placental explants, and samples (in which hCG was measured) were collected every 3 min. Leptin significantly stimulated hCG secretion by explants and induced a pulse of hCG immediately (within 6 min) after its injection, increasing significantly the area under the curve (P=0.04) and the amplitude (P=0.02) of hCG pulses. We conclude that GnRH-II is more effective than GnRH-I in stimulating leptin secretion. This difference could be explained by the existence of two different types of placental GnRH receptors or two different pathways of GnRH degradation. Furthermore, we observe that leptin has a significant stimulatory effect on hCG pulsatility. PMID:12969578

  13. Subjective Quantitative Studies of Human Agency

    ERIC Educational Resources Information Center

    Alkire, Sabina

    2005-01-01

    Amartya Sen's writings have articulated the importance of human agency, and identified the need for information on agency freedom to inform our evaluation of social arrangements. Many approaches to poverty reduction stress the need for empowerment. This paper reviews "subjective quantitative measures of human agency at the individual level." It…

  14. Profiles of cytokines secreted by isolated human endometrial cells under the influence of chorionic gonadotropin during the window of embryo implantation

    PubMed Central

    2013-01-01

    Background Several studies have indicated that human pre-implantation embryo-derived chorionic gonadotropin (hCG) may influence the implantation process by its action on human endometrial epithelial and stromal cells. Despite reports indicating that hCG acts on these cells to affect the production of several cytokines and growth factors (e.g., MIF, IGF-I, VEGF, LIF, IL-11, GMCSF, CXL10 and FGF2), our understanding of the integral influence of hCG on paracrine interactions between endometrial stromal and epithelial cells during implantation is very limited. Methods In the present study, we examined the profile of 48 cytokines in the conditioned media of primary cell cultures of human implantation stage endometrium. Endometrial epithelial cells (group 1; n = 20), stromal cells (group 2; n = 20), and epithelial plus stromal cells (group 3; n = 20) obtained from mid-secretory stage endometrial samples (n = 60) were grown on collagen and exposed to different doses (0, 1, 10 and 100 IU/ml) of rhCG for 24 h in vitro. Immunochemical and qRT-PCR methods were used to determine cytokine profiles. Enrichment and process networks analyses were implemented using a list of cytokines showing differential secretion in response to hCG. Results Under basal conditions, endometrial epithelial and stromal cells exhibited cell type-specific profiles of secreted cytokines. Administration of hCG (100 IU) resulted in significantly (P < 0.05) different cytokine secretion profiles indicative of macropinocytic transport (HGF, MCSF) in epithelial cells, signal transduction (CCL4, FGF2, IL-1b, IL-6, IL-17, VEGF) in stromal cells, and epithelial-mesenchymal transition (FGF2, HGF, IL-1b, TNF) in mixed cells. Overall, the administration of hCG affected cytokines involved in the immune response, chemotaxis, inflammatory changes, proliferation, cell adhesion and apoptosis. Conclusions CG can influence the function of the endometrium during blastocyst implantation via its

  15. Human umbilical cord blood mononuclear cells and chorionic plate-derived mesenchymal stem cells promote axon survival in a rat model of optic nerve crush injury

    PubMed Central

    CHUNG, SOKJOONG; RHO, SEUNGSOO; KIM, GIJIN; KIM, SO-RA; BAEK, KWANG-HYUN; KANG, MYUNGSEO; LEW, HELEN

    2016-01-01

    The use of mesenchymal stem cells (MSCs) in cell therapy in regenerative medicine has great potential, particularly in the treatment of nerve injury. Umbilical cord blood (UCB) reportedly contains stem cells, which have been widely used as a hematopoietic source and may have therapeutic potential for neurological impairment. Although ongoing research is dedicated to the management of traumatic optic nerve injury using various measures, novel therapeutic strategies based on the complex underlying mechanisms responsible for optic nerve injury, such as inflammation and/or ischemia, are required. In the present study, a rat model of optic nerve crush (ONC) injury was established in order to examine the effects of transplanting human chorionic plate-derived MSCs (CP-MSCs) isolated from the placenta, as well as human UCB mononuclear cells (CB-MNCs) on compressed rat optic nerves. Expression markers for inflammation, apoptosis, and optic nerve regeneration were analyzed, as well as the axon survival rate by direct counting. Increased axon survival rates were observed following the injection of CB-MNCs at at 1 week post-transplantation compared with the controls. The levels of growth-associated protein-43 (GAP-43) were increased after the injection of CB-MNCs or CP-MSCs compared with the controls, and the expression levels of hypoxia-inducible factor-1α (HIF-1α) were also significantly increased following the injection of CB-MNCs or CP-MSCs. ERM-like protein (ERMN) and SLIT-ROBO Rho GTPase activating protein 2 (SRGAP2) were found to be expressed in the optic nerves of the CP-MSC-injected rats with ONC injury. The findings of our study suggest that the administration of CB-MNCs or CP-MSCs may promote axon survival through systemic concomitant mechanisms involving GAP-43 and HIF-1α. Taken together, these findings provide further understanding of the mechanisms repsonsible for optic nerve injury and may aid in the development of novel cell-based therapeutic strategies with

  16. Localization of major histocompatibility complex class I and II mRNA in human first-trimester chorionic villi by in situ hybridization.

    PubMed

    Lata, J A; Tuan, R S; Shepley, K J; Mulligan, M M; Jackson, L G; Smith, J B

    1992-04-01

    Maternal immune recognition of pregnancy occurs despite the nonexpression of classical major histocompatibility complex (MHC) antigenic determinants by chorionic villous trophoblast, which comprise the major surface area where maternal blood contacts fetal-derived cells. cDNA-mRNA in situ hybridization was used to probe expression of transcripts corresponding to nonpolymorphic MHC determinants in first-trimester chorionic villus samples. The HLA-B7 probe hybridization signals were localized to syncytiotrophoblast and to cells of the mesenchyme but not to villous cytotrophoblast. HLA-G mRNA was found only in syncytiotrophoblast. A DR beta clone hybridized to both villous cytotrophoblast and syncytiotrophoblast. The results suggest that expression of trophoblast class I and class II determinants early in gestation (10 wk) may be regulated by posttranscriptional events. This also suggests the potential for maternal antifetal alloimmune responses. PMID:1552281

  17. Morphologic features of human chorionic gonadotropin- or alpha-fetoprotein-producing germ cell tumors of the central nervous system: histological heterogeneity and surgical meaning.

    PubMed

    Sugiyama, K; Arita, K; Tominaga, A; Hanaya, R; Taniguchi, E; Okamura, T; Itoh, Y; Yamasaki, F; Kurisu, K

    2001-01-01

    Our study of germ cell tumors (GCT) of the central nervous system (CNS) investigated the relationship between tumor histology and patient serum titers of human chorionic gonadotropin (HGC) and alpha-fetoprotein (AFP). Thirty-five patients were enrolled. Their serum titers of HCG (mlU/ml) and/or AFP (ng/ml) before initial treatment were available, as were tumor specimens obtained before the administration of adjuvant therapy. They were divided into three groups, depending on whether HCG alone (group H), AFP alone (group A), or both HCG and AFP (group HA) were detected. Each group was subdivided into three groups: patients in group I had H, A, and/or HA titers below 9.9; patients in group II/III had titers from 10.0 to 999; and those in group IV had titers of 1000 or more. Serial sections of tissue specimens were repeatedly stained, mainly with hematoxylin and eosin (H-E) stain, HCG immunostain, and AFP immunostain. There were seven patients in the H-I group and five in H-II/III. Of these 12 patients, 11 had germinomas (G) and one had an embryonal carcinoma (EC). Five patients were included in group A: one was classified as A-II/III and had a germinoma, and the remaining four patients were in A-IV and had yolk sac tumors (YST) or mixed GCT consisting mainly of YST or EC (MXGCT-YST, EC). The HA group consisted of 18 patients. Three were classified as HA-I and had germinomas; nine HA-II/III patients had T or MXGCT-T; and six HA-IV patients had choriocarcinoma (CC), YST, MXGCT-CC, or MXGCT-YST. Throughout the study, the situations for the elevated serum titers could be elucidated in only four cases (three in group A-IV and one in group HA-IV). These results led to the conclusion that serologic evaluation is superior to morphologic evaluation in diagnosing marker-producing GCTs. From a diagnostic perspective, the role of surgery is to verify the HCG- and AFP-immunonegative tissue in patients with G, T, and EC. PMID:11908867

  18. Maternal serum hyperglycosylated human chorionic gonadotrophin (HhCG) in the first trimester of pregnancies affected by Down syndrome, using a sialic acid-specific lectin immunoassay.

    PubMed

    Spencer, K; Talbot, J A; Abushoufa, R A

    2002-08-01

    In a series of 54 cases of pregnancies complicated by Down syndrome and 224 unaffected pregnancies we examined maternal serum levels of hyperglycosylated human chorionic gonadotrophin (HhCG) in samples collected in the first trimester (11-13 weeks) using a sialic acid-specific lectin immunoassay. We compared these levels with those of other potential first trimester serum markers [free beta-hCG, pregnancy-associated plasma protein A (PAPP-A) and total hCG (ThCG)] and modeled detection rates and false-positive rates of various biochemical markers in conjunction with fetal nuchal translucency (NT) and maternal age using an maternal age standardized population. Maternal serum HhCG in cases of Down syndrome were significantly elevated (median MoM 1.97) with 24/54 (44%) of cases above the 95th centile for unaffected pregnancies. Free beta-hCG was also elevated (median MoM 2.09) with 33% of cases above the 95th centile. PAPP-A levels were reduced (median MoM 0.47) with 38% below the 5th centile. ThCG levels, whilst elevated (median MoM 1.34), had only 20% of cases above the 95th centile. Maternal serum HhCG levels were not correlated with fetal NT but showed significant correlation with ThCG and free beta-hCG and with PAPP-A in the Down syndrome group (r=0.536). Maternal serum HhCG levels in cases with Down syndrome had a significant correlation with gestational age, increasing as the gestation increased. When HhCG was combined together with fetal NT, PAPP-A and maternal age, at a 5% false-positive rate the modeled detection rate was 83%, some 6% lower than when free beta-hCG was used and some 4% better than when ThCG was used. Maternal serum HhCG is unlikely to be of additional value when screening for Down syndrome in the first trimester. PMID:12210572

  19. Down-regulation of messenger ribonucleic acid encoding an importer of sulfoconjugated steroids during human chorionic gonadotropin-induced follicular luteinization in vivo.

    PubMed

    Brown, Kristy A; Bouchard, Nadine; Lussier, Jacques G; Sirois, Jean

    2007-01-01

    Members of the organic anion transporting polypeptide (SLCO/OATP) superfamily are capable of importing anionic compounds across the lipid bilayer in a sodium-independent manner. Member 2B1 has been shown to transport few substrates, two of which are dihydroepiandrosterone-3-sulfate (DHEA-S) and estrone-3-sulfate. Steroid sulfatase (STS) catalyses the hydrolysis of these steroids into their unconjugated counterparts. The objective of this study was to investigate the regulation of SLCO2B1 and STS mRNAs during human chorionic gonadotropin (hCG)-induced ovulation/luteinization. The equine SLCO2B1 cDNA was cloned and shown to encode a 709-amino acid protein (OATP2B1) that is highly conserved when compared to mammalian orthologs. RT-PCR/Southern blot analyses were performed to study the regulation of SLCO2B1 and STS transcripts in equine preovulatory follicles isolated between 0 and 39h after hCG treatment. Results showed high levels of SLCO2B1 mRNA expression before hCG, with a marked decrease observed in follicles obtained 24-39h post-hCG (P<0.05). Analyses of isolated granulosa and theca interna cells identified high mRNA expression in both cell types prior to hCG treatment, with granulosa cells showing a more rapid SLCO2B1 mRNA down-regulation. No significant change in STS mRNA was observed in intact follicle walls. However, when both cell types were isolated, a significant decrease in STS mRNA was observed in granulosa cells 24-39h post-hCG. Collectively, these results demonstrate that the hCG-dependent induction of follicular luteinization is accompanied by the down-regulation of SLCO2B1 and STS transcripts. Considering that OATP2B1 can import sulfoconjugated DHEA and estrogens, and that STS can remove the sulfonate moiety from these steroids, their down-regulation in luteinizing preovulatory follicles may provide an additional biochemical basis for the decrease in ovarian 17beta-estradiol biosynthesis after the LH surge. PMID:17049229

  20. Plasma prorenin response to human chorionic gonadotropin in ovarian-hyperstimulated women: correlation with the number of ovarian follicles and steroid hormone concentrations.

    PubMed Central

    Itskovitz, J; Sealey, J E; Glorioso, N; Rosenwaks, Z

    1987-01-01

    Plasma prorenin and active renin were measured before and after human chorionic gonadotropin (hCG) administration in two groups of patients undergoing ovarian stimulation for 4-6 days with follicle-stimulating hormone alone or in combination with luteinizing hormone, for in vitro fertilization. Baseline total plasma renin (prorenin plus active renin; n = 12) averaged 25 +/- 8 ng/ml per hr (mean +/- SD). Total renin did not change during ovarian stimulation but it increased to 46 +/- 16 ng/ml per hr (P less than 0.05) 1 or 2 days later, just before hCG administration. Thirty-six hours after hCG administration, just before laparoscopy and egg retrieval, total renin was 123 +/- 97 ng/ml per hr; a peak of 182 +/- 143 ng/ml per hr occurred 2-6 days later--i.e., during the luteal phase of the menstrual cycle. In eight of the patients who did not conceive, total renin returned to baseline 14 days after hCG administration. In four who conceived, a nadir was reached (57 +/- 13 ng/ml per hr) 8-12 days after hCG administration and then total renin increased again as the plasma beta hCG measurement began to rise. By day 16 it averaged 225 +/- 157 ng/ml per hr. In a second group of five patients active renin and prorenin were measured separately. Active renin comprised less than 20% of the total renin at all times. It was unchanged until day 4 after hCG administration and then increased significantly only when plasma progesterone was high. Thus, the initial response to hCG was entirely due to an increase in prorenin. A highly significant correlation was observed between the number of follicles and the total renin increases on the day of aspiration (r = 0.93, P less than 0.001) and at the peak (r = 0.89, P less than 0.001). After hCG administration, a temporal relationship was observed between the rise in total renin and plasma estradiol and progesterone levels. These results demonstrate that plasma prorenin increases markedly after administration of hCG and that the rise is

  1. Effects of two human chorionic gonadotropin doses administered to the ovarian states during the in vitro fertilization and embryo transfer program

    PubMed Central

    MA, MINGXING; WANG, JIALIN; XU, LIJUN; ZHANG, QINXI; DU, BOTAO; JIANG, XIAOYING; SHI, QINGLI; ZHOU, LILI; LI, BAOXIN; SAITO, HIDEKAZU; KURACHI, HIROHISA

    2015-01-01

    The aim of the present study was to examine the effects of the human chorionic gonadotropin (hCG) dose on the pulsatility indices (PI) of the intraovarian artery on the day of follicle aspiration and the oocyte quality, intrafollicular oxidative stress and luteinization. PI was also measured on the day of hCG administration. A total of 15 patients were undergoing the in vitro fertilization and embryo transfer (IVF-ET) program. To estimate whether there was any difference between the intraovarian artery blood flow and oocyte development of the same patients treated with 5,000 or 10,000 IU hCG, the intraovarian artery blood flow was measured by transvaginal color ultrasonography pulsed wave Doppler, and the follicular fluids and the granulosa cells were collected at follicle aspiration. There were statistically significant differences between the same patients undergoing the two different hCG-dose treatments in which the first protocol included 10,000 IU and the second protocol included 5,000 IU hCG treatment. These differences were apparent in the PI of intraovarian artery blood flow on the day of follicle aspiration (P=0.0023), in the incidence of apoptosis in cumulus (ApoC) and mural (ApoM) granulosa cells (ApoC, P=0.0077; ApoM, P=0.0128), in the total oocytes retrieved (P=0.0342) and in the follicle fluid progesterone concentration (P=0.0044). There were no significant differences between the two protocols in the PI of intraovarian artery blood flow on the day of hCG administration (P=0.4326), serum steroid on the day of follicle aspiration [serum P, P>0.9999; serum estradiol (E2), P=0.8589], follicle fluid E2 concentration (P=0.8939), mature oocyte rate (P=0.3743) and total mature oocytes retrieved (P=0.2026). In conclusion, the dose of hCG administration can significantly affect the intraovarian artery blood flow and the development of follicles and oocytes in an IVF-ET program. PMID:26075075

  2. Reproductive outcomes of Alpine goats primed with progesterone and treated with human chorionic gonadotropin during the anestrus-to-estrus transition season.

    PubMed

    Alvarado-Espino, A S; Meza-Herrera, C A; Carrillo, E; González-Álvarez, V H; Guillen-Muñoz, J M; Ángel-García, O; Mellado, M; Véliz-Deras, F G

    2016-04-01

    This study aimed to determine the possible effects of a single injection of human chorionic gonadotropin (hCG) as a means for estrus induction in acyclic French-Alpine goats during the reproductive transition period at 25°N, 103°W. The potential effects of hCG upon ovarian function and reproductive performance of goats were also assessed. Multiparous acyclic French-Alpine goats (n = 39; 37.4 ± 8 .5 kg) were primed with 20mg progesterone (P4) 1 day prior to hCG administration. Thereafter, does were treated either with saline (hCG-0; n = 10), 50 (hCG-50; n = 9), 100 (hCG-100; n = 10), or 300 IU of hCG (hCG-300; n = 10). Ovarian structures and pregnancy were monitored by transrectal ultrasonography. In addition, after hCG application, goats were monitored twice daily (0800 and 1800 h) to detect estrus signs, with the use of aproned, sexually active bucks treated with testosterone. Goats were bred 12h after the onset of estrus. Two days after hCG administration, the number of large follicles was higher (P < 0.05) in the hCG-50 and hCG-300 groups (1.7 ± 0.1 and 1.8 ± 0.2, respectively) compared with the hCG-100 and hCG-0 groups (1.4 ± 0.2 and 1.1 ± 0.1, respectively). Although none of the hCG-0-goats depicted estrus, the estrus response from the hCG-50, hCG-100, and hCG-300 groups over the 7-d breeding period was 67%, 100%, and 90%, respectively (P > 0.05), being always accompanied by ovulation. Pregnancy rate (67, 100, and 70%), kidding rate (55%, 80%, and 70%), and litter size (1.6 ± 0.5, 1.5 ± 0.5, and 1.5 ± 0.5) for hCG-50, hCG-100, and hCG-300, respectively, did not differ among the hCG-treated does. Therefore, the combined use of P4-priming plus a 100-IU hCG injection is an effective protocol for inducing estrus in non-cycling Alpine goats during the anestrus-to-estrus transition period, which is of key importance for both goat producers and industrializers. PMID:26944772

  3. Differential expression of human chorionic gonadotropin (hCG) glycosylation isoforms in failing and continuing pregnancies: preliminary characterization of the hyperglycosylated hCG epitope.

    PubMed

    Kovalevskaya, G; Birken, S; Kakuma, T; Ozaki, N; Sauer, M; Lindheim, S; Cohen, M; Kelly, A; Schlatterer, J; O'Connor, J F

    2002-03-01

    Human chorionic gonadotropin (hCG) glycoforms change as pregnancy progresses. We have developed an antibody (B152) which can measure a hyperglycosylated early pregnancy isoform of hCG. This putative hyperglycosylated form of hCG arises very early in pregnancies and is rapidly replaced by an isoform that predominates for the remainder of the pregnancy. The profiles of these hCG glycoforms are measured as a ratio of values of two immunometric assays. The profiles of these ratios differ between pregnancies which persist and those which will experience early failure. In this report, daily urine hCG isoform ratios from donor eggs (no exogenous hCG pretreatment), in vitro fertilization pregnancies were profiled and analyzed from the first day following embryo transfer (ET). Significant differences were found between continuing pregnancy and pregnancy loss throughout days 5-20 post-ET. When hCG isoform ratios were analyzed from the first day of detectable hCG, pregnancy loss could be predicted in the case of a single fetus both during the 5- to 10-day time segment (P=0.018) and the 10- to 15-day time segment (P=0.045). When single and multiple fetus pregnancies were analyzed together significance was approached in the 10- to 15-day time period (P=0.058). In a second population of pregnant women who conceived naturally, in whom urine samples were collected at approximately weekly intervals to either term birth or clinical spontaneous abortion, the ratio could discriminate between miscarriages and normal term pregnancies (P=0.043). In later pregnancy, the ratio of hCG isoforms declined more rapidly in miscarriages than in term pregnancy. Antibody B152 was produced using a choriocarcinoma-derived hCG (C5), which was hyperglycosylated at both N- and O-linked sites and was 100% nicked at position beta(47-48). Western blot analyses supported the assay results showing that early pregnancy urine does not contain nicked C5-like hCG. Also, the early pregnancy hCG appeared to be the

  4. Estrus synchronization and fertility in post-partum dairy cattle after administration of human chorionic gonadotrophin (HCG) and prostaglandin F2 alpha analog.

    PubMed

    De Rensis, F; Allegri, M; Seidel, G E

    1999-07-15

    Human chorionic gonadotrophin (hCG) plus PGF2 alpha was compared with GnRH plus PGF2 alpha for estrus synchronization of dairy cows. There were 3 treatments: GnRH analog (Buserelin, 12.6 micrograms) plus PGF2 alpha analog (Cloprostenol, 150 micrograms) 6 d later (GnRH + PGF[Day 6]); hCG (2000 IU) plus PGF2 alpha 9 d later (hCG + PGF[Day 9]); and hCG plus PGF2 alpha 6 d later (hCG + PGF[Day 6]). Treatment occurred either Days 55 to 90 or Days 91 to 135 post partum. For responses during the first 10 d after PGF2 alpha administration, estrus synchronization (P = 0.24), efficacy (percentage of treated pregnant; P = 0.20) and conception (percentage of inseminated pregnant; P = 0.23) rates were not different among the 3 treatments. Cows treated between Days 55 and 90 had a higher rate (P < 0.05) of detected estrus during this period (69% for GnRH + PG [Day 6], 70% for hCG + PGF[Day 9] and 72% for hCG + PGF[Day 6]) compared with cows treated between Days 91 and 135 (52% for GnRH + PGF[Day 6], 50% for hCG + PGF[Day 9] and 57% for hCG + PGF[Day 6]). Efficacy of treatment was higher (P < 0.05) in animals treated between Days 55 and 90 (54% for GnRH + PGF[Day 6], 56% for hCG + PGF[Day 9] and 63% for hCG + PGF [Day 6]) compared to animals treated between Days 91 and 135 (36% for GnRH + PGF[Day 6], 35% for hCG + PGF[Day 9] and 47% for hCG + PGF[Day 6]). There were no significant differences in conception between Days 51 and 90 and Days 91 and 135. The interval between parturition-first AI with conception was significantly (P < 0.001) shorter in GnRH + PGF (Day 6; 106 d), hCG + PGF (Day 9; 109 d) and hCG + PGF (Day 6; 103 d) treated cattle than in 106 untreated animals (136 d). Thus, GnRH plus PGF2 alpha or hCG plus PGF2 alpha treatments elicited similar effects in estrus synchronization, treatment efficacy, and conception rate in post-partum dairy cows. PMID:10734393

  5. Role of early serum beta human chorionic gonadotropin measurement in predicting multiple pregnancy and pregnancy wastage in an in vitro ET fertilization cycle

    PubMed Central

    Singh, Neeta; Begum, Anjuman Ara; Malhotra, Neena; Bahadur, Anupama; Vanamail, P.

    2013-01-01

    OBJECTIVE: This study was performed to assess the prognostic value of serum beta human chorionic gonadotropin (βhCG), measured on day 14 post embryo transfer (ET) for predicting multiple gestation and pregnancy wastage in women undergoing in vitro fertilization ET (IVF-ET). MATERIALS AND METHODS: This retrospective study was performed between May 2009 and November 2012. Out of the 181 women who conceived, 168 were included and the remaining 13 were excluded as their pregnancy was biochemical. Serum βhCG was measured using a chemiluminescent enzyme immunometric assay. The predictive values of serum βhCG for establishing multiple pregnancy and pregnancy wastages were calculated by receiver operating characteristic (ROC) curve analysis. Median values of serum βhCG and outcome of all pregnancies were compared. RESULTS: Out of the 168 patients who conceived after IVF treatment, 114 (68%) were viable pregnancies (delivered/ongoing). Among the viable pregnancies, 97 (85%) had a successful pregnancy outcome and the remaining 17 patients are ongoing pregnancies. Median values of βhCG (625 IU/L) among viable pregnancies was significantly (P < 0.05) higher than that of nonviable pregnancies (174 IU/L). The median values of βhCG for singleton (502 IU/L), twins (1093 IU/L), and triplets (2160 IU/L) was statistically significant (P < 0.05). Using ROC curve it was predicted that for a value of βhCG at 375 IU/L, the sensitivity of viable pregnancy was 65% and specificity of viable pregnancy was also 65%, with positive and negative predictive values of 65 and 68%, respectively. Similarly for multiple pregnancy and pregnancy wastage the predictive values of βhCG were 808 and 375 IU/L, respectively; while the sensitivity and specificity is more than 65% each. CONCLUSION: βhCG cutoff values determined on day 14 post ET by ROC curve analysis are useful in discriminating between multiple pregnancy and pregnancy losses. The cutoff value might aid in the prognosis, clinical

  6. A comparison of human chorionic gonadotropin and luteinizing hormone releasing hormone on the induction of spermiation and amplexus in the American toad (Anaxyrus americanus)

    PubMed Central

    2012-01-01

    Background Captive breeding programs for endangered amphibian species often utilize exogenous hormones for species that are difficult to breed. The purpose of our study was to compare the efficacy of two different hormones at various concentrations on sperm production, quantity and quality over time in order to optimize assisted breeding. Methods Male American toads (Anaxyrus americanus) were divided into three separate treatment groups, with animals in each group rotated through different concentrations of luteinizing hormone releasing hormone analog (LHRH; 0.1, 1.0, 4.0 and 32 micrograms/toad), human chorionic gonadotropin (hCG; 50, 100, 200, and 300 IU), or the control over 24 hours. We evaluated the number of males that respond by producing spermic urine, the sperm concentration, percent motility, and quality of forward progression. We also evaluated the effects of hCG and LHRH on reproductive behavior as assessed by amplexus. Data were analyzed using the Generalized Estimating Equations incorporating repeated measures over time and including the main effects of treatment and time, and the treatment by time interaction. Results The hormone hCG was significantly more effective at stimulating spermiation in male Anaxyrus americanus than LHRH and showed a dose-dependent response in the number of animals producing sperm. At the most effective hCG dose (300 IU), 100% of the male toads produced sperm, compared to only 35% for the best LHRH dose tested (4.0 micrograms). In addition to having a greater number of responders (P < 0.05), the 300 IU hCG treatment group had a much higher average sperm concentration (P < 0.05) than the treatment group receiving 4.0 micrograms LHRH. In contrast, these two treatments did not result in significant differences in sperm motility or quality of forward progressive motility. However, more males went into amplexus when treated with LHRH vs. hCG (90% vs. 75%) by nine hours post-administration. Conclusion There is a clear

  7. Quantitation of phosphorothioate oligonucleotides in human plasma.

    PubMed

    Leeds, J M; Graham, M J; Truong, L; Cummins, L L

    1996-03-01

    Methods are presented for the extraction of phosphorothioate oligonucleotides from human plasma to permit quantitation by capillary gel electrophoresis. Extraction of the phosphorothioate oligonucleotides from plasma was accomplished using two solid-phase extraction columns, a strong anion-exchange column to remove plasma proteins and lipids, followed by a reverse-phase column to remove salts. A second desalting step, achieved by dialysis utilizing a membrane with a molecular weight cutoff of 2500 Da floating on distilled water, was required to remove residual ionic material from the extracted sample. This method should be generally applicable to the analysis and quantitation of phosphorothioate oligonucleotides. PMID:8850544

  8. Quantitative deconvolution of human thermal infrared emittance.

    PubMed

    Arthur, D T J; Khan, M M

    2013-01-01

    The bioheat transfer models conventionally employed in etiology of human thermal infrared (TIR) emittance rely upon two assumptions; universal graybody emissivity and significant transmission of heat from subsurface tissue layers. In this work, a series of clinical and laboratory experiments were designed and carried out to conclusively evaluate the validity of the two assumptions. Results obtained from the objective analyses of TIR images of human facial and tibial regions demonstrated significant variations in spectral thermophysical properties at different anatomic locations on human body. The limited validity of the two assumptions signifies need for quantitative deconvolution of human TIR emittance in clinical, psychophysiological and critical applications. A novel approach to joint inversion of the bioheat transfer model is also introduced, levering the deterministic temperature-dependency of proton resonance frequency in low-lipid human soft tissue for characterizing the relationship between subsurface 3D tissue temperature profiles and corresponding TIR emittance. PMID:23086533

  9. Germ cell quantitation in human testicular biopsy.

    PubMed

    Sinha Hikim, A P; Chakraborty, J; Jhunjhunwala, J S

    1985-01-01

    Quantitative analysis of human seminiferous epithelium was carried out using an improved method of glutaraldehyde and osmium fixation with plastic embedding. Part of each biopsy specimen was fixed in Bouin's fixative and embedded in paraffin for comparison. Epon embedded tissue had very little artifactual damage compared with paraffin embedded tissue sections. The germ cell to Sertoli cell ratios were determined by counting the various germ cells per "unit" tubular area. Data obtained by this method reflect a remarkable stability of Sertoli cell number and germ cell-Sertoli cell ratios both between biopsies from different individuals and between biopsies from right and left testes from the same individual. Agreement between the present results and those of earlier studies based on paraffin embedded testicular specimens supports the validity of this method of germ cell quantitation of human testicular biopsy samples. PMID:3927550

  10. 21 CFR 522.1079 - Serum gonadotropin and chorionic gonadotropin.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Serum gonadotropin and chorionic gonadotropin. 522.1079 Section 522.1079 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN.... (i) Gilts. For induction of fertile estrus (heat) in healthy prepuberal (noncycling) gilts. (ii)...

  11. 21 CFR 522.1079 - Serum gonadotropin and chorionic gonadotropin.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Serum gonadotropin and chorionic gonadotropin. 522.1079 Section 522.1079 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN.... (i) Gilts. For induction of fertile estrus (heat) in healthy prepuberal (noncycling) gilts. (ii)...

  12. 21 CFR 522.1079 - Serum gonadotropin and chorionic gonadotropin.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Serum gonadotropin and chorionic gonadotropin. 522.1079 Section 522.1079 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN.... (i) Gilts. For induction of fertile estrus (heat) in healthy prepuberal (noncycling) gilts. (ii)...

  13. Quantitative Proteomic Analysis of the Human Nucleolus.

    PubMed

    Bensaddek, Dalila; Nicolas, Armel; Lamond, Angus I

    2016-01-01

    Recent years have witnessed spectacular progress in the field of mass spectrometry (MS)-based quantitative proteomics, including advances in instrumentation, chromatography, sample preparation methods, and experimental design for multidimensional analyses. It is now possible not only to identify most of the protein components of a cell proteome in a single experiment, but also to describe additional proteome dimensions, such as protein turnover rates, posttranslational modifications, and subcellular localization. Furthermore, by comparing the proteome at different time points, it is possible to create a "time-lapse" view of proteome dynamics. By combining high-throughput quantitative proteomics with detailed subcellular fractionation protocols and data analysis techniques it is also now possible to characterize in detail the proteomes of specific subcellular organelles, providing important insights into cell regulatory mechanisms and physiological responses. In this chapter we present a reliable workflow and protocol for MS-based analysis and quantitation of the proteome of nucleoli isolated from human cells. The protocol presented is based on a SILAC analysis of human MCF10A-Src-ER cells with analysis performed on a Q-Exactive Plus Orbitrap MS instrument (Thermo Fisher Scientific). The subsequent chapter describes how to process the resulting raw MS files from this experiment using MaxQuant software and data analysis procedures to evaluate the nucleolar proteome using customized R scripts. PMID:27576725

  14. The quantitative modelling of human spatial habitability

    NASA Technical Reports Server (NTRS)

    Wise, J. A.

    1985-01-01

    A model for the quantitative assessment of human spatial habitability is presented in the space station context. The visual aspect assesses how interior spaces appear to the inhabitants. This aspect concerns criteria such as sensed spaciousness and the affective (emotional) connotations of settings' appearances. The kinesthetic aspect evaluates the available space in terms of its suitability to accommodate human movement patterns, as well as the postural and anthrometric changes due to microgravity. Finally, social logic concerns how the volume and geometry of available space either affirms or contravenes established social and organizational expectations for spatial arrangements. Here, the criteria include privacy, status, social power, and proxemics (the uses of space as a medium of social communication).

  15. Oocyte and cumulus cell transcripts from cultured mouse follicles are induced to deviate from normal in vivo conditions by combinations of insulin, follicle-stimulating hormone, and human chorionic gonadotropin.

    PubMed

    Sánchez, Flor; Romero, Sergio; Smitz, Johan

    2011-09-01

    Gonadotropins and insulin are major regulators of cell proliferation, differentiation, and survival in cultured mouse ovarian follicles. Applications of variable doses of insulin in combination with follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were studied at the gene expression level in oocytes and cumulus cells. Early preantral follicles grown over 9 days were sequentially exposed to combinations of doses of insulin, FSH, and human chorionic gonadotropin (hCG). From culture Day 1 to 6 (preantral stage), two insulin concentrations (5 ng/ml and 5 μg/ml) were combined with 10 mIU/ml FSH. From Days 6 to 9 (antral stage), the three variable gonadotropin treatments set under each insulin condition were 10 mIU/ml FSH, 25 mIU/ml FSH, and 25 mIU/ml FSH plus 3 mIU/ml hCG. The Gdf9, Bmp15, Fgf8, Dazl, Pou5f1, and Pik3ca mRNA transcripts were quantified in oocytes, and the Amh, Lhcgr, Hsd3b1, Vegfa, and Insig1 mRNA transcripts were quantified in cumulus cells. In vivo controls were unprimed and eCG (equine chorionic gonadotropin)-primed prepubertal female mice. During the preantral stage, none except the Amh transcripts was regulated by insulin. Oocyte transcripts were not affected by the variable gonadotropin treatments on the last culture day but were upregulated in the combination of high insulin plus 25 mIU/ml FSH. Under low insulin conditions, high FSH levels increased levels of Lhcgr and Vegfa expression, and hCG abated this effect. However, under high insulin conditions, hCG upregulated levels of Lhcgr, Vegfa, and Insig1 mRNA. High insulin concentrations upregulated Hsd3b1 transcripts. These results demonstrate that in an in vitro follicle culture, a near physiological insulin background yields oocyte and cumulus cell transcript levels that are more similar to those in vivo. PMID:21565993

  16. Quantitation of vitamin K in human milk

    SciTech Connect

    Canfield, L.M.; Hopkinson, J.M.; Lima, A.F.; Martin, G.S.; Sugimoto, K.; Burr, J.; Clark, L.; McGee, D.L. )

    1990-07-01

    A quantitative method was developed for the assay of vitamin K in human colostrum and milk. The procedure combines preparative and analytical chromatography on silica gel in a nitrogen atmosphere followed by reversed phase high performance liquid chromatography (HPLC). Two HPLC steps were used: gradient separation with ultraviolet (UV) detection followed by isocratic separation detected electrochemically. Due to co-migrating impurities, UV detection alone is insufficient for identification of vitamin K. Exogenous vitamin K was shown to equilibrate with endogenous vitamin K in the samples. A statistical method was incorporated to control for experimental variability. Vitamin K1 was analyzed in 16 pooled milk samples from 7 donors and in individual samples from 15 donors at 1 month post-partum. Vitamin K1 was present at 2.94 +/- 1.94 and 3.15 +/- 2.87 ng/mL in pools and in individuals, respectively. Menaquinones, the bacterial form of the vitamin, were not detected. The significance of experimental variation to studies of vitamin K in individuals is discussed.

  17. Increased SCF/c-kit by hypoxia promotes autophagy of human placental chorionic plate-derived mesenchymal stem cells via regulating the phosphorylation of mTOR.

    PubMed

    Lee, Youjin; Jung, Jieun; Cho, Kyung Jin; Lee, Seoung-Kwan; Park, Jong-Wan; Oh, Il-Hoan; Kim, Gi Jin

    2013-01-01

    Hypoxia triggers physiological and pathological cellular processes, including proliferation, differentiation, and death, in several cell types. Mesenchymal stem cells (MSCs) derived from various tissues have self-renewal activity and can differentiate towards multiple lineages. Recently, it has been reported that hypoxic conditions tip the balance between survival and death by hypoxia-induced autophagy, although the underlying mechanism is not clear. The objectives of this study are to compare the effect of hypoxia on the self-renewal of bone marrow-derived mesenchymal stem cells (BM-MSCs) and placental chorionic plate-derived mesenchymal stem cells (CP-MSCs) and to investigate the regulatory mechanisms of self-renewal in each MSC type during hypoxia. The expression of self-renewal markers (e.g., Oct4, Nanog, Sox2) was assessed in both cell lines. PI3K and stem cell factor (SCF) expression gradually increased in CP-MSCs but were markedly downregulated in BM-MSCs by hypoxia. The phosphorylation of ERK and mTOR was augmented by hypoxia in CP-MSCs compared to control. Also, the expression of LC3 II, a component of the autophagosome and the hoof-shaped autophagosome was detected more rapidly in CP-MSCs than in BM-MSCs under hypoxia. Hypoxia induced the expression of SCF in CP-MSCs and increased SCF/c-kit pathway promotes the self-renewal activities of CP-MSCs via an autocrine/paracrine mechanism that balances cell survival and cell death events by autophagy. These activities occur to a greater extent in CP-MSCs than in BM-MSCs through regulating the phosphorylation of mTOR. These findings will provide useful guidelines for better understanding the function of SCF/c-kit in the self-renewal and autophagy-regulated mechanisms that promote of MSC survival. PMID:22833529

  18. Quantitative PCR for Genetic Markers of Human Fecal Pollution

    EPA Science Inventory

    Assessment of health risk and fecal bacteria loads associated with human fecal pollution requires reliable host-specific analytical methods and a rapid quantificationapproach. We report the development of quantitative PCR assays for quantification of two recently described human-...

  19. Treatment of cystic ovarian disease in dairy cattle. Comparative observation of the effects of an intramuscular injection of corticosteroids and an intravenous injection of a combination of human chorionic gonadotropin and progesterone.

    PubMed

    Nakao, T; Ono, H

    1977-01-01

    Of 67 cows with cystic ovarian disease (COD), 34 were injected intramuscularly with 20 mg of betamethasone of 10 mg of dexamethasone (CC) and 33 intravenously with a combination of 3,000 IU of human chorionic gonadotropin and 125 mg of progesterone (HCG -P). Pregnancy rates and intervals between treatment and conception were 32.4% and 32.1 +/- 30.5 days, respectively, in the CC-treated cows and 30.3% and 51.6 +/- 29.5 days in the HCG -P-treated ones. Cows which had not responded to gonadotropin treatment showed a considerably higher pregnancy rate when treated with CC and a lower pregnancy rate when administered with HCG -P than those which had received no treatment before. There was a trend that the earlier a cow was treated, the more readily she recovered. CC injection gave a satisfactory result even when performed long after calving. The estrous behaviour seemed to be related with prognosis in cows with COD. In both CC-treated and HCG -P-treated cows, the pregnancy rate was the highest in cows showing anestrus and the lowest in those exhibiting irregular estrus. PMID:64331

  20. A comprehensive analysis of the chorion locus in silkmoth

    PubMed Central

    Chen, Zhiwei; Nohata, Junko; Guo, Huizhen; Li, Shenglong; Liu, Jianqiu; Guo, Youbing; Yamamoto, Kimiko; Kadono-Okuda, Keiko; Liu, Chun; Arunkumar, Kallare P.; Nagaraju, Javaregowda; Zhang, Yan; Liu, Shiping; Labropoulou, Vassiliki; Swevers, Luc; Tsitoura, Panagiota; Iatrou, Kostas; Gopinathan, Karumathil P.; Goldsmith, Marian R.; Xia, Qingyou; Mita, Kazuei

    2015-01-01

    Despite more than 40 years of intense study, essential features of the silkmoth chorion (eggshell) are still not fully understood. To determine the precise structure of the chorion locus, we performed extensive EST analysis, constructed a bacterial artificial chromosome (BAC) contig, and obtained a continuous genomic sequence of 871,711 base pairs. We annotated 127 chorion genes in two segments interrupted by a 164 kb region with 5 non-chorion genes, orthologs of which were on chorion bearing scaffolds in 4 ditrysian families. Detailed transcriptome analysis revealed expression throughout choriogenesis of most chorion genes originally categorized as “middle”, and evidence for diverse regulatory mechanisms including cis-elements, alternative splicing and promoter utilization, and antisense RNA. Phylogenetic analysis revealed multigene family associations and faster evolution of early chorion genes and transcriptionally active pseudogenes. Proteomics analysis identified 99 chorion proteins in the eggshell and micropyle localization of 1 early and 6 Hc chorion proteins. PMID:26553298

  1. A comprehensive analysis of the chorion locus in silkmoth.

    PubMed

    Chen, Zhiwei; Nohata, Junko; Guo, Huizhen; Li, Shenglong; Liu, Jianqiu; Guo, Youbing; Yamamoto, Kimiko; Kadono-Okuda, Keiko; Liu, Chun; Arunkumar, Kallare P; Nagaraju, Javaregowda; Zhang, Yan; Liu, Shiping; Labropoulou, Vassiliki; Swevers, Luc; Tsitoura, Panagiota; Iatrou, Kostas; Gopinathan, Karumathil P; Goldsmith, Marian R; Xia, Qingyou; Mita, Kazuei

    2015-01-01

    Despite more than 40 years of intense study, essential features of the silkmoth chorion (eggshell) are still not fully understood. To determine the precise structure of the chorion locus, we performed extensive EST analysis, constructed a bacterial artificial chromosome (BAC) contig, and obtained a continuous genomic sequence of 871,711 base pairs. We annotated 127 chorion genes in two segments interrupted by a 164 kb region with 5 non-chorion genes, orthologs of which were on chorion bearing scaffolds in 4 ditrysian families. Detailed transcriptome analysis revealed expression throughout choriogenesis of most chorion genes originally categorized as "middle", and evidence for diverse regulatory mechanisms including cis-elements, alternative splicing and promoter utilization, and antisense RNA. Phylogenetic analysis revealed multigene family associations and faster evolution of early chorion genes and transcriptionally active pseudogenes. Proteomics analysis identified 99 chorion proteins in the eggshell and micropyle localization of 1 early and 6 Hc chorion proteins. PMID:26553298

  2. Type I and II Diabetic Adipose-Derived Stem Cells Respond In Vitro to Dehydrated Human Amnion/Chorion Membrane Allograft Treatment by Increasing Proliferation, Migration, and Altering Cytokine Secretion

    PubMed Central

    Massee, Michelle; Chinn, Kathryn; Lim, Jeremy J.; Godwin, Lisa; Young, Conan S.; Koob, Thomas J.

    2016-01-01

    Objective: Human amniotic membranes have been shown to be effective for healing diabetic foot ulcers clinically and to regulate stem cell activity in vitro and in vivo; however, diabetic stem cells may be impaired as a sequela of the disease. In this study, dehydrated human amnion/chorion membrane (dHACM) allografts (EpiFix®; MiMedx Group) were evaluated for their ability to regulate diabetic stem cells in vitro. Approach: Human adipose-derived stem cells (ADSCs) from normal, type I diabetic, and type II diabetic donors were treated with soluble extracts of dHACM and evaluated for proliferation after 3 days by DNA assay, chemotactic migration after 1 day by transwell assay, cytokine secretion after 3 days by multiplex ELISA, and gene expression after 5 days by reverse transcription–polymerase chain reaction. Results: Although diabetic ADSCs demonstrated decreased responses compared to normal ADSCs, dHACM treatment stimulated diabetic ADSCs to proliferate after 3 days and enhanced migration over 24 h, similar to normal ADSCs. dHACM-treated diabetic ADSCs modulated secretion of soluble signals, including regulators of inflammation, angiogenesis, and healing. All ADSCs evaluated also responded to dHACM treatment with altered expression of immunomodulatory genes, including interleukins (IL)-1α, IL-1β, and IL-1RA. Innovation: This is the first reported case demonstrating that diabetic ADSCs respond to novel amniotic membrane therapies, specifically treatment with dHACM. Conclusion: dHACM stimulated diabetic ADSCs to migrate, proliferate, and alter cytokine expression suggesting that, despite their diabetic origin, ADSCs may respond to dHACM to accelerate diabetic wound healing. PMID:26862462

  3. Quantitative PCR for genetic markers of human fecal pollution

    EPA Science Inventory

    Assessment of health risk and fecal bacteria loads associated with human fecal pollution requires reliable host-specific analytical methods and a rapid quantification approach. We report the development of quantitative PCR assays for enumeration of two recently described hum...

  4. MicroRNA miR-513a-3p acts as a co-regulator of luteinizing hormone/chorionic gonadotropin receptor gene expression in human granulosa cells.

    PubMed

    Troppmann, B; Kossack, N; Nordhoff, V; Schüring, A N; Gromoll, J

    2014-06-01

    The luteinizing hormone/chorionic gonadotropin receptor (LHCGR) is essential for normal male and female reproductive processes. The spatial and temporal LHCGR gene expression is controlled by a complex system of regulatory mechanisms which are crucial for normal physiological function, especially during the female cycle. In this study, we aimed to elucidate whether microRNAs are involved in this network and play a role in regulating LHCGR expression. Computational analysis predicted that miR-513a-3p interacts with the LHCGR mRNA via three binding sites located in the 3'UTR region, enabling a synergistic action. Moreover, using a luciferase-based reporter assay we found that miR-513a-3p targets the LHCGR, resulting in a significant down-regulation of its expression. In human primary granulosa cell cultures we detected a dynamic, inversely associated expression pattern of miR-513a-3p and the LHCGR. In addition, transfection with miR-513a-3p or its specific inhibitor led to a down- or up-regulation at the LHCGR mRNA level, respectively. An increased amount of miR-513a-3p resulted in the down-regulation of the LHCGR mRNA, reflected by the attenuation of cAMP synthesis after hormonal stimulation. In conclusion, these data demonstrate that miR-513a-3p is involved in the control of the LHCGR expression by an inversely regulated mechanism at the post-transcriptional level and show for the first time that this kind of post-transcriptional process contributes to the multifaceted system of the human LHCGR regulation. PMID:24747085

  5. The quantitative failure of human reliability analysis

    SciTech Connect

    Bennett, C.T.

    1995-07-01

    This philosophical treatise argues the merits of Human Reliability Analysis (HRA) in the context of the nuclear power industry. Actually, the author attacks historic and current HRA as having failed in informing policy makers who make decisions based on risk that humans contribute to systems performance. He argues for an HRA based on Bayesian (fact-based) inferential statistics, which advocates a systems analysis process that employs cogent heuristics when using opinion, and tempers itself with a rational debate over the weight given subjective and empirical probabilities.

  6. Plasma progesterone profile and conception rate following exogenous supplementation of gonadotropin-releasing hormone, human chorionic gonadotropin, and progesterone releasing intra-vaginal device in repeat-breeder crossbred cows

    PubMed Central

    Pandey, N. K. J.; Gupta, H. P.; Prasad, Shiv; Sheetal, S. K.

    2016-01-01

    Aim: This study was designed to evaluate the effect of gonadotropin-releasing hormone (GnRH), human chorionic gonadotropin (hCG), and progesterone impregnated intra-vaginal device on progesterone profile and conception rate in repeat-breeding crossbred cows. Materials and Methods: Repeat-breeding crossbred cows aged 3-8 years (n=32), lactating and negative to white side test were randomly divided into four groups: Group 1 (Control, n=8), Group 2 (GnRH at 10 µg i.m, n=8), Group 3 (hCG at 1500 IU i.m., n=8), and Group 4 (progesterone impregnated intra-vaginal device at 958 mg, n=8). All the treatme nts were given on 5th daypostbreeding and in Group 4 intra-vaginally implanted device was withdrawn on 9th day (i.e., implant inserted for total 4 days) of the estrous cycle. Blood samples were collected on day 0, 5, 10, 15, and day 20 of estrous cycle, and plasma was separated for progesterone estimation. Results: Accessory corpus luteum was not formed in crossbred cows of Group4 and control group. However, total 6 and 8 accessory corpora lutea were found in Group 2 and Group 3, respectively. In pregnant cows, the plasma progesterone concentration increased continuously from day 0 to day 20. In non-pregnant cows, it increased from day 0 to day 15 and then declined. The conception rate on day 60 in Group 1, Group 2, Group 3, and Group 4 was 37.5%, 50%, 75%, and 37.5%, respectively. Conclusions: Treating repeat-breeder cows with hCG is effective in increasing conception rate by developing accessory corpora lutea and higher progesterone level. PMID:27397976

  7. Human Chorionic Gonadotropin Has Anti-Inflammatory Effects at the Maternal-Fetal Interface and Prevents Endotoxin-Induced Preterm Birth, but Causes Dystocia and Fetal Compromise in Mice.

    PubMed

    Furcron, Amy-Eunice; Romero, Roberto; Mial, Tara N; Balancio, Amapola; Panaitescu, Bogdan; Hassan, Sonia S; Sahi, Aashna; Nord, Claire; Gomez-Lopez, Nardhy

    2016-06-01

    Human chorionic gonadotropin (hCG) is implicated in the maintenance of uterine quiescence by down-regulating myometrial gap junctions during pregnancy, and it was considered as a strategy to prevent preterm birth after the occurrence of preterm labor. However, the effect of hCG on innate and adaptive immune cells implicated in parturition is poorly understood. Herein, we investigated the immune effects of hCG at the maternal-fetal interface during late gestation, and whether this hormone can safely prevent endotoxin-induced preterm birth. Using immunophenotyping, we demonstrated that hCG has immune effects at the maternal-fetal interface (decidual tissues) by: 1) increasing the proportion of regulatory T cells; 2) reducing the proportion of macrophages and neutrophils; 3) inducing an M1 → M2 macrophage polarization; and 4) increasing the proportion of T helper 17 cells. Next, ELISAs were used to determine whether the local immune changes were associated with systemic concentrations of progesterone, estradiol, and/or cytokines (IFNgamma, IL1beta, IL2, IL4, IL5, IL6, IL10, IL12p70, KC/GRO, and TNFalpha). Plasma concentrations of IL1beta, but not progesterone, estradiol, or any other cytokine, were increased following hCG administration. Pretreatment with hCG prevented endotoxin-induced preterm birth by 44%, proving the effectiveness of this hormone as an anti-inflammatory agent. However, hCG administration alone caused dystocia and fetal compromise, as proven by Doppler ultrasound. These results provide insight into the mechanisms whereby hCG induces an anti-inflammatory microenvironment at the maternal-fetal interface during late gestation, and demonstrate its effectiveness in preventing preterm labor/birth. However, the deleterious effects of this hormone on mothers and fetuses warrant caution. PMID:27146032

  8. Effect of a single injection of gonadotropin-releasing hormone (GnRH) and human chorionic gonadotropin (hCG) on testicular blood flow measured by color doppler ultrasonography in male Shiba goats.

    PubMed

    Samir, Haney; Sasaki, Kazuaki; Ahmed, Eman; Karen, Aly; Nagaoka, Kentaro; El Sayed, Mohamed; Taya, Kazuyoshi; Watanabe, Gen

    2015-05-01

    Although color Doppler ultrasonography has been used to evaluate testicular blood flow in many species, very little has been done in goat. Eight male Shiba goats were exposed to a single intramuscular injection of either gonadotropin-releasing hormone (GnRH group; 1 µg/kg BW) or human chorionic gonadotropin (hCG group; 25 IU/kg BW). Plasma testosterone (T), estradiol (E2) and inhibin (INH) were measured just before (0 hr) and at different intervals post injection by radioimmunoassay. Testis volume (TV) and Doppler indices, such as resistive index (RI) and pulsatility index (PI) of the supratesticular artery, were measured by B-mode and color Doppler ultrasonography, respectively. The results indicated an increase in testicular blood flow in both groups, as RI and PI decreased significantly (P<0.05), but this increase was significant higher and earlier in hCG group (1 hr) than in the GnRH group (2 hr). A high correlation was found for RI and PI with both T (RI, r= -0.862; PI, r= -0.707) and INH in the GnRH group (RI, r=0.661; PI, r=0.701). However, a significant (P<0.05) correlation was found between E2 and both RI (r= -0.610) and PI (r= -0.763) in hCG group. In addition, TV significantly increased and was highly correlated with RI in both groups (GnRH, r= -0.718; hCG, r= -0.779). In conclusion, hCG and GnRH may improve testicular blood flow and TV in Shiba goats. PMID:25715956

  9. The chorionic bump: Etiologic insights from two pathologic pregnancies.

    PubMed

    Wax, Joseph R; Blaszyk, Hagen; Jones, Michael; Cartin, Angelina; Pinette, Michael G

    2016-09-01

    The clinical significance and etiology of the chorionic bump remain unclear. We describe two pregnancies characterized by chorionic bumps, which subsequently were diagnosed with a complete mole and trisomy 18, respectively. We hypothesize that placental pathology, including edema and hydropic villi, may contribute to or cause the sonographic finding of some chorionic bumps. An association between chorionic bumps and aneuploidy awaits future study. © 2016 Wiley Periodicals, Inc. J Clin Ultrasound 44:452-454, 2016. PMID:27220064

  10. The quantitative modelling of human spatial habitability

    NASA Technical Reports Server (NTRS)

    Wise, James A.

    1988-01-01

    A theoretical model for evaluating human spatial habitability (HuSH) in the proposed U.S. Space Station is developed. Optimizing the fitness of the space station environment for human occupancy will help reduce environmental stress due to long-term isolation and confinement in its small habitable volume. The development of tools that operationalize the behavioral bases of spatial volume for visual kinesthetic, and social logic considerations is suggested. This report further calls for systematic scientific investigations of how much real and how much perceived volume people need in order to function normally and with minimal stress in space-based settings. The theoretical model presented in this report can be applied to any size or shape interior, at any scale of consideration, for the Space Station as a whole to an individual enclosure or work station. Using as a point of departure the Isovist model developed by Dr. Michael Benedikt of the U. of Texas, the report suggests that spatial habitability can become as amenable to careful assessment as engineering and life support concerns.

  11. Quantitative PIXE analysis of human uterine myoma

    NASA Astrophysics Data System (ADS)

    Yeh, S. C.; Chu, T. C.; Lin, H. J.; Hsu, C. C.

    1986-11-01

    Twenty-two samples, taken from eight pathological proved uterine myoma patients, were embedded in paraffin and cut into slices of identical thickness (4.0 μm). After deparaffinization, washing and drying, the slices of myomal tissue and their neighboring myometrial tissue were bombarded by 2.0 MeV proton beams from a 3 MV Van de Graaff accelerator. The induced characteristic X-rays were then detected and analyzed using a HPGe detector system. The absolute concentrations of trace elements contained in tumors and normal tissues of human myomal uterus were determined, in reference to a known concentration of doped yttrium. Significant correlations between the concentration of elements, both in tumors and in normal tissues, were found.

  12. SMAD1/5 Signaling in the Early Equine Placenta Regulates Trophoblast Differentiation and Chorionic Gonadotropin Secretion

    PubMed Central

    Cabrera-Sharp, Victoria; Read, Jordan E.; Richardson, Stephanie; Kowalski, Alycia A.; Antczak, Douglas F.; Cartwright, Judith E.; Mukherjee, Abir

    2014-01-01

    TGFβ superfamily proteins, acting via SMAD (Sma- and Mad-related protein)2/3 pathways, regulate placental function; however, the role of SMAD1/5/8 pathway in the placenta is unknown. This study investigated the functional role of bone morphogenetic protein (BMP)4 signaling through SMAD1/5 in terminal differentiation of primary chorionic gonadotropin (CG)-secreting trophoblast. Primary equine trophoblast cells or placental tissues were isolated from day 27–34 equine conceptuses. Detected by microarray, RT-PCR, and quantitative RT-PCR, equine chorionic girdle trophoblast showed increased gene expression of receptors that bind BMP4. BMP4 mRNA expression was 20- to 60-fold higher in placental tissues adjacent to the chorionic girdle compared with chorionic girdle itself, suggesting BMP4 acts primarily in a paracrine manner on the chorionic girdle. Stimulation of chorionic girdle-trophoblast cells with BMP4 resulted in a dose-dependent and developmental stage-dependent increase in total number and proportion of terminally differentiated binucleate cells. Furthermore, BMP4 treatment induced non-CG-secreting day 31 chorionic girdle trophoblast cells to secrete CG, confirming a specific functional response to BMP4 stimulation. Inhibition of SMAD2/3 signaling combined with BMP4 treatment further enhanced differentiation of trophoblast cells. Phospho-SMAD1/5, but not phospho-SMAD2, expression as determined by Western blotting was tightly regulated during chorionic girdle trophoblast differentiation in vivo, with peak expression of phospho-SMAD1/5 in vivo noted at day 31 corresponding to maximal differentiation response of trophoblast in vitro. Collectively, these experiments demonstrate the involvement of BMP4-dependent pathways in the regulation of equine trophoblast differentiation in vivo and primary trophoblast differentiation in vitro via activation of SMAD1/5 pathway, a previously unreported mechanism of TGFβ signaling in the mammalian placenta. PMID:24848867

  13. Follicle-stimulating hormone potentiates the steroidogenic activity of chorionic gonadotropin and the anti-apoptotic activity of luteinizing hormone in human granulosa-lutein cells in vitro.

    PubMed

    Casarini, Livio; Riccetti, Laura; De Pascali, Francesco; Nicoli, Alessia; Tagliavini, Simonetta; Trenti, Tommaso; La Sala, Giovanni Battista; Simoni, Manuela

    2016-02-15

    Luteinizing hormone (LH) and choriogonadotropin (hCG) are glycoprotein hormones regulating ovarian function and pregnancy, respectively. Since these molecules act on the same receptor (LHCGR), they were traditionally assumed as equivalent in assisted reproduction techniques (ART), although differences between LH and hCG were demonstrated at molecular and physiological level. In this study, we demonstrated for the first time that co-treatment with a follicle-stimulating hormone (FSH) dose in the ART therapeutic range potentiates different LH- and hCG-dependent responses in vitro, measured in terms of cAMP, phospho-CREB, -ERK1/2 and -AKT activation, gene expression, progesterone and estradiol production in human granulosa-lutein cells (hGLC). We show that in the presence of FSH, hCG biopotency is about 5-fold increased, in the presence of FSH, in terms of cAMP activation. Accordingly, CREB phosphorylation and steroid production is increased under hCG and FSH co-treatment. LH effects, evaluated as steroidogenic cAMP/PKA pathway activation, do not change in the presence of FSH, which, however, increases LH-dependent ERK1/2 and AKT, but not CREB phosphorylation, resulting in anti-apoptotic effects. The different modulatory activity of FSH on LH and hCG action in vitro corresponds to their different physiological functions, reflecting proliferative effects exerted by LH during the follicular phase and before trophoblast development, and the high steroidogenic potential of hCG requested to sustain pregnancy from the luteal phase onwards. PMID:26690776

  14. A double-blind, placebo-controlled, randomized clinical trial of recombinant human chorionic gonadotropin on muscle strength and physical function and activity in older men with partial age-related androgen deficiency.

    PubMed

    Liu, Peter Y; Wishart, Susan M; Handelsman, David J

    2002-07-01

    Despite partial androgen deficiency, the safety and efficacy of androgen therapy in older men remains controversial because controlled studies of testosterone have given equivocal results. Human chorionic gonadotropin (hCG) can be conveniently and infrequently self-administered, and it increases not only circulating testosterone but also estradiol and other testicular steroids. We evaluated the efficacy and safety of 3 months of treatment with sc recombinant hCG (r-hCG, Ovidrel) on muscle mass, strength, mobility, and physical activity in ambulant, community-dwelling men more than 60 yr old having partial androgen deficiency (testosterone < or = 15 nmol/liter, twice). Forty eligible men (mean age, 67 yr; range, 60-85 yr) were randomized to receive r-hCG (5000 IU, 250 microg) or placebo by twice weekly sc self-injection and were studied before treatment, monthly during treatment, and 1 month after treatment. All completed the study, and treatment groups were well matched. r-hCG significantly increased body weight (approximately 1 kg; P < 0.05) and lean body mass ( approximately 2 kg; P < 0.001) and reduced fat mass (approximately 1 kg, P < 0.05). However, anthropometric measures of skinfold thickness (biceps, triceps, subscapular, suprailiac) and circumferences (midarm, waist, hip, and midthigh), including the waist-hip ratio, did not change significantly. Shoulder and knee strength (peak torque), as measured by isokinetic and isometric dynamometry, was not significantly increased, nor was physical activity (accelerometry and Physical Activity Scale for Elderly self-report) or gait and balance (modified Guralnik and Frailty and Injuries: Cooperative Studies of Intervention Techniques performance batteries) altered. Total and free testosterone and estradiol were markedly (150%; P < 0.001) and stably increased, whereas LH, FSH, and urea were significantly decreased. Testis volume was significantly decreased (approximately 5 ml; P < 0.05). There were no significant

  15. Amnion and Chorion Allografts in Combination with Coronally Advanced Flap in the Treatment of Gingival Recession: A Clinical Study

    PubMed Central

    Chakraborthy, Sonali; Sambashivaiah, Savita; Bilchodmath, Shivaprasad

    2015-01-01

    Background Guided tissue regeneration (GTR) based root coverage using different allograft membranes has been utilized to correct gingival recession defects with promising results. Amnion and chorion allograft membranes of alternative origin derived from human placental tissue has been advocated in the treatment of gingival recession. However, chorion membrane has been used in combination with amnion membrane no study has compared these allograft membranes in the treatment of gingival recession. Therefore, the purpose of this study was to clinically evaluate and compare the efficacy of amnion membrane and chorion membrane in combination with coronally advanced flap in the treatment of gingival recessions. Materials and Methods Twelve systemically healthy patients having at least 2 bilateral Miller’s Class I or Class II gingival recession were recruited and coronally advanced flap was performed with amnion membrane or chorion membrane. Clinical parameters such as gingival Index, plaque index, length of the recession, width of the recession, width of keratinized gingiva, relative attachment level were evaluated at baseline, 3 and 6 months post-surgery. Results The mean decrease in length of recession (LR) for Chorion site was 2.00±1.54mm and amnion site was 1.58±1.14mm. The gain in attachment level for amnion site was 2.17±1.53mm and for chorion site was 1.58±1.22mm. The total mean percentage of root coverage was 34% for chorion site and 22% for amnion site. Conclusion Both amnion membrane and chorion membrane has shown to be versatile allograft material to be used in the treatment of root coverage. PMID:26501023

  16. Precision mapping of multiple quantitative trait loci in humans

    SciTech Connect

    Guo, S.W.

    1994-09-01

    Mapping complex quantitative traits in humans is in general a difficult task. Parametric methods are very efficient in mapping a single major quantitative trait locus but they are sensitive to violations in modeling assumptions such non-normality, absence of major genes, and biased sampling. Semi-parametric methods such as Haseman-Elston sib-pair method are more robust but they are less efficient than parametric ones. We have extended the sib-pair method to (1) incorporate information from multiple markers based on the gene identical-by-descent proportions and the proportion of allele sharing at the trait locus, all based on flanking markers; (2) to include sibship data; and (3) to allow for precise localization of multiple quantitative trait loci. We show through simulation that the power of the proposed approach is greater than the sib-pair method. Further extension to include other types of relative pairs is possible.

  17. Quantitative determination of fissionable materials in human hair

    SciTech Connect

    Bentley, K.W.; Wyatt, J.H.

    1980-04-01

    A technique for quantitative neutron-induced autoradiography (NIAR) of human hair samples containing small amounts of plutonium or uranium is described. It requires irradiation of hairs in intimate contact with two thin mica sheets by a suitable dose of thermal neutrons to provide adequate numbers of fission fragments. The technique has been used to examine scalp hair voluntarily provided by 35 humans intermittently exposed to both soluble and insoluble uranium materials. The results indicate that in situations of low intermittent exposure the method is preferable to presently used blood, urine, or whole body monitoring methods. Limitations of the method are discussed.

  18. Quantitative PCR for genetic markers of human fecal pollution.

    PubMed

    Shanks, Orin C; Kelty, Catherine A; Sivaganesan, Mano; Varma, Manju; Haugland, Richard A

    2009-09-01

    Assessment of health risk and fecal bacterial loads associated with human fecal pollution requires reliable host-specific analytical methods and a rapid quantification approach. We report the development of quantitative PCR assays for quantification of two recently described human-specific genetic markers targeting Bacteroidales-like cell surface-associated genes. Each assay exhibited a range of quantification from 10 to 1 x 10(6) copies of target DNA. For each assay, internal amplification controls were developed to detect the presence or absence of amplification inhibitors. The assays predominantly detected human fecal specimens and exhibited specificity levels greater than 97% when tested against 265 fecal DNA extracts from 22 different animal species. The abundance of each human-specific genetic marker in primary effluent wastewater samples collected from 20 geographically distinct locations was measured and compared to quantities estimated by real-time PCR assays specific for rRNA gene sequences from total Bacteroidales and enterococcal fecal microorganisms. Assay performances combined with the prevalence of DNA targets in sewage samples provide experimental evidence supporting the potential application of these quantitative methods for monitoring fecal pollution in ambient environmental waters. PMID:19592537

  19. Quantitative analysis of laminin 5 gene expression in human keratinocytes.

    PubMed

    Akutsu, Nobuko; Amano, Satoshi; Nishiyama, Toshio

    2005-05-01

    To examine the expression of laminin 5 genes (LAMA3, LAMB3, and LAMC2) encoding the three polypeptide chains alpha3, beta3, and gamma2, respectively, in human keratinocytes, we developed novel quantitative polymerase chain reaction (PCR) methods utilizing Thermus aquaticus DNA polymerase, specific primers, and fluorescein-labeled probes with the ABI PRISM 7700 sequence detector system. Gene expression levels of LAMA3, LAMB3, and LAMC2 and glyceraldehyde-3-phosphate dehydrogenase were quantitated reproducibly and sensitively in the range from 1 x 10(2) to 1 x 10(8) gene copies. Basal gene expression level of LAMB3 was about one-tenth of that of LAMA3 or LAMC2 in human keratinocytes, although there was no clear difference among immunoprecipitated protein levels of alpha3, beta3, and gamma2 synthesized in radio-labeled keratinocytes. Human serum augmented gene expressions of LAMA3, LAMB3, and LAMC2 in human keratinocytes to almost the same extent, and this was associated with an increase of the laminin 5 protein content measured by a specific sandwich enzyme-linked immunosorbent assay. These results demonstrate that the absolute mRNA levels generated from the laminin 5 genes do not determine the translated protein levels of the laminin 5 chains in keratinocytes, and indicate that the expression of the laminin 5 genes may be controlled by common regulation mechanisms. PMID:15854126

  20. Quantitative analysis of astrogliosis in drug-dependent humans.

    PubMed

    Weber, Marco; Scherf, Nico; Kahl, Thomas; Braumann, Ulf-Dietrich; Scheibe, Patrick; Kuska, Jens-Peer; Bayer, Ronny; Büttner, Andreas; Franke, Heike

    2013-03-15

    Drug addiction is a chronic, relapsing disease caused by neurochemical and molecular changes in the brain. In this human autopsy study qualitative and quantitative changes of glial fibrillary acidic protein (GFAP)-positive astrocytes in the hippocampus of 26 lethally intoxicated drug addicts and 35 matched controls are described. The morphological characterization of these cells reflected alterations representative for astrogliosis. But, neither quantification of GFAP-positive cells nor the Western blot analysis indicated statistical significant differences between drug fatalities versus controls. However, by semi-quantitative scoring a significant shift towards higher numbers of activated astrocytes in the drug group was detected. To assess morphological changes quantitatively, graph-based representations of astrocyte morphology were obtained from single cell images captured by confocal laser scanning microscopy. Their underlying structures were used to quantify changes in astroglial fibers in an automated fashion. This morphometric analysis yielded significant differences between the investigated groups for four different measures of fiber characteristics (Euclidean distance, graph distance, number of graph elements, fiber skeleton distance), indicating that, e.g., astrocytes in drug addicts on average exhibit significant elongation of fiber structures as well as two-fold increase in GFAP-positive fibers as compared with those in controls. In conclusion, the present data show characteristic differences in morphology of hippocampal astrocytes in drug addicts versus controls and further supports the involvement of astrocytes in human pathophysiology of drug addiction. The automated quantification of astrocyte morphologies provides a novel, testable way to assess the fiber structures in a quantitative manner as opposed to standard, qualitative descriptions. PMID:23337617

  1. Quantitative Structural Insight into Human Variegate Porphyria Disease*

    PubMed Central

    Wang, Baifan; Wen, Xin; Qin, Xiaohong; Wang, Zhifang; Tan, Ying; Shen, Yuequan; Xi, Zhen

    2013-01-01

    Defects in the human protoporphyrinogen oxidase (hPPO) gene, resulting in ∼50% decreased activity of hPPO, is responsible for the dominantly inherited disorder variegate porphyria (VP). To understand the molecular mechanism of VP, we employed the site-directed mutagenesis, biochemical assays, structural biology, and molecular dynamics simulation studies to investigate VP-causing hPPO mutants. We report here the crystal structures of R59Q and R59G mutants in complex with acifluorfen at a resolution of 2.6 and 2.8 Å. The r.m.s.d. of the Cα atoms of the active site structure of R59G and R59Q with respect to the wild-type was 0.20 and 0.15 Å, respectively. However, these highly similar static crystal structures of mutants with the wild-type could not quantitatively explain the observed large differences in their enzymatic activity. To understand how the hPPO mutations affect their catalytic activities, we combined molecular dynamics simulation and statistical analysis to quantitatively understand the molecular mechanism of VP-causing mutants. We have found that the probability of the privileged conformations of hPPO can be correlated very well with the kcat/Km of PPO (correlation coefficient, R2 > 0.9), and the catalytic activity of 44 clinically reported VP-causing mutants can be accurately predicted. These results indicated that the VP-causing mutation affect the catalytic activity of hPPO by affecting the ability of hPPO to sample the privileged conformations. The current work, together with our previous crystal structure study on the wild-type hPPO, provided the quantitative structural insight into human variegate porphyria disease. PMID:23467411

  2. The quantitation of C6 in rabbit and human sera

    PubMed Central

    Tedesco, F.; Lachmann, P. J.

    1971-01-01

    C6 quantitation was carried out in rabbit and human sera by the single radial immunodiffusion technique. The C6 content of the rabbit and human sera used as standards was estimated by precipitin analysis, using an anti-C6 antiserum labelled with 125I. The mean C6 level in normal human serum was 11 μg/ml, whereas in normal rabbit serum it was 35 μg/ml. Sera from forty rabbits heterozygous for C6 deficiency were found to have a mean concentration of C6 of 14 μg/ml. The C6 level was estimated in sera from patients with various immunological disorders and found to be significantly higher in the sera from patients with rheumatoid arthritis and normal in the sera from patients with SLE, glomerulonephritis, nephrotic syndrome and myeloma. C6 haemolytic assays were found to correlate well with the antigenic assays only in fresh sera. In various circumstances this correlation breaks down, presumably because of C6 inactivator. This inactivator, in contrast to C3-inactivator, appears to be bound to antigen–antibody complement complexes. ImagesFig. 2Fig. 3 PMID:4998970

  3. Quantitation of human in vitro megakaryocytopoiesis by radioimmunoassay

    SciTech Connect

    Grant, B.W.; Nichols, W.L.; Solberg, L.A.; Yachimiak, D.J.; Mann, K.G.

    1987-05-01

    The isolation and characterization of human megakaryocyte growth factors has been hampered because evaluation of megakaryocyte growth in semisolid medium requires both lengthy incubation and visual quantitation. In addition, colony formation requires cell division, while most regulation of platelet production may involve individual, nonproliferating differentiating megakaryocytes. We have developed a radioimmunoassay (RIA) that makes use of an iodinated murine monoclonal antibody (MoAb) specific for platelet/megakaryocyte glycoprotein IIb/IIIa (GPIIb/IIIa) to measure megakaryocyte production in liquid marrow culture. This assay is sensitive to 3 X 10(3) platelets (roughly 30 megakaryocytes) and linear up to 1 X 10(6) platelets, and thus it provides a useful range for quantitating megakaryocyte production in in vitro marrow culture. Significant differences (threefold to fivefold) in megakaryocyte/platelet-specific GPIIb/IIIa complex are detected between stimulated and unstimulated marrow cultures by day 7, although antigen accrual in stimulated cultures continues through at least day 16. Conditions that promote megakaryocyte growth in semisolid medium (ie, aplastic plasma and PHA-LCM) have also been facilitory in liquid culture. This rapid and sensitive assay for cell-bound GPIIb/IIIa should facilitate recognition and isolation of megakaryocyte and platelet growth factors.

  4. Optical spectroscopy for quantitative sensing in human pancreatic tissues

    NASA Astrophysics Data System (ADS)

    Wilson, Robert H.; Chandra, Malavika; Lloyd, William; Chen, Leng-Chun; Scheiman, James; Simeone, Diane; McKenna, Barbara; Mycek, Mary-Ann

    2011-07-01

    Pancreatic adenocarcinoma has a five-year survival rate of only 6%, largely because current diagnostic methods cannot reliably detect the disease in its early stages. Reflectance and fluorescence spectroscopies have the potential to provide quantitative, minimally-invasive means of distinguishing pancreatic adenocarcinoma from normal pancreatic tissue and chronic pancreatitis. The first collection of wavelength-resolved reflectance and fluorescence spectra and time-resolved fluorescence decay curves from human pancreatic tissues was acquired with clinically-compatible instrumentation. Mathematical models of reflectance and fluorescence extracted parameters related to tissue morphology and biochemistry that were statistically significant for distinguishing between pancreatic tissue types. These results suggest that optical spectroscopy has the potential to detect pancreatic disease in a clinical setting.

  5. A quantitative transcriptome reference map of the normal human hippocampus.

    PubMed

    Caracausi, Maria; Rigon, Vania; Piovesan, Allison; Strippoli, Pierluigi; Vitale, Lorenza; Pelleri, Maria Chiara

    2016-01-01

    We performed an innovative systematic meta-analysis of 41 gene expression profiles of normal human hippocampus to provide a quantitative transcriptome reference map of it, i.e. a reference typical value of expression for each of the 30,739 known mapped and the 16,258 uncharacterized (unmapped) transcripts. For this aim, we used the software called TRAM (Transcriptome Mapper), which is able to generate transcriptome maps based on gene expression data from multiple sources. We also analyzed differential expression by comparing the hippocampus with the whole brain transcriptome map to identify a typical expression pattern of this subregion compared with the whole organ. Finally, due to the fact that the hippocampus is one of the main brain region to be severely affected in trisomy 21 (the best known genetic cause of intellectual disability), a particular attention was paid to the expression of chromosome 21 (chr21) genes. Data were downloaded from microarray databases, processed, and analyzed using TRAM software. Among the main findings, the most over-expressed loci in the hippocampus are the expressed sequence tag cluster Hs.732685 and the member of the calmodulin gene family CALM2. The tubulin folding cofactor B (TBCB) gene is the best gene at behaving like a housekeeping gene. The hippocampus vs. the whole brain differential transcriptome map shows the over-expression of LINC00114, a long non-coding RNA mapped on chr21. The hippocampus transcriptome map was validated in vitro by assaying gene expression through several magnitude orders by "Real-Time" reverse transcription polymerase chain reaction (RT-PCR). The highly significant agreement between in silico and experimental data suggested that our transcriptome map may be a useful quantitative reference benchmark for gene expression studies related to human hippocampus. Furthermore, our analysis yielded biological insights about those genes that have an intrinsic over-/under-expression in the hippocampus. PMID

  6. Human lymphocyte polymorphisms detected by quantitative two-dimensional electrophoresis

    SciTech Connect

    Goldman, D.; Merril, C.R.

    1983-09-01

    A survey of 186 soluble lymphocyte proteins for genetic polymorphism was carried out utilizing two-dimensional electrophoresis of /sup 14/C-labeled phytohemagglutinin (PHA)-stimulated human lymphocyte proteins. Nineteen of these proteins exhibited positional variation consistent with independent genetic polymorphism in a primary sample of 28 individuals. Each of these polymorphisms was characterized by quantitative gene-dosage dependence insofar as the heterozygous phenotype expressed approximately 50% of each allelic gene product as was seen in homozygotes. Patterns observed were also identical in monozygotic twins, replicate samples, and replicate gels. The three expected phenotypes (two homozygotes and a heterozygote) were observed in each of 10 of these polymorphisms while the remaining nine had one of the homozygous classes absent. The presence of the three phenotypes, the demonstration of gene-dosage dependence, and our own and previous pedigree analysis of certain of these polymorphisms supports the genetic basis of these variants. Based on this data, the frequency of polymorphic loci for man is: P . 19/186 . .102, and the average heterozygosity is .024. This estimate is approximately 1/3 to 1/2 the rate of polymorphism previously estimated for man in other studies using one-dimensional electrophoresis of isozyme loci. The newly described polymorphisms and others which should be detectable in larger protein surveys with two-dimensional electrophoresis hold promise as genetic markers of the human genome for use in gene mapping and pedigree analyses.

  7. Quantitative proteomic profiling of human articular cartilage degradation in osteoarthritis.

    PubMed

    Lourido, Lucía; Calamia, Valentina; Mateos, Jesús; Fernández-Puente, Patricia; Fernández-Tajes, Juan; Blanco, Francisco J; Ruiz-Romero, Cristina

    2014-12-01

    Osteoarthritis (OA) is the most common rheumatic pathology and is characterized primarily by articular cartilage degradation. Despite its high prevalence, there is no effective therapy to slow disease progression or regenerate the damaged tissue. Therefore, new diagnostic and monitoring tests for OA are urgently needed, which would also promote the development of alternative therapeutic strategies. In the present study, we have performed an iTRAQ-based quantitative proteomic analysis of secretomes from healthy human articular cartilage explants, comparing their protein profile to those from unwounded (early disease) and wounded (advanced disease) zones of osteoarthritic tissue. This strategy allowed us to identify a panel of 76 proteins that are distinctively released by the diseased tissue. Clustering analysis allowed the classification of proteins according to their different profile of release from cartilage. Among these proteins, the altered release of osteoprotegerin (decreased in OA) and periostin (increased in OA), both involved in bone remodelling processes, was verified in further analyses. Moreover, periostin was also increased in the synovial fluid of OA patients. Altogether, the present work provides a novel insight into the mechanisms of human cartilage degradation and a number of new cartilage-characteristic proteins with possible biomarker value for early diagnosis and prognosis of OA. PMID:25383958

  8. Quantitation of small intestinal permeability during normal human drug absorption

    PubMed Central

    2013-01-01

    Background Understanding the quantitative relationship between a drug’s physical chemical properties and its rate of intestinal absorption (QSAR) is critical for selecting candidate drugs. Because of limited experimental human small intestinal permeability data, approximate surrogates such as the fraction absorbed or Caco-2 permeability are used, both of which have limitations. Methods Given the blood concentration following an oral and intravenous dose, the time course of intestinal absorption in humans was determined by deconvolution and related to the intestinal permeability by the use of a new 3 parameter model function (“Averaged Model” (AM)). The theoretical validity of this AM model was evaluated by comparing it to the standard diffusion-convection model (DC). This analysis was applied to 90 drugs using previously published data. Only drugs that were administered in oral solution form to fasting subjects were considered so that the rate of gastric emptying was approximately known. All the calculations are carried out using the freely available routine PKQuest Java (http://www.pkquest.com) which has an easy to use, simple interface. Results Theoretically, the AM permeability provides an accurate estimate of the intestinal DC permeability for solutes whose absorption ranges from 1% to 99%. The experimental human AM permeabilities determined by deconvolution are similar to those determined by direct human jejunal perfusion. The small intestinal pH varies with position and the results are interpreted in terms of the pH dependent octanol partition. The permeability versus partition relations are presented separately for the uncharged, basic, acidic and charged solutes. The small uncharged solutes caffeine, acetaminophen and antipyrine have very high permeabilities (about 20 x 10-4 cm/sec) corresponding to an unstirred layer of only 45 μm. The weak acid aspirin also has a large AM permeability despite its low octanol partition at pH 7.4, suggesting

  9. Assessment of early chorionic circulation by three-dimensional power Doppler.

    PubMed

    Hafner, Tomislav; Kurjak, Asim; Funduk-Kurjak, Biserka; Bekavac, Ivanka

    2002-01-01

    Three-dimensional power Doppler sonography is a unique instrument that enables assessment of vascular signals within the whole investigated area. Hemodynamical changes included in the process of early placentation are one of the most exciting topics in investigation of early human development. This investigation was designed as an observational cross-sectional study. A group of 25 patients in gestational age five to eleven weeks were recruited for the study. After acquirement of the volume containing three-dimensional power Doppler data of the pregnant uterus, the signals belonging to the chorion were isolated. Vascular 3D measurements were undertaken through 3D color/power histogram and expressed by Vascularization Index (VI) and Vascularization Flow Index (VFI). Volume of the chorion increased exponentially throughout the observation period. The VI and VFI positively correlated with the crown-rump length and chorion volume, and showed gradual increment through the investigation period. This investigation produced results confirming gradual augmentation of the loci and intensity of the intervillous flow in pregnancies between five and eleven gestational weeks. PMID:11933653

  10. Quantitative Analysis of 3′-Hydroxynorcotinine in Human Urine

    PubMed Central

    Upadhyaya, Pramod

    2015-01-01

    Introduction: Based on previous metabolism studies carried out in patas monkeys, we hypothesized that urinary 3′-hydroxynorcotinine could be a specific biomarker for uptake and metabolism of the carcinogen N′-nitrosonornicotine in people who use tobacco products. Methods: We developed a method for quantitation of 3′-hydroxynorcotinine in human urine. [Pyrrolidinone-13C4]3′-hydroxynorcotinine was added to urine as an internal standard, the samples were treated with β-glucuronidase, partially purified by solid supported liquid extraction and quantified by liquid chromatography–electrospray ionization–tandem mass spectrometry. Results: The method was accurate (average accuracy = 102%) and precise (coefficient of variation = 5.6%) in the range of measurement. 3′-Hydroxynorcotinine was detected in 48 urine samples from smokers (mean 393±287 pmol/ml urine) and 12 samples from individuals who had stopped smoking and were using the nicotine patch (mean 658±491 pmol/ml urine), but not in any of 10 samples from nonsmokers. Conclusions: Since the amounts of 3′-hydroxynorcotinine found in smokers’ urine were approximately 50 times greater than the anticipated daily dose of N′-nitrosonornicotine, we concluded that it is a metabolite of nicotine or one of its metabolites, comprising perhaps 1% of nicotine intake in smokers. Therefore, it would not be suitable as a specific biomarker for uptake and metabolism of N′-nitrosonornicotine. Since 3′-hydroxynorcotinine has never been previously reported as a constituent of human urine, further studies are required to determine its source and mode of formation. PMID:25324430

  11. Quantitative Mapping of Human Cartilage at 3.0T

    PubMed Central

    Wang, Ligong; Regatte, Ravinder R.

    2014-01-01

    Rationale and Objectives The objectives of this study were to measure the parallel changes of transverse relaxation times (T2), spin-lattice relaxation time in the rotating frame (T1ρ), and the delayed gadolinium-enhanced magnetic resonance imaging of cartilage (dGEMRIC)-T1 mapping of human knee cartilage in detecting cartilage degeneration at 3.0T. Materials and Methods Healthy volunteers (n = 10, mean age 35.6 years) and patients (n = 10, mean age 65 years) with early knee osteoarthritis (OA) were scanned at 3.0T MR using an 8-channel phased array knee coil (transmit–receive). Quantitative assessment of T2, T1ρ, and dGEMRIC-T1 values (global and regional) were correlated between asymptomatic subjects and patients with OA. Results The average T2 (39 ± 2 milliseconds [mean ± standard deviation] vs. 47 ± 6 milliseconds, P < .0007) and T1ρ (48 ± 3 vs. 62 ± 8 milliseconds, P < .0002) values were all markedly increased in all patients with OA when compared to healthy volunteers. The average dGEMRIC-T1 (1244 ± 134 vs. 643 ± 227 milliseconds, P < .000002) value was sharply decreased after intravenous administration of gadolinium contrast agent in all patients with OA. Conclusions The research results showed that all the T2, T1ρ, and dGEMRIC-T1 relaxation times varied with the cartilage degeneration. The dGEMRIC-T1 and T1ρ relaxation times seem to be more sensitive than T2 in detecting early cartilage degeneration. The preliminary study demonstrated that the early biochemical changes in knee osteoarthritic patients could be detected noninvasively in in vivo using T1ρ and dGEMRIC-T1 mapping. PMID:24594416

  12. Quantitation of the human basal ganglia with Positron Emission Tomography

    SciTech Connect

    Bendriem, B.; Dewey, S.L.; Schlyer, D.J.; Wolf, A.P.; Volkow, N.D.

    1990-01-01

    The accurate measurement of the concentration of a radioisotope in small structures with PET requires a correction for quantitation loss due to the partial volume effect and the effect of scattered radiation. To evaluate errors associated with measures in the human basal ganglia (BG) we have built a unilateral model of the BG that we have inserted in a 20 cm cylinder. The recovery coefficient (RC = measured activity/true activity) for our BG phantom has been measured on a CTI tomograph (model 931-08/12) with different background concentrations (contrast) and at different axial locations in the gantry. The BG was visualized on 4 or 5 slices depending on its position in the gantry and on the contrast used. The RC was 0.75 with no background (contrast equal to 1.0). Increasing the relative radioactivity concentration in the background increased the RC from 0.75 to 2.00 when the contrast was {minus}0.7 (BG < Background). The RC was also affected by the size and the shape of the region of interest (ROI) used (RC from 0.75 to 0.67 with ROI size from 0.12 to 1.41 cm{sup 2}). These results show that accurate RC correction depends not only on the volume of the structure but also on its contrast with its surroundings as well as on the selection of the ROI. They also demonstrate that the higher the contrast the more sensitive to axial positioning PET measurements in the BG are. These data provide us with some information about the variability of PET measurements in small structure like the BG and we have proposed some strategies to improve the reproducibility. 18 refs., 3 figs., 5 tabs.

  13. Quantitative Analysis of Human Cancer Cell Extravasation Using Intravital Imaging.

    PubMed

    Willetts, Lian; Bond, David; Stoletov, Konstantin; Lewis, John D

    2016-01-01

    Metastasis, or the spread of cancer cells from a primary tumor to distant sites, is the leading cause of cancer-associated death. Metastasis is a complex multi-step process comprised of invasion, intravasation, survival in circulation, extravasation, and formation of metastatic colonies. Currently, in vitro assays are limited in their ability to investigate these intricate processes and do not faithfully reflect metastasis as it occurs in vivo. Traditional in vivo models of metastasis are limited by their ability to visualize the seemingly sporadic behavior of where and when cancer cells spread (Reymond et al., Nat Rev Cancer 13:858-870, 2013). The avian embryo model of metastasis is a powerful platform to study many of the critical steps in the metastatic cascade including the migration, extravasation, and invasion of human cancer cells in vivo (Sung et al., Nat Commun 6:7164, 2015; Leong et al., Cell Rep 8, 1558-1570, 2014; Kain et al., Dev Dyn 243:216-28, 2014; Leong et al., Nat Protoc 5:1406-17, 2010; Zijlstra et al., Cancer Cell 13:221-234, 2008; Palmer et al., J Vis Exp 51:2815, 2011). The chicken chorioallantoic membrane (CAM) is a readily accessible and well-vascularized tissue that surrounds the developing embryo. When the chicken embryo is grown in a shell-less, ex ovo environment, the nearly transparent CAM provides an ideal environment for high-resolution fluorescent microcopy approaches. In this model, the embryonic chicken vasculature and labeled cancer cells can be visualized simultaneously to investigate specific steps in the metastatic cascade including extravasation. When combined with the proper image analysis tools, the ex ovo chicken embryo model offers a cost-effective and high-throughput platform for the quantitative analysis of tumor cell metastasis in a physiologically relevant in vivo setting. Here we discuss detailed procedures to quantify cancer cell extravasation in the shell-less chicken embryo model with advanced fluorescence

  14. Vitelline envelope, chorion, and micropyle of Fundulus heteroclitus eggs

    SciTech Connect

    Dumont, J.N.; Brummet, A.R.

    1980-01-01

    The architecture and transformation of the vitelline envelope of the developing oocyte into the chorion of the mature egg of Fundulus heteroclitus have been examined by scanning and transmission electron microscopy. The mature vitelline envelope is structurally complex and consists of about nine strata. The envelope is penetrated by pore canals that contain microvilli arising from the oocyte and macrovilli from follicle cells. During the envelope's transformation into the chorion, the pore canals are lost and the envelope becomes more fibrous and compact and its stratified nature less apparent. The micropyle, or pore, through which the sperm gains access to the enclosed egg is located at the bottom of a small funnel-shaped depression in the envelope. Internally, the micropyle opens on the apex of a cone-like elevation of the chorion. During the development of the envelope, structured chorionic fibrils, the components of which are presumed to be synthesized by the follicle cells, become attached to its surface. These chorionic fibrils are thought to aid in the attachment of the egg to the substratum and perhaps to help prevent water loss during low tides when the egg may be exposed.

  15. Visualization of Drosophila melanogaster chorion genes undergoing amplification

    SciTech Connect

    Osheim, Y.N.; Miller, O.L. Jr.; Beyer, A.L.

    1988-07-01

    The authors visualized by electron microscopy the preferential amplification of Drosophila chorion genes in late-stage follicle cells. Chromatin spreads revealed large clusters of actively transcribed genes of the appropriate size, spacing, and orientation for chorion genes that were expressed with the correct temporal specificity. Occasionally the active genes were observed within or contiguous with intact replicons and replication forks. In every case, our micrographs are consistent with the hypothesis that the central region of each chorion domain contains a replication origin(s) used during the amplification event. In one case, a small replication bubble was observed precisely at the site of the essential region of the X chromosome amplification control element. The micrographs also suggest that forks at either end of a replicon frequently progress very different distances, presumably due to different times in initiation or different rates of movement. It appears that all chorion genes (even those coding for minor proteins) are transcribed in a ''fully on'' condition, albeit for varied durations, and that if replication fork passage does inactivate a promoter, it does so very transiently. Furthermore, a DNA segment containing one active gene is likely to have an additional active gene(s). Surprisingly, during the time frame of expected maximum activity, approximately half of the chorion sequences appear transciptionally inactive.

  16. Quantitative assessment of human motion using video motion analysis

    NASA Technical Reports Server (NTRS)

    Probe, John D.

    1990-01-01

    In the study of the dynamics and kinematics of the human body, a wide variety of technologies was developed. Photogrammetric techniques are well documented and are known to provide reliable positional data from recorded images. Often these techniques are used in conjunction with cinematography and videography for analysis of planar motion, and to a lesser degree three-dimensional motion. Cinematography has been the most widely used medium for movement analysis. Excessive operating costs and the lag time required for film development coupled with recent advances in video technology have allowed video based motion analysis systems to emerge as a cost effective method of collecting and analyzing human movement. The Anthropometric and Biomechanics Lab at Johnson Space Center utilizes the video based Ariel Performance Analysis System to develop data on shirt-sleeved and space-suited human performance in order to plan efficient on orbit intravehicular and extravehicular activities. The system is described.

  17. Quantitative mapping of intracellular cations in the human amniotic membrane

    NASA Astrophysics Data System (ADS)

    Moretto, Ph.; Llabador, Y.; Simonoff, M.; Razafindrabe, L.; Bara, M.; Guiet-Bara, A.

    1993-05-01

    The effect of magnesium and taurine on the permeability of cell membranes to monovalent cations has been investigated using the Bordeaux nuclear microprobe. PIXE and RBS techniques have been used to provide quantitative measurements and ion distributions in the isolated amniotic membrane. This physiological model for cellular exchanges allowed us to reveal the distribution of most elements involved in cellular pathways and the modifications under different experimental conditions of incubation in physiological fluids. The PIXE microanalysis provided an original viewpoint on these mechanisms. Following this first study, the amnion compact lamina was found to play a role which was not, up to now, taken into account in the interpretation of electrophysiological experimentations. The release of some ionic species, such as K +, from the epithelial cells, during immersion in isotonic fluids, could have been hitherto underestimated.

  18. Quantitative assessment of human motion using video motion analysis

    NASA Technical Reports Server (NTRS)

    Probe, John D.

    1993-01-01

    In the study of the dynamics and kinematics of the human body a wide variety of technologies has been developed. Photogrammetric techniques are well documented and are known to provide reliable positional data from recorded images. Often these techniques are used in conjunction with cinematography and videography for analysis of planar motion, and to a lesser degree three-dimensional motion. Cinematography has been the most widely used medium for movement analysis. Excessive operating costs and the lag time required for film development, coupled with recent advances in video technology, have allowed video based motion analysis systems to emerge as a cost effective method of collecting and analyzing human movement. The Anthropometric and Biomechanics Lab at Johnson Space Center utilizes the video based Ariel Performance Analysis System (APAS) to develop data on shirtsleeved and space-suited human performance in order to plan efficient on-orbit intravehicular and extravehicular activities. APAS is a fully integrated system of hardware and software for biomechanics and the analysis of human performance and generalized motion measurement. Major components of the complete system include the video system, the AT compatible computer, and the proprietary software.

  19. Raman spectroscopy of human skin: looking for a quantitative algorithm to reliably estimate human age.

    PubMed

    Pezzotti, Giuseppe; Boffelli, Marco; Miyamori, Daisuke; Uemura, Takeshi; Marunaka, Yoshinori; Zhu, Wenliang; Ikegaya, Hiroshi

    2015-06-01

    The possibility of examining soft tissues by Raman spectroscopy is challenged in an attempt to probe human age for the changes in biochemical composition of skin that accompany aging. We present a proof-of-concept report for explicating the biophysical links between vibrational characteristics and the specific compositional and chemical changes associated with aging. The actual existence of such links is then phenomenologically proved. In an attempt to foster the basics for a quantitative use of Raman spectroscopy in assessing aging from human skin samples, a precise spectral deconvolution is performed as a function of donors' ages on five cadaveric samples, which emphasizes the physical significance and the morphological modifications of the Raman bands. The outputs suggest the presence of spectral markers for age identification from skin samples. Some of them appeared as authentic "biological clocks" for the apparent exactness with which they are related to age. Our spectroscopic approach yields clear compositional information of protein folding and crystallization of lipid structures, which can lead to a precise identification of age from infants to adults. Once statistically validated, these parameters might be used to link vibrational aspects at the molecular scale for practical forensic purposes. PMID:26112367

  20. Raman spectroscopy of human skin: looking for a quantitative algorithm to reliably estimate human age

    NASA Astrophysics Data System (ADS)

    Pezzotti, Giuseppe; Boffelli, Marco; Miyamori, Daisuke; Uemura, Takeshi; Marunaka, Yoshinori; Zhu, Wenliang; Ikegaya, Hiroshi

    2015-06-01

    The possibility of examining soft tissues by Raman spectroscopy is challenged in an attempt to probe human age for the changes in biochemical composition of skin that accompany aging. We present a proof-of-concept report for explicating the biophysical links between vibrational characteristics and the specific compositional and chemical changes associated with aging. The actual existence of such links is then phenomenologically proved. In an attempt to foster the basics for a quantitative use of Raman spectroscopy in assessing aging from human skin samples, a precise spectral deconvolution is performed as a function of donors' ages on five cadaveric samples, which emphasizes the physical significance and the morphological modifications of the Raman bands. The outputs suggest the presence of spectral markers for age identification from skin samples. Some of them appeared as authentic "biological clocks" for the apparent exactness with which they are related to age. Our spectroscopic approach yields clear compositional information of protein folding and crystallization of lipid structures, which can lead to a precise identification of age from infants to adults. Once statistically validated, these parameters might be used to link vibrational aspects at the molecular scale for practical forensic purposes.

  1. PCR detection and quantitation of predominant anaerobic bacteria in human and animal fecal samples

    SciTech Connect

    Wang, Rong-Fu; Cao, Wei-Wen; Cerniglia, C.E.

    1996-04-01

    PCR procedures based on 16S rRNA genen sequence specific for 12 anaerobic bacteria that predominate in the human intestinal tract were developed and used for quantitative detection of these species in human feces and animal feces. The reported PCR procedure including the fecal sample preparation method is simplified and rapid and eliminates the DNA isolation steps.

  2. A quantitative method for studying human arterial baroreflexes

    NASA Technical Reports Server (NTRS)

    Eckberg, Dwain L.; Fritsch, Janice M.; Goble, Ross L.

    1991-01-01

    A new system is described that delivers precise, stereotyped pressure changes to the human neck and elicits neurally-mediated heart rate changes. The centerpiece of this system is a Silastic chamber that is strapped to the anterior neck. This chamber is connected to a stepping-motor-controlled bellows assembly. A strain-gauge transducer measures the intensity of pressure changes. The entire system is controlled by microprocessors, and both stimuli and responses are displayed on a digital oscilloscope. The end-product of this system is a reproducible baroreceptor stimulus-cardiac response relation that can be recorded rapidly and safely in astronauts in space.

  3. Sensitivity of quantitative sensory models to morphine analgesia in humans

    PubMed Central

    Olesen, Anne Estrup; Brock, Christina; Sverrisdóttir, Eva; Larsen, Isabelle Myriam; Drewes, Asbjørn Mohr

    2014-01-01

    Introduction Opioid analgesia can be explored with quantitative sensory testing, but most investigations have used models of phasic pain, and such brief stimuli may be limited in the ability to faithfully simulate natural and clinical painful experiences. Therefore, identification of appropriate experimental pain models is critical for our understanding of opioid effects with the potential to improve treatment. Objectives The aim was to explore and compare various pain models to morphine analgesia in healthy volunteers. Methods The study was a double-blind, randomized, two-way crossover study. Thirty-nine healthy participants were included and received morphine 30 mg (2 mg/mL) as oral solution or placebo. To cover both tonic and phasic stimulations, a comprehensive multi-modal, multi-tissue pain-testing program was performed. Results Tonic experimental pain models were sensitive to morphine analgesia compared to placebo: muscle pressure (F=4.87, P=0.03), bone pressure (F=3.98, P=0.05), rectal pressure (F=4.25, P=0.04), and the cold pressor test (F=25.3, P<0.001). Compared to placebo, morphine increased tolerance to muscle stimulation by 14.07%; bone stimulation by 9.72%; rectal mechanical stimulation by 20.40%, and reduced pain reported during the cold pressor test by 9.14%. In contrast, the more phasic experimental pain models were not sensitive to morphine analgesia: skin heat, rectal electrical stimulation, or rectal heat stimulation (all P>0.05). Conclusion Pain models with deep tonic stimulation including C fiber activation and and/or endogenous pain modulation were more sensitive to morphine analgesia. To avoid false negative results in future studies, we recommend inclusion of reproducible tonic pain models in deep tissues, mimicking clinical pain to a higher degree. PMID:25525384

  4. 21 CFR 522.1081 - Chorionic gonadotropin.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... fish intended for human consumption, the total dose administered per fish (all injections combined... repeated in 14 days if the animal's behavior or examination of the ovaries per rectum indicates...

  5. 21 CFR 522.1081 - Chorionic gonadotropin.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... fish intended for human consumption, the total dose administered per fish (all injections combined... repeated in 14 days if the animal's behavior or examination of the ovaries per rectum indicates...

  6. 21 CFR 522.1081 - Chorionic gonadotropin.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... fish intended for human consumption, the total dose administered per fish (all injections combined... repeated in 14 days if the animal's behavior or examination of the ovaries per rectum indicates...

  7. Development of ovine chorionic somatomammotropin hormone-deficient pregnancies.

    PubMed

    Baker, Callie M; Goetzmann, Lindsey N; Cantlon, Jeremy D; Jeckel, Kimberly M; Winger, Quinton A; Anthony, Russell V

    2016-05-01

    Intrauterine growth restriction (IUGR) is a leading cause of neonatal mortality and morbidity. Chorionic somatomammotropin hormone (CSH), a placenta-specific secretory product found at high concentrations in maternal and fetal circulation throughout gestation, is significantly reduced in human and sheep IUGR pregnancies. The objective of this study was to knock down ovine CSH (oCSH) expression in vivo using lentiviral-mediated short-hairpin RNA to test the hypothesis that oCSH deficiency would result in IUGR of near-term fetal lambs. Three different lentiviral oCSH-targeting constructs were used and compared with pregnancies (n = 8) generated with a scrambled control (SC) lentiviral construct. Pregnancies were harvested at 135 days of gestation. The most effective targeting sequence, "target 6" (tg6; n = 8), yielded pregnancies with significant reductions (P ≤ 0.05) in oCSH mRNA (50%) and protein (38%) concentrations, as well as significant reductions (P ≤ 0.05) in placental (52%) and fetal (32%) weights compared with the SC pregnancies. Fetal liver weights were reduced 41% (P ≤ 0.05), yet fetal liver insulin-like growth factor-I (oIGF1) and -II mRNA concentrations were reduced (P ≤ 0.05) 82 and 71%, respectively, and umbilical artery oIGF1 concentrations were reduced 62% (P ≤ 0.05) in tg6 pregnancies. Additionally, fetal liver oIGF-binding protein (oIGFBP) 2 and oIGFBP3 mRNA concentrations were reduced (P ≤ 0.05), whereas fetal liver oIGFBP1 mRNA concentration was not impacted nor was maternal liver oIGF and oIGFBP mRNA concentrations or uterine artery oIGF1 concentrations (P ≥ 0.10). Based on our results, it appears that oCSH deficiency does result in IUGR, by impacting placental development as well as fetal liver development and function. PMID:26887431

  8. Quantitative Anatomy of the Growing Lungs in the Human Fetus

    PubMed Central

    Szpinda, Michał; Siedlaczek, Waldemar; Szpinda, Anna; Woźniak, Alina; Mila-Kierzenkowska, Celestyna; Badura, Mateusz

    2015-01-01

    Using anatomical, digital, and statistical methods we examined the three-dimensional growth of the lungs in 67 human fetuses aged 16–25 weeks. The lung dimensions revealed no sex differences. The transverse and sagittal diameters and the base circumference were greater in the right lungs while the lengths of anterior and posterior margins and the lung height were greater in the left lungs. The best-fit curves for all the lung parameters were natural logarithmic models. The transverse-to-sagittal diameter ratio remained stable and averaged 0.56 ± 0.08 and 0.52 ± 0.08 for the right and left lungs, respectively. For the right and left lungs, the transverse diameter-to-height ratio significantly increased from 0.74 ± 0.09 to 0.92 ± 0.08 and from 0.56 ± 0.07 to 0.79 ± 0.09, respectively. The sagittal diameter-to-height ratio significantly increased from 1.41 ± 0.23 to 1.66 ± 0.18 in the right lung, and from 1.27 ± 0.17 to 1.48 ± 0.22 in the left lung. In the fetal lungs, their proportionate increase in transverse and sagittal diameters considerably accelerates with relation to the lung height. The lung dimensions in the fetus are relevant in the evaluation of the normative pulmonary growth and the diagnosis of pulmonary hypoplasia. PMID:26413517

  9. Low reversibility of intracellular cAMP accumulation in mouse Leydig tumor cells (MLTC-1) stimulated by human Luteinizing Hormone (hLH) and Chorionic Gonadotropin (hCG).

    PubMed

    Klett, Danièle; Meslin, Philippine; Relav, Lauriane; Nguyen, Thi Mong Diep; Mariot, Julie; Jégot, Gwenhaël; Cahoreau, Claire; Combarnous, Yves

    2016-10-15

    In order to study the intracellular cAMP response kinetics of Leydig cells to hormones with LH activity, we used MLTC-1 cells transiently expressing a chimeric cAMP-responsive luciferase so that real-time variations of intracellular cAMP concentration could be followed using oxiluciferin luminescence produced from catalyzed luciferin oxidation. The potencies of the different LHs and CGs were evaluated using areas under the curves (AUC) of their kinetics over 60 min stimulation. All mammalian LHs and CGs tested were found to stimulate cAMP accumulation in these cells. The reversibility of this stimulation was studied by removing the hormone from the culture medium after 10 min of incubation. The ratios of kinetics AUC after removing or not the hormone were used to evaluate the stimulation reversibility of each hormone. Natural and recombinant hLHs and hCGs were found to exhibit slowly reversible activation compared to pituitary rat, ovine, porcine, camel and equine LHs, serum-derived eCG (PMSG) and recombinant eLH/CGs. Carbohydrate side chains are not involved in this phenomenon since natural and recombinant homologous hormones exhibit the same reversibility rates. It is still unknown whether only one human subunit, α or β, is responsible for this behaviour or whether it is due to a particular feature of the hLH and hCG quaternary structure. PMID:27373440

  10. New quantitative patterns of the growing trachea in human fetuses

    PubMed Central

    Szpinda, Michał; Daroszewski, Marcin; Szpinda, Anna; WoŸniak, Alina; Wiśniewski, Marcin; Mila-Kierzenkowska, Celestyna; Baumgart, Mariusz; Paruszewska-Achtel, Monika

    2012-01-01

    Summary Background Rapid progress in perinatal medicine has resulted in numerous tracheo-bronchial interventions on fetal and neonatal airways. The present study was performed to compile normative data for tracheal dimensions at varying gestational ages. Material/Methods Using anatomical dissection, digital image analysis (NIS-Elements BR 3.0) and statistical analysis (Wilcoxon signed-rank test, Student’s t test, one-way ANOVA, post-hoc Bonferroni test, linear and nonlinear regression analysis) a range of the 4 variables (length in mm, middle external transverse diameter in mm, proximal internal cross-sectional area in mm2, internal volume in mm3) for the trachea in 73 spontaneously aborted human fetuses (39 male, 34 female) aged 14–25 weeks was examined. Results No significant male-female differences were found (P>0.05). The length ranged from 10.37±2.15 to 26.54±0.26 mm as y=−65.098 + 28.796 × ln (Age) ±1.794 (R2=0.82). The middle external transverse diameter varied from 2.53±0.09 to 5.09±0.42 mm with the model y=−11.020 + 5.049 × ln (Age) ±0.330 (R2=0.81). The trachea indicated a proportional evolution because the middle external transverse diameter-to-length ratio was stable (0.23±0.03). The proximal internal cross-sectional area rose from 1.46±0.04 to 5.76±1.04 mm2 as y=−3.562 + 0.352 × Age ±0.519 (R2=0.76). The internal volumetric growth from 11.89±2.49 to 119.63±4.95 mm3 generated the function y=−135.248 + 9.919 × Age ±10.478 (R2=0.86). Conclusions The growth in both length and middle external transverse diameter of the trachea follows logarithmic functions, whereas growth of both its proximal internal cross-sectional area and internal volume follow linear functions. The length and middle external transverse diameter of the trachea develop proportionally to each other. The tracheal dimensions may be helpful in the prenatal diagnosis and monitoring of tracheal malformations and obstructive anomalies of the upper respiratory tract

  11. Regulation of 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4 isomerase expression and activity in the hypophysectomized rat ovary: Interactions between the stimulatory effect of human chorionic gonadotropin and the luteolytic effect of prolactin

    SciTech Connect

    Martel, C.; Labrie, C.; Dupont, E.; Couet, J.; Trudel, C.; Rheaume, E.; Simard, J.; Luu-The, V.; Pelletier, G.; Labrie, F. )

    1990-12-01

    The enzyme 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4 isomerase (3 beta-HSD) catalyzes an obligatory step in the conversion of pregnenolone and other 5-ene-3 beta-hydroxysteroids into progesterone as well as precursors of all androgens and estrogens in the ovary. Since 3 beta-HSD is likely to be an important target for regulation by pituitary hormones, we have studied the effect of chronic treatment with LH (hCG), FSH, and PRL on ovarian 3 beta-HSD expression and activity in hypophysectomized adult female rats. Human CG (hCG) (10 IU, twice a day (bid)), ovine FSH (0.5 microgram, bid), and ovine PRL (1 mg, bid) were administered, singly or in combination, for a period of 10 days starting 15 days after hypophysectomy. In hypophysectomized rats, PRL exerted a potent inhibitory effect on all the parameters studied. In fact, PRL caused a 81% decrease in ovarian 3 beta-HSD mRNA content accompanied by a similar decrease in 3 beta-HSD activity and protein levels. In addition, ovarian weight decreased by 40% whereas serum progesterone fell dramatically from 1.92 nmol/liter to undetectable levels after treatment with PRL. Whereas hCG alone had only slight stimulatory effects on 3 beta-HSD mRNA, protein content and activity levels, treatment with the gonadotropin partially or completely reversed the potent inhibitory effects of oPRL on all the parameters measured. FSH, on the other hand, had no significant effect on 3 beta-HSD expression and activity. In situ hybridization experiments using the 35S-labeled rat ovary 3 beta-HSD cDNA probe show that the inhibitory effect of PRL is exerted primarily on luteal cell 3 beta-HSD expression and activity. On the other hand, it can be seen that hCG stimulates 3 beta-HSD mRNA accumulation in interstitial cells.

  12. Recombinant thyrotropin containing a beta-subunit chimera with the human chorionic gonadotropin-beta carboxy-terminus is biologically active, with a prolonged plasma half-life: role of carbohydrate in bioactivity and metabolic clearance.

    PubMed

    Joshi, L; Murata, Y; Wondisford, F E; Szkudlinski, M W; Desai, R; Weintraub, B D

    1995-09-01

    Recombinant TSH is now successfully being used in clinical studies of thyroid cancer. Because of its therapeutic potential, we have constructed a longer acting analog of TSH by fusing the carboxy-terminal extension peptide (CTEP) of hCG beta onto TSH beta. When coexpressed either with alpha-subunit complementary DNA or alpha minigene in African green monkey (COS-7) and human embryonic kidney (293) cells, the chimera was fully bioactive in vitro and exhibited enhanced in vivo potency associated with a prolonged plasma half-life. The addition of 25 amino acids with 4 O-linked oligosaccharide chains did not affect the assembly and secretion of chimeric TSH. Wild-type (WT) and chimeric TSH secreted by COS-7 and 293 cells displayed wide differences in their plasma half-lives, presumably due to the presence of terminal sialic acid and SO4 on their oligosaccharide chains, respectively. Chimeric and WT TSH secreted by both cell lines demonstrated similar bioactivity in cAMP production, with some differences in [3H]thymidine incorporation. Chimeric TSH appears to be more effective in COS-7 cells than in 293 cells, as judged by growth assay. COS-7-produced chimeric TSH showed the maximum increase in half-life, indicating the importance of sialic acid in prolonging half-life and in vivo potency. Sulfation of both subunits, predominantly beta and to a lesser extent alpha, appears to be responsible at least in part for the increased metabolic clearance of WT and chimeric TSH secreted by 293 cells. Apart from its therapeutic potential, chimeric TSH produced in various cell lines can be used as a tool to delineate the roles of sulfate and sialic acid in the in vivo clearance and, thereby, the in vivo bioactivity. PMID:7544273

  13. 77 FR 55481 - Withdrawal of Approval of New Animal Drug Applications; Chorionic Gonadotropin; Naloxone...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-09-10

    ...; Chorionic Gonadotropin; Naloxone; Oxymorphone; Oxytocin AGENCY: Food and Drug Administration, HHS. ACTION... Pharmaceuticals hydrochloride) Inc., 100 Painters Injection. Dr., Chadds Ford, PA 19317. 046-822 VETOCIN...

  14. QUANTITATIVE EVALUATION OF BROMODICHLOROMETHANE METABOLISM BY RECOMBINANT RAT AND HUMAN CYTOCHROME P450S

    EPA Science Inventory

    ABSTRACT
    We report quantitative estimates of the parameters for metabolism of bromodichloromethane (BDCM) by recombinant preparations of hepatic cytochrome P450s (CYPs) from rat and human. BDCM is a drinking water disinfectant byproduct that has been implicated in liver, kidn...

  15. WORKSHOP ON THE QUALITATIVE AND QUANTITATIVE COMPARABILITY OF HUMAN AND ANIMAL DEVELOPMENTAL NEUROTOXICITY: SUMMARY AND IMPLICATIONS

    EPA Science Inventory

    The Workshop on the Qualitative and Quantitative Comparability of Human and Animal Developmental Neurotoxicity was convened by the U.S. Environmental Protection Agency (EPA) and the National Institute on Drug Abuse to address issues related to when testing should be required, wha...

  16. A Quantitative Literacy Course for Humanities and Law Students: The Challenges of a Context-Based Curriculum

    ERIC Educational Resources Information Center

    Frith, Vera

    2012-01-01

    This article examines some aspects of the effectiveness of a first-year course in quantitative literacy for Humanities and Law students at a South African university. This intervention is necessary to assist students in developing the appropriate quantitative competencies because there is an articulation gap between the quantitative literacy of…

  17. Prediction of human pharmacokinetics from preclinical information: comparative accuracy of quantitative prediction approaches.

    PubMed

    Hosea, Natilie A; Collard, Wendy T; Cole, Susan; Maurer, Tristan S; Fang, Rick X; Jones, Hannah; Kakar, Shefali M; Nakai, Yasuhiro; Smith, Bill J; Webster, Rob; Beaumont, Kevin

    2009-05-01

    Quantitative prediction of human pharmacokinetics is critical in assessing the viability of drug candidates and in determining first-in-human dosing. Numerous prediction methodologies, incorporating both in vitro and preclinical in vivo data, have been developed in recent years, each with advantages and disadvantages. However, the lack of a comprehensive data set, both preclinical and clinical, has limited efforts to evaluate the optimal strategy (or strategies) that results in quantitative predictions of human pharmacokinetics. To address this issue, the authors conducted a retrospective analysis using 50 proprietary compounds for which in vitro, preclinical pharmacokinetic data and oral single-dose human pharmacokinetic data were available. Five predictive strategies, involving either allometry or use of unbound intrinsic clearance from microsomes or hepatocytes, were then compared for their ability to predict human oral clearance, half-life through predictions of systemic clearance, volume of distribution, and bioavailability. Use of a single-species scaling approach with rat, dog, or monkey was as accurate as or more accurate than using multiple-species allometry. For those compounds cleared almost exclusively by P450-mediated pathways, scaling from human liver microsomes was as predictive as single-species scaling of clearance based on data from rat, dog, or monkey. These data suggest that use of predictive methods involving either single-species in vivo data or in vitro human liver microsomes can quantitatively predict human in vivo pharmacokinetics and suggest the possibility of streamlining the predictive methodology through use of a single species or use only of human in vitro microsomal preparations. PMID:19299532

  18. Primary human chorionic gonadotropin secreting germinoma of the corpus callosum

    PubMed Central

    Chuan Aaron, Foo Song; Dawn, Chong Q. Q.; Kenneth, Chang T. E.; Hoe, Ng Wai; Yen, Soh Shui; Chee Kian, Tham

    2013-01-01

    Background: Primary intracranial germinomas are a rare subset of intracranial tumors derived from mis-incorporated germ cells within the folding neural plate during embryogenesis. Though known to arise from midline structures in the central nervous system (CNS), occurrence within the corpus callosum is exceedingly rare. Case Description: We present a rare case of secreting primary intracranial germinoma with extensive intraventricular metastasis presenting as a multi-cystic butterfly lesion in the genu of the corpus callosum in a young boy. Conclusion: Intracranial germ cell tumors must be considered for any multi-cystic lesion arising from midline structures in the CNS in the preadult population. PMID:24233184

  19. [Demonstration and evolution of chorionic gonadotropin in ewes].

    PubMed

    Lacroix, M C; Martal, J

    1979-02-26

    Occurrence of ovine Chorionic Gonadotropin (oCG) is demonstrated in placenta and amniotic fluid with the use of a radioreceptor assay (corpus luteum membranes) in ewes. Identification of oCG is possible as early as 15th day of pregnancy. It should be secreted at a constant rate, and its maximum concentration is recorded on the 130th day. This hormonal factor might be one of the major components accountable for high progesteronemia observed during the 100 last days of pregnancy. PMID:110484

  20. Initial description of a quantitative, cross-species (chimpanzee-human) social responsiveness measure

    PubMed Central

    Marrus, Natasha; Faughn, Carley; Shuman, Jeremy; Petersen, Steve; Constantino, John; Povinelli, Daniel; Pruett, John R.

    2011-01-01

    Objective Comparative studies of social responsiveness, an ability that is impaired in autistic spectrum disorders, can inform our understanding of both autism and the cognitive architecture of social behavior. Because there is no existing quantitative measure of social responsiveness in chimpanzees, we generated a quantitative, cross-species (human-chimpanzee) social responsiveness measure. Method We translated the Social Responsiveness Scale (SRS), an instrument that quantifies human social responsiveness, into an analogous instrument for chimpanzees. We then retranslated this "Chimp SRS" into a human "Cross-Species SRS" (XSRS). We evaluated three groups of chimpanzees (n=29) with the Chimp SRS and typical and autistic spectrum disorder (ASD) human children (n=20) with the XSRS. Results The Chimp SRS demonstrated strong inter-rater reliability at the three sites (ranges for individual ICCs: .534–.866 and mean ICCs: .851–.970). As has been observed in humans, exploratory principal components analysis of Chimp SRS scores supports a single factor underlying chimpanzee social responsiveness. Human subjects' XSRS scores were fully concordant with their SRS scores (r=.976, p=.001) and distinguished appropriately between typical and ASD subjects. One chimpanzee known for inappropriate social behavior displayed a significantly higher score than all other chimpanzees at its site, demonstrating the scale's ability to detect impaired social responsiveness in chimpanzees. Conclusion Our initial cross-species social responsiveness scale proved reliable and discriminated differences in social responsiveness across (in a relative sense) and within (in a more objectively quantifiable manner) humans and chimpanzees. PMID:21515200

  1. Selection and sequence analysis of a cDNA clone encoding a known chorion protein of the A family.

    PubMed Central

    Tsitilou, S G; Regier, J C; Kafatos, F C

    1980-01-01

    Using as criteria the size, abundance and developmental specificity of hybridizing mRNA sequences, we have selected from our chorion cDNA library a clone corresponding to a specific chorion protein, A4--cl. Comparison between the clone sequence and the largely known sequence of A4--cl validates the use of the cDNA library for sequence analysis of the chorion multigene families. The two major chorion protein families, A and B, share certain structural similarities. Images PMID:7433133

  2. A qualitative and quantitative comparison of the fat in human, feline and canine kidneys.

    PubMed Central

    Bell, J.; Scott, G. B.

    1977-01-01

    While xanthogranulomatous pyelonephritis in humans tends to be rich in birefringent fat, this variety cannot be demonstrated microscopically in the feline morphological counterpart of the condition, which is, however, rich in readily stainable fat. Since normal feline kidney is rich in such lipid, a qualitative and quantitative comparison of the lipid in human, feline and canine kidney was carried out in an attempt to throw further light on the possible origin of the birefringent fat in the human disease. No significant difference could be found in the amount of cholesterol in the 3 species. Despite its visual prominence in feline kidneys, human kidney was richer in neutral fat and the percentage of total lipid formed by cholesterol was greater in humans than in the other species. The results suggested that cases of xanthogranulomatous pyelonephritis may occur in those kidneys unusually rich in cholesterol. Images Fig. 1 Fig. 2 PMID:836763

  3. Identification of Novel Tumor-Associated Cell Surface Sialoglycoproteins in Human Glioblastoma Tumors Using Quantitative Proteomics

    PubMed Central

    Autelitano, François; Loyaux, Denis; Roudières, Sébastien; Déon, Catherine; Guette, Frédérique; Fabre, Philippe; Ping, Qinggong; Wang, Su; Auvergne, Romane; Badarinarayana, Vasudeo; Smith, Michael; Guillemot, Jean-Claude; Goldman, Steven A.; Natesan, Sridaran; Ferrara, Pascual; August, Paul

    2014-01-01

    Glioblastoma multiform (GBM) remains clinical indication with significant “unmet medical need”. Innovative new therapy to eliminate residual tumor cells and prevent tumor recurrences is critically needed for this deadly disease. A major challenge of GBM research has been the identification of novel molecular therapeutic targets and accurate diagnostic/prognostic biomarkers. Many of the current clinical therapeutic targets of immunotoxins and ligand-directed toxins for high-grade glioma (HGG) cells are surface sialylated glycoproteins. Therefore, methods that systematically and quantitatively analyze cell surface sialoglycoproteins in human clinical tumor samples would be useful for the identification of potential diagnostic markers and therapeutic targets for malignant gliomas. In this study, we used the bioorthogonal chemical reporter strategy (BOCR) in combination with label-free quantitative mass spectrometry (LFQ-MS) to characterize and accurately quantify the individual cell surface sialoproteome in human GBM tissues, in fetal, adult human astrocytes, and in human neural progenitor cells (NPCs). We identified and quantified a total of 843 proteins, including 801 glycoproteins. Among the 843 proteins, 606 (72%) are known cell surface or secreted glycoproteins, including 156 CD-antigens, all major classes of cell surface receptor proteins, transporters, and adhesion proteins. Our findings identified several known as well as new cell surface antigens whose expression is predominantly restricted to human GBM tumors as confirmed by microarray transcription profiling, quantitative RT-PCR and immunohistochemical staining. This report presents the comprehensive identification of new biomarkers and therapeutic targets for the treatment of malignant gliomas using quantitative sialoglycoproteomics with clinically relevant, patient derived primary glioma cells. PMID:25360666

  4. QUANTITATIVE EVALUATION OF PUTATIVE HUMAN CARCINOGENS AND RELATED CHEMICALS ON HUMAN FORESKIN FIBROBLASTS

    EPA Science Inventory

    We have evaluated 12 compounds representative of diverse classes of chemicals for their cytotoxicity and transforming ability of human skin fibroblasts in vitro in the presence and absence of human liver S-9 mix. n the absence of the human liver S-9 mix, only seven of the 12 comp...

  5. Quantitative expression profile of hepatobiliary transporters in sandwich cultured rat and human hepatocytes.

    PubMed

    Li, Na; Bi, Yi-An; Duignan, David B; Lai, Yurong

    2009-01-01

    As sandwich cultured (SC) hepatocytes can repolarize to form bile canalicular networks, allowing active excretion of compounds in a vectorial manner, the model has been widely used for assessing the transporter related complexity of ADME/tox issues. A lack of quantitative information on transporter expression during cell culture has made in vitro to in vivo extrapolation of hepatobiliary transport difficult. In the present study, using our newly developed LC-MS/MS absolute quantitative methods, we determined the quantitative expression profile of three biliary transporters in SC rat and human hepatocytes. A significant shift of hepatobiliary transporter proteins was observed both in human and rat sandwich cultures. A decrease of BSEP/Bsep protein and an increase of BCRP/Bcrp protein were detected in both rat and human hepatocytes over time in culture. Interestingly, Mrp2 in rat hepatocytes was significantly diminished, while MRP2 constantly increased in human hepatocytes during the cell culture. Consequently, the interspecies difference between rat and human in absolute amount of MRP2/Mrp2 was minimized over time in culture. Following the sandwich culture, the species difference of hepatobiliary transporter protein between human and rat at day 5 post SC was diminished (MRP2/Mrp2), identical (BSEP/Bsep) or reversed (BCRP/Bcrp), compared to the in vivo situation. In addition, the absolute protein amount of BCRP/Bcrp or MRP2/Mrp2 was proportionally correlated with the intrinsic biliary clearance estimated in various lots of SC rat and human hepatocytes. The results revealed that absolute protein amount is a key determinant for hepatobiliary clearance and could provide fundamental support on extrapolation of biliary secretion from in vitro to in vivo. PMID:19545175

  6. Quantitative Risk Assessment of Human Trichinellosis Caused by Consumption of Pork Meat Sausages in Argentina.

    PubMed

    Sequeira, G J; Zbrun, M V; Soto, L P; Astesana, D M; Blajman, J E; Rosmini, M R; Frizzo, L S; Signorini, M L

    2016-03-01

    In Argentina, there are three known species of genus Trichinella; however, Trichinella spiralis is most commonly associated with domestic pigs and it is recognized as the main cause of human trichinellosis by the consumption of products made with raw or insufficiently cooked pork meat. In some areas of Argentina, this disease is endemic and it is thus necessary to develop a more effective programme of prevention and control. Here, we developed a quantitative risk assessment of human trichinellosis following pork meat sausage consumption, which may be used to identify the stages with greater impact on the probability of acquiring the disease. The quantitative model was designed to describe the conditions in which the meat is produced, processed, transported, stored, sold and consumed in Argentina. The model predicted a risk of human trichinellosis of 4.88 × 10(-6) and an estimated annual number of trichinellosis cases of 109. The risk of human trichinellosis was sensitive to the number of Trichinella larvae that effectively survived the storage period (r = 0.89), the average probability of infection (PPinf ) (r = 0.44) and the storage time (Storage) (r = 0.08). This model allowed assessing the impact of different factors influencing the risk of acquiring trichinellosis. The model may thus help to select possible strategies to reduce the risk in the chain of by-products of pork production. PMID:26227185

  7. Quantitative 3D Tracing of Gene-delivery Viral Vectors in Human Cells and Animal Tissues

    PubMed Central

    Xiao, Ping-Jie; Li, Chengwen; Neumann, Aaron; Samulski, R Jude

    2012-01-01

    Trafficking through a variety of cellular structures and organelles is essential for the interaction between gene-delivery vectors (i.e., adeno-associated virus (AAV) and liposomes) and host cells/tissues. Here, we present a method of computer-assisted quantitative 3D biodistribution microscopy that samples the whole population of fluorescently-labeled vectors and document their trafficking routes. Using AAV as a working model, we first experimentally defined numerical parameters for the singularity of Cy5-labeled particles by combining confocal microscopy and atomic force microscopy (AFM). We then developed a robust approach that integrates single-particle fluorescence imaging with 3D deconvolution and isosurface rendering to quantitate viral distribution and trafficking in human cells as well as animal tissues at the single-particle level. Using this quantitative method, we uncovered an as yet uncharacterized rate-limiting step during viral cell entry, while delineating nuclear accumulation of virions during the first 8 hours postinfection. Further, our studies revealed for the first time that following intramuscular injection, AAV spread progressively across muscle tissues through endomysium between myofibers instead of traversing through target cells. Such 3D resolution and quantitative dissection of vector–host interactions at the subcellular level should significantly improve our ability to resolve trafficking mechanisms of gene-delivery particles and facilitate the development of enhanced viral vectors. PMID:22108857

  8. Sources of Technical Variability in Quantitative LC-MS Proteomics: Human Brain Tissue Sample Analysis.

    SciTech Connect

    Piehowski, Paul D.; Petyuk, Vladislav A.; Orton, Daniel J.; Xie, Fang; Moore, Ronald J.; Ramirez Restrepo, Manuel; Engel, Anzhelika; Lieberman, Andrew P.; Albin, Roger L.; Camp, David G.; Smith, Richard D.; Myers, Amanda J.

    2013-05-03

    To design a robust quantitative proteomics study, an understanding of both the inherent heterogeneity of the biological samples being studied as well as the technical variability of the proteomics methods and platform is needed. Additionally, accurately identifying the technical steps associated with the largest variability would provide valuable information for the improvement and design of future processing pipelines. We present an experimental strategy that allows for a detailed examination of the variability of the quantitative LC-MS proteomics measurements. By replicating analyses at different stages of processing, various technical components can be estimated and their individual contribution to technical variability can be dissected. This design can be easily adapted to other quantitative proteomics pipelines. Herein, we applied this methodology to our label-free workflow for the processing of human brain tissue. For this application, the pipeline was divided into four critical components: Tissue dissection and homogenization (extraction), protein denaturation followed by trypsin digestion and SPE clean-up (digestion), short-term run-to-run instrumental response fluctuation (instrumental variance), and long-term drift of the quantitative response of the LC-MS/MS platform over the 2 week period of continuous analysis (instrumental stability). From this analysis, we found the following contributions to variability: extraction (72%) >> instrumental variance (16%) > instrumental stability (8.4%) > digestion (3.1%). Furthermore, the stability of the platform and its’ suitability for discovery proteomics studies is demonstrated.

  9. How to perform transabdominal chorionic villus sampling: a practical guideline.

    PubMed

    Monni, Giovanni; Pagani, Giorgio; Stagnati, Valentina; Iuculano, Ambra; Ibba, Rosa Maria

    2016-05-01

    The spread of both first trimester screening for chromosomal abnormalities and the possibility to check for single gene disorders at DNA-analysis has increased the request for chorionic villus sampling (CVS) in the first trimester. In order to perform placental biopsy, two routes are possible: the transcervical (TC) and the transabdominal (TA). In early days, the trancervical technique was the most diffused, but since its introduction into clinical practice, the TA technique has become the approach of choice in detriment of the TC technique. In our institution, we have a 30-year experience in TA-CVS with more than 26 000 procedures performed. Considering the expertise and the volume of procedures undertaken at our unit, we suggest a practical guideline for novel operators in TA-CVS. PMID:26372474

  10. Bleeding as a consequence of chorion villus sampling.

    PubMed

    Liu, D T; Jeavons, B; Preston, C; Slater, E; Symonds, E M

    1989-03-01

    A series of 4 separate studies were conducted to assess the incidence and short term consequence of bleeding associated with chorion villus sampling. Results support previous reports that risk of foetal-maternal transfusion as suggested by a rise in maternal serum alpha-fetoprotein (MSAFP) can occur. This occurrence is not consistent and need not be obvious even after therapeutic abortion. It is also transient and did not complicate mid-trimester neural tube screening or subsequent course of pregnancy. Eighty-seven percent of blood contaminating villus samples are of maternal origin. Following diagnosis 37% of patients reported some vaginal bleeding. This is mainly in the form of spotting which did not preclude normal pregnancy. Foetal loss occurred in 4 of the patients when bleeding considered heavier than spotting continued. In rhesus negative patients prophylactic anti-D gamma-globulin is advised, since neither Kleihauer counts nor MSAFP estimation reliably detect all foetal-maternal transfusions. PMID:2472129

  11. A quantitative collagen fibers orientation assessment using birefringence measurements: calibration and application to human osteons.

    PubMed

    Spiesz, Ewa M; Kaminsky, Werner; Zysset, Philippe K

    2011-12-01

    Even though mechanical properties depend strongly on the arrangement of collagen fibers in mineralized tissues, it is not yet well resolved. Only a few semi-quantitative evaluations of the fiber arrangement in bone, like spectroscopic techniques or circularly polarized light microscopy methods are available. In this study the out-of-plane collagen arrangement angle was calibrated to the linear birefringence of a longitudinally fibered mineralized turkey leg tendon cut at variety of angles to the main axis. The calibration curve was applied to human cortical bone osteons to quantify the out-of-plane collagen fibers arrangement. The proposed calibration curve is normalized to sample thickness and wavelength of the probing light to enable a universally applicable quantitative assessment. This approach may improve our understanding of the fibrillar structure of bone and its implications on mechanical properties. PMID:21970947

  12. Characterizing Aging in the Human Brainstem Using Quantitative Multimodal MRI Analysis

    PubMed Central

    Lambert, Christian; Chowdhury, Rumana; FitzGerald, Thomas H. B.; Fleming, Stephen M.; Lutti, Antoine; Hutton, Chloe; Draganski, Bogdan; Frackowiak, Richard; Ashburner, John

    2013-01-01

    Aging is ubiquitous to the human condition. The MRI correlates of healthy aging have been extensively investigated using a range of modalities, including volumetric MRI, quantitative MRI (qMRI), and diffusion tensor imaging. Despite this, the reported brainstem related changes remain sparse. This is, in part, due to the technical and methodological limitations in quantitatively assessing and statistically analyzing this region. By utilizing a new method of brainstem segmentation, a large cohort of 100 healthy adults were assessed in this study for the effects of aging within the human brainstem in vivo. Using qMRI, tensor-based morphometry (TBM), and voxel-based quantification (VBQ), the volumetric and quantitative changes across healthy adults between 19 and 75 years were characterized. In addition to the increased R2* in substantia nigra corresponding to increasing iron deposition with age, several novel findings were reported in the current study. These include selective volumetric loss of the brachium conjunctivum, with a corresponding decrease in magnetization transfer and increase in proton density (PD), accounting for the previously described “midbrain shrinkage.” Additionally, we found increases in R1 and PD in several pontine and medullary structures. We consider these changes in the context of well-characterized, functional age-related changes, and propose potential biophysical mechanisms. This study provides detailed quantitative analysis of the internal architecture of the brainstem and provides a baseline for further studies of neurodegenerative diseases that are characterized by early, pre-clinical involvement of the brainstem, such as Parkinson’s and Alzheimer’s diseases. PMID:23970860

  13. Quantitative imaging of protein targets in the human brain with PET

    NASA Astrophysics Data System (ADS)

    Gunn, Roger N.; Slifstein, Mark; Searle, Graham E.; Price, Julie C.

    2015-11-01

    PET imaging of proteins in the human brain with high affinity radiolabelled molecules has a history stretching back over 30 years. During this period the portfolio of protein targets that can be imaged has increased significantly through successes in radioligand discovery and development. This portfolio now spans six major categories of proteins; G-protein coupled receptors, membrane transporters, ligand gated ion channels, enzymes, misfolded proteins and tryptophan-rich sensory proteins. In parallel to these achievements in radiochemical sciences there have also been significant advances in the quantitative analysis and interpretation of the imaging data including the development of methods for image registration, image segmentation, tracer compartmental modeling, reference tissue kinetic analysis and partial volume correction. In this review, we analyze the activity of the field around each of the protein targets in order to give a perspective on the historical focus and the possible future trajectory of the field. The important neurobiology and pharmacology is introduced for each of the six protein classes and we present established radioligands for each that have successfully transitioned to quantitative imaging in humans. We present a standard quantitative analysis workflow for these radioligands which takes the dynamic PET data, associated blood and anatomical MRI data as the inputs to a series of image processing and bio-mathematical modeling steps before outputting the outcome measure of interest on either a regional or parametric image basis. The quantitative outcome measures are then used in a range of different imaging studies including tracer discovery and development studies, cross sectional studies, classification studies, intervention studies and longitudinal studies. Finally we consider some of the confounds, challenges and subtleties that arise in practice when trying to quantify and interpret PET neuroimaging data including motion artifacts

  14. Quantitative imaging of protein targets in the human brain with PET.

    PubMed

    Gunn, Roger N; Slifstein, Mark; Searle, Graham E; Price, Julie C

    2015-11-21

    PET imaging of proteins in the human brain with high affinity radiolabelled molecules has a history stretching back over 30 years. During this period the portfolio of protein targets that can be imaged has increased significantly through successes in radioligand discovery and development. This portfolio now spans six major categories of proteins; G-protein coupled receptors, membrane transporters, ligand gated ion channels, enzymes, misfolded proteins and tryptophan-rich sensory proteins. In parallel to these achievements in radiochemical sciences there have also been significant advances in the quantitative analysis and interpretation of the imaging data including the development of methods for image registration, image segmentation, tracer compartmental modeling, reference tissue kinetic analysis and partial volume correction. In this review, we analyze the activity of the field around each of the protein targets in order to give a perspective on the historical focus and the possible future trajectory of the field. The important neurobiology and pharmacology is introduced for each of the six protein classes and we present established radioligands for each that have successfully transitioned to quantitative imaging in humans. We present a standard quantitative analysis workflow for these radioligands which takes the dynamic PET data, associated blood and anatomical MRI data as the inputs to a series of image processing and bio-mathematical modeling steps before outputting the outcome measure of interest on either a regional or parametric image basis. The quantitative outcome measures are then used in a range of different imaging studies including tracer discovery and development studies, cross sectional studies, classification studies, intervention studies and longitudinal studies. Finally we consider some of the confounds, challenges and subtleties that arise in practice when trying to quantify and interpret PET neuroimaging data including motion artifacts

  15. Quantitative Determination of Common Urinary Odorants and Their Glucuronide Conjugates in Human Urine

    PubMed Central

    Wagenstaller, Maria; Buettner, Andrea

    2013-01-01

    Our previous study on the identification of common odorants and their conjugates in human urine demonstrated that this substance fraction is a little-understood but nonetheless a promising medium for analysis and diagnostics in this easily accessible physiological medium. Smell as an indicator for diseases, or volatile excretion in the course of dietary processes bares high potential for a series of physiological insights. Still, little is known today about the quantitative composition of odorous or volatile targets, as well as their non-volatile conjugates, both with regard to their common occurrence in urine of healthy subjects, as well as in that of individuals suffering from diseases or other physiological misbalancing. Accordingly, the aim of our study was to develop a highly sensitive and selective approach to determine the common quantitative composition of selected odorant markers in healthy human subjects, as well as their corresponding glucuronide conjugates. We used one- and two-dimensional high resolution gas chromatography-mass spectrometry in combination with stable isotope dilution assays to quantify commonly occurring and potent odorants in human urine. The studies were carried out on both native urine and on urine that had been treated by glucuronidase assays, with analysis of the liberated odor-active compounds using the same techniques. Analytical data are discussed with regard to their potential translation as future diagnostic tool. PMID:24958143

  16. Two-dimensional electrophoresis and computer imaging: quantitation of human milk casein.

    PubMed

    Goldfarb, M

    1999-01-01

    Because human casein does not precipitate from milk at its isoelectric point as does bovine casein, there is no easy method of quantitation. Casein represents only approximately 30% of the protein fraction in human milk, and the complex methods necessary for isolation cannot be used easily with small samples in a survey of a large number of mothers. Two-dimensional electrophoresis coupled with computer imaging has the potential to compare and quantitate proteins expeditiously using a small sample size. IsoDalt, a denaturing methodology, separates the casein micelle into its component parts, beta-casein, kappa-casein, parakappa-casein and casomorphins. Identification of these spots was made by immunoassay of a Western blot with monoclonal anti-human casein. Two spots at 24 kDa and 26 kDa, thought to be phosphorylated isomers of beta casein, were selected for quantitation. Milk samples from 20 mothers, 8 weeks post partum, were run on two-dimensional (2-D) gels; a slide was taken of each silverstained gel with a Kodak control strip; the slide was scanned into powerMac Photoshop 3 with a Polaroid-Sprintscan; spots were isolated using "threshold", "mask" with IPTK (Imaging Processing Tool Kit, Reindeer Games) a Photoshop plug-in, and transferred to the NIH-Image program. Using an NIH-Image gel macro (Thomas Seebacher), the area and integrated density of the spots were measured. The Kodak control scale provided calibration and conversion to OD units. Visual scanning of the gels and computer units indicated a wide range of concentrations. To understand the range in units of weight, a standard was generated using bovine alpha casein (Sigma). Measurements will be used in a statistical program, Statview (Abacus), in an attempt to correlate information from a questionaire with casein concentration. PMID:10344261

  17. Metabolic remodeling of the human red blood cell membrane measured by quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Park, YongKeun; Best, Catherine; Auth, Thorsten; Gov, Nir S.; Safran, Samuel; Popescu, Gabriel

    2011-02-01

    We have quantitatively and systemically measured the morphologies and dynamics of fluctuations in human RBC membranes using a full-field laser interferometry technique that accurately measures dynamic membrane fluctuations. We present conclusive evidence that the presence of adenosine 5'-triphosphate (ATP) facilitates nonequilibrium dynamic fluctuations in the RBC membrane and that these fluctuations are highly correlated with specific regions in the biconcave shape of RBCs. Spatial analysis reveals that these nonequilibrium membrane fluctuations are enhanced at the scale of the spectrin mesh size. Our results indicate the presence of dynamic remodeling in the RBC membrane cortex powered by ATP, which results in nonequilibrium membrane fluctuations.

  18. Quantitative values of blood flow through the human forearm, hand, and finger as functions of temperature

    NASA Technical Reports Server (NTRS)

    Montgomery, L. D.

    1974-01-01

    A literature search was made to obtain values of human forearm, hand and finger blood flow as functions of environmental temperature. The sources used include both government and laboratory reports and the research presented in the open literature. An attempt was made to review many of the more quantitative noninvasive determinations and to collate the results in such a way as to yield blood flow values for each body segment as continuous functions of temperature. A brief review of the various ways used to measure blood flow is included along with an abstract of each work from which data was taken.

  19. Bacteria Localization and Chorion Thinning among Preterm Premature Rupture of Membranes

    PubMed Central

    Fortner, Kimberly B.; Grotegut, Chad A.; Ransom, Carla E.; Bentley, Rex C.; Feng, Liping; Lan, Lan; Heine, R. Phillips; Seed, Patrick C.; Murtha, Amy P.

    2014-01-01

    Objective Bacterial colonization of the fetal membranes and its role in pathogenesis of membrane rupture is poorly understood. Prior retrospective work revealed chorion layer thinning in preterm premature rupture of membranes (PPROM) subjects. Our objective was to prospectively examine fetal membrane chorion thinning and to correlate to bacterial presence in PPROM, preterm, and term subjects. Study Design Paired membrane samples (membrane rupture and membrane distant) were prospectively collected from: PPROM = 14, preterm labor (PTL = 8), preterm no labor (PTNL = 8), term labor (TL = 10), and term no labor (TNL = 8), subjects. Sections were probed with cytokeratin to identify fetal trophoblast layer of the chorion using immunohistochemistry. Fluorescence in situ hybridization was performed using broad range 16 s ribosomal RNA probe. Images were evaluated, chorion and choriodecidua were measured, and bacterial fluorescence scored. Chorion thinning and bacterial presence were compared among and between groups using Student's t-test, linear mixed effect model, and Poisson regression model (SAS Cary, NC). Results In all groups, the fetal chorion cellular layer was thinner at rupture compared to distant site (147.2 vs. 253.7 µm, p<0.0001). Further, chorion thinning was greatest among PPROM subjects compared to all other groups combined, regardless of site sampled [PPROM(114.9) vs. PTL(246.0) vs. PTNL(200.8) vs. TL(217.9) vs. TNL(246.5)]. Bacteria counts were highest among PPROM subjects compared to all other groups regardless of site sampled or histologic infection [PPROM(31) vs. PTL(9) vs. PTNL(7) vs. TL(7) vs. TNL(6)]. Among all subjects at both sites, bacterial counts were inversely correlated with chorion thinning, even excluding histologic chorioamnionitis (p<0.0001 and p = 0.05). Conclusions Fetal chorion was uniformly thinner at rupture site compared to distant sites. In PPROM fetal chorion, we demonstrated pronounced global thinning

  20. A general approach for the purification and quantitative glycomic analysis of human plasma.

    PubMed

    Tep, Samnang; Hincapie, Marina; Hancock, William S

    2012-03-01

    The development of a general method for the purification and quantitative glycomic analysis of human plasma samples to characterize global glycosylation changes shall be presented. The method involves multiple steps, including the depletion of plasma via multi-affinity chromatography to remove high abundant proteins, the enrichment of the lower abundant glycoproteins via multi-lectin affinity chromatography, the isotopic derivatization of released glycans, and quantitative analysis by MALDI-TOF MS. Isotopic derivatization of glycans is accomplished using the well-established chemistry of reductive amination to derivatize glycans with either a light analog ((12)C anthranilic acid) or a heavy analog ((13)C(7) anthranilic acid), which allows for the direct comparison of the alternately labeled glycans by MALDI-TOF MS. The method displays a tenfold linear dynamic range for both neutral and sialylated glycans with sub-picomolar sensitivity. Additionally, by using anthranilic acid, a very sensitive fluorophore, as the derivatization reagent, the glycans can be analyzed by chromatography with fluorescence detection. The utility of this methodology is highlighted by the many diseases and disorders that are known to either show or be the result of changes in glycosylation. A method that provides a generic approach for sample preparation and quantitative data will help to further advance the field of glycomics. PMID:22274286

  1. Dynamic quantitative photothermal monitoring of cell death of individual human red blood cells upon glucose depletion

    NASA Astrophysics Data System (ADS)

    Vasudevan, Srivathsan; Chen, George Chung Kit; Andika, Marta; Agarwal, Shuchi; Chen, Peng; Olivo, Malini

    2010-09-01

    Red blood cells (RBCs) have been found to undergo ``programmed cell death,'' or eryptosis, and understanding this process can provide more information about apoptosis of nucleated cells. Photothermal (PT) response, a label-free photothermal noninvasive technique, is proposed as a tool to monitor the cell death process of living human RBCs upon glucose depletion. Since the physiological status of the dying cells is highly sensitive to photothermal parameters (e.g., thermal diffusivity, absorption, etc.), we applied linear PT response to continuously monitor the death mechanism of RBC when depleted of glucose. The kinetics of the assay where the cell's PT response transforms from linear to nonlinear regime is reported. In addition, quantitative monitoring was performed by extracting the relevant photothermal parameters from the PT response. Twofold increases in thermal diffusivity and size reduction were found in the linear PT response during cell death. Our results reveal that photothermal parameters change earlier than phosphatidylserine externalization (used for fluorescent studies), allowing us to detect the initial stage of eryptosis in a quantitative manner. Hence, the proposed tool, in addition to detection of eryptosis earlier than fluorescence, could also reveal physiological status of the cells through quantitative photothermal parameter extraction.

  2. Rapid quantitation of human epididymis protein 4 in human serum by amplified luminescent proximity homogeneous immunoassay (AlphaLISA).

    PubMed

    Zhao, Hui; Lin, Guanfeng; Liu, Tiancai; Liang, Junyu; Ren, Zhiqi; Liang, Rongliang; Chen, Baihong; Huang, Wenhua; Wu, Yingsong

    2016-10-01

    A sensitive, rapid and homogeneous reaction measurement method for quantitation of human epididymis protein 4 (HE4) in human serum by amplified luminescent proximity homogeneous immunoassay (AlphaLISA) was described. Built on a sandwich-type immunoassay format, analytes in samples were captured by one biotinylated monoclonal antibody combining on the surface of streptavidin coated donor beads, and "sandwiched" by another monoclonal antibody coated on acceptor beads. The coefficient variations of the method were lower than 10%, and the recoveries were in the range of 90-110% for serum samples. A value of 0.88pmol/l was identified as the minimum detectable dose of the present method for HE4. Compared with the results from electrochemiluminescence immunoassay kit (Roche) in 170 serum samples, there was a satisfied correlation coefficient of 0.984. The present assay demonstrated high sensitivity, wider effective detection range and excellent reproducibility for quantitation of HE4 can be useful for early screening and prognosis evaluation of patients with ovarian cancer. PMID:27568283

  3. Placental mesenchymal dysplasia, a case of intrauterine sudden death of fetus with rupture of cirsoid periumbilical chorionic vessels

    PubMed Central

    2011-01-01

    We report a 32-year-old woman (1-gravid, 1-para) with a vesicular lesion in her uterus that was pointed out on ultrasound at 8 weeks' gestation. Amniocentesis at 15 weeks' gestation showed a normal female karyotype, 46XX. As the pregnancy advanced, the mole-like lesion became relatively reduced. Throughout gestation, the maternal human chorionic gonadotropin level was normal, but the serum alpha fetoprotein level rose as her pregnancy progressed. Her fetus did not exhibit any remarkable anomalies. The patient visited our hospital complaining of a diminished feeling of fetal movements at 36 weeks 5 days' gestation, and intrauterine fetal death (IUFD) was confirmed. She delivered a 2336-g female without any definite anomalies. A pathological examination led to a diagnosis of placental mesenchymal dysplasia, and androgenetic/biparental mosaicism in the placenta was identified using p57kip2 immunohistochemical staining. And it also revealed that the rupture of the cirsoid chorionic vessels had led to IUFD. PMID:21513565

  4. Multispectral quantitative phase imaging of human red blood cells using inexpensive narrowband multicolor LEDs.

    PubMed

    Dubey, Vishesh; Singh, Gyanendra; Singh, Veena; Ahmad, Azeem; Mehta, Dalip Singh

    2016-04-01

    We report multispectral phase-shifting interference microscopy for quantitative phase imaging of human red blood cells (RBCs). A wide range of wavelengths are covered by means of using multiple color light emitting diodes (LEDs) with narrow spectral bandwidth ranging from violet to deep red color. The multicolor LED light source was designed and operated sequentially, which works as a multispectral scanning light source. Corresponding to each color LED source, five phase-shifted interferograms were recorded sequentially for the measurement of phase maps, as well as the refractive index of RBCs within the entire visible region. The proposed technique provides information about the effect of wavelengths on the morphology and refractive index of human RBCs. The system does not require expensive multiple color filters or any wavelength scanning mechanism along with broadband light source. PMID:27139652

  5. Absolute Quantitation of Water and Metabolites in the Human Brain. I. Compartments and Water

    NASA Astrophysics Data System (ADS)

    Ernst, T.; Kreis, R.; Ross, B. D.

    A method is presented to determine the compartmentation of a localized region in the human brain in terms of CSF, tissue water, and an NMR-invisible rest, using a PRESS or STEAM sequence. Discrimination between CSF and tissue water is based on differences in their T2 relaxation times. The NMR-invisible compartment is assessed using an external standard. The composition of three regions in the human brain is determined. The CSF content of specific regions can be used to quantify cortical atrophy. The method provides a means for measuring the water content of brain tissue in vivo with a precision of 1.5%. After appropriate corrections, the results are in close agreement with biochemical values. The method has major applications in localized quantitative spectroscopy. The compartmentation model can be used to correct for the CSF content of the selected volume and to properly define and interconvert all major concentration units.

  6. Quantitative 3D molecular cutaneous absorption in human skin using label free nonlinear microscopy.

    PubMed

    Chen, Xueqin; Grégoire, Sébastien; Formanek, Florian; Galey, Jean-Baptiste; Rigneault, Hervé

    2015-02-28

    Understanding the penetration mechanisms of drugs into human skin is a key issue in pharmaceutical and cosmetics research. To date, the techniques available for percutaneous penetration of compounds fail to provide a quantitative 3D map of molecular concentration distribution in complex tissues as the detected microscopy images are an intricate combination of concentration distribution and laser beam attenuation upon deep penetration. Here we introduce and validate a novel framework for imaging and reconstructing molecular concentration within the depth of artificial and human skin samples. Our approach combines the use of deuterated molecular compounds together with coherent anti-Stokes Raman scattering spectroscopy and microscopy that permits targeted molecules to be unambiguously discriminated within skin layers. We demonstrate both intercellular and transcellular pathways for different active compounds, together with in-depth concentration profiles reflecting the detailed skin barrier architecture. This method provides an enabling platform for establishing functional activity of topically applied products. PMID:25550155

  7. Quantitation of Human Metallothionein Isoforms: A Family of Small, Highly Conserved, Cysteine-rich Proteins*

    PubMed Central

    Mehus, Aaron A.; Muhonen, Wallace W.; Garrett, Scott H.; Somji, Seema; Sens, Donald A.; Shabb, John B.

    2014-01-01

    Human metallothioneins (MTs) are important regulators of metal homeostasis and protectors against oxidative damage. Their altered mRNA expression has been correlated with metal toxicity and a variety of cancers. Current immunodetection methods lack the specificity to distinguish all 12 human isoforms. Each, however, can be distinguished by the mass of its acetylated, cysteine-rich, hydrophilic N-terminal tryptic peptides. These properties were exploited to develop a bottom-up MALDI-TOF/TOF-MS-based method for their simultaneous quantitation. Key features included enrichment of N-terminal acetylated peptides by strong cation exchange chromatography, optimization of C18 reversed-phase chromatography, and control of methionine oxidation. Combinations of nine isoforms were identified in seven cell lines and two tissues. Relative quantitation was accomplished by comparing peak intensities of peptides generated from pooled cytosolic proteins alkylated with 14N- or 15N-iodoacetamide. Absolute quantitation was achieved using 15N-iodoacetamide-labeled synthetic peptides as internal standards. The method was applied to the cadmium induction of MTs in human kidney HK-2 epithelial cells expressing recombinant MT-3. Seven isoforms were detected with abundances spanning almost 2 orders of magnitude and inductions up to 12-fold. The protein-to-mRNA ratio for MT-1E was one-tenth that of other MTs, suggesting isoform-specific differences in protein expression efficiency. Differential expression of MT-1G1 and MT-1G2 suggested tissue- and cell-specific alternative splicing for the MT-1G isoform. Protein expression of MT isoforms was also evaluated in human breast epithelial cancer cell lines. Estrogen-receptor-positive cell lines expressed only MT-2 and MT-1X, whereas estrogen-receptor-negative cell lines additionally expressed MT-1E. The combined expression of MT isoforms was 38-fold greater in estrogen-receptor-negative cell lines than in estrogen-receptor-positive cells. These

  8. Quantitative investigations on the human entorhinal area: left-right asymmetry and age-related changes.

    PubMed

    Heinsen, H; Henn, R; Eisenmenger, W; Götz, M; Bohl, J; Bethke, B; Lockemann, U; Püschel, K

    1994-08-01

    The total nerve cell numbers in the right and in the left human entorhinal areas have been calculated by volume estimations with the Cavalieri principle and by cell density determinations with the optical disector. Thick gallocyanin-stained serial frozen sections through the parahippocampal gyrus of 22 human subjects (10 female, 12 male) ranging from 18 to 86 years were analysed. The laminar composition of gallocyanin (Nissl)-stained sections could easily be compared with Braak's (1972, 1980) pigmentoarchitectonic study, and Braak's nomenclature of the entorhinal laminas was adopted. Cell-sparse laminae dissecantes can more clearly be distinguished in Nissl than in aldehydefuchsin preparations. These cell-poor dissecantes, lamina dissecans externa (dis-ext), lamina dissecans 1 (dis-1) and lamina dissecans 2 (dis-2), were excluded from nerve cell number determinations. An exact delineation of the entorhinal area is indispensable for any kind of quantitative investigation. We have defined the entorhinal area by the presence of pre-alpha cell clusters and the deeper layers of lamina principalis externa (pre-beta and gamma) separated from lamina principalis interna (pri) by lamina dissecans 1 (dis-1). The human entorhinal area is quantitatively characterized by a left-sided (asymmetric) higher pre-alpha cell number and an age-related nerve cell loss in pre as well as pri layers. At variance with other CNS cortical and subcortical structures, the neuronal number of the entorhinal area appears to decrease continuously from the earliest stages analysed, although a secular trend has to be considered. The asymmetry in pre-alpha cell number is discussed in the context of higher human mental capabilities, especially language. PMID:7818090

  9. Construction, complete sequence, and annotation of a BAC contig covering the silkworm chorion locus.

    PubMed

    Chen, Zhiwei; Nohata, Junko; Guo, Huizhen; Li, Shenglong; Liu, Jianqiu; Guo, Youbing; Yamamoto, Kimiko; Kadono-Okuda, Keiko; Liu, Chun; Arunkumar, Kallare P; Nagaraju, Javaregowda; Zhang, Yan; Liu, Shiping; Labropoulou, Vassiliki; Swevers, Luc; Tsitoura, Panagiota; Iatrou, Kostas; Gopinathan, Karumathil P; Goldsmith, Marian R; Xia, Qingyou; Mita, Kazuei

    2015-01-01

    The silkmoth chorion was studied extensively by F.C. Kafatos' group for almost 40 years. However, the complete structure of the chorion locus was not obtained in the genome sequence of Bombyx mori published in 2008 due to repetitive sequences, resulting in gaps and an incomplete view of the locus. To obtain the complete sequence of the chorion locus, expressed sequence tags (ESTs) derived from follicular epithelium cells were used as probes to screen a bacterial artificial chromosome (BAC) library. Seven BACs were selected to construct a contig which covered the whole chorion locus. By Sanger sequencing, we successfully obtained complete sequences of the chorion locus spanning 871,711 base pairs on chromosome 2, where we annotated 127 chorion genes. The dataset reported here will recruit more researchers to revisit one of the oldest model systems which has been used to study developmentally regulated gene expression. It also provides insights into egg development and fertilization mechanisms and is relevant to applications related to improvements in breeding procedures and transgenesis. PMID:26594380

  10. Construction, complete sequence, and annotation of a BAC contig covering the silkworm chorion locus

    PubMed Central

    Chen, Zhiwei; Nohata, Junko; Guo, Huizhen; Li, Shenglong; Liu, Jianqiu; Guo, Youbing; Yamamoto, Kimiko; Kadono-Okuda, Keiko; Liu, Chun; Arunkumar, Kallare P.; Nagaraju, Javaregowda; Zhang, Yan; Liu, Shiping; Labropoulou, Vassiliki; Swevers, Luc; Tsitoura, Panagiota; Iatrou, Kostas; Gopinathan, Karumathil P.; Goldsmith, Marian R.; Xia, Qingyou; Mita, Kazuei

    2015-01-01

    The silkmoth chorion was studied extensively by F.C. Kafatos’ group for almost 40 years. However, the complete structure of the chorion locus was not obtained in the genome sequence of Bombyx mori published in 2008 due to repetitive sequences, resulting in gaps and an incomplete view of the locus. To obtain the complete sequence of the chorion locus, expressed sequence tags (ESTs) derived from follicular epithelium cells were used as probes to screen a bacterial artificial chromosome (BAC) library. Seven BACs were selected to construct a contig which covered the whole chorion locus. By Sanger sequencing, we successfully obtained complete sequences of the chorion locus spanning 871,711 base pairs on chromosome 2, where we annotated 127 chorion genes. The dataset reported here will recruit more researchers to revisit one of the oldest model systems which has been used to study developmentally regulated gene expression. It also provides insights into egg development and fertilization mechanisms and is relevant to applications related to improvements in breeding procedures and transgenesis. PMID:26594380

  11. A Quantitative Prioritisation of Human and Domestic Animal Pathogens in Europe

    PubMed Central

    McIntyre, K. Marie; Setzkorn, Christian; Hepworth, Philip J.; Morand, Serge; Morse, Andrew P.; Baylis, Matthew

    2014-01-01

    Disease or pathogen risk prioritisations aid understanding of infectious agent impact within surveillance or mitigation and biosecurity work, but take significant development. Previous work has shown the H-(Hirsch-)index as an alternative proxy. We present a weighted risk analysis describing infectious pathogen impact for human health (human pathogens) and well-being (domestic animal pathogens) using an objective, evidence-based, repeatable approach; the H-index. This study established the highest H-index European pathogens. Commonalities amongst pathogens not included in previous surveillance or risk analyses were examined. Differences between host types (humans/animals/zoonotic) in pathogen H-indices were explored as a One Health impact indicator. Finally, the acceptability of the H-index proxy for animal pathogen impact was examined by comparison with other measures. 57 pathogens appeared solely in the top 100 highest H-indices (1) human or (2) animal pathogens list, and 43 occurred in both. Of human pathogens, 66 were zoonotic and 67 were emerging, compared to 67 and 57 for animals. There were statistically significant differences between H-indices for host types (humans, animal, zoonotic), and there was limited evidence that H-indices are a reasonable proxy for animal pathogen impact. This work addresses measures outlined by the European Commission to strengthen climate change resilience and biosecurity for infectious diseases. The results include a quantitative evaluation of infectious pathogen impact, and suggest greater impacts of human-only compared to zoonotic pathogens or scientific under-representation of zoonoses. The outputs separate high and low impact pathogens, and should be combined with other risk assessment methods relying on expert opinion or qualitative data for priority setting, or could be used to prioritise diseases for which formal risk assessments are not possible because of data gaps. PMID:25136810

  12. Quantitative imaging of the human upper airway: instrument design and clinical studies

    NASA Astrophysics Data System (ADS)

    Leigh, M. S.; Armstrong, J. J.; Paduch, A.; Sampson, D. D.; Walsh, J. H.; Hillman, D. R.; Eastwood, P. R.

    2006-08-01

    Imaging of the human upper airway is widely used in medicine, in both clinical practice and research. Common imaging modalities include video endoscopy, X-ray CT, and MRI. However, no current modality is both quantitative and safe to use for extended periods of time. Such a capability would be particularly valuable for sleep research, which is inherently reliant on long observation sessions. We have developed an instrument capable of quantitative imaging of the human upper airway, based on endoscopic optical coherence tomography. There are no dose limits for optical techniques, and the minimally invasive imaging probe is safe for use in overnight studies. We report on the design of the instrument and its use in preliminary clinical studies, and we present results from a range of initial experiments. The experiments show that the instrument is capable of imaging during sleep, and that it can record dynamic changes in airway size and shape. This information is useful for research into sleep disorders, and potentially for clinical diagnosis and therapies.

  13. Quantitative evaluation of unrestrained human gait on change in walking velocity.

    PubMed

    Makino, Yuta; Tsujiuchi, Nobutaka; Ito, Akihito; Koizumi, Takayuki; Nakamura, Shota; Matsuda, Yasushi; Tsuchiya, Youtaro; Hayashi, Yuichiro

    2014-01-01

    In human gait motion analysis, which is one useful method for efficient physical rehabilitation to define various quantitative evaluation indices, ground reaction force, joint angle and joint loads are measured during gait. To obtain these data as unrestrained gait measurement, a novel gait motion analysis system using mobile force plates and attitude sensors has been developed. On the other hand, a human maintains a high correlation among the motion of all joints during gait. The analysis of the correlation in the recorded joint motion extracts a few simultaneously activating segmental coordination patterns, and the structure of the intersegmental coordination is attracting attention to an expected relationship with a control strategy. However, when the evaluation method using singular value decomposition has been applied to joint angles of the lower limb as representative kinematic parameters, joint moments related to the rotational motion of the joints have not yet been considered. In this paper, joint moments as kinetic parameters applied on the lower limb during gait of a normal subject and a trans-femoral amputee are analyzed under change in walking velocity by the wearable gait motion analysis system, and the effectiveness for quantitatively evaluate the rotational motion pattern in the joints of the lower limb by using joint moments is validated. PMID:25570503

  14. Quantitative BOLD: Mapping of Human Cerebral Deoxygenated Blood Volume and Oxygen Extraction Fraction: Default State

    PubMed Central

    He, Xiang; Yablonskiy, Dmitriy A.

    2014-01-01

    Since Ogawa et al. (Proc Natl Acad Sci USA 1990;87:9868–9872) made the fundamental discovery of blood oxygenation level-dependent (BOLD) contrast in MRI, most efforts have been directed toward the study of dynamic BOLD (i.e., temporal changes in the MRI signal during changes in brain activity). However, very little progress has been made in elucidating the nature of BOLD contrast during the resting or baseline state of the brain, which is important for understanding normal human performance because it accounts for most of the enormous energy budget of the brain. It is also crucial for deciphering the consequences of baseline-state impairment by cerebral vascular diseases. The objective of this study was to develop a BOLD MR-based method that allows quantitative evaluation of tissue hemodynamic parameters, such as the blood volume, deoxyhemoglobin concentration, and oxygen extraction fraction (OEF). The proposed method, which we have termed quantitative BOLD (qBOLD), is based on an MR signal model that incorporates prior knowledge about brain tissue composition and considers signals from gray matter (GM), white matter (WM), cerebrospinal fluid (CSF), and blood. A 2D gradient-echo sampling of spin-echo (GESSE) pulse sequence is used for the acquisition of the MRI signal. The method is applied to estimate the hemodynamic parameters of the normal human brain in the baseline state. PMID:17191227

  15. High-Throughput Quantitative Lipidomics Analysis of Nonesterified Fatty Acids in Human Plasma.

    PubMed

    Christinat, Nicolas; Morin-Rivron, Delphine; Masoodi, Mojgan

    2016-07-01

    We present a high-throughput, nontargeted lipidomics approach using liquid chromatography coupled to high-resolution mass spectrometry for quantitative analysis of nonesterified fatty acids. We applied this method to screen a wide range of fatty acids from medium-chain to very long-chain (8 to 24 carbon atoms) in human plasma samples. The method enables us to chromatographically separate branched-chain species from their straight-chain isomers as well as separate biologically important ω-3 and ω-6 polyunsaturated fatty acids. We used 51 fatty acid species to demonstrate the quantitative capability of this method with quantification limits in the nanomolar range; however, this method is not limited only to these fatty acid species. High-throughput sample preparation was developed and carried out on a robotic platform that allows extraction of 96 samples simultaneously within 3 h. This high-throughput platform was used to assess the influence of different types of human plasma collection and preparation on the nonesterified fatty acid profile of healthy donors. Use of the anticoagulants EDTA and heparin has been compared with simple clotting, and only limited changes have been detected in most nonesterified fatty acid concentrations. PMID:27185515

  16. Changes in transmural distribution of myocardial perfusion assessed by quantitative intravenous myocardial contrast echocardiography in humans

    PubMed Central

    Fukuda, S; Muro, T; Hozumi, T; Watanabe, H; Shimada, K; Yoshiyama, M; Takeuchi, K; Yoshikawa, J

    2002-01-01

    Objective: To clarify whether changes in transmural distribution of myocardial perfusion under significant coronary artery stenosis can be assessed by quantitative intravenous myocardial contrast echocardiography (MCE) in humans. Methods: 31 patients underwent dipyridamole stress MCE and quantitative coronary angiography. Intravenous MCE was performed by continuous infusion of Levovist. Images were obtained from the apical four chamber view with alternating pulsing intervals both at rest and after dipyridamole infusion. Images were analysed offline by placing regions of interest over both endocardial and epicardial sides of the mid-septum. The background subtracted intensity versus pulsing interval plots were fitted to an exponential function, y = A (1 − e−βt), where A is plateau level and β is rate of rise. Results: Of the 31 patients, 16 had significant stenosis (> 70%) in the left anterior descending artery (group A) and 15 did not (group B). At rest, there were no differences in the A endocardial to epicardial ratio (A-EER) and β-EER between the two groups (mean (SD) 1.2 (0.6) v 1.2 (0.8) and 1.2 (0.7) v 1.1 (0.6), respectively, NS). During hyperaemia, β-EER in group A was significantly lower than that in group B (1.0 (0.5) v 1.4 (0.5), p < 0.05) and A-EER did not differ between the two groups (1.0 (0.5) v 1.2 (0.4), NS). Conclusions: Changes in transmural distribution of myocardial perfusion under significant coronary artery stenosis can be assessed by quantitative intravenous MCE in humans. PMID:12231594

  17. Quantitative Models of the Dose-Response and Time Course of Inhalational Anthrax in Humans

    PubMed Central

    Schell, Wiley A.; Bulmahn, Kenneth; Walton, Thomas E.; Woods, Christopher W.; Coghill, Catherine; Gallegos, Frank; Samore, Matthew H.; Adler, Frederick R.

    2013-01-01

    Anthrax poses a community health risk due to accidental or intentional aerosol release. Reliable quantitative dose-response analyses are required to estimate the magnitude and timeline of potential consequences and the effect of public health intervention strategies under specific scenarios. Analyses of available data from exposures and infections of humans and non-human primates are often contradictory. We review existing quantitative inhalational anthrax dose-response models in light of criteria we propose for a model to be useful and defensible. To satisfy these criteria, we extend an existing mechanistic competing-risks model to create a novel Exposure–Infection–Symptomatic illness–Death (EISD) model and use experimental non-human primate data and human epidemiological data to optimize parameter values. The best fit to these data leads to estimates of a dose leading to infection in 50% of susceptible humans (ID50) of 11,000 spores (95% confidence interval 7,200–17,000), ID10 of 1,700 (1,100–2,600), and ID1 of 160 (100–250). These estimates suggest that use of a threshold to human infection of 600 spores (as suggested in the literature) underestimates the infectivity of low doses, while an existing estimate of a 1% infection rate for a single spore overestimates low dose infectivity. We estimate the median time from exposure to onset of symptoms (incubation period) among untreated cases to be 9.9 days (7.7–13.1) for exposure to ID50, 11.8 days (9.5–15.0) for ID10, and 12.1 days (9.9–15.3) for ID1. Our model is the first to provide incubation period estimates that are independently consistent with data from the largest known human outbreak. This model refines previous estimates of the distribution of early onset cases after a release and provides support for the recommended 60-day course of prophylactic antibiotic treatment for individuals exposed to low doses. PMID:24058320

  18. Quantitative Phosphoproteomics Identifies Filaggrin and other Targets of Ionizing Radiation in a Human Skin Model

    SciTech Connect

    Yang, Feng; Waters, Katrina M.; Webb-Robertson, Bobbie-Jo M.; Sowa, Marianne B.; Freiin von Neubeck, Claere H.; Aldrich, Joshua T.; Markillie, Lye Meng; Wirgau, Rachel M.; Gristenko, Marina A.; Zhao, Rui; Camp, David G.; Smith, Richard D.; Stenoien, David L.

    2012-04-17

    Our objective here was to perform a quantitative phosphoproteomic study on a reconstituted human skin tissue to identify low and high dose ionizing radiation dependent signaling in a complex 3-dimensional setting. Application of an isobaric labeling strategy using sham and 3 radiation doses (3, 10, 200 cGy) resulted in the identification of 1113 unique phosphopeptides. Statistical analyses identified 151 phosphopeptides showing significant changes in response to radiation and radiation dose. Proteins responsible for maintaining skin structural integrity including keratins and desmosomal proteins (desmoglein, desmoplakin, plakophilin 1 and 2,) had altered phosphorylation levels following exposure to both low and high doses of radiation. A phosphorylation site present in multiple copies in the linker regions of human profilaggrin underwent the largest fold change. Increased phosphorylation of these sites coincided with altered profilaggrin processing suggesting a role for linker phosphorylation in human profilaggrin regulation. These studies demonstrate that the reconstituted human skin system undergoes a coordinated response to ionizing radiation involving multiple layers of the stratified epithelium that serve to maintain skin barrier functions and minimize the damaging consequences of radiation exposure.

  19. Quantitative study of impacts of climate change and human activities on runoff in Beiluohe River basin

    NASA Astrophysics Data System (ADS)

    Guo, A.; Huang, Q.; Wang, Y.

    2015-05-01

    In the Beiluohe River basin (BRB) runoff has been experiencing a significant reduction induced by climate change and human activities. This paper considers the impacts of climate change and human activities on runoff reduction. An improved empirical method for climate factors and runoff was developed to study the impacts quantitatively. Meanwhile climate elasticity of runoff, based on the Budyko hypothesis, was also adopted to study the impacts. Using runoff change points, series were divided into natural period (1960-1969) and impacted periods (1970-1994, 1995-2010). Results show that the methods used to quantify the contributions obtained different but close conclusions to one another. For 1970-1994, climate was the primary factor impacting runoff, compared with that for 1960-1969, with the contribution reaching around 70.84-83.42%, which was greater than human activities (16.58-29.16%). For 1995-2010, the role of human activities strengthened with the contribution around 62.58-65.07%, greater than climate changes, around 34.93-37.42%.

  20. Effects of Anti-Human Neutrophil Antibodies In Vitro. QUANTITATIVE STUDIES

    PubMed Central

    Boxer, Laurence A.; Stossel, Thomas P.

    1974-01-01

    Opsonic, antiphagocytic, cytotoxic, and metabolic effects of homologous and heterologous antibodies against human neutrophils were analyzed by means of quantitative assays to facilitate detection of antibody activity, and to probe membrane function of these cells. Normal human neutrophils were purified by gradient centrifugation, sensitized with heat-inactivated antineutrophil antisera, and incubated with rabbit alveolar macrophages in balanced salt solution containing nitroblue tetrazolium. The macrophages engulfed sensitized neutrophils and reduced nitroblue tetrazolium to formazan in phagocytic vacuoles. The initial rate of nitroblue tetrazolium reduction by macrophages ingesting the neutrophils was measured spectrophotometrically. Neutrophils treated with rabbit anti-human leukocyte antiserum or IgG, with sera from mothers of infants with neonatal isoimmune neutropenia, and with 27% of sera from frequently transfused patients promoted rapid rates of nitroblue tetrazolium reduction by alveolar macrophages. This indicates that antineutrophil antibodies without added complement opsonized neutrophils for ingestion by the macrophages. Some sera from frequently transfused patients with opsonic activity for certain donors' neutrophils did not agglutinate these neutrophils (44%), did not lyse them in the presence of fresh plasma (47%), and did not inhibit phagocytosis of particles by the neutrophils (26%). The reverse was not observed. The opsonic activity of antineutrophil antiserum appears to be the most sensitive and a quantitative means of detecting antibody activity in vitro. Low concentrations of rabbit anti-human leukocyte antisera or IgG stimulated the ingestion rate of unopsonized or opsonized particles by human neutrophils, and, as previously reported by others, enhanced rates of oxidation of [1-14C]glucose by the cells. High concentrations of the antisera or IgG inhibited ingestion. All concentrations of homologous antineutrophil antisera tested only

  1. A Quantitative Comparison of the Similarity between Genes and Geography in Worldwide Human Populations

    PubMed Central

    Wang, Chaolong; Zöllner, Sebastian; Rosenberg, Noah A.

    2012-01-01

    Multivariate statistical techniques such as principal components analysis (PCA) and multidimensional scaling (MDS) have been widely used to summarize the structure of human genetic variation, often in easily visualized two-dimensional maps. Many recent studies have reported similarity between geographic maps of population locations and MDS or PCA maps of genetic variation inferred from single-nucleotide polymorphisms (SNPs). However, this similarity has been evident primarily in a qualitative sense; and, because different multivariate techniques and marker sets have been used in different studies, it has not been possible to formally compare genetic variation datasets in terms of their levels of similarity with geography. In this study, using genome-wide SNP data from 128 populations worldwide, we perform a systematic analysis to quantitatively evaluate the similarity of genes and geography in different geographic regions. For each of a series of regions, we apply a Procrustes analysis approach to find an optimal transformation that maximizes the similarity between PCA maps of genetic variation and geographic maps of population locations. We consider examples in Europe, Sub-Saharan Africa, Asia, East Asia, and Central/South Asia, as well as in a worldwide sample, finding that significant similarity between genes and geography exists in general at different geographic levels. The similarity is highest in our examples for Asia and, once highly distinctive populations have been removed, Sub-Saharan Africa. Our results provide a quantitative assessment of the geographic structure of human genetic variation worldwide, supporting the view that geography plays a strong role in giving rise to human population structure. PMID:22927824

  2. A male-specific quantitative trait locus on 1p21 controlling human stature

    PubMed Central

    Sammalisto, S; Hiekkalinna, T; Suviolahti, E; Sood, K; Metzidis, A; Pajukanta, P; Lilja, H; Soro-Paavonen, A; Taskinen, M; Tuomi, T; Almgren, P; Orho-Melander, M; Groop, L; Peltonen, L; Perola, M

    2005-01-01

    Background: Many genome-wide scans aimed at complex traits have been statistically underpowered due to small sample size. Combining data from several genome-wide screens with comparable quantitative phenotype data should improve statistical power for the localisation of genomic regions contributing to these traits. Objective: To perform a genome-wide screen for loci affecting adult stature by combined analysis of four previously performed genome-wide scans. Methods: We developed a web based computer tool, Cartographer, for combining genetic marker maps which positions genetic markers accurately using the July 2003 release of the human genome sequence and the deCODE genetic map. Using Cartographer, we combined the primary genotype data from four genome-wide scans and performed variance components (VC) linkage analyses for human stature on the pooled dataset of 1417 individuals from 277 families and performed VC analyses for males and females separately. Results: We found significant linkage to stature on 1p21 (multipoint LOD score 4.25) and suggestive linkages on 9p24 and 18q21 (multipoint LOD scores 2.57 and 2.39, respectively) in males-only analyses. We also found suggestive linkage to 4q35 and 22q13 (multipoint LOD scores 2.18 and 2.85, respectively) when we analysed both females and males and to 13q12 (multipoint LOD score 2.66) in females-only analyses. Conclusions: We strengthened the evidence for linkage to previously reported quantitative trait loci (QTL) for stature and also found significant evidence of a novel male-specific QTL on 1p21. Further investigation of several interesting candidate genes in this region will help towards characterisation of this first sex-specific locus affecting human stature. PMID:15827092

  3. Quantitative Mapping of Collagen Fiber Orientation in Non-glaucoma and Glaucoma Posterior Human Sclerae

    PubMed Central

    Pijanka, Jacek K.; Coudrillier, Baptiste; Ziegler, Kimberly; Sorensen, Thomas; Meek, Keith M.; Nguyen, Thao D.; Quigley, Harry A.; Boote, Craig

    2012-01-01

    Purpose. The posterior sclera has a major biomechanical influence on the optic nerve head, and may therefore be important in glaucoma. Scleral material properties are influenced significantly by collagen fiber architecture. Here we quantitatively map fiber orientation in non-glaucoma and glaucoma posterior human sclerae. Methods. Wide-angle x-ray scattering quantified fiber orientation at 0.5-mm intervals across seven non-glaucoma post-mortem human sclerae, and five sclerae with glaucoma history and confirmed axon loss. Multiphoton microscopy provided semiquantitative depth-profiling in the peripapillary sclera. Results. Midposterior fiber orientation was either uniaxial (one preferred direction) or biaxial (two directions). The peripapillary sclera was characterized by a ring of fibers located mainly in the mid-/outer stromal depth and encompassing ∼50% of the total tissue thickness. Fiber anisotropy was 37% higher in the peripapillary sclera compared with midposterior, varied up to 4-fold with position around the scleral canal, and was consistently lowest in the superior-nasal quadrant. Mean fiber anisotropy was significantly lower in the superior-temporal (P < 0.01) and inferior-nasal (P < 0.05) peripapillary scleral quadrants in glaucoma compared with non-glaucoma eyes. Conclusions. The collagen fiber architecture of the posterior human sclera is highly anisotropic and inhomogeneous. Regional differences in peripapillary fiber anisotropy between non-glaucoma and glaucoma eyes may represent adaptive changes in response to elevated IOP and/or glaucoma, or baseline structural properties that associate with predisposition to glaucomatous axon damage. Quantitative fiber orientation data will benefit numerical eye models aimed at predicting the sclera's influence on nerve head biomechanics, and thereby its possible role in glaucoma. PMID:22786908

  4. Quantitative detection of human spermatogonia for optimization of spermatogonial stem cell culture

    PubMed Central

    Zheng, Y.; Thomas, A.; Schmidt, C.M.; Dann, C.T.

    2014-01-01

    STUDY QUESTION Can human spermatogonia be detected in long-term primary testicular cell cultures using validated, germ cell-specific markers of spermatogonia? SUMMARY ANSWER Germ cell-specific markers of spermatogonia/spermatogonial stem cells (SSCs) are detected in early (1–2 weeks) but not late (> 6 weeks) primary testicular cell cultures; somatic cell markers are detected in late primary testicular cell cultures. WHAT IS KNOWN ALREADY The development of conditions for human SSC culture is critically dependent on the ability to define cell types unequivocally and to quantify spermatogonia/SSCs. Growth by somatic cells presents a major challenge in the establishment of SSC cultures and therefore markers that define spermatogonia/SSCs, but are not also expressed by testicular somatic cells, are essential for accurate characterization of SSC cultures. STUDY DESIGN, SIZE, DURATION Testicular tissue from eight organ donors with normal spermatogenesis was used for assay validation and establishing primary testicular cell cultures. PARTICIPANTS/MATERIALS, SETTING, METHODS Immunofluorescence analysis of normal human testicular tissue was used to validate antibodies (UTF1, SALL4, DAZL and VIM) and then the antibodies were used to demonstrate that primary testicular cells cultured in vitro for 1–2 weeks were composed of somatic cells and rare germ cells. Primary testicular cell cultures were further characterized by comparing to testicular somatic cell cultures using quantitative reverse transcriptase PCR (UTF1, FGFR3, ZBTB16, GPR125, DAZL, GATA4 and VIM) and flow cytometry (CD9 and SSEA4). MAIN RESULTS AND THE ROLE OF CHANCE UTF1, FGFR3, DAZL and ZBTB16 qRT–PCR and SSEA4 flow cytometry were validated for the sensitive, quantitative and specific detection of germ cells. In contrast, GPR125 mRNA and CD9 were found to be not specific to germ cells because they were also expressed in testicular somatic cell cultures. While the germ cell-specific markers were detected in

  5. Quantitative detection of caffeine in human skin by confocal Raman spectroscopy--A systematic in vitro validation study.

    PubMed

    Franzen, Lutz; Anderski, Juliane; Windbergs, Maike

    2015-09-01

    For rational development and evaluation of dermal drug delivery, the knowledge of rate and extent of substance penetration into the human skin is essential. However, current analytical procedures are destructive, labor intense and lack a defined spatial resolution. In this context, confocal Raman microscopy bares the potential to overcome current limitations in drug depth profiling. Confocal Raman microscopy already proved its suitability for the acquisition of qualitative penetration profiles, but a comprehensive investigation regarding its suitability for quantitative measurements inside the human skin is still missing. In this work, we present a systematic validation study to deploy confocal Raman microscopy for quantitative drug depth profiling in human skin. After we validated our Raman microscopic setup, we successfully established an experimental procedure that allows correlating the Raman signal of a model drug with its controlled concentration in human skin. To overcome current drawbacks in drug depth profiling, we evaluated different modes of peak correlation for quantitative Raman measurements and offer a suitable operating procedure for quantitative drug depth profiling in human skin. In conclusion, we successfully demonstrate the potential of confocal Raman microscopy for quantitative drug depth profiling in human skin as valuable alternative to destructive state-of-the-art techniques. PMID:25828208

  6. Comparison of Complications of Chorionic Villus Sampling and Amniocentesis

    PubMed Central

    Shahbazian, Nahid; Barati, Mojgan; Arian, Parvin; Saadati, Najmie

    2012-01-01

    Background A significant number of pregnancies are associated with the cytogenetic abnormalities of the fetus. Amniocentesis and chorionic villus sampling (CVS) are procedures used for prenatal genetic diagnosis. In this study, we compare the safety and complications of mid-trimester amniocentesis and transabdominal CVS. Materials and Methods This analytic cross-sectional study was performed in 308 patients from 2.11.2007 to 26.10.2009. We had 155 cases of amniocentesis, which we performed in weeks 15-23 of pregnancy; and 153 cases of CVS, which we performed during weeks 10-14 of pregnancy. Results There were 2 cases (1.2%) of premature rupture of membrane (PROM) in amniocentesis which occurred 1 and 10 days after the procedure and caused pregnancy loss before 20 weeks. We had 1 case (0.7%) of abortion in CVS, which occurred 10 days after the procedure. Additionally, there was 1 case of amniotic fluid leakage (0.7%) in which, after admission to the hospital and observation, leakage was stopped and the pregnancy continued normally. Conclusion In this study, we had more complications with amniocentesis cases than CVS. CVS is a procedure performed in the earlier stages of pregnancy and its complications are less than amniocentesis. We suggest CVS to be the procedure of choice for genetic diagnosis. PMID:25210610

  7. A human fecal contamination index for ranking impaired recreational watersusing the HF183 quantitative real-time PCR method

    EPA Science Inventory

    Human fecal pollution of surface water remains a public health concern worldwide. As a result, there is a growing interest in the application of human-associated fecal source identification quantitative real-time PCR (qPCR) technologies for recreational water quality risk managem...

  8. MAMMALIAN CELL CULTURE ASSAY TO QUANTITATE CHEMICALLY INDUCED ANEUPLOIDY: USE OF A MONOCHROMOSOMAL HUMAN/MOUSE CELL HYBRID

    EPA Science Inventory

    A short-term assay utilizing a human/mouse monochromosomal hybrid cell line R3-5, to detect chemically induced aneuploidy in mammalian cells is described. A single human chromosome transferred into mouse cells was used as a cytogenetic marker to quantitate abnormal chromosome seg...

  9. Improved HF183 quantitative real-time PCR assay for characterization of human fecal pollution in ambient surface water samples

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Real-time quantitative PCR assays that target the human-associated HF183 bacterial cluster have been found to be some of the top performing methods for the characterization of human fecal pollution in ambient surface waters. The United States Environmental Protection Agency is planning to conduct a ...

  10. Improved HF183 quantitative real-time PCR assay for characterization of human fecal pollution in ambient surface water samples

    EPA Science Inventory

    Real-time quantitative PCR assays that target the human-associated HF183 bacterial cluster are considered to be some of the top performing methods for the characterization of human fecal pollution in ambient surface waters. In response, the United States Environmental Protectio...

  11. Quantitative and Qualitative Analysis of Transient Fetal Compartments during Prenatal Human Brain Development

    PubMed Central

    Vasung, Lana; Lepage, Claude; Radoš, Milan; Pletikos, Mihovil; Goldman, Jennifer S.; Richiardi, Jonas; Raguž, Marina; Fischi-Gómez, Elda; Karama, Sherif; Huppi, Petra S.; Evans, Alan C.; Kostovic, Ivica

    2016-01-01

    The cerebral wall of the human fetal brain is composed of transient cellular compartments, which show characteristic spatiotemporal relationships with intensity of major neurogenic events (cell proliferation, migration, axonal growth, dendritic differentiation, synaptogenesis, cell death, and myelination). The aim of the present study was to obtain new quantitative data describing volume, surface area, and thickness of transient compartments in the human fetal cerebrum. Forty-four postmortem fetal brains aged 13–40 postconceptional weeks (PCW) were included in this study. High-resolution T1 weighted MR images were acquired on 19 fetal brain hemispheres. MR images were processed using in-house software (MNI-ACE toolbox). Delineation of fetal compartments was performed semi-automatically by co-registration of MRI with histological sections of the same brains, or with the age-matched brains from Zagreb Neuroembryological Collection. Growth trajectories of transient fetal compartments were reconstructed. The composition of telencephalic wall was quantitatively assessed. Between 13 and 25 PCW, when the intensity of neuronal proliferation decreases drastically, the relative volume of proliferative (ventricular and subventricular) compartments showed pronounced decline. In contrast, synapse- and extracellular matrix-rich subplate compartment continued to grow during the first two trimesters, occupying up to 45% of telencephalon and reaching its maximum volume and thickness around 30 PCW. This developmental maximum coincides with a period of intensive growth of long cortico-cortical fibers, which enter and wait in subplate before approaching the cortical plate. Although we did not find significant age related changes in mean thickness of the cortical plate, the volume, gyrification index, and surface area of the cortical plate continued to exponentially grow during the last phases of prenatal development. This cortical expansion coincides developmentally with the

  12. Quantitative and Qualitative Analysis of Transient Fetal Compartments during Prenatal Human Brain Development.

    PubMed

    Vasung, Lana; Lepage, Claude; Radoš, Milan; Pletikos, Mihovil; Goldman, Jennifer S; Richiardi, Jonas; Raguž, Marina; Fischi-Gómez, Elda; Karama, Sherif; Huppi, Petra S; Evans, Alan C; Kostovic, Ivica

    2016-01-01

    The cerebral wall of the human fetal brain is composed of transient cellular compartments, which show characteristic spatiotemporal relationships with intensity of major neurogenic events (cell proliferation, migration, axonal growth, dendritic differentiation, synaptogenesis, cell death, and myelination). The aim of the present study was to obtain new quantitative data describing volume, surface area, and thickness of transient compartments in the human fetal cerebrum. Forty-four postmortem fetal brains aged 13-40 postconceptional weeks (PCW) were included in this study. High-resolution T1 weighted MR images were acquired on 19 fetal brain hemispheres. MR images were processed using in-house software (MNI-ACE toolbox). Delineation of fetal compartments was performed semi-automatically by co-registration of MRI with histological sections of the same brains, or with the age-matched brains from Zagreb Neuroembryological Collection. Growth trajectories of transient fetal compartments were reconstructed. The composition of telencephalic wall was quantitatively assessed. Between 13 and 25 PCW, when the intensity of neuronal proliferation decreases drastically, the relative volume of proliferative (ventricular and subventricular) compartments showed pronounced decline. In contrast, synapse- and extracellular matrix-rich subplate compartment continued to grow during the first two trimesters, occupying up to 45% of telencephalon and reaching its maximum volume and thickness around 30 PCW. This developmental maximum coincides with a period of intensive growth of long cortico-cortical fibers, which enter and wait in subplate before approaching the cortical plate. Although we did not find significant age related changes in mean thickness of the cortical plate, the volume, gyrification index, and surface area of the cortical plate continued to exponentially grow during the last phases of prenatal development. This cortical expansion coincides developmentally with the

  13. Quantitative Magnetic Resonance Fat Measurements in Humans correlate With Established Methods but Are Biased

    PubMed Central

    Gallagher, Dympna; Thornton, John C.; He, Qing; Wang, Jack; Yu, Wen; Bradstreet, Thomas E.; Burke, Joanne; Heymsfield, Steven B.; Rivas, Veronica M.; Kaufman, Rhonda

    2012-01-01

    Precision and accuracy of the quantitative magnetic resonance (QMR) system for measuring fat in phantoms and total body fat (TBF) in humans were investigated. Measurements were made using phantoms: oil, beef with water, beef with oil, and humans with oil and water. TBFQMR in humans was compared with TBF by a four-compartment model (TBF4C). The coefficient of variation (CV) for replicate TBFQMR was 0.437%. QMR fat was lower at 23 °C vs. 37 °C. The fat increase in QMR phantom studies was consistent with the oil increase. When oil was added with humans, the increase in TBFQMR was >250 g for the initial 250 g of oil. With additional oil increments, the increase in TBFQMR was consistent with the amount of oil added. When water was added with humans, the TBFQMR increased independent of the amount of water added. TBFQMR was significantly less (mean ± s.e.) than TBF4C (females: −0.68 ± 0.27 kg, males: −4.66 ± 0.62 kg; P = 0.0001), TBFBV (females: −1.90 ± 0.40 kg; males: −5.68 ± 0.75 kg; P = 0.0001), and TBFD2O for males, but greater for females (1.19 ± 0.43 kg vs. −3.69 ± 0.81 kg for males; P = 0.0003). TBFQMR was lower than TBFiDXA with the difference greater in males (P = 0.001) and decreased with age (P = 0.011). The strong linear relationships between TBFQMR and TBF4C, TBFBV, and TBFD2O with slopes consistent with unity suggest that modifications are required to improve the accuracy. Should the latter be accomplished, QMR holds promise as a highly precise, rapid, and safe, noninvasive method for estimating the amount of and changes in TBF in overweight and severely obese persons. PMID:20448539

  14. Human Papillomavirus Load Measured by Linear Array Correlates with Quantitative PCR in Cervical Cytology Specimens

    PubMed Central

    Gravitt, Patti E.; Long, Rodney; Schiffman, Mark; Dunn, S. Terence; Carreon, J. Daniel; Allen, Richard A.; Gunja, Munira; Zuna, Rosemary E.; Sherman, Mark E.; Gold, Michael A.; Walker, Joan L.; Wang, Sophia S.

    2012-01-01

    Carcinogenic human papillomavirus (HPV) infections are necessary causes of most anogenital cancers. Viral load has been proposed as a marker for progression to cancer precursors but has been confirmed only for HPV16. Challenges in studying viral load are related to the lack of validated assays for a large number of genotypes. We compared viral load measured by Linear Array (LA) HPV genotyping with the gold standard, quantitative PCR (Q-PCR). LA genotyping and Q-PCR were performed in 143 cytology specimens from women referred to colposcopy. LA signal strength was measured by densitometry. Correlation coefficients and receiver operating characteristic (ROC) analyses were used to evaluate analytical and clinical performance. We observed a moderate to strong correlation between the two quantitative viral load measurements, ranging from an R value of 0.61 for HPV31 to an R value of 0.86 for HPV52. We also observed agreement between visual LA signal strength evaluation and Q-PCR. Both quantifications agreed on the disease stages with highest viral load, which varied by type (cervical intraepithelial neoplasia grade 2 [CIN2] for HPV52, CIN3 for HPV16 and HPV33, and cancer for HPV18 and HPV31). The area under the curve (AUC) for HPV16 Q-PCR at the CIN3 cutoff was 0.72 (P = 0.004), and the AUC for HPV18 LA at the CIN2 cutoff was 0.78 (P = 0.04). Quantification of LA signals correlates with the current gold standard for viral load, Q-PCR. Analyses of viral load need to address multiple infections and type attribution to evaluate whether viral load has clinical value beyond the established HPV16 finding. Our findings support conducting comprehensive studies of viral load and cervical cancer precursors using quantitative LA genotyping data. PMID:22337992

  15. Direct quantitation of methyl phosphonate adducts to human serum butyrylcholinesterase by immunomagnetic-UHPLC-MS/MS.

    PubMed

    Carter, Melissa D; Crow, Brian S; Pantazides, Brooke G; Watson, Caroline M; Thomas, Jerry D; Blake, Thomas A; Johnson, Rudolph C

    2013-11-19

    Hydrolysis of G- and V-series organophosphorus nerve agents (OPNAs) containing a phosphorus-methyl bond yields a methylphosphonic acid (MeP) product when adducted to human butyrylcholinesterase (BChE). The MeP adduct is considered a sign of "aging" and results in loss of the o-alkyl identifier specific to each nerve agent. After aging has occurred, common therapeutics such as oximes cannot reactivate the cholinesterase enzyme and relieve cholinergic inhibition. Until now, a direct, quantitative method for determination of the MeP adduct to BChE was unavailable. Aged adducts in serum samples were processed by immunomagnetic separation of BChE by antibody conjugated bead, isotope-dilution, pepsin digestion, followed by UHPLC separation and detection by conventional electrospray ionization-tandem mass spectrometry (ESI-MS/MS). Ions were detected in selected reaction monitoring (SRM) mode, and transition m/z 874.3 → 778.3 was used for quantitation. The analytical response ratio was linearly proportional to the serum concentration of MeP-adducted peptide (MeP-P) over the nominal concentration range of 2.0-250 ng/mL, with a coefficient of determination of R(2) ≥ 0.997. Intrarun accuracy, expressed as %Relative Error (%RE), was ≤13.5%, 16.3%, and 3.20% at 2.0, 16, and 250 ng/mL, respectively; the corresponding precision expressed as %RSD was ≤11.9%, 6.15%, and 3.39%. Interday %RSD was ≤7.13%, 5.69%, and 1.91%. Recovery of MeP-P from serum was ≥68% across the validated concentration range, and contributions from matrix effects were minimal. The method provides a direct, quantitative measurement of MeP-P found in clinical samples suspected of nerve agent exposure and subjected to such post-sampling stresses as elevated temperature and extended shipping. PMID:24205842

  16. A quantitative description of Ndc80 complex linkage to human kinetochores

    PubMed Central

    Suzuki, Aussie; Badger, Benjamin L.; Salmon, Edward D.

    2015-01-01

    The Ndc80 complex, which mediates end-on attachment of spindle microtubules, is linked to centromeric chromatin in human cells by two inner kinetochore proteins, CENP-T and CENP-C. Here to quantify their relative contributions to Ndc80 recruitment, we combine measurements of kinetochore protein copy number with selective protein depletion assays. This approach reveals about 244 Ndc80 complexes per human kinetochore (∼14 per kinetochore microtubule), 215 CENP-C, 72 CENP-T and only 151 Ndc80s as part of the KMN protein network (1:1:1 Knl1, Mis12 and Ndc80 complexes). Each CENP-T molecule recruits ∼2 Ndc80 complexes; one as part of a KMN network. In contrast, ∼40% of CENP-C recruits only a KMN network. Replacing the CENP-C domain that binds KMN with the CENP-T domain that recruits both an Ndc80 complex and KMN network yielded functional kinetochores. These results provide a quantitative picture of the linkages between centromeric chromatin and the microtubule-binding Ndc80 complex at the human kinetochore. PMID:26345214

  17. Establishment of two quantitative nested qPCR assays targeting the human EPO transgene.

    PubMed

    Neuberger, E W I; Perez, I; Le Guiner, C; Moser, D; Ehlert, T; Allais, M; Moullier, P; Simon, P; Snyder, R O

    2016-04-01

    For ethical and safety reasons it is critical to develop easily implemented assays with high sensitivity and specificity for gene doping surveillance. Two nested quantitative real-time PCR (qPCR) assays were developed that target the human EPO (hEPO) cDNA sequence in a circular form, representative of recombinant adeno-associated viral (rAAV) vector genomes found in vivo. Through an interlaboratory evaluation, the assays were validated and utilized in an in vitro blinded study. These assays are specific and extremely sensitive with a limit of detection (LOD) of 1 copy of circular plasmid DNA and a limit of quantification (LOQ) of 10 to 20 copies in the presence of 500 ng of human genomic DNA (hgDNA) extracted from WBCs. Additionally, using the two nested qPCR assays in a non-human primate study, where macaques were injected intramuscularly with a rAAV8 vector harboring a promoterless hEPO cDNA sequence, the viral vector was detected 8 to 14 weeks post-injection in macaque WBCs. The high sensitivity of the nested qPCR approach along with the capability of quantifying target DNA, make this approach a reliable tool for gene doping surveillance and the monitoring of exogenous DNA sequences. PMID:26752352

  18. Consensus Genome-Wide Expression Quantitative Trait Loci and Their Relationship with Human Complex Trait Disease.

    PubMed

    Yu, Chen-Hsin; Pal, Lipika R; Moult, John

    2016-07-01

    Most of the risk loci identified from genome-wide association (GWA) studies do not provide direct information on the biological basis of a disease or on the underlying mechanisms. Recent expression quantitative trait locus (eQTL) association studies have provided information on genetic factors associated with gene expression variation. These eQTLs might contribute to phenotype diversity and disease susceptibility, but interpretation is handicapped by low reproducibility of the expression results. To address this issue, we have generated a set of consensus eQTLs by integrating publicly available data for specific human populations and cell types. Overall, we find over 4000 genes that are involved in high-confidence eQTL relationships. To elucidate the role that eQTLs play in human common diseases, we matched the high-confidence eQTLs to a set of 335 disease risk loci identified from the Wellcome Trust Case Control Consortium GWA study and follow-up studies for 7 human complex trait diseases-bipolar disorder (BD), coronary artery disease (CAD), Crohn's disease (CD), hypertension (HT), rheumatoid arthritis (RA), type 1 diabetes (T1D), and type 2 diabetes (T2D). The results show that the data are consistent with ∼50% of these disease loci arising from an underlying expression change mechanism. PMID:27428252

  19. Quantitative Subcellular Proteome and Secretome Profiling of Influenza A Virus-Infected Human Primary Macrophages

    PubMed Central

    Lietzén, Niina; Julkunen, Ilkka; Aittokallio, Tero; Matikainen, Sampsa; Nyman, Tuula A.

    2011-01-01

    Influenza A viruses are important pathogens that cause acute respiratory diseases and annual epidemics in humans. Macrophages recognize influenza A virus infection with their pattern recognition receptors, and are involved in the activation of proper innate immune response. Here, we have used high-throughput subcellular proteomics combined with bioinformatics to provide a global view of host cellular events that are activated in response to influenza A virus infection in human primary macrophages. We show that viral infection regulates the expression and/or subcellular localization of more than one thousand host proteins at early phases of infection. Our data reveals that there are dramatic changes in mitochondrial and nuclear proteomes in response to infection. We show that a rapid cytoplasmic leakage of lysosomal proteins, including cathepsins, followed by their secretion, contributes to inflammasome activation and apoptosis seen in the infected macrophages. Also, our results demonstrate that P2X7 receptor and src tyrosine kinase activity are essential for inflammasome activation during influenza A virus infection. Finally, we show that influenza A virus infection is associated with robust secretion of different danger-associated molecular patterns (DAMPs) suggesting an important role for DAMPs in host response to influenza A virus infection. In conclusion, our high-throughput quantitative proteomics study provides important new insight into host-response against influenza A virus infection in human primary macrophages. PMID:21589892

  20. Detection of human brain tumor infiltration with quantitative stimulated Raman scattering microscopy

    PubMed Central

    Ji, Minbiao; Lewis, Spencer; Camelo-Piragua, Sandra; Ramkissoon, Shakti H.; Snuderl, Matija; Venneti, Sriram; Fisher-Hubbard, Amanda; Garrard, Mia; Fu, Dan; Wang, Anthony C.; Heth, Jason A.; Maher, Cormac O.; Sanai, Nader; Johnson, Timothy D.; Freudiger, Christian W.; Sagher, Oren; Xie, Xiaoliang Sunney; Orringer, Daniel A.

    2016-01-01

    Differentiating tumor from normal brain is a major barrier to achieving optimal outcome in brain tumor surgery. New imaging techniques for visualizing tumor margins during surgery are needed to improve surgical results. We recently demonstrated the ability of stimulated Raman scattering (SRS) microscopy, a non-destructive, label-free optical method, to reveal glioma infiltration in animal models. Here we show that SRS reveals human brain tumor infiltration in fresh, unprocessed surgical specimens from 22 neurosurgical patients. SRS detects tumor infiltration in near-perfect agreement with standard hematoxylin and eosin light microscopy (κ=0.86). The unique chemical contrast specific to SRS microscopy enables tumor detection by revealing quantifiable alterations in tissue cellularity, axonal density and protein:lipid ratio in tumor-infiltrated tissues. To ensure that SRS microscopic data can be easily used in brain tumor surgery, without the need for expert interpretation, we created a classifier based on cellularity, axonal density and protein:lipid ratio in SRS images capable of detecting tumor infiltration with 97.5% sensitivity and 98.5% specificity. Importantly, quantitative SRS microscopy detects the spread of tumor cells, even in brain tissue surrounding a tumor that appears grossly normal. By accurately revealing tumor infiltration, quantitative SRS microscopy holds potential for improving the accuracy of brain tumor surgery. PMID:26468325

  1. Noninvasive quantitation of phosphorus metabolites in human tissue by NMR spectroscopy

    NASA Astrophysics Data System (ADS)

    Roth, K.; Hubesch, B.; Meyerhoff, D. J.; Naruse, S.; Gober, J. R.; Lawry, T. J.; Boska, M. D.; Matson, G. B.; Weiner, M. W.

    Quantitation of metabolite concentrations by NMR spectroscopy is complicated by the need to determine the volume from which signals are detected, and by the need to obtain the relative sensitivity of detection within this volume. The use of coils with inhomogeneous B1 fields further complicates these problems. In order to quantify metabolite concentrations using 31P NMR spectroscopy, an external reference of hexamethyl phosphoroustriamide was used. Studies were performed on phantoms, using either a surface coil or a Helmholtz head coil to confirm the accuracy of both the ISIS volume selection technique and the use of an external reference. The limitations of this method are related to contamination and signal loss inherent in the ISIS technique and difficulties with integration of broad overlapping peaks. The method was applied to seven normal human subjects. The integrals for metabolite signals in normal brain and calf muscle were determined by using NMRI software. The T1 values of the signals of all phosphorus metabolites in the selected volume were measured in order to correct for saturation effects. The concentrations for PCr, P i, and ATP were 4.9, 2.0, and 2.5 m M in brain and 36.5, 5.7, and 7.3 m M in muscle. These results are in good agreement with those reported for animals, demonstrating the validity of this quantitation technique.

  2. Quantitative detection of nitric oxide in exhaled human breath by extractive electrospray ionization mass spectrometry

    NASA Astrophysics Data System (ADS)

    Pan, Susu; Tian, Yong; Li, Ming; Zhao, Jiuyan; Zhu, Lanlan; Zhang, Wei; Gu, Haiwei; Wang, Haidong; Shi, Jianbo; Fang, Xiang; Li, Penghui; Chen, Huanwen

    2015-03-01

    Exhaled nitric oxide (eNO) is a useful biomarker of various physiological conditions, including asthma and other pulmonary diseases. Herein a fast and sensitive analytical method has been developed for the quantitative detection of eNO based on extractive electrospray ionization mass spectrometry (EESI-MS). Exhaled NO molecules selectively reacted with 2-phenyl-4, 4, 5, 5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO) reagent, and eNO concentration was derived based on the EESI-MS response of 1-oxyl-2-phenyl-4, 4, 5, 5-tetramethylimidazoline (PTI) product. The method allowed quantification of eNO below ppb level (~0.02 ppbv) with a relative standard deviation (RSD) of 11.6%. In addition, eNO levels of 20 volunteers were monitored by EESI-MS over the time period of 10 hrs. Long-term eNO response to smoking a cigarette was recorded, and the observed time-dependent profile was discussed. This work extends the application of EESI-MS to small molecules (<30 Da) with low proton affinity and collision-induced dissociation efficiency, which are usually poorly visible by conventional ion trap mass spectrometers. Long-term quantitative profiling of eNO by EESI-MS opens new possibilities for the research of human metabolism and clinical diagnosis.

  3. Hyperspectral and differential CARS microscopy for quantitative chemical imaging in human adipocytes.

    PubMed

    Di Napoli, Claudia; Pope, Iestyn; Masia, Francesco; Watson, Peter; Langbein, Wolfgang; Borri, Paola

    2014-05-01

    In this work, we demonstrate the applicability of coherent anti-Stokes Raman scattering (CARS) micro-spectroscopy for quantitative chemical imaging of saturated and unsaturated lipids in human stem-cell derived adipocytes. We compare dual-frequency/differential CARS (D-CARS), which enables rapid imaging and simple data analysis, with broadband hyperspectral CARS microscopy analyzed using an unsupervised phase-retrieval and factorization method recently developed by us for quantitative chemical image analysis. Measurements were taken in the vibrational fingerprint region (1200-2000/cm) and in the CH stretch region (2600-3300/cm) using a home-built CARS set-up which enables hyperspectral imaging with 10/cm resolution via spectral focussing from a single broadband 5 fs Ti:Sa laser source. Through a ratiometric analysis, both D-CARS and phase-retrieved hyperspectral CARS determine the concentration of unsaturated lipids with comparable accuracy in the fingerprint region, while in the CH stretch region D-CARS provides only a qualitative contrast owing to its non-linear behavior. When analyzing hyperspectral CARS images using the blind factorization into susceptibilities and concentrations of chemical components recently demonstrated by us, we are able to determine vol:vol concentrations of different lipid components and spatially resolve inhomogeneities in lipid composition with superior accuracy compared to state-of-the art ratiometric methods. PMID:24877002

  4. Multiparametric MRI Assessment of Human Articular Cartilage Degeneration: Correlation with Quantitative Histology and Mechanical Properties

    PubMed Central

    Rautiainen, Jari; Nissi, Mikko J.; Salo, Elli-Noora; Tiitu, Virpi; Finnilä, Mikko A.J.; Aho, Olli-Matti; Saarakkala, Simo; Lehenkari, Petri; Ellermann, Jutta; Nieminen, Miika T.

    2014-01-01

    Purpose To evaluate the sensitivity of quantitative MRI techniques (T1, T1,Gd, T2, continous wave (CW) T1ρ dispersion, adiabatic T1ρ, adiabatic T2ρ, RAFF and inversion-prepared magnetization transfer (MT)) for assessment of human articular cartilage with varying degrees of natural degeneration. Methods Osteochondral samples (n = 14) were obtained from the tibial plateaus of patients undergoing total knee replacement. MRI of the specimens was performed at 9.4 T and the relaxation time maps were evaluated in the cartilage zones. For reference, quantitative histology, OARSI grading and biomechanical measurements were performed and correlated with MRI findings. Results All MRI parameters, except T1,Gd, showed statistically significant differences in tangential and full-thickness ROIs between early and advanced osteoarthritis (OA) groups, as classified by OARSI grading. CW-T1ρ showed significant dispersion in all ROIs and featured classical laminar structure of cartilage with spin-lock powers below 1000 Hz. Adiabatic T1ρ, T2ρ, CW-T1ρ, MT and RAFF correlated strongly with OARSI grade and biomechanical parameters. Conclusion MRI parameters were able to differentiate between early and advanced OA. Furthermore, rotating frame methods, namely adiabatic T1ρ, adiabatic T2ρ, CW-T1ρ and RAFF, as well as MT experiment correlated strongly with biomechanical parameters and OARSI grade, suggesting high sensitivity of the parameters for cartilage degeneration. PMID:25104181

  5. A method for the simultaneous identification and quantitation of five superwarfarin rodenticides in human serum.

    PubMed

    Kuijpers, E A; den Hartigh, J; Savelkoul, T J; de Wolff, F A

    1995-01-01

    A high-performance liquid chromatographic method with ultraviolet (UV) and fluorescence detection was developed for the analysis of one indandione and four hydroxycoumarin anticoagulant rodenticides in human serum. The superwarfarin rodenticides, chlorophacinone, bromadiolone, difenacoum, brodifacoum, and difethialone, can be identified and quantitated simultaneously with this method. After adding a buffer (pH 5.5), the anticoagulants were extracted from serum with chloroform-acetone. The organic phase was separated and evaporated to dryness, and the residue was subjected to chromatographic analysis. The anticoagulants were separated by reversed-phase chromatography and detected by UV absorption at 285 nm and by fluorescence at an excitation wavelength of 265 nm and an emission wavelength of 400 nm. Extraction efficiencies from 55 to 131% were obtained. The within-run precision ranged from 2.0 to 7.1% for UV detection and from 0.0 to 4.8% for fluorescence detection. Between-run precision ranged from 1.3 to 16.0% for UV detection and from 1.8 to 9.0% for fluorescence detection. The anticoagulants can be quantitated at serum concentrations down to 3-12 ng/mL for fluorescence detection and down to 20-75 ng/mL for UV detection. No interferences were observed with the related compounds warfarin and vitamin K1. PMID:8577178

  6. Detection of human brain tumor infiltration with quantitative stimulated Raman scattering microscopy.

    PubMed

    Ji, Minbiao; Lewis, Spencer; Camelo-Piragua, Sandra; Ramkissoon, Shakti H; Snuderl, Matija; Venneti, Sriram; Fisher-Hubbard, Amanda; Garrard, Mia; Fu, Dan; Wang, Anthony C; Heth, Jason A; Maher, Cormac O; Sanai, Nader; Johnson, Timothy D; Freudiger, Christian W; Sagher, Oren; Xie, Xiaoliang Sunney; Orringer, Daniel A

    2015-10-14

    Differentiating tumor from normal brain is a major barrier to achieving optimal outcome in brain tumor surgery. New imaging techniques for visualizing tumor margins during surgery are needed to improve surgical results. We recently demonstrated the ability of stimulated Raman scattering (SRS) microscopy, a nondestructive, label-free optical method, to reveal glioma infiltration in animal models. We show that SRS reveals human brain tumor infiltration in fresh, unprocessed surgical specimens from 22 neurosurgical patients. SRS detects tumor infiltration in near-perfect agreement with standard hematoxylin and eosin light microscopy (κ = 0.86). The unique chemical contrast specific to SRS microscopy enables tumor detection by revealing quantifiable alterations in tissue cellularity, axonal density, and protein/lipid ratio in tumor-infiltrated tissues. To ensure that SRS microscopic data can be easily used in brain tumor surgery, without the need for expert interpretation, we created a classifier based on cellularity, axonal density, and protein/lipid ratio in SRS images capable of detecting tumor infiltration with 97.5% sensitivity and 98.5% specificity. Quantitative SRS microscopy detects the spread of tumor cells, even in brain tissue surrounding a tumor that appears grossly normal. By accurately revealing tumor infiltration, quantitative SRS microscopy holds potential for improving the accuracy of brain tumor surgery. PMID:26468325

  7. Quantitative remineralization evolution kinetics of artificially demineralized human enamel using photothermal radiometry and modulated luminescence.

    PubMed

    Hellen, Adam; Mandelis, Andreas; Finer, Yoav; Amaechi, Bennett T

    2011-11-01

    Human molars were subjected to demineralization in acid gel followed by incubation in remineralization solutions without or with fluoride (1 or 1000 ppm). Photothermal radiometry (PTR) and modulated luminescence (LUM) frequency scans were performed prior to and during de/remineralization treatments. Transverse Micro-Radiography (TMR) analysis followed at treatment conclusion to determine mineral loss and lesion depth. The remineralization process illustrated a complex interplay between surface and subsurface mineral deposition, confining the thermal-wave centroid toward the dominating layer. Experimental amplitudes and phases were fitted to a coupled diffuse-photon-density-wave and thermal-wave theoretical model used to quantitatively evaluate evolving changes in thermal and optical properties of de/remineralized enamel lesions. Additional information obtained from the LUM data corroborated the remineralization kinetics affecting the PTR signals. The results pointed to enhanced effectiveness of subsurface lesion remineralization in the presence of fluoride. PMID:21761572

  8. Quantitative visualization of DNA G-quadruplex structures in human cells

    NASA Astrophysics Data System (ADS)

    Biffi, Giulia; Tannahill, David; McCafferty, John; Balasubramanian, Shankar

    2013-03-01

    Four-stranded G-quadruplex nucleic acid structures are of great interest as their high thermodynamic stability under near-physiological conditions suggests that they could form in cells. Here we report the generation and application of an engineered, structure-specific antibody employed to quantitatively visualize DNA G-quadruplex structures in human cells. We show explicitly that G-quadruplex formation in DNA is modulated during cell-cycle progression and that endogenous G-quadruplex DNA structures can be stabilized by a small-molecule ligand. Together these findings provide substantive evidence for the formation of G-quadruplex structures in the genome of mammalian cells and corroborate the application of stabilizing ligands in a cellular context to target G-quadruplexes and intervene with their function.

  9. Quantitative Proteomic Profiling of Low Dose Ionizing Radiation Effects in a Human Skin Model

    SciTech Connect

    Hengel, Shawna; Aldrich, Joshua T.; Waters, Katrina M.; Pasa-Tolic, Ljiljana; Stenoien, David L.

    2014-07-29

    To assess molecular responses to low doses of radiation that may be encountered during medical diagnostic procedures, nuclear accidents, or terrorist acts, a quantitative global proteomic approach was used to identify protein alterations in a reconstituted human skin tissue treated with 10 cGy of ionizing radiation. Subcellular fractionation was employed to remove highly abundant structural proteins and provide insight on radiation induced alterations in protein abundance and localization. In addition, peptides were post-fractionated using high resolution 2-dimensional liquid chromatography to increase the dynamic range of detection of protein abundance and translocation changes. Quantitative data was obtained by labeling peptides with 8-plex isobaric iTRAQ tags. A total of 207 proteins were detected with statistically significant alterations in abundance and/or subcellular localization compared to sham irradiated tissues. Bioinformatics analysis of the data indicated that the top canonical pathways affected by low dose radiation are related to cellular metabolism. Among the proteins showing alterations in abundance, localization and proteolytic processing was the skin barrier protein filaggrin which is consistent with our previous observation that ionizing radiation alters profilaggrin processing with potential effects on skin barrier functions. In addition, a large number of proteases and protease regulators were affected by low dose radiation exposure indicating that altered proteolytic activity may be a hallmark of low dose radiation exposure. While several studies have demonstrated altered transcriptional regulation occurs following low dose radiation exposures, the data presented here indicates post-transcriptional regulation of protein abundance, localization, and proteolytic processing play an important role in regulating radiation responses in complex human tissues.

  10. Rapid and efficacious real-time quantitative PCR assay for quantitation of human DNA in forensic samples.

    PubMed

    Tringali, G; Barbaro, A; Insirello, E; Cormaci, P; Roccazzello, A M

    2004-12-01

    A reliable quantification of human DNA is necessary in the process of routine forensic human identification. When DNA is degraded, contaminated or associated with inhibitors, is important to accurately estimate the concentration of extracted DNA prior to perform an analysis based on nuclear markers amplified. In this work, a new approaches to DNA quantification employees the use of a real-time fluorescence probe system. The real-time PCR assay is highly sensitive, specific, rapid, cost-effective and flexible assay for analysis of forensic casework samples, can perform with DNA of poor quality and detect specifically amplifiable DNA rather than total DNA for STR analysis. PMID:15639571

  11. Quantitative assessment of human and pet exposure to Salmonella associated with dry pet foods.

    PubMed

    Lambertini, Elisabetta; Buchanan, Robert L; Narrod, Clare; Ford, Randall M; Baker, Robert C; Pradhan, Abani K

    2016-01-01

    Recent Salmonella outbreaks associated with dry pet foods and treats highlight the importance of these foods as previously overlooked exposure vehicles for both pets and humans. In the last decade efforts have been made to raise the safety of this class of products, for instance by upgrading production equipment, cleaning protocols, and finished product testing. However, no comprehensive or quantitative risk profile is available for pet foods, thus limiting the ability to establish safety standards and assess the effectiveness of current and proposed Salmonella control measures. This study sought to develop an ingredients-to-consumer quantitative microbial exposure assessment model to: 1) estimate pet and human exposure to Salmonella via dry pet food, and 2) assess the impact of industry and household-level mitigation strategies on exposure. Data on prevalence and concentration of Salmonella in pet food ingredients, production process parameters, bacterial ecology, and contact transfer in the household were obtained through literature review, industry data, and targeted research. A probabilistic Monte Carlo modeling framework was developed to simulate the production process and basic household exposure routes. Under the range of assumptions adopted in this model, human exposure due to handling pet food is null to minimal if contamination occurs exclusively before extrusion. Exposure increases considerably if recontamination occurs post-extrusion during coating with fat, although mean ingested doses remain modest even at high fat contamination levels, due to the low percent of fat in the finished product. Exposure is highly variable, with the distribution of doses ingested by adult pet owners spanning 3Log CFU per exposure event. Child exposure due to ingestion of 1g of pet food leads to significantly higher doses than adult doses associated with handling the food. Recontamination after extrusion and coating, e.g., via dust or equipment surfaces, may also lead to

  12. Perceptions of climate change and its impact on human health: an integrated quantitative and qualitative approach

    PubMed Central

    Toan, Do Thi Thanh; Kien, Vu Duy; Bao Giang, Kim; Van Minh, Hoang

    2014-01-01

    Background The World Health Organization emphasized that climate change is a significant and emerging threat to public health, especially in lower income populations and tropical/subtropical countries. However, people in Asia and Africa were the least likely to perceive global warming as a threat. In Vietnam, little research has been conducted concerning the perceptions of effects of climate change on human health. Objective The aim of this study was to explore the perceptions on climate change and its impact on human health among people in Hanoi. Design We applied a combined quantitative and qualitative approach to study perceptions on climate change among people in Hanoi. A total of 1,444 people were recruited, including 754 people living in non-slum areas and 690 people living in slum areas of Hanoi. A structured questionnaire was used to collect quantitative data on their perceptions. In a parallel qualitative study, two focus group discussions and 12 in-depth interviews (IDs) were carried out involving 24 people from both slum and non-slum areas. Results The majority of the respondents in the study had heard about climate change and its impact on human health (79.3 and 70.1% in non-slum and slum areas, respectively). About one third of the respondents reported that members of their family had experienced illness in the recent summer and winter compared to the same seasons 5 years ago. The most common symptoms reported during hot weather were headaches, fatigue, and dizziness; hypertension and other cardiovascular diseases were also reported. During cold weather, people reported experiencing cough, fever, and influenza, as well as pneumonia and emerging infectious diseases such as dengue and Japanese encephalitis. Conclusions The observed high level of awareness on the links between climate change and human health may help to increase the success of the National Prevention Program on Climate Change. Moreover, understanding the concerns of the people may help

  13. Interleukin-10 modifies the effects of interleukin-1beta and tumor necrosis factor-alpha on the activity and expression of prostaglandin H synthase-2 and the NAD+-dependent 15-hydroxyprostaglandin dehydrogenase in cultured term human villous trophoblast and chorion trophoblast cells.

    PubMed

    Pomini, F; Caruso, A; Challis, J R

    1999-12-01

    The concentrations of tumor necrosis factor-alpha (TNFalpha) and interleukin-1beta (IL-1beta), two inflammatory cytokines in amniotic fluid, have been shown to rise during chorioamnionitis. This is probably related to activation of the immune system in order to intensify the inflammatory process and to protect the maternal and fetal organism from infectious agents. These cytokines activate the PG biosynthetic pathway in several tissues, but few studies have examined effects on PG-metabolizing enzymes. When PGs are produced by increased synthesis and/or decreased metabolism at the chorio-decidual interface, labor can be induced. Interleukin-10 (IL-10) is known to act as an antiinflammatory cytokine. The goals of this study were to evaluate the interaction of IL-10 with IL-1beta and TNFalpha on PG synthesis and to determine the effects of IL-10, IL-1beta, and TNFalpha on PG metabolism using purified cultures of villous trophoblast and chorion trophoblast cells prepared from placentas of patients at term. Cells were treated with IL-1beta and TNFalpha with or without IL-10 for various times up to 24 h. Levels of messenger ribonucleic acid (mRNA) encoding PGH synthase-2 (PGHS-2) and NAD+-dependent 15-hydroxyprostaglandin dehydrogenase (PGDH) were quantified by Northern blotting, and PGE2 and 13,14-dihydro-15-keto-PGF2alpha (PGFM) output in the medium was measured by RIA. IL-1beta increased PGHS-2 mRNA and PGE2 output from villous and chorion trophoblasts and decreased PGDH mRNA in villous trophoblasts (all P < 0.05). These effects were reversed by IL-10. We found no change in PGHS-2 mRNA or PGE2 output in either trophoblast type treated with TNFalpha, but TNFalpha reduced PGDH mRNA in villous trophoblast, and this effect was reversed by IL-10 (both P < 0.05). We conclude that proinflammatory cytokines can influence PG output through effects on PG synthesis and metabolism and that these effects may be opposed by an antiinflammatory cytokine. These interactions may be

  14. Quantitative Analysis of Human Salivary Gland-Derived Intact Proteome Using Top-Down Mass Spectrometry

    SciTech Connect

    Wu, Si; Brown, Joseph N.; Tolic, Nikola; Meng, Da; Liu, Xiaowen; Zhang, Haizhen; Zhao, Rui; Moore, Ronald J.; Pevzner, Pavel A.; Smith, Richard D.; Pasa-Tolic, Ljiljana

    2014-05-31

    There are several notable challenges inherent to fully characterizing the entirety of the human saliva proteome using bottom-up approaches, including polymorphic isoforms, post-translational modifications, unique splice variants, deletions, and truncations. To address these challenges, we have developed a top-down based liquid chromatography-mass spectrometry (LC-MS) approach, which cataloged 20 major human salivary proteins with a total of 83 proteoforms, containing a broad range of post-translational modifications. Among these proteins, several previously reported disease biomarker proteins were identified at the intact protein level, such as beta-2 microglobulin (B2M). In addition, intact glycosylated proteoforms of several saliva proteins were also characterized, including intact N-glycosylated protein prolactin inducible protein (PIP) and O-glycosylated acidic protein rich protein (aPRP). These characterized proteoforms constitute an intact saliva proteoform database, which was used for quantitative comparison of intact salivary proteoforms among six healthy individuals. Human parotid (PS) and submandibular/sublingual gland (SMSL) secretion samples (2 μg of protein each) from six healthy individuals were compared using RPLC coupled with the 12T FTICR mass spectrometer. Significantly different protein and PTM patterns were resolved with high reproducibility between PS and SMSL glands. The results from this study provide further insight into the potential mechanisms of PTM pathways in oral glandular secretion, expanding our knowledge of this complex yet easily accessible fluid. Intact protein LC-MS approach presented herein can potentially be applied for rapid and accurate identification of biomarkers from only a few microliters of human glandular saliva.

  15. The role of chorion on toxicity of silver nanoparticles in the embryonic zebrafish assay

    PubMed Central

    Kim, Ki-Tae; Tanguay, Robert L.

    2014-01-01

    Objectives This study was designed to investigate how the size- and surface coating-dependent toxicity of silver nanoparticles (AgNPs) is influenced by the presence and absence of the chorion in an embryonic zebrafish assay. Methods Normal and dechorinated embryos were exposed to four different AgNPs, 20 or 110 nm in size, with polypyrrolidone (PVP) or citrate surface coatings in a standard zebrafish embryo medium (EM). This was then compared to a 62.5 μM calcium chloride (CaCl2) solution where agglomeration was controlled. Results Embryonic toxicity in the absence of the chorion was greater than in its presence. The smaller 20 nm AgNPs were more toxic than the larger 110 nm AgNPs, regardless of the chorion and test media. However, surface coating affected toxicity, since PVPcoated AgNPs were more toxic than citrate-coated AgNPs; this was strongly affected by the presence of the chorion in both EM and CaCl2. Conclusions Our results demonstrate the permeability function of the chorion on the size- and surface coating-dependent toxicity of AgNPs. Thereafter, careful experiment should be conducted to assess nanoparticle toxicity in zebrafish embryos. PMID:25518841

  16. Analysis of developmentally regulated chorion gene promoter architecture via electroporation of silk moth follicles.

    PubMed

    Tsatsarounos, S P; Rodakis, G C; Lecanidou, R

    2015-02-01

    In the silk moth Bombyx mori, chorion genes of the same developmental specificity are organized in divergently transcribed α/β gene pairs, sharing a common 5' flanking promoter region. This bidirectional promoter contains a complete set of cis-elements responsible for developmentally accurate gene expression. In the present paper, based on the observation that Bombyx chorion gene promoters contain cis-elements for the same transcription factors without concrete evidence on which of them are essential, we address the question as to how promoter architecture (number, orientation and position of common factor binding sites) facilitates developmentally accurate chorion gene regulation. To this end, we constructed several mutated promoter regions of an early-middle gene pair and cloned them upstream of a reporter gene to introduce these plasmid constructs into silk moth follicle epithelial cells via electroporation as an efficient and quick method for transient expression. This is the first time that an ex vivo method had been applied to test the impact of systematic cis-element mutations on a chorion gene promoter. Our results confirmed the importance of the HMGA factor and the role of the GATA factor as an early repressor, and led to a more detailed understanding of which C/EBP sites participate in the regulation of early-middle chorion gene expression. PMID:25256090

  17. Quantitative analysis of MDR1 (multidrug resistance) gene expression in human tumors by polymerase chain reaction

    SciTech Connect

    Noonan, K.E.; Beck, C.; Holzmayer, T.A.; Chin, J.E.; Roninson, I.B. ); Wunder, J.S.; Andrulis, I.L. ); Gazdar, A.F. ); Willman, C.L.; Griffith, B. ); Von Hoff, D.D. )

    1990-09-01

    The resistance of tumor cells ot chemotheraprutic drugs is a major obstacle to successful cancer chemotherapy. In human cells, expression of the MDR1 gene, encoding a transmembrane efflux pump (P-glycoprotein), leads to decreased intracellular accumulation and resistance to a variety of lipophilic drugs (multidrug resistance; MDR). The levels of MDR in cell lines selected in bitro have been shown to correlate with the steady-state levels of MDR1 mRNA and P-glycoprotein. In cells with a severalfold increase in cellular drug resistance, MDR1 expression levels are close to the limits of detection by conventional assays. MDR1 expression has been frequently observed in human tumors after chemotherapy and in some but not all types of clinically refactory tumors untreated with chemotherapeutic drugs. The authors have devised a highly sensitive, specific, and quantitative protocol for measuring the levels of MDR1 mRNA in clincal samples, based on the polymerase chain reaction. They have used this assay to measure MDR1 gene expression in MDR cell lines and >300 normal tissues, tumor-derived cell lines, and clinical specimens of untreated tumors of the types in which MDR1 expression was rarely observed by standard assays. Low levels of MDR1 expression were found by polymerase chain reaction in most solid tumors and leukemias tested. The frequency of samples without detectable MDR1 expression varied among different types of tumors; MDR1-negative samples were ost common among tumor types known to be relatively responsive to chemotherapy.

  18. Nonmydriatic fluorescence-based quantitative imaging of human macular pigment distributions

    PubMed Central

    Sharifzadeh, Mohsen; Bernstein, Paul S.; Gellermann, Werner

    2011-01-01

    We have developed a CCD-camera-based nonmydriatic instrument that detects fluorescence from retinal lipofuscin chromophores (“autofluorescence”) as a means to indirectly quantify and spatially image the distribution of macular pigment (MP). The lipofuscin fluorescence intensity is reduced at all retinal locations containing MP, since MP has a competing absorption in the blue–green wavelength region. Projecting a large diameter, 488 nm excitation spot onto the retina, centered on the fovea, but extending into the macular periphery, and comparing lipofuscin fluorescence intensities outside and inside the foveal area, it is possible to spatially map out the distribution of MP. Spectrally selective detection of the lipofuscin fluorescence reveals an important wavelength dependence of the obtainable image contrast and deduced MP optical density levels, showing that it is important to block out interfering fluorescence contributions in the detection setup originating from ocular media such as the lens. Measuring 70 healthy human volunteer subjects with no ocular pathologies, we find widely varying spatial extent of MP, distinctly differing distribution patterns of MP, and strongly differing absolute MP levels among individuals. Our population study suggests that MP imaging based on lipofuscin fluorescence is useful as a relatively simple, objective, and quantitative noninvasive optical technique suitable to rapidly screen MP levels and distributions in healthy humans with undilated pupils. PMID:16985523

  19. Nonmydriatic fluorescence-based quantitative imaging of human macular pigment distributions

    NASA Astrophysics Data System (ADS)

    Sharifzadeh, Mohsen; Bernstein, Paul S.; Gellermann, Werner

    2006-10-01

    We have developed a CCD-camera-based nonmydriatic instrument that detects fluorescence from retinal lipofuscin chromophores ("autofluorescence") as a means to indirectly quantify and spatially image the distribution of macular pigment (MP). The lipofuscin fluorescence intensity is reduced at all retinal locations containing MP, since MP has a competing absorption in the blue-green wavelength region. Projecting a large diameter, 488 nm excitation spot onto the retina, centered on the fovea, but extending into the macular periphery, and comparing lipofuscin fluorescence intensities outside and inside the foveal area, it is possible to spatially map out the distribution of MP. Spectrally selective detection of the lipofuscin fluorescence reveals an important wavelength dependence of the obtainable image contrast and deduced MP optical density levels, showing that it is important to block out interfering fluorescence contributions in the detection setup originating from ocular media such as the lens. Measuring 70 healthy human volunteer subjects with no ocular pathologies, we find widely varying spatial extent of MP, distinctly differing distribution patterns of MP, and strongly differing absolute MP levels among individuals. Our population study suggests that MP imaging based on lipofuscin fluorescence is useful as a relatively simple, objective, and quantitative noninvasive optical technique suitable to rapidly screen MP levels and distributions in healthy humans with undilated pupils.

  20. Quantitative high-throughput identification of drugs as modulators of human constitutive androstane receptor.

    PubMed

    Lynch, Caitlin; Zhao, Jinghua; Huang, Ruili; Xiao, Jingwei; Li, Linhao; Heyward, Scott; Xia, Menghang; Wang, Hongbing

    2015-01-01

    The constitutive androstane receptor (CAR, NR1I3) plays a key role in governing the transcription of numerous hepatic genes that involve xenobiotic metabolism/clearance, energy homeostasis, and cell proliferation. Thus, identification of novel human CAR (hCAR) modulators may not only enhance early prediction of drug-drug interactions but also offer potentially novel therapeutics for diseases such as metabolic disorders and cancer. In this study, we have generated a double stable cell line expressing both hCAR and a CYP2B6-driven luciferase reporter for quantitative high-throughput screening (qHTS) of hCAR modulators. Approximately 2800 compounds from the NIH Chemical Genomics Center Pharmaceutical Collection were screened employing both the activation and deactivation modes of the qHTS. Activators (115) and deactivators (152) of hCAR were identified from the primary qHTS, among which 10 agonists and 10 antagonists were further validated in the physiologically relevant human primary hepatocytes for compound-mediated hCAR nuclear translocation and target gene expression. Collectively, our results reveal that hCAR modulators can be efficiently identified through this newly established qHTS assay. Profiling drug collections for hCAR activity would facilitate the prediction of metabolism-based drug-drug interactions, and may lead to the identification of potential novel therapeutics. PMID:25993555

  1. Quantitative analysis of intraneuronal transport in human iPS neurons.

    PubMed

    Nakamura, Haruko; Yamashita, Naoya; Kanamaru, Yuri; Tachibana, Takahiko; Sekino, Yuko; Chen, Sandy; Gotoh, Toshiyuki; Tanaka, Fumiaki; Goshima, Yoshio

    2015-08-01

    Induced pluripotent stem (iPS) cells are promising tools to investigate disease mechanism and develop new drugs. Intraneuronal transport, which is fundamental for neuronal survival and function, is vulnerable to various pharmacological and chemical agents and is disrupted in some neurodegenerative disorders. We applied a quantification method for axonal transport by counting CM-DiI-labeled particles traveling along the neurite, which allowed us to monitor and quantitate, for the first time, intraneuronal transport in human neurons differentiated from iPS cells (iCell neurons). We evaluated the acute effects of several anti-neoplastic agents that have been previously shown to affect intraneuronal transport. Vincristine, paclitaxel and oxaliplatin decreased the number of moving particle along neurites. Cisplatin, however, produced no effect on intraneuronal transport, which is in contrast to our previous report indicating that it inhibits transport in chick dorsal root ganglion neurons. Our system may be a useful method for assessing intraneuronal transport and neurotoxicity in human iPS neurons. PMID:26232861

  2. Quantitative proteomic analysis of human breast epithelial cells with differential telomere length

    SciTech Connect

    Yu, Li-Rong . E-mail: lyu@ncifcrf.gov; Chan, King C.; Tahara, Hidetoshi; Lucas, David A.; Chatterjee, Koushik; Issaq, Haleem J.; Veenstra, Timothy D. . E-mail: veenstra@ncifcrf.gov

    2007-05-18

    Telomeres play important functional roles in cell proliferation, cell cycle regulation, and genetic stability, in which telomere length is critical. In this study, quantitative proteome comparisons for the human breast epithelial cells with short and long telomeres (184-hTERT{sub L} vs. 184-hTERT{sub S} and 90P-hTERT{sub L} vs. 90P-hTERT{sub S}), resulting from transfection of the human telomerase reverse transcriptase (hTERT) gene, were performed using cleavable isotope-coded affinity tags. More than 2000 proteins were quantified in each comparative experiment, with approximately 77% of the proteins identified in both analyses. In the cells with long telomeres, significant and consistent alterations were observed in metabolism (amino acid, nucleotide, and lipid metabolism), genetic information transmission (transcription and translation regulation, spliceosome and ribosome complexes), and cell signaling. Interestingly, the DNA excision repair pathway is enhanced, while integrin and its ligands are downregulated in the cells with long telomeres. These results may provide valuable information related to telomere functions.

  3. Quantitative High-Throughput Identification of Drugs as Modulators of Human Constitutive Androstane Receptor

    PubMed Central

    Lynch, Caitlin; Zhao, Jinghua; Huang, Ruili; Xiao, Jingwei; Li, Linhao; Heyward, Scott; Xia, Menghang; Wang, Hongbing

    2015-01-01

    The constitutive androstane receptor (CAR, NR1I3) plays a key role in governing the transcription of numerous hepatic genes that involve xenobiotic metabolism/clearance, energy homeostasis, and cell proliferation. Thus, identification of novel human CAR (hCAR) modulators may not only enhance early prediction of drug-drug interactions but also offer potentially novel therapeutics for diseases such as metabolic disorders and cancer. In this study, we have generated a double stable cell line expressing both hCAR and a CYP2B6-driven luciferase reporter for quantitative high-throughput screening (qHTS) of hCAR modulators. Approximately 2800 compounds from the NIH Chemical Genomics Center Pharmaceutical Collection were screened employing both the activation and deactivation modes of the qHTS. Activators (115) and deactivators (152) of hCAR were identified from the primary qHTS, among which 10 agonists and 10 antagonists were further validated in the physiologically relevant human primary hepatocytes for compound-mediated hCAR nuclear translocation and target gene expression. Collectively, our results reveal that hCAR modulators can be efficiently identified through this newly established qHTS assay. Profiling drug collections for hCAR activity would facilitate the prediction of metabolism-based drug-drug interactions, and may lead to the identification of potential novel therapeutics. PMID:25993555

  4. Myelin and iron concentration in the human brain: a quantitative study of MRI contrast.

    PubMed

    Stüber, Carsten; Morawski, Markus; Schäfer, Andreas; Labadie, Christian; Wähnert, Miriam; Leuze, Christoph; Streicher, Markus; Barapatre, Nirav; Reimann, Katja; Geyer, Stefan; Spemann, Daniel; Turner, Robert

    2014-06-01

    During the last five years ultra-high-field magnetic resonance imaging (MRI) has enabled an unprecedented view of living human brain. Brain tissue contrast in most MRI sequences is known to reflect mainly the spatial distributions of myelin and iron. These distributions have been shown to overlap significantly in many brain regions, especially in the cortex. It is of increasing interest to distinguish and identify cortical areas by their appearance in MRI, which has been shown to be feasible in vivo. Parcellation can benefit greatly from quantification of the independent contributions of iron and myelin to MRI contrast. Recent studies using susceptibility mapping claim to allow such a separation of the effects of myelin and iron in MRI. We show, using post-mortem human brain tissue, that this goal can be achieved. After MRI scanning of the block with appropriate T1 mapping and T2* weighted sequences, we section the block and apply a novel technique, proton induced X-ray emission (PIXE), to spatially map iron, phosphorus and sulfur elemental concentrations, simultaneously with 1μm spatial resolution. Because most brain phosphorus is located in myelin phospholipids, a calibration step utilizing element maps of sulfur enables semi-quantitative ex vivo mapping of myelin concentration. Combining results for iron and myelin concentration in a linear model, we have accurately modeled MRI tissue contrasts. Conversely, iron and myelin concentrations can now be estimated from appropriate MRI measurements in post-mortem brain samples. PMID:24607447

  5. Quantitative Structure-Pharmacokinetic Relationships for the Prediction of Renal Clearance in Humans

    PubMed Central

    Dave, Rutwij A.

    2015-01-01

    Renal clearance (CLR), a major route of elimination for many drugs and drug metabolites, represents the net result of glomerular filtration, active secretion and reabsorption, and passive reabsorption. The aim of this study was to develop quantitative structure-pharmacokinetic relationships (QSPKR) to predict CLR of drugs or drug-like compounds in humans. Human CLR data for 382 compounds were obtained from the literature. Step-wise multiple linear regression was used to construct QSPKR models for training sets and their predictive performance was evaluated using internal validation (leave-one-out method). All qualified models were validated externally using test sets. QSPKR models were also constructed for compounds in accordance with their 1) net elimination pathways (net secretion, extensive net secretion, net reabsorption, and extensive net reabsorption), 2) net elimination clearances (net secretion clearance, CLSEC; or net reabsorption clearance, CLREAB), 3) ion status, and 4) substrate/inhibitor specificity for renal transporters. We were able to predict 1) CLREAB (Q2 = 0.77) of all compounds undergoing net reabsorption; 2) CLREAB (Q2 = 0.81) of all compounds undergoing extensive net reabsorption; and 3) CLR for substrates and/or inhibitors of OAT1/3 (Q2 = 0.81), OCT2 (Q2 = 0.85), MRP2/4 (Q2 = 0.78), P-gp (Q2 = 0.71), and MATE1/2K (Q2 = 0.81). Moreover, compounds undergoing net reabsorption/extensive net reabsorption predominantly belonged to Biopharmaceutics Drug Disposition Classification System classes 1 and 2. In conclusion, constructed parsimonious QSPKR models can be used to predict CLR of compounds that 1) undergo net reabsorption/extensive net reabsorption and 2) are substrates and/or inhibitors of human renal transporters. PMID:25352657

  6. Detection of human brain cancer infiltration ex vivo and in vivo using quantitative optical coherence tomography.

    PubMed

    Kut, Carmen; Chaichana, Kaisorn L; Xi, Jiefeng; Raza, Shaan M; Ye, Xiaobu; McVeigh, Elliot R; Rodriguez, Fausto J; Quiñones-Hinojosa, Alfredo; Li, Xingde

    2015-06-17

    More complete brain cancer resection can prolong survival and delay recurrence. However, it is challenging to distinguish cancer from noncancer tissues intraoperatively, especially at the transitional, infiltrative zones. This is especially critical in eloquent regions (for example, speech and motor areas). This study tested the feasibility of label-free, quantitative optical coherence tomography (OCT) for differentiating cancer from noncancer in human brain tissues. Fresh ex vivo human brain tissues were obtained from 32 patients with grade II to IV brain cancer and 5 patients with noncancer brain pathologies. On the basis of volumetric OCT imaging data, pathologically confirmed brain cancer tissues (both high- and low-grade) had significantly lower optical attenuation values at both cancer core and infiltrated zones when compared with noncancer white matter, and OCT achieved high sensitivity and specificity at an attenuation threshold of 5.5 mm(-1) for brain cancer patients. We also used this attenuation threshold to confirm the intraoperative feasibility of performing in vivo OCT-guided surgery using a murine model harboring human brain cancer. Our OCT system was capable of processing and displaying a color-coded optical property map in real time at a rate of 110 to 215 frames per second, or 1.2 to 2.4 s for an 8- to 16-mm(3) tissue volume, thus providing direct visual cues for cancer versus noncancer areas. Our study demonstrates the translational and practical potential of OCT in differentiating cancer from noncancer tissue. Its intraoperative use may facilitate safe and extensive resection of infiltrative brain cancers and consequently lead to improved outcomes when compared with current clinical standards. PMID:26084803

  7. Quantitative comparison of cerebral artery development in metatherians and monotremes with non-human eutherians.

    PubMed

    Ashwell, Ken W S; Shulruf, Boaz

    2016-03-01

    A quantitative comparison of the internal diameters of cerebral feeder arteries (internal carotid and vertebral) and the aorta in developing non-human eutherians, metatherians and monotremes has been made, with the aim of determining if there are differences in cerebral arterial flow between the three infraclasses of mammals such as might reflect differences in metabolism of the developing brain. There were no significant differences between eutherians and metatherians in the internal radius of the aorta or the thickness of the aortic wall, but aortic internal radius was significantly smaller in developing monotremes than therians at the < 10 mm body length range. Aortic thickness in the developing monotremes also rose at a slower rate relative to body length than in metatherians or eutherians. The sums of the internal calibres of the internal carotid and vertebral arteries were significantly lower in metatherians as a group and monotremes compared with non-human eutherians at body lengths up to 20 mm and in metatherians at > 20 mm body length. The internal calibre of the internal carotids relative to the sum of all cerebral feeder arteries was also significantly lower in monotremes at < 10 mm body length compared with eutherians. It was noted that dasyurids differed from other metatherians in several measures of cerebral arterial calibre and aortic internal calibre. The findings suggest that: (i) both aortic outflow and cerebral arterial inflow may be lower in developing monotremes than in therians, particularly at small body size (< 20 mm); (ii) cerebral inflow may be lower in some developing metatherians than non-human eutherians; and (iii) dasyurids have unusual features of cerebral arteries possibly related to the extreme immaturity and small size at which they are born. The findings have implications for nutritional sourcing of the developing brain in the three infraclasses of mammals. PMID:26644330

  8. Quantitative determination of fluoxetine in human serum by high performance thin layer chromatography.

    PubMed

    Mennickent, Sigrid; Fierro, Ricardo; Vega, Mario; De Diego, Marta; Godoy, C Gloria

    2010-07-01

    A high performance thin layer chromatographic method was developed and validated for the quantification of fluoxetine in human serum. Fluoxetine was extracted by liquid-liquid extraction method with diethyl ether as extraction solvent. Imipramine was used as internal standard. The chromatographic separation was achieved on precoated silica gel F 254 high performance thin layer chromatographic plates using a mixture of toluene/acetic acid glacial (4:5 v/v) as mobile phase. 4-Dimethylamino-azobenzene-4-sulphonyl chloride was used as derivatization reagent. Densitometric detection was done at 272 nm. The method was linear between 12.5 and 87.5 ng/spot, corresponding to 0.05 and 0.35 ng/microL of fluoxetine in human serum after extraction process and applying 25 microL to the chromatographic plates. The method correlation coefficient was 0.999. The intra-assay and inter-assay precisions, expressed as the RSD, were in the range of 0.70-2.01% (n=3) and 0.81-3.90% (n=9), respectively. The LOD was 0.23 ng, and the LOQ was 0.70 ng. The method proved be accurate, with a recovery between 94.75 and 98.95%, with a RSD not higher than 3.61% and was selective for the active principle tested. This method was successfully applied to quantify fluoxetine in patient serum samples. In conclusion, the method is useful for quantitative determination of fluoxetine in human serum. PMID:20533339

  9. Detection of Human Brain Cancer Infiltration ex vivo and in vivo Using Quantitative Optical Coherence Tomography*

    PubMed Central

    Kut, Carmen; Chaichana, Kaisorn L.; Xi, Jiefeng; Raza, Shaan M.; Ye, Xiaobu; McVeigh, Elliot R.; Rodriguez, Fausto J.; Quinones-Hinojosa, Alfredo; Li, Xingde

    2015-01-01

    More complete brain cancer resection can prolong survival and delay recurrence. However, it is challenging to distinguish cancer from non-cancer tissues intraoperatively, especially at the transitional, infiltrative zones. This is especially critical in eloquent regions (e.g. speech and motor areas). This study tested the feasibility of label-free, quantitative optical coherence tomography (OCT) for differentiating cancer from non-cancer in human brain tissues. Fresh ex vivo human brain tissues were obtained from 32 patients with grades II-IV brain cancer and 5 patients with non-cancer brain pathologies. Based on volumetric OCT imaging data, pathologically confirmed brain cancer tissues (both high-grade and low-grade) had significantly lower optical attenuation values at both cancer core and infiltrated zones when compared with non-cancer white matter, and OCT achieved high sensitivity and specificity at an attenuation threshold of 5.5 mm-1 for brain cancer patients. We also used this attenuation threshold to confirm the intraoperative feasibility of performing in vivo OCT-guided surgery using a murine model harboring human brain cancer. Our OCT system was capable of processing and displaying a color-coded optical property map in real time at a rate of 110-215 frames per second, or 1.2-2.4 seconds for an 8-16 mm3 tissue volume, thus providing direct visual cues for cancer versus non-cancer areas. Our study demonstrates the translational and practical potential of OCT in differentiating cancer from non-cancer tissue. Its intraoperative use may facilitate safe and extensive resection of infiltrative brain cancers and consequently lead to improved outcomes when compared with current clinical standards. PMID:26084803

  10. Quantitative analysis of intact apolipoproteins in human HDL by top-down differential mass spectrometry

    PubMed Central

    Mazur, Matthew T.; Cardasis, Helene L.; Spellman, Daniel S.; Liaw, Andy; Yates, Nathan A.; Hendrickson, Ronald C.

    2010-01-01

    Top-down mass spectrometry holds tremendous potential for the characterization and quantification of intact proteins, including individual protein isoforms and specific posttranslationally modified forms. This technique does not require antibody reagents and thus offers a rapid path for assay development with increased specificity based on the amino acid sequence. Top-down MS is efficient whereby intact protein mass measurement, purification by mass separation, dissociation, and measurement of product ions with ppm mass accuracy occurs on the seconds to minutes time scale. Moreover, as the analysis is based on the accurate measurement of an intact protein, top-down mass spectrometry opens a research paradigm to perform quantitative analysis of “unknown” proteins that differ in accurate mass. As a proof of concept, we have applied differential mass spectrometry (dMS) to the top-down analysis of apolipoproteins isolated from human HDL3. The protein species at 9415.45 Da demonstrates an average fold change of 4.7 (p-value 0.017) and was identified as an O-glycosylated form of apolipoprotein C-III [NANA-(2 → 3)-Gal-β(1 → 3)-GalNAc, +656.2037 Da], a protein associated with coronary artery disease. This work demonstrates the utility of top-down dMS for quantitative analysis of intact protein mixtures and holds potential for facilitating a better understanding of HDL biology and complex biological systems at the protein level. PMID:20388904

  11. Solid-State Quantitative (1)H and (31)P MRI of Cortical Bone in Humans.

    PubMed

    Seifert, Alan C; Wehrli, Felix W

    2016-06-01

    Magnetic resonance imaging (MRI) plays a pivotal role for assessment of the musculoskeletal system. It is currently the clinical modality of choice for evaluation of soft tissues including cartilage, ligaments, tendons, muscle, and bone marrow. By comparison, the study of calcified tissue by MRI is still in its infancy. In this article, we review the potential of the modality for assessment of cortical bone properties known to be affected in degenerative bone disease, with focus on parameters related to matrix and mineral densities, and porosity, by means of emerging solid-state (1)H and (31)P MRI techniques. In contrast to soft tissues, the MRI signal in calcified tissues has very short lifetime, on the order of 100 μs to a few milliseconds, demanding customized imaging approaches that allow capture of the signal almost immediately after excitation. The technologies described are suited for quantitatively imaging human cortical bone in specimens as well as in vivo in patients on standard clinical imagers, yielding either concentrations in absolute units when measured against a reference standard, or more simply, in the form of surrogate biomarkers. The two major water fractions in cortical bone are those of collagen-bound and pore water occurring at an approximately 3:1 ratio. Collagen-bound water density provides a direct quantitative measure of osteoid density. While at an earlier stage of development, quantification of mineral phosphorus by (31)P MRI yields mineral density and, together with knowledge of matrix density, should allow quantification of the degree of bone mineralization. PMID:27048472

  12. Quantitative analysis of endovenous laser ablation based on human vein optical properties

    NASA Astrophysics Data System (ADS)

    Nozoe, Saki; Honda, Norihiro; Ishii, Katsunori; Awazu, Kunio

    2011-07-01

    Endovenous laser ablation (EVLA) is a common treatment method for varicose vein. However, the precise irradiation dose for EVLA is not understood quantitatively. The objective of this study is to evaluate EVLA quantitatively based on optical properties of the varicose vein tissue, and compare the efficacy and the safety at wavelengths of 980 nm and 1470 nm. A human varicose vein tissue was used as a sample. The samples were irradiated by using the 980 nm and 1470 nm laser diodes in various irradiation parameters. The power density was varied from 260 to 1710 W/cm2 and the irradiation time was varied from 3 to 10 s. The optical properties of samples were determined by using a double integrating sphere and an inverse Monte Carlo method. The optical penetration depth of samples was estimated from the optical properties. In the 980 nm laser irradiation, the initial shrinkage of the tissue was observed during laser irradiation conducted at the average energy density of 3630 J/cm2 (1210 W/cm2, 3 s). In the 1470 nm laser irradiation, the initial shrinkage of the tissue was observed during laser irradiation conducted at the average energy density of 2600 J/cm2 (260 W/cm2, 10 s). Penetration depth of the vein wall at the wavelength of 980 nm and 1470 nm were 1.3 mm and 0.22 mm, respectively. The sample irradiated with the 1470 nm laser diode showed vein shrinkage in lower energy density than the 980 nm laser irradiation. The penetration depth at the wavelength of 1470 nm was smaller than the sample thickness about 0.8 mm. These data indicate that EVLA with the 1470 nm laser diode may be more effective and safer than EVLA with the 980 nm laser diode.

  13. Quantitative analysis of glycerol levels in human urine by liquid chromatography-tandem mass spectrometry.

    PubMed

    Dong, Ying; Ma, Yanhua; Yan, Kuan; Shen, Li; Wang, Xiaobing; Xu, Youxuan; He, Genye; Wu, Yun; Lu, Jianghai; Yang, Zhiyong; Feng, Feifei

    2014-04-15

    Glycerol has the latent capacity to act as a plasma volume expander and disguise blood doping practices. Therefore, it has been prohibited in sports as a masking agent by the World Anti-Doping Agency (WADA) since January 2010 and a urinary threshold (1mg/mL) was recommended recently [1]. The purpose of this study was to establish and validate a novel quantitative method for the determination of urinary glycerol concentrations using a liquid chromatography-tandem mass spectrometry approach. This simple yet highly specific method made use of the derivatization of glycerol by benzoyl chloride in aqueous solution at 40°C followed by analysis via LC-ESI-MS/MS without sample pre-concentration or cleanup. The assay was linear over the concentration range of 1.0-1000μg/mL for glycerol in human urine. The lower limit of detection (LLOD) and lower limit of quantitation (LLOQ) were 0.3μg/mL and 1.0μg/mL, respectively. The intra- and inter-day precision of the method at three concentration levels (3, 500 and 900μg/mL) was less than 12.2%. The method also afforded satisfactory results in terms of accuracy, derivatization yield, extraction recovery, matrix effect and specificity. The method has been successfully applied to the detection of glycerol in "Quality Assurance Program" samples provided by the World Association of Anti-Doping Scientists (WAADS) and routine doping-control samples in our laboratory. PMID:24657408

  14. A quantitative and high-throughput assay of human papillomavirus DNA replication.

    PubMed

    Gagnon, David; Fradet-Turcotte, Amélie; Archambault, Jacques

    2015-01-01

    Replication of the human papillomavirus (HPV) double-stranded DNA genome is accomplished by the two viral proteins E1 and E2 in concert with host DNA replication factors. HPV DNA replication is an established model of eukaryotic DNA replication and a potential target for antiviral therapy. Assays to measure the transient replication of HPV DNA in transfected cells have been developed, which rely on a plasmid carrying the viral origin of DNA replication (ori) together with expression vectors for E1 and E2. Replication of the ori-plasmid is typically measured by Southern blotting or PCR analysis of newly replicated DNA (i.e., DpnI digested DNA) several days post-transfection. Although extremely valuable, these assays have been difficult to perform in a high-throughput and quantitative manner. Here, we describe a modified version of the transient DNA replication assay that circumvents these limitations by incorporating a firefly luciferase expression cassette in cis of the ori. Replication of this ori-plasmid by E1 and E2 results in increased levels of firefly luciferase activity that can be accurately quantified and normalized to those of Renilla luciferase expressed from a control plasmid, thus obviating the need for DNA extraction, digestion, and analysis. We provide a detailed protocol for performing the HPV type 31 DNA replication assay in a 96-well plate format suitable for small-molecule screening and EC50 determinations. The quantitative and high-throughput nature of the assay should greatly facilitate the study of HPV DNA replication and the identification of inhibitors thereof. PMID:25348316

  15. Quantitation of Human Papillomavirus DNA in Plasma of Oropharyngeal Carcinoma Patients

    SciTech Connect

    Cao Hongbin; Banh, Alice; Kwok, Shirley; Shi Xiaoli; Wu, Simon; Krakow, Trevor; Khong, Brian; Bavan, Brindha; Bala, Rajeev; Pinsky, Benjamin A.; Colevas, Dimitrios; Pourmand, Nader; Koong, Albert C.; Kong, Christina S.; Le, Quynh-Thu

    2012-03-01

    Purpose: To determine whether human papillomavirus (HPV) DNA can be detected in the plasma of patients with HPV-positive oropharyngeal carcinoma (OPC) and to monitor its temporal change during radiotherapy. Methods and Materials: We used polymerase chain reaction to detect HPV DNA in the culture media of HPV-positive SCC90 and VU147T cells and the plasma of SCC90 and HeLa tumor-bearing mice, non-tumor-bearing controls, and those with HPV-negative tumors. We used real-time quantitative polymerase chain reaction to quantify the plasma HPV DNA in 40 HPV-positive OPC, 24 HPV-negative head-and-neck cancer patients and 10 non-cancer volunteers. The tumor HPV status was confirmed by p16{sup INK4a} staining and HPV16/18 polymerase chain reaction or HPV in situ hybridization. A total of 14 patients had serial plasma samples for HPV DNA quantification during radiotherapy. Results: HPV DNA was detectable in the plasma samples of SCC90- and HeLa-bearing mice but not in the controls. It was detected in 65% of the pretreatment plasma samples from HPV-positive OPC patients using E6/7 quantitative polymerase chain reaction. None of the HPV-negative head-and-neck cancer patients or non-cancer controls had detectable HPV DNA. The pretreatment plasma HPV DNA copy number correlated significantly with the nodal metabolic tumor volume (assessed using {sup 18}F-deoxyglucose positron emission tomography). The serial measurements in 14 patients showed a rapid decline in HPV DNA that had become undetectable at radiotherapy completion. In 3 patients, the HPV DNA level had increased to a discernable level at metastasis. Conclusions: Xenograft studies indicated that plasma HPV DNA is released from HPV-positive tumors. Circulating HPV DNA was detectable in most HPV-positive OPC patients. Thus, plasma HPV DNA might be a valuable tool for identifying relapse.

  16. Locating human quantitative trait loci: guidelines for the selection of sibling pairs for genotyping.

    PubMed

    Eaves, L; Meyer, J

    1994-09-01

    Simulation studies were conducted to assess the relative merits of different nonrandom sampling strategies for the selection of sibling pairs for genotyping in the attempt to locate individual loci (QTLs) contributing to variation in human quantitative traits. For a constant amount of variation contributed by a QTL (25% of the total) the frequencies and dominance relationships of a trait increasing allele were varied. Three strategies for selection of pairs for genotyping were based on the phenotypic values of the siblings: "Concordant sib pairs" (CSP) are pairs in which both individuals exceed a given threshold value; "discordant sib pairs" (DSP) are pairs in which one member exceeds a given upper threshold and the other is below a specified lower threshold; and "most similar pairs" (MSP) are pairs selected for falling below a specified percentile ranking of the within-pair mean square for the quantitative trait. Tests for linkage with markers at 1, 2, 5, 10, and 20 cM from each of the QTLs were conducted for each of the selected samples and compared with tests based on the regression, in the entire sample, of within pair variation on the proportion of alleles identical by descent (IBD) at each marker locus. Tests for the effect of the increasing allele at the QTL ("candidate gene") were also conducted for the DSP pairs. No single nonrandom selection procedure yields as much as half the information realized in the total sample. However, a combined strategy which involves genotyping the 5% of MSP and DSP for the upper and lower quintiles of values of the quantitative trait (a further 3% of the sample approximately) yields lod scores which are usually more than 65% of the values realized for the entire sample. Tests comparing the proportion of increasing alleles in high- and low-scoring siblings from DSP samples are uniformly very powerful for detecting candidate loci. Even when it is not possible to measure the entire range of the phenotype with uniform precision

  17. Quantitation of leukotriene B(4) in human sputum as a biomarker using UPLC-MS/MS.

    PubMed

    Jian, Wenying; Edom, Richard W; Xue, Xiaohua; Huang, Mike-Qingtao; Fourie, Anne; Weng, Naidong

    2013-08-01

    Leukotriene B4 (LTB4) is a potent mediator of inflammation and has been recognized as an important target for therapeutic intervention for treatment of diseases such as asthma. In the current work, a highly selective and sensitive UPLC-MS/MS assay was developed for quantitation of LTB4 in human sputum as a biomarker for LTB4 biosynthesis inhibition. A fit-for-purpose strategy for method development, assay qualification, and study support was adopted for this biomarker project. A surrogate matrix (protein buffer) was used for preparation of calibration samples and certain levels of quality control (QC) samples to avoid interference from endogenous analyte, while the low QC was prepared in authentic matrix, human sputum. The analytical methodology utilized a liquid-liquid extraction procedure in 96-well plate format. Chromatographic separation was achieved with a reversed-phase ultra high pressure liquid chromatography (UPLC) column using gradient elution, and the run time was 4.5min per sample. The lower limit of quantitation (LLOQ) was 0.2ng/mL, and the calibration curve range was 0.2-20ng/mL. Acceptable accuracy, precision, linearity, specificity, recovery, and matrix effect was obtained. Bench-top stability (6h), freeze-thaw stability (3 cycles at -20°C), and autosampler stability (97h at ambient temperature) all met acceptance criteria. Frozen long-term stability for 166 days at -20°C in sputum did not meet acceptance criteria by showing only ≥75% of nominal concentration and the information was taken into consideration for study support. Two important observations in the current work were: (1) LTB4 was unstable in sputum in the presence of liquification reagent dithiothreitol (DTT). Therefore, a non-DTT treatment method for sputum processing was developed and applied to the bioanalytical assay and clinical study support; and (2) chromatographic separation of LTB4 from its three non-enzymatically derived isomers, i.e. 6-trans-LTB4, 12-epi-LTB4, and 6-trans

  18. GWAS implicates a role for quantitative immune traits and threshold effects in risk for human autoimmune disorders

    PubMed Central

    Gregersen, Peter K.; Diamond, Betty; Plenge, Robert M.

    2016-01-01

    Genome wide association studies in human autoimmune disorders has provided a long list of alleles with rather modest degrees of risk. A large fraction of these associations are likely due to either quantitative differences in gene expression or amino acid changes that regulate quantitative aspects of the immune response. While functional studies are still lacking for most of these associations, we present examples of autoimmune disease risk alleles that influence quantitative changes in lymphocyte activation, cytokine signaling and dendritic cell function. The analysis of immune quantitative traits associated with autoimmune loci is clearly going to be an important component of understanding the pathogenesis of autoimmunity. This will require both new and more efficient ways of characterizing the normal immune system, as well as large population resources with which genotype-phenotype correlations can be convincingly demonstrated. Future development of new therapies will depend on understanding the mechanistic underpinnings of immune regulation by these new risk loci. PMID:23026397

  19. Examination of Information Technology (IT) Certification and the Human Resources (HR) Professional Perception of Job Performance: A Quantitative Study

    ERIC Educational Resources Information Center

    O'Horo, Neal O.

    2013-01-01

    The purpose of this quantitative survey study was to test the Leontief input/output theory relating the input of IT certification to the output of the English-speaking U.S. human resource professional perceived IT professional job performance. Participants (N = 104) rated their perceptions of IT certified vs. non-IT certified professionals' job…

  20. Single Laboratory Comparison of Quantitative Real-Time PCR Assays for the Detection of Human Fecal Pollution

    EPA Science Inventory

    There are numerous quantitative real-time PCR (qPCR) methods available to detect and enumerate human fecal pollution in ambient waters. Each assay employs distinct primers and/or probes and many target different genes and microorganisms leading to potential variations in method ...

  1. Single Laboratory Comparison of Quantitative Real-Time PCR Assays for the Detection of Human Fecal Pollution - Poster

    EPA Science Inventory

    There are numerous quantitative real-time PCR (qPCR) methods available to detect and enumerate human fecal pollution in ambient waters. Each assay employs distinct primers and/or probes and many target different genes and microorganisms leading to potential variations in method p...

  2. Data Acceptance Criteria for Standardized Human-Associated Fecal Source Identification Quantitative Real-Time PCR Methods

    EPA Science Inventory

    There is a growing interest in the application of human-associated fecal sourceidentification quantitative real-time PCR (qPCR) technologies for water quality management. The transition from a research tool to a standardized protocol requires a high degree of confidence in data q...

  3. 3-D imaging and quantitative comparison of human dentitions and simulated bite marks.

    PubMed

    Blackwell, S A; Taylor, R V; Gordon, I; Ogleby, C L; Tanijiri, T; Yoshino, M; Donald, M R; Clement, J G

    2007-01-01

    This study presents a technique developed for 3-D imaging and quantitative comparison of human dentitions and simulated bite marks. A sample of 42 study models and the corresponding bites, made by the same subjects in acrylic dental wax, were digitised by laser scanning. This technique allows image comparison of a 3-D dentition with a 3-D bite mark, eliminating distortion due to perspective as experienced in conventional photography. Cartesian co-ordinates of a series of landmarks were used to describe the dentitions and bite marks, and a matrix was created to compare all possible combinations of matches and non-matches using cross-validation techniques. An algorithm, which estimated the probability of a dentition matching its corresponding bite mark, was developed. A receiver operating characteristic graph illustrated the relationship between values for specificity and sensitivity. This graph also showed for this sample that 15% of non-matches could not be distinguished from the true match, translating to a 15% probability of falsely convicting an innocent person. PMID:16391946

  4. Development of phantom for quantitative analyses of human dentin mineral density.

    PubMed

    Hayashi-Sakai, Sachiko; Kondo, Tatsuya; Kasuga, Yuto; Sakamoto, Makoto; Endo, Hideaki; Sakai, Jun

    2015-01-01

    The purpose of the present study was to develop a novel-designed phantom that could be scanned with a sample in the same image, that specialize for quantitative analyses of human dentin mineral density using the X-ray attenuation method. A further attempt was made to demonstrate the intracoronal dentin mineral density using this phantom in mandibular incisors. The phantom prepared with a 15 mm hole in the center of an acrylic resin bar having an outside diameter of 25 mm and 8 small holes (diameter, 3 mm) were made at equal intervals around the center. Liquid dipotassium hydrogen phosphate (K2HPO4) solutions were established at 0.4, 0.6, 0.8 and 1.0 g/cm3, and were arranged to these holes. The mean value of the intracoronal dentin mineral density was 1.486 ± 0.016 g/cm3 in the present study. As the results of the present study corresponded to previous reports, this new phantom was considered to be useful. This phantom enables the analysis of samples that are not readily available by conventional mechanical tests and may facilitate biomechanical investigations using X-ray images. It was suggested that this system is a simple, accurate and novel mineralization measuring system. PMID:26484556

  5. Quantitative analysis of miRNA expression in seven human foetal and adult organs.

    PubMed

    Tang, Yanping; Liu, Dong; Zhang, Lijie; Ingvarsson, Sigurdur; Chen, Huiping

    2011-01-01

    miRNAs have been found to repress gene expression at posttranscriptional level in cells. Studies have shown that expression of miRNAs is tissue-specific and developmental-stage-specific. The mechanism behind this could be explained by miRNA pathways. In this study, totally 54 miRNAs were analysed in 7 matched human foetal and adult organs (brain, colon, heart, kidney, liver, lung and spleen) using real-time PCR. Quantitative analysis showed that a big proportion of the 54 miRNAs have higher general expression in the organs of the foetal period than the adult period, with the exception of the heart. The miRNA gene promoter methylation level in the adult stages was higher than in the foetal stages. Moreover, there is a high general expression level of several miRNAs in both stages of brain, kidney, liver, lung and spleen, but not seen in colon and heart. Our results indicate that the miRNAs may play a bigger role in the foetal stage than the adult stage of brain, colon, kidney, liver, lung and spleen. The majority of the miRNAs analysed may play an important role in the growth and development of brain, kidney, liver, lung and spleen. However, a minority of the miRNAs may be functional in colon and heart. PMID:22194897

  6. Quantitative Phosphoproteomic Profiling of Human Non-Small Cell Lung Cancer Tumors

    PubMed Central

    Schweppe, Devin K.; Rigas, James R.; Gerber, Scott A.

    2013-01-01

    Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related deaths worldwide. Within the molecular scope of NCSLC, a complex landscape of dysregulated cellular signaling has emerged, defined largely by mutations in select mediators of signal transduction, including the epidermal growth factor receptor (EGFR) and anaplastic lymphoma (ALK) kinases. Consequently, these mutant kinases become constitutively activated and targets for chemotherapeutic intervention. Encouragingly, small molecule inhibitors of these pathways have shown promise in clinical trials or are approved for clinical use. However, many protein kinases are dysregulated in NSCLC without genetic mutations. To quantify differences in tumor cell signaling that are transparent to genomic methods, we established a super-SILAC internal standard derived from NSCLC cell lines grown in vitro and labeled with heavy lysine and arginine, and deployed them in a phosphoproteomics workflow. We identified 9019 and 8753 phosphorylation sites in two separate tumors. Relative quantification of phosphopeptide abundance between tumor samples allowed for the determination of specific hubs and pathways differing between each tumor. Sites downstream of Ras showed decreased inhibitory phosphorylation (Raf/Mek) and increased activating phosphorylation (Erk1/2) in one tumor versus another. In this way, we were able to quantitatively access oncogenic kinase signaling in primary human tumors. PMID:23911959

  7. Quantitative measurements of human sperm nuclei using automated microscopy and image analysis

    SciTech Connect

    Wyrobek, A.J.; Firpo, M. ); Sudar, D. )

    1993-01-01

    A package of computer codes, called Morphometry Automation Program (MAP), was developed to (a) detect human sperm smeared onto glass slides, (b) measure more than 30 aspects of the size, shape, texture, and staining of their nuclei, and (c) retain operator evaluation of the process. MAP performs the locating and measurement functions automatically, without operator assistance. In addition to standard measurements, MAP utilizes axial projections of nuclear area and stain intensity to detect asymmetries. MAP also stores for each cell the gray-scale images for later display and evaluation, and it retains coordinates for optional relocation and inspection under the microscope. MAP operates on the Quantitative Image Processing System (QUIPS) at LLNL. MAP has potential applications in the evaluation of infertility and in reproductive toxicology, such as (a) classifying sperm into clinical shape categories for assessing fertility status, (b) identifying subtle effects of host factors (diet, stress, etc.), (c) assessing the risk of potential spermatogenic toxicants (tobacco, drugs, etc.), and (d) investigating associations with abnormal pregnancy outcomes (time to pregnancy, early fetal loss, etc.).

  8. Quantitative comparison of a human cancer cell surface proteome between interphase and mitosis

    PubMed Central

    Özlü, Nurhan; Qureshi, Mohammad H; Toyoda, Yusuke; Renard, Bernhard Y; Mollaoglu, Gürkan; Özkan, Nazlı E; Bulbul, Selda; Poser, Ina; Timm, Wiebke; Hyman, Anthony A; Mitchison, Timothy J; Steen, Judith A

    2015-01-01

    The cell surface is the cellular compartment responsible for communication with the environment. The interior of mammalian cells undergoes dramatic reorganization when cells enter mitosis. These changes are triggered by activation of the CDK1 kinase and have been studied extensively. In contrast, very little is known of the cell surface changes during cell division. We undertook a quantitative proteomic comparison of cell surface-exposed proteins in human cancer cells that were tightly synchronized in mitosis or interphase. Six hundred and twenty-eight surface and surface-associated proteins in HeLa cells were identified; of these, 27 were significantly enriched at the cell surface in mitosis and 37 in interphase. Using imaging techniques, we confirmed the mitosis-selective cell surface localization of protocadherin PCDH7, a member of a family with anti-adhesive roles in embryos. We show that PCDH7 is required for development of full mitotic rounding pressure at the onset of mitosis. Our analysis provided basic information on how cell cycle progression affects the cell surface. It also provides potential pharmacodynamic biomarkers for anti-mitotic cancer chemotherapy. PMID:25476450

  9. Quantitative Microbial Risk Assessment in Occupational Settings Applied to the Airborne Human Adenovirus Infection.

    PubMed

    Carducci, Annalaura; Donzelli, Gabriele; Cioni, Lorenzo; Verani, Marco

    2016-01-01

    Quantitative Microbial Risk Assessment (QMRA) methodology, which has already been applied to drinking water and food safety, may also be applied to risk assessment and management at the workplace. The present study developed a preliminary QMRA model to assess microbial risk that is associated with inhaling bioaerosols that are contaminated with human adenovirus (HAdV). This model has been applied to air contamination data from different occupational settings, including wastewater systems, solid waste landfills, and toilets in healthcare settings and offices, with different exposure times. Virological monitoring showed the presence of HAdVs in all the evaluated settings, thus confirming that HAdV is widespread, but with different average concentrations of the virus. The QMRA results, based on these concentrations, showed that toilets had the highest probability of viral infection, followed by wastewater treatment plants and municipal solid waste landfills. Our QMRA approach in occupational settings is novel, and certain caveats should be considered. Nonetheless, we believe it is worthy of further discussions and investigations. PMID:27447658

  10. Identification of Mediator Kinase Substrates in Human Cells using Cortistatin A and Quantitative Phosphoproteomics.

    PubMed

    Poss, Zachary C; Ebmeier, Christopher C; Odell, Aaron T; Tangpeerachaikul, Anupong; Lee, Thomas; Pelish, Henry E; Shair, Matthew D; Dowell, Robin D; Old, William M; Taatjes, Dylan J

    2016-04-12

    Cortistatin A (CA) is a highly selective inhibitor of the Mediator kinases CDK8 and CDK19. Using CA, we now report a large-scale identification of Mediator kinase substrates in human cells (HCT116). We identified over 16,000 quantified phosphosites including 78 high-confidence Mediator kinase targets within 64 proteins, including DNA-binding transcription factors and proteins associated with chromatin, DNA repair, and RNA polymerase II. Although RNA-seq data correlated with Mediator kinase targets, the effects of CA on gene expression were limited and distinct from CDK8 or CDK19 knockdown. Quantitative proteome analyses, tracking around 7,000 proteins across six time points (0-24 hr), revealed that CA selectively affected pathways implicated in inflammation, growth, and metabolic regulation. Contrary to expectations, increased turnover of Mediator kinase targets was not generally observed. Collectively, these data support Mediator kinases as regulators of chromatin and RNA polymerase II activity and suggest their roles extend beyond transcription to metabolism and DNA repair. PMID:27050516

  11. Quantitative Microbial Risk Assessment in Occupational Settings Applied to the Airborne Human Adenovirus Infection

    PubMed Central

    Carducci, Annalaura; Donzelli, Gabriele; Cioni, Lorenzo; Verani, Marco

    2016-01-01

    Quantitative Microbial Risk Assessment (QMRA) methodology, which has already been applied to drinking water and food safety, may also be applied to risk assessment and management at the workplace. The present study developed a preliminary QMRA model to assess microbial risk that is associated with inhaling bioaerosols that are contaminated with human adenovirus (HAdV). This model has been applied to air contamination data from different occupational settings, including wastewater systems, solid waste landfills, and toilets in healthcare settings and offices, with different exposure times. Virological monitoring showed the presence of HAdVs in all the evaluated settings, thus confirming that HAdV is widespread, but with different average concentrations of the virus. The QMRA results, based on these concentrations, showed that toilets had the highest probability of viral infection, followed by wastewater treatment plants and municipal solid waste landfills. Our QMRA approach in occupational settings is novel, and certain caveats should be considered. Nonetheless, we believe it is worthy of further discussions and investigations. PMID:27447658

  12. Quantitative proteomics analyses of activation states of human THP-1 macrophages.

    PubMed

    Meijer, Kees; Weening, Desiree; de Vries, Marcel P; Priebe, Marion G; Vonk, Roel J; Roelofsen, Han

    2015-10-14

    Macrophages display large functional and phenotypical plasticity. They can adopt a broad range of activation states depending on their microenvironment. Various surface markers are used to characterize these differentially polarized macrophages. However, this is not informative for the functions of the macrophage. In order to have a better understanding of the functional changes of macrophages upon differential polarization, we studied differences in LPS- and IL4-stimulated macrophages. The THP-1 human monocytic cell line, was used as a model system. Cells were labeled, differentiated and stimulated with either LPS or IL-4 in a quantitative SILAC proteomics set-up. The resulting sets of proteins were functionally clustered. LPS-stimulated macrophages show increased secretion of proinflammatory peptides, leading to increased pressure on protein biosynthesis and processing. IL4-stimulated macrophages show upregulation of cell adhesion and extracellular matrix remodeling. Our approach provides an integrated view of polarization-induced functional changes and proves useful for studying functional differences between subsets of macrophages. Moreover, the identified polarization specific proteins may contribute to a better characterization of different activation states in situ and their role in various inflammatory processes. PMID:26200757

  13. Quantitative gas chromatographic mass spectrometric determination of pinaverium-bromide in human serum.

    PubMed

    de Weerdt, G A; Beke, R P; Verdievel, H G; Barbier, F; Jonckheere, J A; de Leenheer, A P

    1983-03-01

    A method has been developed for quantitative determination of pinaverium-bromide, a quaternary ammonium derivative with papaverine-like activity, in human serum. The method involves a chloroform extraction of serum spiked with N-(6,6-dimethyl bicyclo[3.1.1]2-heptenyl-ethoxyethyl) perhydro-1,4-oxazine as internal standard. After evaporation of the solvent, and reduction of the residue with Raney-Nickel, the internal standard and the reduced pinaverium-bromide are re-extracted from the reaction mixture with toluene and analysed isothermally on a fused silica column coated with OV-101. Although chemical ionization with methane revealed intense protonated molecular ions for both pinaverium-bromide and the internal standard, selectivity and sensitivity were significantly lower in comparison with electron impact ionization at 70 e V. Therefore, quantification was performed in the electron impact mode by single ion monitoring of the common fragment ion at m/z 100.2. A linear detector response was observed up to 160 ng ml-1. A within-run assay precision better than 2% CV (n = 5) was found, and a detection limit of 1 ng pinaverium-bromide ml-1 of serum was attained. PMID:6850068

  14. Quantitative Validation of a Human Body Finite Element Model Using Rigid Body Impacts.

    PubMed

    Vavalle, Nicholas A; Davis, Matthew L; Stitzel, Joel D; Gayzik, F Scott

    2015-09-01

    Validation is a critical step in finite element model (FEM) development. This study focuses on the validation of the Global Human Body Models Consortium full body average male occupant FEM in five localized loading regimes-a chest impact, a shoulder impact, a thoracoabdominal impact, an abdominal impact, and a pelvic impact. Force and deflection outputs from the model were compared to experimental traces and corridors scaled to the 50th percentile male. Predicted fractures and injury severity measures were compared to evaluate the model's injury prediction capabilities. The methods of ISO/TS 18571 were used to quantitatively assess the fit of model outputs to experimental force and deflection traces. The model produced peak chest, shoulder, thoracoabdominal, abdominal, and pelvis forces of 4.8, 3.3, 4.5, 5.1, and 13.0 kN compared to 4.3, 3.2, 4.0, 4.0, and 10.3 kN in the experiments, respectively. The model predicted rib and pelvic fractures related to Abbreviated Injury Scale scores within the ranges found experimentally all cases except the abdominal impact. ISO/TS 18571 scores for the impacts studied had a mean score of 0.73 with a range of 0.57-0.83. Well-validated FEMs are important tools used by engineers in advancing occupant safety. PMID:25739950

  15. Multiplexed femtomolar quantitation of human cytokines in a fluoropolymer microcapillary film.

    PubMed

    Castanheira, Ana P; Barbosa, Ana I; Edwards, Alexander D; Reis, Nuno M

    2015-08-21

    Sensitive quantitation of multiple cytokines can provide important diagnostic information during infection, inflammation and immunopathology. In this study sensitive immunoassay detection of human cytokines IL-1β, IL-6, IL-12p70 and TNFα is shown for singleplex and multiplex formats using a novel miniaturized ELISA platform. The platform uses a disposable plastic multi-syringe aspirator (MSA) integrating 8 disposable fluoropolymer microfluidic test strips, each containing an array of ten 200 μm mean i.d. microcapillaries coated with a set of monoclonal antibodies. Each MSA device thus performs 10 tests on 8 samples, delivering 80 measurements. Unprecedented levels of sensitivity were obtained with the novel fluoropolymer microfluidic material and simple colorimetric detection in a flatbed scanner. The limit of detection for singleplex detection ranged from 2.0 to 15.0 pg ml(-1), i.e. 35 and 713 femtomolar for singleplex cytokine detection, and the intra- and inter-assay coefficient of variation (CV) remained within 10%. In addition, a triplex immunoassay was developed for measuring IL-1β, IL-12p70 and TNFα simultaneously from a given sample in the pg ml(-1) range. These assays permit high sensitivity measurement with rapid <15 min assay or detection from undiluted blood serum. The portability, speed and low-cost of this system are highly suited to point-of-care testing and field diagnostics applications. PMID:26120601

  16. New Real-Time Quantitative PCR Procedure for Quantification of Bifidobacteria in Human Fecal Samples

    PubMed Central

    Gueimonde, Miguel; Tölkkö, Satu; Korpimäki, Teemu; Salminen, Seppo

    2004-01-01

    The application of a real-time quantitative PCR method (5′ nuclease assay), based on the use of a probe labeled at its 5′ end with a stable, fluorescent lanthanide chelate, for the quantification of human fecal bifidobacteria was evaluated. The specificities of the primers and the primer-probe combination were evaluated by conventional PCR and real-time PCR, respectively. The results obtained by real-time PCR were compared with those obtained by fluorescent in situ hybridization, the current gold standard for intestinal microbiota quantification. In general, a good correlation between the two methods was observed. In order to determine the detection limit and the accuracy of the real-time PCR procedure, germfree rat feces were spiked with known amounts of bifidobacteria and analyzed by both methods. The detection limit of the method used in this study was found to be about 5 × 104 cells per g of feces. Both methods, real-time PCR and fluorescent in situ hybridization, led to an accurate quantification of the spiked samples with high levels of bifidobacteria, but real-time PCR was more accurate for samples with low levels. We conclude that the real-time PCR procedure described here is a specific, accurate, rapid, and easy method for the quantification of bifidobacteria in feces. PMID:15240297

  17. Quantitation of intracellular Mac-1 (CD11b/CD18) pools in human neutrophils.

    PubMed

    Jones, D H; Anderson, D C; Burr, B L; Rudloff, H E; Smith, C W; Krater, S S; Schmalstieg, F C

    1988-12-01

    The adhesive glycoprotein Mac-1 (CD11b/CD18) of the CD11/CD18 complex contributes to multiple neutrophil inflammatory functions. Activation of neutrophils by chemotactic stimuli results in a rapid, protein synthesis-independent increase in surface Mac-1 derived from incompletely defined intracellular compartments. Therefore, we developed a novel quantitative lectin immunoblot technique to define intracellular pools of Mac-1 in subcellular neutrophil fractions resolved on discontinuous Percoll gradients. In cavitates of unstimulated neutrophils, 30% and 26% of total Mac-1 was identified in beta [1.10 gm/ml; vitamin B12 binding protein (vit B12 B.P.)-rich] or pre-gamma (1.07 gm/ml; vit B12 B.P.-poor) granular fractions, respectively, whereas 24% was associated with the plasma membrane-rich gamma (1.06 gm/ml) fractions. N-formyl-methionyl-leucyl-phenylalanine (fMLP) stimulation (10(-8) M, 15 min, 37 degrees C) significantly diminished Mac-1 in pre-gamma (-18% of total, P less than 0.05) but not beta fractions (+6% of total). Under these conditions, the content of Mac-1 in gamma fractions increased 13% in association with four- to eightfold increase in surface Mac-1 expression (OKM-1 binding). These findings suggest that chemotactic stimuli increase plasma membrane and/or surface Mac-1 on human neutrophils by mobilizing a novel intracellular granule pool. PMID:2903896

  18. Bromodichloromethane inhibits human placental trophoblast differentiation.

    PubMed

    Chen, Jiangang; Thirkill, Twanda L; Lohstroh, Peter N; Bielmeier, Susan R; Narotsky, Michael G; Best, Deborah S; Harrison, Randy A; Natarajan, Kala; Pegram, Rex A; Overstreet, James W; Lasley, Bill L; Douglas, Gordon C

    2004-03-01

    Epidemiological data suggest an association between exposures to bromodichloromethane (BDCM), a trihalomethane found in drinking water as a result of drinking water disinfection, and an increased risk of spontaneous abortion. We previously hypothesized that BDCM targets the placenta and showed that the secretion of chorionic gonadotrophin (CG) was reduced in primary cultures of human term syncytiotrophoblasts exposed to BDCM. In the present study we extend this observation by evaluating the effects of BDCM on the morphological differentiation of mononucleated cytotrophoblast cells to multinucleated syncytiotrophoblast-like colonies. Addition of BDCM to cytotrophoblast cultures inhibited the subsequent formation of multinucleated colonies in a dose-dependent manner, as determined by immunocytochemical staining for desmosomes and nuclei. The effect was seen at BDCM concentrations between 0.02 and 2 mM and was confirmed by quantitative image analysis. Secretion of bioactive and immunoreactive chorionic gonadotropin was also significantly inhibited in a dose-dependent manner under these culture conditions, and cellular levels of CG were also reduced. Trophoblast viability was not compromised by exposure to BDCM. We conclude that BDCM disrupts syncytiotrophoblast formation and inhibits CG secretion in vitro. Although other tissue targets are not ruled out, these data substantiate the idea that BDCM targets the placenta and could have implications for understanding the adverse pregnancy outcomes associated with BDCM exposure in humans. PMID:14691210

  19. The Proteome of Human Liver Peroxisomes: Identification of Five New Peroxisomal Constituents by a Label-Free Quantitative Proteomics Survey

    PubMed Central

    Ofman, Rob; Bunse, Christian; Pawlas, Magdalena; Hayen, Heiko; Eisenacher, Martin; Stephan, Christian; Meyer, Helmut E.; Waterham, Hans R.; Erdmann, Ralf; Wanders, Ronald J.; Warscheid, Bettina

    2013-01-01

    The peroxisome is a key organelle of low abundance that fulfils various functions essential for human cell metabolism. Severe genetic diseases in humans are caused by defects in peroxisome biogenesis or deficiencies in the function of single peroxisomal proteins. To improve our knowledge of this important cellular structure, we studied for the first time human liver peroxisomes by quantitative proteomics. Peroxisomes were isolated by differential and Nycodenz density gradient centrifugation. A label-free quantitative study of 314 proteins across the density gradient was accomplished using high resolution mass spectrometry. By pairing statistical data evaluation, cDNA cloning and in vivo colocalization studies, we report the association of five new proteins with human liver peroxisomes. Among these, isochorismatase domain containing 1 protein points to the existence of a new metabolic pathway and hydroxysteroid dehydrogenase like 2 protein is likely involved in the transport or β-oxidation of fatty acids in human peroxisomes. The detection of alcohol dehydrogenase 1A suggests the presence of an alternative alcohol-oxidizing system in hepatic peroxisomes. In addition, lactate dehydrogenase A and malate dehydrogenase 1 partially associate with human liver peroxisomes and enzyme activity profiles support the idea that NAD+ becomes regenerated during fatty acid β-oxidation by alternative shuttling processes in human peroxisomes involving lactate dehydrogenase and/or malate dehydrogenase. Taken together, our data represent a valuable resource for future studies of peroxisome biochemistry that will advance research of human peroxisomes in health and disease. PMID:23460848

  20. Quantitative analysis of amino acid oxidation and related gluconeogenesis in humans.

    PubMed

    Jungas, R L; Halperin, M L; Brosnan, J T

    1992-04-01

    Significant gaps remain in our knowledge of the pathways of amino acid catabolism in humans. Further quantitative data describing amino acid metabolism in the kidney are especially needed as are further details concerning the pathways utilized for certain amino acids in liver. Sufficient data do exist to allow a broad picture of the overall process of amino acid oxidation to be developed along with approximate quantitative assessments of the role played by liver, muscle, kidney, and small intestine. Our analysis indicates that amino acids are the major fuel of liver, i.e., their oxidative conversion to glucose accounts for about one-half of the daily oxygen consumption of the liver, and no other fuel contributes nearly so importantly. The daily supply of amino acids provided in the diet cannot be totally oxidized to CO2 in the liver because such a process would provide far more ATP than the liver could utilize. Instead, most amino acids are oxidatively converted to glucose. This results in an overall ATP production during amino acid oxidation very nearly equal to the ATP required to convert amino acid carbon to glucose. Thus gluconeogenesis occurs without either a need for ATP from other fuels or an excessive ATP production that could limit the maximal rate of the process. The net effect of the oxidation of amino acids to glucose in the liver is to make nearly two-thirds of the total energy available from the oxidation of amino acids accessible to peripheral tissues, without necessitating that peripheral tissues synthesize the complex array of enzymes needed to support direct amino acid oxidation. As a balanced mixture of amino acids is oxidized in the liver, nearly all carbon from glucogenic amino acids flows into the mitochondrial aspartate pool and is actively transported out of the mitochondria via the aspartate-glutamate antiport linked to proton entry. In the cytoplasm the aspartate is converted to fumarate utilizing urea cycle enzymes; the fumarate flows via

  1. Quantitative proton MR spectroscopic imaging of normal human cerebellum and brain stem.

    PubMed

    Jacobs, M A; Horská, A; van Zijl, P C; Barker, P B

    2001-10-01

    Quantitative, multislice proton MR spectroscopic imaging (MRSI) was used to investigate regional metabolite levels and ratios in the normal adult human posterior fossa. Six normal volunteers (36 +/- 3 years, five male, one female) were scanned on a 1.5 T scanner using multislice MRSI at long echo time (TE 280 msec). The entire cerebellum was covered using three oblique-axial slice locations, which also included the pons, mid-brain, insular cortex, and parieto-occipital lobe. Concentrations of N-acetylaspartate (NAA), choline (Cho), and creatine (Cr) were estimated using the phantom replacement technique. Regional variations of the concentrations were assessed using ANOVA (P < 0.05). High-resolution MRSI data was obtained in all subjects and brain regions examined. Metabolite concentrations (mM) (mean +/- SD) were as follows: cerebellar vermis: 2.3 +/- 0.4, 8.8 +/- 1.7 and 7.6 +/- 1.0 for Cho, Cr, and NAA respectively; cerebellar hemisphere: 2.2 +/- 0.6, 8.9 +/- 2.1, 7.5 +/- 0.8; pons 2.2 +/- 0.5, 4.3 +/- 1.1, 8.3 +/- 0.9; insular cortex, 1.8 +/- 0.5, 7.8 +/- 2, 8.0 +/- 1.1, parieto-occipital gray matter, 1.3 +/- 0.3, 5.7 +/- 1.1, 7.2 +/- 0.9, and occipital white matter, 1.4 +/- 0.3, 5.3 +/- 1.3, 7.5 +/- 0.8. Consistent with previous reports, significantly higher levels of Cr were found in the cerebellum compared to parieto-occipital gray and occipital white matter, and pons (P < 0.0001). NAA was essentially uniformly distributed within the regions chosen for analysis, with the highest level in the pons (P < 0.04). Cho was significantly higher in the cerebellum and pons than parieto-occipital gray and occipital white matter (P < 0.002) and was also higher in the pons than in the insular cortex (P < 0.05). Quantitative multislice MRSI of the posterior fossa is feasible and significant regional differences in metabolite concentrations were found. PMID:11590646

  2. Measurements of morphology and refractive indexes on human downy hairs using three-dimensional quantitative phase imaging.

    PubMed

    Lee, SangYun; Kim, Kyoohyun; Lee, Yuhyun; Park, Sungjin; Shin, Heejae; Yang, Jongwon; Ko, Kwanhong; Park, HyunJoo; Park, YongKeun

    2015-01-01

    We present optical measurements of morphology and refractive indexes (RIs) of human downy arm hairs using three-dimensional (3-D) quantitative phase imaging techniques. 3-D RI tomograms and high-resolution two-dimensional synthetic aperture images of individual downy arm hairs were measured using a Mach–Zehnder laser interferometric microscopy equipped with a two-axis galvanometer mirror. From the measured quantitative images, the RIs and morphological parameters of downy hairs were noninvasively quantified including the mean RI, volume, cylinder, and effective radius of individual hairs. In addition, the effects of hydrogen peroxide on individual downy hairs were investigated. PMID:26205909

  3. On the quantitative evaluation of edge detection schemes and their comparison with human performance. [image processing of satellite photographs

    NASA Technical Reports Server (NTRS)

    Fram, J. R.; Deutsch, E. S.

    1975-01-01

    A technique for the quantitative evaluation of edge detection schemes is presented. It is used to assess the performance of three such schemes using a specially-generated set of images containing noise. The ability of human subjects to distinguish the edges in the presence of noise is also measured and compared with that of the edge detection schemes. The edge detection schemes are used on a high-resolution satellite photograph with varying degrees of noise added in order to relate the quantitative comparison to real-life imagery.

  4. Measurements of morphology and refractive indexes on human downy hairs using three-dimensional quantitative phase imaging

    NASA Astrophysics Data System (ADS)

    Lee, SangYun; Kim, Kyoohyun; Lee, Yuhyun; Park, Sungjin; Shin, Heejae; Yang, Jongwon; Ko, Kwanhong; Park, HyunJoo; Park, YongKeun

    2015-11-01

    We present optical measurements of morphology and refractive indexes (RIs) of human downy arm hairs using three-dimensional (3-D) quantitative phase imaging techniques. 3-D RI tomograms and high-resolution two-dimensional synthetic aperture images of individual downy arm hairs were measured using a Mach-Zehnder laser interferometric microscopy equipped with a two-axis galvanometer mirror. From the measured quantitative images, the RIs and morphological parameters of downy hairs were noninvasively quantified including the mean RI, volume, cylinder, and effective radius of individual hairs. In addition, the effects of hydrogen peroxide on individual downy hairs were investigated.

  5. Human figure drawings and house tree person drawings as indicators of self-esteem: a quantitative approach.

    PubMed

    Groth-Marnat, G; Roberts, L

    1998-02-01

    This study assessed the concurrent validity of Human Figure Drawings (HFD) and House-Tree-Person (HTP) drawings as measures of self-esteem. Adult subjects were requested to make HFD and HTP drawings and to complete measures of psychological adjustment which included the Coopersmith Self Esteem Inventory and Tennessee Self Concept Scale. The drawings were scored using a quantitative, composite rating scale derived from HFD and HTP empirical and theoretical literature on psychological health. Results indicated that neither the HFD nor the HTP quantitative composite ratings of psychological health related to the formal measures of self-esteem. PMID:9467766

  6. Egg morphology and chorionic ultrastructure of key stored product insect pests of the United States

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Eggs of Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) and Plodia interpunctella (Hübner) (Lepidoptera: Pyralidae) were imaged with scanning electron microscopy to explore how respiratory openings on the chorion surface may be related to the efficacy of fumigants. Each P. interpunctella eg...

  7. Quantitation of Murine Stroma and Selective Purification of the Human Tumor Component of Patient-Derived Xenografts for Genomic Analysis.

    PubMed

    Schneeberger, Valentina E; Allaj, Viola; Gardner, Eric E; Poirier, J T; Rudin, Charles M

    2016-01-01

    Patient-derived xenograft (PDX) mouse models are increasingly used for preclinical therapeutic testing of human cancer. A limitation in molecular and genetic characterization of PDX tumors is the presence of integral murine stroma. This is particularly problematic for genomic sequencing of PDX models. Rapid and dependable approaches for quantitating stromal content and purifying the malignant human component of these tumors are needed. We used a recently developed technique exploiting species-specific polymerase chain reaction (PCR) amplicon length (ssPAL) differences to define the fractional composition of murine and human DNA, which was proportional to the fractional composition of cells in a series of lung cancer PDX lines. We compared four methods of human cancer cell isolation: fluorescence-activated cell sorting (FACS), an immunomagnetic mouse cell depletion (MCD) approach, and two distinct EpCAM-based immunomagnetic positive selection methods. We further analyzed DNA extracted from the resulting enriched human cancer cells by targeted sequencing using a clinically validated multi-gene panel. Stromal content varied widely among tumors of similar histology, but appeared stable over multiple serial tumor passages of an individual model. FACS and MCD were superior to either positive selection approach, especially in cases of high stromal content, and consistently allowed high quality human-specific genomic profiling. ssPAL is a dependable approach to quantitation of murine stromal content, and MCD is a simple, efficient, and high yield approach to human cancer cell isolation for genomic analysis of PDX tumors. PMID:27611664

  8. Initial Description of a Quantitative, Cross-Species (Chimpanzee-Human) Social Responsiveness Measure

    ERIC Educational Resources Information Center

    Marrus, Natasha; Faughn, Carley; Shuman, Jeremy; Petersen, Steve E.; Constantino, John N.; Povinelli, Daniel J.; Pruett, John R., Jr.

    2011-01-01

    Objective: Comparative studies of social responsiveness, an ability that is impaired in autism spectrum disorders, can inform our understanding of both autism and the cognitive architecture of social behavior. Because there is no existing quantitative measure of social responsiveness in chimpanzees, we generated a quantitative, cross-species…

  9. A Humanized Clinically Calibrated Quantitative Systems Pharmacology Model for Hypokinetic Motor Symptoms in Parkinson's Disease.

    PubMed

    Roberts, Patrick; Spiros, Athan; Geerts, Hugo

    2016-01-01

    The current treatment of Parkinson's disease with dopamine-centric approaches such as L-DOPA and dopamine agonists, although very successful, is in need of alternative treatment strategies, both in terms of disease modification and symptom management. Various non-dopaminergic treatment approaches did not result in a clear clinical benefit, despite showing a clear effect in preclinical animal models. In addition, polypharmacy is common, sometimes leading to unintended effects on non-motor cognitive and psychiatric symptoms. To explore novel targets for symptomatic treatment and possible synergistic pharmacodynamic effects between different drugs, we developed a computer-based Quantitative Systems Pharmacology (QSP) platform of the closed cortico-striatal-thalamic-cortical basal ganglia loop of the dorsal motor circuit. This mechanism-based simulation platform is based on the known neuro-anatomy and neurophysiology of the basal ganglia and explicitly incorporates domain expertise in a formalized way. The calculated beta/gamma power ratio of the local field potential in the subthalamic nucleus correlates well (R (2) = 0.71) with clinically observed extra-pyramidal symptoms triggered by antipsychotics during schizophrenia treatment (43 drug-dose combinations). When incorporating Parkinsonian (PD) pathology and reported compensatory changes, the computer model suggests a major increase in b/g ratio (corresponding to bradykinesia and rigidity) from a dopamine depletion of 70% onward. The correlation between the outcome of the QSP model and the reported changes in UPDRS III Motor Part for 22 placebo-normalized drug-dose combinations is R (2) = 0.84. The model also correctly recapitulates the lack of clinical benefit for perampanel, MK-0567 and flupirtine and offers a hypothesis for the translational disconnect. Finally, using human PET imaging studies with placebo response, the computer model predicts well the placebo response for chronic treatment, but not for acute

  10. Quantitative evaluation of simulated human enamel caries kinetics using photothermal radiometry and modulated luminescence

    NASA Astrophysics Data System (ADS)

    Hellen, Adam; Mandelis, Andreas; Finer, Yoav; Amaechi, Bennett T.

    2011-03-01

    Photothermal radiometry and modulated luminescence (PTR-LUM) is a non-destructive methodology applied toward the detection, monitoring and quantification of dental caries. The purpose of this study was to evaluate the efficacy of PTRLUM to detect incipient caries lesions and quantify opto-thermophysical properties as a function of treatment time. Extracted human molars (n=15) were exposed to an acid demineralization gel (pH 4.5) for 10 or 40 days in order to simulate incipient caries lesions. PTR-LUM frequency scans (1 Hz - 1 kHz) were performed prior to and during demineralization. Transverse Micro-Radiography (TMR) analysis followed at treatment conclusion. A coupled diffusephoton- density-wave and thermal-wave theoretical model was applied to PTR experimental amplitude and phase data across the frequency range of 4 Hz - 354 Hz, to quantitatively evaluate changes in thermal and optical properties of sound and demineralized enamel. Excellent fits with small residuals were observed experimental and theoretical data illustrating the robustness of the computational algorithm. Increased scattering coefficients and poorer thermophysical properties were characteristic of demineralized lesion bodies. Enhanced optical scattering coefficients of demineralized lesions resulted in poorer luminescence yield due to scattering of both incident and converted luminescent photons. Differences in the rate of lesion progression for the 10-day and 40-day samples points to a continuum of surface and diffusion controlled mechanism of lesion formation. PTR-LUM sensitivity to changes in tooth mineralization coupled with opto-thermophysical property extraction illustrates the technique's potential for non-destructive quantification of enamel caries.

  11. Pharmacology-based toxicity assessment: towards quantitative risk prediction in humans.

    PubMed

    Sahota, Tarjinder; Danhof, Meindert; Della Pasqua, Oscar

    2016-05-01

    Despite ongoing efforts to better understand the mechanisms underlying safety and toxicity, ~30% of the attrition in drug discovery and development is still due to safety concerns. Changes in current practice regarding the assessment of safety and toxicity are required to reduce late stage attrition and enable effective development of novel medicines. This review focuses on the implications of empirical evidence generation for the evaluation of safety and toxicity during drug development. A shift in paradigm is needed to (i) ensure that pharmacological concepts are incorporated into the evaluation of safety and toxicity; (ii) facilitate the integration of historical evidence and thereby the translation of findings across species as well as between in vitro and in vivo experiments and (iii) promote the use of experimental protocols tailored to address specific safety and toxicity questions. Based on historical examples, we highlight the challenges for the early characterisation of the safety profile of a new molecule and discuss how model-based methodologies can be applied for the design and analysis of experimental protocols. Issues relative to the scientific rationale are categorised and presented as a hierarchical tree describing the decision-making process. Focus is given to four different areas, namely, optimisation, translation, analytical construct and decision criteria. From a methodological perspective, the relevance of quantitative methods for estimation and extrapolation of risk from toxicology and safety pharmacology experimental protocols, such as points of departure and potency, is discussed in light of advancements in population and Bayesian modelling techniques (e.g. non-linear mixed effects modelling). Their use in the evaluation of pharmacokinetics (PK) and pharmacokinetic-pharmacodynamic relationships (PKPD) has enabled great insight into the dose rationale for medicines in humans, both in terms of efficacy and adverse events. Comparable benefits

  12. Evaluation of Iron Content in Human Cerebral Cavernous Malformation using Quantitative Susceptibility Mapping

    PubMed Central

    Tan, Huan; Liu, Tian; Wu, Ying; Thacker, Jon; Shenkar, Robert; Mikati, Abdul Ghani; Shi, Changbin; Dykstra, Conner; Wang, Yi; Prasad, Pottumarthi V.; Edelman, Robert R.; Awad, Issam A.

    2014-01-01

    Objectives To investigate and validate quantitative susceptibility mapping (QSM) for lesional iron quantification in cerebral cavernous malformations (CCM). Materials and Methods Magnetic resonance imaging (MRI) studies were performed in phantoms and 16 patients on a 3T scanner. QSM, susceptibility weighted imaging (SWI), and R2* maps were reconstructed from in vivo data acquired with a three-dimensional, multi-echo, and T2*-weighted gradient echo sequence. Magnetic susceptibility measurements were correlated to SWI and R2* results. In addition, iron concentrations from surgically excised CCM lesion specimens were determined using inductively coupled plasma mass spectrometry and correlated with QSM measurements. Results The QSM images demonstrated excellent image quality for depicting CCM lesions in both sporadic and familial cases. Susceptibility measurements revealed a positive linear correlation with R2* values (R2 = 0.99 for total, R2 = 0.69 for mean; p < 0.01). QSM values of known iron-rich brain regions matched closely with previous studies and in interobserver consistency. A strong correlation was found between QSM and the concentration of iron phantoms (0.925, p < 0.01), as well as between QSM and mass spectroscopy estimation of iron deposition (0.999 for total iron, 0.86 for iron concentration; p < 0.01) in 18 fragments of 4 excised human CCM lesion specimens. Conclusions The ability of QSM to evaluate iron deposition in CCM lesions was illustrated via phantom, in vivo and ex vivo validation studies. QSM may be a potential biomarker for monitoring CCM disease activity and response to treatments. PMID:24619210

  13. A quantitative methodology to assess the risks to human health from CO 2 leakage into groundwater

    NASA Astrophysics Data System (ADS)

    Siirila, Erica R.; Navarre-Sitchler, Alexis K.; Maxwell, Reed M.; McCray, John E.

    2012-02-01

    Leakage of CO 2 and associated gases into overlying aquifers as a result of geologic carbon capture and sequestration may have adverse impacts on aquifer drinking-water quality. Gas or aqueous-phase leakage may occur due to transport via faults and fractures, through faulty well bores, or through leaky confining materials. Contaminants of concern include aqueous salts and dissolved solids, gaseous or aqueous-phase organic contaminants, and acidic gas or aqueous-phase fluids that can liberate metals from aquifer minerals. Here we present a quantitative risk assessment framework to predict potential human health risk from CO 2 leakage into drinking water aquifers. This framework incorporates the potential release of CO 2 into the drinking water aquifer; mobilization of metals due to a decrease in pH; transport of these metals down gradient to municipal receptors; distributions of contaminated groundwater to multiple households; and exposure and health risk to individuals using this water for household purposes. Additionally, this framework is stochastic, incorporates detailed variations in geological and geostatistical parameters and discriminates between uncertain and variable parameters using a two-stage, or nested, Monte Carlo approach. This approach is demonstrated using example simulations with hypothetical, yet realistic, aquifer characteristics and leakage scenarios. These example simulations show a greater risk for arsenic than for lead for both cancer and non-cancer endpoints, an unexpected finding. Higher background groundwater gradients also yield higher risk. The overall risk and the associated uncertainty are sensitive to the extent of aquifer stratification and the degree of local-scale dispersion. These results all highlight the importance of hydrologic modeling in risk assessment. A linear relationship between carcinogenic and noncarcinogenic risk was found for arsenic and suggests action levels for carcinogenic risk will be exceeded in exposure

  14. Quantitative Autofluorescence and Cell Density Maps of the Human Retinal Pigment Epithelium

    PubMed Central

    Ach, Thomas; Huisingh, Carrie; McGwin, Gerald; Messinger, Jeffrey D.; Zhang, Tianjiao; Bentley, Mark J.; Gutierrez, Danielle B.; Ablonczy, Zsolt; Smith, R. Theodore; Sloan, Kenneth R.; Curcio, Christine A.

    2014-01-01

    Purpose. Lipofuscin (LF) accumulation within RPE cells is considered pathogenic in AMD. To test whether LF contributes to RPE cell loss in aging and to provide a cellular basis for fundus autofluorescence (AF) we created maps of human RPE cell number and histologic AF. Methods. Retinal pigment epithelium–Bruch's membrane flat mounts were prepared from 20 donor eyes (10 ≤ 51 and 10 > 80 years; postmortem: ≤4.2 hours; no retinal pathologies), preserving foveal position. Phalloidin-binding RPE cytoskeleton and LF-AF (488-nm excitation) were imaged at up to 90 predefined positions. Maps were assembled from 83,330 cells in 1470 locations. From Voronoi regions representing each cell, the number of neighbors, cell area, and total AF intensity normalized to an AF standard was determined. Results. Highly variable between individuals, RPE-AF increases significantly with age. A perifoveal ring of high AF mirrors rod photoreceptor topography and fundus-AF. Retinal pigment epithelium cell density peaks at the fovea, independent of age, yet no net RPE cell loss is detectable. The RPE monolayer undergoes considerable lifelong re-modeling. The relationship of cell size and AF, a surrogate for LF concentration, is orderly and linear in both groups. Autofluorescence topography differs distinctly from the topography of age-related rod loss. Conclusions. Digital maps of quantitative AF, cell density, and packing geometry provide metrics for cellular-resolution clinical imaging and model systems. The uncoupling of RPE LF content, cell number, and photoreceptor topography in aging challenges LF's role in AMD. PMID:25034602

  15. Quantitative determination of percutaneous absorption of radiolabeled drugs in vitro and in vivo by human skin.

    PubMed

    Schaefer, H; Stüttgen, G; Zesch, A; Schalla, W; Gazith, J

    1978-01-01

    We have measured concentrations of about 30 drugs in the living layers of the skin under conditions which provide data which are applicable in therapeutic treatment. Since the skin is a thin organ and small amounts of drug represent high target concentrations, it is necessary to select a sensitive quantitative method; observation of the kinetics of absorption using radiolabeled drugs is the method of choice. Because of possible hazards--and legal and ethical problems--absorption studies in human skin are commonly performed in vitro. Related in vivo investigations demonstrate the relevance and the limitations of the in vitro experiments. The main hindrance against penetration of drugs is by the horny layer. The barrier-function of this layer--if it is undisturbed--may be described by a multilayer model. The reciprocal function, the reservoir function, is important for the efficiency of topical treatment; it also plays a role in determining the unique pharmacokinetics of drug absorption in the skin and percutaneous resorption. If the horny layer is injured, i.e. in diseased skin, both the barrier and the reservoir functions are disturbed. In consequence, drug concentrations in the skin--and percutaneous resorption--may be greatly enhanced, and topically applied drugs may enter preferentially into diseased areas. The form of application, such as ointment, solution, etc. influences the penetration kinetics in such a specific manner that a specific vehicle for a specific drug should always be postulated. The frequently discussed hazards of side effects due to percutaneous resorption of drugs like corticosteroids are a function of the treated area rather than of its penetration capacity. Thus the indication for local or oral treatment of severe dermatoses should be considered in terms of the affected area. The relatively frequent side effects in the skin itself which originate from unnecessarily high drug concentrations and long term treatment must also be taken into

  16. Quantitative ultrashort echo time (UTE) MRI of human cortical bone: correlation with porosity and biomechanical properties.

    PubMed

    Bae, Won C; Chen, Peter C; Chung, Christine B; Masuda, Koichi; D'Lima, Darryl; Du, Jiang

    2012-04-01

    In this study we describe the use of ultrashort echo time (UTE) magnetic resonance imaging (MRI) to evaluate short and long T2* components as well as the water content of cortical bone. Fourteen human cadaveric distal femur and proximal tibia were sectioned to produce 44 rectangular slabs of cortical bone for quantitative UTE MR imaging, microcomputed tomography (µCT), and biomechanical testing. A two-dimensional (2D) UTE pulse sequence with a minimal nominal TE of 8 µseconds was used together with bicomponent analysis to quantify the bound and free water in cortical bone using a clinical 3T scanner. Total water concentration was measured using a 3D UTE sequence together with a reference water phantom. UTE MR measures of water content (total, free, and bound), T2* (short and long), and short and long T2* fractions were compared with porosity assessed with µCT, as well as elastic (modulus, yield stress, and strain) and failure (ultimate stress, failure strain, and energy) properties, using Pearson correlation. Porosity significantly correlated positively with total (R(2)  = 0.23; p < 0.01) and free (R(2)  = 0.31; p < 0.001) water content as well as long T2* fraction (R(2)  = 0.25; p < 0.001), and negatively with short T2* fraction and short T2* (R(2)  = 0.24; p < 0.01). Failure strain significantly correlated positively with short T2* (R(2)  = 0.29; p < 0.001), ultimate stress significantly correlated negatively with total (R(2)  = 0.25; p < 0.001) and bound (R(2)  = 0.22; p < 0.01) water content, and failure energy significantly correlated positively with both short (R(2)  = 0 30; p < 0.001) and long (R(2)  = 0.17; p < 0.01) T2* values. These results suggest that UTE MR measures are sensitive to the structure and failure properties of human cortical bone, and may provide a novel way of evaluating cortical bone quality. PMID:22190232

  17. Quantitative immunological determination of 12 plasma proteins excreted in human urine collected before and after exercise

    PubMed Central

    Poortmans, Jacques; Jeanloz, Roger W.

    1968-01-01

    Urine was collected from 6 healthy male adults at rest and from 20 male adults after a marathon race (25 miles). The concentrated urines were quantitatively analyzed, by single radial immunodiffusion, for their content in 12 different plasma proteins: tryptophan-rich prealbumin, albumin, α1-acid glycoprotein, α1-antitrypsin, ceruloplasmin, haptoglobin, Gc-globulin, transferrin, hemopexin, β2-glycoprotein I, γA-globulin, and γG-globulin. Albumin, γA-globulin, and γG-globulin represent the major part of the plasma proteins detected in normal urine excreted by humans at rest (12, 0.5, and 2.5 mg respectively, out of a total excretion of 17.5 mg of plasma proteins per 24 hr). The other plasma proteins were excreted at a lower rate (< 0.4 mg/24 hr). The relative content of tryptophan-rich prealbumin, α1-antitrypsin, Gc-globulin, transferrin, and γG-globulin was lower in normal urine than in normal serum, whereas that of α1-acid glycoprotein, β2-glycoprotein I, and γA-globulin was higher. The ratio of γG-globulin to γA-globulin was 4.9:1. When plotted on a logarithmic scale, no direct relationship between the molecular weight of a protein and the value of its renal clearance could be observed. Strenuous exercise increased (up to 50-fold) the excretion of plasma proteins which represent 82% of the total proteins found in urine, instead of 57% in urine collected from humans at rest. There was particularly a significant rise of tryptophan-rich albumin, albumin, α1-acid glycoprotein, transferrin, γA-globulin, and γG-globulin (0.26, 127, 11.8, 3.3, 1.2, and 2.0 μg respectively, out of a total excretion of 167 μg of plasma proteins per min). The ratio of γG-globulin to γA-globulin was 16:1. After exercise, the renal clearance of proteins increased from 2 to 40 times, but, as for the urine of normal subjects at rest, no direct relationship between molecular weight and renal clearance could be observed. Images PMID:4170390

  18. Zebrafish (Danio rerio) as a possible bioindicator of epigenetic factors present in drinking water that may affect reproductive function: is chorion an issue?

    PubMed

    Martinez-Sales, M; García-Ximénez, F; Espinós, F J

    2015-06-01

    Emerging organic contaminants have been monitored in stream waters, raw and finished waters and wastewater effluents. Most of these contaminants, such as epigenetic substances, have been detected at very low levels. Unfortunately, their complete monitoring and/or removal are very difficult, given the increasing presence of new contaminants and due to analytical and economic considerations. For this reason, bioindicators are used as an alternative to monitor their presence. To this end, zebrafish is being used to assess certain contaminants in water quality studies. As our long-term aim is to determine if zebrafish (Danio rerio) can be used to detect environmental epigenetic factors in drinking waters with effects on human reproduction, an initial question is whether the chorion could interfere with the possible action of epigenetic factors in two reproductive events: genital ridge formation and migration of the primordial germ cells (PGCs) to these genital ridges. In the first experiment, we attempted to partially degrade the chorion of mid blastula transition (MBT) embryos with pronase, with acceptable survival rates at 5 days post fertilisation (dpf), with the group exposed for 15 min giving the best survival results. As denuded early embryos require a specific culture medium, in the next experiment embryo survival was evaluated when they were cultured up to 5 dpf in drinking waters from six different sources. Results showed a negative effect on embryo survival at 5 dpf from several waters but not in others, thus distorting the survival outcomes. These results suggest using embryos with the chorion intact from the outset when drinking waters from different sources are to be tested. PMID:24598355

  19. Brief Communication: Quantitative- and molecular-genetic differentiation in humans and chimpanzees: implications for the evolutionary processes underlying cranial diversification.

    PubMed

    Weaver, Timothy D

    2014-08-01

    Estimates of the amount of genetic differentiation in humans among major geographic regions (e.g., Eastern Asia vs. Europe) from quantitative-genetic analyses of cranial measurements closely match those from classical- and molecular-genetic markers. Typically, among-region differences account for ∼10% of the total variation. This correspondence is generally interpreted as evidence for the importance of neutral evolutionary processes (e.g., genetic drift) in generating among-region differences in human cranial form, but it was initially surprising because human cranial diversity was frequently assumed to show a strong signature of natural selection. Is the human degree of similarity of cranial and DNA-sequence estimates of among-region genetic differentiation unusual? How do comparisons with other taxa illuminate the evolutionary processes underlying cranial diversification? Chimpanzees provide a useful starting point for placing the human results in a broader comparative context, because common chimpanzees (Pan troglodytes) and bonobos (Pan paniscus) are the extant species most closely related to humans. To address these questions, I used 27 cranial measurements collected on a sample of 861 humans and 263 chimpanzees to estimate the amount of genetic differentiation between pairs of groups (between regions for humans and between species or subspecies for chimpanzees). Consistent with previous results, the human cranial estimates are quite similar to published DNA-sequence estimates. In contrast, the chimpanzee cranial estimates are much smaller than published DNA-sequence estimates. It appears that cranial differentiation has been limited in chimpanzees relative to humans. PMID:24827671

  20. Classification of histologically scored human knee osteochondral plugs by quantitative analysis of magnetic resonance images at 3T.

    PubMed

    Lukas, Vanessa A; Fishbein, Kenneth W; Lin, Ping-Chang; Schär, Michael; Schneider, Erika; Neu, Corey P; Spencer, Richard G; Reiter, David A

    2015-05-01

    This work evaluates the ability of quantitative MRI to discriminate between normal and pathological human osteochondral plugs characterized by the Osteoarthritis Research Society International (OARSI) histological system. Normal and osteoarthritic human osteochondral plugs were scored using the OARSI histological system and imaged at 3 T using MRI sequences producing T1 and T2 contrast and measuring T1, T2, and T2* relaxation times, magnetization transfer, and diffusion. The classification accuracies of quantitative MRI parameters and corresponding weighted image intensities were evaluated. Classification models based on the Mahalanobis distance metric for each MRI measurement were trained and validated using leave-one-out cross-validation with plugs grouped according to OARSI histological grade and score. MRI measurements used for classification were performed using a region-of-interest analysis which included superficial, deep, and full-thickness cartilage. The best classifiers based on OARSI grade and score were T1- and T2-weighted image intensities, which yielded accuracies of 0.68 and 0.75, respectively. Classification accuracies using OARSI score-based group membership were generally higher when compared with grade-based group membership. MRI-based classification--either using quantitative MRI parameters or weighted image intensities--is able to detect early osteoarthritic tissue changes as classified by the OARSI histological system. These findings suggest the benefit of incorporating quantitative MRI acquisitions in a comprehensive clinical evaluation of OA. PMID:25641500

  1. Quantitative determination of terbutaline and orciprenaline in human plasma by gas chromatography/negative ion chemical ionization/mass spectrometry.

    PubMed

    Leis, H J; Gleispach, H; Nitsche, V; Malle, E

    1990-06-01

    A method for the determination of unconjugated terbutaline and orciprenaline in human plasma is described. The assay is based on stable isotope dilution gas chromatography/negative ion chemical ionization/mass spectrometry. An inexpensive and rapid method for preparation of stable isotope labelled analogues as well as their use in quantitative gas chromatography/mass spectrometry is shown. A highly efficient sample work-up procedure with product recoveries of more than 95% is presented. The method developed permits quantitative measurement of terbutaline and orciprenaline in human plasma down to 100 pg ml-1, using 1 ml of sample. Plasma levels of terbutaline after oral administration of 5 mg of terbutaline sulphate were estimated. PMID:2357489

  2. TeratoScore: Assessing the Differentiation Potential of Human Pluripotent Stem Cells by Quantitative Expression Analysis of Teratomas.

    PubMed

    Avior, Yishai; Biancotti, Juan Carlos; Benvenisty, Nissim

    2015-06-01

    Teratoma formation is the gold standard assay for testing the capacity of human pluripotent stem cells to differentiate into all embryonic germ layers. Although widely used, little effort has been made to transform this qualitative assay into a quantitative one. Using gene expression data from a wide variety of cells, we created a scorecard representing tissues from all germ layers and extraembryonic tissues. TeratoScore, an online, open-source platform based on this scorecard, distinguishes pluripotent stem cell-derived teratomas from malignant tumors, translating cell potency into a quantitative measure (http://benvenisty.huji.ac.il/teratoscore.php). The teratomas used for the algorithm also allowed us to examine gene expression differences between tumors with a diploid karyotype and those initiated by aneuploid cells. Chromosomally aberrant teratomas show a significantly different gene expression signature from that of teratomas originating from diploid cells, particularly in central nervous system-specific genes, congruent with human chromosomal syndromes. PMID:26070610

  3. Quantitative and qualitative effects of N10-methylfolate on high-affinity folate binding in human leukocytes.

    PubMed

    Holm, J; Hansen, S I; Lyngbye, J

    1984-01-01

    N10-methylfolate acted as a potent competitive inhibitor of high-affinity [3H] folate binding in human leukocytes, while methotrexate had no effect. Furthermore, folate binding changed into a non-cooperative type in the presence of N10-methylfolate. Hence, in qualitative and quantitative respects, the substrate specificity characteristics of leukocyte folate binding resemble those of other high-affinity folate binding systems. PMID:6500843

  4. Rapid quantitative method for the detection of phenylalanine and tyrosine in human plasma using pillar array columns and gradient elution.

    PubMed

    Song, Yanting; Takatsuki, Katsuya; Sekiguchi, Tetsushi; Funatsu, Takashi; Shoji, Shuichi; Tsunoda, Makoto

    2016-07-01

    This study reports a fast and quantitative determination method for phenylalanine (Phe) and tyrosine (Tyr) in human plasma using on-chip pressure-driven liquid chromatography. A pillar array column with low-dispersion turns and a gradient elution system was used. The separation of fluorescent derivatives of Phe, Tyr, and other hydrophobic amino acids was successfully performed within 140 s. Under the optimized conditions, Phe and Tyr in human plasma were quantified. The developed method is promising for rapid diagnosis in the clinical field. PMID:27209196

  5. Quantitation of 4,4′-methylene diphenyl diisocyanate human serum albumin adducts

    PubMed Central

    Luna, Leah G.; Green, Brett J.; Zhang, Fagen; Arnold, Scott M.; Siegel, Paul D.; Bartels, Michael J.

    2016-01-01

    4,4′-Methylene diphenyl diisocyanate (herein 4,4′-MDI) is used in the production of polyurethane foams, elastomers, coatings, adhesives and the like for a wide range of commercial products. Occupational exposure to MDI levels above current airborne exposure limits can elicit immune mediated hypersensitivity reactions such as occupational asthma in sensitive individuals. To accurately determine exposure, there has been increasing interest in developing analytical methods to measure internal biomarkers of exposure to MDI. Previous investigators have reported methodologies for measuring MDI diamine metabolites and MDI-Lysine (4,4′-MDI-Lys) adducts. The purpose of this study was to develop and validate an ultra performance liquid chromatography isotope dilution tandem mass spectrometry (UPLC-ID/MS/MS) quantitation method via a signature peptide approach to enable biomonitoring of 4,4′-MDI adducted to human serum albumin (HSA) in plasma. A murine, anti-4,4′-MDI monoclonal IgM antibody was bound to magnetic beads and utilized for enrichment of the MDI adducted HSA. Following enrichment, trypsin digestion was performed to generate the expected 414 site (primary site of adduction) 4,4′-MDI-adducted HSA signature peptide that was quantified by UPLC-ID/MS/MS. An Agilent 6530 UPLC/quadrupole time of flight MS (QTOF) system was utilized for intact adducted protein analysis and an Agilent 6490 UPLC/MS/MS system operated in multiple reaction monitoring (MRM) mode was utilized for quantification of the adducted signature peptide biomarker both for in chemico and worker serum samples. Worker serum samples were initially screened utilizing the previously developed 4,4′-MDI-Lys amino acid method and results showed that 12 samples were identified as quantifiable for 4,4′-MDI-Lys adducts. The signature peptide adduct approach was applied to the 12 worker samples identified as quantifiable for 4,4′-MDI-Lys adducts. Results indicated no positive results were obtained

  6. A quantitative exposure model simulating human norovirus transmission during preparation of deli sandwiches.

    PubMed

    Stals, Ambroos; Jacxsens, Liesbeth; Baert, Leen; Van Coillie, Els; Uyttendaele, Mieke

    2015-03-01

    Human noroviruses (HuNoVs) are a major cause of food borne gastroenteritis worldwide. They are often transmitted via infected and shedding food handlers manipulating foods such as deli sandwiches. The presented study aimed to simulate HuNoV transmission during the preparation of deli sandwiches in a sandwich bar. A quantitative exposure model was developed by combining the GoldSim® and @Risk® software packages. Input data were collected from scientific literature and from a two week observational study performed at two sandwich bars. The model included three food handlers working during a three hour shift on a shared working surface where deli sandwiches are prepared. The model consisted of three components. The first component simulated the preparation of the deli sandwiches and contained the HuNoV reservoirs, locations within the model allowing the accumulation of NoV and the working of intervention measures. The second component covered the contamination sources being (1) the initial HuNoV contaminated lettuce used on the sandwiches and (2) HuNoV originating from a shedding food handler. The third component included four possible intervention measures to reduce HuNoV transmission: hand and surface disinfection during preparation of the sandwiches, hand gloving and hand washing after a restroom visit. A single HuNoV shedding food handler could cause mean levels of 43±18, 81±37 and 18±7 HuNoV particles present on the deli sandwiches, hands and working surfaces, respectively. Introduction of contaminated lettuce as the only source of HuNoV resulted in the presence of 6.4±0.8 and 4.3±0.4 HuNoV on the food and hand reservoirs. The inclusion of hand and surface disinfection and hand gloving as a single intervention measure was not effective in the model as only marginal reductions of HuNoV levels were noticeable in the different reservoirs. High compliance of hand washing after a restroom visit did reduce HuNoV presence substantially on all reservoirs. The

  7. Protein profiling of human lung telocytes and microvascular endothelial cells using iTRAQ quantitative proteomics

    PubMed Central

    Zheng, Yonghua; Cretoiu, Dragos; Yan, Guoquan; Cretoiu, Sanda Maria; Popescu, Laurentiu M; Fang, Hao; Wang, Xiangdong

    2014-01-01

    Telocytes (TCs) are described as a particular type of cells of the interstitial space (www.telocytes.com). Their main characteristics are the very long telopodes with alternating podoms and podomers. Recently, we performed a comparative proteomic analysis of human lung TCs with fibroblasts, demonstrating that TCs are clearly a distinct cell type. Therefore, the present study aims to reinforce this idea by comparing lung TCs with endothelial cells (ECs), since TCs and ECs share immunopositivity for CD34. We applied isobaric tag for relative and absolute quantification (iTRAQ) combined with automated 2-D nano-ESI LC-MS/MS to analyse proteins extracted from TCs and ECs in primary cell cultures. In total, 1609 proteins were identified in cell cultures. 98 proteins (the 5th day), and 82 proteins (10th day) were confidently quantified (screened by two-sample t-test, P < 0.05) as up- or down-regulated (fold change >2). We found that in TCs there are 38 up-regulated proteins at the 5th day and 26 up-regulated proteins at the 10th day. Bioinformatics analysis using Panther revealed that the 38 proteins associated with TCs represented cellular functions such as intercellular communication (via vesicle mediated transport) and structure morphogenesis, being mainly cytoskeletal proteins and oxidoreductases. In addition, we found 60 up-regulated proteins in ECs e.g.: cell surface glycoprotein MUC18 (15.54-fold) and von Willebrand factor (5.74-fold). The 26 up-regulated proteins in TCs at 10th day, were also analysed and confirmed the same major cellular functions, while the 56 down-regulated proteins confirmed again their specificity for ECs. In conclusion, we report here the first extensive comparison of proteins from TCs and ECs using a quantitative proteomics approach. Our data show that TCs are completely different from ECs. Protein expression profile showed that TCs play specific roles in intercellular communication and intercellular signalling. Moreover, they might

  8. A quantitative methodology to assess the risks to human health from CO2 leakage into groundwater

    NASA Astrophysics Data System (ADS)

    Siirila, E.; Sitchler, A.; Maxwell, R. M.; McCray, J. E.

    2010-12-01

    Leakage of CO2 and associated gases into overlying aquifers as a result of geologic carbon capture and sequestration may have adverse impacts on aquifer drinking-water quality. Gas or aqueous-phase leakage may occur due to transport via faults and fractures, through faulty well bores, or through leaky confining materials. Contaminants of concern include aqueous salts and dissolved solids, gaseous or aqueous-phase organic contaminants, and acidic gas or aqueous-phase fluids that can liberate metals from aquifer minerals. Here we present a quantitative risk assessment framework to predict potential human health risk from CO2 leakage into drinking water aquifers. This framework incorporates the potential release of CO2 into the drinking water aquifer; mobilization of metals due to a decrease in pH; transport of these metals down gradient to municipal receptors; distributions of contaminated groundwater to multiple households; and exposure and health risk to individuals using this water for household purposes. Additionally, this framework is stochastic, incorporates detailed variations in geological and geostatistical parameters and discriminates between uncertain and variable parameters using a two-stage, or nested, Monte Carlo approach. This approach is demonstrated using example simulations with hypothetical, yet realistic, aquifer characteristics and leakage scenarios. These example simulations show a greater risk for arsenic than for lead for both cancer and non-cancer endpoints, an unexpected finding given greater toxicity of lead at lower doses than arsenic. It was also found that higher background groundwater gradients also yield higher risk. The overall risk and the associated uncertainty are sensitive to the extent of aquifer stratification and the degree of local-scale dispersion. These results all highlight the importance of hydrologic modeling in risk assessment. A linear relationship between carcinogenic and noncarcinogenic risk was found for arsenic and

  9. Quantitative high-throughput gene expression profiling of human striatal development to screen stem cell–derived medium spiny neurons

    PubMed Central

    Straccia, Marco; Garcia-Diaz Barriga, Gerardo; Sanders, Phil; Bombau, Georgina; Carrere, Jordi; Mairal, Pedro Belio; Vinh, Ngoc-Nga; Yung, Sun; Kelly, Claire M; Svendsen, Clive N; Kemp, Paul J; Arjomand, Jamshid; Schoenfeld, Ryan C; Alberch, Jordi; Allen, Nicholas D; Rosser, Anne E; Canals, Josep M

    2015-01-01

    A systematic characterization of the spatio-temporal gene expression during human neurodevelopment is essential to understand brain function in both physiological and pathological conditions. In recent years, stem cell technology has provided an in vitro tool to recapitulate human development, permitting also the generation of human models for many diseases. The correct differentiation of human pluripotent stem cell (hPSC) into specific cell types should be evaluated by comparison with specific cells/tissue profiles from the equivalent adult in vivo organ. Here, we define by a quantitative high-throughput gene expression analysis the subset of specific genes of the whole ganglionic eminence (WGE) and adult human striatum. Our results demonstrate that not only the number of specific genes is crucial but also their relative expression levels between brain areas. We next used these gene profiles to characterize the differentiation of hPSCs. Our findings demonstrate a temporal progression of gene expression during striatal differentiation of hPSCs from a WGE toward an adult striatum identity. Present results establish a gene expression profile to qualitatively and quantitatively evaluate the telencephalic hPSC-derived progenitors eventually used for transplantation and mature striatal neurons for disease modeling and drug-screening. PMID:26417608

  10. Quantitative high-throughput gene expression profiling of human striatal development to screen stem cell-derived medium spiny neurons.

    PubMed

    Straccia, Marco; Garcia-Diaz Barriga, Gerardo; Sanders, Phil; Bombau, Georgina; Carrere, Jordi; Mairal, Pedro Belio; Vinh, Ngoc-Nga; Yung, Sun; Kelly, Claire M; Svendsen, Clive N; Kemp, Paul J; Arjomand, Jamshid; Schoenfeld, Ryan C; Alberch, Jordi; Allen, Nicholas D; Rosser, Anne E; Canals, Josep M

    2015-01-01

    A systematic characterization of the spatio-temporal gene expression during human neurodevelopment is essential to understand brain function in both physiological and pathological conditions. In recent years, stem cell technology has provided an in vitro tool to recapitulate human development, permitting also the generation of human models for many diseases. The correct differentiation of human pluripotent stem cell (hPSC) into specific cell types should be evaluated by comparison with specific cells/tissue profiles from the equivalent adult in vivo organ. Here, we define by a quantitative high-throughput gene expression analysis the subset of specific genes of the whole ganglionic eminence (WGE) and adult human striatum. Our results demonstrate that not only the number of specific genes is crucial but also their relative expression levels between brain areas. We next used these gene profiles to characterize the differentiation of hPSCs. Our findings demonstrate a temporal progression of gene expression during striatal differentiation of hPSCs from a WGE toward an adult striatum identity. Present results establish a gene expression profile to qualitatively and quantitatively evaluate the telencephalic hPSC-derived progenitors eventually used for transplantation and mature striatal neurons for disease modeling and drug-screening. PMID:26417608

  11. Setting health research priorities using the CHNRI method: V. Quantitative properties of human collective knowledge

    PubMed Central

    Rudan, Igor; Yoshida, Sachiyo; Wazny, Kerri; Chan, Kit Yee; Cousens, Simon

    2016-01-01

    Introduction The CHNRI method for setting health research priorities has crowdsourcing as the major component. It uses the collective opinion of a group of experts to generate, assess and prioritize between many competing health research ideas. It is difficult to compare the accuracy of human individual and collective opinions in predicting uncertain future outcomes before the outcomes are known. However, this limitation does not apply to existing knowledge, which is an important component underlying opinion. In this paper, we report several experiments to explore the quantitative properties of human collective knowledge and discuss their relevance to the CHNRI method. Methods We conducted a series of experiments in groups of about 160 (range: 122–175) undergraduate Year 2 medical students to compare their collective knowledge to their individual knowledge. We asked them to answer 10 questions on each of the following: (i) an area in which they have a degree of expertise (undergraduate Year 1 medical curriculum); (ii) an area in which they likely have some knowledge (general knowledge); and (iii) an area in which they are not expected to have any knowledge (astronomy). We also presented them with 20 pairs of well–known celebrities and asked them to identify the older person of the pair. In all these experiments our goal was to examine how the collective answer compares to the distribution of students’ individual answers. Results When answering the questions in their own area of expertise, the collective answer (the median) was in the top 20.83% of the most accurate individual responses; in general knowledge, it was in the top 11.93%; and in an area with no expertise, the group answer was in the top 7.02%. However, the collective answer based on mean values fared much worse, ranging from top 75.60% to top 95.91%. Also, when confronted with guessing the older of the two celebrities, the collective response was correct in 18/20 cases (90%), while the 8 most

  12. Human pituitary and placental hormones control human insulin-like growth factor II secretion in human granulosa cells

    SciTech Connect

    Ramasharma, K.; Li, C.H.

    1987-05-01

    Human granulosa cells cultured with calf serum actively proliferated for 18-20 generations and secreted progesterone into the medium; progesterone levels appeared to decline with increase in generation number. Cells cultured under serum-free conditions secreted significant amounts of progesterone and insulin-like growth factor II (IGF-II). The progesterone secretion was enhanced by the addition of human follitropin, lutropin, and chorionic gonadotropin but not by growth hormone. These cells, when challenged to varying concentrations of human growth hormone, human chorionic somatomammotropin, human prolactin, chorionic gonadotropin, follitropin, and lutropin, secreted IGF-II into the medium as measured by specific IGF-II RIA. Among these human hormones, chorionic gonadotropin, follitropin, and lutropin were most effective in inducing IGF-II secretion from these cells. When synthetic lutropin-releasing hormone and ..cap alpha..-inhibin-92 were tested, only lutropin-releasing hormone was effective in releasing IGF-II. The results described suggest that cultured human granulosa cells can proliferate and actively secrete progesterone and IGF-II into the medium. IGF-II production in human granulosa cells was influenced by a multi-hormonal complex including human growth hormone, human chorionic somatomammotropin, and prolactin.

  13. A MALDI-MS-based quantitative analytical method for endogenous estrone in human breast cancer cells.

    PubMed

    Kim, Kyoung-Jin; Kim, Hee-Jin; Park, Han-Gyu; Hwang, Cheol-Hwan; Sung, Changmin; Jang, Kyoung-Soon; Park, Sung-Hee; Kim, Byung-Gee; Lee, Yoo-Kyung; Yang, Yung-Hun; Jeong, Jae Hyun; Kim, Yun-Gon

    2016-01-01

    The level of endogenous estrone, one of the three major naturally occurring estrogens, has a significant correlation with the incidence of post-menopausal breast cancer. However, it is challenging to quantitatively monitor it owing to its low abundance. Here, we develop a robust and highly sensitive mass-assisted laser desorption/ionization mass spectrometry (MALDI-MS)-based quantitative platform to identify the absolute quantities of endogenous estrones in a variety of clinical specimens. The one-step modification of endogenous estrone provided good linearity (R(2) > 0.99) and significantly increased the sensitivity of the platform (limit of quantitation: 11 fmol). In addition, we could identify the absolute amount of endogenous estrones in cells of the breast cancer cell line MCF-7 (34 fmol/10(6) cells) by using a deuterated estrone as an internal standard. Finally, by applying the MALDI-MS-based quantitative method to endogenous estrones, we successfully monitored changes in the metabolic expression level of estrones (17.7 fmol/10(6) letrozole-treated cells) in MCF-7 cells resulting from treatment with an aromatase inhibitor. Taken together, these results suggest that this MALDI-MS-based quantitative approach may be a general method for the targeted metabolomics of ketone-containing metabolites, which can reflect clinical conditions and pathogenic mechanisms. PMID:27091422

  14. A MALDI-MS-based quantitative analytical method for endogenous estrone in human breast cancer cells

    PubMed Central

    Kim, Kyoung-Jin; Kim, Hee-Jin; Park, Han-Gyu; Hwang, Cheol-Hwan; Sung, Changmin; Jang, Kyoung-Soon; Park, Sung-Hee; Kim, Byung-Gee; Lee, Yoo-Kyung; Yang, Yung-Hun; Jeong, Jae Hyun; Kim, Yun-Gon

    2016-01-01

    The level of endogenous estrone, one of the three major naturally occurring estrogens, has a significant correlation with the incidence of post-menopausal breast cancer. However, it is challenging to quantitatively monitor it owing to its low abundance. Here, we develop a robust and highly sensitive mass-assisted laser desorption/ionization mass spectrometry (MALDI-MS)-based quantitative platform to identify the absolute quantities of endogenous estrones in a variety of clinical specimens. The one-step modification of endogenous estrone provided good linearity (R2 > 0.99) and significantly increased the sensitivity of the platform (limit of quantitation: 11 fmol). In addition, we could identify the absolute amount of endogenous estrones in cells of the breast cancer cell line MCF-7 (34 fmol/106 cells) by using a deuterated estrone as an internal standard. Finally, by applying the MALDI-MS-based quantitative method to endogenous estrones, we successfully monitored changes in the metabolic expression level of estrones (17.7 fmol/106 letrozole-treated cells) in MCF-7 cells resulting from treatment with an aromatase inhibitor. Taken together, these results suggest that this MALDI-MS-based quantitative approach may be a general method for the targeted metabolomics of ketone-containing metabolites, which can reflect clinical conditions and pathogenic mechanisms. PMID:27091422

  15. A human interactome in three quantitative dimensions organized by stoichiometries and abundances.

    PubMed

    Hein, Marco Y; Hubner, Nina C; Poser, Ina; Cox, Jürgen; Nagaraj, Nagarjuna; Toyoda, Yusuke; Gak, Igor A; Weisswange, Ina; Mansfeld, Jörg; Buchholz, Frank; Hyman, Anthony A; Mann, Matthias

    2015-10-22

    The organization of a cell emerges from the interactions in protein networks. The interactome is critically dependent on the strengths of interactions and the cellular abundances of the connected proteins, both of which span orders of magnitude. However, these aspects have not yet been analyzed globally. Here, we have generated a library of HeLa cell lines expressing 1,125 GFP-tagged proteins under near-endogenous control, which we used as input for a next-generation interaction survey. Using quantitative proteomics, we detect specific interactions, estimate interaction stoichiometries, and measure cellular abundances of interacting proteins. These three quantitative dimensions reveal that the protein network is dominated by weak, substoichiometric interactions that play a pivotal role in defining network topology. The minority of stable complexes can be identified by their unique stoichiometry signature. This study provides a rich interaction dataset connecting thousands of proteins and introduces a framework for quantitative network analysis. PMID:26496610

  16. Correlation of host immune response with quantitative recovery of Chlamydia trachomatis from the human endocervix.

    PubMed Central

    Brunham, R C; Kuo, C C; Cles, L; Holmes, K K

    1983-01-01

    We studied 95 women with uncomplicated Chlamydia trachomatis cervical infection. Quantitative isolation of C. trachomatis was performed in HeLa 229 cells, and the results were correlated with serum immunoglobulin M and immunoglobulin G antibody to the organism. We found that quantitative cultures for C. trachomatis can provide a meaningful measurement by which to evaluate the effect of the acquired immune response. In particular, secretory immunoglobulin A antibody to C. trachomatis in cervical secretion demonstrated a striking and inverse correlation with recovery of the organism from the cervix. It is suggested that this component of the immune response may regulate shedding of the organism. PMID:6840846

  17. Performance Assessment of Human and Cattle Associated Quantitative Real-time PCR Assays - slides

    EPA Science Inventory

    The presentation overview is (1) Single laboratory performance assessment of human- and cattle associated PCR assays and (2) A Field Study: Evaluation of two human fecal waste management practices in Ohio watershed.

  18. Quantitation of N,N-dimethyltryptamine and harmala alkaloids in human plasma after oral dosing with ayahuasca.

    PubMed

    Callaway, J C; Raymon, L P; Hearn, W L; McKenna, D J; Grob, C S; Brito, G S; Mash, D C

    1996-10-01

    Harmine, harmaline, tetrahydroharmine (THH), and N,N-dimethyltryptamine (DMT) were quantitated in plasma from 15 healthy male volunteers after the ingestion of ayahuasca, a beverage that has been used for religious purposes in Brazil since pre-Columbian times. A growing awareness of the interest in this ancient shamanistic practice in modern urban cultures and the widespread popular dissemination of the inebriant effects and type and sources of the plant admixtures used to prepare the beverage have provided additional impetus for this study. The three harmala alkaloids were quantitated from protein-precipitated plasma by high-performance liquid chromatography using fluorescence detection. Recovery from blank human plasma was quantitative, and the limit of quantitation (LOQ) was below 2 ng/mL of plasma for each of the harmala alkaloids. Standard concentrations ranged from 10 to 250 ng/mL for harmine and THH and from 1.0 to 25.0 ng/mL for harmaline, respectively. Linearity was observed for harmine, harmaline, and THH within these respective ranges. The highest concentrations of harmala alkaloids in human plasma were found to be 222.3 ng/mL for harmine, 134.5 ng/mL for THH, and 9.4 ng/mL for harmaline. DMT was quantitated by gas chromatography using nitrogen-phosphorus detection after liquid-liquid extraction with diphenhydramine as an internal standard. DMT recovery was quantitative, and the limit of detection and LOQ were 0.5 and 5 ng/mL, respectively. Linearity for DMT was observed from 5 to 1000 ng/mL. The one-step extraction method for DMT and the protein precipitation method for the three harmala alkaloids afford rapid, sensitive, and quantitative analyses of these alkaloids with minimal analyte loss. The analytical methods also may be applicable to other matrices, including whole blood and urine samples and homogenized tissue specimens. These are the first reported observations of DMT and harmala alkaloids in plasma after ritual ingestion of ayahuasca. PMID

  19. The State of Humanities in Post-Apartheid South Africa--A Quantitative Story

    ERIC Educational Resources Information Center

    Pillay, V.; Yu, K.

    2010-01-01

    This article depicts the state of Humanities in post-apartheid South Africa by examining HEMIS enrolment and graduation data from 1999 to 2007. It demonstrates that although the decline in student enrolment and graduation in Humanities has not been severe; read in the context of substantial growth of all other disciplines, Humanities is in a…

  20. A genome-wide linkage scan for quantitative trait loci influencing the craniofacial complex in humans(Homo sapiens sapiens)

    PubMed Central

    Sherwood, Richard J.; Duren, Dana L.; Mahaney, Michael C.; Blangero, John; Dyer, Thomas D.; Cole, Shelley A.; Czerwinski, Stefan A.; Chumlea, Wm. Cameron; Siervogel, Roger M.; Choh, Audrey C.; Nahhas, Ramzi W.; Lee, Miryoung; Towne, Bradford

    2011-01-01

    The genetic architecture of the craniofacial complex has been the subject of intense scrutiny because of the high frequency of congenital malformations. Numerous animal models have been used to document the early development of the craniofacial complex, but few studies have focused directly on the genetic underpinnings of normal variation in the human craniofacial complex. The current study examines 80 quantitative traits derived from lateral cephalographs of 981 participants in the Fels Longitudinal Study, Wright State University, Dayton, Ohio. Quantitative genetic analyses were conducted using the SOLAR analytic platform, a maximum-likelihood variance components method that incorporates all familial information for parameter estimation. Heritability estimates were significant and of moderate to high magnitude for all craniofacial traits. Additionally, significant quantitative trait loci (QTL) were identified for 10 traits from the three developmental components (basicranium, splanchnocranium, and neurocranium) of the craniofacial complex. These QTL were found on chromosomes 3, 6, 11, 12, and 14. This study of the genetic architecture of the craniofacial complex elucidates fundamental information of the genetic architecture of the craniofacial complex in humans. PMID:21328561

  1. Quantitative Clinical Diagnostic Analysis of Acetone in Human Blood by HPLC: A Metabolomic Search for Acetone as Indicator

    PubMed Central

    Akgul Kalkan, Esin; Sahiner, Mehtap; Ulker Cakir, Dilek; Alpaslan, Duygu; Yilmaz, Selehattin

    2016-01-01

    Using high-performance liquid chromatography (HPLC) and 2,4-dinitrophenylhydrazine (2,4-DNPH) as a derivatizing reagent, an analytical method was developed for the quantitative determination of acetone in human blood. The determination was carried out at 365 nm using an ultraviolet-visible (UV-Vis) diode array detector (DAD). For acetone as its 2,4-dinitrophenylhydrazone derivative, a good separation was achieved with a ThermoAcclaim C18 column (15 cm × 4.6 mm × 3 μm) at retention time (tR) 12.10 min and flowrate of 1 mL min−1 using a (methanol/acetonitrile) water elution gradient. The methodology is simple, rapid, sensitive, and of low cost, exhibits good reproducibility, and allows the analysis of acetone in biological fluids. A calibration curve was obtained for acetone using its standard solutions in acetonitrile. Quantitative analysis of acetone in human blood was successfully carried out using this calibration graph. The applied method was validated in parameters of linearity, limit of detection and quantification, accuracy, and precision. We also present acetone as a useful tool for the HPLC-based metabolomic investigation of endogenous metabolism and quantitative clinical diagnostic analysis. PMID:27298750

  2. Thyrotropin-luteinizing hormone/chorionic gonadotropin receptor extracellular domain chimeras as probes for thyrotropin receptor function

    SciTech Connect

    Nagayama, Yuji; Wadsworth, H.L.; Chazenbalk, G.D.; Russo, D.; Seto, Pui; Rapoport, B. Univ. of California, San Francisco )

    1991-02-01

    To define the sites in the extracellular domain of the human thyrotropin (TSH) receptor that are involved in TSH binding and signal transduction the authors constructed chimeric thyrotropin-luteinizing hormone/chorionic gonadotropin (TSH-LH/CG) receptors. The extracellular domain of the human TSH receptor was divided into five regions that were replaced, either singly or in various combinations, with homologous regions of the rat LH/CG receptor. The chimeric receptors were stably expressed in Chinese hamster ovary cells. The data obtained suggest that the carboxyl region of the extracellular domain (amino acid residues 261-418) and particularly the middle region (residues 171-260) play a role in signal transduction. The possibility is also raised of an interaction between the amino and carboxyl regions of the extracellular domain in the process of signal transduction. In summary, these studies suggest that the middle region and carboxyl half of the extracellular domain of the TSH receptor are involved in signal transduction and that the TSH-binding region is likely to span the entire extracellular domain, with multiple discontinuous contact sites.

  3. Effects of flucycloxuron, a chitin synthesis inhibitor, on reproductive events and thickness of chorion in mealworms.

    PubMed

    Hami, M; Taibi, F; Soltani-Mazouni, N

    2004-01-01

    Flucycloxuron (FCX), a benzoylphenylurea derivative, was evaluated on Tenebrio molitor. The compound was incorporated into the diet and administrated to newly emerged females at various doses (2, 5 and 10 mg/kg). FCX was found to affect several reproductive events such as the duration of preovipostion and oviposition period, the fecundity, the viability of eggs and the duration of embryonic development, respectively. Morphological study of ovaries showed that FCX reduced both oocytes number, the ovaries weight and the size and the volume of the basal oocyte during the sexual maturation. In addition, it reduced the thickness of chorion from freshly laid eggs. However, electron microscopic study revealed that this compound had no significant effect on the fine structure of chorion. Finally, measurements of ovarian ecdysteroids production by an enzyme immunoassay indicated a reduction in the hormonal amounts recorded. PMID:15759421

  4. Chorionic villus sampling and materno-fetal transfusions: an immunological pathogenesis of vascular disruptive syndromes?

    PubMed

    Los, F J; Noomen, P; Vermeij-Keers, C; Gaillard, J L; Brandenburg, H; Jahoda, M G; Luider, T M

    1996-03-01

    Experimental materno-embryonic transfusions with serum that is immunologically active against blood group antigens cause congenital malformations in the rat embryo. In view of the possible increased incidence of vascular disruptive syndromes after chorionic villus sampling (CVS), we investigated the occurrence of materno-fetal transfusions (MFTs) in this procedure. In 18 pregnant women experiencing two needle introductions at CVS, we looked immunohistochemically at the presence of haemoglobin A1-containing maternal erythrocytes in the fetal circulation of the separately collected first and second chorionic villus samples. In 4 of 18 patients (22 per cent), a significant increase of maternal cells was observed in the second sample compared with the first sample, indicating the occurrence of MFT by CVS. On the rare occasion of maternal immunization against fetal antigens, a CVS-associated MFT might provoke immunological damage to the fetus. PMID:8710770

  5. Tissue-specific expression of silkmoth chorion genes in vivo using Bombyx mori nuclear polyhedrosis virus as a transducing vector.

    PubMed Central

    Iatrou, K; Meidinger, R G

    1990-01-01

    A pair of silkmoth chorion chromosomal genes, HcA.12-HcB.12, was inserted into a baculovirus transfer vector, pBmp2, derived from the nuclear polyhedrosis virus of Bombyx mori. This vector, which permits the insertion of foreign genetic material in the vicinity of a mutationally inactivated polyhedrin gene, was used to acquire the corresponding recombinant virus. Injection of mutant silkmoth pupae that lack all Hc chorion genes with the recombinant virus resulted in the infection of all internal organs including follicular tissue. Analysis of RNA from infected tissues has demonstrated that the two chorion genes present in the viral genome are correctly transcribed under the control of their own promoter in follicular cells, the tissue in which chorion genes are normally expressed. The chorion primary transcripts are also correctly processed in the infected follicular cells and yield mature mRNAs indistinguishable from authentic chorion mRNAs present in wild-type follicles. These results demonstrate that recombinant nuclear polyhedrosis viruses can be used as transducing vectors for introducing genetic material of host origin into the cells of the organism and that the transduced genes are transiently expressed in a tissue-specific manner under the control of their resident regulatory sequences. Thus we show the in vivo expression of cloned genes under cellular promoter control in an insect other than Drosophila melanogaster. The approach should be applicable to all insect systems that are subject to nuclear polyhedrosis virus infection. Images PMID:2187186

  6. Quantitative Proteomic Analysis of Human Lung Tumor Xenografts Treated with the Ectopic ATP Synthase Inhibitor Citreoviridin

    PubMed Central

    Wu, Yi-Hsuan; Hu, Chia-Wei; Chien, Chih-Wei; Chen, Yu-Ju; Huang, Hsuan-Cheng; Juan, Hsueh-Fen

    2013-01-01

    ATP synthase is present on the plasma membrane of several types of cancer cells. Citreoviridin, an ATP synthase inhibitor, selectively suppresses the proliferation and growth of lung cancer without affecting normal cells. However, the global effects of targeting ectopic ATP synthase in vivo have not been well defined. In this study, we performed quantitative proteomic analysis using isobaric tags for relative and absolute quantitation (iTRAQ) and provided a comprehensive insight into the complicated regulation by citreoviridin in a lung cancer xenograft model. With high reproducibility of the quantitation, we obtained quantitative proteomic profiling with 2,659 proteins identified. Bioinformatics analysis of the 141 differentially expressed proteins selected by their relative abundance revealed that citreoviridin induces alterations in the expression of glucose metabolism-related enzymes in lung cancer. The up-regulation of enzymes involved in gluconeogenesis and storage of glucose indicated that citreoviridin may reduce the glycolytic intermediates for macromolecule synthesis and inhibit cell proliferation. Using comprehensive proteomics, the results identify metabolic aspects that help explain the antitumorigenic effect of citreoviridin in lung cancer, which may lead to a better understanding of the links between metabolism and tumorigenesis in cancer therapy. PMID:23990911

  7. Application of quantitative PCR assays to detection of human Bacteroides species in the intestines of pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    When evaluating the efficacy of probiotic bacteria, it is beneficial to know whether a fed bacterium reaches the appropriate location in the digestive tract. Use of quantitative PCR could detect specific bacteria in a sample with a complex microbial community. In order to determine whether three Bac...

  8. Microscopic chorionic pseudocysts in placental membranes: a histologic lesion of in utero hypoxia.

    PubMed

    Stanek, Jerzy; Weng, Eric

    2007-01-01

    Grossly apparent and microscopically intermediate trophoblast-lined subchorionic, septal, and cell island cysts are relatively common placental findings. To analyze the clinicopathologic correlations of histologically similar but grossly inapparent microscopic chorionic pseudocysts (lakes) arising in the chorion laeve of placental membranes (mccpm), selected placental and clinical parameters of all 172 consecutive placentas with mccpm (study group, sg) and all consecutive 3743 placentas without mccpm (comparative group, cg) from years 1994 through 2005 were statistically compared; mccpm were observed in 4.3% of all placentas and in 14.9% of placentas from preeclamptic mothers from 24- to 42-week pregnancies, their gestational weeks' distribution almost mirroring that of the distribution of preeclampsia, with a peak in the middle of the 3rd trimester. Microscopic chorionic pseudocysts (lakes) arising in the chorion laeve of placental membranes were statistically significantly more common in patients with preeclampsia and maternal diabetes mellitus. In placentas with mccpm, decidual arteriolopathy, homogeneous placental maturation, global hypoxic pattern of placental injury, chorangiosis, placental infarction, laminar necrosis of membranes, stem obliterative endarteritis, erythroblasts of fetal blood, and decidual hemosiderosis were statistically significantly more common, while acute chorioamnionitis, villous fibrosis, and villous edema were less common (P < or = 0.05). There were no statistically significant differences between sg and cg in meconium staining, retroplacental hematoma, perivillous fibrin deposition, intervillous thrombi, chronic villitis, chorangiomas, placenta accreta, amnion nodosum, and marginate/vallate placenta. Highly statistically significant associations of mccpm with preeclampsia and a cluster of placental lesions known to be linked to placental hypoxia indicate that the mccpm form in response to hypoxia, particularly in patients with

  9. Clinical outcomes after assisted reproductive technology in twin pregnancies: chorionicity-based comparison.

    PubMed

    Sun, Luming; Zou, Gang; Wei, Xing; Chen, Yan; Zhang, Jun; Okun, Nanette; Duan, Tao

    2016-01-01

    The chorionicity-based evaluation of the perinatal risk in twin pregnancies after assisted reproductive technology (ART) is lacking. A retrospective review was performed of all twin pregnancies monitored prenatally and delivered at our hospital between 2010 and 2014. Chorionicity was diagnosed by ultrasound examination at first trimester and confirmed by postnatal pathology. Pregnancy and perinatal outcomes were prospectively recorded. Adjusted odds ratios (aOR) with 95% confidence intervals (CI) were calculated in a logistic regression model. A total of 1153 twin pregnancies were analyzed. The occurrence of preterm premature rupture of membranes (PPROM) was 3 times as frequent in monochorionic diamniotic (MCDA) twin pregnancies after ART as in those spontaneous counterparts (aOR 3.0; 95%CI 1.1-3.2). The prevalence of intrahepatic cholestasis of pregnancies (ICP) was significantly higher in dichorionic diamniotic (DCDA) twin pregnancies following ART compared to spontaneous DCDA pregnancies (aOR 3.3; 95%CI 1.3-5.6). Perinatal outcomes did not differ between two conception methods, either in MCDA or DCDA twin pregnancies. Based on differentiation of chorionicity, ART is associated with the increased risk of PPROM in MCDA twin pregnancies and with a higher rate of ICP in DCDA twin gestations. ART does not increase adversity of perinatal outcomes in twin pregnancies. PMID:27243373

  10. QUANTITATIVE TOXICOPROTEOMIC ANALYSIS OF CARCINOGEN-TREATED ANIMAL TISSUES AND HUMAN CELLS FOR HUMAN HEALTH RISK ASSESSMENT

    EPA Science Inventory

    Humans are exposed to a variety of environmental toxicants, and this together with a large number of interacting factors can contribute to an individual's risk for health. To understand the toxic mechanisms and/or modes of action for human health risk assessment, molecular charac...

  11. Automated quantitative analysis of 3D morphology and mean corpuscular hemoglobin in human red blood cells stored in different periods.

    PubMed

    Moon, Inkyu; Yi, Faliu; Lee, Yeon H; Javidi, Bahram; Boss, Daniel; Marquet, Pierre

    2013-12-16

    Quantitative phase (QP) images of red blood cells (RBCs), which are obtained by off-axis digital holographic microscopy, can provide quantitative information about three-dimensional (3D) morphology of human RBCs and the characteristic properties such as mean corpuscular hemoglobin (MCH) and MCH surface density (MCHSD). In this paper, we investigate modifications of the 3D morphology and MCH in RBCs induced by the period of storage time for the purpose of classification of RBCs with different periods of storage by using off-axis digital holographic microscopy. The classification of RBCs based on the duration of storage is highly relevant because a long storage of blood before transfusion may alter the functionality of RBCs and, therefore, cause complications in patients. To analyze any changes in the 3D morphology and MCH of RBCs due to storage, we use data sets from RBC samples stored for 8, 13, 16, 23, 27, 30, 34, 37, 40, 47, and 57 days, respectively. The data sets consist of more than 3,300 blood cells in eleven classes, with more than 300 blood cells per class. The classes indicate the storage period of RBCs and are listed in chronological order. Using the RBCs donated by healthy persons, the off-axis digital holographic microscopy reconstructs several quantitative phase images of RBC samples stored for eleven different periods. We employ marker-controlled watershed transform to remove the background in the RBC quantitative phase images obtained by the off-axis digital holographic microscopy. More than 300 single RBCs are extracted from the segmented quantitative phase images for each class. Such a large number of RBC samples enable us to obtain statistical distributions of the characteristic properties of RBCs after a specific period of storage. Experimental results show that the 3D morphology of the RBCs, in contrast to MCH, is essentially related to the aging of the RBCs. PMID:24514667

  12. [Quantitative determination of IgA from human bronchial mucus (author's transl)].

    PubMed

    Hayem, A; Laine, A; Bailleul, V; Lebas, J

    1975-05-15

    Before quantitative analysis of bronchial mucus proteins, sputum must be transformed into a homogeneous medium: this is done by ultrasonic action and contact with cationic resin AG 50 W X 2. Then, acid mucins are fixed on Ecteola-cellulose, and proteins extracted by a phosphate/saline buffer at pH 5. IgA is estimated by electroimmunodiffusion. Reproducibility of the method is discussed: the accuracy for IgA values is 4%. PMID:238765

  13. Quantitative comparisons of the actute neurotoxicity of toulene in rats and humans.

    EPA Science Inventory

    The behavioral and neurophysiological effects of acute exposure to toluene are the most thoroughly explored of all the hydrocarbon solvents. Behavioral effects have been experimentally studied in humans and other species, for example, rats. The existence of both rat and human dos...

  14. Quantitative determination of the human breast milk macronutrients by near-infrared Raman spectroscopy

    NASA Astrophysics Data System (ADS)

    Motta, Edlene d. C. M.; Zângaro, Renato A.; Silveira, Landulfo, Jr.

    2012-03-01

    This work proposes the evaluation of the macronutrient constitution of human breast milk based on the spectral information provided by near-infrared Raman spectroscopy. Human breast milk (5 mL) from a subject was collected during the first two weeks of breastfeeding and stocked in -20°C freezer. Raman spectra were measured using a Raman spectrometer (830 nm excitation) coupled to a fiber based Raman probe. Spectra of human milk were dominated by bands of proteins, lipids and carbohydrates in the 600-1800 cm-1 spectral region. Raman spectroscopy revealed differences in the biochemical constitution of human milk depending on the time of breastfeeding startup. This technique could be employed to develop a classification routine for the milk in Human Milk Banking (HMB) depending on the nutritional facts.

  15. Quantitative phase imaging of human red blood cells using phase-shifting white light interference microscopy with colour fringe analysis

    NASA Astrophysics Data System (ADS)

    Singh Mehta, Dalip; Srivastava, Vishal

    2012-11-01

    We report quantitative phase imaging of human red blood cells (RBCs) using phase-shifting interference microscopy. Five phase-shifted white light interferograms are recorded using colour charge coupled device camera. White light interferograms were decomposed into red, green, and blue colour components. The phase-shifted interferograms of each colour were then processed by phase-shifting analysis and phase maps for red, green, and blue colours were reconstructed. Wavelength dependent refractive index profiles of RBCs were computed from the single set of white light interferogram. The present technique has great potential for non-invasive determination of refractive index variation and morphological features of cells and tissues.

  16. Luteinizing hormone/chorionic gonadotrophin receptor overexpressed in granulosa cells from polycystic ovary syndrome ovaries is functionally active.

    PubMed

    Kanamarlapudi, Venkateswarlu; Gordon, Uma D; López Bernal, Andrés

    2016-06-01

    Polycystic ovarian syndrome (PCOS) is associated with anovulatory infertility. Luteinizing hormone/chorionic gonadotrophin receptor (LHCGR), which is critical for ovulation, has been suggested to be expressed prematurely in the ovarian follicles of women with PCOS. This study aimed to analyse the expression and activity of LHCGR in ovarian granulosa cells from PCOS patients and the involvement of ARF6 small GTPase in LHCGR internalization. Granulosa cells (GC) isolated from follicular fluid collected during oocyte retrieval from normal women (n = 19) and women with PCOS (n = 17) were used to study differences in LHCGR protein expression and activity between normal and PCOS patients. LHCGR expression is up-regulated in GC from PCOS women. LHCGR in PCOS GC is functionally active, as shown by increased cAMP production upon human gonadotrophin (HCG)-stimulation. Moreover, ARF6 is highly expressed in GC from PCOS patients and HCG-stimulation increases the concentrations of active ARF6. The inhibition of ARF6 activation attenuates HCG-induced LHCGR internalization in both normal and PCOS GC, indicating that there are no alterations in LHCGR internalisation in GC from PCOS. In conclusion, the expression and activation of LHCGR and ARF6 are up-regulated in GC from PCOS women but the mechanism of agonist-induced LHCGR internalization is unaltered. PMID:27061682

  17. The Use of a Dehydrated Amnion/Chorion Membrane Allograft in Patients Who Subsequently Undergo Reexploration after Posterior Lumbar Instrumentation

    PubMed Central

    Subach, Brian R.; Copay, Anne G.

    2015-01-01

    Background Context. Products that can reduce development of epidural fibrosis may reduce risk for ongoing pain associated with development of scar tissue and make subsequent epidural reexploration easier. Purpose. To evaluate the use of dehydrated human amnion/chorion membrane (dHACM) on the formation of soft tissue scarring in the epidural space. Study Design. Case series. Patient Sample. Five patients having transforaminal lumbar interbody lumbar fusion (TLIF) with posterior instrumentation and implantation of dHACM in the epidural space and subsequent epidural reexploration. Outcome Measures. Degree of scar tissue adjacent to the epidural space at reexploration. Intraoperative and postoperative complications related to dHACM and patient reported outcomes. Methods. The degree of scar tissue adjacent to the epidural space was assessed during the reexploration surgery. Patients' outcomes were collected using standard validated questionnaires. Results. Four of 5 cases had easily detachable tissue during epidural reexploration. Angiolipoma of 10% was noted in 1 case and 5% in 2 cases. Significant improvements in patient reported outcomes were observed. No intraoperative or postoperative complications occurred. Conclusions. Our findings suggest that dHACM implant during TLIF may have favorable effects on epidural fibrosis and is well tolerated. Further studies with larger cohorts are required to prove our results. PMID:25653880

  18. Human Fecal Source Identification: Real-Time Quantitative PCR Method Standardization

    EPA Science Inventory

    Method standardization or the formal development of a protocol that establishes uniform performance benchmarks and practices is necessary for widespread adoption of a fecal source identification approach. Standardization of a human-associated fecal identification method has been...

  19. HUMAN FECAL SOURCE IDENTIFICATION: REAL-TIME QUANTITATIVE PCR METHOD STANDARDIZATION - abstract

    EPA Science Inventory

    Method standardization or the formal development of a protocol that establishes uniform performance benchmarks and practices is necessary for widespread adoption of a fecal source identification approach. Standardization of a human-associated fecal identification method has been...

  20. Approaches to advancing quantitative human health risk assessment of environmental chemicals in the post-genomic era

    SciTech Connect

    Chiu, Weihsueh A.; Euling, Susan Y.; Scott, Cheryl Siegel; Subramaniam, Ravi P.

    2013-09-15

    The contribution of genomics and associated technologies to human health risk assessment for environmental chemicals has focused largely on elucidating mechanisms of toxicity, as discussed in other articles in this issue. However, there is interest in moving beyond hazard characterization to making more direct impacts on quantitative risk assessment (QRA) — i.e., the determination of toxicity values for setting exposure standards and cleanup values. We propose that the evolution of QRA of environmental chemicals in the post-genomic era will involve three, somewhat overlapping phases in which different types of approaches begin to mature. The initial focus (in Phase I) has been and continues to be on “augmentation” of weight of evidence — using genomic and related technologies qualitatively to increase the confidence in and scientific basis of the results of QRA. Efforts aimed towards “integration” of these data with traditional animal-based approaches, in particular quantitative predictors, or surrogates, for the in vivo toxicity data to which they have been anchored are just beginning to be explored now (in Phase II). In parallel, there is a recognized need for “expansion” of the use of established biomarkers of susceptibility or risk of human diseases and disorders for QRA, particularly for addressing the issues of cumulative assessment and population risk. Ultimately (in Phase III), substantial further advances could be realized by the development of novel molecular and pathway-based biomarkers and statistical and in silico models that build on anticipated progress in understanding the pathways of human diseases and disorders. Such efforts would facilitate a gradual “reorientation” of QRA towards approaches that more directly link environmental exposures to human outcomes.

  1. Quantitative podocyte parameters predict human native kidney and allograft half-lives

    PubMed Central

    Cibrik, Diane; Hodgin, Jeffrey B.; Wu, Fan; Zhang, Min; Kikuchi, Masao; Wickman, Larysa; Samaniego, Milagros; Bitzer, Markus; Wiggins, Jocelyn E.; Ojo, Akinlolu; Li, Yi; Wiggins, Roger C.

    2016-01-01

    BACKGROUND Kidney function decreases with age. A potential mechanistic explanation for kidney and allograft half-life has evolved through the realization that linear reduction in glomerular podocyte density could drive progressive glomerulosclerosis to impact both native kidney and allograft half-lives. METHODS Predictions from podometrics (quantitation of podocyte parameters) were tested using independent pathologic, functional, and outcome data for native kidneys and allografts derived from published reports and large registries. RESULTS With age, native kidneys exponentially develop glomerulosclerosis, reduced renal function, and end-stage kidney disease, projecting a finite average kidney life span. The slope of allograft failure rate versus age parallels that of reduction in podocyte density versus age. Quantitative modeling projects allograft half-life at any donor age, and rate of podocyte detachment parallels the observed allograft loss rate. CONCLUSION Native kidneys are designed to have a limited average life span of about 100–140 years. Allografts undergo an accelerated aging-like process that accounts for their unexpectedly short half-life (about 15 years), the observation that older donor age is associated with shorter allograft half-life, and the fact that long-term allograft survival has not substantially improved. Podometrics provides potential readouts for these processes, thereby offering new approaches for monitoring and intervention. FUNDING National Institutes of Health. PMID:27280173

  2. Quantitation of human MAO A and B in liver, intestine and placenta: Reassessment of activity

    SciTech Connect

    Riley, L.A.

    1989-01-01

    Monoamine oxidases (MAO) oxidize a variety of exogenous and endogenous amines including neurotransmitters such as serotonin, dopamine and norepinephrine as well as the potent dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP). The two forms of MAO (A and B) differ in molecular weight and inhibitor selectivity, and are differentially expressed in the nervous system and many other tissues. Although some substrates are preferentially oxidized by one form of MAO, substrates that can be oxidized by only one MAO form have not been reported. How well each MAO oxidizes various substrates has not been thoroughly characterized because of difficulties in separating and quantitating MAO A and B active sites. By immunoblotting SDS-polyacrylamide gels of mitochondrial extracts with monoclonal antibodies specific for each form of MAO, MAO B protein was detected in intestine and placenta, tissues that have been reported to contain MAO A activity. An improved procedure was developed for quantitating the ratio and amounts of MAO A and B active sites, using the ligand ({sup 3}H)-pargyline to label MAO and specific monoclonal antibodies to separate MAO A from B. Data from liver, placenta and platelets were used to re-evaluate the molecular activity of both MAO A and B for six commonly studied substrates.

  3. On the quantitative characterization of human body sway in experiments with long-term performance.

    PubMed

    Seidel, H; Bräuer, D; Bastek, R; Issel, I

    1978-01-01

    Two different conditions of standing were examined in young male subjects with an intact vestibular system by means of stabilography. Low-frequency sways were eliminated by means of a digital high-pass filter. The parameters of the histograms of amplitudes, the autocorrelation functions, and the power-spectral density were estimated. The quantitative parameters of the stabilograms are compared for the two standing positions, both for the frontal and sagittal planes. Quotients of estimates of variance components calculated on the basis of analysis of variance are stated as measures of reliability for the parameters of histograms of amplitudes and power-spectral density in defined frequency bands. The reliability of the standard deviations of amplitudes was much better than the reliability of skewness and excess. The reliability of spectral density differed for frequency ranges investigated. Generally, the quotients of estimates of variance components were higher for the spectral density in the frequency bands of sagittal stabilograms than in those of frontal ones. Performance significantly affected several parameters of histograms and the distributions of power-spectral density. Standard deviation of amplitudes and power-spectral density proved to be suitable for the quantitative characterization of stabilograms. PMID:752208

  4. Semi-quantitative evaluation of fecal contamination potential by human and ruminant sources using multiple lines of evidence

    USGS Publications Warehouse

    Stoeckel, D.M.; Stelzer, E.A.; Stogner, R.W.; Mau, D.P.

    2011-01-01

    Protocols for microbial source tracking of fecal contamination generally are able to identify when a source of contamination is present, but thus far have been unable to evaluate what portion of fecal-indicator bacteria (FIB) came from various sources. A mathematical approach to estimate relative amounts of FIB, such as Escherichia coli, from various sources based on the concentration and distribution of microbial source tracking markers in feces was developed. The approach was tested using dilute fecal suspensions, then applied as part of an analytical suite to a contaminated headwater stream in the Rocky Mountains (Upper Fountain Creek, Colorado). In one single-source fecal suspension, a source that was not present could not be excluded because of incomplete marker specificity; however, human and ruminant sources were detected whenever they were present. In the mixed-feces suspension (pet and human), the minority contributor (human) was detected at a concentration low enough to preclude human contamination as the dominant source of E. coli to the sample. Without the semi-quantitative approach described, simple detects of human-associated marker in stream samples would have provided inaccurate evidence that human contamination was a major source of E. coli to the stream. In samples from Upper Fountain Creek the pattern of E. coli, general and host-associated microbial source tracking markers, nutrients, and wastewater-associated chemical detections-augmented with local observations and land-use patterns-indicated that, contrary to expectations, birds rather than humans or ruminants were the predominant source of fecal contamination to Upper Fountain Creek. This new approach to E. coli allocation, validated by a controlled study and tested by application in a relatively simple setting, represents a widely applicable step forward in the field of microbial source tracking of fecal contamination. ?? 2011 Elsevier Ltd.

  5. /sup 99m/Tc-labeled solid-phase meal: a quantitative clinical measurement of human gastric emptying

    SciTech Connect

    Martin, J.L.; Beck, W.J.; McDonald, A.P.; Carlson, G.M.; Mathias, J.R.

    1983-08-01

    A solid-phase meal labeled with /sup 99m/Tc-sulfur colloid provides an improved clinical test for the quantitative evaluation of human gastric emptying. We studied 12 healthy male controls and five male patients with known gastric stasis secondary to a vagotomy and drainage procedure. All subjects were fasted for 8 hours before the study, and each consumed an unbuttered biscuit and a poached egg white containing 1 mCi of /sup 99m/Tc-sulfur colloid. For 2 hours, 60-second counts were measured every 10 minutes by a Pho Gamma III scintillation camera. The t/sup 1///sup 2/ for control subjects was 60 minutes, at which time patients with gastric stasis had retained 98% of the test meal. At 120 minutes, control subjects and patients with gastric stasis had 4.7% and 89%, respectively, of the meal remaining in the stomach. The solid-phase test meal labeled with /sup 99m/Tc-sulfur colloid is easy to perform and can be used clinically to quantitatively measure gastric emptying in humans. This test can discriminate between control subjects and patients with known gastric stasis.

  6. Quantitation of Gingerols in Human Plasma by Newly Developed Stable Isotope Dilution Assays and Assessment of Their Immunomodulatory Potential.

    PubMed

    Schoenknecht, Carola; Andersen, Gaby; Schmidts, Ines; Schieberle, Peter

    2016-03-23

    In a pilot study with two volunteers, the main pungent and bioactive ginger (Zingiber officinale Roscoe) compounds, the gingerols, were quantitated in human plasma after ginger tea consumption using a newly established HPLC-MS/MS(ESI) method on the basis of stable isotope dilution assays. Limits of quantitation for [6]-, [8]-, and [10]-gingerols were determined as 7.6, 3.1, and 4.0 nmol/L, respectively. The highest plasma concentrations of [6]-, [8]-, and [10]-gingerols (42.0, 5.3, and 4.8 nmol/L, respectively) were reached 30-60 min after ginger tea intake. Incubation of activated human T lymphocytes with gingerols increased the intracellular Ca(2+) concentration as well as the IFN-γ secretion by about 20-30%. This gingerol-induced increase of IFN-γ secretion could be blocked by the specific TRPV1 antagonist SB-366791. The results of the present study point to an interaction of gingerols with TRPV1 in activated T lymphocytes leading to an augmentation of IFN-γ secretion. PMID:26939769

  7. Visualization and quantitative analysis of extrachromosomal telomere-repeat DNA in individual human cells by Halo-FISH

    PubMed Central

    Komosa, Martin; Root, Heather; Meyn, M. Stephen

    2015-01-01

    Current methods for characterizing extrachromosomal nuclear DNA in mammalian cells do not permit single-cell analysis, are often semi-quantitative and frequently biased toward the detection of circular species. To overcome these limitations, we developed Halo-FISH to visualize and quantitatively analyze extrachromosomal DNA in single cells. We demonstrate Halo-FISH by using it to analyze extrachromosomal telomere-repeat (ECTR) in human cells that use the Alternative Lengthening of Telomeres (ALT) pathway(s) to maintain telomere lengths. We find that GM847 and VA13 ALT cells average ∼80 detectable G/C-strand ECTR DNA molecules/nucleus, while U2OS ALT cells average ∼18 molecules/nucleus. In comparison, human primary and telomerase-positive cells contain <5 ECTR DNA molecules/nucleus. ECTR DNA in ALT cells exhibit striking cell-to-cell variations in number (<20 to >300), range widely in length (<1 to >200 kb) and are composed of primarily G- or C-strand telomere-repeat DNA. Halo-FISH enables, for the first time, the simultaneous analysis of ECTR DNA and chromosomal telomeres in a single cell. We find that ECTR DNA comprises ∼15% of telomere-repeat DNA in GM847 and VA13 cells, but <4% in U2OS cells. In addition to its use in ALT cell analysis, Halo-FISH can facilitate the study of a wide variety of extrachromosomal DNA in mammalian cells. PMID:25662602

  8. Quantitative GSL-glycome analysis of human whole serum based on an EGCase digestion and glycoblotting method.

    PubMed

    Furukawa, Jun-ichi; Sakai, Shota; Yokota, Ikuko; Okada, Kazue; Hanamatsu, Hisatoshi; Kobayashi, Takashi; Yoshida, Yasunobu; Higashino, Kenichi; Tamura, Tomohiro; Igarashi, Yasuyuki; Shinohara, Yasuro

    2015-12-01

    Glycosphingolipids (GSLs) are lipid molecules linked to carbohydrate units that form the plasma membrane lipid raft, which is clustered with sphingolipids, sterols, and specific proteins, and thereby contributes to membrane physical properties and specific recognition sites for various biological events. These bioactive GSL molecules consequently affect the pathophysiology and pathogenesis of various diseases. Thus, altered expression of GSLs in various diseases may be of importance for disease-related biomarker discovery. However, analysis of GSLs in blood is particularly challenging because GSLs are present at extremely low concentrations in serum/plasma. In this study, we established absolute GSL-glycan analysis of human serum based on endoglycoceramidase digestion and glycoblotting purification. We established two sample preparation protocols, one with and the other without GSL extraction using chloroform/methanol. Similar amounts of GSL-glycans were recovered with the two protocols. Both protocols permitted absolute quantitation of GSL-glycans using as little as 20 μl of serum. Using 10 healthy human serum samples, up to 42 signals corresponding to GSL-glycan compositions could be quantitatively detected, and the total serum GSL-glycan concentration was calculated to be 12.1-21.4 μM. We further applied this method to TLC-prefractionated serum samples. These findings will assist the discovery of disease-related biomarkers by serum GSL-glycomics. PMID:26420879

  9. Visualization and quantitative analysis of extrachromosomal telomere-repeat DNA in individual human cells by Halo-FISH.

    PubMed

    Komosa, Martin; Root, Heather; Meyn, M Stephen

    2015-02-27

    Current methods for characterizing extrachromosomal nuclear DNA in mammalian cells do not permit single-cell analysis, are often semi-quantitative and frequently biased toward the detection of circular species. To overcome these limitations, we developed Halo-FISH to visualize and quantitatively analyze extrachromosomal DNA in single cells. We demonstrate Halo-FISH by using it to analyze extrachromosomal telomere-repeat (ECTR) in human cells that use the Alternative Lengthening of Telomeres (ALT) pathway(s) to maintain telomere lengths. We find that GM847 and VA13 ALT cells average ∼80 detectable G/C-strand ECTR DNA molecules/nucleus, while U2OS ALT cells average ∼18 molecules/nucleus. In comparison, human primary and telomerase-positive cells contain <5 ECTR DNA molecules/nucleus. ECTR DNA in ALT cells exhibit striking cell-to-cell variations in number (<20 to >300), range widely in length (<1 to >200 kb) and are composed of primarily G- or C-strand telomere-repeat DNA. Halo-FISH enables, for the first time, the simultaneous analysis of ECTR DNA and chromosomal telomeres in a single cell. We find that ECTR DNA comprises ∼15% of telomere-repeat DNA in GM847 and VA13 cells, but <4% in U2OS cells. In addition to its use in ALT cell analysis, Halo-FISH can facilitate the study of a wide variety of extrachromosomal DNA in mammalian cells. PMID:25662602

  10. Quantitative gene expression profiling of mouse brain regions reveals differential transcripts conserved in human and affected in disease models.

    PubMed

    Brochier, Camille; Gaillard, Marie-Claude; Diguet, Elsa; Caudy, Nicolas; Dossat, Carole; Ségurens, Béatrice; Wincker, Patrick; Roze, Emmanuel; Caboche, Jocelyne; Hantraye, Philippe; Brouillet, Emmanuel; Elalouf, Jean-Marc; de Chaldée, Michel

    2008-04-22

    Using serial analysis of gene expression, we collected quantitative transcriptome data in 11 regions of the adult wild-type mouse brain: the orbital, prelimbic, cingulate, motor, somatosensory, and entorhinal cortices, the caudate-putamen, the nucleus accumbens, the thalamus, the substantia nigra, and the ventral tegmental area. With >1.2 million cDNA tags sequenced, this database is a powerful resource to explore brain functions and disorders. As an illustration, we performed interregional comparisons and found 315 differential transcripts. Most of them are poorly characterized and 20% lack functional annotation. For 78 differential transcripts, we provide independent expression level measurements in mouse brain regions by real-time quantitative RT-PCR. We also show examples where we used in situ hybridization to achieve infrastructural resolution. For 30 transcripts, we next demonstrated that regional enrichment is conserved in the human brain. We then quantified the expression levels of region-enriched transcripts in the R6/2 mouse model of Huntington disease and the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of Parkinson disease and observed significant alterations in the striatum, cerebral cortex, thalamus and substantia nigra of R6/2 mice and in the striatum of MPTP-treated mice. These results show that the gene expression data provided here for the mouse brain can be used to explore pathophysiological models and disclose transcripts differentially expressed in human brain regions. PMID:18252803

  11. Simultaneous quantitative determination of alpha-ketoglutaric acid and 5-hydroxymethylfurfural in human plasma by gas chromatography-mass spectrometry.

    PubMed

    Wagner, Bernhard M; Donnarumma, Fabrizio; Wintersteiger, Reinhold; Windischhofer, Werner; Leis, Hans J

    2010-04-01

    Alpha-ketoglutaric acid (alpha-KG) and 5-hydroxymethylfurfural (5-HMF) are currently under investigation as promising cancer cell damaging agents. A method for the simultaneous quantitative determination of alpha-KG and 5-HMF in human plasma was established for screening these compounds in human plasma. Plasma samples were directly treated with O-(2,3,4,5,6-pentafluorobenzyl) hydroxylamine hydrochloride to form the corresponding oximes, thus facilitating subsequent liquid-liquid extraction. After formation of the trimethylsilyl ethers, samples were analyzed by gas chromatography with electron ionization mass spectrometry. Stable isotope labeled standards were used, the preparation of (13)C(6)-5-HMF is described. Limits of quantitation were set to 0.938 microg/mL for alpha-KG and 0.156 microg/mL for 5-HMF. Inter-day accuracy was < or = 93.7% (alpha-KG) and < or = 92.8% (5-HMF). Inter-day precision was < or = 6.0% (alpha-KG) and < or = 4.6% (5-HMF). The method has been successfully applied to pharmacokinetic profiling of the compounds after intravenous application. PMID:20155414

  12. Age- and gender-dependent heterogeneous proportion of variation explained by SNPs in quantitative traits reflecting human health.

    PubMed

    Lee, Dain; Lee, Chaeyoung

    2015-01-01

    Age-related effects are often included as covariates in the analytical model for genome-wide association analysis of quantitative traits reflecting human health. Nevertheless, previous studies have hardly examined the effects of age on the proportion of variation explained by single nucleotide polymorphisms (PVSNP) in these traits. In this study, the PVSNP estimates of body mass index (BMI), waist-to-hip ratio, pulse pressure, high-density lipoprotein cholesterol level, triglyceride level (TG), low-density lipoprotein cholesterol level, and glucose level were obtained from Korean consortium metadata partitioned by gender or by age. Restricted maximum likelihood estimates of the PVSNP were obtained in a mixed model framework. Previous studies using pedigree data suggested possible differential heritability of certain traits with regard to gender, which we observed in our current study (BMI and TG; P < 0.05). However, the PVSNP analysis based on age revealed that, with respect to every trait tested, individuals aged 40 to 49 exhibited significantly lower PVSNP estimates than individuals aged 50 to 59 or 60 to 69 (P < 0.05). The consistent heterogeneous PVSNP with respect to age may be due to degenerated genetic functions in individuals between the ages of 50 and 69. Our results suggest the genetic mechanism of age- and gender-dependent PVSNP of quantitative traits related to human health should be further examined. PMID:25701395

  13. Quantitative analysis of squalamine, a self-ionization-suppressing aminosterol sulfate, in human plasma by LC-MS/MS.

    PubMed

    Li, Austin C; Sabo, Andrew M; McCormick, Timothy; Johnston, Sean M

    2004-02-18

    The special physico-chemical property of squalamine enables the formation of intra- or inter-molecular non-volatile strong salt, which is difficult to ionize in a mass spectrometer's interface. A sensitive, accurate, precise, and specific method for the quantitative determination of this self ion-suppressing compound in human plasma has been developed and validated using high performance liquid chromatography (HPLC) coupled with positive electrospray tandem mass spectrometry (MS/MS). Solid phase extraction (SPE) technique was utilized to extract human plasma samples using the Waters Oasis HLB cartridges. Deuterated squalamine was used as the internal standard (IS). Positive multiple reaction monitoring (MRM) mode was used to achieve both sensitivity and selectivity. A quadratic linearity range over 5-1000 ng/ml, R > 0.999 was achieved. Performance of the method has been validated and met all the specifications set forth in the US Food and Drug Administration's May 2001 "Bioanalytical Method Validation Guidance for Industry". Different sample reconstitution solutions were found to have dramatic impact on sensitivity of mass spectrometer used to squalamine. This is the first quantitation method using a positive and true multiple reaction monitoring mode detection for squalamine. PMID:15127819

  14. QUANTITATIVE MORPHOLOGICAL AND MOLECULAR PATHOLOGY OF THE HUMAN THYMUS CORRELATE WITH INFANT CAUSE OF DEATH

    PubMed Central

    Lloyd, Mark C.; Burke, Nancy; Kalantarpour, Fatemeh; Niesen, Melissa I.; Hall, Aaron; Pennypacker, Keith; Citron, Bruce; Pick, Chaim G.; Adams, Vernard; Das, Mahasweta; Mohapatra, Shyam; Cualing, Hernani; Blanck, George

    2014-01-01

    The objective of this study was to investigate and quantify the morphological and molecular changes in the thymus for common causes of human infant death. Thymic architecture and molecular changes apparent in human infant head trauma victims were assessed by microscopy and quantified by image analysis of digital whole slide images. Thymuses from victims of SIDS and suffocated infants displaying normal thymus architecture were used for comparison. Molecular expression of proliferation and serotonin receptor and transporter protein markers was evaluated. Duplicate morphological and molecular studies of rodent thymuses were completed with both mouse and rat models. Quantification of novel parameters of digital images of thymuses from human infants suffering mortal head trauma revealed a disruption of the corticomedullary organization of the thymus, particularly involving dissolution of the corticomedullary border. A similar result was obtained for related mouse and rat models. The human thymuses from head trauma cases also displayed a higher percentage of Ki-67-positive thymocytes. Finally, we determined that thymus expression of the human serotonin receptor, and the serotonin transporter, occur almost exclusively in the thymic medulla. Head trauma leads to a disruption of the thymic, corticomedullary border, and molecular expression patterns in a robust and quantifiable manner. PMID:25309682

  15. Quantitative evaluation of p53 as a new indicator of DNA damage in human spermatozoa

    PubMed Central

    Raimondo, Salvatore; Gentile, Tommaso; Cuomo, Felice; De Filippo, Stefania; Aprea, Gilda E.; Guida, John

    2014-01-01

    BACKGROUND: Sperm DNA integrity is considered an important parameter to assess seminal fluid quality and can be used as a predictive test of potential fertility. Amongst the various tests to determine sperm DNA integrity, one is the Acridine Orange test. Recent studies have demonstrated the importance of p53 in maintaining sperm DNA integrity. The aim of this study was to assess if a p53 ELISA assay could be a new indicator of DNA damage in human spermatozoa. MATERIALS AND METHODS: 103 human semen samples were evaluated using both Acridine Orange test and p53 ELISA and results were compared. RESULTS: A clear correlation between the values measured by two methods was obtained. CONCLUSIONS: If this hypothesis will be confirmed by further studies, the p53 ELISA assay could become a new and more precise indicator of DNA damage in human spermatozoa. PMID:25395748

  16. Quantitative Determination of Lethal Toxin Proteins in Culture Supernatant of Human Live Anthrax Vaccine Bacillus anthracis A16R.

    PubMed

    Zai, Xiaodong; Zhang, Jun; Liu, Ju; Liu, Jie; Li, Liangliang; Yin, Ying; Fu, Ling; Xu, Junjie; Chen, Wei

    2016-03-01

    Bacillus anthracis (B. anthracis) is the etiological agent of anthrax affecting both humans and animals. Anthrax toxin (AT) plays a major role in pathogenesis. It includes lethal toxin (LT) and edema toxin (ET), which are formed by the combination of protective antigen (PA) and lethal factor (LF) or edema factor (EF), respectively. The currently used human anthrax vaccine in China utilizes live-attenuated B. anthracis spores (A16R; pXO1+, pXO2-) that produce anthrax toxin but cannot produce the capsule. Anthrax toxins, especially LT, have key effects on both the immunogenicity and toxicity of human anthrax vaccines. Thus, determining quantities and biological activities of LT proteins expressed by the A16R strain is meaningful. Here, we explored LT expression patterns of the A16R strain in culture conditions using another vaccine strain Sterne as a control. We developed a sandwich ELISA and cytotoxicity-based method for quantitative detection of PA and LF. Expression and degradation of LT proteins were observed in culture supernatants over time. Additionally, LT proteins expressed by the A16R and Sterne strains were found to be monomeric and showed cytotoxic activity, which may be the main reason for side effects of live anthrax vaccines. Our work facilitates the characterization of anthrax vaccines components and establishment of a quality control standard for vaccine production which may ultimately help to ensure the efficacy and safety of the human anthrax vaccine A16R. PMID:26927174

  17. Quantitative Determination of Lethal Toxin Proteins in Culture Supernatant of Human Live Anthrax Vaccine Bacillus anthracis A16R

    PubMed Central

    Zai, Xiaodong; Zhang, Jun; Liu, Ju; Liu, Jie; Li, Liangliang; Yin, Ying; Fu, Ling; Xu, Junjie; Chen, Wei

    2016-01-01

    Bacillus anthracis (B. anthracis) is the etiological agent of anthrax affecting both humans and animals. Anthrax toxin (AT) plays a major role in pathogenesis. It includes lethal toxin (LT) and edema toxin (ET), which are formed by the combination of protective antigen (PA) and lethal factor (LF) or edema factor (EF), respectively. The currently used human anthrax vaccine in China utilizes live-attenuated B. anthracis spores (A16R; pXO1+, pXO2−) that produce anthrax toxin but cannot produce the capsule. Anthrax toxins, especially LT, have key effects on both the immunogenicity and toxicity of human anthrax vaccines. Thus, determining quantities and biological activities of LT proteins expressed by the A16R strain is meaningful. Here, we explored LT expression patterns of the A16R strain in culture conditions using another vaccine strain Sterne as a control. We developed a sandwich ELISA and cytotoxicity-based method for quantitative detection of PA and LF. Expression and degradation of LT proteins were observed in culture supernatants over time. Additionally, LT proteins expressed by the A16R and Sterne strains were found to be monomeric and showed cytotoxic activity, which may be the main reason for side effects of live anthrax vaccines. Our work facilitates the characterization of anthrax vaccines components and establishment of a quality control standard for vaccine production which may ultimately help to ensure the efficacy and safety of the human anthrax vaccine A16R. PMID:26927174

  18. Development of quantitative structure activity relationships for the binding affinity of methoxypyridinium cations for human acetylcholinesterase.

    PubMed

    Morrill, Jason A; Topczewski, Joseph J; Lodge, Alexander M; Yasapala, Nilanthi; Quinn, Daniel M

    2015-11-01

    Among the most toxic substances known are the organophosphorus (OP) compounds used as pesticides and chemical warfare agents. Owing to their high toxicity there is a number of efforts underway to develop effective therapies for OP agent exposure. To date all therapies in use treat inhibited acetylcholinesterase (AChE), but are ineffective for the treatment of inhibited AChE, which has undergone a subsequent hydrolysis process, referred to as aging. Toward developing a therapy for treating victims of OP intoxication in the aged state we have developed Quantitative Structure-Activity Relationships (QSARs) based on the AM1 semiempirical quantum mechanical method using the program, CODESSA (COmprehensive Descriptors for Structural and Statistical Analysis). Using this methodology we obtained a multiple correlation QSAR equation which gave R(2)=0.9359 for a random training set of 38 ligands and R(2)=0.9236 for prediction on a random test set of 9 ligands. PMID:26454505

  19. Quantitative non-invasive intracellular imaging of Plasmodium falciparum infected human erythrocytes

    NASA Astrophysics Data System (ADS)

    Edward, Kert; Farahi, Faramarz

    2014-05-01

    Malaria is a virulent pathological condition which results in over a million annual deaths. The parasitic agent Plasmodium falciparum has been extensively studied in connection with this epidemic but much remains unknown about its development inside the red blood cell host. Optical and fluorescence imaging are among the two most common procedures for investigating infected erythrocytes but both require the introduction of exogenous contrast agents. In this letter, we present a procedure for the non-invasive in situ imaging of malaria infected red blood cells. The procedure is based on the utilization of simultaneously acquired quantitative phase and independent topography data to extract intracellular information. Our method allows for the identification of the developmental stages of the parasite and facilitates in situ analysis of the morphological changes associated with the progression of this disease. This information may assist in the development of efficacious treatment therapies for this condition.

  20. Quantitative Structure Activity Relationship for Inhibition of Human Organic Cation/Carnitine Transporter (OCTN2)

    PubMed Central

    Diao, Lei; Ekins, Sean; Polli, James E.

    2010-01-01

    Organic cation/carnitine transporter (OCTN2; SLC22A5) is an important transporter for L-carnitine homeostasis, but can be inhibited by drugs, which may cause L-carnitine deficiency and possibly other OCTN2-mediated drug-drug interactions. One objective was to develop a quantitative structure–activity relationship (QSAR) of OCTN2 inhibitors, in order to predict and identify other potential OCTN2 inhibitors and infer potential clinical interactions. A second objective was to assess two high renal clearance drugs that interact with OCTN2 in vitro (cetirizine and cephaloridine) for possible OCTN2-mediated drug-drug interactions. Using previously generated in vitro data of 22 drugs, a 3D quantitative pharmacophore model and a Bayesian machine learning model were developed. The four pharmacophore features include two hydrophobic groups, one hydrogen-bond acceptor, and one positive ionizable center. The Bayesian machine learning model was developed using simple interpretable descriptors and function class fingerprints of maximum diameter 6 (FCFP_6). An external test set of 27 molecules, including 15 newly identified OCTN2 inhibitors, and a literature test set of 22 molecules were used to validate both models. The computational models afforded good capability to identify structurally diverse OCTN2 inhibitors, providing a valuable tool to predict new inhibitors efficiently. Inhibition results confirmed our previously observed association between rhabdomyolysis and Cmax/Ki ratio. The two high renal clearance drugs cetirizine and cephaloridine were found not to be OCTN2 substrates and their diminished elimination by other drugs is concluded not to be mediated by OCTN2. PMID:20831193

  1. Accuracy and precision of quantitative 31P-MRS measurements of human skeletal muscle mitochondrial function.

    PubMed

    Layec, Gwenael; Gifford, Jayson R; Trinity, Joel D; Hart, Corey R; Garten, Ryan S; Park, Song Y; Le Fur, Yann; Jeong, Eun-Kee; Richardson, Russell S

    2016-08-01

    Although theoretically sound, the accuracy and precision of (31)P-magnetic resonance spectroscopy ((31)P-MRS) approaches to quantitatively estimate mitochondrial capacity are not well documented. Therefore, employing four differing models of respiratory control [linear, kinetic, and multipoint adenosine diphosphate (ADP) and phosphorylation potential], this study sought to determine the accuracy and precision of (31)P-MRS assessments of peak mitochondrial adenosine-triphosphate (ATP) synthesis rate utilizing directly measured peak respiration (State 3) in permeabilized skeletal muscle fibers. In 23 subjects of different fitness levels, (31)P-MRS during a 24-s maximal isometric knee extension and high-resolution respirometry in muscle fibers from the vastus lateralis was performed. Although significantly correlated with State 3 respiration (r = 0.72), both the linear (45 ± 13 mM/min) and phosphorylation potential (47 ± 16 mM/min) models grossly overestimated the calculated in vitro peak ATP synthesis rate (P < 0.05). Of the ADP models, the kinetic model was well correlated with State 3 respiration (r = 0.72, P < 0.05), but moderately overestimated ATP synthesis rate (P < 0.05), while the multipoint model, although being somewhat less well correlated with State 3 respiration (r = 0.55, P < 0.05), most accurately reflected peak ATP synthesis rate. Of note, the PCr recovery time constant (τ), a qualitative index of mitochondrial capacity, exhibited the strongest correlation with State 3 respiration (r = 0.80, P < 0.05). Therefore, this study reveals that each of the (31)P-MRS data analyses, including PCr τ, exhibit precision in terms of mitochondrial capacity. As only the multipoint ADP model did not overstimate the peak skeletal muscle mitochondrial ATP synthesis, the multipoint ADP model is the only quantitative approach to exhibit both accuracy and precision. PMID:27302751

  2. A quantitative evaluation of human coordination interfaces for computer assisted surgery.

    PubMed

    Cardin, M A; Wang, J X; Lobaugh, N J; Guimont, I; Plewes, D B

    2007-03-01

    Computer assisted surgery (CAS) for tumor resection can assist the surgeon in locating the tumor margin accurately via some form of guidance method. A wide array of guidance methods can be considered, including model-based visual representations, symbolic graphical interfaces, and those based on other sensory cues such as sound. Given the variety of these guidance methods, it becomes increasingly important to test and analyze guidance methods for CAS in a quantitative and context-dependent manner to determine which is most suitable for a given surgical task. In this paper, we present a novel experimental methodology and analysis framework to test candidate guidance methods for CAS. Different viewpoints and stereographic, symbolic and auditory cues were tested in isolation or in combination in a set of virtual surgery experiments. A total of 28 participants were asked to circumscribe a virtual tumor with a magnetically tracked scalpel while measuring the surgical trajectory. This allowed measurement of surgical accuracy, speed, and the frequency with which the tumor margin was intersected, and enabled a quantitative comparison of guidance approaches. This study demonstrated that adding sound to pictorial guidance methods consistently improved accuracy, speed and margin intersection of the virtual surgery. However, the use of stereovision showed less benefit than expected. While guidance based on a combination of symbolic and pictorial cues enhanced accuracy, we found that speed could be substantially impaired. These studies demonstrate that optimal guidance combinations exist which would not be apparent by studying individual guidance methods in isolation. Our findings suggest that care is needed when using expensive and sometimes cumbersome virtual visualization technologies for CAS, and that simpler, non-stereo presentation may be sufficient for specific surgical tasks. PMID:17487657

  3. A Quantitative ADME-base Tool for Exploring Human Exposure to Consumer Product Ingredients

    EPA Science Inventory

    Exposure to a wide range of chemicals through our daily habits and routines is ubiquitous and largely unavoidable within modern society. The potential for human exposure, however, has not been quantified for the vast majority of chemicals with wide commercial use. Creative advanc...

  4. A QUANTITATIVE COMPARISON OF THE EFFECTS OF ACUTE INHALED TOLUENE IN HUMAN RATS

    EPA Science Inventory

    The effects of acute exposure to toluene have been explored more thoroughly than other hydrocarbon solvents. These effects have been experimentally studied in humans and other species, e.g., rats, as well as in a number of in vitro preparations. The existence ofdosimetric and eff...

  5. Success Factors of Human Resource Profession in Malaysia: A Quantitative Study

    ERIC Educational Resources Information Center

    Long, Choi Sang

    2013-01-01

    Human Resource (HR) profession is moving toward being more aligned with business outcomes. To function constructively and to play critical roles more effectively, HR professionals must master the necessary competencies. The competencies that are examined in this study are business knowledge, strategy architect, HR delivery, credible activist and…

  6. Social Indicators of Basic Needs: Quantitative Data for Human Rights Policy.

    ERIC Educational Resources Information Center

    de Neufville, Judith Innes

    Developing social indicators of basic human needs involves (1) recognizing the problems in selection, (2) identifying the criteria for making selections, (3) choosing which basic needs to cover, and (4) selecting the indicators. The social indicators are to help formulate U.S. foreign policy and will be used by the State Department's Bureau of…

  7. Effect of lufenuron on chorionic and cuticular structure of unhatched larval Ctenocephalides felis (Siphonaptera: Pulicidae).

    PubMed

    Meola, R W; Dean, S R; Meola, S M; Sittertz-Bhatkar, H; Schenker, R

    1999-01-01

    When adult cat fleas, Ctenocephalides felis (Bouché), were fed concentrations of < or = 0.08 ppm lufenuron in cattle blood, egg hatch did not differ significantly from the controls. However, as the concentration of lufenuron in blood increased from 0.125 to 1.0 ppm egg hatch decreased to 64 and 2%, respectively. Most of these eggs contained fully developed larvae. Microscopic examination of unhatched larvae, revealed that the cuticle epidermal cells, chorion, and vitelline membrane all were affected by lufenuron treatment. Larvae often produced 2 separate cuticles in response to treatment. The 1st cuticle consisted of an indistinct layer of epicuticle and a procuticle composed of randomly deposited chitin microfibrils. After the 1st layer of procuticle separated from the epidermal cells, a 2nd layer of procuticle was deposited. It was not possible to determine whether the egg tooth was functional during larval hatch. The surface of the egg tooth appeared normal, but the cuticle may have had structural abnormalities similar to those seen in other areas of the exoskeleton. Structural defects appeared to be due to the cytotoxic effects of lufenuron. The epidermal cells of treated larvae showed evidence of disintegration (i.e., the nuclei and mitochondria appeared to be degenerating and the amount of endoplasmic reticulum and other cytoplasmic organelles was decreased). The chorion of lufenuron-treated larvae consisted of an outer layer, middle and inner layers that were thinner and less electron dense than those of controls, and lacked the innermost chorionic layer found in the control larvae. The vitelline membrane also was thinner than that of the controls. Larval hatching was prevented by ruptures in the cuticle, which opened during eclosion resulting in the loss of hemolymph and desiccation of the larva. Evidently, tearing of the cuticle was caused by abnormal formation of the procuticle that was not strong enough to withstand the cuticular expansion and muscular

  8. Quantitative assay for mutation in diploid human lymphoblasts using microtiter plates

    SciTech Connect

    Furth, E.A.; Thilly, W.G.; Penman, B.W.; Liber, H.L.; Rand, W.M.

    1981-01-01

    A microtiter plating technique which eliminates the need for soft agar and fibroblast feeder layers to determine the colony-forming ability of diploid human lymphoblast lines was described. The calculation of cloning efficiency is based on the Poisson distribution, and a statistical method for calculating confidence intervals is presented. This technique has been applied to the comcomitant examination of induced mutation at the putative loci for hypoxanthine guanine phosphoribosyl transferase, thymidine, kinase, and Na/sup +//K/sup +/ adenosine triphosphatase.

  9. Gonadotropin-regulated Testicular RNA Helicase (GRTH/DDX25), a Negative Regulator of Luteinizing/Chorionic Gonadotropin Hormone-induced Steroidogenesis in Leydig Cells

    PubMed Central

    Fukushima, Masato; Villar, Joaquin; Tsai-Morris, Chon-Hwa; Dufau, Maria L.

    2011-01-01

    Gonadotropin-regulated testicular RNA helicase (GRTH/DDX25) is a testis-specific gonadotropin-regulated RNA helicase that is present in Leydig cells (LCs) and germ cells and is essential for spermatid development and completion of spermatogenesis. Normal basal levels of testosterone in serum and LCs were observed in GRTH null (GRTH−/−) mice. However, testosterone production was enhanced in LCs of GRTH−/− mice compared with WT mice by both in vivo and in vitro human chorionic gonadotropin stimulation. LCs of GRTH−/− mice had swollen mitochondria with a significantly increased cholesterol content in the inner mitochondrial membrane. Basal protein levels of SREBP2, HMG-CoA reductase, and steroidogenic acute regulatory protein (StAR; a protein that transports cholesterol to the inner mitochondrial membrane) were markedly increased in LCs of GRTH−/− mice compared with WT mice. Gonadotropin stimulation caused an increase in StAR mRNA levels and protein expression in GRTH−/− mice versus WT mice, with no further increase in SREBP2 and down-regulation of HMG-CoA reductase protein. The half-life of StAR mRNA was significantly increased in GRTH−/− mice. Moreover, association of StAR mRNA with GRTH protein was observed in WT mice. Human chorionic gonadotropin increased GRTH gene expression and its associated StAR protein at cytoplasmic sites. Taken together, these findings indicate that, through its negative role in StAR message stability, GRTH regulates cholesterol availability at the mitochondrial level. The finding of an inhibitory action of GRTH associated with gonadotropin-mediated steroidogenesis has provided insights into a novel negative autocrine molecular control mechanism of this helicase in the regulation of steroid production in the male. PMID:21719703

  10. [Exclusion of Sandhoff disease (Tay-Sachs 0 variant) by chorion biopsy].

    PubMed

    Veszprémi, B; Baranyai, Z; Klujber, L; Arany, A

    1992-04-01

    Transcervical chorionic villus sampling with ultrasound guidance at the 11-th week of pregnancy was made at a woman with the history of one lethal case of Sandhoff disease. The total hexosaminidase and the hexosaminidase A were determined. At the 16-th week amniocentesis was performed and the characteristic enzymes were determined from the amniotic cell culture. The results of the examinations made possible to advise the patient to carry out the pregnancy. The examinations after delivery confirmed the newborn to be a carrier. PMID:1522989

  11. First-trimester prenatal diagnosis of mucolipidosis II (I-cell disease) by chorionic biopsy.

    PubMed Central

    Poenaru, L; Castelnau, L; Dumez, Y; Thepot, F

    1984-01-01

    We investigated the possibility of mucolipidosis type II (ML II) prenatal diagnosis by lysosomal enzyme determination on trophoblast biopsy obtained at 10 weeks of gestation in two pregnancies at risk. Diagnosis of ML II was made in both cases on fresh chorionic villi on the basis of depressed beta-galactosidase activity, and after abortion, the diagnosis was confirmed on fresh fetal tissues and on cells cultured from trophoblast and fetuses. We stress the importance of culturing cells from the trophoblast biopsy to ensure a reliable diagnosis. PMID:6440435

  12. Amnion and Chorion Membranes: Potential Stem Cell Reservoir with Wide Applications in Periodontics

    PubMed Central

    2015-01-01

    The periodontal therapy usually aims at elimination of disease causing bacteria and resolution of inflammation. It involves either resective or regenerative surgery to resolve the inflammation associated defects. Over the years, several methods have been used for achievement of periodontal regeneration. One of the oldest biomaterials used for scaffolds is the fetal membrane. The amniotic membranes of developing embryo, that is, amnion (innermost lining) and chorion (a layer next to it), have the properties with significant potential uses in dentistry. This paper reviews the properties, mechanism of action, and various applications of these placental membranes in general and specifically in Periodontics. PMID:26770199

  13. Integrative Quantitative Proteomics Unveils Proteostasis Imbalance in Human Hepatocellular Carcinoma Developed on Nonfibrotic Livers*

    PubMed Central

    Negroni, Luc; Taouji, Said; Arma, Daniela; Pallares-Lupon, Nestor; Leong, Kristen; Beausang, Lee Anne; Latterich, Martin; Bossé, Roger; Balabaud, Charles; Schmitter, Jean-Marie; Bioulac-Sage, Paulette; Zucman-Rossi, Jessica; Rosenbaum, Jean; Chevet, Eric

    2014-01-01

    Proteomics-based clinical studies represent promising resources for the discovery of novel biomarkers or for unraveling molecular mechanisms underlying particular diseases. Here, we present a discovery study of hepatocellular carcinoma developed on nonfibrotic liver (nfHCC) that combines complementary quantitative iTRAQ-based proteomics and phosphoproteomics approaches. Using both approaches, we compared a set of 24 samples (18 nfHCC versus six nontumor liver tissue). We identified 43 proteins (67 peptides) differentially expressed and 32 peptides differentially phosphorylated between the experimental groups. The functional analysis of the two data sets pointed toward the deregulation of a protein homeostasis (proteostasis) network including the up-regulation of the Endoplasmic Reticulum (ER) resident HSPA5, HSP90B1, PDIA6, and P4HB and of the cytosolic HSPA1B, HSP90AA1, HSPA9, UBC, CNDP2, TXN, and VCP as well as the increased phosphorylation of the ER resident calnexin at Ser583. Antibody-based validation approaches (immunohistochemistry, immunoblot, Alphascreen®, and AMMP®) on independent nfHCC tumor sets (up to 77 samples) confirmed these observations, thereby indicating a common mechanism occurring in nfHCC tumors. Based on these results we propose that adaptation to proteostasis imbalance in nfHCC tumors might confer selective advantages to those tumors. As such, this model could provide an additional therapeutic opportunity for those tumors arising on normal liver by targeting the tumor proteostasis network. Data are available via ProteomeXchange with identifier PXD001253. PMID:25225353

  14. High-performance liquid chromatographic assay for the quantitation of L-arginine in human plasma.

    PubMed

    Gopalakrishnan, V; Burton, P J; Blaschke, T F

    1996-10-01

    L-Arginine is metabolized to nitric oxide by nitric oxide synthase, and abnormalities in nitric oxide production have been implicated in the pathogenesis of some diseases involving the vasculature. Thus, there has been interest in the effects of pharmacologic doses of L-arginine in patients with cardiovascular and renal diseases. To study the disposition of exogenous doses, an HPLC method was developed to analyze plasma samples for L-arginine. The assay involves precolumn derivatization of arginine with naphthalenedicarboxaldehyde and cyanide followed by HPLC with UV detection. Only a simple deproteinization of the plasma samples was required. The derivatized arginine was stable (less than 5% degradation in 20 h), facilitating batch sample processing and analysis in an autosampler. Calibration curves were generated in Ringer's lactate solution instead of plasma to correct for endogenous plasma L-arginine. Recovery in plasma, compared to Ringer's solution (n = 4), was 103%. Mean intraday assay precision (n = 6), expressed as coefficient of variation, was 3.4%. Interassay precision (n = 6) was 7%. The assay was applied for the quantitation of L-arginine in plasma samples from a normal subject who had been given a single oral (10 g) and a single intravenous dose (30 g) of exogenous L-arginine. PMID:8843144

  15. Quantitation and pharmacokinetic modeling of therapeutic antibody quality attributes in human studies.

    PubMed

    Li, Yinyin; Monine, Michael; Huang, Yu; Swann, Patrick; Nestorov, Ivan; Lyubarskaya, Yelena

    2016-01-01

    A thorough understanding of drug metabolism and disposition can aid in the assessment of efficacy and safety. However, analytical methods used in pharmacokinetics (PK) studies of protein therapeutics are usually based on ELISA, and therefore can provide a limited perspective on the quality of the drug in concentration measurements. Individual post-translational modifications (PTMs) of protein therapeutics are rarely considered for PK analysis, partly because it is technically difficult to recover and quantify individual protein variants from biological fluids. Meanwhile, PTMs may be directly linked to variations in drug efficacy and safety, and therefore understanding of clearance and metabolism of biopharmaceutical protein variants during clinical studies is an important consideration. To address such challenges, we developed an affinity-purification procedure followed by peptide mapping with mass spectrometric detection, which can profile multiple quality attributes of therapeutic antibodies recovered from patient sera. The obtained data enable quantitative modeling, which allows for simulation of the PK of different individual PTMs or attribute levels in vivo and thus facilitate the assessment of quality attributes impact in vivo. Such information can contribute to the product quality attribute risk assessment during manufacturing process development and inform appropriate process control strategy. PMID:27216574

  16. Application of quantitative uncertainty analysis for human health risk assessment at Rocky Flats

    SciTech Connect

    Duncan, F.L.W.; Gordon, J.W. ); Smith, D. ); Singh, S.P. )

    1993-01-01

    The characterization of uncertainty is an important component of the risk assessment process. According to the U.S. Environmental Protection Agency's (EPA's) [open quotes]Guidance on Risk Characterization for Risk Managers and Risk Assessors,[close quotes] point estimates of risk [open quotes]do not fully convey the range of information considered and used in developing the assessment.[close quotes] Furthermore, the guidance states that the Monte Carlo simulation may be used to estimate descriptive risk percentiles. To provide information about the uncertainties associated with the reasonable maximum exposure (RME) estimate and the relation of the RME to other percentiles of the risk distribution for Operable Unit 1 (OU-1) at Rocky Flats, uncertainties were identified and quantitatively evaluated. Monte Carlo simulation is a technique that can be used to provide a probability function of estimated risk using random values of exposure factors and toxicity values in an exposure scenario. The Monte Carlo simulation involves assigning a joint probability distribution to the input variables (i.e., exposure factors) of an exposure scenario. Next, a large number of independent samples from the assigned joint distribution are taken and the corresponding outputs calculated. Methods of statistical inference are used to estimate, from the output sample, some parameters of the output distribution, such as percentiles and the expected value.

  17. Involution of human fetal Leydig cells. An immunohistochemical, ultrastructural and quantitative study.

    PubMed Central

    Codesal, J; Regadera, J; Nistal, M; Regadera-Sejas, J; Paniagua, R

    1990-01-01

    The testes of stillborn fetuses (from 13 to 28 weeks of gestational age), fetuses born alive (from 29 weeks of gestational age) who died a few days later, and infants dying 1 to 8 months after birth were processed for light and electron microscopy. Paraffin-embedded material was stained with the avidin-biotin peroxidase complex (ABC) method for immunohistochemical detection of testosterone (T) in order to quantify the age-related changes in the number of T-positive interstitial cells. This number decreased progressively from the 24th week of gestation up to birth and remained unchanged up to the second month of postnatal life. During the third month of age, the number of T-positive cells rose markedly but fell again from the fourth month to the end of the study. The ultrastructural study revealed the following types of interstitial cells at all ages studied: fibroblast-like cells, myofibroblast-like cells, developed fetal Leydig cells, degenerating fetal Leydig cells and infantile Leydig cells with a multilobed nucleus and focal cytoplasmic accumulations of smooth endoplasmic reticulum and lipid droplets. Quantitative ultrastructural studies revealed that the changes in the number of fetal Leydig cells with age were similar to those found in the number of T-positive cells although, for each age studied, absolute values were higher in the ultrastructural study. The number of infantile Leydig cells increased with age. Images Figs. 1-4 Figs. 5-9 Figs. 10-11 PMID:2272896

  18. Quantitative Determination of the Human Immune Response to Immunization with Meningococcal Vaccines

    PubMed Central

    Gotschlich, Emil C.; Rey, Michel; Triau, Rene; Sparks, Kenneth J.

    1972-01-01

    Radioactive antigen binding tests have been developed to measure quantitatively the antibody response of 167 adults, 84 children, and 51 infants to several different preparations of group A and group C meningococcal polysaccharides. Almost all the adults injected responded and the geometric mean responses were approximately 15 μg/ml of antibody protein in individuals vaccinated subcutaneously with two preparations of group A vaccine. The geometric mean antibody concentration after immunization with two preparations of group C vaccine was approximately 35 μg/ml. Most children aged 7 yr responded to immunization with two group A vaccines, and their mean response was only slightly less than that seen in adults. There was no difference between the subcutaneous and the intradermal route if both were given with jet gun. The majority of infants aged 6-13 months responded to a preparation of group A vaccine and the geometric mean titer was approximately 1.2 μg/ml. Adults, children, and infants responded significantly less to a preparation of group A polysaccharide which was of low molceular weight. PMID:4621363

  19. Challenges of interdisciplinary research: reconciling qualitative and quantitative methods for understanding human-landscape systems.

    PubMed

    Lach, Denise

    2014-01-01

    While interdisciplinary research is increasingly practiced as a way to transcend the limitations of individual disciplines, our concepts, and methods are primarily rooted in the disciplines that shape the way we think about the world and how we conduct research. While natural and social scientists may share a general understanding of how science is conducted, disciplinary differences in methodologies quickly emerge during interdisciplinary research efforts. This paper briefly introduces and reviews different philosophical underpinnings of quantitative and qualitative methodological approaches and introduces the idea that a pragmatic, realistic approach may allow natural and social scientists to work together productively. While realism assumes that there is a reality that exists independently of our perceptions, the work of scientists is to explore the mechanisms by which actions cause meaningful outcomes and the conditions under which the mechanisms can act. Our task as interdisciplinary researchers is to use the insights of our disciplines in the context of the problem to co-produce an explanation for the variables of interest. Research on qualities necessary for successful interdisciplinary researchers is also discussed along with recent efforts by funding agencies and academia to increase capacities for interdisciplinary research. PMID:23892682

  20. Challenges of Interdisciplinary Research: Reconciling Qualitative and Quantitative Methods for Understanding Human-Landscape Systems

    NASA Astrophysics Data System (ADS)

    Lach, Denise

    2014-01-01

    While interdisciplinary research is increasingly practiced as a way to transcend the limitations of individual disciplines, our concepts, and methods are primarily rooted in the disciplines that shape the way we think about the world and how we conduct research. While natural and social scientists may share a general understanding of how science is conducted, disciplinary differences in methodologies quickly emerge during interdisciplinary research efforts. This paper briefly introduces and reviews different philosophical underpinnings of quantitative and qualitative methodological approaches and introduces the idea that a pragmatic, realistic approach may allow natural and social scientists to work together productively. While realism assumes that there is a reality that exists independently of our perceptions, the work of scientists is to explore the mechanisms by which actions cause meaningful outcomes and the conditions under which the mechanisms can act. Our task as interdisciplinary researchers is to use the insights of our disciplines in the context of the problem to co-produce an explanation for the variables of interest. Research on qualities necessary for successful interdisciplinary researchers is also discussed along with recent efforts by funding agencies and academia to increase capacities for interdisciplinary research.

  1. Quantitation and pharmacokinetic modeling of therapeutic antibody quality attributes in human studies

    PubMed Central

    Li, Yinyin; Monine, Michael; Huang, Yu; Swann, Patrick; Nestorov, Ivan; Lyubarskaya, Yelena

    2016-01-01

    ABSTRACT A thorough understanding of drug metabolism and disposition can aid in the assessment of efficacy and safety. However, analytical methods used in pharmacokinetics (PK) studies of protein therapeutics are usually based on ELISA, and therefore can provide a limited perspective on the quality of the drug in concentration measurements. Individual post-translational modifications (PTMs) of protein therapeutics are rarely considered for PK analysis, partly because it is technically difficult to recover and quantify individual protein variants from biological fluids. Meanwhile, PTMs may be directly linked to variations in drug efficacy and safety, and therefore understanding of clearance and metabolism of biopharmaceutical protein variants during clinical studies is an important consideration. To address such challenges, we developed an affinity-purification procedure followed by peptide mapping with mass spectrometric detection, which can profile multiple quality attributes of therapeutic antibodies recovered from patient sera. The obtained data enable quantitative modeling, which allows for simulation of the PK of different individual PTMs or attribute levels in vivo and thus facilitate the assessment of quality attributes impact in vivo. Such information can contribute to the product quality attribute risk assessment during manufacturing process development and inform appropriate process control strategy. PMID:27216574

  2. A device for in vivo measurements of quantitative ultrasound variables at the human proximal femur.

    PubMed

    Barkmann, Reinhard; Laugier, Pascal; Moser, Urs; Dencks, Stefanie; Klausner, Michael; Padilla, Frédéric; Haïat, Guilleaume; Glüer, Claus-C

    2008-01-01

    Quantitative ultrasound (QUS) at the calcaneus has similar power as a bone mineral density (BMD)- measurement using DXA for the prediction of osteoporotic fracture risk. Ultrasound equipment is less expensive than DXA and free of ionizing radiation. As a mechanical wave, QUS has the potential of measuring different bone properties than dual X-ray absorptiometry (DXA,) which depends on X-ray attenuation and might be developed into a tool of comprehensive assessment of bone strength. However, site-specific DXA at the proximal femur shows best performance in the prediction of hip fractures. To combine the potential of QUS with measurements directly at the femur, we developed a device for in vivo QUS measurements at this site. Methods comprise ultrasound transmission through the bone, reflection from the bone surface, and backscatter from the inner trabecular structure. The complete area of the proximal femur can be scanned except at the femoral head, which interferes with the ilium. To avoid edge artifacts, a subregion of the proximal femur in the trochanteric region was selected as measurement region. First, in vivo measurements demonstrate a good signal to noise ratio and proper depiction of the proximal femur on an attenuation image. Our results demonstrate the feasibility of in vivo measurements. Further improvements can be expected by refinement of the scanning technique and data evaluation method to enhance the potential of the new method for the estimation of bone strength. PMID:18599408

  3. Quantitative microbial risk assessment of human illness from exposure to marine beach sand.

    PubMed

    Shibata, Tomoyuki; Solo-Gabriele, Helena M

    2012-03-01

    Currently no U.S. federal guideline is available for assessing risk of illness from sand at recreational sites. The objectives of this study were to compute a reference level guideline for pathogens in beach sand and to compare these reference levels with measurements from a beach impacted by nonpoint sources of contamination. Reference levels were computed using quantitative microbial risk assessment (QMRA) coupled with Monte Carlo simulations. In order to reach an equivalent level of risk of illness as set by the U.S. EPA for marine water exposure (1.9 × 10(-2)), levels would need to be at least about 10 oocysts/g (about 1 oocyst/g for a pica child) for Cryptosporidium, about 5 MPN/g (about 1 MPN/g for pica) for enterovirus, and less than 10(6) CFU/g for S. aureus. Pathogen levels measured in sand at a nonpoint source recreational beach were lower than the reference levels. More research is needed in evaluating risk from yeast and helminth exposures as well as in identifying acceptable levels of risk for skin infections associated with sand exposures. PMID:22296573

  4. Quantitative optical coherence tomography angiography of vascular abnormalities in the living human eye.

    PubMed

    Jia, Yali; Bailey, Steven T; Hwang, Thomas S; McClintic, Scott M; Gao, Simon S; Pennesi, Mark E; Flaxel, Christina J; Lauer, Andreas K; Wilson, David J; Hornegger, Joachim; Fujimoto, James G; Huang, David

    2015-05-01

    Retinal vascular diseases are important causes of vision loss. A detailed evaluation of the vascular abnormalities facilitates diagnosis and treatment in these diseases. Optical coherence tomography (OCT) angiography using the highly efficient split-spectrum amplitude decorrelation angiography algorithm offers an alternative to conventional dye-based retinal angiography. OCT angiography has several advantages, including 3D visualization of retinal and choroidal circulations (including the choriocapillaris) and avoidance of dye injection-related complications. Results from six illustrative cases are reported. In diabetic retinopathy, OCT angiography can detect neovascularization and quantify ischemia. In age-related macular degeneration, choroidal neovascularization can be observed without the obscuration of details caused by dye leakage in conventional angiography. Choriocapillaris dysfunction can be detected in the nonneovascular form of the disease, furthering our understanding of pathogenesis. In choroideremia, OCT's ability to show choroidal and retinal vascular dysfunction separately may be valuable in predicting progression and assessing treatment response. OCT angiography shows promise as a noninvasive alternative to dye-based angiography for highly detailed, in vivo, 3D, quantitative evaluation of retinal vascular abnormalities. PMID:25897021

  5. Temporal lobe in human aging: A quantitative protein profiling study of samples from Chinese Human Brain Bank.

    PubMed

    Xu, Benhong; Xiong, Feng; Tian, Rui; Zhan, Shaohua; Gao, Yanpan; Qiu, Wenying; Wang, Renzhi; Ge, Wei; Ma, Chao

    2016-01-01

    The temporal lobe is a portion of the cerebral cortex with critical functionality. The age-related protein profile changes in the human temporal lobe have not been previously studied. This 4-plex tandem mass tag labeled proteomic study was performed on samples of temporal lobe from Chinese donors. Tissue samples were assigned to four age groups: Group A (the young, age: 34±13 years); Group B (the elderly, 62±5 years); Group C (the aged, 84±4 years) and Group D (the old, 95±1 years). Pooled samples from the different groups were subjected to proteomics and bioinformatics analysis to identify age-related changes in protein expression and associated pathways. We isolated 5072 proteins, and found that 67 proteins were downregulated and 109 proteins were upregulated in one or more groups during the aging process. Western blotting assays were performed to verify the proteomic results. Bioinformatic analysis identified proteins involved in neuronal degeneration, including proteins involved in neuronal firing, myelin sheath damage, and cell structure stability. We also observed the accumulation of extracellular matrix and lysosomal proteins which imply the occurrence of fibrosis and autophagy. Our results suggest a series of changes across a wide range of proteins in the human temporal lobe that may relate to aging and age-related neurodegenerative disorders. PMID:26631761

  6. Quantitative Evaluation of the Expression and Activity of Five Major Sulfotransferases (SULTs) in Human Tissues: The SULT “Pie”

    PubMed Central

    Riches, Zoe; Stanley, Emma L.; Bloomer, Jackie C.

    2009-01-01

    Expression levels of the major human sulfotransferases (SULTs) involved in xenobiotic detoxification in a range of human tissues (i.e., SULT “pies”) are not available in a form allowing comparison between tissues and individuals. Here we have determined, by quantitative immunoblotting, expression levels for the five principal human SULTs—SULT1A1, SULT1A3/4, SULT1B1, SULT1E1, and SULT2A1—and determined the kinetic properties toward probe substrates, where available, for these enzymes in cytosol samples from a bank of adult human liver, small intestine, kidney, and lung. We produced new isoform-selective antibodies against SULT1B1 and SULT2A1, which were used alongside antibodies against SULT1A3 and SULT1A1 previously produced in our laboratory or available commercially (SULT1E1). Expression levels were derived using purified recombinant enzymes to construct standard curves for each individual isoform and immunoblot. Substantial intertissue and interindividual differences in expression were observed. SULT1A1 was the major enzyme (>50% of total, range 420-4900 ng/mg cytosol protein) in the liver, followed by SULT2A1, SULT1B1, and SULT1E1. SULT1A3 was completely absent from this tissue. In contrast, the small intestine contained the largest overall amount of SULT of any of the tissues, with SULT1B1 the major enzyme (36%), closely followed by SULT1A3 (31%), and SULT1A1, SULT1E1, and SULT2A1 more minor forms (19, 8, and 6% of total, respectively). The kidney and lung contained low levels of SULT. We provide a unique data set that will add value to the study of the role and contribution of sulfation to drug and xenobiotic metabolism in humans. PMID:19679676

  7. Quantitative Cross-Species Extrapolation between Humans and Fish: The Case of the Anti-Depressant Fluoxetine

    PubMed Central

    Margiotta-Casaluci, Luigi; Owen, Stewart F.; Cumming, Rob I.; de Polo, Anna; Winter, Matthew J.; Panter, Grace H.; Rand-Weaver, Mariann; Sumpter, John P.

    2014-01-01

    Fish are an important model for the pharmacological and toxicological characterization of human pharmaceuticals in drug discovery, drug safety assessment and environmental toxicology. However, do fish respond to pharmaceuticals as humans do? To address this question, we provide a novel quantitative cross-species extrapolation approach (qCSE) based on the hypothesis that similar plasma concentrations of pharmaceuticals cause comparable target-mediated effects in both humans and fish at similar level of biological organization (Read-Across Hypothesis). To validate this hypothesis, the behavioural effects of the anti-depressant drug fluoxetine on the fish model fathead minnow (Pimephales promelas) were used as test case. Fish were exposed for 28 days to a range of measured water concentrations of fluoxetine (0.1, 1.0, 8.0, 16, 32, 64 µg/L) to produce plasma concentrations below, equal and above the range of Human Therapeutic Plasma Concentrations (HTPCs). Fluoxetine and its metabolite, norfluoxetine, were quantified in the plasma of individual fish and linked to behavioural anxiety-related endpoints. The minimum drug plasma concentrations that elicited anxiolytic responses in fish were above the upper value of the HTPC range, whereas no effects were observed at plasma concentrations below the HTPCs. In vivo metabolism of fluoxetine in humans and fish was similar, and displayed bi-phasic concentration-dependent kinetics driven by the auto-inhibitory dynamics and saturation of the enzymes that convert fluoxetine into norfluoxetine. The sensitivity of fish to fluoxetine was not so dissimilar from that of patients affected by general anxiety disorders. These results represent the first direct evidence of measured internal dose response effect of a pharmaceutical in fish, hence validating the Read-Across hypothesis applied to fluoxetine. Overall, this study demonstrates that the qCSE approach, anchored to internal drug concentrations, is a powerful tool to guide the

  8. Quantitative Spatiotemporal Chemical Profiling of Individual Lipid Droplets by Hyperspectral CARS Microscopy in Living Human Adipose-Derived Stem Cells.

    PubMed

    Di Napoli, Claudia; Pope, Iestyn; Masia, Francesco; Langbein, Wolfgang; Watson, Pete; Borri, Paola

    2016-04-01

    There is increasing evidence showing that cytosolic lipid droplets, present in all eukaryotic cells, play a key role in many cellular functions. Yet their composition at the individual droplet level and how it evolves over time in living cells is essentially unknown due to the lack of suitable quantitative nondestructive measurement techniques. In this work, we demonstrate the ability of label-free hyperspectral coherent anti-Stokes Raman scattering (CARS) microscopy, together with a quantitative image analysis algorithm developed by us, to quantify the lipid type and content in vol/vol concentration units of individual lipid droplets in living human adipose-derived stem cells during differentiation over 9 days in media supplemented with different fatty acids. Specifically, we investigated the addition of the polyunsaturated linoleic and alpha-linolenic fatty acids into the normal differentiation medium (mostly containing monounsaturated fatty acids). We observe a heterogeneous uptake which is droplet-size dependent, time dependent, and lipid dependent. Cells grown in linoleic-acid-supplemented medium show the largest distribution of lipid content across different droplets at all times during differentiation. When analyzing the average lipid content, we find that adding linoleic or alpha-linolenic fatty acids at day 0 results in uptake of the new lipid components with an exponential time constant of 22 ± 2 h. Conversely, switching lipids at day 3 results in an exponential time constant of 60 ± 5 h. These are unprecedented findings, exemplifying that the quantitative imaging method demonstrated here could open a radically new way of studying and understanding cytosolic lipid droplets in living cells. PMID:26937957

  9. New method to quantitate platelets adhered on biomaterials using monoclonal antibodies to human platelet membrane glycoprotein SZ-21

    SciTech Connect

    Xi, T.F.; Zhang, J.C.; Tian, W.H.; Wang, C.R.; Lei, X.H.; Wai, H.Y.; Ruan, C.G. )

    1990-01-01

    This study developed a new technique to quantitate platelets adhered on biomaterials surfaces in vitro, based on a surface phased radioimmunoassay using a monoclonal antibody SZ-21, directed specifically against the membrane glycoprotein complex IIIa of human platelets. In vitro perfusion is performed in system which consists of testing tubes and infusion pump. After 5 minutes perfusion with fresh ACD anticoagulated human whole blood at 2,000s-1 platelets deposition on surface precoated with proteins determined using anti-human platelet antibody (125 I-SZ-21) are 4,173 +/- 932 (Albumin), 59,032 +/- 25,554 (Fibrinogen), and 71,253 +/- 11,484 (Collagen). Meanwhile, platelets adhered on surfaces of four polymers were determined (platelet/mm2): 19,493 +/- 2,050 (Silicone), 48,193 +/- 4,055 (Polytetrafluoroethylene), 50,375 +/- 8,675 (Polyvinyl chloride) and 101,906 +/- 5,916 (Polyethylene). These results were confirmed by SEM. This method is not only applied for evaluating rapidly and reliably blood compatibility of biomaterials in vitro, but will be used at basic study for interaction of blood materials.

  10. New insights on human T cell development by quantitative T cell receptor gene rearrangement studies and gene expression profiling

    PubMed Central

    Dik, Willem A.; Pike-Overzet, Karin; Weerkamp, Floor; de Ridder, Dick; de Haas, Edwin F.E.; Baert, Miranda R.M.; van der Spek, Peter; Koster, Esther E.L.; Reinders, Marcel J.T.; van Dongen, Jacques J.M.; Langerak, Anton W.; Staal, Frank J.T.

    2005-01-01

    To gain more insight into initiation and regulation of T cell receptor (TCR) gene rearrangement during human T cell development, we analyzed TCR gene rearrangements by quantitative PCR analysis in nine consecutive T cell developmental stages, including CD34+ lin− cord blood cells as a reference. The same stages were used for gene expression profiling using DNA microarrays. We show that TCR loci rearrange in a highly ordered way (TCRD-TCRG-TCRB-TCRA) and that the initiating Dδ2-Dδ3 rearrangement occurs at the most immature CD34+CD38−CD1a− stage. TCRB rearrangement starts at the CD34+CD38+CD1a− stage and complete in-frame TCRB rearrangements were first detected in the immature single positive stage. TCRB rearrangement data together with the PTCRA (pTα) expression pattern show that human TCRβ-selection occurs at the CD34+CD38+CD1a+ stage. By combining the TCR rearrangement data with gene expression data, we identified candidate factors for the initiation/regulation of TCR recombination. Our data demonstrate that a number of key events occur earlier than assumed previously; therefore, human T cell development is much more similar to murine T cell development than reported before. PMID:15928199

  11. Improved HF183 quantitative real-time PCR assay for characterization of human fecal pollution in ambient surface water samples.

    PubMed

    Green, Hyatt C; Haugland, Richard A; Varma, Manju; Millen, Hana T; Borchardt, Mark A; Field, Katharine G; Walters, William A; Knight, R; Sivaganesan, Mano; Kelty, Catherine A; Shanks, Orin C

    2014-05-01

    Quantitative real-time PCR (qPCR) assays that target the human-associated HF183 bacterial cluster within members of the genus Bacteroides are among the most widely used methods for the characterization of human fecal pollution in ambient surface waters. In this study, we show that a current TaqMan HF183 qPCR assay (HF183/BFDrev) routinely forms nonspecific amplification products and introduce a modified TaqMan assay (HF183/BacR287) that alleviates this problem. The performance of each qPCR assay was compared in head-to-head experiments investigating limits of detection, analytical precision, predicted hybridization to 16S rRNA gene sequences from a reference database, and relative marker concentrations in fecal and sewage samples. The performance of the modified HF183/BacR287 assay is equal to or improves upon that of the original HF183/BFDrev assay. In addition, a qPCR chemistry designed to combat amplification inhibition and a multiplexed internal amplification control are included. In light of the expanding use of PCR-based methods that rely on the detection of extremely low concentrations of DNA template, such as qPCR and digital PCR, the new TaqMan HF183/BacR287 assay should provide more accurate estimations of human-derived fecal contaminants in ambient surface waters. PMID:24610857

  12. Quantitative and kinetic profile of Wnt/β-catenin signaling components during human neural progenitor cell differentiation.

    PubMed

    Mazemondet, Orianne; Hubner, Rayk; Frahm, Jana; Koczan, Dirk; Bader, Benjamin M; Weiss, Dieter G; Uhrmacher, Adelinde M; Frech, Moritz J; Rolfs, Arndt; Luo, Jiankai

    2011-12-01

    ReNcell VM is an immortalized human neural progenitor cell line with the ability to differentiate in vitro into astrocytes and neurons, in which the Wnt/β-catenin pathway is known to be involved. However, little is known about kinetic changes of this pathway in human neural progenitor cell differentiation. In the present study, we provide a quantitative profile of Wnt/β-catenin pathway dynamics showing its spatio-temporal regulation during ReNcell VM cell differentiation. We show first that T-cell factor dependent transcription can be activated by stabilized β-catenin. Furthermore, endogenous Wnt ligands, pathway receptors and signaling molecules are temporally controlled, demonstrating changes related to differentiation stages. During the first three hours of differentiation the signaling molecules LRP6, Dvl2 and β-catenin are spatio-temporally regulated between distinct cellular compartments. From 24 h onward, components of the Wnt/β-catenin pathway are strongly activated and regulated as shown by mRNA up-regulation of Wnt ligands (Wnt5a and Wnt7a), receptors including Frizzled-2, -3, -6, -7, and -9, and co-receptors, and target genes including Axin2. This detailed temporal profile of the Wnt/β-catenin pathway is a first step to understand, control and to orientate, in vitro, human neural progenitor cell differentiation. PMID:21805133

  13. Characterization of human metabolism and disposition of levo-tetrahydropalmatine: Qualitative and quantitative determination of oxidative and conjugated metabolites.

    PubMed

    Xiao, Weibin; Shen, Guolin; Zhuang, Xiaomei; Ran, Xiaorong; Zhu, Mingshe; Li, Hua

    2016-09-01

    Levo-tetrahydropalmatine (l-THP) is a tetrahydroprotoberberine isoquinoline alkaloid and has been used as an analgesic agent in China for over 50 years. Recent studies revealed that l-THP was effective in the treatment of drug addiction. However, the plasma metabolic profile, mass balance and clearance pathways of l-THP in human remain unknown. In the present study, an analytical strategy was developed for qualitative and quantitative investigation of metabolism and disposition of l-THP in human. Detection and structural characterization of l-THP metabolites were performed using liquid chromatography-quadrupole time-of-flight mass spectrometry. Selected major metabolites in plasma, urine and feces determined by liquid chromatography with UV detection were further quantified using a triple quadruple mass spectrometry and reference standards. A total of 20 metabolites were identified, most of which were formed via demethylation, mono-hydroxylation, and glucuronidation and sulfonation of desmethyl metabolites. Five major metabolites accounted for over 10% of the parent drug concentration in plasma. Major urinary and fecal metabolites and the parent drug that were monitored for 72h accounted for 46.3% of the dose, while only 0.16% of the dose was the unchanged drug. Multiple demethylations followed by glucuronide and sulfate conjugations and renal excretion were the major drug clearance pathways of l-THP in human. PMID:27343900

  14. Qualitative and quantitative determination of human biomarkers by laser photoacoustic spectroscopy methods

    NASA Astrophysics Data System (ADS)

    Popa, C.; Bratu, A. M.; Matei, C.; Cernat, R.; Popescu, A.; Dumitras, D. C.

    2011-07-01

    The hypothesis that blood, urine and other body fluids and tissues can be sampled and analyzed to produce clinical information for disease diagnosis or therapy monitoring is the basis of modern clinical diagnosis and medical practice. The analysis of breath air has major advantages because it is a non-invasive method, represents minimal risk to personnel collecting the samples and can be often sampled. Breath air samples from the human subjects were collected using aluminized bags from QuinTron and analyzed using the laser photoacoustic spectroscopy (LPAS) technique. LPAS is used to detect traces of ethylene in breath air resulting from lipid peroxidation in lung epithelium following the radiotherapy and also traces of ammonia from patients subjected to hemodialysis for treatment of renal failure. In the case of patients affected by cancer and treated by external radiotherapy, all measurements were done at 10P(14) CO2 laser line, where the ethylene absorption coefficient has the largest value (30.4 cm-1 atm-1), whereas for patients affected by renal failure and treated by standard dialysis, all measurements were performed at 9R(30) CO2 laser line, where the ammonia absorption coefficient has the maximum value of 57 cm-1 atm-1. The levels of ethylene and ammonia in exhaled air, from patients with cancer and renal failure, respectively, were measured and compared with breath air contents from healthy humans. Human gas biomarkers were measured at sub-ppb (parts per billion) concentration sensitivities. It has been demonstrated that LPAS technique will play an important role in the future of exhaled breath air analysis. The key attributes of this technique are sensitivity, selectivity, fast and real time response, as well as its simplicity.

  15. Human in vivo regional intestinal permeability: quantitation using site-specific drug absorption data.

    PubMed

    Sjögren, Erik; Dahlgren, David; Roos, Carl; Lennernäs, Hans

    2015-06-01

    Application of information on regional intestinal permeability has been identified as a key aspect of successful pharmaceutical product development. This study presents the results and evaluation of an approach for the indirect estimation of site-specific in vivo intestinal effective permeability (Peff) in humans. Plasma concentration-time profiles from 15 clinical studies that administered drug solutions to specific intestinal regions were collected and analyzed. The intestinal absorption rate for each drug was acquired by deconvolution, using historical intravenous data as reference, and used with the intestinal surface area and the dose remaining in the lumen to estimate the Peff. Forty-three new Peff values were estimated (15 from the proximal small intestine, 11 from the distal small intestine, and 17 from the large intestine) for 14 active pharmaceutical ingredients representing a wide range of biopharmaceutical properties. A good correlation (r(2) = 0.96, slope = 1.24, intercept = 0.030) was established between these indirect jejunal Peff estimates and jejunal Peff measurements determined directly using the single-pass perfusion double balloon technique. On average, Peff estimates from the distal small intestine and large intestine were 90% and 40%, respectively, of those from the proximal small intestine. These results support the use of the evaluated deconvolution method for indirectly estimating regional intestinal Peff in humans. This study presents the first comprehensive data set of estimated human regional intestinal permeability values for a range of drugs. These biopharmaceutical data can be used to improve the accuracy of gastrointestinal absorption predictions used in drug development decision-making. PMID:25919764

  16. Dopamine Receptors in Human Lymphocytes: Radioligand Binding and Quantitative RT-PCR Assays

    PubMed Central

    Kirillova, Galina P.; Hrutkay, Rebecca J.; Shurin, Michael R.; Shurin, Galina V.; Tourkova, Irina L.; Vanyukov, Michael M.

    2008-01-01

    Analysis of dopamine receptors (DR) in lymphocytes of the human peripheral blood mononuclear cell (PBMC) fraction is an attractive tool for evaluation of functional properties of dopaminergic function underlying variation in complex psychological/psychopathological traits. Receptor binding assays (RBA) with selective radioligands, which are widely used in CNS studies, have not produced consistent results when applied to isolated PBMC. We tested the assay conditions that could be essential for detection of DR in human PBMC and their membrane preparations. Using [3H]SCH23390, a dopamine D1-like receptor antagonist, we demonstrated the presence of two binding sites in PBMC-derived membrane fraction. One of them is characterized by the Kd value consistent with that reported for D5 dopamine receptors in human lymphocytes, whereas the other Kd value possibly corresponds to serotonin receptor(s). Although D5 receptor binding sites in PBMC membranes could be characterized by binding assays, the low protein expression and the large volume of blood needed for membrane preparation render the binding method impracticable for individual phenotyping. In contrast, real-time RT-PCR may be used for this purpose, contingent on the relationship between DR expression in the brain and in lymphocytes. The expression of the DRD2-DRD5 genes, as detected by this method, varied widely among samples, whereas the DRD1 expression was not detected. The expression levels were comparable with those in the brain for DRD3 and DRD4, and were significantly lower for DRD2 and DRD5. PMID:18721826

  17. Datasets from label-free quantitative proteomic analysis of human glomeruli with sclerotic lesions

    PubMed Central

    Zhang, Ying; Xu, Bo; Kinoshita, Naohiko; Yoshida, Yutaka; Tasaki, Masayuki; Fujinaka, Hidehiko; Magdeldin, Sameh; Yaoita, Eishin; Yamamoto, Tadashi

    2015-01-01

    Human glomeruli with intermediate (i-GS) and advanced (GS) sclerotic lesions as well as the normal control (Nor) were captured from laser microdissection, digested by trypsin and subjected to shotgun LC-MS/MS analysis (LTQ-Orbitrap XL). The label-free quantification was performed using the Normalized Spectral Index (SIN) to assess the relative molar concentration of each protein identified in a sample. All the experimental data are shown in this article. The data is associated to the research article submitted to Journal of Proteomics [1]. PMID:26217785

  18. Separation and quantitation of polyethylene glycols 400 and 3350 from human urine by high-performance liquid chromatography.

    PubMed

    Ryan, C M; Yarmush, M L; Tompkins, R G

    1992-04-01

    Polyethylene glycol 3350 (PEG 3350) is useful as an orally administered probe to measure in vivo intestinal permeability to macromolecules. Previous methods to detect polyethylene glycol (PEG) excreted in the urine have been hampered by inherent inaccuracies associated with liquid-liquid extraction and turbidimetric analysis. For accurate quantitation by previous methods, radioactive labels were required. This paper describes a method to separate and quantitate PEG 3350 and PEG 400 in human urine that is independent of radioactive labels and is accurate in clinical practice. The method uses sized regenerated cellulose membranes and mixed ion-exchange resin for sample preparation and high-performance liquid chromatography with refractive index detection for analysis. The 24-h excretion for normal individuals after an oral dose of 40 g of PEG 3350 and 5 g of PEG 400 was 0.12 +/- 0.04% of the original dose of PEG 3350 and 26.3 +/- 5.1% of the original dose of PEG 400. PMID:1501072

  19. Three-Dimensional Neuroepithelial Culture from Human Embryonic Stem Cells and Its Use for Quantitative Conversion to Retinal Pigment Epithelium

    PubMed Central

    Zhu, Yu; Carido, Madalena; Meinhardt, Andrea; Kurth, Thomas; Karl, Mike O.; Ader, Marius; Tanaka, Elly M.

    2013-01-01

    A goal in human embryonic stem cell (hESC) research is the faithful differentiation to given cell types such as neural lineages. During embryonic development, a basement membrane surrounds the neural plate that forms a tight, apico-basolaterally polarized epithelium before closing to form a neural tube with a single lumen. Here we show that the three-dimensional epithelial cyst culture of hESCs in Matrigel combined with neural induction results in a quantitative conversion into neuroepithelial cysts containing a single lumen. Cells attain a defined neuroepithelial identity by 5 days. The neuroepithelial cysts naturally generate retinal epithelium, in part due to IGF-1/insulin signaling. We demonstrate the utility of this epithelial culture approach by achieving a quantitative production of retinal pigment epithelial (RPE) cells from hESCs within 30 days. Direct transplantation of this RPE into a rat model of retinal degeneration without any selection or expansion of the cells results in the formation of a donor-derived RPE monolayer that rescues photoreceptor cells. The cyst method for neuroepithelial differentiation of pluripotent stem cells is not only of importance for RPE generation but will also be relevant to the production of other neuronal cell types and for reconstituting complex patterning events from three-dimensional neuroepithelia. PMID:23358448

  20. Direct and rapid quantitation of ephedrines in human urine by paper spray ionization/high resolution mass spectrometry.

    PubMed

    Jeong, Eun Sook; Kim, Ki Hun; Cha, Eunju; Kwon, Oh-Seung; Cha, Sangwon; Lee, Jaeick

    2016-08-15

    A rapid and direct paper spray ionization/mass spectrometry (PSI/MS) method was developed for quantitative analysis of ephedrine, pseudoephedrine, norpseudoephedrine, and methylephedrine in human urine. This method involves the use of a triangular filter paper and high-resolution mass spectrometry, where the molecular ions of ephedrines were generated by applying high voltage after loading the spray solvent to the paper which urine sample was pre-loaded. Small amounts (2μL) of urine spiked with an internal standard were directly analyzed for ephedrines. The PSI/MS method was validated for linearity, within- and between-run precision, accuracy, and limit of detection. The results showed good linearity (R(2)≥0.9928) and acceptable precision and accuracy. Furthermore, the accuracy of the method was assessed by analyzing a blind urine sample from World Anti-Doping Agency and comparing the measured concentrations with the nominal concentrations. This test resulted in accuracies ranging from 96.4 to 106.1%, indicating that the PSI/MS method has the potential to be an alternative technique for the fast quantitation of ephedrines in doping control analysis. PMID:27393909

  1. Nested-PCR and TaqMan real-time quantitative PCR assays for human adenoviruses in environmental waters.

    PubMed

    Huang, Wen-Chien; Chou, Yi-Pen; Kao, Po-Min; Hsu, Tsui-Kang; Su, Hung-Chang; Ho, Ying-Ning; Yang, Yi-Chun; Hsu, Bing-Mu

    2016-01-01

    Human adenovirus (HAdV) infections can occur throughout the year. Cases of HAdV-associated respiratory disease have been more common in the late winter, spring, and early summer. In this study, to provide viral pollution data for further epidemiological studies and governmental actions, the presence of HAdV in the aquatic environment was quantitatively surveyed in the summer. This study was conducted to compare the efficiencies of nested-PCR (polymerase chain reaction) and qPCR (quantitative PCR) for detecting HAdV in environmental waters. A total of 73 water samples were collected from Puzi River in Taiwan and subjected to virus concentration methods. In the results, qPCR had much better efficiency for specifying the pathogen in river sample. HAdV41 was detected most frequently in the river water sample (10.9%). The estimated HAdV concentrations ranged between 6.75 × 10(2) and 2.04 × 10(9) genome copies/L. Significant difference was also found in heterotrophic plate counts, conductivity, water temperature, and water turbidity between presence/absence of HAdV. HAdV in the Puzi River may pose a significant health risk. PMID:27120637

  2. Mapping of HPV transcripts in four human cervical lesions using RNAseq suggests quantitative rearrangements during carcinogenic progression.

    PubMed

    Chen, Jinmiao; Xue, Yuezhen; Poidinger, Michael; Lim, Thimothy; Chew, Sung Hock; Pang, Chai Ling; Abastado, Jean-Pierre; Thierry, Françoise

    2014-08-01

    Two classes of Human papillomaviruses (HPV) infect the anogenital track: high risk viruses that are associated with risk of cervical cancer and low risk types that drive development of benign lesions, such as condylomas. In the present study, we established quantitative transcriptional maps of the viral genome in clinical lesions associated with high risk HPV16 or low risk HPV6b. Marked qualitative and quantitative changes in the HPV16 transcriptome were associated with progression from low to high grade lesions. Specific transcripts encoding essential regulatory proteins such as E7, E2, E1^E4 and E5 were identified. We also identified intrinsic differences between the HPV6b-associated condyloma transcript map and that of the HPV16-associated low grade CIN specifically regarding promoter usage. Characterization and quantification of HPV transcripts in patient samples thus establish the impact of viral transcriptional regulation on the status of HPV-associated lesions and may therefore help in defining new biologically-relevant prognosis markers. PMID:25092457

  3. Differential permeability and quantitative MR imaging of a human lung carcinoma brain xenograft in the nude rat.

    PubMed Central

    Barnett, P. A.; Roman-Goldstein, S.; Ramsey, F.; McCormick, C. I.; Sexton, G.; Szumowski, J.; Neuwelt, E. A.

    1995-01-01

    This study characterized agent differential permeability, three-dimensional tumor volume, and survival in an LX-1 human small cell lung carcinoma intracerebral xenograft model in the nude rat. The percent accessible tissue space (distribution volume) and the permeability x capillary surface product for aminoisobutyric acid (M(r) 103), methotrexate (M(r) 454), dextran 10 (M(r) 10,000), and dextran 70 (M(r) 70,000) were measured between 8 and 16 days after inoculation of tumor. Magnetic resonance imaging and histology were used to quantitate intracerebral tumor volume (mm3). Accessible tissue space (ml/g) and permeability x capillary surface product in intracranial tumor, surrounding brain, and subcutaneous tumor decreased with increasing molecular weight of the agent, regardless of the number of days after inoculation. Accessible tissue space in intracranial tumor increased between 8 and 16 days for all agents except dextran 70. There was little change in the subcutaneous tumor or other tissues with time. Tumor volume calculations from imaging studies correlated with volumetric measurements from histological sections (r2 = 98.5%) and illustrated natural tumor progression (9 to 225 mm3). These results provide a basis for therapeutic design based on differential permeability of specific agents and the ability to quantitatively measure brain tumor volume for accessing tumor response. Images Figure 4 PMID:7856753

  4. Determination of permethrin resistance allele frequency of human head louse populations by quantitative sequencing.

    PubMed

    Kwon, Deok Ho; Yoon, Kyong Sup; Strycharz, Joseph P; Clark, J Marshall; Lee, Si Hyeock

    2008-09-01

    A quantitative sequencing (QS) protocol that detects the frequencies of sodium channel mutations (M815I, T917I, and L920F) responsible for knockdown resistance in permethrin-resistant head lice (Pediculus humanus capitis De Geer) was tested as a population genotyping method for use as a preliminary resistance monitoring tool. Genomic DNA fragments of the sodium channel a-subunit gene that encompass the three mutation sites were polymerase chain reaction (PCR)-1 amplified from individual head lice with either resistant or susceptible genotypes, and combined in various ratios to generate standard DNA template mixtures for QS. After sequencing, the signal ratios between resistant and susceptible nucleotides were calculated and plotted against the corresponding resistance allele frequencies. Quadratic regression coefficients of the plots were close to 1, demonstrating that the signal ratios are highly correlated with the resistance allele frequencies. Resistance allele frequencies predicted by QS, using either "pooled DNA" (DNA extracted from individual louse specimens and pooled) or "pooled specimen DNA" (DNA simultaneously extracted from multiple louse specimens), agreed well with those determined by individual sequencing, confirming the reliability and accuracy of QS as a population genotyping method and validating our approach of using the pooled specimen DNA as the DNA template for QS. Our protocol for QS was determined to be highly reliable for the prediction of resistance allele frequencies higher than approximately 7.4% at the 95% confidence level. According to the resistance allele frequencies determined by QS, pyrethroid resistance varies substantially among different geographical regions, emphasizing the importance of early resistance detection and proper management strategies. PMID:18826035

  5. Quantitative Percussion Diagnostics and Bone Density Analysis of the Implant-Bone Interface in a Pre- and Postmortem Human Subject

    PubMed Central

    Hui, DiDi; Bajaj, Vaibhav; Earthman, James C.

    2014-01-01

    Purpose It has been hypothesized that a correlation exists between the density of surrounding cortical bone and the stability of an implant under percussion loading that can be used to quantify the implant's osseointegration. The purpose of the present research was to explore whether quantitative percussion testing of dental implants gives reasonable indications of the level of osseointegration that are consistent with bone configuration and its influence on osseointegration quality. Material and Methods Data from percussion testing of a live human subject, obtained using the Periometer®, were compared with corresponding bone density estimates from high-resolution computer tomography images and postmortem percussion probe data. Results The results confirm the hypothesis that the nature of an implant’s response to percussion is determined by its cortical bone support. Conclusions The findings suggest that the cortical bone supporting the crestal and apical regions of the implant is primarily responsible for structural stability. PMID:24278927

  6. Age, gender, and hemispheric differences in human striatum: a quantitative review and new data from in vivo MRI morphometry.

    PubMed

    Raz, N; Torres, I J; Acker, J D

    1995-03-01

    We reviewed postmortem and neuroimaging studies of striatal neuroanatomy in humans. The quantitative review revealed evidence of moderate age-related shrinkage of the caudate nucleus and the putamen and consistent leftward asymmetry of the globus pallidus. The data on sex differences were very sparse. We examined neostriatal structures in two samples (healthy volunteers and patients with negative radiological findings) using in vivo magnetic resonance imaging. In both samples, bilateral age-related shrinkage of the caudate nucleus was found, although among the patients the effect was significant only for males. The putamen was measured only in the second sample, and age-related reduction in its volume was found also only among males. A trend for rightward asymmetry in the volume of caudate nucleus was observed in both samples, although it reached statistical significance only among the patients. Putative pathological and physiological mechanisms underlying the observed differences in teh neostriatum are discussed. PMID:7663886

  7. A quantitative immunohistochemical study on the time-dependent course of acute inflammatory cellular response to human brain injury.

    PubMed

    Hausmann, R; Kaiser, A; Lang, C; Bohnert, M; Betz, P

    1999-01-01

    The time-dependent inflammatory cell reaction in human cortical contusions has been investigated during the first 30 weeks after blunt head injury. Immunohistochemical staining was carried out using CD 15 for granulocytes and LCA, CD 3 and UCHL-1 for mononuclear leucocytes. In order to provide reliable data for a forensic wound age estimation, the intensity of the cellular reaction was evaluated with a quantitative image analysis system. CD 15-labelled granulocytes were detectable earliest 10 min after brain injury, whereas significantly increased numbers of mononuclear leucocytes occurred in cortical contusions after a postinfliction interval of at least 1.1 days (LCA), 2 days (CD 3) or 3.7 days (UCHL-1), respectively. PMID:10433032

  8. A Quantitative Proteomic Approach for Detecting Protein Profiles of Activated Human Myeloid Dendritic Cells

    PubMed Central

    Schlatzer, Daniela M; Sugalski, Julia; Dazard, Jean-Eudes; Chance, Mark R; Anthony, Donald D.

    2011-01-01

    Dendritic cells (DC) direct the magnitude, polarity and effector function of the adaptive immune response. DC express toll-like receptors (TLR), antigen capturing and processing machinery, and costimulatory molecules, which facilitate innate sensing and T cell activation. Once activated, DC can efficiently migrate to lymphoid tissue and prime T cell responses. Therefore, DC play an integral role as mediators of the immune response to multiple pathogens. Elucidating the molecular mechanisms involved in DC activation is therefore central in gaining an understanding of host response to infection. Unfortunately, technical constraints have limited system-wide ‘omic’ analysis of human DC subsets collected ex vivo. Here we have applied novel proteomic approaches to human myeloid dendritic cells (mDCs) purified from 100 milliliters of peripheral blood to characterize specific molecular networks of cell activation at the individual patient level, and have successfully quantified over 700 proteins from individual samples containing as little as 200,000 mDCs. The proteomic and network readouts after ex vivo stimulation of mDCs with TLR3 agonists is measured and verified using flow cytometry. PMID:21945394

  9. Liposome-based immunoaffinity chromatographic assay for the quantitation of immunoglobulin E in human serum.

    PubMed

    Annie Ho, Ja-an; Wu, Li-Chen; Chang, Li-Hui; Hwang, Kuo-Chu; Reuben Hwu, Jih-Ru

    2010-01-15

    Immunoglobulin E (IgE)-mediated type I allergies affect over 25% of the world's population; they are among the most common diseases in developed countries. Therefore, simple and rapid in vivo and in vitro methods for diagnosing allergies are becoming increasingly important. In this paper, we demonstrate the feasibility of using sulforhodamine B, a fluorescent dye, entrapped inside immunoliposomes, the outer surfaces of which were sensitized with IgE, as a signal amplifier for the development of a simple, rapid, and inexpensive colorimetric affinity chromatographic immunoassay for the detection of total IgE in serum. This assay operates based on competition between standards (or human serum samples) containing IgE and IgE-sensitized immunoliposomes for the limited number of antigen binding sites of immobilized anti-IgE antibodies at the antigen capture (AC) zone on the nitrocellulose membranes. The color density of the AC zone is indirectly proportional to the number of IgE units present in the test sample. The detection limit of this liposome-based immunoaffinity chromatographic assay was 0.37ng in IgE-free serum solution (equivalent to 20microL of a 18.5ngmL(-1) solution). A commercially available ELISA kit was used as a reference method to validate the proposed assay through the analysis of three human serum samples. PMID:19683481

  10. Quantitative Gene Expression of ERG9 in Model Saccharomyces cerevisiae: Chamomile Extract For Human Cancer Treatment

    PubMed Central

    Hosseinpour, Maryam; Mobini-Dehkordi, Mohsen

    2016-01-01

    Introduction Over expression of squalene synthase gene causes induction of growth tumour and reduction of apoptosis. This gene which is conserved between Saccharomyces cerevisiae yeast and humans, is named (ERG9). Aim In this work, we studied the effect of Matricaria recutita extract on ERG9 gene (squalene synthase) expression in S.cerevisiae which was used as organism model in cancer therapy. Materials and Methods S. cerevisiae was cultured in YPD medium plus 0,250, 1000 and 3000 μg/ml of Matricaria recutita extract and we evaluated the (ERG9) gene expression by Real-time RT-PCR method after 24 hours. Statistical analysis used At least 3 independent experiments were done. Data were analyzed using One-way ANOVA and Dunnett’s test. A p-value of less than 0.01 was considered as significant. Results We found that 250, 1000 and 3000 μg/ml of Matricaria recutita extract could reduce expression of ERG9 gene significantly (p<0.01). Interestingly, the expression of this gene was completely inhibited in 1000 and 3000 μg/ml concentrations. Conclusion This study predicted that Matricaria recutita extract produced anti-cancer effects in humans, because it could inhibit the expression of an analogue key gene in this malignant disease. Further investigations should be made, to study its molecular mechanism of action at the mammal cell level.

  11. Quantitative evaluation of the kinetics of human enamel simulated caries using photothermal radiometry and modulated luminescence.

    PubMed

    Hellen, Adam; Mandelis, Andreas; Finer, Yoav; Amaechi, Bennett T

    2011-07-01

    Photothermal radiometry and modulated luminescence (PTR-LUM) is an emerging nondestructive methodology applied toward the characterization and quantification of dental caries. We evaluate the efficacy of PTR-LUM in vitro to detect, monitor, and quantify human enamel caries. Artificial caries are created in extracted human molars (n = 15) using an acidified gel system (pH 4.5) for 10 or 40 days. PTR-LUM frequency scans (1 Hz-1 kHz) are performed before and during demineralization. Transverse microradiography (TMR) analysis, the current gold standard, follows at treatment conclusion to determine the mineral loss and depth of the artificially demineralized lesions. A theoretical model is applied to PTR experimental data to evaluate the changes in optothermophysical properties of demineralized enamel as a function of time. Higher optical scattering coefficients and poorer thermophysical properties are characteristic of the growing demineralized lesions, as verified by TMR, where the generated microporosities of the subsurface lesion confine the thermal-wave centroid. Enhanced optical scattering coefficients of demineralized lesions result in poorer luminescence yield due to scattering of both incident and converted luminescent photons. PTR-LUM sensitivity to changes in tooth mineralization coupled with opto-thermophysical property extraction illustrates the technique's potential for nondestructive quantification of enamel caries. PMID:21806252

  12. Quantitative Proteomics Reveals a Role for Epigenetic Reprogramming During Human Monocyte Differentiation*

    PubMed Central

    Nicholas, Dequina; Tang, Hui; Zhang, Qiongyi; Rudra, Jai; Xu, Feng; Langridge, William; Zhang, Kangling

    2015-01-01

    The differentiation of monocytes into macrophages and dendritic cells involves mechanisms for activation of the innate immune system in response to inflammatory stimuli, such as pathogen infection and environmental cues. Epigenetic reprogramming is thought to play an important role during monocyte differentiation. Complementary to cell surface markers, the characterization of monocytic cell lineages by mass spectrometry based protein/histone expression profiling opens a new avenue for studying immune cell differentiation. Here, we report the application of mass spectrometry and bioinformatics to identify changes in human monocytes during their differentiation into macrophages and dendritic cells. Our data show that linker histone H1 proteins are significantly down-regulated during monocyte differentiation. Although highly enriched H3K9-methyl/S10-phos/K14-acetyl tri-modification forms of histone H3 were identified in monocytes and macrophages, they were dramatically reduced in dendritic cells. In contrast, histone H4 K16 acetylation was found to be markedly higher in dendritic cells than in monocytes and macrophages. We also found that global hyperacetylation generated by the nonspecific histone deacetylase HDAC inhibitor Apicidin induces monocyte differentiation. Together, our data suggest that specific regulation of inter- and intra-histone modifications including H3 K9 methylation, H3 S10 phosphorylation, H3 K14 acetylation, and H4 K16 acetylation must occur in concert with chromatin remodeling by linker histones for cell cycle progression and differentiation of human myeloid cells into macrophages and dendritic cells. PMID:25316709

  13. Quantitative proteomics reveals a role for epigenetic reprogramming during human monocyte differentiation.

    PubMed

    Nicholas, Dequina; Tang, Hui; Zhang, Qiongyi; Rudra, Jai; Xu, Feng; Langridge, William; Zhang, Kangling

    2015-01-01

    The differentiation of monocytes into macrophages and dendritic cells involves mechanisms for activation of the innate immune system in response to inflammatory stimuli, such as pathogen infection and environmental cues. Epigenetic reprogramming is thought to play an important role during monocyte differentiation. Complementary to cell surface markers, the characterization of monocytic cell lineages by mass spectrometry based protein/histone expression profiling opens a new avenue for studying immune cell differentiation. Here, we report the application of mass spectrometry and bioinformatics to identify changes in human monocytes during their differentiation into macrophages and dendritic cells. Our data show that linker histone H1 proteins are significantly down-regulated during monocyte differentiation. Although highly enriched H3K9-methyl/S10-phos/K14-acetyl tri-modification forms of histone H3 were identified in monocytes and macrophages, they were dramatically reduced in dendritic cells. In contrast, histone H4 K16 acetylation was found to be markedly higher in dendritic cells than in monocytes and macrophages. We also found that global hyperacetylation generated by the nonspecific histone deacetylase HDAC inhibitor Apicidin induces monocyte differentiation. Together, our data suggest that specific regulation of inter- and intra-histone modifications including H3 K9 methylation, H3 S10 phosphorylation, H3 K14 acetylation, and H4 K16 acetylation must occur in concert with chromatin remodeling by linker histones for cell cycle progression and differentiation of human myeloid cells into macrophages and dendritic cells. PMID:25316709

  14. Quantitative changes in human epithelial cancers and osteogenesis imperfecta disease detected using nonlinear multicontrast microscopy

    NASA Astrophysics Data System (ADS)

    Adur, Javier; Pelegati, Vitor B.; de Thomaz, Andre A.; D'Souza-Li, Lilia; Assunção, Maria do Carmo; Bottcher-Luiz, Fátima; Andrade, Liliana A. L. A.; Cesar, Carlos L.

    2012-08-01

    We show that combined multimodal nonlinear optical (NLO) microscopies, including two-photon excitation fluorescence, second-harmonic generation (SHG), third harmonic generation, and fluorescence lifetime imaging microscopy (FLIM) can be used to detect morphological and metabolic changes associated with stroma and epithelial transformation during the progression of cancer and osteogenesis imperfecta (OI) disease. NLO microscopes provide complementary information about tissue microstructure, showing distinctive patterns for different types of human breast cancer, mucinous ovarian tumors, and skin dermis of patients with OI. Using a set of scoring methods (anisotropy, correlation, uniformity, entropy, and lifetime components), we found significant differences in the content, distribution and organization of collagen fibrils in the stroma of breast and ovary as well as in the dermis of skin. We suggest that our results provide a framework for using NLO techniques as a clinical diagnostic tool for human cancer and OI. We further suggest that the SHG and FLIM metrics described could be applied to other connective or epithelial tissue disorders that are characterized by abnormal cells proliferation and collagen assembly.

  15. Quantitative evaluation of the kinetics of human enamel simulated caries using photothermal radiometry and modulated luminescence

    NASA Astrophysics Data System (ADS)

    Hellen, Adam; Mandelis, Andreas; Finer, Yoav; Amaechi, Bennett T.

    2011-07-01

    Photothermal radiometry and modulated luminescence (PTR-LUM) is an emerging nondestructive methodology applied toward the characterization and quantification of dental caries. We evaluate the efficacy of PTR-LUM in vitro to detect, monitor, and quantify human enamel caries. Artificial caries are created in extracted human molars (n = 15) using an acidified gel system (pH 4.5) for 10 or 40 days. PTR-LUM frequency scans (1 Hz-1 kHz) are performed before and during demineralization. Transverse microradiography (TMR) analysis, the current gold standard, follows at treatment conclusion to determine the mineral loss and depth of the artificially demineralized lesions. A theoretical model is applied to PTR experimental data to evaluate the changes in optothermophysical properties of demineralized enamel as a function of time. Higher optical scattering coefficients and poorer thermophysical properties are characteristic of the growing demineralized lesions, as verified by TMR, where the generated microporosities of the subsurface lesion confine the thermal-wave centroid. Enhanced optical scattering coefficients of demineralized lesions result in poorer luminescence yield due to scattering of both incident and converted luminescent photons. PTR-LUM sensitivity to changes in tooth mineralization coupled with opto-thermophysical property extraction illustrates the technique's potential for nondestructive quantification of enamel caries.

  16. Demonstration and quantitation of phosphoglycolate in human red cells: a potential regulator of 2,3-DPG metabolism

    SciTech Connect

    Spear, D.; Vora, S.

    1986-05-01

    Phosphoglycolate (G-2-P), an important intermediate in the photorespiratory carbon oxidation cycle in plants, is a potent activator of 2,3-DPG phosphatase (DPGP), the enzyme that degrades 2,3-DPG. This has led to the speculation regarding its role in the regulation of red cell 2,3-DPG level, although its existence in red cells has been controversial. Therefore, the authors set out to isolate and quantitate G-2-P, if any, from human red cells. They purified endogenous G-2-P by Dowex-1-Cl/sup -/ chromatography of perchloric acid extracts of fresh, heparinized blood using (/sup 14/C)-G-2-P (2.5 nmoles) as an internal standard of recovery. The putative purified compound and appropriate controls were assayed for G-2-P using (a) a spectrophotometric assay that measured the rate of degradation of 2,3-DPG by DPGP in the presence of G-2-P, linked to the oxidation of NADH via phosphoglycerate kinase and glyceraldehyde-3-P dehydrogenase; and (b) a radiometric assay in which (/sup 32/Pi) is released quantitatively from (/sup 32/P)-2,3-DPG by DPGP in the presence of G-2-P. The lowest limits of detection of these assays are 3 nmoles and 50 pmoles of G-2-P respectively. The results by both assays consistently demonstrated the presence of an average 1.3 ..mu..M (range 0.7-2.7 ..mu..M) G-2-P in red cells from several different individuals. These results confirm those of Rose and Salon and implicate G-2-P in the regulation of red cell 2,3-DPG level. The investigations of the metabolism of G-2-P and its biosynthesis are in progress to define the role(s) of the compound, if any, in human physiology.

  17. A novel uHPLC-MS/MS method for the quantitation of AZD7451 (AZ12607092) in human plasma.

    PubMed

    Peer, Cody J; Brown, Jeffrey L; Martin, Timothy J; Roth, Jeffrey; Spencer, Shawn D; Brassil, Patrick; McNeill, Katharine A; Kreisl, Teri N; Fine, Howard A; Figg, William D

    2013-12-30

    Tropomyosin-related kinases (Trk) are tyrosine kinase receptors implicated in tumor proliferation, invasion, and survival signaling across a number of tumors, making them potentially attractive targets