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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. Determination of human chorionic gonadotropin.

    PubMed

    Stenman, Ulf-Håkan; Alfthan, Henrik

    2013-12-01

    Determination of human chorionic gonadotropin (hCG) is used for diagnosis and monitoring of pregnancy, pregnancy related disorders, for trophoblastic and some nontrophoblastic tumors. In addition, hCG is determined for doping control in males. Assay of hCG is complicated by the occurrence of different molecular forms, which are detected to various degrees by different assays. The main form of hCG in circulation and in patients with trophoblastic tumors is intact heterodimeric hCG. The free β subunit (hCGβ) is a minor form in plasma in both conditions, but it may be the major form aggressive trophoblastic cancer. Therefore, assays measuring hCG and hCGβ together are mainly used for diagnosis of pregnancy and trophoblastic diseases. When excreted into urine, most of hCG (and hCGβ) is broken down to the core fragment of hCGβ (hCGβcf), which is the main immunoreactive form of hCG in urine during pregnancy. Specific determination of hCGβ is of value in screening for Down's syndrome and diagnosis of nontrophoblastic cancer. hCGbcf is of limited utility but it is important because it may disturb assay of hCG in pregnancy.

  3. Suppression of mixed lymphocyte reactivity by human chorionic gonadotrophin

    PubMed Central

    Beling, C. G.; Weksler, M. E.

    1974-01-01

    Highly purified human chorionic gonadotrophin inhibits the response of lymphocytes from both male and female subjects to allogeneic cells in mixed lymphocyte culture. Human chorionic gonadotrophin is not cytotoxic for human lymphocytes. PMID:4283122

  4. Paradoxical consequence of human chorionic gonadotropin misuse.

    PubMed

    Pektezel, Mehmet Yasir; Bas, Demet Funda; Topcuoglu, Mehmet Akif; Arsava, Ethem Murat

    2015-01-01

    Recombinant human chorionic gonadotropin (hCG) is commonly misused as a weight reducing or performance enhancing agent but is associated with increased risk of thromboembolic events. A 29-year-old female with a history of obesity was admitted to our center with a diagnosis of ischemic stroke. Etiologic workup revealed a large patent foramen ovale and history of recent use of hCG as part of a weight loss regimen. This report highlights the potential complications of hCG therapy, particularly when used for unapproved indications and without medical supervision.

  5. Withholding gonadotropins until human chorionic gonadotropin administration.

    PubMed

    Abdallah, Rony; Kligman, Isaac; Davis, Owen; Rosenwaks, Zev

    2010-11-01

    Withholding gonadotropins in women who exhibit high estradiol responses before follicles reach full maturation is called "coasting." Coasting, or suspending gonadotropin administration, can be an effective strategy for decreasing the risk of ovarian hyperstimulation syndrome (OHSS) while reducing cancelation rates. In in vitro fertilization cycles, mechanistically it is believed that withholding gonadotropins starves smaller follicles, induces apoptosis, and decreases the potential for these follicles to elaborate vascular endothelial growth factor, a known mediator of OHSS. It is generally accepted that coasting should be initiated when the estradiol (E₂) level is >3000 pg/mL in the setting of immature follicles. The human chorionic gonadotropin (hCG) trigger should be administered when the E₂ level subsequently drops to a "safe" level. Cycle cancellation should be considered if, after 3 to 4 days of coasting, the E₂ level remains excessively elevated. Oocyte retrieval may also be cancelled if the E₂ level on the day after hCG trigger drops precipitously. In gonadotropin-releasing hormone agonist (GnRHa)-based protocols, one can consider withholding GnRHa administration if the E₂ level continues to increase after a few days of coasting. Current data seem to show that the coasting period is short and/or is less likely to be required in GnRH-antagonist protocols as compared with GnRHa-based protocols. Large randomized control trials are still needed to establish the relative efficacy of coasting versus embryo cryopreservation in the context of OHSS prevention.

  6. Rapid one step urine test for human chorionic gonadotrophin in evaluating suspected complications of early pregnancy.

    PubMed Central

    Kingdom, J C; Kelly, T; MacLean, A B; McAllister, E J

    1991-01-01

    OBJECTIVE--To determine the ability of a sensitive one step urine test to detect human chorionic gonadotrophin in women with suspected complications of early pregnancy. DESIGN--Test on women presenting to accident and emergency department with gynaecological problems over six months. Results were validated using a quantitative assay for human chorionic gonadotrophin in serum and urine. SETTING--Accident and emergency department and gynaecology wards of a university teaching hospital. SUBJECTS--130 unselected women. MAIN OUTCOME MEASURES--Detection of human chorionic gonadotrophin by one step test, presence of ectopic pregnancy, and results of quantitative analysis of chorionic gonadotrophin in serum and urine. RESULTS--79 women had a positive urine test result and 51 a negative result. All 12 women with ectopic pregnancy had a positive test result, although urinary concentration varied from 191 IU/l to 47,800 IU/l. Only one woman, who had a faintly positive result, was found not to be pregnant on subsequent examination. The sensitivity and negative predictive values of the urine test were 100% respectively. 33 women were sent home from the accident and emergency department with normal clinical findings after a negative urine test result. All these women had undetectable concentrations of chorionic gonadotrophin in matched samples of urine and serum. CONCLUSIONS--A simple, rapid one step test for chorionic gonadotrophin should be available for the initial evaluation of emergency gynaecological problems. The additional cost of the test is offset by not admitting those patients whose clinical findings are normal and who have a negative urine test result and by reducing the number of women requiring quantitative assays of chorionic gonadotrophin. Images FIG 1 PMID:2059687

  7. 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.

  8. 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...

  9. Human chorionic gonadotropin and CA 15-3 producing adenocarcinoma.

    PubMed

    Uçkaya, G; Ozet, A; Arpaci, A; Kömürcü, S

    1998-01-01

    50 years old man suffering from primary lung adenocarcinoma presented with high levels of both beta subunit human chorionic gonadotropin (beta HCG) and cancer antigen 15-3 (CA 15-3) in the absence of elevated carcinoembrionic antigen (CEA), alfa fetoprotein (AFP) and carbohydrate antigen 19-9 (CA 19-9). Although beta HCG or CA 15-3 high levels were reported in adenocarcinoma of lung, this is the first report of a patient with high levels of both markers.

  10. 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.

  11. 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... Test Systems § 862.1155 Human chorionic gonadotropin (HCG) test system. (a) Human...

  12. 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.

  13. Potential Therapy for Neisseria Gonorrhoeae Infections With Human Chorionic Gonadotropin.

    PubMed

    Rao, C V

    2015-12-01

    The scientific evidence suggests that Neisseria gonorrhoeae (NG) infects human fallopian tubes by molecular mimicry in which pathogens act like a ligand to bind to epithelial cell surface human chorionic gonadotrophin (hCG)/luteinizing hormone (LH) receptors. The hCG-like molecule has been identified as ribosomal protein L12 in NG coat surface. Human fallopian tube epithelial cells have been shown to contain functional hCG/LH receptors. As previously shown in human fallopian tube organ and cell culture studies, cellular invasion and infection can be prevented by exposing the cells to excess hCG, which would outnumber and outcompete NG for receptor binding. Based on these data, we suggest testing hCG in clinical trials on infected women.

  14. Syndecan expressions in the human amnion and chorionic plate.

    PubMed

    Lorenzi, T; Turi, A; Crescimanno, C; Morroni, M; Castellucci, M; David, G; Tranquilli, A L; Marzioni, D

    2010-10-27

    The syndecan family consists of four distinct membrane glycoproteins in mammals. Syndecans control cell proliferation, differentiation, adhesion and migration through participation in cell-cell interactions, anchorage of cells to the extracellular environment, and modulation of multiple growth factors. Therefore, syndecans may play a pivotal role in the regulation of cell behaviour depending on the cellular microenvironment. Here, we demonstrate that syndecan-1, syndecan-2 and syndecan-4 are expressed in fetal membrane tissue with different immunolocalizations. Syndecan-1 is expressed in the amniotic epithelium, localizing at basolateral cell surfaces. Syndecan-2 and syndecan-4, in contrast, are mostly localized in intracellular compartments, in the extravillous cytotrophoblastic cells and in some fibroblasts of the chorionic plate as well as in the amniotic epithelial cells. In the latter, syndecan-4 is mainly localized in the apical part of the cells. Our results strongly suggest a key role of syndecan-1, syndecan-2 and syndecan-4 in the determination of structural and functional characteristics of human amnion and chorionic plate. Since the solute exchanges between fetus and mother take place in fetal membranes, our data suggest that syndecans are important players in the placenta for the establishment of the fetal-maternal inter-communication.

  15. Luteinizing hormone and human chorionic gonadotropin: origins of difference.

    PubMed

    Choi, Janet; Smitz, Johan

    2014-03-05

    Luteinizing hormone (LH) and human chorionic gonadotropin (hCG) are widely recognized for their roles in ovulation and the support of early pregnancy. Aside from the timing of expression, however, the differences between LH and hCG have largely been overlooked in the clinical realm because of their similar molecular structures and shared receptor. With technologic advancements, including the development of highly purified and recombinant gonadotropins, researchers now appreciate that these hormones are not as interchangeable as once believed. Although they bind to a common receptor, emerging evidence suggests that LH and hCG have disparate effects on downstream signaling cascades. Increased understanding of the inherent differences between LH and hCG will foster more effective diagnostic and prognostic assays for use in a variety of clinical contexts and support the individualization of treatment strategies for conditions such as infertility.

  16. Production of human chorionic gonadotrophin by a hepatoblastoma resulting in precocious puberty.

    PubMed Central

    Beach, R; Betts, P; Radford, M; Millward-Sadler, H

    1984-01-01

    A two year old boy presented with precocious puberty associated with hepatoblastoma. Serum concentrations of beta human chorionic gonadotrophin and alpha-fetoprotein were raised. An aggressive chemotherapeutic regimen resulted in useful palliation and interesting changes in the beta human chorionic gonadotrophin and alpha-fetoprotein concentrations. In contrast to a previous report, the ultrastructure of the tumour showed frequent Golgi apparatus but no other electron dense membrane bound vesicles. Images PMID:6205022

  17. Apoptosis in human chorionic villi and decidua in normal and ectopic pregnancy.

    PubMed

    Kokawa, K; Shikone, T; Nakano, R

    1998-01-01

    To investigate possible effects of implantation on apoptosis, we examined the cleavage of DNA in human chorionic villi and decidua in intrauterine and ectopic pregnancy. Very limited but detectable cleavage of DNA was recognized in the chorionic villi and decidua in normal pregnancy. A ladder pattern, characteristic of the apoptotic breakdown of DNA, was present in the villi in tubal pregnancy. High molecular weight DNA was predominant in the decidua in tubal pregnancy. Quantitative analysis of low molecular weight fragments of DNA revealed a significant increase in the villous tissue, together with a significant decrease in the decidual tissue, in tubal pregnancy as compared to those in normal pregnancy. An analysis in situ revealed that apoptotic cells were predominant in the syncytiotrophoblast in tubal pregnancy. In decidual tissue, labelled cells were occasionally seen in normal pregnancy, and their numbers decreased in tubal pregnancy. The present study demonstrates that apoptosis occurs in the villi, but not in the decidua in tubal pregnancy, unlike the situation in normal pregnancy. Our results suggest that the implantation site might affect the occurrence of apoptotic changes in early pregnancy of humans.

  18. Determination of Human Chorionic Gonadotropin in Postmortem Samples in Ectopic Pregnancies.

    PubMed

    Palmiere, Cristian; Lesta, Maria del Mar; Fanton, Laurent; Ventura, Francesco; Bonsignore, Alessandro; Reggiani Bonetti, Luca

    2016-01-01

    Increased human chorionic gonadotropin levels (HCG) can be detected in femoral blood, bile, and vitreous humor collected during autopsy of pregnant women using a standard kit designed for living patients. In the study herein, the concentrations of HCG were measured in postmortem serum, vitreous, bile, cerebrospinal, and pericardial fluids in 4 cases of fatal ectopic pregnancy and 40 controls using a quantitative electrochemiluminescence immunoassay designed for living patients. No false-negative cases were identified in any of the analyzed samples in any of the ectopic pregnancy cases. No correlations were found between total HCG levels in postmortem serum and the other tested specimens. The results of this study would suggest that higher HCG in bile, vitreous, pericardial, and cerebrospinal fluids may confirm the existence of ectopic pregnancy and therefore identify other situations in which this hormone is increased, although gestational age cannot be reliably estimated using these values.

  19. Urinary human chorionic gonadotropin isoform concentrations in doping control samples.

    PubMed

    Butch, Anthony W; Woldemariam, Getachew A

    2016-11-01

    Anti-doping laboratories routinely use immunoassays to measure urinary concentrations of human chorionic gonadotropin (hCG). To minimize immunoassay differences and false positive screen results from inactive isoforms (free β-subunit (hCGβ), β-subunit core fragment (hCGβcf)) laboratories now use intact hCG instead of total hCG immunoassays to measure hCG. To determine the distribution of hCG isoforms in urine, we determined the concentrations of intact hCG, hCGβ, and hCGβcf in male urine samples based on immunoassay total hCG concentrations using a sequential immunoextraction and a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. hCG was isolated using antibody-conjugated magnetic beads and unique tryptic peptides were quantified by LC-MS/MS. Negative samples with detectable but low total hCG concentrations (1.2-3.5 pmol/L) had intact and hCGβ concentrations <1.2 pmol/L, and hCGβcf concentrations <2.3 pmol/L by LC-MS/MS. Urine samples from an athlete receiving hCG had intact hCG concentrations ranging from 18.8 to 57.6 pmol/L, hCGβ concentrations <0.7 pmol/L, and hCGβcf concentrations ranging from 94 to 243% of the intact hCG concentration. In 27 atypical samples with total hCG concentrations ranging from 16.7 to 412.7 pmol/L with intact hCG <2.7 pmol/L by immunoassay, all samples had intact hCG concentrations <3.8 pmol/L and hCGβ concentrations <6.2 pmol/L by LC-MS/MS. hCGβcf concentrations by LC-MS/MS varied widely and ranged from 1.03 to 21.9 pmol/L. In summary, total hCG immunoassays significantly overestimate hCG concentrations and can produce false positive results. Although the intact hCG immunoassay slightly overestimates hCG concentrations compared to LC-MS/MS, it can distinguish between cases of hCG use and atypical cases with elevated total hCG concentrations. Copyright © 2016 John Wiley & Sons, Ltd.

  20. 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...

  1. 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...

  2. 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 ...

  3. The human chorion contains definitive hematopoietic stem cells from the fifteenth week of gestation.

    PubMed

    Muench, Marcus O; Kapidzic, Mirhan; Gormley, Matthew; Gutierrez, Alan G; Ponder, Kathryn L; Fomin, Marina E; Beyer, Ashley I; Stolp, Haley; Qi, Zhongxia; Fisher, Susan J; Bárcena, Alicia

    2017-04-15

    We examined the contribution of the fetal membranes, amnion and chorion, to human embryonic and fetal hematopoiesis. A population of cells displaying a hematopoietic progenitor phenotype (CD34(++) CD45(low)) of fetal origin was present in the chorion at all gestational ages, associated with stromal cells or near blood vessels, but was absent in the amnion. Prior to 15 weeks of gestation, these cells lacked hematopoietic in vivo engraftment potential. Differences in the chemokine receptor and β1 integrin expression profiles of progenitors between the first and second trimesters suggest that these cells had gestationally regulated responses to homing signals and/or adhesion mechanisms that influenced their ability to colonize the stem cell niche. Definitive hematopoietic stem cells, capable of multilineage and long-term reconstitution when transplanted in immunodeficient mice, were present in the chorion from 15-24 weeks gestation, but were absent at term. The second trimester cells also engrafted secondary recipients in serial transplantation experiments. Thus, the human chorion contains functionally mature hematopoietic stem cells at mid-gestation.

  4. 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

  5. Avoidance of Maternal Cell Contamination and Overgrowth in Isolating Fetal Chorionic Villi Mesenchymal Stem Cells from Human Term Placenta.

    PubMed

    Sardesai, Varda S; Shafiee, Abbas; Fisk, Nicholas M; Pelekanos, Rebecca A

    2017-04-01

    Human placenta is rich in mesenchymal stem/stromal cells (MSC), with their origin widely presumed fetal. Cultured placental MSCs are confounded by a high frequency of maternal cell contamination. Our recent systematic review concluded that only a small minority of placental MSC publications report fetal/maternal origin, and failed to discern a specific methodology for isolation of fetal MSC from term villi. We determined isolation conditions to yield fetal and separately maternal MSC during ex vivo expansion from human term placenta. MSCs were isolated via a range of methods in combination; selection from various chorionic regions, different commercial media, mononuclear cell digest and/or explant culture. Fetal and maternal cell identities were quantitated in gender-discordant pregnancies by XY chromosome fluorescence in situ hybridization. We first demonstrated reproducible maternal cell contamination in MSC cultures from all chorionic anatomical locations tested. Cultures in standard media rapidly became composed entirely of maternal cells despite isolation from fetal villi. To isolate pure fetal cells, we validated a novel isolation procedure comprising focal dissection from the cotyledonary core, collagenase/dispase digestion and explant culture in endothelial growth media that selected, and provided a proliferative environment, for fetal MSC. Comparison of MSC populations within the same placenta confirmed fetal to be smaller, more osteogenic and proliferative than maternal MSC. We conclude that in standard media, fetal chorionic villi-derived MSC (CV-MSC) do not grow readily, whereas maternal MSC proliferate to result in maternal overgrowth during culture. Instead, fetal CV-MSCs require isolation under specific conditions, which has implications for clinical trials using placental MSC. Stem Cells Translational Medicine 2017;6:1070-1084.

  6. 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.

  7. 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

  8. Successful quadruplet pregnancy following ovulation induced with human menopausal gonadotropin and human chorionic gonadotropin.

    PubMed

    Lauersen, N H; Buchman, M; Beling, C G

    1974-01-01

    A case report of a quadruplet pregnancy that followed the induction of ovulation by human chorionic gonadotropin and human menopausal gonadotropin is presented. Examination revealed 4 separate placentas, indicating development from 4 different ova. The infants all did well at term, with no signs of respiratory distress syndrome, and have developed normally. Early diagnosis by ultrasonography and complete early bedrest are important for fetal survival. Hospitalization at Week 27-28 of pregnancy is essential, and a complete, competent staff able to handle high-risk patients should be available. Intravenous ethanol infusion is useful during early labor. The patient must be carefully observed for postpartum hemorrhage and should be followed in the recovery room for 24 hours.

  9. Beta-human chorionic gonadotropin producing osteosarcoma of the sacrum in a 26-year-old woman: a case report and review of the literature.

    PubMed

    Glass, Ryan; Asirvatham, Jaya Ruth; Kahn, Leonard; Aziz, Mohamed

    2015-01-01

    Ectopic secretion of beta-human chorionic gonadotropin is considered a poor prognostic marker in epithelial tumors. However, very few cases have been reported in sarcomas. We present the case of a 26-year-old female who presented with a metastatic osteosarcoma. She underwent usual testing prior to starting treatment and was found to have elevated levels of beta-human chorionic gonadotropin. As the patient was not pregnant, another source of beta-human chorionic gonadotropin secretion had to be considered. The tumor cells demonstrated positive staining for beta-human chorionic gonadotropin by immunohistochemistry, and serum levels of beta-human chorionic gonadotropin were used to monitor tumor progression and response to chemotherapy. We review the literature and discuss a potential role of beta-human chorionic gonadotropin in the treatment of such patients.

  10. Human chorionic gonadotropin is expressed virtually in all intracranial germ cell tumors.

    PubMed

    Takami, Hirokazu; Fukushima, Shintaro; Fukuoka, Kohei; Suzuki, Tomonari; Yanagisawa, Takaaki; Matsushita, Yuko; Nakamura, Taishi; Arita, Hideyuki; Mukasa, Akitake; Saito, Nobuhito; Kanamori, Masayuki; Kumabe, Toshihiro; Tominaga, Teiji; Kobayashi, Keiichi; Nagane, Motoo; Iuchi, Toshihiko; Tamura, Kaoru; Maehara, Taketoshi; Sugiyama, Kazuhiko; Nakada, Mitsutoshi; Kanemura, Yonehiro; Nonaka, Masahiro; Yokogami, Kiyotaka; Takeshima, Hideo; Narita, Yoshitaka; Shibui, Soichiro; Nakazato, Yoichi; Nishikawa, Ryo; Ichimura, Koichi; Matsutani, Masao

    2015-08-01

    Human chorionic gonadotropin (hCG) production has been utilized as a diagnostic marker for germinoma with syncytiotrophoblastic giant cells (STGC) and choriocarcinoma. Elevated hCG in germinoma is considered to predict less favorable prognosis, and an intensive treatment strategy may accordingly be applied. However, there is some evidence that any germinoma may produce hCG to varying extent. We investigated mRNA expression of the hCG β subunit (hCGβ) using real time quantitative polymerase chain reaction in 94 germ cell tumors (GCTs). Most (93.3 %) GCTs showed higher expression levels compared with that of normal brain tissue (1.09 × 10(0)-1.40 × 10(5) fold). The expression was the highest in GCTs which harbor choriocarcinoma or STGC components. The expression level of hCGβ in germinoma was highly variable (1.09 × 10(0)-5.88 × 10(4) fold) in linear but not bimodal distribution. hCG concentrations in serum and CSF correlated with gene expression, especially when GCTs with single histological component were analyzed separately. The expression was not significantly associated with recurrence in pure germinoma. These results suggest that the serum/CSF hCG levels may need to be interpreted with caution as most GCTs appear to have the capacity of producing hCG irrespective of their histology. The clinical significance of ubiquitous hCG expression in GCTs needs further investigation.

  11. 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.

  12. 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.

  13. Beta Human Chorionic Gonadotropin - Induction of Apoptosis in Breast Cancer

    DTIC Science & Technology

    2006-01-01

    rehydrated, and digested with proteinase K (25 ug/ml in TBS) using standard 19 methods. After quenching with 3% hydrogen peroxide , sections were...the 19 Chemicon Mouse to Mouse detection kit. Endogenous peroxidase was blocked with 3% aqueous 20 hydrogen peroxide . Slides were incubated with...Agwarwal, M.L., Das, T., Sa, G., 2002. Curcumin induces apoptosis in human breast cancer cells through p53-dependent Bax induction. FEBS Lett. 512

  14. Gastric cancer in the setting of persistently elevated human chorionic gonadotropin: a case report.

    PubMed

    Walker, Latoya R; Erler, Brian

    2011-01-01

    A 35-year-old woman presented to the emergency room for the evaluation of failed surgical and medical management of a suspected ectopic pregnancy. When imaging studies were performed, she had lymphadenopathy and diffuse sclerosis of the osseous framework. Multiple biopsies were performed and revealed poorly differentiated metastatic carcinoma with signet ring features. Esophagogastroduodenoscopy confirmed the findings of a Stage IV gastric adenocarcinoma. Signs and symptoms of gastric carcinoma are vague. However, to our knowledge, an elevation in human chorionic gonadotropin (hCG) is not an associated finding. Persistence of hCG has many causes from abnormal pregnancy to menopause and other forms of cancer.

  15. Effects of human chorionic gonadotropin on testicular interstitial tissues in men with non-obstructive azoospermia.

    PubMed

    Oka, S; Shiraishi, K; Matsuyama, H

    2016-11-16

    Non-obstructive azoospermia is a severe condition because spermatogenesis per se is disrupted. Although microdissection testicular sperm extraction is the standard therapy for non-obstructive azoospermia, sperm retrieval is unsuccessful in approximately 50% of patients. For these patients, we conducted human chorionic gonadotropin-based salvage hormonal therapy, and sperm retrieval was possible in 10-20% of patients. The objectives of this study were to assess the changes in interstitial lesions in patients with non-obstructive azoospermia and to evaluate the effects of human chorionic gonadotropin on these tissues. Testicular biopsy specimens were obtained from 10 non-obstructive azoospermia patients who failed to obtain spermatozoa and from 10 obstructive azoospermia patients. All non-obstructive azoospermia patients received salvage hormonal therapy after microdissection testicular sperm extraction. Hematoxylin and eosin (H.E.) staining and immunohistochemical staining for steroidogenic acute regulatory protein antibody, the Leydig cell marker, and TE-7 antibody, the fibroblast marker, as well as picrosirius red staining to detect collagen fibers, were performed. We measured interstitial lesions, as Leydig cell area and the other area, with ImageJ software. Interstitial area, excluding Leydig cells, increased up to 12.5% in non-obstructive azoospermia compared with 1.2% in obstructive azoospermia (p < 0.01), which was mainly because of fibrosis with TE-7-positive fibroblasts. The increase in interstitial lesions was correlated with Johnsen scores. Interstitial area, excluding the Leydig cells, decreased by 29% after salvage hormonal therapy (p < 0.05), indicating improvement of interstitial fibrosis in non-obstructive azoospermia. There were no significant difference in total Leydig cell area and size of each Leydig cells between obstructive azoospermia and non-obstructive azoospermia. After the salvage hormonal therapy, a portion of the Leydig cells became

  16. The secretory patterns of relaxin and human chorionic gonadotropin in human pregnancy.

    PubMed

    Seki, K; Uesato, T; Tabei, T; Kato, K

    1985-10-01

    Relaxin and human chorionic gonadotropin (hCG) were simultaneously determined in the same serum samples obtained from pregnant women. Although the secretory pattern of relaxin, in general, appeared to parallel that of hCG during human pregnancy, several discrepancies were discerned in the secretory patterns of the two hormones. The mean hCG concentration significantly differed between weeks 4-7 and 8-11 of pregnancy, but the mean relaxin concentration did not. The mean relaxin concentration began to decrease at weeks 16-19 whereas that of hCG did so at weeks 12-15. The mean relaxin concentration at weeks 4-7 was significantly higher than that at weeks 24-27, though there was no significant difference between the mean hCG concentrations in the two periods. These differences in the secretory pattern of relaxin from that of hCG indicate that relaxin secretion in pregnancy is not determined only by the circulating level of hCG. The responsiveness of the corpus luteum of pregnancy to hCG stimulation of relaxin secretion may vary as a function of the age of the corpus luteum, and this may partially account for the differences between the secretory pattern of relaxin and that of hCG observed in the present study.

  17. Giant cell tumor of bone with secondary aneurysmal bone cyst-like change producing β-human chorionic gonadotropin.

    PubMed

    Fitzhugh, Valerie A; Katava, Gordana; Wenokor, Cornelia; Roche, Natalie; Beebe, Kathleen S

    2014-06-01

    Giant cell tumor of bone is a benign, locally aggressive neoplasm that is composed of sheets of neoplastic mononuclear cells interspersed amongst non-neoplastic, uniformly distributed, osteoclast-like giant cells. They represent approximately 4-5% of primary bone tumors. Rarely, bone tumors have been noted to produce human chorionic gonadotropin, a finding most often reported in osteosarcoma. We present the case of a young woman who presented with a low-level human chorionic gonadotropin level which, after resection of her recurrent giant cell tumor of bone with secondary aneurysmal bone cyst-like change, became undetectable in her blood. Furthermore, cells within the aneurysmal bone cyst component were immunohistochemically positive for β-human chorionic gonadotropin. This is the first report of such a finding in the literature.

  18. Human chorionic gonadotropin decreases human breast cancer cell proliferation and promotes differentiation.

    PubMed

    Liao, Xing-Hua; Wang, Yue; Wang, Nan; Yan, Ting-Bao; Xing, Wen-Jing; Zheng, Li; Zhao, Dong-Wei; Li, Yan-Qi; Liu, Long-Yue; Sun, Xue-Guang; Hu, Peng; Zhang, Tong-Cun

    2014-05-01

    Human chorionic gonadotropin (hCG) is a glycoprotein produced by placental trophoblasts. Previous studies indicated that hCG could be responsible for the pregnancy-induced protection against breast cancer in women. It is reported that hCG decreases proliferation and invasion of breast cancer MCF-7 cells. Our research also demonstrates that hCG can reduce the proliferation of MCF-7 cells by downregulating the expression of proliferation markers, proliferating cell nuclear antigen (PCNA), and proliferation-related Ki-67 antigen (Ki-67). Interestingly, we find here that hCG elevates the state of cellular differentiation, as characterized by the upregulation of differentiation markers, β-casein, cytokeratin-18 (CK-18), and E-cadherin. Inhibition of hCG secretion or luteinizing hormone/hCG receptors (LH/hCGRs) synthesis can weaken the effect of hCG on the induction of cell differentiation. Furthermore, hCG can suppress the expression of estrogen receptor alpha. hCG activated receptor-mediated cyclic adenosine monophosphate/protein kinase A signaling pathway. These findings indicated that a protective effect of hCG against breast cancer may be associated with its growth inhibitory and differentiation induction function in breast cancer cells.

  19. A bacterial protein has homology with human chorionic gonadotropin (hCG).

    PubMed

    Grover, S; Woodward, S R; Odell, W D

    1993-06-30

    Studies from our laboratory have demonstrated the presence of a 48.5 kD cell wall protein in the bacterium, Xanthomonas maltophilia, which immunologically resembles the beta subunit of human chorionic gonadotropin. Primers were designed from the amino acid sequences of enzymatically cleaved peptide fragments of this protein. These primers were used to obtain PCR amplified products, which were subsequently cloned in a PCR11TA cloning vector, and a 492 base pair nucleotide sequence was obtained with a 164 amino acid open reading frame. When this nucleotide sequence was aligned with exon 2 of genes 5 and 6 of the beta hCG gene, a 53% homology was observed. The translated protein sequence had a 35% homology with hCG and a 25% homology with human luteinizing hormone.

  20. Biological properties of dehydrated human amnion/chorion composite graft: implications for chronic wound healing.

    PubMed

    Koob, Thomas J; Rennert, Robert; Zabek, Nicole; Massee, Michelle; Lim, Jeremy J; Temenoff, Johnna S; Li, William W; Gurtner, Geoffrey

    2013-10-01

    Human amnion/chorion tissue derived from the placenta is rich in cytokines and growth factors known to promote wound healing; however, preservation of the biological activities of therapeutic allografts during processing remains a challenge. In this study, PURION® (MiMedx, Marietta, GA) processed dehydrated human amnion/chorion tissue allografts (dHACM, EpiFix®, MiMedx) were evaluated for the presence of growth factors, interleukins (ILs) and tissue inhibitors of metalloproteinases (TIMPs). Enzyme-linked immunosorbent assays (ELISA) were performed on samples of dHACM and showed quantifiable levels of the following growth factors: platelet-derived growth factor-AA (PDGF-AA), PDGF-BB, transforming growth factor α (TGFα), TGFβ1, basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), placental growth factor (PLGF) and granulocyte colony-stimulating factor (GCSF). The ELISA assays also confirmed the presence of IL-4, 6, 8 and 10, and TIMP 1, 2 and 4. Moreover, the relative elution of growth factors into saline from the allograft ranged from 4% to 62%, indicating that there are bound and unbound fractions of these compounds within the allograft. dHACM retained biological activities that cause human dermal fibroblast proliferation and migration of human mesenchymal stem cells (MSCs) in vitro. An in vivo mouse model showed that dHACM when tested in a skin flap model caused mesenchymal progenitor cell recruitment to the site of implantation. The results from both the in vitro and in vivo experiments clearly established that dHACM contains one or more soluble factors capable of stimulating MSC migration and recruitment. In summary, PURION® processed dHACM retains its biological activities related to wound healing, including the potential to positively affect four distinct and pivotal physiological processes intimately involved in wound healing: cell proliferation, inflammation, metalloproteinase activity and recruitment of progenitor cells. This suggests

  1. The Role of Human Chorionic Gonadotropin as Tumor Marker: Biochemical and Clinical Aspects.

    PubMed

    Sisinni, Lorenza; Landriscina, Matteo

    2015-01-01

    Tumor markers are biological substances that are produced/released mainly by malignant tumor cells, enter the circulation in detectable amounts and are potential indicators of the presence of a tumor. The most useful biochemical markers are the tumor-specific molecules, i.e., receptors, enzymes, hormones, growth factors or biological response modifiers that are specifically produced by tumor cells and not, or minimally, by the normal counterpart (Richard et al. Principles and practice of gynecologic oncology. Wolters Kluwer Health, Philadelphia, 2009). Based on their specificity and sensitivity in each malignancy, biomarkers are used for screening, diagnosis, disease monitoring and therapeutic response assessment in clinical management of cancer patients.This chapter is focused on human chorionic gonadotropin (hCG), a hormone with a variety of functions and widely used as a tumor biomarker in selected tumors. Indeed, hCG is expressed by both trophoblastic and non-trophoblastic human malignancies and plays a role in cell transformation, angiogenesis, metastatization, and immune escape, all process central to cancer progression. Of note, hCG testing is crucial for the clinical management of placental trophoblastic malignancies and germ cell tumors of the testis and the ovary. Furthermore, the production of hCG by tumor cells is accompanied by varying degrees of release of the free subunits into the circulation, and this is relevant for the management of cancer patients (Triozzi PL, Stevens VC, Oncol Rep 6(1):7-17, 1999).The name chorionic gonadotropin was conceived: chorion derives from the latin chordate meaning afterbirth, gonadotropin indicates that the hormone is a gonadotropic molecule, acting on the ovaries and promoting steroid production (Cole LA, Int J Endocrinol Metab 9(2):335-352, 2011). The function, the mechanism of action and the interaction between hCG and its receptor continue to be the subject of intensive investigation, even though many issues about

  2. Human chorionic gonadotropin promotes expression of protein absorption factors in the intestine of goldfish (Carassius auratus).

    PubMed

    Zhou, Y; Hao, G; Zhong, H; Wu, Q; Lu, S Q; Zhao, Q; Liu, Z

    2015-07-27

    Protein use is crucial for the ovulation and spawning of fish. Currently, limited information is available regarding the expression of protein absorption factors during the breeding seasons of teleosts and thus how various proteins involved in this process is not well-understood. The expression of CDX2, CREB, gluatamate dehydrogenase, LAT2, aminopeptidase N, PepT1, and SP1 were significantly elevated from the non-breeding season to the breeding season in female goldfish, and all proteins except PepT1 and SP1 were elevated in male goldfish. Injection of human chorionic gonadotropin upregulated the expression of all proteins except for aminopeptidase N in female goldfish and SP1 in male goldfish, suggesting a luteinizing hormone-inductive effect on protein absorption factors. Protein use in the intestine is increased during the breeding seasons as a result of increased luteinizing hormone.

  3. Potential Therapy for Rheumatoid Arthritis and Sjögren Syndrome With Human Chorionic Gonadotropin.

    PubMed

    Rao, C V

    2016-05-01

    Autoimmune diseases such as rheumatoid arthritis (RA) and Sjögren syndrome (SS) ameliorate during pregnancy, through dampening (immunotolerance) of the maternal immune system which protects the fetus from rejection. A large number of studies have shown that human chorionic gonadotropin (hCG) contributes to this tolerance. Studies on animal models have reaffirmed that hCG treatment mimics the benefits of pregnancy. Based on the scientific evidence, randomized clinical trials comparing hCG with current therapies and/or placebo are recommended for RA, SS, and for other autoimmune diseases such as, type 1 diabetes and ankylosing spondylitis, which also get better during pregnancy and hCG treatment seems to help.

  4. 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.

  5. The suitability of human chorionic gonadotropin (hCG)-based birth-control vaccines.

    PubMed

    Dirnhofer, S; Wick, G; Berger, P

    1994-10-01

    It has been widely hoped that immunological methods of fertility regulation by active immunization against specific antigens of the oocyte, sperm, zygote and early embryo, and the placental pregnancy hormone human chorionic gonadotropin (hCG), will provide a means to control the problem of worldwide population growth. The most advanced candidate vaccines are based on hCG immunogens and have entered clinical trials. However, during the past few years, increasing evidence has emerged that the current approaches using hCG as the target molecule may have some major drawbacks. On the basis of their recent findings, Stephan Dirnhofer and colleagues raise doubts on the suitability, safety and efficacy of gonadotropin-based immunological contraceptive vaccines.

  6. Expression of the alpha subunit of human chorionic gonadotropin is specifically correlated with tumorigenic expression in human cell hybrids.

    PubMed Central

    Stanbridge, E J; Rosen, S W; Sussman, H H

    1982-01-01

    The expression of HeLa parent phenotype protein markers, the alpha subunit of human chorionic gonadotropin and placental alkaline phosphatase isoenzymes, has been evaluated in paired tumorigenic and nontumorigenic HeLa-fibroblast human cell hybrids. Both of these proteins have been used clinically as markers of malignancy. The results showed that both are expressed in the hybrids. Expression of the gonadotropin subunit in the hybrids is specifically correlated with tumorigenicity; the placental alkaline phosphatase isoenzyme showed no such correlation and was expressed in both tumorigenic and nontumorigenic hybrids. PMID:6959112

  7. [Human chorionic gonadotropin--a well-known hormone with unknown functions].

    PubMed

    Głodek, Aleksandra; Kubiczak, Marta; Urbaniak, Paulina; Walkowiak, Grzegorz; Nowak-Markwitz, Ewa; Jankowska, Anna

    2012-10-01

    Human chorionic gonadotropin (CG) belongs to the glycoprotein family consisting of LH, FSH and TSH. All of these hormones are composed of two subunits: common to the whole family alpha subunit and hormone-specific beta subunit CG has paracrine effects on several processes such as placentation, implantation, angiogenesis and delaying the apoptosis of corpus luteum. Serum level of CG is used to monitor pregnancy and pregnancy disorders. Recent studies have shown that the synthesis of CG is a characteristic feature of a wide variety of malignant and non-malignant tumors. The role of CG in cancerogensis remains unclear but the main hypothesis concerns its antiapoptotic impact of the hormone on the neoplastic cells. The synthesis of functional CG requires the activity of separate genes encoding both hormone's subunits, but it is the beta subunit accessibility which controls the process. The protein synthesis must be followed by proper folding and posttranslational modifications of the molecule. Particularly glycosylation of human chorionic gonadotropin was shown to have an impact on the hormone's function. The amount and the structure of carbohydrate residuals attached to CG may be different and lead to the formation of hormone variants, which vary in molecular mass. Normal CG with a molecular mass of about 37.5 kDa is produced by the syncytiotrophoblast, while the variant with higher molecular mass - 38.5-40 kDa, described as hyperglicosylated CG, is secreted by undifferentiated trophoblast cells and some cancers. It is suggested that those forms have different but complementary biological functions. However the mechanism of the action of particular variants and signaling pathways activated by those forms are still obscure.

  8. Ontological Differences in First Compared to Third Trimester Human Fetal Placental Chorionic Stem Cells

    PubMed Central

    Jones, Gemma N.; Moschidou, Dafni; Puga-Iglesias, Tamara-Isabel; Kuleszewicz, Katarzyna; Vanleene, Maximilien; Shefelbine, Sandra J.; Bou-Gharios, George; Fisk, Nicholas M.; David, Anna L.; De Coppi, Paolo; Guillot, Pascale V.

    2012-01-01

    Human mesenchymal stromal/stem cells (MSC) isolated from fetal tissues hold promise for use in tissue engineering applications and cell-based therapies, but their collection is restricted ethically and technically. In contrast, the placenta is a potential source of readily-obtainable stem cells throughout pregnancy. In fetal tissues, early gestational stem cells are known to have advantageous characteristics over neonatal and adult stem cells. Accordingly, we investigated whether early fetal placental chorionic stem cells (e-CSC) were physiologically superior to their late gestation fetal chorionic counterparts (l-CSC). We showed that e-CSC shared a common phenotype with l-CSC, differentiating down the osteogenic, adipogenic and neurogenic pathways, and containing a subset of cells endogenously expressing NANOG, SOX2, c-MYC, and KLF4, as well as an array of genes expressed in pluripotent stem cells and primordial germ cells, including CD24, NANOG, SSEA4, SSEA3, TRA-1-60, TRA-1-81, STELLA, FRAGILIS, NANOS3, DAZL and SSEA1. However, we showed that e-CSC have characteristics of an earlier state of stemness compared to l-CSC, such as smaller size, faster kinetics, uniquely expressing OCT4A variant 1 and showing higher levels of expression of NANOG, SOX2, c-MYC and KLF4 than l-CSC. Furthermore e-CSC, but not l-CSC, formed embryoid bodies containing cells from the three germ layer lineages. Finally, we showed that e-CSC demonstrate higher tissue repair in vivo; when transplanted in the osteogenesis imperfecta mice, e-CSC, but not l-CSC increased bone quality and plasticity; and when applied to a skin wound, e-CSC, but not l-CSC, accelerated healing compared to controls. Our results provide insight into the ontogeny of the stemness phenotype during fetal development and suggest that the more primitive characteristics of early compared to late gestation fetal chorionic stem cells may be translationally advantageous. PMID:22962584

  9. 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.

  10. 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.

  11. Gap junctional communication during human trophoblast differentiation: influence of human chorionic gonadotropin.

    PubMed

    Cronier, L; Bastide, B; Hervé, J C; Délèze, J; Malassiné, A

    1994-07-01

    During pregnancy, the trophoblast develops from the fusion of cytotrophoblastic cells into a syncytiotrophoblast. As the exchange of molecules through gap junctions is considered to play a role in the control of cell and tissue differentiation, the cell to cell diffusion of a fluorescent dye was investigated in human trophoblastic cells differentiating in culture. The fluorescence recovery after photobleaching technique was used to estimate the transfer of 6-carboxyfluorescein from contiguous cellular elements into photobleached cells. Fluorescence recovery follows a slow exponential time course when the cell to cell exchange process is rate limited by the presence of gap junctional channels between contiguous cells, contrasting with a much faster step-like course in the case of fusion of the plasma membranes. In the presence of 10% fetal calf serum, Percoll-purified cytotrophoblastic cells develop into cellular aggregates, then into a syncytium, within 24-48 h after plating. During this in vitro differentiation, fluorescence recoveries after photobleaching with a time course typical for gap junctions were observed between aggregated cytotrophoblastic cells, between cytotrophoblastic cells and syncytiotrophoblasts, and between contiguous syncytiotrophoblasts. The maximum percentage of gap junctional coupling occurs on the fourth day. This fluorescence recovery is attributed to the diffusion of dye through gap junctions, because it can be interrupted by exposure to a known junctional uncoupler (3 mM heptanol). The effects of hCG on this gap junctional communication during trophoblast differentiation were investigated. In the presence of 500 mIU/ml hCG in the culture medium, the percentage of coupled cells was increased at all stages of culture, and the highest proportion of coupled cells was observed after 2 days of culture vs. 4 days in control medium. Moreover, the diffusion rate constant k (the inverse value of the time constant measured on recovery curves) was

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

    PubMed

    Ezcurra, Diego; Humaidan, Peter

    2014-10-03

    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.

  13. Potential of human fetal chorionic stem cells for the treatment of osteogenesis imperfecta.

    PubMed

    Jones, Gemma N; Moschidou, Dafni; Abdulrazzak, Hassan; Kalirai, Bhalraj Singh; Vanleene, Maximilien; Osatis, Suchaya; Shefelbine, Sandra J; Horwood, Nicole J; Marenzana, Massimo; De Coppi, Paolo; Bassett, J H Duncan; Williams, Graham R; Fisk, Nicholas M; Guillot, Pascale V

    2014-02-01

    Osteogenesis imperfecta (OI) is a genetic bone pathology with prenatal onset, characterized by brittle bones in response to abnormal collagen composition. There is presently no cure for OI. We previously showed that human first trimester fetal blood mesenchymal stem cells (MSCs) transplanted into a murine OI model (oim mice) improved the phenotype. However, the clinical use of fetal MSC is constrained by their limited number and low availability. In contrast, human fetal early chorionic stem cells (e-CSC) can be used without ethical restrictions and isolated in high numbers from the placenta during ongoing pregnancy. Here, we show that intraperitoneal injection of e-CSC in oim neonates reduced fractures, increased bone ductility and bone volume (BV), increased the numbers of hypertrophic chondrocytes, and upregulated endogenous genes involved in endochondral and intramembranous ossification. Exogenous cells preferentially homed to long bone epiphyses, expressed osteoblast genes, and produced collagen COL1A2. Together, our data suggest that exogenous cells decrease bone brittleness and BV by directly differentiating to osteoblasts and indirectly stimulating host chondrogenesis and osteogenesis. In conclusion, the placenta is a practical source of stem cells for the treatment of OI.

  14. Immunological studies of human placentae: the distribution and character of immunoglobulins in chorionic villi.

    PubMed Central

    Johnson, P M; Natvig, J B; Ystehede, U A; Faulk, W P

    1977-01-01

    All four human IgG subclasses, and both kappa and lambda light chains, were detected by immunofluorescence in similar distributions in chorionic villi of human placentae. IgG1 and IgG3 were the predominant subclasses. No evidence was obtained for local enzymatic digestion of IgG during placental transfer. Most of the IgG on the trophoblastic basement membrane (TBM) was loosely bound and could be removed by prolonged washing, although some appeared to be more tightly bound to small segments of the TBM. IgM, but not IgA, was present in small amounts in placental villous structures. Immunoglobulin was never observed within the syncytiotrophoblast. Antisera to IgG genetic (Gm) markers were used to locate IgG thought to be of foetal or maternal origin. The presence of paternal Gm markers not carried by the mother was taken as evidence for foetal IgG. Foetal (paternal) Gm markers were observed in placentae, although maternal IgG was the major immunoglobulin present in placental villi. Both maternal and foetal IgG were demonstrated in fibrinoid deposits, vessel walls and the cytoplasm of some stromal cells. Only foetal IgG was definitively observed in the immunoglobulin that is tightly bound to the TBM. PMID:342151

  15. Paper-based microfluidic devices for electrochemical immunofiltration analysis of human chorionic gonadotropin.

    PubMed

    Cao, Liangli; Fang, Cheng; Zeng, Ruosheng; Zhao, Xiongjie; Jiang, Yuren; Chen, Zhencheng

    2017-02-02

    An electrochemical immunofiltration analysis was introduced into microfluidic paper-based analytical devices (μPADs) for the first time, which was based on photolithography and screen-printing technology. The hydrophilic test zones of the aldehyde-functionalized screen-printed electrodes (SPEs) were biofunctionalized with capture antibodies (Ab1). A sensitive immune detection method was developed by using primary signal antibody functionalized gold nanoparticles (GNPs/Ab2) and alkaline phosphatase conjugated secondary antibody (ALP-IgG). Differential pulse voltammetry (DPV) was performed to detect the electrochemical response. The microfluidic paper-based electrochemical immunosensor (μ-PEI) was optimized and characterized for the detection of human chorionic gonadotropin (HCG), a model analyte, in a linear range from 1.0mIUmL(-1) to 100.0 IU mL(-1) with a detection limit of 0.36mIUmL(-1). Additionally, the proposed μ-PEI was used to test HCG in real human serum and obtained satisfactory results. The disposable, efficient, sensitive and low-cost μ-PEI has exhibited great potential for the development of point-of-care testing (POCT) devices that can be applicated in healthcare monitoring.

  16. Human chorionic gonadotrophin: embryonic secretion is a time-dependent phenomenon.

    PubMed

    Woodward, B J; Lenton, E A; Turner, K

    1993-09-01

    Of 48 spare human pre-embryos achieving the expanded blastocyst stage, 22 (45.6%) secreted significant amounts of human chorionic gonadotrophin (HCG) (> 5 IU/l/day). Of these, nine remained intrazonal, seven partially hatched and six fully hatched. Embryonic production of HCG in vitro appeared to be time-dependent, starting after a certain minimum time (approximately 160 h post-insemination) and rising exponentially, with maximal HCG production around day 10. Hatching was not a prerequisite for HCG secretion, since similar amounts were produced by intrazonal blastocysts. Blastocysts derived from abnormally fertilized oocytes also began secreting HCG exponentially but secretion was delayed and the upper limit of maximum HCG secretion rate was comparatively low. The actual amount of HCG is thought to reflect the number of viable trophectoderm cells producing the hormone. HCG doubling times for blastocysts in vitro were rapid when compared to implanting blastocysts of a similar age in vivo, with 19/22 (86.4%) blastocysts having a doubling time of < 10 h. Provided a pre-embryo can secrete HCG and maintain an adequate doubling time, sufficient HCG should be produced for initial stages of embryonic recognition in vivo. Since intrazonal blastocysts are capable of fulfilling both of these criteria, the limiting factor in realizing their full potential may be escaping from the zona pellucida.

  17. Characterization of bmp15 and its regulation by human chorionic gonadotropin in the follicle of gibel carp (Carassius auratus gibelio).

    PubMed

    Chen, A-Qin; Liu, Zhi-Wei; Yang, Zhi-Gang; Leng, Xiang-Jun

    2012-09-01

    Bone morphogenetic protein (BMP15) is a member of the transforming growth factor β (TGF-β) superfamily with a key role in regulating follicle development in mammals and birds. However, potential ovarian roles of BMPs remain unexplored in teleosts. In this study, the full-length sequences of bmp15 were obtained using rapid-amplification of cDNA ends (RACE). The full-length cDNA sequence of bmp15 is 2217 bp which contained 214 bp 5'-UTR and 845 bp 3'-UTR. The open reading frame (ORF) sequence of bmp15 is 1158 bp, encoding a predicted protein of 385 amino acid residues. BMP15 has a specific RXXR protease cleavage site of TGF-β superfamily (is RIRR) and six conserved cysteine residues. Using real-time quantitative PCR revealed that bmp15 mRNA was largely expressed in the ovary and testis and mostly in oocytes within the follicle, slightly expressed in muscle, liver and pituitary. BMP15 is mainly present at stage I follicles by real-time quantitative PCR and immunohistochemistry. Phylogenetic analysis showed that gibel carp bmp15 was similar to bmp15 of zebrafish and other fish species. Treatment with human chorionic gonadotropin (hCG) in isolated follicles of gibel carp in vitro showed altered bmp15 mRNA expression: when treated with 10 ng/mL hCG for 10h, the expression level of bmp15 was significantly increased. However, with proceeding cultivation, the expression level of BMP15 mRNA decreased. The results of this study indicate that bmp15 may play a key role during development of follicles in gibel carp, especially in early stage follicles.

  18. The prognostic significance of beta human chorionic gonadotrophin and its metabolites in women with cervical carcinoma.

    PubMed Central

    Crawford, R A; Iles, R K; Carter, P G; Caldwell, C J; Shepherd, J H; Chard, T

    1998-01-01

    AIMS: To examine long term survival of women with primary and recurrent cervical carcinoma in relation to (1) excretion of beta-core (a urinary metabolite of beta human chorionic gonadotrophin (beta hCG)) and (2) beta hCG immunostaining of the tumours, to determine the suitability of these markers for assessing prognosis. METHODS: This was a prospective observational study undertaken in a gynaecological oncology centre: 57 women with primary cervical cancer and 42 with recurrent disease were recruited between January 1990 and September 1992. Kaplan-Meier survival analysis with the log rank test was used to assess survival differences with survival rate given per year of follow up. RESULTS: In primary disease, the four year survival for the beta-core negative group was 79%, compared with 14% for the beta-core positive group (p = 0.001). This was still significant for early stage disease or squamous lesions alone. In recurrent disease, beta-core positivity was not prognostically significant. Immunohistochemistry was of no prognostic significance in either group. CONCLUSIONS: beta-core excretion appears to be useful in assessing prognosis of primary cervical cancer but not of recurrent disease. A large prospective study of urinary beta-core in early stage cervical cancer is needed to determine whether it can be used as an index for modifying treatment. PMID:9930074

  19. Pregnancy rates with recombinant versus urinary human chorionic gonadotropin in in vitro fertilization: an observational study.

    PubMed

    Zeke, József; Kanyó, Katalin; Zeke, Helga; Cseh, Aron; Vásárhelyi, Barna; Szilágyi, András; Konc, János

    2011-01-01

    Randomized clinical trials (RCTs) demonstrated the equal efficacy of urinary human chorionic gonadotropin (uhCG) and recombinant hCG (rhCG) products in in vitro fertilisation (IVF). However, limitations inherent with RCTs necessitate the reinforcement of RCT results in real-life. We retrospectively analyzed pregnancies after treatment with rhCG and uhCG products (n = 391, and 96, resp.). We found that laboratory-verified pregnancy occurred more frequently in rhCG patients than in those on uhCG (43% versus 30%, P = 0.02). The association remains significant (P = 0.002) after its adjustment for clinical characteristics. The prevalence of laboratory-verified pregnancies was higher with GnRH agonist use (P = 0.012) and BMI under 30 kg/m(2) (P = 0.053) while decreased the age (P = 0.014) and the number of previous failed attempts (P = 0.08). Similar (but not significant) trends were observed with rates of pregnancy filled the 24th week. These results reinforce RCTs supporting the notion that rhCG is more efficient as uhCG during IVF.

  20. Persistent Human Chorionic Gonadotropin After Methotrexate Treatment and an Emergency Surgical Procedure for Ectopic Pregnancy.

    PubMed

    Kurt-Mangold, Michelle; Van Voorhis, Bradley J; Krasowski, Matthew D

    2015-01-01

    The case study is a 33-year-old white female with persistently elevated serum human chorionic gonadotropin (hCG) levels following methotrexate treatment and emergency surgery for ectopic pregnancy. At the time of the first methotrexate dose, the serum hCG concentration was 27,995 IU/L. The laboratory was consulted 3.5 months after the surgery, because serum hCG levels had stopped declining and had leveled off to around 80 to 90 IU/L but with negative urine pregnancy tests. Laboratory studies ruled out heterophile antibody interference and hook effect by multiple methods including analysis by different serum hCG assays, treatment with heterophile antibody blocking agents, and dilution studies. Three additional doses of methotrexate over six months were required for serum hCG concentrations to decline to undetectable levels. This case illustrates challenges that may arise with serum hCG measurements in management of ectopic pregnancies. Close collaboration between the laboratory and clinical service is key for optimal patient care.

  1. A Graphene Oxide-Based Fluorescent Method for the Detection of Human Chorionic Gonadotropin

    PubMed Central

    Xia, Ning; Wang, Xin; Liu, Lin

    2016-01-01

    Human chorionic gonadotropin (hCG) has been regarded as a biomarker for the diagnosis of pregnancy and some cancers. Because the currently used methods (e.g., disposable Point of Care Testing (POCT) device) for hCG detection require the use of many less stable antibodies, simple and cost-effective methods for the sensitive and selective detection of hCG have always been desired. In this work, we have developed a graphene oxide (GO)-based fluorescent platform for the detection of hCG using a fluorescein isothiocyanate (FITC)-labeled hCG-specific binding peptide aptamer (denoted as FITC-PPLRINRHILTR) as the probe, which can be manufactured cheaply and consistently. Specifically, FITC-PPLRINRHILTR adsorbed onto the surface of GO via electrostatic interaction showed a poor fluorescence signal. The specific binding of hCG to FITC-PPLRINRHILTR resulted in the release of the peptide from the GO surface. As a result, an enhanced fluorescence signal was observed. The fluorescence intensity was directly proportional to the hCG concentration in the range of 0.05–20 IU/mL. The detection limit was found to be 20 mIU/mL. The amenability of the strategy to hCG analysis in biological fluids was demonstrated by assaying hCG in the urine samples. PMID:27754379

  2. 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.

  3. Human chorionic gonadotropin β subunit affects the expression of apoptosis-regulating factors in ovarian cancer.

    PubMed

    Szczerba, Anna; Śliwa, Aleksandra; Kubiczak, Marta; Nowak-Markwitz, Ewa; Jankowska, Anna

    2016-01-01

    Expression of human chorionic gonadotropin, especially its free β subunit (hCGβ) were shown to play an important role in cancer growth, invasion and metastasis. It is postulated that hCGβ is one of the factors determining cancer cell survival. To test this hypothesis, we applied two models: an in vitro model of ovarian cancer using OVCAR-3 and SKOV-3 cell lines transfected with the CGB5 gene and an in vivo model of ovarian cancer tissues. The material was tested against changes in expression level of genes encoding factors involved in apoptosis: BCL2, BAX and BIRC5. Overexpression of hCGβ was found to cause a decrease in expression of the analyzed genes in the transfected cells compared with the control cells. In ovarian cancer tissues, high expression of CGB was related to significantly lower BCL2 but higher BAX and BIRC5 transcript levels. Moreover, a low BCL2/BAX ratio, characteristic of advanced stages of ovarian cancer, was revealed. Since tumors were discriminated by a significantly lower LHCGR level than the level noted in healthy fallopian tubes and ovaries, it may be stated that the effect of hCGβ on changes in the expression of apoptosis-regulating agents observed in ovarian cancer is LHCGR-independent. The results of the study suggest that the biological effects evoked by hCGβ are related to apoptosis suppression.

  4. Pleomorphic Carcinoma of the Lung with High Serum Beta-human Chorionic Gonadotropin Level and Gynecomastia

    PubMed Central

    Hasbal, Baris; Aydin, Kubra; Bozkurt, Mustafa; Namal, Esat; Oz, Buge; Kaynak, Kamil; Demir, Gokhan

    2010-01-01

    Although gynecomastia is a well-defined paraneoplastic syndrome in patients with non-small cell lung cancer, the association with pleomorphic carcinoma has not been reported. A 50-yr-old man presented with bilateral gynecomastia and elevated serum beta-human chorionic gonadotropin (βhCG) level. Chest tomography showed a mass in the right middle lobe. Right middle lobectomy and mediastinal lymph node dissection were performed. βhCG levels decreased rapidly after surgery. Histological examination revealed pleomorphic carcinoma with positive immunostaining for βhCG. Serum βhCG levels began to increase gradually on postoperatively 4th month. Computed tomography detected recurrence and chemotherapy was started. After second cycle of chemotherapy, βhCG levels decreased dramatically again and tomography showed regression in mass. Patient died 6 months later due to brain metastasis. βhCG expression may be associated with aggressive clinical course and increased risk of recurrence, also βhCG levels may be used to evaluate therapy response in patients with pleomorphic carcinoma. PMID:21165299

  5. Sensitive immunoassay of human chorionic gonadotrophin based on multi-walled carbon nanotube-chitosan matrix.

    PubMed

    Li, Na; Yuan, Ruo; Chai, Yaqin; Chen, Shihong; An, Haizhen

    2008-10-01

    A novel amperometric immunosensor for human chorionic gonadotropin (HCG) assay has been fabricated through incorporating toluidine blue (TB) and hemoglobin (Hb) on the multiwall carbon nanotube (MWNT)-chitosan (CS) modified glassy carbon electrode, followed by electrostatic adsorption of a conducting gold nanoparticles (nanogold) film as sensing interface. The MWNT-CS matrix provided a congenial microenvironment for the immobilization of biomolecules and promoted the electron transfer to enhance the sensitivity of the immunosensor. Due to the strong electrocatalytic properties of Hb and MWNT toward H(2)O(2), the Hb and MWNT significantly amplified the current signal of the antigen-antibody reaction. The immobilized toluidine blue as an electron transfer mediator exhibited excellent electrochemical redox property. After the immunosensor was incubated with HCG solution, the access of activity center of the Hb to toluidine blue was partly inhibited, which leaded to a linear decrease in the catalytic efficiency of the Hb to the oxidation of immobilized toluidine blue by H(2)O(2) over HCG concentration ranges from 0.8 to 500 mIU/mL. Under optimal condition, the detection limit for the HCG immunoassay was 0.3 mIU/mL estimated at a signal-to-noise ratio of 3. Moreover, the proposed immunosensor displayed a satisfactory stability and reproducibility.

  6. Human Chorionic Stem Cells: Podocyte Differentiation and Potential for the Treatment of Alport Syndrome.

    PubMed

    Moschidou, Dafni; Corcelli, Michelangelo; Hau, Kwan-Leong; Ekwalla, Victoria J; Behmoaras, Jacques V; De Coppi, Paolo; David, Anna L; Bou-Gharios, George; Cook, H Terence; Pusey, Charles D; Fisk, Nicholas M; Guillot, Pascale V

    2016-03-01

    Alport syndrome (AS) is a hereditary glomerulopathy caused by a mutation in type IV collagen genes, which disrupts glomerular basement membrane, leading to progressive glomerulosclerosis and end-stage renal failure. There is at present no cure for AS, and cell-based therapies offer promise to improve renal function. In this study, we found that human first trimester fetal chorionic stem cells (CSC) are able to migrate to glomeruli and differentiate down the podocyte lineage in vitro and in vivo. When transplanted into 7-week-old Alport 129Sv-Col4α3(tm1Dec)/J (-/-) mice, a single intraperitoneal injection of CSC significantly lowered blood urea and urine proteinuria levels over the ensuing 2 weeks. In addition, nearly two-thirds of transplanted -/- mice maintained their weight above the 80% welfare threshold, with both males and females weighing more than age-matched nontransplanted -/- mice. This was associated with less renal cortical fibrosis and interstitial inflammation compared to nontransplanted mice as shown by reduction in murine CD4, CD68, and CD45.2 cells. Transplanted CSC homed to glomeruli, where they expressed CR1, VEGFA, SYNAPTOPODIN, CD2AP, and PODOCIN at the RNA level and produced PODOCIN, CD2AP, and COLIVα3 proteins in nontransplanted -/- mice, indicating that CSC have adopted a podocyte phenotype. Together, these data indicate that CSC may be used to delay progression of renal pathology by a combination of anti-inflammatory effects and replacement of the defective resident podocytes.

  7. Quantitative fluorescent polymerase chain reaction for rapid prenatal diagnosis of fetal aneuploidies in chorionic villus sampling in a single institution

    PubMed Central

    Shin, You Jung; Kim, Do Jin; Ryu, Hyun Mee; Kim, Moon Young; Han, Jung Yeol; Choi, June Seek

    2016-01-01

    Objective To validate quantitative fluorescent polymerase chain reaction (QF-PCR) via chorionic villus sampling (CVS) for the diagnosis of fetal aneuploidies. Methods We retrospectively reviewed the medical records of consecutive pregnant women who had undergone CVS at Cheil General Hospital between December 2009 and June 2014. Only cases with reported QF-PCR before long-term culture (LTC) for conventional cytogenetic analysis were included, and the results of these two methods were compared. Results A total of 383 pregnant women underwent QF-PCR and LTC via CVS during the study period and 403 CVS specimens were collected. The indications of CVS were as follows: abnormal first-trimester ultrasonographic findings, including increased fetal nuchal translucency (85.1%), advanced maternal age (6.8%), previous history of fetal anomalies (4.2%), and positive dual test results for trisomy 21 (3.9%). The results of QF-PCR via CVS were as follows: 76 (18.9%) cases were identified as trisomy 21 (36 cases), 18 (33 cases), or 13 (seven cases), and 4 (1.0%) cases were suspected to be mosaicism. All results of common autosomal trisomies by QF-PCR were consistent with those of LTC and there were no false-positive findings. Four cases suspected as mosaicism in QF-PCR were confirmed as non-mosaic trisomies of trisomy 21 (one case) or trisomy 18 (three cases) in LTC. Conclusion QF-PCR via CVS has the advantage of rapid prenatal screening at an earlier stage of pregnancy for common chromosomal trisomies and thus can reduce the anxiety of parents. In particular, it can be helpful for pregnant women with increased fetal nuchal translucency or abnormal first-trimester ultrasonographic findings. PMID:27896246

  8. 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

  9. 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

    2016-01-01

    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.

  10. 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.

  11. Identification of Extracellular Matrix Components and Biological Factors in Micronized Dehydrated Human Amnion/Chorion Membrane.

    PubMed

    Lei, Jennifer; Priddy, Lauren B; Lim, Jeremy J; Massee, Michelle; Koob, Thomas J

    2017-02-01

    Objective: The use of bioactive extracellular matrix (ECM) grafts such as amniotic membranes is an attractive treatment option for enhancing wound repair. In this study, the concentrations, activity, and distribution of matrix components, growth factors, proteases, and inhibitors were evaluated in PURION(®) Processed, micronized, dehydrated human amnion/chorion membrane (dHACM; MiMedx Group, Inc.). Approach: ECM components in dHACM tissue were assessed by using immunohistochemical staining, and growth factors, cytokines, proteases, and inhibitors were quantified by using single and multiplex ELISAs. The activities of proteases that were native to the tissue were determined via gelatin zymography and EnzChek(®) activity assay. Results: dHACM tissue contained the ECM components collagens I and IV, hyaluronic acid, heparin sulfate proteoglycans, fibronectin, and laminin. In addition, numerous growth factors, cytokines, chemokines, proteases, and protease inhibitors that are known to play a role in the wound-healing process were quantified in dHACM. Though matrix metalloproteinases (MMPs) were present in dHACM tissues, inhibitors of MMPs overwhelmingly outnumbered the MMP enzymes by an overall molar ratio of 28:1. Protease activity assays revealed that the MMPs in the tissue existed primarily either in their latent form or complexed with inhibitors. Innovation: This is the first study to characterize components that function in wound healing, including inhibitor and protease content and activity, in micronized dHACM. Conclusion: A variety of matrix components and growth factors, as well as proteases and their inhibitors, were identified in micronized dHACM, providing a better understanding of how micronized dHACM tissue can be used to effectively promote wound repair.

  12. Pharmacokinetics of human chorionic gonadotropin after i.m. administration in goats (Capra hircus).

    PubMed

    Saleh, M; Shahin, M; Wuttke, W; Gauly, M; Holtz, W

    2012-07-01

    The present investigation addresses the pharmacokinetics of human chorionic gonadotropin (hCG), intramuscularly (i.m.) administered to goats. Nine pluriparous does of the Boer goat breed, 2-6 years of age and weighing 45-60 kg, were administered 500 IU hCG (2 ml Chorulon) deep into the thigh musculature 18 h after superovulatory FSH treatment. Blood samples were drawn from the jugular vein at 2  h intervals for the first 24h, at 6 h intervals until 42 h, and at 12 h intervals until 114 h after administration. After centrifugation, plasma hCG concentrations were determined by electrochemiluminescence immunoassay. Pharmacokinetical parameters were as follows: lag time, 0.4 (s.e.m. 0.1) h; absorption rate constant, 0.34 (s.e.m. 0.002) h; absorption half-life, 2.7 (s.e.m. 0.5) h; elimination rate constant, 0.02 (s.e.m. 0.002) h; biological half-life, 39.4 (s.e.m. 5.1) h; and apparent volume of distribution, 16.9 (s.e.m. 4.3) l. The plasma hCG profile was characterized by an absorption phase of 11.6 (s.e.m. 1.8) h and an elimination phase of 70.0 (s.e.m. 9.8) h, with considerable individual variation in bioavailability and pharmacokinetical parameters. Biological half-life was negatively correlated (P<0.05) with peak concentration (r=-0.76), absorption rate constant (r=-0.78), and elimination rate constant (r=-0.87). The results indicate that after rapid absorption, hCG remains in the circulation for an extended period. This has to be taken into account when assessing the stimulatory response to hCG treatment on an ovarian level.

  13. Gonadotropin-releasing hormone/human chorionic gonadotropin beta based recombinant antibodies and vaccines.

    PubMed

    Talwar, G P; Vyas, Hemant K; Purswani, Shilpi; Gupta, Jagdish C

    2009-12-01

    Gonadotropin-releasing hormone (GnRH) and human chorionic gonadotropin (hCG) are unique targets for the control of fertility. Immunological approaches to neutralizing these hormones have additional utility in cancer treatment. Vaccines have been developed against both GnRH and hCG and these have undergone Phase I/II clinical trials documenting their safety, reversibility and efficacy. The heterospecies dimer hCG vaccine prevented pregnancy in women of proven fertility without impairment of ovulation or derangement of menstrual regularity and bleeding profiles. The protective threshold of antibody titers to achieve efficacy was determined in these first-ever trials. Recently, a recombinant vaccine against the beta subunit of hCG linked to the B subunit of heat labile enterotoxin has been made and expressed as a glycosylated conjugate in Pichia pastoris. Experiments indicate its ability to generate antibodies above the protective threshold in all immunized Balb/c mice. Ectopic expression of hCG/hCGbeta is observed in many advanced stage cancers of various origins. A chimeric high affinity and specific recombinant antibody against hCGbeta linked to curcumin kills hCGbeta expressing T lymphoblastic leukemia cells without any deleterious effect. Several synthetic and recombinant vaccines have been developed against GnRH. These reduce serum testosterone to castration levels causing atrophy of the prostate. Three Phase I/II clinical trials conducted in India and Austria have shown that these vaccines elicit non-surgical reduction of testosterone, a fall in prostate specific antigen and clinical improvement of prostate carcinoma patients. A multimer recombinant vaccine against GnRH has high efficacy for sterilization of pigs and other animals.

  14. Effects of 5-hydroxytryptamine on human isolated placental chorionic arteries and veins.

    PubMed Central

    Reviriego, J.; Marín, J.

    1989-01-01

    1. Effects of 5-hydroxytrypamine (5-HT) on cylindrical segments of human chorionic arteries and veins were investigated. Concentrations of 5-HT (up to 3 x 10(-6) M) produced concentration-dependent contractions; higher concentrations induced a reduction of the maximal response. These responses were antagonized by methysergide and ketanserin in a non-competitive manner. The contractions elicited by low 5-HT concentrations were more affected by methysergide (10(-7) M) than by ketanserin (10(-7) M). Ketanserin apparently increased the responses to high 5-HT concentrations in veins. Arteries appeared to be more sensitive to both drugs than veins. Single concentrations of 5-HT elicited transient contractions in both kinds of vessel. 2. Marked tachyphylaxis was seen in segments exposed to high concentrations of 5-HT or in which a concentration-response curve was determined. 3. Contractions induced by 5-HT were reduced in a Ca2+-free medium. Veins were more affected by the Ca2+ antagonists, nifedipine (10(-7) M), nicardipine (10(-5) M) and diltiazem (10(-5) M) than arteries. 4. 5-HT (10(-6) M) enhanced 45Ca2+ uptake in those vessels in which a concentration-response curve had not been previously determined. In veins, this increase was reduced by the three Ca2+ antagonists. 5. The results indicate that 5-HT responses in these vessels were greatly dependent on extracellular Ca2+. A type of 5-HT1-receptor may mediate responses to low 5-HT concentrations, while higher concentrations may activate 5-HT2-receptors. 5-HT may desensitize the latter by interconversion between a high affinity and low affinity state, as suggested by others, a change prevented in part by ketanserin. PMID:2743086

  15. Immune Modulatory Effects of Human Chorionic Gonadotropin on Dendritic Cells Supporting Fetal Survival in Murine Pregnancy

    PubMed Central

    Dauven, Dominique; Ehrentraut, Stefanie; Langwisch, Stefanie; Zenclussen, Ana Claudia; Schumacher, Anne

    2016-01-01

    Dendritic cells (DCs) are critically involved in the determination of immunity vs. tolerance. Hence, DCs are key regulators of immune responses either favoring or disfavoring fetal survival. Several factors were proposed to modulate DC phenotype and function during pregnancy. Here, we studied whether the pregnancy hormone human chorionic gonadotropin (hCG) is involved in DC regulation. In vitro, bone marrow-derived DCs (BMDCs) were stimulated in the presence or absence of urine-purified or recombinant hCG (rhCG) preparations. Subsequently, BMDC maturation was assessed. Cytokine secretion of activated BMDCs and their capability to enforce TH1, TH2, TH17, or Treg cell differentiation was determined after rhCG treatment. Moreover, the in vivo potential of hCG-modulated BMDCs to influence pregnancy outcome, Treg cell number, and local cytokine expression was evaluated after adoptive transfer in a murine abortion-prone model before and after conception. Both hCG preparations impaired the maturation process of BMDCs. rhCG treatment did neither alter cytokine secretion by BMDCs nor their ability to drive TH1, TH2, or TH17 differentiation. rhCG-treated BMDCs augmented the number of Treg cells within the T cell population. Adoptive transfer of rhCG-treated BMDCs after conception did not influence pregnancy outcome. However, transfer of hCG-treated BMDCs prior to mating had a protective effect on pregnancy. This positive effect was accompanied by increased Treg cell numbers and decidual IL-10 and TGF-β expression. Our results unveil the importance of hCG in retaining DCs in a tolerogenic state, thereby promoting Treg cell increment and supporting fetal survival. PMID:27895621

  16. Identification of Extracellular Matrix Components and Biological Factors in Micronized Dehydrated Human Amnion/Chorion Membrane

    PubMed Central

    Lei, Jennifer; Priddy, Lauren B.; Lim, Jeremy J.; Massee, Michelle; Koob, Thomas J.

    2017-01-01

    Objective: The use of bioactive extracellular matrix (ECM) grafts such as amniotic membranes is an attractive treatment option for enhancing wound repair. In this study, the concentrations, activity, and distribution of matrix components, growth factors, proteases, and inhibitors were evaluated in PURION® Processed, micronized, dehydrated human amnion/chorion membrane (dHACM; MiMedx Group, Inc.). Approach: ECM components in dHACM tissue were assessed by using immunohistochemical staining, and growth factors, cytokines, proteases, and inhibitors were quantified by using single and multiplex ELISAs. The activities of proteases that were native to the tissue were determined via gelatin zymography and EnzChek® activity assay. Results: dHACM tissue contained the ECM components collagens I and IV, hyaluronic acid, heparin sulfate proteoglycans, fibronectin, and laminin. In addition, numerous growth factors, cytokines, chemokines, proteases, and protease inhibitors that are known to play a role in the wound-healing process were quantified in dHACM. Though matrix metalloproteinases (MMPs) were present in dHACM tissues, inhibitors of MMPs overwhelmingly outnumbered the MMP enzymes by an overall molar ratio of 28:1. Protease activity assays revealed that the MMPs in the tissue existed primarily either in their latent form or complexed with inhibitors. Innovation: This is the first study to characterize components that function in wound healing, including inhibitor and protease content and activity, in micronized dHACM. Conclusion: A variety of matrix components and growth factors, as well as proteases and their inhibitors, were identified in micronized dHACM, providing a better understanding of how micronized dHACM tissue can be used to effectively promote wound repair. PMID:28224047

  17. The biochemical properties of urinary human chorionic gonadotropin from the patients with trophoblastic disease.

    PubMed

    Nishimura, R; Endo, Y; Tanabe, K; Ashitaka, Y; Tojo, S

    1981-01-01

    Human chorionic gonadotropin (hCG) was extracted and purified from urine of normal pregnant women and patients with hydatidiform mole and choriocarcinoma using the sam methods. Both hCG-hydatidiform mole and hCG-choriocarcinoma as well as hCG-normal pregnancy was separated into alpha and beta subunits by SDS disc electrophoresis upon treatment with 2-mercaptoethanol and showed the same immunoreactivities against anti-hCG, -alpha hCG, and -beta hCG as hCG in each radioimmunoassay. In vivo bioassay, bioactivities of hCG- normal pregnancy and hCG-hydatidiform mole were approximately 7,000 IU/mg (2nd IS), while that of hCG--choriocarcinoma was only 400 IU/mg. Conversely, the receptor binding activities in vitro of hCG-chorio carcinoma was about 3 times more effective than the other 2. Although the amino acid composition of these hCG preparations were practically identical, a great difference in the carbohydrate composition was observed. The significant difference was that while sialic acid was undetectable in hCG-choriocarcinoma approximately 8.5% of sialic acid was found in hCG-normal pregnancy and hCG-hydatidiform mole. A parallel finding was that iodinated hCG-choriocarcinoma was taken up in large quantities by the liver in comparison to the ovary which differed from that observed with hCG-normal pregnancy and hCG-hydatidiform mole in Parlow rats. The present findings support the thesis that neoplastic or malignant transformation of trophoblasts may result in an alteration of the glycosylation process, especially the sialylation, in the biosynthesis of hCG rather than the translation steps.

  18. 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.

  19. Late-onset hypogonadism: the advantages of treatment with human chorionic gonadotropin rather than testosterone.

    PubMed

    La Vignera, Sandro; Condorelli, Rosita Angela; Cimino, Laura; Russo, Giorgio Ivan; Morgia, Giuseppe; Calogero, Aldo E

    2016-01-01

    The traditional pharmacological treatment of patients with late onset hypogonadism (LOH) is represented by different formulations of testosterone (T) or alternatively by the extractive human chorionic gonadotropin (HCG). The hormone replacement treatment (HRT) is associated with the potential increase of hematocrit, serum concentrations of prostate-specific antigen (PSA) and prostate volume. Moreover, the gynecomastia represent a condition frequently associated with HRT. Recent evidences showed the role of leydig cells in the 25-hydroxylation of vitamin D and the elevated frequency of hypovitaminosis D among LOH patients. Finally, another important aspect of LOH is represented by the frequency of secondary infertility due to age or to traditional HRT. This study evaluated 40 LOH patients treated for 6 months with extractive HCG (n = 10 patients) and three different formulations of T: transdermal (n = 10 patients), undecaonate (n = 10 patients) and enantate (n = 10 patients). Hormonal, anthropometric, metabolic and sperm parameters were evaluated and compared. Moreover, the main safety parameters and the results of the main questionnaires were evaluated. After treatment, HCG group showed serum concentrations of 25-OH-vitamin D significantly higher (p < 0.05) and serum concentrations of oestrogens significantly lower (p < 0.05) compared with other groups. Moreover, they showed a mean value of hematocrit, PSA and prostate volume significantly lower (p < 0.05) compared with other groups. Finally, all the groups treated with T showed a significant reduction (p < 0.05) of sperm density and of percentage of spermatozoa with progressive motility compared with HCG group.

  20. 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

  1. Analysis of human luteinizing hormone and human chorionic gonadotropin preparations of different origins by reversed-phase high-performance liquid chromatography.

    PubMed

    Almeida, B E; Oliveira, J E; Carvalho, C M; Dalmora, S L; Bartolini, P; Ribela, M T C P

    2010-09-21

    Specific reversed-phase high-performance liquid chromatography conditions are reported for the analysis of recombinant and native human luteinizing hormone (hLH) and human chorionic gonadotropin (hCG) preparations. Heterodimeric hLH, hCG and their alpha- and beta-subunits migrated with significantly different retention times (t(R)) in the following order of increasing hydrophobicity: alpha-hCGQuantitative analysis was validated for the homogeneous preparations and a highly linear dose-response curve (r=0.99998; p<0.0001; n=20) used to assess the accuracy, precision and sensitivity of the analysis. Quantification of the different gonadotropins in the heterogeneous preparations was also carried out, but with limitations in accuracy.

  2. Intratubular trophoblasts in the contralateral testis caused elevation of serum human chorionic gonadotropin following complete remission of stage II testicular tumor: a case report.

    PubMed

    Nitta, Satoshi; Kawai, Koji; Onozawa, Mizuki; Ando, Satoshi; Miyazaki, Jun; Nagata, Chigusa; Noguchi, Masayuki; Yamasaki, Kazumitsu; Uchida, Katsunori; Iwamoto, Teruaki; Nishiyama, Hiroyuki

    2013-01-01

    We report the case of a 22-year-old male who had a history of metastatic right testicular tumor successfully treated with chemotherapy and surgery. Twenty-one months after the initial treatment, the serum human chorionic gonadotropin started to increase gradually, but whole body imaging including the left testis revealed no abnormal finding except testicular microlithiasis. A biopsy of the left testis revealed intratubular germ cell neoplasia, unclassified type. After the human chorionic gonadotropin level reached 6.6 mIU/ml, he underwent left high orchiectomy. Histology demonstrated a small malignant germ cell tumor as well as intratubular germ cell neoplasia, unclassified type, both of which were negative for human chorionic gonadotropin staining. Besides these lesions, there were tiny foci of human chorionic gonadotropin-immunoreactive intratubular trophoblasts. Serum human chorionic gonadotropin normalized immediately after the orchiectomy, and he had no sign of recurrence at 6 months. The present case will provide new insight into the diagnosis of testicular tumor recurrence with isolated elevation of a serum tumor marker.

  3. 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

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

    PubMed

    Cole, Laurence A

    2009-01-26

    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

  5. 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.

  6. Development of dual-enzyme-based simultaneous immunoassay for measurement of progesterone and human chorionic gonadotropin.

    PubMed

    Basu, Anupam; Maitra, Saumen Kumar; Shrivastav, Tulsidas G

    2007-07-15

    The development of a simultaneous multianalyte immunoassay for the detection of progesterone and human chorionic gonadotropin (hCG) in serum is described. In this simultaneous multianalyte assay, two different enzymes, viz. horse radish peroxidase (HRP) and alkaline phosphatase (ALP), were used as markers. To the simultaneous immobilized progesterone and hCG antibody microwells, 50 microL of different concentrations of combined standards or serum samples was added in duplicate and then 100 microL of combined conjugate reagent, composed of 17-alpha-OH-P-ALP and hCG-biotin was added to all the wells and incubated for 1h at 37 degrees C. After incubation, the contents of the wells were decanted and washed thoroughly with running tap water. After washing, 100 microL alkaline phosphatase substrate along with streptavidin-horseradish peroxidase was added to all the wells and incubated for 0.5 h at 37 degrees C. After incubation, the developed color was measured at 405 nm. The absorbency at this stage provides the result for the progesterone assay. The contents of the wells were decanted and washed. In the next step, 100 microL of tetramethylbenzidene/H2O2 reagent was added to all the wells. After 15 min of incubation, 100 microL of 0.5 M H2SO4 was added to all the wells and the color was read at 450 nm. The absorbency at this stage provides the result for the hCG assay. Sensitivity of the progesterone and hCG assays were 0.118 ng/ml and 0.124 IU/ml respectively. Intra- and inter assay percentage coefficients of variation ranged from 1.8 to 7.1 and 9.1 to 11.5 for progesterone and from 2.1 to 10.4 and 7.2 to 11.3 for hCG. There was good correlation between the discrete and the simultaneous assays. For progesterone assay, R2 was 0.99 and for hCG R2 was also 0.99. The developed dual assay for progesterone and hCG may be useful for the diagnosis of abnormal pregnancies such as miscarriages and ectopic pregnancies.

  7. 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.

  8. 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.

  9. Gossypol inhibits human chorionic gonadotropin-stimulated testosterone production by cultured canine testicular interstitial cells.

    PubMed

    Mushtaq, M; Kulp, S; Chang, W; Lin, Y C

    1996-03-01

    Gossypol (GP) is a natural polyphenolic compound that possesses antifertility and antisteroidogenic activities in both males and females. The dog is highly sensitive to GP toxicity, yet GP's effect on canine testicular steroidogenesis has never been reported. Thus, the present study examines GP's effects on human chorionic gonadotropin (hCG)-induced testosterone (T) production by primary cultured canine testicular interstitial cells. After decapsulation and enzymatic dissociation of canine testes in Dulbecco's Modified Eagle Medium with Ham's Nutrient Mixture F-12 (1:1; DME/F-12) containing 0.1% collagenase, 0.1% BSA, and 10 micrograms/ml DNase 1 (37 degrees C, 20 min), interstitial cells were isolated by sedimentation and filtration (140 microns) and then cultured in supplemented DME/F-12 medium (5 micrograms/ml insulin, 5 micrograms/ml transferrin, 5 ng/ml sodium selenite; DME/F-12/S) containing 0.1% fetal bovine serum (FBS). FBS was used to enhance cell attachment during the first 24 hours of culture. After 24 hours, the medium was replaced with serum-free DME/F-12/S and the cells were cultured for an additional 24 hours. Thereafter, cells were treated with hCG (0.1 IU/ml) alone and in combination with GP (0.05, 0.5, 2.5 and 5.0 microM). Media were collected for T radioimmunoassay and cells for protein estimation after 8, 16 and 24 hours of treatment. Treatment with hCG significantly (p < 0.05) stimulated T production over that of controls at all treatment times examined. At 8, 16 and 24 hours, T secretion was elevated from 0.91 +/- 0.25, 1.32 +/- 0.42, and 1.41 +/- 0.40 pg/microgram protein to 2.36 +/- 0.50, 2.84 +/- 0.60, and 2.82 +/- 0.43 pg/microgram protein, respectively. At 0.5, 2.5 and 5.0 microM, GP significantly (p < 0.05) reduced hCG-induced T secretion at 16 and 24 hours of treatment to 1.79 +/- 0.50, 1.62 +/- 0.12, 1.34 +/- 0.16 (16 hr), and 1.53 +/- 0.38, 1.43 +/- 0.11, 1.42 +/- 0.32 (24 hr) pg/microgram protein, respectively. At 8 hours, T

  10. Differential expression profile of long noncoding RNAs in human chorionic villi of early recurrent miscarriage.

    PubMed

    Wang, Leilei; Tang, Huaiyun; Xiong, Yun; Tang, Lisha

    2017-01-01

    Miscarriage is the spontaneous loss of a pregnancy before the fetus reaching viability, including all pregnancy losses from the time of conception until 24weeks of gestation. Three or more times of consecutive spontaneous miscarriages is defined as recurrent miscarriage (RM). The underlying causes cannot be confirmed in nearly 50% of RM patients. We investigated the differential expression of long noncoding RNAs (lncRNAs) in chorionic villi tissues of RM patients compared with normal women, and their possible involvement in the pathogenic pathways leading to RM. A total number of 1449 differentially expressed lncRNAs were identified from chorionic villi tissues of RM patients compared to normal women. KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis revealed that these differentially expressed lncRNAs were involved in 26 biological pathways including 11 up-regulated and 15 down-regulated ones. Functional analysis showed that pathways of endocrine, immunity, ECM-receptor interactions and apoptosis are the major pathogenic mechanisms involved in the development of RM. Characterization of these lncRNAs in different pathways opened a new starting point for further investigating the epigenetic regulation mechanisms of lncRNAs in RM.

  11. Oxygen tension and normalisation pressure modulate nifedipine-sensitive relaxation of human placental chorionic plate arteries.

    PubMed

    Cooper, E J; Wareing, M; Greenwood, S L; Baker, P N

    2006-01-01

    Fetoplacental blood vessel constriction in response to reduced oxygenation has been demonstrated in placenta perfused in vitro. In pulmonary vessels, hypoxic vasoconstriction involves Ca2+ influx into smooth muscle through membrane ion channels including voltage-gated Ca2+ channels (VGCCs). We hypothesised that VGCCs are involved in agonist-induced constriction of fetoplacental resistance vessels and that their contribution is modulated by oxygen. Chorionic plate small arteries were studied using wire myography. Arteries were normalised at high (0.9 of L(13.3 kPa)) or low (0.9 of L(5.1 kPa)) stretch and experiments performed at 156, 38 or 15 mmHg oxygen. At low stretch, U46619 (thromboxane-mimetic) or KCl (smooth muscle depolarisation) constriction was greater at 38 than 156 or 15 mmHg oxygen. An L-type VGCC blocker nifedipine, inhibited KCl constriction by >85% but was less effective in U46619 constrictions (43-67%). At high stretch, nifedipine inhibition of KCl- and U46619-induced constriction was less at 15 than 38 or 156 mmHg oxygen. Oxygen did not affect constriction to U46619 or nifedipine-induced relaxation when vessels were normalised at high stretch. In conclusion, oxygen modulates chorionic plate arterial constriction at low stretch but regulation is lost at high stretch. U46619 constriction is underlain by VGCCs and nifedipine-insensitive processes; their relative contribution is influenced by oxygen.

  12. 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.

  13. Segregation patterns of polymorphic restriction sites of the gene encoding the alpha subunit of human chorionic gonadotropin in trophoblastic disease.

    PubMed Central

    Hoshina, M; Boothby, M R; Hussa, R D; Pattillo, R A; Camel, H M; Boime, I

    1984-01-01

    The gene encoding the alpha subunit of human chorionic gonadotropin contains at least two polymorphic sites in its 3' flanking region detected by restriction enzymes HindIII and EcoRI. We used these polymorphic sites as markers of tissue genotype in normal placenta, hydatidiform mole, choriocarcinoma, and peripheral leukocytes. As expected, inheritance patterns of most hydatidiform moles showed only a paternal genetic contribution. However, one uncommon DNA polymorphism pattern, homozygosity for the absence of the EcoRI site and the presence of the HindIII site, predominated in choriocarcinoma. Thus, our results suggest that moles which have this uncommon polymorphism pattern appear particularly likely to develop into choriocarcinoma. Images PMID:6201859

  14. A novel reverse fluorescent immunoassay approach for sensing human chorionic gonadotropin based on silver-gold nano-alloy and magnetic nanoparticles.

    PubMed

    Huang, Xiaopeng; Li, Yuqin; Huang, Xiang; Xie, Xiaona; Xu, Yanping; Chen, Yaowen; Gao, Wenhua

    2016-01-01

    A novel and environmentally friendly reverse fluorescent immunoassay approach was proposed and utilized for sensing human chorionic gonadotropin (HCG) in human serum by coupling a newly prepared and highly fluorescent glutathione-stabilized silver-gold nano-alloy (GSH-AgAuNAs) with magnetic nanoparticles (MNPs). To construct such a reverse system, fluorescent GSH-AgAuNAs and MNPs were first prepared and bio-functionalized with monoclonal antibodies (Mab-I and Mab-II) toward HCG antigen, respectively. Then, the GSH-AgAuNAs functionalized with Mab-I were incubated with HCG, followed by the addition of MNPs attached to Mab-II. Thereafter, a sandwich-type immunoassay could be constructed for determination of HCG owing to the antibody-antigen recognition between the functionalized GSH-AgAuNAs and MNPs. Afterwards, a magnetic collection was employed. Hence, the amount of GSH-AgAuNAs would be reduced through an immuno-magnetic separation, thus weakening the fluorescent intensity. Different from conventional immunoassay, our work determined the quantitative signal by measuring the decreasing gradient fluorescent intensity. Under optimal conditions, the developed reverse method exhibited a wide linear range of 0.5-600 ng mL(-1) toward HCG with a detection limit of 0.25 ng mL(-1). Additionally, the proposed immunoassay was validated using spiked samples, illustrating a satisfactory result in practical application.

  15. Label-free electrochemical immunosensor based on gold-silicon carbide nanocomposites for sensitive detection of human chorionic gonadotrophin.

    PubMed

    Yang, Long; Zhao, Hui; Fan, Shuangmei; Deng, Shuangsheng; Lv, Qi; Lin, Jie; Li, Can-Peng

    2014-07-15

    Uniform and highly dispersed gold-silicon carbide (Au@SiC) nanocomposites were prepared via simple way and used for fabrication of label-free electrochemical immunosensor for sensitive detection of human chorionic gonadotrophin (hCG). Using Au@SiC as electrode material and using ferricyanide as mediator, the proposed immunosensor provides a simple and economic method with higher sensitivity and a wider concentration range for detection of hCG. Under the optimal condition, the approach provided a good linear response range from 0.1 to 5 IU/L and 5 to 1000 IU/L with a low detection limit of 0.042 IU/L. The immunosensor showed good selectivity, acceptable stability and reproducibility. Satisfactory results were obtained for determination of hCG in human serum samples. The proposed method provides a promising platform of clinical immunoassay for other biomolecules. In addition, the bio-functionalization of SiC combined with other nanomaterials will provide promising approach for electrochemical sensing and biosensing platform.

  16. 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.

  17. Human chorionic gonadotropin and its free β-subunit stimulate trophoblast invasion independent of LH/hCG receptor.

    PubMed

    Lee, Cheuk-Lun; Chiu, Philip C N; Hautala, Laura; Salo, Tuula; Yeung, William S B; Stenman, Ulf-Håkan; Koistinen, Hannu

    2013-08-15

    Both paracrine and autocrine factors are involved in the regulation of trophoblast invasion. One of these factors is human chorionic gonadotropin (hCG), which stimulates trophoblast invasion. The stimulatory activity has especially been ascribed to a hyperglycosylated form of hCG (hCG-h) that is expressed in early pregnancy. We compared the stimulatory activities of different forms of hCG and its free β-subunit (hCGβ) on trophoblast invasion. hCG, hCG-h, hCGβ, and its hyperglycosylated form (hCGβ-h) stimulated the invasion of JEG-3 choriocarcinoma cells. The stimulatory effect of hCGβ was also confirmed with primary human trophoblasts. Down-regulation of the LH/hCG receptor by RNA-interference did not significantly reduce the effect of hCGβ and hCG on cell invasion. Increased invasion was associated with increased levels of MMP-2, MMP-9 and activity of uPA. Our findings suggest that hCG, hCGβ and their hyperglycosylated forms stimulate the invasion of trophoblast cells independent of the classical LH/hCG-receptor.

  18. 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.

  19. 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.

  20. 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

  1. 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.

  2. A 15-year-old female with amenorrhea, abdominal distention, and elevated human chorionic gonadotropin: pregnancy, right? Not so fast….

    PubMed

    Aggarwal, Arun; Ocon, Anthony J; Nibhanipudi, Kumara

    2012-10-01

    Nongestational choriocarcinoma, a rare ovarian tumor, may present in young women with amenorrhea, abdominal distention, and elevated urine human chorionic gonadotropin (hCG), all of which may be mistaken for pregnancy. A 15-year-old Hispanic female, who reported no sexual activity, presented with 6 months of amenorrhea, abdominal pain, and progressive abdominal distension. Initially, suspicion of pregnancy was considered. Physical examination was significant for abdominal distension, but no uterine fundus or fetal anatomy could be palpated, and auscultation did not reveal any fetal heart sounds or bruits. Laboratory values showed elevated urine hCG, cancer antigen 125, and cancer antigen 19.9 levels but normal serum hCG level and was inconsistent with pregnancy. Computed tomographic scans revealed a large abdominal heterogeneous mass and pleural effusions. Salpingo-oophorectomy with total omentectomy and inversion appendectomy removed a 21 × 20.5 × 16.5-cm tumor. Pathological testing determined it to be a nongestational choriocarcinoma. This rare tumor is more common in the pediatric adolescent population than in adults. Surgical resection and chemotherapy often result in a positive prognosis. In female adolescent patients presenting with elevated hCG level, amenorrhea, and abdominal distention, choriocarcinoma should be considered, especially in those with no history of sexual activity or before menarche.

  3. 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

  4. A simple and sensitive immunoassay for the determination of human chorionic gonadotropin by graphene-based chemiluminescence resonance energy transfer.

    PubMed

    Lei, Jiuqian; Jing, Tao; Zhou, Tingting; Zhou, Yusun; Wu, Wei; Mei, Surong; Zhou, Yikai

    2014-04-15

    In this study, we report a strategy of chemiluminescence resonance energy transfer (CRET) using graphene as an efficient long-range energy acceptor. Magnetic nanoparticles were also used in CRET for simple magnetic separation and immobilization of horseradish peroxidase (HRP)-labeled anti-HCG antibody. In the design of CRET system, the sandwich-type immunocomplex was formed between human chorionic gonadotropin (HCG, antigen) and two different antibodies bridged the magnetic nanoparticles and graphene (acceptors), which led to the occurrence of CRET from chemiluminescence light source to graphene. After optimizing the experimental conditions, the quenching of chemiluminescence signal depended linearly on the concentration of HCG in the range of 0.1 mIU mL(-1)-10 mIU mL(-1) and the detection limit was 0.06 mIU mL(-1). The proposed method was successfully applied for the determination of HCG levels in saliva and serum samples, and the results were in good agreement with the plate ELISA with colorimetric detection. It could also be developed for detection of other antigen-antibody immune complexes by using the corresponding antigens and respective antibodies.

  5. Elevated hypothalamic aromatization at the onset of precocious puberty in transgenic female mice hypersecreting human chorionic gonadotropin: effect of androgens.

    PubMed

    Gonzalez, Betina; Ratner, Laura D; Scerbo, María J; Di Giorgio, Noelia P; Poutanen, Matti; Huhtaniemi, Ilpo T; Calandra, Ricardo S; Lux-Lantos, Victoria A R; Cambiasso, María J; Rulli, Susana B

    2014-06-05

    Transgenic female mice overexpressing the α- and β- subunits of human chorionic gonadotropin (hCGαβ+) exhibited precocious puberty, as evidenced by early vaginal opening. Chronically elevated hCG in 21-day-old hCGαβ+ females stimulated gonadal androgen production, which exerted negative feedback over the endogenous gonadotropin synthesis, and activated the hypothalamic GnRH pulsatility and gene expression. Transgenic females also exhibited elevated hypothalamic aromatization in the preoptic area (POA), which is the sexually-differentiated area that controls the LH surge in adulthood. Ovariectomy at 14 days of age was unable to rescue this phenotype. However, the blockade of androgen action by flutamide from postnatal day 6 onwards reduced the aromatase levels in the POA of hCGαβ+ females. Our results suggest that early exposure of females to androgen action during a critical period between postnatal days 6-14 induces sex-specific organizational changes of the brain, which affect the aromatase expression in the POA at the onset of precocious puberty.

  6. 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.

  7. No evidence of the human chorionic gonadotropin-beta gene 5 betaV79M polymorphism in Mexican women.

    PubMed

    Dominguez-Lopez, Pablo; Diaz-Cueto, Laura; Ulloa-Aguirre, Alfredo; Lopez-Valle, Miguel Angel; Arechavaleta-Velasco, Fabian

    2008-01-01

    Human chorionic gonadotropin (hCG) is a placental hormone essential for the maintenance of pregnancy. Previous studies have shown a G to A transition in exon 3 of the hCGbeta gene 5, which changes the naturally occurring valine to methionine in codon 79. The frequency of this transition varies among different ethnic groups, being high in USA women, and less common, or absent, in various European populations. The purpose of the present study was to determine the frequency of the betaV79M allelic variant of the beta-subunit of hCG in a Mexican population, and to compare this frequency with those found in other ethnic groups. Placental DNA from 161 pregnant Mexican women was genotyped for the betaV79M by polymerase chain reaction (PCR)-restriction fragments length polymorphism analysis. No polymorphic betaV79M alleles were identified in the population studied. The allele and genotypic frequencies of betaV79M polymorphism in Mexican Mestizo women were significantly different from those reported for the US population, but not from five different European populations. In contrast to what has been found in women from the USA, it seems that the hCGbeta V79M polymorphism is absent or extremely rare in Mexican Mestizo women.

  8. Human chorionic gonadotropin administration is associated with high pregnancy rates during ovarian stimulation and timed intercourse or intrauterine insemination

    PubMed Central

    Mitwally, Mohamed F; Abdel-Razeq, Sonya; Casper, Robert F

    2004-01-01

    Background There are different factors that influence treatment outcome after ovarian stimulation and timed-intercourse or intrauterine insemination (IUI). After patient age, it has been suggested that timing of insemination in relation to ovulation is probably the most important variable affecting the success of treatment. The objective of this study is to study the value of human chorionic gonadotropin (hCG) administration and occurrence of luteinizing hormone (LH) surge in timing insemination on the treatment outcome after follicular monitoring with timed-intercourse or intrauterine insemination, with or without ovarian stimulation. Methods Retrospective analysis of 2000 consecutive completed treatment cycles (637 timed-intercourse and 1363 intrauterine insemination cycles). Stimulation protocols included clomiphene alone or with FSH injection, letrozole (an aromatase inhibitor) alone or with FSH, and FSH alone. LH-surge was defined as an increase in LH level ≥200% over mean of preceding two days. When given, hCG was administered at a dose of 10,000 IU. The main outcome was clinical pregnancy rate per cycle. Results Higher pregnancy rates occurred in cycles in which hCG was given. Occurrence of an LH-surge was associated with a higher pregnancy rate with clomiphene treatment, but a lower pregnancy rate with FSH treatment. Conclusions hCG administration is associated with a favorable outcome during ovarian stimulation. Awaiting occurrence of LH-surge is associated with a better outcome with CC but not with FSH treatment. PMID:15239837

  9. Final Oocyte Maturation in Assisted Reproduction with Human Chorionic Gonadotropin and Gonadotropin-releasing Hormone agonist (Dual Trigger)

    PubMed Central

    de Oliveira, Sofia Andrade; Calsavara, Vinícius Fernando; Cortés, Gemma Castillón

    2016-01-01

    Final oocyte maturation with Human Chorionic Gonadotropin (hCG) and ovarian stimulation with Follicle Stimulation Hormone (FSH) combined with Gonadotrophin-releasing Hormone (GnRH) antagonist to block Luteinizing hormone (LH) surge is a standard procedure of in vitro Fertilization (IVF) and Intracytoplasmic Sperm Injection (ICSI). However, GnRH agonist has been replacing the use of hCG in certain situations, especially in patients at risk of Ovarian Hyperstimulation Syndrome (OHSS). Some studies have also shown advantages in the combined use of GnRH agonist concurrently with hCG in inducing final oocyte maturation, a treatment known as "Dual Trigger". In theory, this method combines the advantages of both induction regimens, and it has brought promising results. The objective of this study is to compare Dual Trigger with the use of hCG alone or the use of GnRH agonist alone. A systematic review of articles on Dual Trigger and a retrospective cohort study comparing the three methods of induction of final oocyte maturation have been conducted. It has been found that Dual Triggering for poor responder patients had a statistically significant increase in the number of retrieved oocytes, mature oocytes, and fertilized embryos in the positive beta hCG rate, implantation rate, and newborn/transferred embryo (TE) rate. PMID:28050961

  10. Final Oocyte Maturation in Assisted Reproduction with Human Chorionic Gonadotropin and Gonadotropin-releasing Hormone agonist (Dual Trigger).

    PubMed

    Oliveira, Sofia Andrade de; Calsavara, Vinícius Fernando; Cortés, Gemma Castillón

    2016-12-01

    Final oocyte maturation with Human Chorionic Gonadotropin (hCG) and ovarian stimulation with Follicle Stimulation Hormone (FSH) combined with Gonadotrophin-releasing Hormone (GnRH) antagonist to block Luteinizing hormone (LH) surge is a standard procedure of in vitro Fertilization (IVF) and Intracytoplasmic Sperm Injection (ICSI). However, GnRH agonist has been replacing the use of hCG in certain situations, especially in patients at risk of Ovarian Hyperstimulation Syndrome (OHSS). Some studies have also shown advantages in the combined use of GnRH agonist concurrently with hCG in inducing final oocyte maturation, a treatment known as "Dual Trigger". In theory, this method combines the advantages of both induction regimens, and it has brought promising results. The objective of this study is to compare Dual Trigger with the use of hCG alone or the use of GnRH agonist alone. A systematic review of articles on Dual Trigger and a retrospective cohort study comparing the three methods of induction of final oocyte maturation have been conducted. It has been found that Dual Triggering for poor responder patients had a statistically significant increase in the number of retrieved oocytes, mature oocytes, and fertilized embryos in the positive beta hCG rate, implantation rate, and newborn/transferred embryo (TE) rate.

  11. Toxic and therapeutic effects of Nifurtimox and Benznidazol on Trypanosoma cruzi ex vivo infection of human placental chorionic villi explants.

    PubMed

    Rojo, Gemma; Castillo, Christian; Duaso, Juan; Liempi, Ana; Droguett, Daniel; Galanti, Norbel; Maya, Juan Diego; López-Muñoz, Rodrigo; Kemmerling, Ulrike

    2014-04-01

    Nifurtimox (Nfx) and Benznidazole (Bnz) are the only available drugs in use for the treatment of Chagas disease. These drugs are recommended but not fully validated in evidence-based medicine and reports about the differential toxicity of both drugs are controversial. Here, we evaluated the toxic and therapeutic effects of Nfx and Bnz on human placental chorionic villi explants (HPCVE) during ex vivo infection of Trypanosoma cruzi, performing histopathological, histochemical, immunohistochemical as well as immunofluorescence analysis of the tissue. Additionally, we determined the effect of both drugs on parasite load by real time PCR. Bnz prevents the parasite induced tissue damage in ex vivo infected HPCVE compared to Nfx, which is toxic per se. The presence of T. cruzi antigens and DNA in infected explants suggests that these drugs do not impair parasite invasion into the HPCVE. Additionally, our results confirm reports suggesting that Bnz is less toxic than Nfx and support the need for the development of more effective and better-tolerated drugs.

  12. 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

  13. Endometrial cysteine-rich secretory protein 3 is inhibited by human chorionic gonadotrophin, and is increased in the decidua of tubal ectopic pregnancy

    PubMed Central

    Horne, A.W.; Duncan, W.C.; King, A.E.; Burgess, S.; Lourenco, P.C.; Cornes, P.; Ghazal, P.; Williams, A.R.; Udby, L.; Critchley, H.O.D.

    2009-01-01

    Ectopic pregnancy (EP) remains a considerable cause of morbidity and occasional mortality. Currently, there is no reliable test to differentiate ectopic from intrauterine gestation. We have previously used array technology to demonstrate that differences in gene expression in decidualized endometrium from women with ectopic and intrauterine gestations could be used to identify candidate diagnostic biomarkers for EP. The aim of this study was to further investigate the decidual gene with the highest fold increase in EP, cysteine-rich secretory protein-3 (CRISP-3). Decidualized endometrium from gestation-matched women undergoing surgical termination of pregnancy (n = 8), evacuation of uterus for miscarriage (n = 6) and surgery for EP (n = 11) was subjected to quantitative RT–PCR, morphological assessment, immunohistochemistry and western blot analysis. Sera were analysed for progesterone and human chorionic gonadotrophin (hCG) levels. Immortalized endometrial epithelial cells were cultured with physiological concentrations of hCG. CRISP-3 mRNA and protein expression were greater in endometrium from ectopic when compared with intrauterine pregnancies (P < 0.05). CRISP-3 protein was localized to epithelium and granulocytes of endometrium. CRISP-3 serum concentrations were not different in women with ectopic compared with intrauterine pregnancies. CRISP-3 expression in endometrium was not related to the degree of decidualization or to serum progesterone levels. Endometrial CRISP-3 expression was inversely proportional to serum hCG concentrations (P < 0.001). Stimulation of endometrial epithelial cells with hCG in vitro caused a reduction in CRISP-3 expression (P < 0.01). The measurement of CRISP-3 in endometrium could provide an additional tool in the diagnosis of failing early pregnancy of unknown location. The absence of a local reduction in expression of CRISP-3 in decidualized endometrium of women with EP may be due to reduced exposure to hCG due to the ectopic

  14. Endometrial cysteine-rich secretory protein 3 is inhibited by human chorionic gonadotrophin, and is increased in the decidua of tubal ectopic pregnancy.

    PubMed

    Horne, A W; Duncan, W C; King, A E; Burgess, S; Lourenco, P C; Cornes, P; Ghazal, P; Williams, A R; Udby, L; Critchley, H O D

    2009-05-01

    Ectopic pregnancy (EP) remains a considerable cause of morbidity and occasional mortality. Currently, there is no reliable test to differentiate ectopic from intrauterine gestation. We have previously used array technology to demonstrate that differences in gene expression in decidualized endometrium from women with ectopic and intrauterine gestations could be used to identify candidate diagnostic biomarkers for EP. The aim of this study was to further investigate the decidual gene with the highest fold increase in EP, cysteine-rich secretory protein-3 (CRISP-3). Decidualized endometrium from gestation-matched women undergoing surgical termination of pregnancy (n = 8), evacuation of uterus for miscarriage (n = 6) and surgery for EP (n = 11) was subjected to quantitative RT-PCR, morphological assessment, immunohistochemistry and western blot analysis. Sera were analysed for progesterone and human chorionic gonadotrophin (hCG) levels. Immortalized endometrial epithelial cells were cultured with physiological concentrations of hCG. CRISP-3 mRNA and protein expression were greater in endometrium from ectopic when compared with intrauterine pregnancies (P < 0.05). CRISP-3 protein was localized to epithelium and granulocytes of endometrium. CRISP-3 serum concentrations were not different in women with ectopic compared with intrauterine pregnancies. CRISP-3 expression in endometrium was not related to the degree of decidualization or to serum progesterone levels. Endometrial CRISP-3 expression was inversely proportional to serum hCG concentrations (P < 0.001). Stimulation of endometrial epithelial cells with hCG in vitro caused a reduction in CRISP-3 expression (P < 0.01). The measurement of CRISP-3 in endometrium could provide an additional tool in the diagnosis of failing early pregnancy of unknown location. The absence of a local reduction in expression of CRISP-3 in decidualized endometrium of women with EP may be due to reduced exposure to hCG due to the ectopic

  15. 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...

  16. 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...

  17. Associations between maternal serum free beta human chorionic gonadotropin (β-hCG) levels and adverse pregnancy outcomes.

    PubMed

    Sirikunalai, P; Wanapirak, C; Sirichotiyakul, S; Tongprasert, F; Srisupundit, K; Luewan, S; Traisrisilp, K; Tongsong, T

    2016-01-01

    The objective was to determine the strength of relationship between maternal free beta human chorionic gonadotropin (β-hCG) concentrations and rates of adverse pregnancy outcomes. Consecutive records of the database of our Down screening project were assessed for free β-hCG levels and pregnancy outcomes. Pregnancies with foetal chromosomal or structural anomalies and those with underlying disease were excluded. Free β-hCG levels of < 0.5, > 0.5 and < 2.0, and ≥ 2.0 MoM were categorised as low, normal and high, respectively. Of 17,082 screened women, 13,620 were available for analysis. In the first trimester (n = 8150), low β-hCG levels significantly increased risk for intrauterine growth restriction (IUGR), preterm birth, low birth weight (LBW) and low Apgar score with relative risk of 1.66, 1.43, 1.83 and 2.89; whereas high β-hCG group had a significant decreased risk of preterm birth and GDM with relative risk of 0.73 and 0.62. In the second trimester (n = 5470), both low and high β-hCG groups had significant increased risks of the most common adverse outcomes, i.e. spontaneous abortion, IUGR and preterm birth. In conclusion, abnormally low (< 0.5MoM) or high (> 2.0 MoM) free β-hCG levels are generally associated with an increased risk of adverse pregnancy outcomes. Nevertheless, high free β-hCG levels in the first trimester may possibly decrease risk of preterm delivery and GDM.

  18. Association between altered placental human chorionic gonadotrophin (hCG) production and the occurrence of cryptorchidism: a retrospective study

    PubMed Central

    2014-01-01

    Background An increase in cryptorchidism has been reported in many countries. One mechanism could be low fetal testosterone production possibly secondary to altered placental human chorionic gonadotrophin (hCG) release. Our Objective was to compare hCG values from maternal blood between boys with cryptorchidism and normal boys. Methods Total hCG and α-fetoprotein (AFP) values [12–16 weeks of gestation; from the double test for Down syndrome screening) were compared between cases of cryptorchidism and normal control boys who were matched for maternal age, maternal smoking, gestational age at time of hCG measurement (±1 day), birth weight and birth term. Measurements were performed in a single laboratory; values were expressed as absolute values (KU/L) and multiples of the median (MoM). Boys whose mothers had had a complicated pregnancy were excluded. Groups were compared using the Student’s t test. Log transformation was used to normalize hCG, MoM hCG, AFP and MoM AFP distribution, and values were expressed as geometric means (-1, + 1 tolerance factor). Results Total hCG and MoM hCG levels were significantly lower in the 51 boys with cryptorchidism compared to 306 controls (21.4 (12.3; 37) KU/L vs 27.7 (15.9; 47.9) KU/L and 0.8 (0.5; 1.2) MoM vs 1.0 (0.6; 1.6) MoM, respectively, p < 0.01). By contrast, AFP and MoM AFP levels were similar between groups. Conclusion This study showed a link between low maternal serum hCG levels and cryptorchidism in boys from uncomplicated pregnancy, while normal AFP levels indicated a normal fetoplacental unit. Whether these abnormalities were due to endogenous or exogenous factors remains to be determined. PMID:25064170

  19. Effect of intrauterine injection of human chorionic gonadotropin before embryo transfer on pregnancy rate: A prospective randomized study

    PubMed Central

    Mostajeran, Fatemeh; Godazandeh, Farzaneh; Ahmadi, Sayed Mehdi; Movahedi, Minoo; Jabalamelian, Seyed Abolfazl

    2017-01-01

    Background: Human chorionic gonadotropin (hCG) as the most important factor to controlled implantation is one of the early embryonic signals in primates that is secreted by the embryo before its implantation. This study was designed to assess the effects of intrauterine injection of hCG before the embryo transfer in an in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) cycle on pregnancy rate in infertile patients. Materials and Methods: This randomized study was done on 100 infertile patients in two groups: intervention group received injection of 700 IU of intrauterine hCG 10 min before embryo transfer and control group did not receive hCG. The pregnancy rate was tested 2 weeks after embryo transfer, and if the pregnancy test was positive, a transvaginal ultrasound was performed 3 weeks later to search for signs of pregnancy, such as the presence of a gestational sac, embryo, and fetal heart rate, and confirmed as successful pregnancy. Results: Pregnancy test was positive in 13 (28.6%) of 46 patients in hCG group and in control group was positive in 6 (12.5%) of 48 patients. The pregnancy rate between hCG group and control group was not significantly different (P = 0.54). The pregnancy rate in hCG group with IVF fertilization was 20.8% and in their controls was 7.4% (P = 0.51). The pregnancy rate in hCG group with ICSI fertilization was 36.4% and in their controls was 19% (P = 0.16). Conclusion: The intrauterine injection of 700 IU of hCG before embryo transfer improved pregnancy rate compared to control group but was not significantly different.

  20. 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.

  1. Absence of human chorionic somatomammotropin during pregnancy associated with two types of gene deletion.

    PubMed

    Simon, P; Decoster, C; Brocas, H; Schwers, J; Vassart, G

    1986-11-01

    Complete absence of human somatomammotropin (hCS) was demonstrated in two patients experiencing an otherwise uneventful pregnancy. After delivery, DNA was prepared from the neonate blood or from the placenta and the integrity of the hCS-hGH gene cluster was investigated by Southern blotting and hybridization with an hCS cDNA probe. Patient 1 was found to be homozygous for a deletion involving hCS-A, hGH-V, and hCS-B. Patient 2 was a double heterozygote, with one chromosome bearing the same deletion as that of patient 1, while in the other, only the hCS-A gene was missing. Considerations relative to the frequency of the defect are derived from the present results.

  2. Uterine arteriovenous malformation with positive serum beta-human chorionic gonadotropin: Embolization of both uterine arteries and extra-uterine feeding arteries

    PubMed Central

    Kim, Su Mi; Ahn, Hee Young; Choi, Min Jeong; Kang, Yun Dan; Park, Jin Wan; Park, Choong Hak

    2016-01-01

    The incidence of uterine arteriovenous malformation (AVM) is rare. However, it is clinically significant in that it can cause life-threatening vaginal bleeding. We report a case of a large uterine AVM with positive serum beta-human chorionic gonadotropin. A presumptive diagnosis was made; a uterine AVM accompanied by, early pregnancy or retained product of conception. Because this uterine AVM was extensive, transcatheter arterial embolization of both uterine arteries and extra-uterine feeding arteries was performed. Three months after undergoing transcatheter arterial embolization, complete resolution of the uterine AVM was confirmed without major complication. PMID:27896262

  3. Chorionic villus sampling

    MedlinePlus

    ... procedure, its risks, and alternative procedures such as amniocentesis . You will be asked to sign a consent ... CVS can be done sooner in pregnancy than amniocentesis, usually at about 10 to 12 weeks. Chorionic ...

  4. 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

  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. Human chorionic gonadotropin suppresses human breast cancer cell growth directly via p53-mediated mitochondrial apoptotic pathway and indirectly via ovarian steroid secretion.

    PubMed

    Yuri, Takashi; Kinoshita, Yuichi; Emoto, Yuko; Yoshizawa, Katsuhiko; Tsubura, Airo

    2014-03-01

    The tumor-suppressive effects of human chorionic gonadotropin (hCG) against human breast cancer cells were examined. In cell viability assays, hCG inhibited the growth of three human breast cancer cell lines (estrogen receptor (ER)-positive KPL-1 and MCF-7, and ER-negative MKL-F cells), and the growth inhibition activity of hCG was most pronounced against KPL-1 cells (luteinizing hormone/chorionic gonadotropin receptor (LHCGR)-positive and luminal-A subtype). In hCG-treated KPL-1 cells, immunoblotting analysis revealed the expression of tumor suppressor protein p53 peaking at 12 h following treatment, followed by cleavage of caspase-9 and caspase-3 at 24 h and 48 h, respectively. KPL-1-transplanted athymic mice were divided into 3 groups: a sham-treated group that received an inoculation of KPL-1 cells at 6 weeks of age followed by daily intraperitoneal (i.p.) injection of saline; an in vitro hCG-treated KPL-1 group that received an inoculation of KPL-1 cells pre-treated with 100 IU/ml hCG in vitro for 48 h at 6 weeks of age, followed by daily i.p. injection of saline; and an in vivo hCG-treated group that received an KPL-1 cell inoculation at 6 weeks of age, followed by daily i.p. injection of 100 IU hCG. The daily injections of saline or hCG continued until the end of the experiment when mice reached 11 weeks of age. KPL-1 tumor growth was retarded in in vitro and in vivo hCG-treated mice compared to sham-treated controls, and the final tumor volume and tumor weight tended to be suppressed in the in vitro hCG-treated group and were significantly suppressed in the in vivo hCG-treated group. In vivo 100-IU hCG injections for 5 weeks elevated serum estradiol levels (35.7 vs. 23.5 pg/ml); thus, the mechanisms of hCG action may be directly coordinated via the p53-mediated mitochondrial apoptotic pathway and indirectly through ovarian steroid secretion that elevates estrogen levels. It is thus concluded that hCG may be an attractive agent for treating human breast

  7. Human chorionic gonadotropin β induces migration and invasion via activating ERK1/2 and MMP-2 in human prostate cancer DU145 cells.

    PubMed

    Li, Zongwen; Li, Chunliu; Du, Lianlian; Zhou, Yan; Wu, Wei

    2013-01-01

    We previously demonstrated that human chorionic gonadotropin β (hCGβ) induced migration and invasion in human prostate cancer cells. However, the involved molecular mechanisms are unclear. Here, we established a stable prostate cancer cell line overexpressing hCGβ and tested hCGβ-triggered signaling pathways causing cell migration and invasion. ELISA showed that the hCGβ amount secreted into medium increased with culture time after the hCGβ-transfected cells were incubated for 3, 6, 9, 12 and 24 h. More, hCGβ standards promoted MAPK (ERK1/2) phosphorylation and increased MMP-2 expression and activity in both dose- and time-dependent manners in hCGβ non-transfected cells. In addition, hCGβ promoted ERK1/2 phosphorylation and increased MMP-2 expression and activity significantly in hCGβ transfected DU145 cells. Whereas ERK1/2 blocker PD98059 (25 µM) significantly downregulated phosphorylated ERK1/2 and MMP-2. Particularly, hCGβ promoted cell migration and invasion, yet the PD98059 diminished the hCGβ-induced cell motility under those conditions. These results indicated that hCGβ induced cell motility via promoting ERK1/2 phosphorylation and MMP-2 upregulation in human prostate cancer DU145 cells.

  8. Changes in chronic low back pain and cardiovascular risk factors using a homeopathic human chorionic gonadotropin–based weight loss program: a case report

    PubMed Central

    Morningstar, Mark W.; Strauchman, Megan N.

    2011-01-01

    Objective The purpose of this case report is to describe the changes in body weight and biochemical markers in a patient who completed a homeopathic human chorionic gonadotropin protocol. Case Report A 52-year-old man reported to an integrative medical center (including chiropractic and osteopathic physicians) for chronic low back pain. The patient reported a 20-year history of chronic, episodic low back pain. A course of spinal manipulative therapy was delivered; however, because of the lack of resolution of symptoms, a radiographic examination was performed, the result of which was essentially normal. Laboratory studies demonstrated hypercholesterolemia, hyperlipidemia, uricemia, and elevated blood glucose. A dietary change in treatment approach was selected. Intervention and Outcome The patient was instructed to take 10 drops of a homeopathic human chorionic gonadotropin product under the tongue 5 times daily. His total daily energy (calorie) was limited for the first 30 days of the program while on the homeopathic product. After 4 months, the patient lost a total of 71 lb, pain and disability scores improved, and reductions in serum cardiovascular markers were noted. Conclusion The findings of this study showed that weight loss seemed to affect the patient's chronic low back pain and cardiovascular risk factors. PMID:22654693

  9. Differentiation Potential of Human Chorion-Derived Mesenchymal Stem Cells into Motor Neuron-Like Cells in Two- and Three-Dimensional Culture Systems.

    PubMed

    Faghihi, Faezeh; Mirzaei, Esmaeil; Ai, Jafar; Lotfi, Abolfazl; Sayahpour, Forough Azam; Ebrahimi-Barough, Somayeh; Barough, Somayeh Ebrahimi; Joghataei, Mohammad Taghi

    2016-04-01

    Many people worldwide suffer from motor neuron-related disorders such as amyotrophic lateral sclerosis and spinal cord injuries. Recently, several attempts have been made to recruit stem cells to modulate disease progression in ALS and also regenerate spinal cord injuries. Chorion-derived mesenchymal stem cells (C-MSCs), used to be discarded as postpartum medically waste product, currently represent a class of cells with self renewal property and immunomodulatory capacity. These cells are able to differentiate into mesodermal and nonmesodermal lineages such as neural cells. On the other hand, gelatin, as a simply denatured collagen, is a suitable substrate for cell adhesion and differentiation. It has been shown that electrospinning of scaffolds into fibrous structure better resembles the physiological microenvironment in comparison with two-dimensional (2D) culture system. Since there is no report on potential of human chorion-derived MSCs to differentiate into motor neuron cells in two- and three-dimensional (3D) culture systems, we set out to determine the effect of retinoic acid (RA) and sonic hedgehog (Shh) on differentiation of human C-MSCs into motor neuron-like cells cultured on tissue culture plates (2D) and electrospun nanofibrous gelatin scaffold (3D).

  10. 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.

  11. Human chorionic gonadotropin therapy in adolescent boys with constitutional delayed puberty vs those with beta-thalassemia major.

    PubMed

    Soliman, Ashraf T; Nasr, Ibrahim; Thabet, Alaa; Rizk, Mustafa M; El Matary, Wael

    2005-01-01

    We studied 12 adolescent boys with beta-thalassemia major and delayed puberty (age, 15.8 +/- 1 years) with Tanner I sexual development treated with a long-term low-transfusion regimen. Ten nonthalassemic adolescents (> 14 years) with constitutional delay of growth and puberty (CDGP) served as controls. Auxologic parameters and testicular size were measured, and bone age was determined. Measurement of basal gonadotropin (luteinizing hormone [LH] and follicle-stimulating hormone [FSH]) and testosterone (T) levels taken at 8 am revealed prepubertal levels in both groups of patients. Human chorionic gonadotropin (hCG, 2500 U/m(2)) was injected intramuscularly twice weekly for 6 months, and anthropometric data, testicular diameter, and serum T concentrations were remeasured after 1 and 6 months. The testicular diameter after 6 month of hCG therapy was significantly correlated with the testicular diameter and T level after 1 month of therapy (r = 0.93 and 0.39, respectively, P < .01). After 6 months of hCG therapy, the mean growth velocity (GV) increased from 4.1 to 8.6 cm/y in thalassemic patients and from 4.6 to 10.3 cm/y in those with CDGP during hCG therapy. In thalassemic boys, the mean T concentration increased from 0.93 to 2.7 nmol/L (mean increase = 1.8 nmol/L) vs an increase from 0.47 to 4.81 nmol/L (mean increase = 4.32 nmol/L) in those with CDGP. All adolescents with CDGP, but only 7 the 12 thalassemic adolescents, had T secretion above 2 nmol/L after 6 months of hCG therapy and maintained their growth and pubertal development for a year after stopping hCG. The 5 thalassemic patients with defective T secretion after hCG therapy had significantly higher ferritin level (1985 +/- 658 ng/mL) vs the other 7 patients (1100 +/- 425 ng/mL). These findings denoted significant testicular dysfunction in those patients with higher iron overload (testicular siderosis). Statural GV was significantly correlated with insulin-like growth factor 1 (IGF-1) concentrations and

  12. Birth after human chorionic gonadotropin-primed oocyte in vitro maturation and fertilization with testicular sperm in a normo-ovulatory patient

    PubMed Central

    González-Ortega, Claudia; Piña-Aguilar, Raul Eduardo; Cancino-Villareal, Patricia; Gutiérrez-Gutiérrez, Antonio Martin

    2016-01-01

    In this report, we present a case of in vitro maturation (IVM) with surgical retrieved testicular sperm in a normo-ovulatory female. Human chorionic gonadotropin-primed IVM, testicular biopsy for sperm retrieval and intracytoplasmic sperm injection with fresh sperm were performed. Fourteen cumulus-oocyte complexes were obtained in germinal vesicle or metaphase I stage, eight oocytes reached metaphase II, seven presumptive zygotes were obtained, and three cleavage stages embryos in day 2 were transferred producing a singleton pregnancy. A single healthy newborn was obtained. Our results suggest that IVM may be an alternative for in vitro fertilization in normo-ovulatory women even if surgical retrieval of sperm is needed. Further research is required to depict contributing factors to the success of IVM in indications different from polycystic ovaries syndrome and the role of male gamete. PMID:27803591

  13. Human chorionic gonadotropin β induces cell motility via ERK1/2 and MMP-2 activation in human glioblastoma U87MG cells.

    PubMed

    Li, Zongwen; Du, Lianlian; Li, Chunliu; Wu, Wei

    2013-02-01

    Human chorionic gonadotropin β (hCGβ) promotes tumorigenesis in a variety of tumors including glioblastoma, breast and prostate cancer cells, etc. However, the involved mechanisms remain elusive. Distinct from the other tumors, glioblastoma is a highly invasive brain tumor; invasion causes high recurrence and mortality. Characterization of hCGβ signaling is to determine therapeutic targets to inhibit invasion and lower recurrence. Through both a stable cell line over-expressing hCGβ and hCGβ standards, we tested hCGβ signaling, migration and invasion in human glioblastoma U87MG cells. ELISA showed that hCGβ secreted into culture medium at an amount of 237.8 ± 7.8 ng/10(7) cells in hCGβ transfected stable cells after the cells were grown for 24 h. Through Western blot and Gelatin zymography, we found that hCGβ standards phosphorylated ERK1/2 and upregulated MMP-2 expression in dose- and time-dependent manners. Meanwhile, overexpressed hCGβ phosphorylated ERK1/2, and upregulated MMP-2 expression and activity, whereas ERK1/2 blocker PD98059 (25 μM) significantly decreased both ERK1/2 and MMP-2 expression and activity. In addition, in the same conditions as the signaling test, hCGβ promoted cell migration and invasion, whereas the PD98059 diminished these effects. These findings demonstrated that hCGβ phosphorylated ERK1/2 upregulating MMP-2 expression and activity leading to cell migration and invasion, suggesting that hCGβ, ERK1/2 and MMP-2 are the potential targets to inhibit glioblastoma invasion.

  14. 21 CFR 522.1081 - Chorionic gonadotropin.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Chorionic gonadotropin. 522.1081 Section 522.1081 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM NEW ANIMAL DRUGS §...

  15. 21 CFR 522.1081 - Chorionic gonadotropin.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Chorionic gonadotropin. 522.1081 Section 522.1081 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM NEW ANIMAL DRUGS §...

  16. 21 CFR 522.1081 - Chorionic gonadotropin.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Chorionic gonadotropin. 522.1081 Section 522.1081 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM NEW ANIMAL DRUGS §...

  17. 21 CFR 522.1081 - Chorionic gonadotropin.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Chorionic gonadotropin. 522.1081 Section 522.1081 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM NEW ANIMAL DRUGS §...

  18. 21 CFR 522.1081 - Chorionic gonadotropin.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Chorionic gonadotropin. 522.1081 Section 522.1081 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM NEW ANIMAL DRUGS §...

  19. The influence of mode of delivery, obstetric analgesia and anaesthesia on the response of isolated human chorionic plate arteries to angiotensin II.

    PubMed

    Odum, C U; Pipkins, F B

    1989-01-01

    A six point concentration: response curved of the contractile effect of angiotensin II (AII) on helically-cut strips of human chorionic plate artery strips were established at final concentrations of between 10(-14) to 10(-9) M. The tissues were obtained from the placentae of primigravid patients who had normal pregnancy, and also from those with pregnancy induced hypertension (PIH). The tissue response were then related to mode of delivery, obstetric analgesia, and anaesthesia. A total of 36 chorionic plate arteries from 12 primigravid patients were studied. i) The overall initial EC50s of the tissues ranged from 8.0 x 10(-13) M and 4.5 x 10(-13) M. The tissues from PIH patients were significantly more sensitive to AII, when compared with tissues from the normotensive subjects (P greater than or equal to 0.01 less than or equal to 0.05). ii) The tissues from epidural vaginal deliveries were also significantly more sensitive to AII, than those from both normal vaginal deliveries and caesarean deliveries respectively. The median gradients of the semi-log transformed concentration response curved were 2.4 +/- 0.18; 1.27 +/- 0.37, and 1.5 +/- 0.49, for epidural, Caeserean and normal vaginal deliveries respectively. iii) It is suggested that whilst Lumbar epidural analgesia may be of great value in pain relief in labour and in the control of intrapartum hypertension in pre-eclampsia, this procedure may be associated with hypersensitivity and perhaps vasospasm of the placental vasculature to vasoactive agents invivo.

  20. 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

  1. [Stimulating effect of human chorionic gonadotropin on the activity of guanidinoacetate-N-methyltransferase of rat testes: role of cyclic AMP].

    PubMed

    Karelin, A A; Mardashev, S R

    1977-01-01

    A study was made of the influence of human chorionic gonadotropin (HCGT) on the activity of guanidineacetate-N-methyltranspherase in human testes. Guanidineacetate-methyltranspherase proved to be stimulated in the testes of the sexually immature rats in vivo under the action of HCGT on the 19th day after birth and from the 24th to the 27th day after birth. The activity of the enzyme was also increased in adult rats weighing 35--40 g. The activity of guanidineacetate-methyltranspherase in the testes of rats was particularly sharply stimulated after a single administration of N6-O2'-dibutiryladenosine-3',5'-cyclophosphate against the background of a preliminary treatment of rats with HCGT for a period of 5 days. The enzyme stimulation induced by combined treatment of rats with HCGT and N6-O2'-dibutiryladenosine-3',5'-cyclophosphate was prevented by the administration of cycloheximide and actinomycine D. The activity of the enzyme decreased after the incubation of the rat testes tissue homogenate with HCGT.

  2. Layer-by-layer assembly of gold nanoparticles and cysteamine on gold electrode for immunosensing of human chorionic gonadotropin at picogram levels.

    PubMed

    Roushani, Mahmoud; Valipour, Akram; Valipour, Mehdi

    2016-04-01

    The development of an electrochemical immunosensor for the detection of human chorionic gonadotropin (hCG) is described with a limit of detection as low as 0.3 pg mL(-1) in phosphate buffer. In this immunosensor, cysteamine (Cys) and gold nanoparticles (AuNPs) were used to immobilize an anti-hCG monoclonal antibody onto a gold electrode (GE). The structure of AuNPs has been confirmed by EDS, SEM, and TEM analysis. Due to the large specific surface area and excellent electrical conductivity of AuNPs, electron transfer was promoted and the amount of hCG antibody was enhanced significantly. A systematic study on the effects of experimental parameters such as pH, incubation time in the hCG solution and urea solution used for experiments on the binding between the immobilized antibody and hCG has been carried out. Under optimal experimental parameters, differential pulse voltammetry (DPV) signal changes of the [Fe(CN)6](3-/4-) are used to detect hCG with two broad linear ranges: 0.001 to 0.2 and 0.2 to 60.7 ng mL(-1). The LOD value proves more sensitive in comparison with previously reported methods. The prepared immunosensor showed high sensitivity and stability. In addition, the immunosensor was successfully used for the determination of hCG in human serum.

  3. 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

  4. Ovarian stimulation using human chorionic gonadotrophin impairs blastocyst implantation and decidualization by altering ovarian hormone levels and downstream signaling in mice.

    PubMed

    Ezoe, Kenji; Daikoku, Takiko; Yabuuchi, Akiko; Murata, Nana; Kawano, Hiroomi; Abe, Takashi; Okuno, Takashi; Kobayashi, Tamotsu; Kato, Keiichi

    2014-11-01

    Ovarian stimulation induced by follicle-stimulating hormone and human chorionic gonadotrophin (hCG) is commonly used in assisted reproductive technology to increase embryo production. However, recent clinical and animal studies have shown that ovarian stimulation disrupts endometrial function and embryo development and adversely affects pregnancy outcomes. How ovarian stimulation impairs pregnancy establishment and the precise mechanisms by which this stimulation reduces the chances of conception remain unclear. In this study, we first demonstrated that ovarian stimulation using hCG alone impairs implantation, decidualization and fetal development of mice by generating abnormal ovarian hormone levels. We also showed that ovarian hormone levels were altered because of changes in the levels of the enzymes involved in their synthesis in the follicles and corpora lutea. Furthermore, we determined that anomalous ovarian hormone secretion induced by ovarian stimulation alters the spatiotemporal expression of progesterone receptors and their downstream genes, especially in the uterine epithelium. Epithelial estrogenic signaling and cell proliferation were promoted on the day of implantation in stimulated mice and these changes led to the failure of uterine transition from the prereceptive to the receptive state. Collectively, our findings indicate that ovarian stimulation using hCG induces an imbalance in steroid hormone secretion, which causes a failure of the development of uterine receptivity and subsequent implantation and decidualization by altering the expression of steroid receptors and their downstream signaling associated with embryo implantation.

  5. Induction of spermatogenesis and spermiation by a single injection of human chorionic gonadotropin in intact and hypophysectomized immature European eel (Anguilla anguilla L.).

    PubMed

    Khan, I A; Lopez, E; Leloup-Hâtey, J

    1987-10-01

    Intact and hypophysectomized male silver eels (Anguilla anguilla) in fresh water received a single injection of human chorionic gonadotropin (hCG) (250 C) or solvent (0.15 M NaCl). No effect of solvent was observed. Spermatogonia proliferated in testis of hCG-treated intact or hypophysectomized eels. One month after the injection, primary and secondary spermatocytes were found. After 3 months, numerous spermatozoa were present. In hypophysectomized eels, hCG was also effective even though maturing germ cells were less numerous and spermiation was less frequent than in intact animals. Within 1 week after hCG injection, plasma levels of free and glucuroconjugated androgens (testosterone and 11-oxotestosterone) rose significantly in intact and hypophysectomized fish. The highest values were observed within 1 month, and then plasma levels decreased to pretreatment values. The most important changes were observed in the case of free 11-oxotestosterone. The long-term effects of hCG can be explained partly by the long half-life of this hormone. The effects of hypophysectomy on the response of testis to hCG caused us to think that some endogenous pituitary secretions must interfere in the intact fish so that maximal effects of hCG, especially on the induction of spermiation, are obtained.

  6. The in vitro effects of steroids, human chorionic gonadotropin and cyanoketone on germinal vesicle breakdown of striped mullet ( Mugil cephalus L.) oocytes

    NASA Astrophysics Data System (ADS)

    Hong, Wanshu; Thomas, Peter

    1987-03-01

    The in vitro effects of steroids, human chorionic gonadotropin (HCG) and cyanoketone on germinal vesicle breakdown (GVBD) of striped mullet ( Mugil cephalus L.) oocytes were investigated. All concentrations of HCG (5,10,50 I.U./ml), progesterone and pregnenolone at the highest concentrations(lug/ml) were moderately effective in inducing GVBD, whereas 17β-estrodiol, cortisol, testosterone, 11β-hydroxyandrostenedione and 11-ketotestosterone did not stimulate GVBD. 17α, 20βdihydroxy-4-pregnen-3-one (17α, 20βdiOHprog) and deoxycorticosterone (DOC) were the most potent steroids in stimulating final oocyte maturation. The results indicate that C21 hydroxylated steroids are potent inducers of final maturation in mullet. Further, co-incubations with 17β-estradiol, cortisol and testosterone did not alter the maturation-inducing effects of HCG or 17α,20βdiOHprog. Cyanoketone, a blocker of 3βHSD activity, was only partially effective in blocking GVBD induced by HCG. This suggests that Δ5 (pregnenolone derived) and Δ4 steroids may be involved in final oocyte maturation in this species.

  7. The effect of intrauterine human chorionic gonadotropin injection before embryo transfer on the implantation and pregnancy rate in infertile patients: A randomized clinical trial

    PubMed Central

    Dehghani Firouzabadi, Razieh; Janati, Sima; Razi, Mohammad Hossein

    2016-01-01

    Background: Implantation is one of the essential steps for the success of assisted reproductive techniques (ART). Their success depends on three main factors: embryo quality, endometrial receptivity (ER), and synchrony between embryo and endometrium. There are various factors that regulate the complex process of implantation. In this regard, one may refer to human chorionic gonadotropin (hCG) as the most important factor. Objective: This study aims to investigate the effect of intrauterine hCG injection before embryo transfer (ET) on pregnancy outcome in infertile couples. Materials and Methods: A total of 159 patients undergone In vitro Fertilization/ Intracytoplasmic Sperm Injection (IVF/ICSI) with an antagonist protocol were evaluated. Patients were divided into three groups (n=53). Group 1 received 500 IU of hCG, group 2 received 1000 IU of hCG intrauterine injection before ET, and the control group underwent ET without hCG preceding intrauterine injection. Results: There was no significant difference among the groups. The implantation rates were 18.86%, 13.52%, and 14.37%, chemical pregnancy rates were 34%, 32.1%, and 35.3%, and clinical pregnancy rates were 34%, 32.1%, and 31.4% respectively. Conclusion: The pregnancy outcome in IVF/ICSI /ET cycles cannot be improved through hCG intrauterine injections before ET. PMID:27921090

  8. Analysis of human chorionic gonadotropin (hCG): application of routine immunological methods for initial testing and confirmation analysis in doping control.

    PubMed

    Kuuranne, Tiia; Ahola, Liisa; Pussinen, Christel; Leinonen, Antti

    2013-08-01

    Human chorionic gonadotropin (hCG) is dimeric glycoprotein produced by placenta in pregnancy and also in low levels by pituitary gland. The main clinical use for exogenous hCG-administration is typically linked to infertility. The desired effect of hCG misuse in sport is due to the enhancement of testicular production of testosterone. Therefore, hCG is listed by the World Anti-Doping Agency (WADA) as a prohibited substance in male athletes and according to the recently published WADA guideline urinary concentrations of hCG > 5 IU/L may be an indicator of doping. In this study two independent immunoassays were used to implement the new WADA guideline. The assay for initial testing (Siemens Immulite 2000 XPi hCG assay) recognizes various hCG variants (e.g. hCG and β-core fragment of hCG) whereas the confirmatory assay (PerkinElmer DELFIA Xpress hCG) is sensitive to intact and nicked hCG only. Both assays showed adequate sensitivity and were proven fit-for-purpose in routine doping control. Population-based distribution of the assays was in good agreement with results of earlier studies and supported well the current threshold of 5 IU/L.

  9. 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.

  10. Beta-human chorionic gonadotropin expression in recurrent and metastatic giant cell tumors of bone: a potential mimicker of germ cell tumor.

    PubMed

    Lawless, Margaret E; Jour, George; Hoch, Benjamin L; Rendi, Mara H

    2014-10-01

    Giant cell tumors of bone (GCTs) are generally benign, locally aggressive neoplasms that rarely metastasize. The beta subunit of human chorionic gonadotropin (beta-hCG) is expressed in syncytiotrophoblasts and several nongynecologic neoplasms but has not been described in GCT. At our institution, we observed cases of elevated beta-hCG in patients with GCT leading to diagnostic difficulty and in one case, concern for metastatic choriocarcinoma. This study aims to determine the frequency of beta-hCG expression in GCT and any relationship to clinical aggressiveness. We evaluated tissue expression of beta-hCG by immunohistochemistry with 58% of cases staining for beta-hCG. Additionally, 2 of 11 patients with available serum and/or urine beta-hCG measurements demonstrated elevated beta-hCG due to tumor. It is important to be aware of beta-hCG expression by GCT and the potential for elevated urine and serum beta-hCG levels in patients with GCT so as to avoid misdiagnosis of pregnancy or gestational trophoblastic disease.

  11. Pelvic pain, free fluid in pelvis, and human chorionic gonadotropin serum elevation: recurrence of malignant ovarian germ-cell tumor or early pregnancy?

    PubMed

    Barczyński, B; Rogala, E; Nowicka, A; Nurzyńska-Flak, J; Kotarski, J

    2013-01-01

    Conservative treatment of metastatic germ-cell tumor of the ovary does not exclude the possibility of pregnancy in the future. Serum beta-human chorionic gonadotropin (beta-hCG) serves as pregnancy test, and has also been proven to be a useful marker for ovarian germ-cell tumors. This paper is a case report of a 19-year-old patient who was admitted to a district hospital in emergency due to pelvic pain, amenorrhoea, and free fluid in the pelvis. Laboratory tests demonstrated slight increase in beta-hCG serum concentration and transvaginal ultrasound (TVUS) showed no evidence of gestational sac in the uterus. At the age of 14, the patient was diagnosed with malignant germ-cell tumor of the left ovary in FIGO Stage IV and was treated with four courses of chemotherapy according to TGM-95 protocol with etoposide, ifosfamide, and cisplatin, followed by conservative surgery and adjuvant two courses of cytostatics. The initial diagnosis was recurrence of ovarian malignancy and the patient was referred to an oncology center. Wait-and-see approach and repeated ultrasound examination confirmed a normal intrauterine pregnancy which concluded with the delivery of a healthy newborn through cesarean section.

  12. Chemical synthesis of the β-subunit of human luteinizing (hLH) and chorionic gonadotropin (hCG) glycoprotein hormones.

    PubMed

    Fernández-Tejada, Alberto; Vadola, Paul A; Danishefsky, Samuel J

    2014-06-11

    Human luteinizing hormone (hLH) and human chorionic gonadotropin (hCG) are human glycoprotein hormones each consisting of two subunits, an identical α-subunit and a unique β-subunit, that form noncovalent heterodimers. Structurally, β-hCG shares a high degree of sequence similarity with β-hLH, including a common N-glycosylation site at the N-terminus but differs mainly in the presence of an extended C-terminal portion incorporating four closely spaced O-linked glycans. These glycoproteins play important roles in reproduction and are used clinically in the treatment of infertility. In addition, the role of hCG as a tumor marker in a variety of cancers has also attracted significant interest for the development of cancer vaccines. In clinical applications, these hormones are administered as mixtures of glycoforms due to limitations of biological methods in producing homogeneous samples of these glycoproteins. Using the powerful tools of chemical synthesis, the work presented herein focuses on the highly convergent syntheses of homogeneous β-hLH and β-hCG bearing model glycans at all native glycosylation sites. Key steps in these syntheses include a successful double Lansbury glycosylation en route to the N-terminal fragment of β-hCG and the sequential installation of four O-linked glycosyl-amino acid cassettes into closely spaced O-glycosylation sites in a single, high-yielding solid-supported synthesis to access the C-terminal portion of the molecule. The final assembly of the individual glycopeptide fragments involved a stepwise native chemical ligation strategy to provide the longest and most complex human glycoprotein hormone (β-hCG) as well as its closely related homologue (β-hLH) as discrete glycoforms.

  13. Complement C3 and Decay-Accelerating Factor Expression Levels Are Modulated by Human Chorionic Gonadotropin in Endometrial Compartments During the Implantation Window

    PubMed Central

    Argandoña, Felipe; Azúa, Rodrigo; Kohen, Paulina; Devoto, Luigi

    2013-01-01

    The control of complement activation in the embryo–maternal environment has been demonstrated to be critical for embryo survival. Complement proteins are expressed in the human endometrium; however, the modulation of this expression by embryo signals has not been explored. To assess the expression of complement proteins in response to human chorionic gonadotropin (hCG), we designed an experimental study using in vivo and in vitro models. Twelve fertile women were treated with hCG or left untreated during the mid-luteal phase, and an endometrial biopsy was performed 24 hours later. The localizations of C3, membrane cofactor protein (MCP; CD46), decay-accelerating factor (DAF; CD55), and protectin (CD59) were assessed by immunohistochemistry, and the messenger RNA (mRNA) levels of these proteins were quantified by real-time reverse transcriptase–polymerase chain reaction (RT-PCR) in cells harvested from endometrial compartments using laser capture microdissection. Endometrial explants were cultured with or without hCG for 24 hours, and the C3 and DAF protein levels were measured by Western blotting. Elevated C3 mRNA levels in stromal cells and elevated DAF levels in epithelial luminal cells were detected after hCG treatment. In the endometrial explant model, the progesterone receptor antagonist RU486 inhibited the increases in the levels of C3 and DAF in response to hCG. The findings of this study indicate that hCG plays a role in embryo–endometrium communication and affects the expression of complement proteins in endometrial compartments during the implantation window. PMID:23427180

  14. Downregulation of A(1) and A(2B) adenosine receptors in human trisomy 21 mesenchymal cells from first-trimester chorionic villi.

    PubMed

    Gessi, Stefania; Merighi, Stefania; Stefanelli, Angela; Mirandola, Prisco; Bonfatti, Alessandra; Fini, Sergio; Sensi, Alberto; Marci, Roberto; Varani, Katia; Borea, Pier Andrea; Vesce, Fortunato

    2012-11-01

    Human reproduction is complex and prone to failure. Though causes of miscarriage remain unclear, adenosine, a proangiogenic nucleoside, may help determine pregnancy outcome. Although adenosine receptor (AR) expression has been characterized in euploid pregnancies, no information is available for aneuploidies, which, as prone to spontaneous abortion (SA), are a potential model for shedding light on the mechanism regulating this event. AR expression was investigated in 71 first-trimester chorionic villi (CV) samples and cultured mesenchymal cells (MC) from euploid and TR21 pregnancies, one of the most frequent autosomal aneuploidy, with a view to elucidating their potential role in the modulation of vascular endothelial growth factor (VEGF) and nitric oxide (NO). Compared to euploid cells, reduced A(1) and A(2B) expression was revealed in TR21 CV and MCs. The non-selective adenosine agonist 5'-N-ethylcarboxamidoadenosine (NECA) increased NO, by activating, predominantly, A(1)AR and A(2A)AR through a molecular pathway involving hypoxia-inducible-factor-1 (HIF-1α), and increased VEGF, mainly through A(2B). In conclusion the adenosine transduction cascade appears to be disturbed in TR21 through reduced expression of A(2B) and A(1)ARs. These anomalies may be implicated in complications such as fetal growth restriction, malformation and/or SA, well known features of aneuploid pregnancies. Therefore A(1) and A(2B)ARs could be potential biomarkers able to provide an early indication of SA risk and their stimulation may turn out to improve fetoplacental perfusion by increasing NO and VEGF.

  15. β-human chorionic gonadotropin assay in vaginal washing fluid for the accurate diagnosis of premature rupture of membranes during late pregnancy

    PubMed Central

    Temel, Orhan; Çöğendez, Ebru; Selçuk, Selçuk; Asoğlu, Mehmet Reşit; Kaya, Erdal

    2013-01-01

    Objective To determine whether the measurement of beta-human chorionic gonadotropin (β-hCG) levels in vaginal fluid is useful for the diagnosis of premature rupture of membranes (PROM). Material and Methods A total of 92 pregnant women between 24 and 40 weeks gestation participated in this study. The patients with fluid leaking from the vagina were designated Group 1, the patients with no fluid leaking from the vagina were Group 2, and those with a suspicion of fluid leaking from the vagina were classified as Group 3. Irrigating the posterior vaginal fornix with 5 mL sterile saline was used to measure β-hCG levels of the patients. Receiver operator curve (ROC) analysis was used to determine the cut-off value for a positive diagnosis. Results The β-hCG levels of vaginal fluid were measured as 20.5±25.0 mIU/mL, 254.6±346.8 mIU/mL, and 74.3±100.8 mIU/mL in Group 1, Group 2, and Group 3, respectively. Vaginal β-hCG level was higher statistically significantly in Group 2 than Group 1 and 3 (p<0.001). 100 mIU/mL was accepted as a cut-off value by using the receiver operating characteristic curve. According to 100 mIU/mL, sensitivity, specificity, positive predictive and negative predictive values were calculated as 71.2, 100, 100, and 65.1%, respectively. Conclusion The study showed that the measurement of β-hCG level in vaginal washing fluid is an efficient and easy diagnostic test for predicting the amount of fluid leaking from the vagina. However, due to the low negative predictive value of the test, it would not be convenient in daily practice. PMID:24592106

  16. Immature human chorionic gonadotropin (hCG) in first trimester placental cells is bound to an ATP-binding protein forming high-molecular-weight hCG.

    PubMed

    Shimojo, M; Sakakibara, R; Ishiguro, M

    1993-07-01

    Human chorionic gonadotropin (hCG) in first trimester placental cells is made up of immature alpha- and beta-subunits containing only N-linked high-mannose sugar chains, which are of 21 kDa for the alpha-subunit and 23 and 19 kDa for the beta-subunit. However, the apparent molecular weight of immature hCG from placental cell extracts has been estimated from gel filtration to be much higher (100-200 kDa; high molecular weight-hCG, HMW-hCG) based on gel filtration than the theoretical value (approximately 44 kDa) of the alpha beta dimer (alpha beta-hCG). We prepared a gel-filtered fraction containing HMW-hCG and investigated treatments for converting it to alpha beta-hCG. We found that the molecular weight of HMW-hCG was decreased to close to that of alpha beta-hCG by treatment with acetone, proteases, or chelating agents. These treatments also shifted the isoelectric point of HMW-hCG from the acidic region (pI = 4-6) to the alkaline (pI = 9-11), approximating to that of alpha beta-hCG. We also found that HMW-hCG, but not acetone-treated HMW-hCG, bound to ATP-agarose resin. These results suggested that the immature alpha beta-hCG molecule in placental cells may be bound to an acidic ATP-binding protein to form HMW-hCG.

  17. 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.

  18. Low-Dose Urinary Human Chorionic Gonadotropin Is Effective for Oocyte Maturation in In Vitro Fertilization/ Intracytoplasmic Sperm Injection Cycles Independent of Body Mass Index

    PubMed Central

    R. Hoyos, Luis; Khan, Sana; Dai, Jing; Singh, Manvinder; P. Diamond, Michael; E. Puscheck, Elizabeth; O. Awonuga, Awoniyi

    2017-01-01

    Background: Currently, there is no agreement on the optimal urinary derived human chorionic gonadotropin (u-hCG) dose requirement for initiating final oocyte maturation prior to oocyte collection in in vitro fertilization (IVF), but doses that range from 2500- 15000 IU have been used. We intended to determine whether low dose u-hCG was effective for oocyte maturation in IVF/intracytoplasmic sperm injection (ICSI) cycles independent of body mass index (BMI). Materials and Methods: We retrospectively evaluated a cohort of 295 women who underwent their first IVF/ICSI cycles between January 2003 and December 2010 at the Division of Reproductive Endocrinology and Infertility, Wayne State University, Detroit, MI, USA. Treatment cycles were divided into 3 groups based on BMI (kg/ m2): <25 (n=136), 25- <30 (n=84), and ≥30 (n=75) women. Patients received 5000, 10000 or 15000 IU u-hCG for final maturation prior to oocyte collection. The primary outcome was clinical pregnancy rates (CPRs) and secondary outcome was live birth rates (LBRs). Results: Only maternal age negatively impacted (P<0.001) CPR [odds ratio (OR=0.85, confidence interval (CI: 0.79-0.91)] and LBR (OR=0.84, CI: 0.78-0.90). Conclusion: Administration of lower dose u-hCG was effective for oocyte maturation in IVF and did not affect the CPRs and LBRs irrespective of BMI. Women’s BMI need not be taken into consideration in choosing the appropriate dose of u-hCG for final oocyte maturation prior to oocyte collection in IVF. Only maternal age at the time of IVF negatively influenced CPRs and LBRs in this study. PMID:28367299

  19. Differences in testosterone, androstenone, and skatole levels in plasma and fat between pubertal purebred Duroc and Landrace boars in response to human chorionic gonadotrophin stimulation.

    PubMed

    Oskam, I C; Lervik, S; Tajet, H; Dahl, E; Ropstad, E; Andresen, Ø

    2010-10-01

    The concentrations of the boar taint compounds androstenone and skatole in plasma and fat, together with those of testosterone in plasma, were investigated in pubertal purebred Duroc and Landrace boars following stimulation with human chorionic gonadotrophin (hCG). Higher initial levels of androstenone and testosterone were found in Duroc than Landrace boars. Duroc boars, which were approximately ten days older than the Landrace boars, also showed a more advanced stage of spermatogenesis than Landrace boars. While Landrace boars had the highest skatole levels. Following stimulation with hCG the relative increases in testosterone, androstenone, and skatole concentrations were highest in Landrace boars. The level of androstenone in fat three days after hCG stimulation exceeded 1 microg/g fat in all stimulated boars. The decreases in plasma levels of androstenone and testosterone on Days 2 and 3 after hCG stimulation were more pronounced in Landrace than Duroc boars. However, unlike the plasma androstenone and testosterone levels, the plasma concentrations of skatole did not decrease on Days 2 and 3 following stimulation, but remained elevated on Day 3. These results indicate that the lower levels of testicular steroids in Landrace boars compared with Duroc boars was not due to a lower production capacity, but more likely to a faster disappearance of steroids in Landrace boars. In the present study, age, live weight, and testicular development did not significantly contribute to the variation in fat androstenone. The present data and previous reports on candidate genes related to androstenone biosynthesis and metabolism suggests that future selection against factors associated with boar taint remains a possible solution for the problem of boar taint in the swine industry.

  20. Significant immunohistochemical expression of human chorionic gonadotropin in high-grade osteosarcoma is rare, but may be associated with clinically elevated serum levels.

    PubMed

    Lee, Anna F; Pawel, Bruce R; Sullivan, Lisa M

    2014-01-01

    Survival rates have plateaued at 70% for osteosarcoma. Proteins ectopically produced by malignant tumors may provide insight into new therapeutic targets. Osteosarcomas secreting human chorionic gonadotropin (hCG) have been suggested to have a worse prognosis. We examined the frequency of expression of β-subunit of hCG (β-hCG) in pretreatment osteosarcoma biopsies, and asked if it was associated with various clinical prognostic parameters, and the development of metastases. We subjected 51 pretreatment biopsies of high-grade osteosarcoma, from 51 patients, to β-hCG immunohistochemistry. In 19 of these patients, postchemotherapy metastatic biopsies also were examined for β-hCG expression. Clinical information (patient age, sex, survival status, and serum hCG in females only), and tumor characteristics (site, size, and presence of metastases) were recorded. The β-hCG positive and negative biopsies were separated and compared. Of 49 interpretable pretreatment biopsies, 28 (57%) showed positive cytoplasmic β-hCG expression: 27 with sparse positivity (1% of tumor cells) and 1 with frequent positivity (10% of tumor cells). The patient with frequent β-hCG positivity in her pretreatment biopsy had elevated serum hCG (88.2 mIU/mL) at diagnosis, decreasing to undetectable following chemotherapy and definitive resection. There was no difference in clinical parameters or rate of metastasis between β-hCG positive versus negative groups. Expression of β-hCG may be seen in high-grade osteosarcoma, but frequent β-hCG immunohistochemical expression by tumor cells, associated with clinically elevated serum β-hCG, is rare. Recognition that some nongerm cell tumors may produce β-hCG can prevent confusion with malignancies containing neoplastic syncytiotrophoblast cells, including germ cell and trophoblastic tumors.

  1. Human chorionic gonadotropin detection in cerebrospinal fluid of patients with a germinoma and its prognostic significance: assessment by using a highly sensitive enzyme immunoassay.

    PubMed

    Fukuoka, Kohei; Yanagisawa, Takaaki; Suzuki, Tomonari; Shirahata, Mitsuaki; Adachi, Jun-Ichi; Mishima, Kazuhiko; Fujimaki, Takamitsu; Katakami, Hideki; Matsutani, Masao; Nishikawa, Ryo

    2016-11-01

    OBJECTIVE Human chorionic gonadotropin (HCG) can be detected in a certain population of patients with a germinoma, but the frequency of germinoma HCG secretion and the prognostic value of HCG in the CSF are unknown. METHODS The authors measured HCG levels in sera and CSF in patients with a histologically confirmed germinoma by using a highly sensitive assay known as an immune complex transfer enzyme immunoassay (EIA), which is more than 100 times as sensitive as the conventional method, and they analyzed the correlation between HCG levels and the prognoses of patients with a germinoma. RESULTS HCG levels in sera and CSF of 35 patients with a germinoma were examined with the immune complex transfer EIA. The median CSF HCG levels in patients with a germinoma during the pretreatment and posttreatment evaluations were 192.5 pg/ml (range 1.2-13,116.5 pg/ml) and 18.7 pg/ml (1.2-283.9 pg/ml), respectively. Before treatment, the CSF HCG level was greater than the cutoff value in 85.7% of the patients with a germinoma. The authors compared survival rates among the patients by using a CSF HCG cutoff level of 1000 pg/ml, and the difference was statistically significant between the groups (p = 0.029, log-rank test). CONCLUSIONS Results of this study demonstrate that most germinomas secrete HCG. Patients with a germinoma that secretes higher amounts of HCG in their CSF experienced recurrence more frequently than those with lower CSF HCG levels.

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

    PubMed

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

    2016-01-01

    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.

  3. 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.

  4. Pretreatment serum human chorionic gonadotropin cutoff value for medical treatment success with single-dose and multi-dose regimen of methotrexate in tubal ectopic pregnancy

    PubMed Central

    Kim, Junhwan; Jung, Young Mi; Lee, Da Yong

    2017-01-01

    Objective To investigate individual pretreatment serum human chorionic gonadotropin (hCG) cutoff value for medical treatment success with single-dose and multi-dose regimen of methotrexate in tubal ectopic pregnancy. Methods Eighty-five women who received methotrexate for the treatment of tubal ectopic pregnancy during 2003 to 2015 were selected. Fifty-three women received a single-dose regimen and 32 women received a multi-dose regimen. Medical treatment failure was defined as necessity of surgical treatment. The medical treatment success rate was estimated in both regimens and the pretreatment serum hCG titer to predict the success was assessed by receiver operating characteristics curve analysis. Results Pretreatment clinical and laboratory parameters were similar between group of single-dose regimen and multi-dose regimen. Treatment success rate was 64.2% in the single-dose regimen group and 71.9% in the multi-dose regimen group (P>0.05). Pretreatment serum hCG titer was an independent prognostic factor for treatment success in each regimen. Serum hCG cutoff value to predict the treatment success was 3,026 IU/L in single-dose regimen group and 3,711 IU/L in multi-dose regimen group. Conclusion We recommend use of single-dose regimen when pretreatment serum hCG <3,026 IU/L but multi-dose regimen may be favored when initial serum hCG level between 3,026 and 3,711 IU/L. PMID:28217676

  5. In anesthetized pigs human chorionic gonadotropin increases myocardial perfusion and function through a β-adrenergic-related pathway and nitric oxide.

    PubMed

    Grossini, Elena; Surico, Daniela; Mary, David A S G; Molinari, Claudio; Surico, Nicola; Vacca, Giovanni

    2013-08-15

    Human chorionic gonadotropin (hCG) is not only responsible for numerous pregnancy-related processes, but can affect the cardiovascular system as well. So far, however, information about any direct effect elicited by hCG on cardiac function, perfusion, and the mechanisms involved has remained scarce. Therefore, the present study aimed to determine the primary in vivo effect of hCG on cardiac contractility and coronary blood flow and the involvement of autonomic nervous system and nitric oxide (NO). Moreover, in coronary endothelial cells (CEC), the intracellular pathways involved in the effects of hCG on NO release were also examined. In 25 anesthetized pigs, intracoronary 500 mU/ml hCG infusion at constant heart rate and aortic blood pressure increased coronary blood flow, maximum rate of change of left ventricular systolic pressure, segmental shortening, cardiac output, and coronary NO release (P < 0.0001). These hemodynamic responses were graded in a further five pigs. Moreover, while blockade of muscarinic cholinoceptors (n = 5) and of α-adrenoceptors (n = 5) did not abolish the observed responses, β1-adrenoceptors blocker (n = 5) prevented the effects of hCG on cardiac function. In addition, β2-adrenoceptors (n = 5) and NO synthase inhibition (n = 5) abolished the coronary response and the effect of hCG on NO release. In CEC, hCG induced the phosphorylation of endothelial NO synthase through cAMP/PKA signaling and ERK1/2, Akt, p38 MAPK involvement, which were activated as downstream effectors of β2-adrenoceptor stimulation. In conclusion, in anesthetized pigs, hCG primarily increased cardiac function and perfusion through the involvement of β-adrenoceptors and NO release. Moreover, cAMP/PKA-dependent kinases phosphorylation was found to play a role in eliciting the observed NO production in CEC.

  6. In vivo induction of human chorionic gonadotropin by osmotic pump advances sexual maturation during pre-spawning phase in adult catfish.

    PubMed

    Murugananthkumar, Raju; Prathibha, Yarikipati; Senthilkumaran, Balasubramanian; Rajakumar, Anbazhagan; Kagawa, Hirohiko

    2016-10-05

    Gonadal maturation is a critical event wherein gonads, under the influence of several hormones and factors, undergo cyclic morphological and physiological changes to produce functional gametes during the spawning phase. However, artificial induction can be effectively used to advance the maturation of gonad vis-à-vis spawning like behavior in seasonal breeders during the off-breeding season. In the present study, osmotic pumps loaded with 5000IU of human chorionic gonadotropin (hCG) or saline as control were implanted intraperitoneally for 21days during the pre-spawning phase (May-June) in catfish Clarias batrachus and C. gariepinus. Significant increase in gonado-somatic index and sperm motility, and in the levels of certain sex steroids were observed in the hCG treated catfish when compared to control while estradiol-17β (E2) was low. Histological analysis in hCG treated testis revealed densely packed sperm and/or spermatids inside the lumen wherein the control testis displayed normal characteristics of the pre-spawning phase. In females, histological analysis showed a significant increase in post-vitellogenic full-grown immature follicles as seen in the spawning phase. In accordance with this, the steroid hormone profile correlated well with steroidogenic shift from E2 to 17α,20β-DP indicating oocyte maturation. However, in the control ovaries of C. batrachus, perinucleolar and pre-vitellogenic oocytes were seen to be predominant. In addition, when compared with the control, the hCG treated group displayed a significant increase in the transcripts of several genes associated with gonadal growth. Taken together, artificial induction by slow release of hCG is an effective strategy to advance sexual maturation in catfish in a programmed manner.

  7. 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

  8. Expression of luteinizing hormone and chorionic gonadotropin receptor messenger ribonucleic acid in human corpora lutea during menstrual cycle and pregnancy.

    PubMed

    Nishimori, K; Dunkel, L; Hsueh, A J; Yamoto, M; Nakano, R

    1995-04-01

    In the present study, we examined the expression of LH and CG receptor messenger RNA (mRNA) in human corpora lutea (CL) during the menstrual cycle and pregnancy. Poly(A)-enriched RNA was extracted from CL and analyzed by Northern and slot blots, using a radiolabeled complementary RNA probe derived from the human LH receptor complementary DNA. Northern blot analysis indicated the presence of multiple LH receptor mRNA transcripts with molecular sizes of 8.0, 7.0 and 4.5 kilobases in human CL during the menstrual cycle. The predominant transcript was 4.5 kilobases in size. However, no hybridization signals were observed in nongonadal tissues (heart, liver, and kidney). Densitometric analyses revealed that the levels of LH receptor mRNA increased from early luteal phase to midluteal phase and subsequently decreased during late luteal phase. After the onset of menstruation, the LH receptor mRNA level was undetectable in the regressing CL. Moreover, radioligand receptor assay (RRA) showed a close parallelism between LH receptor mRNA levels and LH receptor content in CL throughout the menstrual cycle. LH receptor mRNA expression was also found in CL during early pregnancy. The level of LH receptor mRNA was relatively high in early pregnancy CL, whereas LH receptor content was low. Using in situ hybridization, LH receptor mRNAs were uniformly expressed in both large and small luteal cells during early and midluteal phase and early pregnancy, but not in regressing CL. In conclusion, these data demonstrate that the regulation of LH receptor content in human CL during luteal phase is associated with similar changes in the receptor message levels, suggesting the physiological roles for LH receptor mRNA during the menstrual cycle in the human. In addition, the expression of LH receptor mRNA was demonstrated in human CL during early pregnancy.

  9. 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

    Chung, Sokjoong; Rho, Seungsoo; Kim, Gijin; Kim, So-Ra; Baek, Kwang-Hyun; Kang, Myungseo; Lew, Helen

    2016-05-01

    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

  10. 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

  11. Chorionic gonadotropin and its receptor are both expressed in human retina, possible implications in normal and pathological conditions.

    PubMed

    Dukic-Stefanovic, Sladjana; Walther, Jan; Wosch, Sebastian; Zimmermann, Gerolf; Wiedemann, Peter; Alexander, Henry; Claudepierre, Thomas

    2012-01-01

    Extra-gonadal role of gonadotropins has been re-evaluated over the last 20 years. In addition to pituitary secretion of luteinizing hormone (LH) and follicle stimulating hormone (FSH), the CNS has been clearly identified as a source of hCG acting locally to influence behaviour. Here we demonstrated that human retina is producing this gonadotropin that acts as a neuroactive molecule. Müller glial and retinal pigmented epithelial (RPE) cells are producing hCG that may affects neighbour cells expressing its receptor, namely cone photoreceptors. It was previously described that amacrine and retinal ganglion (RGC) cells are targets of the gonadotropin releasing hormone that control the secretion of all gonadotropins. Therefore our findings suggest that a complex neuroendocrine circuit exists in the retina, involving hCG secreting cells (glial and RPE), hCG targets (photoreceptors) and hCG-release controlling cells (amacrine and RGC). The exact physiological functions of this circuit have still to be identified, but the proliferation of photoreceptor-derived tumor induced by hCG demonstrated the need to control this neuroendocrine loop.

  12. Specific binding of /sup 125/I-labeled human chorionic gonadotropin to gonadal tissue: comparison of limited-point saturation analyses to Scatchard analyses for determining binding capacities and factors affecting estimates of binding capacity

    SciTech Connect

    Spicer, L.J.; Ireland, J.J.

    1986-07-01

    Experiments were conducted to compare gonadotropin binding capacity calculated from limited-point saturation analyses to those obtained from Scatchard analyses, and to test the effects of membrane purity and source of gonadotropin receptors on determining the maximum percentage of radioiodinated hormone bound to receptors (maximum bindability). One- to four-point saturation analyses gave results comparable to results by Scatchard analyses when examining relative binding capacities of receptors. Crude testicular homogenates had lower estimates of maximum bindability of /sup 125/I-labeled human chorionic gonadotropin than more purified gonadotropin receptor preparations. Under similar preparation techniques, some gonadotropin receptor sources exhibited low maximum bindability.

  13. 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

  14. 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

  15. Ultra-high sensitivity of the non-immunological affinity of graphene oxide-peptide-based surface plasmon resonance biosensors to detect human chorionic gonadotropin.

    PubMed

    Chiu, Nan-Fu; Kuo, Chia-Tzu; Lin, Ting-Li; Chang, Chia-Chen; Chen, Chen-Yu

    2017-03-07

    Specific peptide aptamers can be used in place of expensive antibody proteins, and they are gaining increasing importance as sensing probes due to their potential in the development of non-immunological assays with high sensitivity, affinity and specificity for human chorionic gonadotropin (hCG) protein. We combined graphene oxide (GO) sheets with a specific peptide aptamer to create a novel, simple and label-free tool to detect abnormalities at an early stage of pregnancy, a GO-peptide-based surface plasmon resonance (SPR) biosensor. This is the first binding interface experiment to successfully demonstrate binding specificity in kinetic analysis biomechanics in peptide aptamers and GO sheets. In addition to the improved affinity offered by the high compatibility with the target hCG protein, the major advantage of GO-peptide-based SPR sensors was their reduced nonspecific adsorption and enhanced sensitivity. The calculation of total electric field intensity (ΔE) in the GO-based sensing interfaces was significantly enhanced by up to 1.2 times that of a conventional SPR chip. The GO-peptide-based chip (1mM) had a high affinity (KA) of 6.37×10(12)M(-1), limit of detection of 0.065nM and ultra-high sensitivity of 16 times that of a conventional SPR chip. The sensitivity of the slope ratio of the low concentration hCG protein assay in linear regression analysis was GO-peptide (1mM): GO-peptide (0.1mM): conventional chip (8-mercaptooctanoic acid)-peptide (0.1mM)=8.6: 3.3: 1. In summary, the excellent binding affinity, low detection limit, high sensitivity, good stability and specificity suggest the potential of this GO-peptide-based SPR chip detection method in clinical application. The development of real-time whole blood analytic and diagnostic tools to detect abnormalities at an early stage of pregnancy is a promising technique for future clinical application.

  16. Gonadotropin-releasing hormone agonist trigger is a better alternative than human chorionic gonadotropin in PCOS undergoing IVF cycles for an OHSS Free Clinic: A Randomized control trial

    PubMed Central

    Krishna, Deepika; Dhoble, Snehal; Praneesh, Gautham; Rathore, Suvarna; Upadhaya, Amit; Rao, Kamini

    2016-01-01

    OBJECTIVE: The objective of this study is to evaluate if gonadotropin-releasing hormone agonist (GnRHa) trigger is a better alternative to human chorionic gonadotropin (hCG) in polycystic ovary syndrome (PCOS) of Indian origin undergoing in vitro fertilization (IVF) cycles with GnRH antagonist for the prevention of ovarian hyperstimulation syndrome (OHSS). DESIGN: Prospective randomized control trial. SETTING: Tertiary care center. MATERIALS AND METHODS: A total of 227 patients diagnosed with PCOS, undergoing IVF in an antagonist protocol were recruited and randomly assigned into two groups: Group A (study group): GnRHa trigger 0.2 mg (n = 92) and Group B (control group): 250 μg of recombinant hCG as trigger (n = 101) 35 h before oocyte retrieval. We chose segmentation strategy, freezing all embryos in both the groups. STATISTICAL ANALYSIS: Continuous variables were expressed as mean ± standard deviation independent sample t-test and Kolmogorov-Smirnov test were used for continuous variables which were normally distributed and Mann-Whitney U-test for data not normally distributed. MAIN OUTCOME MEASURES: Primary outcome: OHSS (mild, moderate, and severe) rates. Secondary outcomes: Maturity rate of the oocytes, fertilization rate, availability of top quality embryos on day 3 (Grade 1 and Grade 2). RESULTS The incidence of moderate to severe OHSS in the hCG group was 37.6% and 0% in the GnRHa group with P < 0.001. The GnRHa group had significantly more mature oocytes retrieved (19.1 ± 11.7 vs. 14.1 ± 4.3), more fertilized oocytes (15.6 ± 5.6 vs. 11.7 ± 3.6), and a higher number of top quality cleavage embryos on day 3 (12.9 ± 4.7 vs. 7.5 ± 4.3) than the hCG group. CONCLUSIONS: The most effective strategy which significantly eliminates the occurrence of OHSS in PCOS following ovarian stimulation in antagonist IVF cycles is the use of GnRHa trigger yielding more mature oocytes and good quality embryos when compared with hCG trigger. PMID:27803584

  17. 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.

  18. Amniocentesis and chorionic villus sampling.

    PubMed Central

    Shulman, L P; Elias, S

    1993-01-01

    Amniocentesis and chorionic villus sampling have been shown through prospective, multicenter trials to be safe and effective methods of prenatal diagnosis; accordingly, a knowledge of these tests is important for those physicians who care for women during their childbearing years. We review the indications, techniques, safety, accuracy, and efficacy of amniocentesis and chorionic villus sampling and compare the advantages and disadvantages of each diagnostic test. This review should enable physicians to provide appropriate counseling and information to women at increased risk for fetal abnormalities detectable by either of these procedures. Images PMID:8236967

  19. Pharmacological identification of P2X1, P2X4 and P2X7 nucleotide receptors in the smooth muscles of human umbilical cord and chorionic blood vessels.

    PubMed

    Valdecantos, P; Briones, R; Moya, P; Germain, A; Huidobro-Toro, J P

    2003-01-01

    To ascertain the role of extracellular adenosine 5'-triphosphate (ATP) receptors in human placenta circulation, we identified and pharmacologically characterized the P2X receptor population in its superficial vessels. Total RNA was extracted from segments of chorionic and umbilical arteries and veins of terminal placentae delivered by vaginal or Caesarian births. Polymerase chain reaction (PCR), followed by sequencing of the products, identified the presence of P2X 1, 4, 5, 6, and 7mRNAs in smooth muscle from chorionic and umbilical arteries and veins. Umbilical vessels proximal to the fetus expressed the same population of P2X subtypes, except for the P2X(5), but additionally expressed the P2X(2). Rings of chorionic vessels contracted upon addition of nucleotides and analogs with the following relative rank order of potencies in arteries and veins: alpha,beta-methyleneATP>beta,gamma-methyleneATP>PNP>ATP=diBzATP>2-MeSATP>ADP>AMP; in umbilical vessels alpha,beta-methyleneATP was at least 100-fold more potent than ATP. Nucleotide potency was less than that of PGF(2alpha) or endothelin-2, but had the same magnitude as serotonin. ATP-desensitized receptors evidenced cross desensitization to alpha,beta-methyleneATP, 2-MeSATP and diBzATP, effect not observed when desensitization was elicited by alpha,beta-methyleneATP, confirming the presence of various P2X receptor subtypes in the smooth muscles of these vessels. The vasocontractile efficacy of alpha,beta-methyleneATP was unaltered by endothelium removal, while that of ATP was significantly attenuated and those elicited by 2-MeSATP were blunted, indicating the presence of additional endothelial nucleotide receptors. These results suggest that P2X receptors participate in the humoral regulation of placental blood flow.

  20. 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 SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM...

  1. 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 SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM...

  2. 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 SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM...

  3. 21 CFR 522.1079 - Serum gonadotropin and chorionic gonadotropin.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Serum gonadotropin and chorionic gonadotropin. 522.1079 Section 522.1079 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM...

  4. 21 CFR 522.1079 - Serum gonadotropin and chorionic gonadotropin.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Serum gonadotropin and chorionic gonadotropin. 522.1079 Section 522.1079 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS IMPLANTATION OR INJECTABLE DOSAGE FORM...

  5. 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).

  6. Integrated and Quantitative Proteomics of Human Tumors.

    PubMed

    Yakkioui, Y; Temel, Y; Chevet, E; Negroni, L

    2017-01-01

    Quantitative proteomics represents a powerful approach for the comprehensive analysis of proteins expressed under defined conditions. These properties have been used to investigate the proteome of disease states, including cancer. It has become a major subject of studies to apply proteomics for biomarker and therapeutic target identification. In the last decades, technical advances in mass spectrometry have increased the capacity of protein identification and quantification. Moreover, the analysis of posttranslational modification (PTM), especially phosphorylation, has allowed large-scale identification of biological mechanisms. Even so, increasing evidence indicates that global protein quantification is often insufficient for the explanation of biology and has shown to pose challenges in identifying new and robust biomarkers. As a consequence, to improve the accuracy of the discoveries made using proteomics in human tumors, it is necessary to combine (i) robust and reproducible methods for sample preparation allowing statistical comparison, (ii) PTM analyses in addition to global proteomics for additional levels of knowledge, and (iii) use of bioinformatics for decrypting protein list. Herein, we present technical specificities for samples preparation involving isobaric tag labeling, TiO2-based phosphopeptides enrichment and hydrazyde-based glycopeptides purification as well as the key points for the quantitative analysis and interpretation of the protein lists. The method is based on our experience with tumors analysis derived from hepatocellular carcinoma, chondrosarcoma, human embryonic intervertebral disk, and chordoma experiments.

  7. JEG-3 Trophoblast Cells Producing Human Chorionic Gonadotropin Promote Conversion of Human CD4+FOXP3- T Cells into CD4+FOXP3+ Regulatory T Cells and Foster T Cell Suppressive Activity.

    PubMed

    Poloski, Eileen; Oettel, Anika; Ehrentraut, Stefanie; Luley, Lydia; Costa, Serban Dan; Zenclussen, Ana Claudia; Schumacher, Anne

    2016-03-09

    The pregnancy hormone human Chorionic Gonadotropin (hCG) reportedly modulates innate and adaptive immune responses and contributes thereby to fetal survival. More precisely, hCG has been shown to support human Treg cell homing into the fetal-maternal interface and enhance number and function of Treg cells in murine pregnancy. Here, we aimed to study whether hCG and hCG-producing human trophoblast cell lines induce Treg cells from CD4(+)FOXP3(-) T cells and promote T cell suppressive activity. CD4(+)FOXP3(-) T cells were isolated from peripheral blood of normal pregnant women and cultured in the presence of hCG-producing (JEG-3, HTR-8) and non-producing (SWAN-71) cell lines. To confirm the participation of hCG in Treg cell conversion, the experiments were performed in the presence of anti-hCG and additional experiments were run with recombinant or urine-purified hCG. After culture the number of CD4(+)FOXP3(+) Treg cells as well as the suppressive capacity of total T cells was assessed. hCG-producing JEG-3 cells as well as recombinant and urine-purified hCG induced CD4(+)FOXP3(+) Treg cells from CD4(+)FOXP3(-) T cells. Blockage of hCG impaired Treg cell induction. Moreover, hCG-producing JEG-3 cells increased suppressive activity of CD4(+)FOXP3(-) T cells through an antigen-independent pathway. Our results propose another mechanism through which hCG modulates the female immune system during pregnancy in favor of the fetus.

  8. Constitutive production of multiple cytokines and a human chorionic gonadotrophin beta-subunit by a human bladder cancer cell line (KU-19-19): possible demonstration of totipotential differentiation.

    PubMed Central

    Tachibana, M.; Miyakawa, A.; Nakashima, J.; Murai, M.; Nakamura, K.; Kubo, A.; Hata, J. I.

    1997-01-01

    Bladder cancer cells have been shown to secrete a variety of factors that are not related to cells of urothelial origin. The histogenesis of these tumour developments is uncertain, and a variety of theories have been previously reported. In the present manuscript, we identify the factors constitutively produced by a human bladder cancer cell line (KU-19-19) that was found to produce beta human chorionic gonadotrophin (beta-hCG), granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin 1alpha (IL-1alpha), interleukin 6 (IL-6) and interleukin 8 (IL-8). The cells were obtained from a case of metastatic carcinoma that was originally diagnosed to be a grade 3 (WHO classification), invasive transitional cell carcinoma of the bladder. On microscopic observation, the cultured cells exhibited an epithelial appearance with vacuole formation in their cytoplasm. Ultrastructural observations revealed relatively marked microvilli and a tight junction. Significant amounts of beta-hCG, G-CSF, GM-CSF, IL-1alpha, IL-6 and IL-8 concentrations in the supernatant from cultured cells were demonstrated by enzyme-linked immunosorbent assays, while the expression of mRNA of these marker proteins in cancer cells was also significantly exhibited by reverse transcription polymerase chain reaction (RT-PCR). In addition, the expression of G-CSF receptor and IL-6 receptor mRNA was also shown by RT-PCR. Xenograft transplantability using nude mice was observed in association with the presence of severe neutrophilia in the peripheral blood. These results indicate that this cell line appears to be an effective model for the study of transitional cell carcinoma of the bladder with multipotent differentiation potentials. Images Figure 1 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 PMID:9231915

  9. Fetal blood flow in branching models of the chorionic arterial vasculature.

    PubMed

    Gordon, Zoya; Eytan, Osnat; Jaffa, Ariel J; Elad, David

    2007-04-01

    Fetal development depends on adequate exchange of materials between the fetus and maternal circulatory systems, which requires optimal distribution of blood vessels over the chorionic plate to ensure perfusion of the whole placental volume. Based on a previous investigation of the architecture of the chorionic vessels in the human placenta, we developed in this study typical models for the dichotomous and monopodial segments of the chorionic arteries of a mature placenta. Each model also included some intraplacental (IP) vessels that branch off into the cotyledons perpendicular to the chorionic arteries. Computational analysis of steady blood flow through these models was performed to explore the distribution of fetal blood over the chorionic plate. The results demonstrated that energy losses are small in the monopodial model, which explains their efficient delivery of fetal blood over the chorionic plate in cases of a marginal cord insertion. On the other hand, the dichotomous model is efficient in distributing a relatively large volume of blood over large areas near the bifurcation. Accordingly, the combination of dichotomous and monopodial bifurcation in a normal chorionic plate ensures a uniform blood perfusion of the placenta. Simulations with narrow daughter and IP vessels did not result in significant changes in the main mother tubes, supporting clinical observations in which umbilical blood flow remains normal although some peripheral vessels may be occluded.

  10. [Prenatal diagnosis using chorionic villi].

    PubMed

    Vega Hernández, M E; Hicks, J J; González-Angulo, J

    1991-07-01

    Chorionic villus sampling (CVS) has a promising future about early detection of fetal abnormalities. It has the potential to become a major tool in the prenatal diagnosis and therapy of genetic disorders. Villus samples can be analyzed by means of cytogenetic, biochemical or molecular technics. Information available at present indicates fetal loss rate should be in the same proportion than amniocentesis. CVS appears to be a reasonably safe and reliable method of prenatal diagnosis in the first trimester of pregnancy. This procedure is setting as fast as it is possible like an excellent alternative to amniocentesis.

  11. 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.

  12. 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-...

  13. Measurement of plasma human chorionic gonadotropin (hCG) and beta-hCG activities in the late luteal phase: evidence of the occurrence of spontaneous menstrual abortions in infertile women.

    PubMed

    Chartier, M; Roger, M; Barrat, J; Michelon, B

    1979-02-01

    Plasma luteinizing hormone (LH-human chorionic gonadotropin (hCG) and beta-hCG activities were measured during the late luteal phase in 321 cycles of 147 infertile women. In 71 cycles the hCG measurement permitted the diagnosis of pregnancy between the 10th and 14th days after the thermal nadir. The slope of the regression line derived from hCG levels during the first 22 days of pregnancy was significantly lower in pregnancies which aborted before the 60th day than in normal pregnancies (P less than 0.01). Among 72 cycles ended by apparently normal menses which exhibited an LH-hCG activity at least equal to 7 mIU of hCG/ml during the late luteal phase, the beta-hCG activity was measured in 49 cycles during which hCG had not been given. Significant beta-hCG activity (greater than or equal to 4 mIU of hCG/ml) was detected in 19 cases. This finding supports the assumption that secretory trophoblastic tissue had been present and that spontaneous menstrual abortions had occurred in these women.

  14. Human chorionic gonadotropin (hCG) in the secretome of cultured embryos: hyperglycosylated hCG and hCG-free beta subunit are potential markers for infertility management and treatment.

    PubMed

    Butler, Stephen A; Luttoo, Jameel; Freire, Maísa O T; Abban, Thomas K; Borrelli, Paola T A; Iles, Ray K

    2013-09-01

    Human chorionic gonadotropin (hCG) is produced by trophoblast cells throughout pregnancy, and gene expression studies have indicated that hCG-beta subunit (hCGβ) expression is active at the 2 blastomere stage. Here, we investigated the qualitative hCG output of developing embryos in culture and hCG isoforms expressed in the secretome as a novel sensitive method for detecting hCG. Culture media was collected from the culture plates of 118 embryos in culture (including controls and embryos at different stages of culture) from 16 patients undergoing routine fertility treatment. The hCGβ was detectable in media from 2 pronuclear (2PN) stage embryos through to the blastocyst stage. The hCGβ was absent in 1PN and arrested embryos as well as all media controls. Prior to hatching, hyperglycosylated hCG (hCGh) was observed selectively in 3PN embryos, but after hatching, along with hCG, became the dominant hCG molecule observed. We have reported at the 2PN stage the earliest evidence of hCGβ expression in embryos. There is a suggestion this may be indicative of quality in early embryos, and hCGh seen at the pronuclear stage may suggest triploid abnormality. The dominance of hCG, and hCGh expression, seen after blastocyst hatching may be indicative of potential implantation success. Thus, hCG isoforms have potential roles as biomarkers of embryo viability for embryo/blastocyst transfer.

  15. Maternal and Fetal Mechanisms of B Cell Regulation during Pregnancy: Human Chorionic Gonadotropin Stimulates B Cells to Produce IL-10 While Alpha-Fetoprotein Drives Them into Apoptosis

    PubMed Central

    Fettke, Franziska; Schumacher, Anne; Canellada, Andrea; Toledo, Natalia; Bekeredjian-Ding, Isabelle; Bondt, Albert; Wuhrer, Manfred; Costa, Serban-Dan; Zenclussen, Ana Claudia

    2016-01-01

    Maternal immune tolerance toward the fetus is an essential requisite for pregnancy. While T cell functions are well documented, little is known about the participation of B cells. We have previously suggested that IL-10-producing B cells are involved in pregnancy tolerance in mice and humans. By employing murine and human systems, we report now that fetal trophoblasts positively regulate the generation of IL-10-producing B cells. We next studied the participation of hormones produced by the placenta as well as the fetal protein alpha-fetoprotein (AFP) in B cell modulation. Human chorionic gonadotropin (hCG), but not progesterone, estrogen, or a combination of both, was able to promote changes in B cell phenotype and boost their IL-10 production, which was abolished after blocking hCG. The hCG-induced B cell phenotype was not associated with augmented galactosylation, sialylation, or fucosylation of IgG subclasses in their Fc. In vitro, hCG induced the synthesis of asymmetrically glycosylated antibodies in their Fab region. Interestingly, AFP had dual effects depending on the concentration. At concentrations corresponding to maternal serum levels, it did not modify the phenotype or IL-10 secretion of B cells. At fetal concentrations, however, AFP was able to drive B cells into apoptosis, which may indicate a protective mechanism to avoid maternal B cells to reach the fetus. Our data suggest that the fetus secrete factors that promote a pregnancy-friendly B cell phenotype, unraveling interesting aspects of B cell function, and modulation by pregnancy hormones and fetal proteins. PMID:28008329

  16. 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

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

    2016-02-01

    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.

  17. 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...

  18. Expression and production of human chorionic gonadotropin (hCG) in the normal secretory endometrium: evidence of CGB7 and/or CGB6 beta hCG subunit gene expression.

    PubMed

    Zimmermann, Gerolf; Ackermann, Wilfried; Alexander, Henry

    2012-03-01

    We have previously confirmed glandular cell CGB and CGA subunit mRNA gene expression as well as the expression of their dimeric and single-subunit human chorionic gonadotropin (hCG) proteins in normal secretory transformed endometrium. The objective of this study was to investigate the endometrial epithelial gene locus of the human hCG/LH gene cluster from CGB genes responsible for gene expression. For this study, endometrial specimens were selected from women characterized using our endometrium score and hCG staining index that had normal secretory transformed endometrium and optimal hCG staining. Using full-length CGB mRNA sequence analysis, we found that epithelial CGB is (co)expressed as the product of gene locus CGB7 and CGB6 (48%), as single CGB7 (42%), or to a lower percentage as single CGB6 (10%). In addition to known differences between these genes and CGB5, the nucleotide sequence of the mRNA differs between CGB7 and CGB6 in the untranslated promoter region and in translated exon 2. Immunohistochemical results show that endometrial joint CGB7 and CGB6, single CGB7, and single CGB6 mRNA expression lead to the release of endometrial hCG. Gene-specific antibodies for CGB7 reveal secretory endometrial hCG production, which is not observed for gene-specific CGB5 antibodies, whereas the placenta is positive for CGB5 and negative for CGB7 antibody as revealed by immunohistochemistry and Western blot hCG isoform analysis. Only endometrial CGB7 expression seems to be supported specifically by secretory endometrial transcription factors. In conclusion, epithelial hCG is expressed and produced as CGB7 and/or CGB6 but not CGB5, and it is produced together with CGA as a secretory transformation marker in the normal secretory phase endometrium.

  19. 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.

  20. Radioimmunochemical quantitation of human adenosine deaminase.

    PubMed Central

    Daddona, P E; Frohman, M A; Kelley, W N

    1979-01-01

    Markedly reduced or absent adenosine deaminase activity in man is associated with an autosomal recesive form of severe conbined immunodeficiency disease. To further define the genetic nature of this enzyme defect, we have quantitated immunologically active adenosine deaminase (CRM) in the hemolysate of homozygous deficient patients and their heterozygous parents. A highly specific radioimmunoassay was developed capable of detecting 0.05% of normal erythrocyte adenosine deaminase. Hemolysates from nine heterozygotes (five families) showed a wide range in CRM (32--100% of normal) and variable absolute specific activities with several being at least 1 SD BELOW THE NORMAL MEAN. Hemolysates from four unrelated patients showed less than 0.09% adenosine deaminase activity with CRM ranging from less than 0.06 to 5.6% of the normal mean. In conclusion, heterozygote and homozygote hemolysates from five of the eight families analyzed revealed variable levels of CRM suggesting heterogeneous genetic alteration or expression of the silent or defective allele(s) of adenosine deaminase. PMID:468994

  1. 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.

  2. 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

  3. A quantitative theory of human color choices.

    PubMed

    Komarova, Natalia L; Jameson, Kimberly A

    2013-01-01

    The system for colorimetry adopted by the Commission Internationale de l'Eclairage (CIE) in 1931, along with its subsequent improvements, represents a family of light mixture models that has served well for many decades for stimulus specification and reproduction when highly controlled color standards are important. Still, with regard to color appearance many perceptual and cognitive factors are known to contribute to color similarity, and, in general, to all cognitive judgments of color. Using experimentally obtained odd-one-out triad similarity judgments from 52 observers, we demonstrate that CIE-based models can explain a good portion (but not all) of the color similarity data. Color difference quantified by CIELAB ΔE explained behavior at levels of 81% (across all colors), 79% (across red colors), and 66% (across blue colors). We show that the unexplained variation cannot be ascribed to inter- or intra-individual variations among the observers, and points to the presence of additional factors shared by the majority of responders. Based on this, we create a quantitative model of a lexicographic semiorder type, which shows how different perceptual and cognitive influences can trade-off when making color similarity judgments. We show that by incorporating additional influences related to categorical and lightness and saturation factors, the model explains more of the triad similarity behavior, namely, 91% (all colors), 90% (reds), and 87% (blues). We conclude that distance in a CIE model is but the first of several layers in a hierarchy of higher-order cognitive influences that shape color triad choices. We further discuss additional mitigating influences outside the scope of CIE modeling, which can be incorporated in this framework, including well-known influences from language, stimulus set effects, and color preference bias. We also discuss universal and cultural aspects of the model as well as non-uniformity of the color space with respect to different

  4. 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.

  5. Differential placental expression profile of human Growth Hormone/Chorionic Somatomammotropin genes in pregnancies with pre-eclampsia and gestational diabetes mellitus

    PubMed Central

    Männik, Jaana; Vaas, Pille; Rull, Kristiina; Teesalu, Pille; Laan, Maris

    2012-01-01

    The human GH/CSH cluster consisting of one pituitary-expressed (GH1) and four placenta-expressed loci has been implicated in maternal metabolic adaptation to pregnancy, regulation of intrauterine and postnatal growth. We investigated how the mRNA expression profile of placental GH2, CSH1 and CSH2 genes and their alternative transcripts correlates with maternal pre-eclampsia (PE) and/or gestational diabetes mellitus (GD). The expression of studied genes in PE placentas (n = 17) compared to controls (n = 17) exhibited a trend for reduced transcript levels. The alternative transcripts retaining intron 4, GH2-2 and CSH1-2 showed significantly reduced expression in PE cases without growth restriction (P = 0.007, P = 0.008, respectively). In maternal GD (n = 23), a tendency of differential expression was detected only for the GH2 gene and in pregnancies with large-for-gestational-age newborns. Our results, together with those reported by others, are consistent with a pleiotropic effect of placental hGH/CSH genes at the maternal-fetal interface relating to the regulation of fetal growth and the risk of affected maternal metabolism. PMID:22387044

  6. In vitro effect of 4-nonylphenol on human chorionic gonadotropin (hCG) stimulated hormone secretion, cell viability and reactive oxygen species generation in mice Leydig cells.

    PubMed

    Jambor, Tomáš; Tvrdá, Eva; Tušimová, Eva; Kováčik, Anton; Bistáková, Jana; Forgács, Zsolt; Lukáč, Norbert

    2017-03-01

    Nonylphenol is considered an endocrine disruptor and has been reported to affect male reproductive functions. In our in vitro study, we evaluated the effects of 4-nonylphenol (4-NP) on cholesterol levels, hormone formation and viability in cultured Leydig cells from adult ICR male mice. We also determined the potential impact of 4-NP on generation of reactive oxygen species (ROS) after 44 h of cultivation. The cells were cultured with addition of 0.04; 0.2; 1.0; 2.5 and 5.0 μg/mL of 4-NP in the present of 1 IU/mL human chorionic gonadotropin (hCG) and compared to the control. The quantity of cholesterol was determined from culture medium using photometry. Determination of hormone production was performed by enzyme-linked immunosorbent assay. Metabolic activity assay was used for quantification of cell viability. The chemiluminescence technique, which uses a luminometer to measure reactive oxygen species, was employed. Applied doses of 4-NP (0.04-5.0 μg/mL) slight increase cholesterol levels and decrease production of dehydroepiandrosterone after 44 h of cultivation, but not significantly. Incubation of 4-NP treated cells with hCG significantly (P < 0.001) inhibited androstenedione, but not testosterone, formation at the highest concentration (5.0 μg/mL). The viability was significantly (P < 0.05); (P < 0.001) increased at 1.0; 2.5 and 5.0 μg/mL of 4-NP after 44 h treatment. Furthermore, 44 h treatment of 4-NP (0.04-5.0 μg/mL) caused significant (P < 0.001) intracellular accumulation of ROS in exposed cells. Taken together, the results of our in vitro study reported herein is consistent with the conclusion that 4-nonylphenol is able to influence hormonal profile, cell viability and generate ROS.

  7. Mobilities of the inner three core residues and the Man(alpha 1--6) branch of the glycan at Asn78 of the alpha-subunit of human chorionic gonadotropin are restricted by the protein.

    PubMed

    van Zuylen, C W; de Beer, T; Leeflang, B R; Boelens, R; Kaptein, R; Kamerling, J P; Vliegenthart, J F

    1998-02-17

    Human chorionic gonadotropin (hCG) is a heterodimeric glycoprotein hormone involved in the maintenance of the corpus luteum in early pregnancy. The free alpha-subunit of hCG has a biological activity of its own, namely, stimulation of prolactin secretion from term pregnancy decidual cells [Blithe, D. L., et al. (1991) Endocrinology 129, 2257-2259]. Glycosylation at Asn78 of the alpha-subunit is required for the stability of the protein, but the exact nature of the stabilizing effect is not known. In our previous study, it was indicated that GlcNAc-1 at Asn78 has a reduced mobility, whereas the glycan at Asn52 is highly mobile [De Beer, T., et al. (1996) Eur. J. Biochem. 241, 229-242]. In the present investigation, it is shown that the PNGase F susceptibility of the Asn52-linked glycan in the free alpha-subunit is absent in the heterodimer. Thus, the high mobility of the glycan at Asn52 may be characteristic for the free alpha-subunit. For accurate modeling of alpha hCG, knowledge of the behavior of each of the glycans is essential. In this context, the mobility of the glycans and their interactions with the protein are explored by NMR spectroscopy using desialylated, partially deglycosylated free alpha-subunit (as-pd alpha) carrying glycans at Asn78 only. NOEs between GlcNAc-2 and several amino acid residues indicate that GlcNAc-2 is involved in stabilizing alpha hCG. From the values of 13C relaxation parameters T2 and T1 rho of the constituting monosaccharide residues, it was concluded that the inner three residues have a severely restricted mobility. The Man-4 and Man-4' residues of the diantennary oligosaccharide exhibit a similar relaxation behavior, suggesting that the Man-4' branch occurs in a single conformation of the C5-C6 linkage of Man-3 instead of in rapidly interconverting conformations that are known to exist for this linkage for the free oligosaccharide.

  8. Transcervical chorionic villus sampling: a practical guide.

    PubMed

    Stergiotou, Iosifina; Borobio, Virginia; Bennasar, Mar; Goncé, Anna; Mula, Raquel; Nuruddin, Mohammed; Soler, Anna; Borrell, Antoni

    2016-01-01

    First trimester screening for fetal aneuploidies has made the implementation of diagnostic techniques essential. Chorionic villus sampling (CVS) is the method of choice for obtaining chorionic villi for molecular and cytogenetic analysis in the first trimester. Two techniques have been developed, a transcervical and a transabdominal. The selection criteria have been based historically on factors, such as placental location, parity, maternal weight and preference of the operator. In our institution, we developed an elevated level of expertise in the field of transcervical approach, resulting in good quality of samples and comparable fetal loss rate to other approaches. Despite three decades of transcervical CVS performance, little consensus in terms of its technique and clinical guidelines exists. Considering the expertise and the volume of procedures performed at our center, we suggest a practical clinical guideline for transcervical CVS.

  9. A rapid chemiluminescent method for quantitation of human DNA.

    PubMed Central

    Walsh, P S; Varlaro, J; Reynolds, R

    1992-01-01

    A sensitive and simple method for the quantitation of human DNA is described. This method is based on probe hybridization to a human alpha satellite locus, D17Z1. The biotinylated probe is hybridized to sample DNA immobilized on nylon membrane. The subsequent binding of streptavidin-horseradish peroxidase to the bound probe allows for chemiluminescent detection using a luminol-based reagent and X-ray film. Less than 150 pg of human DNA can easily be detected with a 15 minute exposure. The entire procedure can be performed in 1.5 hours. Microgram quantities of nonhuman DNA have been tested and the results indicate very high specificity for human DNA. The data on film can be scanned into a computer and a commercially available program can be used to create a standard curve where DNA quantity is plotted against the mean density of each slot blot signal. The methods described can also be applied to the very sensitive determination of quantity and quality (size) of DNA on Southern blots. The high sensitivity of this quantitation method requires the consumption of only a fraction of sample for analysis. Determination of DNA quantity is necessary for RFLP and many PCR-based tests where optimal results are obtained only with a relatively narrow range of DNA quantities. The specificity of this quantitation method for human DNA will be useful for the analysis of samples that may also contain bacterial or other non-human DNA, for example forensic evidence samples, ancient DNA samples, or clinical samples. Images PMID:1408822

  10. A rapid chemiluminescent method for quantitation of human DNA.

    PubMed

    Walsh, P S; Varlaro, J; Reynolds, R

    1992-10-11

    A sensitive and simple method for the quantitation of human DNA is described. This method is based on probe hybridization to a human alpha satellite locus, D17Z1. The biotinylated probe is hybridized to sample DNA immobilized on nylon membrane. The subsequent binding of streptavidin-horseradish peroxidase to the bound probe allows for chemiluminescent detection using a luminol-based reagent and X-ray film. Less than 150 pg of human DNA can easily be detected with a 15 minute exposure. The entire procedure can be performed in 1.5 hours. Microgram quantities of nonhuman DNA have been tested and the results indicate very high specificity for human DNA. The data on film can be scanned into a computer and a commercially available program can be used to create a standard curve where DNA quantity is plotted against the mean density of each slot blot signal. The methods described can also be applied to the very sensitive determination of quantity and quality (size) of DNA on Southern blots. The high sensitivity of this quantitation method requires the consumption of only a fraction of sample for analysis. Determination of DNA quantity is necessary for RFLP and many PCR-based tests where optimal results are obtained only with a relatively narrow range of DNA quantities. The specificity of this quantitation method for human DNA will be useful for the analysis of samples that may also contain bacterial or other non-human DNA, for example forensic evidence samples, ancient DNA samples, or clinical samples.

  11. A Quantitative ADME-base Tool for Exploring Human ...

    EPA Pesticide Factsheets

    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 advances in exposure science are needed to support efficient and effective evaluation and management of chemical risks, particularly for chemicals in consumer products. The U.S. Environmental Protection Agency Office of Research and Development is developing, or collaborating in the development of, scientifically-defensible methods for making quantitative or semi-quantitative exposure predictions. The Exposure Prioritization (Ex Priori) model is a simplified, quantitative visual dashboard that provides a rank-ordered internalized dose metric to simultaneously explore exposures across chemical space (not chemical by chemical). Diverse data streams are integrated within the interface such that different exposure scenarios for “individual,” “population,” or “professional” time-use profiles can be interchanged to tailor exposure and quantitatively explore multi-chemical signatures of exposure, internalized dose (uptake), body burden, and elimination. Ex Priori has been designed as an adaptable systems framework that synthesizes knowledge from various domains and is amenable to new knowledge/information. As such, it algorithmically captures the totality of exposure across pathways. It

  12. Recognition of human IgM subgroups by quantitative measurement

    PubMed Central

    Klein, F.; De Bruyn, A. M.; Radema, H.

    1973-01-01

    By means of the single radial immunodiffusion technique it is possible to detect at least two subgroups of human IgM paraproteins, if the square ring diameter is plotted against absolute concentration. These groups give different calibration lines with the same antiserum, even when no qualitative differences can be detected by the double diffusion technique. The possibility of distinguishing these groups is not limited to the use of a single antiserum. The differences between the groups are abolished by reduction of the IgM paraprotein to 7S subunits and therefore seem to be of conformational nature. No identity with other known classifications has yet been found. In one out of two cases investigated differences in quantitative reactivity were found between 19S and >19S components in the same serum. The results suggest that quantitative methods may be used to detect differences in immunoglobulin structure, when qualitative methods fail. They show that different human IgM paraproteins cannot be directly compared quantitatively by the radial immunodiffusion method. PMID:4128890

  13. A quantitative transcriptome reference map of the normal human brain.

    PubMed

    Caracausi, Maria; Vitale, Lorenza; Pelleri, Maria Chiara; Piovesan, Allison; Bruno, Samantha; Strippoli, Pierluigi

    2014-10-01

    We performed an innovative systematic meta-analysis of 60 gene expression profiles of whole normal human brain, to provide a quantitative transcriptome reference map of it, i.e. a reference typical value of expression for each of the 39,250 known, mapped and 26,026 uncharacterized (unmapped) transcripts. To this aim, we used the software named Transcriptome Mapper (TRAM), which is able to generate transcriptome maps based on gene expression data from multiple sources. We also analyzed differential expression by comparing the brain transcriptome with those derived from human foetal brain gene expression, from a pool of human tissues (except the brain) and from the two normal human brain regions cerebellum and cerebral cortex, which are two of the main regions severely affected when cognitive impairment occurs, as happens in the case of trisomy 21. Data were downloaded from microarray databases, processed and analyzed using TRAM software and validated in vitro by assaying gene expression through several magnitude orders by 'real-time' reverse transcription polymerase chain reaction (RT-PCR). The excellent agreement between in silico and experimental data suggested that our transcriptome maps may be a useful quantitative reference benchmark for gene expression studies related to the human brain. Furthermore, our analysis yielded biological insights about those genes which have an intrinsic over-/under-expression in the brain, in addition offering a basis for the regional analysis of gene expression. This could be useful for the study of chromosomal alterations associated to cognitive impairment, such as trisomy 21, the most common genetic cause of intellectual disability.

  14. Function and failure of the fetal membrane: Modelling the mechanics of the chorion and amnion

    PubMed Central

    Oyen, Michelle L.; Phillips, Andrew T. M.; Nowlan, Niamh C.

    2017-01-01

    The fetal membrane surrounds the fetus during pregnancy and is a thin tissue composed of two layers, the chorion and the amnion. While rupture of this membrane normally occurs at term, preterm rupture can result in increased risk of fetal mortality and morbidity, as well as danger of infection in the mother. Although structural changes have been observed in the membrane in such cases, the mechanical behaviour of the human fetal membrane in vivo remains poorly understood and is challenging to investigate experimentally. Therefore, the objective of this study was to develop simplified finite element models to investigate the mechanical behaviour and rupture of the fetal membrane, particularly its constituent layers, under various physiological conditions. It was found that modelling the chorion and amnion as a single layer predicts remarkably different behaviour compared with a more anatomically-accurate bilayer, significantly underestimating stress in the amnion and under-predicting the risk of membrane rupture. Additionally, reductions in chorion-amnion interface lubrication and chorion thickness (reported in cases of preterm rupture) both resulted in increased membrane stress. Interestingly, the inclusion of a weak zone in the fetal membrane that has been observed to develop overlying the cervix would likely cause it to fail at term, during labour. Finally, these findings support the theory that the amnion is the dominant structural component of the fetal membrane and is required to maintain its integrity. The results provide a novel insight into the mechanical effect of structural changes in the chorion and amnion, in cases of both normal and preterm rupture. PMID:28350838

  15. Quantitative mass spectrometry of unconventional human biological matrices

    NASA Astrophysics Data System (ADS)

    Dutkiewicz, Ewelina P.; Urban, Pawel L.

    2016-10-01

    The development of sensitive and versatile mass spectrometric methodology has fuelled interest in the analysis of metabolites and drugs in unconventional biological specimens. Here, we discuss the analysis of eight human matrices-hair, nail, breath, saliva, tears, meibum, nasal mucus and skin excretions (including sweat)-by mass spectrometry (MS). The use of such specimens brings a number of advantages, the most important being non-invasive sampling, the limited risk of adulteration and the ability to obtain information that complements blood and urine tests. The most often studied matrices are hair, breath and saliva. This review primarily focuses on endogenous (e.g. potential biomarkers, hormones) and exogenous (e.g. drugs, environmental contaminants) small molecules. The majority of analytical methods used chromatographic separation prior to MS; however, such a hyphenated methodology greatly limits analytical throughput. On the other hand, the mass spectrometric methods that exclude chromatographic separation are fast but suffer from matrix interferences. To enable development of quantitative assays for unconventional matrices, it is desirable to standardize the protocols for the analysis of each specimen and create appropriate certified reference materials. Overcoming these challenges will make analysis of unconventional human biological matrices more common in a clinical setting. This article is part of the themed issue 'Quantitative mass spectrometry'.

  16. 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

  17. Identification and quantitation of free ceramides in human platelets.

    PubMed

    Krivit, W; Hammarström, S

    1972-07-01

    Free ceramides were isolated from human platelets. Their structures were unequivocally determined by gas-liquid chromatography-mass spectrometry of the trimethylsilyl ether derivatives. The major components were N-(palmitoyl) sphingosine, N-(stearoyl) sphingosine, N-(eicosanoyl) sphingosine, N-(docosanoyl) sphingosine, N-(tetracosanoyl) sphingosine, and N-(tetracosenoyl) sphingosine. Sphinganine-and sphingadienine-containing ceramides as well as ceramides containing other unsaturated acids were also present. The amount of ceramides was determined by quantitative gas-liquid chromatography, using radioactive ceramide as internal standard and synthetic crystalline ceramides for comparison of peak areas. The concentration of ceramides was found to be 1.31 micro g/10(9) platelets or 0.47 micro g/mg of platelet protein.

  18. 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.

  19. 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.

  20. Quantitative studies of human urinary excretion of uropontin.

    PubMed

    Min, W; Shiraga, H; Chalko, C; Goldfarb, S; Krishna, G G; Hoyer, J R

    1998-01-01

    Uropontin is the urinary form of osteopontin, an aspartic acid-rich phosphorylated glycoprotein. Uropontin has been previously shown to be a potent inhibitor of the nucleation, growth and aggregation of calcium oxalate crystals and the binding of these crystals to renal epithelial cells. Quantitative data defining the excretion of this protein are necessary to determine its role in urinary stone formation. In the present studies, we determined uropontin excretion rates of normal humans. Urine samples were obtained under conditions of known dietary intake from young adult human volunteers with no history, radiographic or laboratory evidence of renal disease. Urinary concentrations of uropontin were measured by a sensitive ELISA employing an affinity purified polyclonal antiserum to uropontin. Thirteen normal subjects ingested a constant diet providing 1 gram of calcium, 1 gram of phosphorus, 150 mEq of sodium and 1 gram of protein per kilogram of body wt per day during an eight day study period. The relationship of urinary volume to uropontin excretion was assessed by varying fluid intake on the last four days of the study to change the mean urine volume/24 hr by > 500 ml. Urine collected in six hour aliquots for eight days was analyzed for uropontin by ELISA, and for calcium, and creatinine. Daily uropontin excretion of 13 individual subjects was 3805 +/- 1805 micrograms/24 hr (mean +/- 1 SD). The mean urinary levels (1.9 micrograms/ml) detected in the present study are sufficient for inhibition of crystallization; our previous studies have demonstrated that the nucleation, growth and aggregation of calcium oxalate crystals and their binding to renal cells in vitro are inhibited by this concentration of purified uropontin. In contrast to the regular pattern of diurnal variation of calcium excretion seen in most subjects, uropontin excretion showed no regularity of diurnal variation and was not directly related to either calcium or creatinine excretion or changes in

  1. Quantitative Real-Time Gene Profiling of Human Alveolar Osteoblasts.

    PubMed

    Coates, Dawn E; Zafar, Sobia; Milne, Trudy J

    2017-01-01

    The use of quantitative real-time reverse transcriptase PCR (qRT(2)-PCR) for the identification of differentially regulated genes is a powerful technology. The protocol presented here uses qRT(2)-PCR gene arrays to investigate the regulation of 84 angiogenic related genes in human primary alveolar osteoblasts following treatment with the bisphosphonate, zoledronic acid (ZA), and geranylgeraniol (GGOH). GGOH has potential as a therapeutic agent for Bisphosphate-Related Osteonecrosis of the Jaw (BRONJ), a serious side-effect resulting from the treatment for metastatic cancer (Zafar et al., J Oral Pathol Med 43:711-721, 2014; Ruggiero, Ann NY Acad Sci 1218:38-46, 2011). The isolation of the primary osteoblast cells follows the methods previously described (Dillon et al., Methods Mol Biol 816:3-18, 2012) with a new RNA extraction technique described fully. The method highlights the importance of obtaining high-quality RNA which is DNA-free. Relative levels of gene expression are normalized against selected housekeeping genes (HKG) and a number of examples of how fold regulation (2(-∆∆Cq)) and gene expression level (2(-∆Cq)) data can be presented are given.

  2. Quantitation of Human Seroresponsiveness to Merkel Cell Polyomavirus

    PubMed Central

    Pastrana, Diana V.; Tolstov, Yanis L.; Becker, Jürgen C.; Moore, Patrick S.; Chang, Yuan; Buck, Christopher B.

    2009-01-01

    Merkel cell carcinoma (MCC) is a relatively uncommon but highly lethal form of skin cancer. A majority of MCC tumors carry DNA sequences derived from a newly identified virus called Merkel cell polyomavirus (MCV or MCPyV), a candidate etiologic agent underlying the development of MCC. To further investigate the role of MCV infection in the development of MCC, we developed a reporter vector-based neutralization assay to quantitate MCV-specific serum antibody responses in human subjects. Our results showed that 21 MCC patients whose tumors harbored MCV DNA all displayed vigorous MCV-specific antibody responses. Although 88% (42/48) of adult subjects without MCC were MCV seropositive, the geometric mean titer of the control group was 59-fold lower than the MCC patient group (p<0.0001). Only 4% (2/48) of control subjects displayed neutralizing titers greater than the mean titer of the MCV-positive MCC patient population. MCC tumors were found not to express detectable amounts of MCV VP1 capsid protein, suggesting that the strong humoral responses observed in MCC patients were primed by an unusually immunogenic MCV infection, and not by viral antigen expressed by the MCC tumor itself. The occurrence of highly immunogenic MCV infection in MCC patients is unlikely to reflect a failure to control polyomavirus infections in general, as seroreactivity to BK polyomavirus was similar among MCC patients and control subjects. The results support the concept that MCV infection is a causative factor in the development of most cases of MCC. Although MCC tumorigenesis can evidently proceed in the face of effective MCV-specific antibody responses, a small pilot animal immunization study revealed that a candidate vaccine based on MCV virus-like particles (VLPs) elicits antibody responses that robustly neutralize MCV reporter vectors in vitro. This suggests that a VLP-based vaccine could be effective for preventing the initial establishment of MCV infection. PMID:19750217

  3. 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.

  4. 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.

  5. Anthropometry of fetal vasculature in the chorionic plate.

    PubMed

    Gordon, Z; Elad, D; Almog, R; Hazan, Y; Jaffa, A J; Eytan, O

    2007-12-01

    Normal fetal development is dependent on adequate placental blood perfusion. The functional role of the placenta takes place mainly in the capillary system; however, ultrasound imaging of fetal blood flow is commonly performed on the umbilical artery, or on its first branches over the chorionic plate. The objective of this study was to evaluate the structural organization of the feto-placental vasculature of the chorionic plate. Casting of the placental vasculature was performed on 15 full-term placentas using a dental polymer mixed with colored ink. Observations of the cast models revealed that the branching architecture of the chorionic vessel is a combination of dichotomous and monopodial patterns, where the first two to three generations are always of a dichotomous nature. Analysis of the daughter-to-mother diameter ratios in the chorionic vessels provided a maximum in the range of 0.6-0.8 for the dichotomous branches, whereas in monopodial branches it was in the range of 0.1-0.3. Similar to previous studies, this study reveals that the vasculature architecture is mostly monopodial for the marginal cord insertion and mostly dichotomous for the central insertion. The more marginal the umbilical cord insertion is on the chorionic plate, the more monopodial branching patterns are created to compensate the dichotomous pattern deficiency to perfuse peripheral placental territories.

  6. Chorionic plate expression patterns of the maspin tumor suppressor protein in preeclamptic and egg donor placentas.

    PubMed

    Taglauer, E S; Gundogan, F; Johnson, K L; Scherjon, S A; Bianchi, D W

    2013-04-01

    Maspin is a serine protease inhibitor involved in regulating human placental trophoblast cell migration. Maspin has not been studied in preeclampsia (PE) or relative to the maternal-fetal immunological relationship, both of which may involve altered trophoblast migration. We examined maspin expression in placentas from in vitro fertilization (IVF) and egg donor (ED) pregnancies with and without PE. Exclusive to the chorionic plate, the number of maspin-positive extravillous trophoblasts was significantly decreased in IVF-PE vs. IVF (p = 0.005) and ED vs. IVF (p = 0.013). These data suggest maspin expression may be influenced by PE and/or the immunological dynamics of pregnancy.

  7. Patterns of inner chorion structure in Anastrepha (Diptera: Tephritidae) eggs.

    PubMed

    Figueiredo, Julia V A; Perondini, André L P; Selivon, Denise

    2017-03-01

    The inner chorion structure of Anastrepha eggs from 16 species of various infrageneric taxonomic groups is described by scanning and transmission electron microscopy. The layers of the chorion, the outer egg membrane, are structurally similar. Furthermore, an additional trabecular layer (ATL) that exists in some species, together with other characteristics, facilitates the recognition of four patterns of chorion structuring: Pattern I, in which the ATL layer is absent, is found in Anastrepha amita, the Anastrepha fraterculus complex, Anastrepha obliqua, Anastrepha sororcula, Anastrepha suspensa and Anastrepha zenildae (fraterculus group), and Anastrepha bistrigata and Anastrepha striata (striata group); Pattern II in Anastrepha serpentina (serpentina group), Anastrepha grandis (grandis group) and Anastrepha pseudoparallela (pseudoparallela group), in which the ATL presents large open spaces with pillars; Pattern III, found in Anastrepha consobrina (pseudoparallela group), in which the ATL is composed of round cavities; and Pattern IV, found in Anastrepha alveata and Anastrepha pickeli (spatulata group), where the large ATL cavities are reticulated. Comparatively, the chorion structure in Anastrepha eggs is more complex than in eggs of other fruit flies, e.g., Bactrocera, Rhagoletis and Ceratitis.

  8. Regional quantitative histological variations in human oral mucosa.

    PubMed

    Ciano, Joseph; Beatty, Brian Lee

    2015-03-01

    Oral mucosa demonstrates regional variations that reflect contact with food during mastication. Though known qualitatively, our aim was to quantitatively assess regions to establish a measurable baseline from which one could compare in pathological and comparative studies, in which the abrasiveness of diets may differ. We assessed variations in the epithelial-connective tissue junction (rete ridges counts), collagen organization within the lamina propria, and elastin composition of the lamina propria of 15 regions of the labial (buccal) gingiva, lingual gingiva, vestibule, and palate. All characteristics varied more between regions within the same individual than between individuals. Lingual gingiva had high rete ridges counts, high level of collagen organization, and moderate elastin composition compared to other regions. The labial gingiva had few rete ridges, high collagen organization, and low elastin. The vestibule had the fewest average of rete ridges, least organized collagen, and high elastin. The hard palate had the highest average of rete ridges, high collagen organization, and the lowest elastin content. The soft palate conversely had the smallest average of rete ridges, moderate collagen organization, and the highest elastin composition. Our results indicate that comparison of these quantitative histological differences is warranted only for collagen organization and elastin composition. Differences in rete ridges counts were not statistically significant. Most histological characteristics observed were not significantly different between dentulous and edentulous cadavers, and the group containing all individuals. An exception was the level of collagen fiber organization within the lamina propria, which was higher in most regions when teeth were present.

  9. 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.

  10. Spectroscopic quantitation of cytochrome P-450 in human lung microsomes.

    PubMed

    Wheeler, C W; Guenthner, T M

    1990-01-01

    The cytochrome P-450 content of human lung microsomes was measured by difference spectroscopy of the carbon monoxide-complexed hemoprotein. These measurements were only possible after the microsome preparation had been subjected to centrifugation over a discontinuous sucrose gradient, to remove an opaque black contaminant. The specific concentration of total cytochrome P-450 in human lung microsomes is essentially identical to that of microsomes prepared under identical conditions from untreated baboon lungs, but is only 0.7% of the specific content found in lung microsomes from untreated rabbits. These measurements correspond well to the observed metabolic capacities of the various microsome samples.

  11. 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.

  12. 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.

  13. Parametric mapping and quantitative analysis of the human calvarium.

    PubMed

    Voie, Arne; Dirnbacher, Maximilian; Fisher, David; Hölscher, Thilo

    2014-12-01

    In this paper we report how thickness and density vary over the calvarium region of a collection of human skulls. Most previous reports involved a limited number of skulls, with a limited number of measurement sites per skull, so data in the literature are sparse. We collected computer tomography (CT) scans of 51 ex vivo human calvaria, and analyzed these in silico using over 2000 measurement sites per skull. Thickness and density were calculated at these sites, for the three skull layers separately and combined, and were mapped parametrically onto the skull surfaces to examine the spatial variations per skull. These were found to be highly variable, and unique descriptors of the individual skulls. Of the three skull layers, the thickness of the inner cortical layer was found to be the most variable, while the least variable was the outer cortical density.

  14. DETECTION AND QUANTITATION OF FALLOUT PARTICLES IN A HUMAN LUNG.

    PubMed

    WEGST, A V; PELLETIER, C A; WHIPPLE, G H

    1964-02-28

    Portions of an adult human lung were studied by autoradiography in order to detect the presence of fallout particles. The radioactivity in the remainder of the tissue was determined with a gamma-ray spectrometer. Four particles were found and their activities were determined. From the measurement for total-fission-product activity in the lung tissue it was calculated that there were approximately 264 particles in the right lung at the time of death.

  15. Relationship between electrical admittivity and quantitative histopathology in human prostate

    NASA Astrophysics Data System (ADS)

    Halter, Ryan; Milone, Michael; Schned, Alan; Heaney, John

    2010-04-01

    Passive bioelectrical properties have been demonstrated to provide sufficient contrast for use in differentiating benign from malignant tissue in a number of different organs including breast, prostate, cervix, bladder, and skin. The underlying microscopic anatomy responsible for these measured differences has been primarily speculative in the past. In this study we recorded electrical conductivity and permittivity spectra (100 Hz - 100 kHz) from 464 three mm diameter circular prostate samples. Each of these tissue specimens were stained with hematoxylin and eosin, processed onto microscopy slides, and digitized using optical microscopy. We used digital imaging processing tools to extract quantitative morphological features including total number of glands, average and total glandular lumen size, shape characteristics of the luminal spaces, and average and total glandular perimeter lengths. Correlative analysis was performed to assess the relationships between the tissue architectural features and the precisely co-registered electrical properties. We report on the findings from this analysis. This statistical assessment aims to provide a valuable piece of new information to help formulate a better understanding of the precise influence morphological architecture has on the flow of current through tissue.

  16. High throughput, quantitative analysis of human osteoclast differentiation and activity.

    PubMed

    Diepenhorst, Natalie A; Nowell, Cameron J; Rueda, Patricia; Henriksen, Kim; Pierce, Tracie; Cook, Anna E; Pastoureau, Philippe; Sabatini, Massimo; Charman, William N; Christopoulos, Arthur; Summers, Roger J; Sexton, Patrick M; Langmead, Christopher J

    2017-02-15

    Osteoclasts are multinuclear cells that degrade bone under both physiological and pathophysiological conditions. Osteoclasts are therefore a major target of osteoporosis therapeutics aimed at preserving bone. Consequently, analytical methods for osteoclast activity are useful for the development of novel biomarkers and/or pharmacological agents for the treatment of osteoporosis. The nucleation state of an osteoclast is indicative of its maturation and activity. To date, activity is routinely measured at the population level with only approximate consideration of the nucleation state (an 'osteoclast population' is typically defined as cells with ≥3 nuclei). Using a fluorescent substrate for tartrate-resistant acid phosphatase (TRAP), a routinely used marker of osteoclast activity, we developed a multi-labelled imaging method for quantitative measurement of osteoclast TRAP activity at the single cell level. Automated image analysis enables interrogation of large osteoclast populations in a high throughput manner using open source software. Using this methodology, we investigated the effects of receptor activator of nuclear factor kappa-B ligand (RANK-L) on osteoclast maturation and activity and demonstrated that TRAP activity directly correlates with osteoclast maturity (i.e. nuclei number). This method can be applied to high throughput screening of osteoclast-targeting compounds to determine changes in maturation and activity.

  17. Quantitative reflection spectroscopy at the human ocular fundus

    NASA Astrophysics Data System (ADS)

    Hammer, Martin; Schweitzer, Dietrich

    2002-01-01

    A new model of the reflection of the human ocular fundus on the basis of the adding-doubling method, an approximate solution of the radiative transport equation, is described. This model enables the calculation of the concentrations of xanthophyll in the retina, of melanin in the retinal pigment epithelium and the choroid, and of haemoglobin in the choroid from fundus reflection spectra. The concentration values found in 12 healthy subjects are in excellent agreement with published data. In individual cases of pathologic fundus alterations, possible benefits to the ophthalmologic diagnostics are demonstrated.

  18. 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.

  19. Quantitative evaluation of the major determinants of human gait.

    PubMed

    Lin, Yi-Chung; Gfoehler, Margit; Pandy, Marcus G

    2014-04-11

    Accurate knowledge of the isolated contributions of joint movements to the three-dimensional displacement of the center of mass (COM) is fundamental for understanding the kinematics of normal walking and for improving the treatment of gait disabilities. Saunders et al. (1953) identified six kinematic mechanisms to explain the efficient progression of the whole-body COM in the sagittal, transverse, and coronal planes. These mechanisms, referred to as the major determinants of gait, were pelvic rotation, pelvic list, stance knee flexion, foot and knee mechanisms, and hip adduction. The aim of the present study was to quantitatively assess the contribution of each major gait determinant to the anteroposterior, vertical, and mediolateral displacements of the COM over one gait cycle. The contribution of each gait determinant was found by applying the concept of an 'influence coefficient', wherein the partial derivative of the COM displacement with respect to a prescribed determinant was calculated. The analysis was based on three-dimensional measurements of joint angular displacements obtained from 23 healthy young adults walking at slow, normal and fast speeds. We found that hip flexion, stance knee flexion, and ankle-foot interaction (comprised of ankle plantarflexion, toe flexion and the displacement of the center of pressure) are the major determinants of the displacements of the COM in the sagittal plane, while hip adduction and pelvic list contribute most significantly to the mediolateral displacement of the COM in the coronal plane. Pelvic rotation and pelvic list contribute little to the vertical displacement of the COM at all walking speeds. Pelvic tilt, hip rotation, subtalar inversion, and back extension, abduction and rotation make negligible contributions to the displacements of the COM in all three anatomical planes.

  20. Quantitative analysis of lymphangiogenic markers in human colorectal cancer.

    PubMed

    Parr, C; Jiang, W G

    2003-08-01

    Lymphatic spread of colorectal cancer cells to regional lymph nodes is one of the early events in metastatic cancer, and is often associated with distant metastatic spread and a poor prognosis. This study examined lymphangiogenic factors, and in particular a panel of newly discovered lymphangiogenic markers, in colorectal cancer tissues from a cohort of patients. Paired samples (background normal mucosa and cancer) of colon tissue were obtained from patients with colorectal cancer. The expression and levels of the VEGF-C and VEGF-D cytokines, the VEGF receptors VEGFR-2 and VEGFR-3, and newly described lymphatic endothelial markers, LYVE-1, Prox-1, podoplanin and 5'-nucleotidase were assessed. RNA was extracted from the frozen colon tissues. The level of expression for each factor/marker was determined using RT-PCR and quantified using a real-time quantitative PCR (RT-QPCR) technique, with respective cloned cDNA plasmids as internal standards. VEGF-D was expressed to a significantly higher degree in the colon tumour tissues. There was no significant difference between the expression levels for both VEGF-C and its receptor, VEGFR-2, in background and cancer tissues. However, levels of the VEGFR-3 receptor were found to be significantly higher in colon cancer than the normal background tissues. LYVE-1 levels were below detection in most cases. There was a significant increase in the degree of Prox-1 and 5'-nucleotidase expression in colon cancer tissue. Podoplanin expression was also increased in the cancer samples. These markers indicate an increase in lymphangiogenesis in colon cancer, and may therefore have prognostic value for colon cancer patients.

  1. Quantitation of human immunodeficiency virus type 1 infection kinetics.

    PubMed Central

    Dimitrov, D S; Willey, R L; Sato, H; Chang, L J; Blumenthal, R; Martin, M A

    1993-01-01

    Tissue culture infections of CD4-positive human T cells by human immunodeficiency virus type 1 (HIV-1) proceed in three stages: (i) a period following the initiation of an infection during which no detectable virus is produced; (ii) a phase in which a sharp increase followed by a peak of released progeny virions can be measured; and (iii) a final period when virus production declines. In this study, we have derived equations describing the kinetics of HIV-1 accumulation in cell culture supernatants during multiple rounds of infection. Our analyses indicated that the critical parameter affecting the kinetics of HIV-1 infection is the infection rate constant k = Inn/ti, where n is the number of infectious virions produced by one cell (about 10(2)) and ti is the time required for one complete cycle of virus infection (typically 3 to 4 days). Of particular note was our finding that the infectivity of HIV-1 during cell-to-cell transmission is 10(2) to 10(3) times greater than the infectivity of cell-free virus stocks, the inocula commonly used to initiate tissue culture infections. We also demonstrated that the slow infection kinetics of an HIV-1 tat mutant is not due to a longer replication time but reflects the small number of infectious particles produced per cycle. PMID:8445728

  2. Chorionic villus sampling in continuing pregnancies. II. Cytogenetic reliability.

    PubMed

    Martin, A O; Simpson, J L; Rosinsky, B J; Elias, S

    1986-06-01

    Cytogenetic analysis was performed on 103 chorionic villus samples. Analysis of the 103 samples revealed six abnormalities. In three of the six the abnormalities were confirmed in fetal or neonatal tissue (47,XY, + 13; 46,XY, t(13q13q); 45,X). In three samples the abnormalities detected were not confirmed; in two of the three the abnormalities were detected only in long-term cultures, whereas in the other samples the abnormality was restricted to direct analysis of the villi after overnight incubation. Our initial experience leads us to conclude that certain abnormalities in chorionic villus sampling may not be indicative of fetal abnormalities; 45,X/46,XX or 45,X/46,XY mosaicism is such a complement. Discrepancies between cytogenetic analysis of intact villi processed soon after sampling and of cells grown in culture can be managed by adhering to several suggested guidelines and by liberal use of confirmatory amniocentesis.

  3. Chorionic plate vessels as an origin of amniotic fluid neutrophils.

    PubMed

    Lee, Soong Deok; Kim, Mi Ran; Hwang, Pil Gyu; Shim, Soon-Sup; Yoon, Bo Hyun; Kim, Chong Jai

    2004-07-01

    The present study was conducted to investigate the potential anatomical source of amniotic fluid neutrophils. Microdissection of neutrophils from the chorioamnion of the fetal membranes and the amnion of the chorionic plates of 10 preterm placentas with acute chorioamnionitis was performed and the genotypes of the neutrophils were compared with those of the mother and fetus using polymerase chain reaction of nine autosomal STR loci. In separate analyses, we reviewed eight cases of fetal autopsies with increased amniotic fluid neutrophils for the presence of neutrophils in the alveoli, and also analyzed the relationship between the amniotic fluid white blood cell (WBC) count and the histological pattern of placental inflammation. The genotypes of all of the neutrophils found in the chorioamnion of the fetal membrane matched those of the mother (n = 10). The genotypes of neutrophils found in the chorionic plate were of mixed maternal and fetal origin (n = 4). In the autopsy series of the fetuses with amniotic fluid WBC (n = 8), only five cases showed neutrophils in the alveolar space, while all the placentas had chorioamnionitis. There was no significant difference in amniotic fluid WBC count between the cases with or without acute membranitis, while among the cases with placental inflammation, those with inflammation of the chorionic plate had a significantly higher amniotic fluid WBC count than both the membranitis-only cases (P < 0.001) and the membranitis and funisitis cases (P < 0.05). These results imply that fetal vasculature at the chorionic plate is the main source of amniotic fluid neutrophils, especially in the cases without funisitis.

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

    PubMed

    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.

  5. Human Spermatozoa Quantitative Proteomic Signature Classifies Normo- and Asthenozoospermia.

    PubMed

    Saraswat, Mayank; Joenväärä, Sakari; Jain, Tushar; Tomar, Anil Kumar; Sinha, Ashima; Singh, Sarman; Yadav, Savita; Renkonen, Risto

    2017-01-01

    Scarcely understood defects lead to asthenozoospermia, which results in poor fertility outcomes. Incomplete knowledge of these defects hinders the development of new therapies and reliance on interventional therapies, such as in vitro fertilization, increases. Sperm cells, being transcriptionally and translationally silent, necessitate the proteomic approach to study the sperm function. We have performed a differential proteomics analysis of human sperm and seminal plasma and identified and quantified 667 proteins in sperm and 429 proteins in seminal plasma data set, which were used for further analysis. Statistical and mathematical analysis combined with pathway analysis and self-organizing maps clustering and correlation was performed on the data set.It was found that sperm proteomic signature combined with statistical analysis as opposed to the seminal plasma proteomic signature can differentiate the normozoospermic versus the asthenozoospermic sperm samples. This is despite the results that some of the seminal plasma proteins have big fold changes among classes but they fall short of statistical significance. S-Plot of the sperm proteomic data set generated some high confidence targets, which might be implicated in sperm motility pathways. These proteins also had the area under the curve value of 0.9 or 1 in ROC curve analysis.Various pathways were either enriched in these proteomic data sets by pathway analysis or they were searched by their constituent proteins. Some of these pathways were axoneme activation and focal adhesion assembly, glycolysis, gluconeogenesis, cellular response to stress and nucleosome assembly among others. The mass spectrometric data is available via ProteomeXchange with identifier PXD004098.

  6. Sequencing human ribs into anatomical order by quantitative multivariate methods.

    PubMed

    Cirillo, John; Henneberg, Maciej

    2012-06-01

    Little research has focussed on methods to anatomically sequence ribs. Correct anatomical sequencing of ribs assists in determining the location and distribution of regional trauma, age estimation, number of puncture wounds, number of individuals, and personal identification. The aim of the current study is to develop a method for placing fragmented and incomplete rib sets into correct anatomical position. Ribs 2-10 were used from eleven cadavers of an Australian population. Seven variables were measured from anatomical locations on the rib. General descriptive statistics were calculated for each variable along with an analysis of variance (ANOVA) and ANOVA with Bonferroni statistics. Considerable overlap was observed between ribs for univariate methods. Bivariate and multivariate methods were then applied. Results of the ANOVA with post hoc Bonferroni statistics show that ratios of various dimensions of a single rib could be used to sequence it within adjacent ribs. Using multiple regression formulae, the most accurate estimation of the anatomical rib number occurs when the entire rib is found in isolation. This however, is not always possible. Even when only the head and neck of the rib are preserved, a modified multivariate regression formula assigned 91.95% of ribs into correct anatomical position or as an adjacent rib. Using multivariate methods it is possible to sequence a single human rib with a high level of accuracy and they are superior to univariate methods. Left and right ribs were found to be highly symmetrical. Some rib dimensions were greater in males than in females, but overall the level of sexual dimorphism was low.

  7. 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.

  8. Novel "omics" approach for study of low-abundance, low-molecular-weight components of a complex biological tissue: regional differences between chorionic and basal plates of the human placenta.

    PubMed

    Kedia, Komal; Nichols, Caitlin A; Thulin, Craig D; Graves, Steven W

    2015-11-01

    Tissue proteomics has relied heavily on two-dimensional gel electrophoresis, for protein separation and quantification, then single protein isolation, trypsin digestion, and mass spectrometric protein identification. Such methods are predominantly used for study of high-abundance, full-length proteins. Tissue peptidomics has recently been developed but is still used to study the most highly abundant species, often resulting in observation and identification of dozens of peptides only. Tissue lipidomics is likewise new, and reported studies are limited. We have developed an "omics" approach that enables over 7,000 low-molecular-weight, low-abundance species to be surveyed and have applied this to human placental tissue. Because the placenta is believed to be involved in complications of pregnancy, its proteomic evaluation is of substantial interest. In previous research on the placental proteome, abundant, high-molecular-weight proteins have been studied. Application of large-scale, global proteomics or peptidomics to the placenta have been limited, and would be challenging owing to the anatomic complexity and broad concentration range of proteins in this tissue. In our approach, involving protein depletion, capillary liquid chromatography, and tandem mass spectrometry, we attempted to identify molecular differences between two regions of the same placenta with only slightly different cellular composition. Our analysis revealed 16 species with statistically significant differences between the two regions. Tandem mass spectrometry enabled successful sequencing, or otherwise enabled chemical characterization, of twelve of these. The successful discovery and identification of regional differences between the expression of low-abundance, low-molecular weight biomolecules reveals the potential of our approach.

  9. Optimization of human dose prediction by using quantitative and translational pharmacology in drug discovery.

    PubMed

    Bueters, Tjerk; Gibson, Christopher; Visser, Sandra A G

    2015-01-01

    In this perspective article, we explain how quantitative and translational pharmacology, when well-implemented, is believed to lead to improved clinical candidates and drug targets that are differentiated from current treatment options. Quantitative and translational pharmacology aims to build and continuously improve the quantitative relationship between drug exposure, target engagement, efficacy, safety and its interspecies relationship at every phase of drug discovery. Drug hunters should consider and apply these concepts to develop compounds with a higher probability of interrogating the clinical biological hypothesis. We offer different approaches to set an initial effective concentration or pharmacokinetic-pharmacodynamic target in man and to predict human pharmacokinetics that determine together the predicted human dose and dose schedule. All concepts are illustrated with ample literature examples.

  10. 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...

  11. Quantitation of 25-hydroxycholecalciferol in human serum by high-pressure liquid chromatography.

    PubMed

    Schaefer, P C; Goldsmith, R S

    1978-01-01

    An HPLC method for assaying 25(OH)D3 in extracts of human serum is described. The method involves extraction of serum with methanol-dichloromethane and chromatography on a column of Sephadex LH-20 prior to HPLC with quantitation by UV absorbance. Comparison with a CBP assay for 25(OH)D3 showed no difference between the two methods.

  12. Production and certification of NIST Standard Reference Material 2372 Human DNA Quantitation Standard.

    PubMed

    Kline, Margaret C; Duewer, David L; Travis, John C; Smith, Melody V; Redman, Janette W; Vallone, Peter M; Decker, Amy E; Butler, John M

    2009-06-01

    Modern highly multiplexed short tandem repeat (STR) assays used by the forensic human-identity community require tight control of the initial amount of sample DNA amplified in the polymerase chain reaction (PCR) process. This, in turn, requires the ability to reproducibly measure the concentration of human DNA, [DNA], in a sample extract. Quantitative PCR (qPCR) techniques can determine the number of intact stretches of DNA of specified nucleotide sequence in an extremely small sample; however, these assays must be calibrated with DNA extracts of well-characterized and stable composition. By 2004, studies coordinated by or reported to the National Institute of Standards and Technology (NIST) indicated that a well-characterized, stable human DNA quantitation certified reference material (CRM) could help the forensic community reduce within- and among-laboratory quantitation variability. To ensure that the stability of such a quantitation standard can be monitored and that, if and when required, equivalent replacement materials can be prepared, a measurement of some stable quantity directly related to [DNA] is required. Using a long-established conventional relationship linking optical density (properly designated as decadic attenuance) at 260 nm with [DNA] in aqueous solution, NIST Standard Reference Material (SRM) 2372 Human DNA Quantitation Standard was issued in October 2007. This SRM consists of three quite different DNA extracts: a single-source male, a multiple-source female, and a mixture of male and female sources. All three SRM components have very similar optical densities, and thus very similar conventional [DNA]. The materials perform very similarly in several widely used gender-neutral assays, demonstrating that the combination of appropriate preparation methods and metrologically sound spectrophotometric measurements enables the preparation and certification of quantitation [DNA] standards that are both maintainable and of practical utility.

  13. Quantitative comparison of cerebral artery development in human embryos with other eutherians.

    PubMed

    Ashwell, Ken W S; Shulruf, Boaz

    2015-09-01

    The embryonic and early fetal human brain is known to undergo extraordinary expansion of its cellular population during embryonic and early fetal life, and is critically dependant on a steady supply of nutrients and oxygen for proper brain development. Quantitative analysis of the internal radius of the aorta and cerebral arteries in a range of eutherian mammals has been used to compare arterial flow to the developing human brain with that to the brains of non-human eutherians. Human embryos showed a much steeper rise of internal radius of the aorta with increasing body size than the embryos of non-human eutherians, but the thickness of the aorta rose at the same pace relative to body size in both humans and non-humans, suggesting that aortic pressure is similar in all eutherian embryos of a similar size. The sums of internal radii of both the internal carotids and vertebral arteries of human embryos raised to the fourth power were much lower at embryonic stages (less than 22 mm body length) than in non-human eutherians, were similar between humans and non-humans at 22-30 mm body length, and exceeded the non-humans at body lengths of more than 30 mm. The relative size of the internal calibre of the cerebral feeder arteries (internal carotid and vertebral) to the aorta did not change between embryonic and fetal sizes in either humans or non-humans. The findings suggest that the developing human brain may actually receive less blood flow at embryonic sizes (less than 22 mm body length) than do other mammalian embryos of a similar body size, but that internal carotid and vertebral flow is higher in human fetuses (body length greater than 30 mm) than in developing non-humans of the same body size. Increased flow to the developing human brain relative to non-humans is achieved by simultaneous increases in both aortic and cerebral feeder artery internal calibre.

  14. Different effects of chorionic gonadotropin on Taenia crassiceps and Taenia solium cysticerci cultured in vitro.

    PubMed

    Díaz-Orea, M A; de Aluja, A S; Erosa, M de L; Gomez-Conde, E; Castellanos Sánchez, V O; Willms, K; Sciutto, E; Fragoso, G

    2007-12-01

    Hormones play a significant role in murine Taenia crassiceps cysticercosis, and they may also participate in the susceptibility to Taenia solium cysticercosis. In the present study, in vitro effects are reported for human chorionic gonadotropin (hCG) on the larval stages of T. crassiceps (WFU strain) and T. solium. hCG effectively promotes parasite reproduction, i.e., it increases the number of buds on T. crassiceps cysticerci and the percentage of evagination and parasite length in T. solium. This is the first report in which a direct effect of hCG is reported for a parasite. hCG or mouse luteinizing hormone could be recognized by the cysticerci as mitogenic factors and contribute to the female and pregnancy bias toward susceptibility to T. crassiceps and T. solium cysticercosis, respectively.

  15. Expanding the Limits of Human Blood Metabolite Quantitation Using NMR Spectroscopy

    PubMed Central

    2015-01-01

    A current challenge in metabolomics is the reliable quantitation of many metabolites. Limited resolution and sensitivity combined with the challenges associated with unknown metabolite identification have restricted both the number and the quantitative accuracy of blood metabolites. Focused on alleviating this bottleneck in NMR-based metabolomics, investigations of pooled human serum combining an array of 1D/2D NMR experiments at 800 MHz, database searches, and spiking with authentic compounds enabled the identification of 67 blood metabolites. Many of these (∼1/3) are new compared with those reported previously as a part of the Human Serum Metabolome Database. In addition, considering both the high reproducibility and quantitative nature of NMR as well as the sensitivity of NMR chemical shifts to altered sample conditions, experimental protocols and comprehensive peak annotations are provided here as a guide for identification and quantitation of the new pool of blood metabolites for routine applications. Further, investigations focused on the evaluation of quantitation using organic solvents revealed a surprisingly poor performance for protein precipitation using acetonitrile. One-third of the detected metabolites were attenuated by 10–67% compared with methanol precipitation at the same solvent-to-serum ratio of 2:1 (v/v). Nearly 2/3 of the metabolites were further attenuated by up to 65% upon increasing the acetonitrile-to-serum ratio to 4:1 (v/v). These results, combined with the newly established identity for many unknown metabolites in the NMR spectrum, offer new avenues for human serum/plasma-based metabolomics. Further, the ability to quantitatively evaluate nearly 70 blood metabolites that represent numerous classes, including amino acids, organic acids, carbohydrates, and heterocyclic compounds, using a simple and highly reproducible analytical method such as NMR may potentially guide the evaluation of samples for analysis using mass spectrometry

  16. Qualitative and quantitative comparability of human and animal developmental neurotoxicants: a workshop summary.

    PubMed

    Rees, D C; Francis, E Z; Kimmel, C A

    1990-01-01

    A Workshop on the Qualitative and Quantitative Comparability of Human and Animal Developmental Neurotoxicity was held in Williamsburg, Va. on April 11-13, 1989. Based upon data presented at the Workshop, the degree of qualitative and quantitative comparability between data obtained from humans and experimental animals is reviewed for several developmental neurotoxicants (lead, agents of abuse, alcohol, PCBs, phenytoin, methylmercury, and ionizing radiation). Qualitative comparability was considered for the following functional categories: motor development and function, cognitive function, sensory function, motivation/arousal behavior, and social behavior. Quantitative comparability was assessed by comparing administered dose as well as measures of internal dose (e.g., blood levels) for selected agents. Comparability of qualitative changes between humans and rodents was most apparent when comparisons were made on the basis of general categories of behavioral function. These data support the use of animal models in assessing risk for developmental neurotoxicants and provide guidance on the types of functional end points that can be incorporated into a developmental neurotoxicity testing battery. Evidence of quantitative comparability was most apparent when an internal measure of dose (e.g., blood level) was used.

  17. Double trouble: the importance of reporting chorionicity and amnionicity in twin pregnancy ultrasound reports.

    PubMed

    Constantine, Sarah; Wilkinson, Chris

    2015-02-01

    An obstetric ultrasound report in a twin pregnancy that does not unambiguously determine chorionicity and amnionicity in the first trimester is substandard. This article will assist radiologists to understand the importance of reporting the chorionicity and amnionicity in all twin obstetric scans.

  18. Quantitative analysis of the expression of ACAT genes in human tissues by real-time PCR.

    PubMed

    Smith, Jeffery L; Rangaraj, Kavitha; Simpson, Robert; Maclean, Donald J; Nathanson, Les K; Stuart, Katherine A; Scott, Shaun P; Ramm, Grant A; de Jersey, John

    2004-04-01

    ACAT (also called sterol o-acyltransferase) catalyzes the esterification of cholesterol by reaction with long-chain acyl-CoA derivatives and plays a pivotal role in the regulation of cholesterol homeostasis. Although two human ACAT genes termed ACAT-1 and ACAT-2 have been reported, prior research on differential tissue expression is qualitative and incomplete. We have developed a quantitative multiplex assay for each ACAT isoform after RT treatment of total RNA using TaqMan real-time quantitative PCR normalized to beta-actin in the same reaction tube. This enabled us to calculate the relative abundance of transcripts in several human tissues as an ACAT-2/ACAT-1 ratio. In liver (n = 17), ACAT-1 transcripts were on average 9-fold (range, 1.7- to 167-fold) more abundant than ACAT-2, whereas in duodenal samples (n = 10), ACAT-2 transcripts were on average 3-fold (range, 0.39- to 12.2-fold) more abundant than ACAT-1. ACAT-2 was detected for the first time in peripheral blood mononuclear cells. Interesting differences in ACAT-2 mRNA expression were evident in subgroup analysis of samples from different sources. These results demonstrate quantitatively that ACAT-1 transcripts predominate in human liver and ACAT-2 transcripts predominate in human duodenum and support the notion that ACAT-2 has an important regulatory role in liver and intestine.

  19. A Network Neuroscience of Human Learning: Potential to Inform Quantitative Theories of Brain and Behavior.

    PubMed

    Bassett, Danielle S; Mattar, Marcelo G

    2017-04-01

    Humans adapt their behavior to their external environment in a process often facilitated by learning. Efforts to describe learning empirically can be complemented by quantitative theories that map changes in neurophysiology to changes in behavior. In this review we highlight recent advances in network science that offer a sets of tools and a general perspective that may be particularly useful in understanding types of learning that are supported by distributed neural circuits. We describe recent applications of these tools to neuroimaging data that provide unique insights into adaptive neural processes, the attainment of knowledge, and the acquisition of new skills, forming a network neuroscience of human learning. While promising, the tools have yet to be linked to the well-formulated models of behavior that are commonly utilized in cognitive psychology. We argue that continued progress will require the explicit marriage of network approaches to neuroimaging data and quantitative models of behavior.

  20. QUANTITATIVE VS. CONVENTIONAL PCR FOR DETECTION OF HUMAN ADENOVIRUSES IN WATER AND SEDIMENT SAMPLES

    PubMed Central

    STAGGEMEIER, Rodrigo; BORTOLUZZI, Marina; HECK, Tatiana Moraes da Silva; SPILKI, Fernando Rosado; ALMEIDA, Sabrina Esteves de Matos

    2015-01-01

    SUMMARY Human Adenoviruses (HAdV) are notably resistant in the environment. These agents may serve as effective indicators of fecal contamination, and may act as causative agents of a number of different diseases in human beings. Conventional polymerase chain reaction (PCR) and, more recently, quantitative PCR (qPCR) are widely used for detection of viral agents in environmental matrices. In the present study PCR and SYBR(r)Green qPCR assays were compared for detection of HAdV in water (55) and sediments (20) samples of spring and artesian wells, ponds and streams, collected from dairy farms. By the quantitative methodology HAdV were detected in 87.3% of the water samples and 80% of the sediments, while by the conventional PCR 47.3% and 35% were detected in water samples and sediments, respectively. PMID:26422153

  1. QUANTITATIVE VS. CONVENTIONAL PCR FOR DETECTION OF HUMAN ADENOVIRUSES IN WATER AND SEDIMENT SAMPLES.

    PubMed

    Staggemeier, Rodrigo; Bortoluzzi, Marina; Heck, Tatiana Moraes da Silva; Spilki, Fernando Rosado; Almeida, Sabrina Esteves de Matos

    2015-01-01

    Human Adenoviruses (HAdV) are notably resistant in the environment. These agents may serve as effective indicators of fecal contamination, and may act as causative agents of a number of different diseases in human beings. Conventional polymerase chain reaction (PCR) and, more recently, quantitative PCR (qPCR) are widely used for detection of viral agents in environmental matrices. In the present study PCR and SYBR(r)Green qPCR assays were compared for detection of HAdV in water (55) and sediments (20) samples of spring and artesian wells, ponds and streams, collected from dairy farms. By the quantitative methodology HAdV were detected in 87.3% of the water samples and 80% of the sediments, while by the conventional PCR 47.3% and 35% were detected in water samples and sediments, respectively.

  2. Comparative assessment of human and farm animal faecal microbiota using real-time quantitative PCR.

    PubMed

    Furet, Jean-Pierre; Firmesse, Olivier; Gourmelon, Michèle; Bridonneau, Chantal; Tap, Julien; Mondot, Stanislas; Doré, Joël; Corthier, Gérard

    2009-06-01

    Pollution of the environment by human and animal faecal pollution affects the safety of shellfish, drinking water and recreational beaches. To pinpoint the origin of contaminations, it is essential to define the differences between human microbiota and that of farm animals. A strategy based on real-time quantitative PCR (qPCR) assays was therefore developed and applied to compare the composition of intestinal microbiota of these two groups. Primers were designed to quantify the 16S rRNA gene from dominant and subdominant bacterial groups. TaqMan probes were defined for the qPCR technique used for dominant microbiota. Human faecal microbiota was compared with that of farm animals using faecal samples collected from rabbits, goats, horses, pigs, sheep and cows. Three dominant bacterial groups (Bacteroides/Prevotella, Clostridium coccoides and Bifidobacterium) of the human microbiota showed differential population levels in animal species. The Clostridium leptum group showed the lowest differences among human and farm animal species. Human subdominant bacterial groups were highly variable in animal species. Partial least squares regression indicated that the human microbiota could be distinguished from all farm animals studied. This culture-independent comparative assessment of the faecal microbiota between humans and farm animals will prove useful in identifying biomarkers of human and animal faecal contaminations that can be applied to microbial source tracking methods.

  3. Quantitation of human milk proteins and their glycoforms using multiple reaction monitoring (MRM).

    PubMed

    Huang, Jincui; Kailemia, Muchena J; Goonatilleke, Elisha; Parker, Evan A; Hong, Qiuting; Sabia, Rocchina; Smilowitz, Jennifer T; German, J Bruce; Lebrilla, Carlito B

    2017-01-01

    Human milk plays a substantial role in the child growth, development and determines their nutritional and health status. Despite the importance of the proteins and glycoproteins in human milk, very little quantitative information especially on their site-specific glycosylation is known. As more functions of milk proteins and other components continue to emerge, their fine-detailed quantitative information is becoming a key factor in milk research efforts. The present work utilizes a sensitive label-free MRM method to quantify seven milk proteins (α-lactalbumin, lactoferrin, secretory immunoglobulin A, immunoglobulin G, immunoglobulin M, α1-antitrypsin, and lysozyme) using their unique peptides while at the same time, quantifying their site-specific N-glycosylation relative to the protein abundance. The method is highly reproducible, has low limit of quantitation, and accounts for differences in glycosylation due to variations in protein amounts. The method described here expands our knowledge about human milk proteins and provides vital details that could be used in monitoring the health of the infant and even the mother. Graphical Abstract The glycopeptides EICs generated from QQQ.

  4. High Throughput Measurement of Extracellular DNA Release and Quantitative NET Formation in Human Neutrophils In Vitro.

    PubMed

    Sil, Payel; Yoo, Dae-Goon; Floyd, Madison; Gingerich, Aaron; Rada, Balazs

    2016-06-18

    Neutrophil granulocytes are the most abundant leukocytes in the human blood. Neutrophils are the first to arrive at the site of infection. Neutrophils developed several antimicrobial mechanisms including phagocytosis, degranulation and formation of neutrophil extracellular traps (NETs). NETs consist of a DNA scaffold decorated with histones and several granule markers including myeloperoxidase (MPO) and human neutrophil elastase (HNE). NET release is an active process involving characteristic morphological changes of neutrophils leading to expulsion of their DNA into the extracellular space. NETs are essential to fight microbes, but uncontrolled release of NETs has been associated with several disorders. To learn more about the clinical relevance and the mechanism of NET formation, there is a need to have reliable tools capable of NET quantitation. Here three methods are presented that can assess NET release from human neutrophils in vitro. The first one is a high throughput assay to measure extracellular DNA release from human neutrophils using a membrane impermeable DNA-binding dye. In addition, two other methods are described capable of quantitating NET formation by measuring levels of NET-specific MPO-DNA and HNE-DNA complexes. These microplate-based methods in combination provide great tools to efficiently study the mechanism and regulation of NET formation of human neutrophils.

  5. Human sensitivity to reinforcement: A comment on Kollins, Newland, and Critchfield's (1997) quantitative literature review

    PubMed Central

    Derenne, Adam; Baron, Alan

    1999-01-01

    In a quantitative review of human operant experiments, Kollins, Newland, and Critchfield (1997) found that humans are less sensitive to reinforcement contingencies than nonhumans are. Human performances were not as consistent with the matching law, and they were more variable from subject to subject. Some of the variables correlated with reduced human sensitivity were surprising. These included collection of the data under more controlled conditions (laboratory rather than naturalistic settings), and inclusions of discriminative stimuli correlated with alternative sources of reinforcement. We discuss these unexpected findings in the light of criticisms that have been leveled against meta-analytic literature reviews (e.g., the wisdom of grouping studies with widely diverse methods), and we suggest ways of improving future analyses of the behavior-analytic literature. PMID:22478319

  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.

  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. Synthesized quantitative assessment of human mental fatigue with EEG and HRV

    NASA Astrophysics Data System (ADS)

    Han, Qingpeng; Wang, Li; Wang, Ping; Wen, Bangchun

    2005-12-01

    The electroencephalograph (EEG) signals and heart rate variable (HRV) signals, which are relative to human body mental stress, are analyzed with the nonlinear dynamics and chaos. Based on calculated three nonlinear parameters, a synthesized quantitative criterion is proposed to assess the body's mental fatigue states. Firstly, the HRV and α wave of EEG from original signals are extracted based on wavelet transform technique. Then, the Largest Lyapunov Exponents, Complexity and Approximate Entropy, are calculated for both HRV and α wave. The three nonlinear parameters reflect quantitatively human physiological activities and can be used to evaluate the mental workload degree. Based on the computation and statistical analysis of practical EEG and HRV data, a synthesized quantitative assessment criterion is induced for mental fatigues with three nonlinear parameters of the above two rhythms. For the known 10 measured data of EEG and HRV signals, the assessment results are obtained with the above laws for different metal fatigue states. To compare with the practical cases, the identification accuracy of mental fatigue or not is up to 100 percent. Furthermore, the accuracies of weak fatigue, middle fatigue and serious fatigue mental workload are all relatively higher; they are about 94.44, 88.89, and 83.33 percent, respectively.

  10. 3-D visualization and quantitation of microvessels in transparent human colorectal carcinoma [corrected].

    PubMed

    Liu, Yuan-An; Pan, Shien-Tung; Hou, Yung-Chi; Shen, Ming-Yin; Peng, Shih-Jung; Tang, Shiue-Cheng; Chung, Yuan-Chiang

    2013-01-01

    Microscopic analysis of tumor vasculature plays an important role in understanding the progression and malignancy of colorectal carcinoma. However, due to the geometry of blood vessels and their connections, standard microtome-based histology is limited in providing the spatial information of the vascular network with a 3-dimensional (3-D) continuum. To facilitate 3-D tissue analysis, we prepared transparent human colorectal biopsies by optical clearing for in-depth confocal microscopy with CD34 immunohistochemistry. Full-depth colons were obtained from colectomies performed for colorectal carcinoma. Specimens were prepared away from (control) and at the tumor site. Taking advantage of the transparent specimens, we acquired anatomic information up to 200 μm in depth for qualitative and quantitative analyses of the vasculature. Examples are given to illustrate: (1) the association between the tumor microstructure and vasculature in space, including the perivascular cuffs of tumor outgrowth, and (2) the difference between the 2-D and 3-D quantitation of microvessels. We also demonstrate that the optically cleared mucosa can be retrieved after 3-D microscopy to perform the standard microtome-based histology (H&E staining and immunohistochemistry) for systematic integration of the two tissue imaging methods. Overall, we established a new tumor histological approach to integrate 3-D imaging, illustration, and quantitation of human colonic microvessels in normal and cancerous specimens. This approach has significant promise to work with the standard histology to better characterize the tumor microenvironment in colorectal carcinoma.

  11. Prenatal Diagnosis by Amniocentesis and Chorionic Villus Biopsy

    PubMed Central

    Reynolds, J. L.

    1986-01-01

    Prenatal diagnosis forms only a small part of day-to-day family practice, but the techniques are of critical importance to couples at risk of having a child affected by genetic disorder. Second trimester amniocentesis will probably be replaced by first trimester chorionic villus biopsy and recombinant DNA technology, but the ethical and moral problems related to prenatal diagnosis are not so easily solved. Family physicians need to examine their own attitudes toward the handicapped before they offer counselling to couples at risk of bearing handicapped children. The controversy over abortion is central to the issue of prenatal diagnosis, and may only be resolved by development of prenatal treatments for certain genetic disorders. Sometimes the only thing we can offer patients is to be with them, in whatever their decisions bring. PMID:21274247

  12. Mapping the developing human brain in utero using quantitative MR imaging techniques

    PubMed Central

    Studholme, Colin

    2015-01-01

    Magnetic resonance imaging of the human fetal brain has been a clinical tool for many years and provides valuable additional information to compliment more common ultrasound studies. Advances in both MRI acquisition and post processing over the last 10 years have enabled full 3D imaging and the accurate combination of data acquired in different head positions to create improved geometric integrity, tissue contrast and resolution. This research is now motivating the development of new quantitative MRI based techniques for clinical imaging that can more accurately characterize brain development and detect abnormalities. In this paper we will review some of the key areas that are driving changes in our understanding of fetal brain growth using quantitative measures derived from inutero MRI, and possible directions for its increased use in improving the evaluation of pregnancies and the accurate characterization of abnormal brain growth. PMID:25813665

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

    PubMed Central

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

    2011-01-01

    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

  14. Mapping the developing human brain in utero using quantitative MR imaging techniques.

    PubMed

    Studholme, Colin

    2015-03-01

    Magnetic resonance imaging of the human fetal brain has been a clinical tool for many years and provides valuable additional information to compliment more common ultrasound studies. Advances in both MRI acquisition and post processing over the last 10 years have enabled full 3D imaging and the accurate combination of data acquired in different head positions to create improved geometric integrity, tissue contrast, and resolution. This research is now motivating the development of new quantitative MRI-based techniques for clinical imaging that can more accurately characterize brain development and detect abnormalities. In this article, we will review some of the key areas that are driving changes in our understanding of fetal brain growth using quantitative measures derived from in utero MRI and the possible directions for its increased use in improving the evaluation of pregnancies and the accurate characterization of abnormal brain growth.

  15. Quantitation and Identification of Thousands of Human Proteoforms below 30 kDa.

    PubMed

    Durbin, Kenneth R; Fornelli, Luca; Fellers, Ryan T; Doubleday, Peter F; Narita, Masashi; Kelleher, Neil L

    2016-03-04

    Top-down proteomics is capable of identifying and quantitating unique proteoforms through the analysis of intact proteins. We extended the coverage of the label-free technique, achieving differential analysis of whole proteins <30 kDa from the proteomes of growing and senescent human fibroblasts. By integrating improved control software with more instrument time allocated for quantitation of intact ions, we were able to collect protein data between the two cell states, confidently comparing 1577 proteoform levels. To then identify and characterize proteoforms, our advanced acquisition software, named Autopilot, employed enhanced identification efficiency in identifying 1180 unique Swiss-Prot accession numbers at 1% false-discovery rate. This coverage of the low mass proteome is equivalent to the largest previously reported but was accomplished in 23% of the total acquisition time. By maximizing both the number of quantified proteoforms and their identification rate in an integrated software environment, this work significantly advances proteoform-resolved analyses of complex systems.

  16. Quantitative comparison of cardiac ventricular myocyte electrophysiology and response to drugs in human and nonhuman species.

    PubMed

    O'Hara, Thomas; Rudy, Yoram

    2012-03-01

    Explanations for arrhythmia mechanisms at the cellular level are usually based on experiments in nonhuman myocytes. However, subtle electrophysiological differences between species may lead to different rhythmic or arrhythmic cellular behaviors and drug response given the nonlinear and highly interactive cellular system. Using detailed and quantitatively accurate mathematical models for human, dog, and guinea pig ventricular action potentials (APs), we simulated and compared cell electrophysiology mechanisms and response to drugs. Under basal conditions (absence of β-adrenergic stimulation), Na(+)/K(+)-ATPase changes secondary to Na(+) accumulation determined AP rate dependence for human and dog but not for guinea pig where slow delayed rectifier current (I(Ks)) was the major rate-dependent current. AP prolongation with reduction of rapid delayed rectifier current (I(Kr)) and I(Ks) (due to mutations or drugs) showed strong species dependence in simulations, as in experiments. For humans, AP prolongation was 80% following I(Kr) block. It was 30% for dog and 20% for guinea pig. Under basal conditions, I(Ks) block was of no consequence for human and dog, but for guinea pig, AP prolongation after I(Ks) block was severe. However, with β-adrenergic stimulation, I(Ks) played an important role in all species, particularly in AP shortening at fast rate. Quantitative comparison of AP repolarization, rate-dependence mechanisms, and drug response in human, dog, and guinea pig revealed major species differences (e.g., susceptibility to arrhythmogenic early afterdepolarizations). Extrapolation from animal to human electrophysiology and drug response requires great caution.

  17. 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

  18. Quantitative analysis of cellular proteome alterations in human influenza A virus-infected mammalian cell lines.

    PubMed

    Vester, Diana; Rapp, Erdmann; Gade, Dörte; Genzel, Yvonne; Reichl, Udo

    2009-06-01

    Over the last years virus-host cell interactions were investigated in numerous studies. Viral strategies for evasion of innate immune response, inhibition of cellular protein synthesis and permission of viral RNA and protein production were disclosed. With quantitative proteome technology, comprehensive studies concerning the impact of viruses on the cellular machinery of their host cells at protein level are possible. Therefore, 2-D DIGE and nanoHPLC-nanoESI-MS/MS analysis were used to qualitatively and quantitatively determine the dynamic cellular proteome responses of two mammalian cell lines to human influenza A virus infection. A cell line used for vaccine production (MDCK) was compared with a human lung carcinoma cell line (A549) as a reference model. Analyzing 2-D gels of the proteomes of uninfected and influenza-infected host cells, 16 quantitatively altered protein spots (at least +/-1.7-fold change in relative abundance, p<0.001) were identified for both cell lines. Most significant changes were found for keratins, major components of the cytoskeleton system, and for Mx proteins, interferon-induced key components of the host cell defense. Time series analysis of infection processes allowed the identification of further proteins that are described to be involved in protein synthesis, signal transduction and apoptosis events. Most likely, these proteins are required for supporting functions during influenza viral life cycle or host cell stress response. Quantitative proteome-wide profiling of virus infection can provide insights into complexity and dynamics of virus-host cell interactions and may accelerate antiviral research and support optimization of vaccine manufacturing processes.

  19. Quantitative analysis of human platelet adhesions under a small-scale flow device.

    PubMed

    Furukawa, Katsuko S; Nakamura, Keigo; Onimura, Yuji; Uchida, Masaki; Ito, Atsuo; Yamane, Takashi; Tamaki, Tamotsu; Ushida, Takashi; Tateishi, Tetsuya

    2010-04-01

    To realize real-time evaluation of human platelet adhesions onto material surfaces with small volumes of human platelet suspensions, we developed an apparatus consisting of a modified cone and plate-type viscometer, combined with an upright epi-fluorescence microscope. The apparatus allowed real-time evaluation of platelet-material interactions and the initial event of thrombus formation, using small platelet suspension volumes (7.5 microL) under shear flow conditions. To study the dynamic behavior of platelet-material interaction, we chose five representative opaque and transparent materials: acrylate resin (AC), polytetrafluoroethylene (PTFE), polyvynylchrolide (PVC), glass, and a monolayer of human normal umbilical cord vein endothelial cells (EC) on glass under shear flow conditions. The values of adhesiveness of human platelets to the test materials in descending order were as follows: AC > PTFE > PVC > glass > human EC. Under this new small-scale flow system, we could obtain highly reproducible data, which were comparable with results from a previously developed large-scale flow system. Therefore, the newly developed cone and plate-type rheometer is a useful instrument for testing and screening materials, and allows precise quantitative evaluation of human platelet adhesion.

  20. 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

  1. Quantitative variability of 342 plasma proteins in a human twin population

    PubMed Central

    Liu, Yansheng; Buil, Alfonso; Collins, Ben C; Gillet, Ludovic CJ; Blum, Lorenz C; Cheng, Lin-Yang; Vitek, Olga; Mouritsen, Jeppe; Lachance, Genevieve; Spector, Tim D; Dermitzakis, Emmanouil T; Aebersold, Ruedi

    2015-01-01

    The degree and the origins of quantitative variability of most human plasma proteins are largely unknown. Because the twin study design provides a natural opportunity to estimate the relative contribution of heritability and environment to different traits in human population, we applied here the highly accurate and reproducible SWATH mass spectrometry technique to quantify 1,904 peptides defining 342 unique plasma proteins in 232 plasma samples collected longitudinally from pairs of monozygotic and dizygotic twins at intervals of 2–7 years, and proportioned the observed total quantitative variability to its root causes, genes, and environmental and longitudinal factors. The data indicate that different proteins show vastly different patterns of abundance variability among humans and that genetic control and longitudinal variation affect protein levels and biological processes to different degrees. The data further strongly suggest that the plasma concentrations of clinical biomarkers need to be calibrated against genetic and temporal factors. Moreover, we identified 13 cis-SNPs significantly influencing the level of specific plasma proteins. These results therefore have immediate implications for the effective design of blood-based biomarker studies. PMID:25652787

  2. Fluorescent nanodiamonds enable quantitative tracking of human mesenchymal stem cells in miniature pigs

    PubMed Central

    Su, Long-Jyun; Wu, Meng-Shiue; Hui, Yuen Yung; Chang, Be-Ming; Pan, Lei; Hsu, Pei-Chen; Chen, Yit-Tsong; Ho, Hong-Nerng; Huang, Yen-Hua; Ling, Thai-Yen; Hsu, Hsao-Hsun; Chang, Huan-Cheng

    2017-01-01

    Cell therapy is a promising strategy for the treatment of human diseases. While the first use of cells for therapeutic purposes can be traced to the 19th century, there has been a lack of general and reliable methods to study the biodistribution and associated pharmacokinetics of transplanted cells in various animal models for preclinical evaluation. Here, we present a new platform using albumin-conjugated fluorescent nanodiamonds (FNDs) as biocompatible and photostable labels for quantitative tracking of human placenta choriodecidual membrane-derived mesenchymal stem cells (pcMSCs) in miniature pigs by magnetic modulation. With this background-free detection technique and time-gated fluorescence imaging, we have been able to precisely determine the numbers as well as positions of the transplanted FND-labeled pcMSCs in organs and tissues of the miniature pigs after intravenous administration. The method is applicable to single-cell imaging and quantitative tracking of human stem/progenitor cells in rodents and other animal models as well. PMID:28358111

  3. 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.

  4. Physical-chemical and biological characterization of different preparations of equine chorionic gonadotropin

    PubMed Central

    Natal, Fabio Luis Nogueira; Ribela, Maria Teresa Carvalho Pinto; de Almeida, Beatriz Elane; de Oliveira, João Ezequiel; Bartolini, Paolo

    2016-01-01

    Ovarian stimulation with commercial preparations of equine chorionic gonadotropin (eCG) produces extremely variable responses in domestic animals, ranging from excessive stimulation to practically no stimulation, when applied on the basis of their declared unitage. This study was conducted to analyze four commercial preparations from different manufacturers via reversed-phase HPLC (RP-HPLC) in comparison with a reference preparation and an official International Standard from the World Health Organization. The peaks obtained by this qualitative and quantitative physical–chemical analysis were compared using an in vivo bioassay based on the ovarian weight gain of prepubertal female rats. The RP-HPLC data showed one or two peaks close to a main peak (tR = 27.9 min), which were related to the in vivo bioactivity. Commercial preparations that have this altered peak showed very little or no in vivo activity, as demonstrated by rat ovarian weight and in peripubertal gilts induced to ovulate. Overall, these findings indicate that RP-HPLC can be a rapid and reliable tool to reveal changes in the physicochemical profile of commercial eCG that is apparently related to decreased biological activity of this hormone. PMID:27297410

  5. Validation of internal reference genes for relative quantitation studies of gene expression in human laryngeal cancer

    PubMed Central

    Wang, Xiaofeng; He, Jinting; Wang, Wei; Ren, Ming; Gao, Sujie; Zhao, Guanjie

    2016-01-01

    Background The aim of this study was to determine the expression stabilities of 12 common internal reference genes for the relative quantitation analysis of target gene expression performed by reverse transcription real-time quantitative polymerase chain reaction (RT-qPCR) in human laryngeal cancer. Methods Hep-2 cells and 14 laryngeal cancer tissue samples were investigated. The expression characteristics of 12 internal reference gene candidates (18S rRNA, GAPDH, ACTB, HPRT1, RPL29, HMBS, PPIA, ALAS1, TBP, PUM1, GUSB, and B2M) were assessed by RT-qPCR. The data were analyzed by three commonly used software programs: geNorm, NormFinder, and BestKeeper. Results The use of the combination of four internal reference genes was more appropriate than the use of a single internal reference gene. The optimal combination was PPIA + GUSB + RPL29 + HPRT1 for both the cell line and tissues; while the most appropriate combination was GUSB + RPL29 + HPRT1 + HMBS for the tissues. Conclusions Our recommended internal reference genes may improve the accuracy of relative quantitation analysis of target gene expression performed by the RT-qPCR method in further gene expression research on laryngeal tumors. PMID:27957397

  6. 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.

  7. Multi-platform characterization of the human cerebrospinal fluid metabolome: a comprehensive and quantitative update

    PubMed Central

    2012-01-01

    Background Human cerebral spinal fluid (CSF) is known to be a rich source of small molecule biomarkers for neurological and neurodegenerative diseases. In 2007, we conducted a comprehensive metabolomic study and performed a detailed literature review on metabolites that could be detected (via metabolomics or other techniques) in CSF. A total of 308 detectable metabolites were identified, of which only 23% were shown to be routinely identifiable or quantifiable with the metabolomics technologies available at that time. The continuing advancement in analytical technologies along with the growing interest in CSF metabolomics has led us to re-visit the human CSF metabolome and to re-assess both its size and the level of coverage than can be achieved with today's technologies. Methods We used five analytical platforms, including nuclear magnetic resonance (NMR), gas chromatography-mass spectrometry (GC-MS), liquid chromatography-mass spectrometry (LC-MS), direct flow injection-mass spectrometry (DFI-MS/MS) and inductively coupled plasma-mass spectrometry (ICP-MS) to perform quantitative metabolomics on multiple human CSF samples. This experimental work was complemented with an extensive literature review to acquire additional information on reported CSF compounds, their concentrations and their disease associations. Results NMR, GC-MS and LC-MS methods allowed the identification and quantification of 70 CSF metabolites (as previously reported). DFI-MS/MS allowed the quantification of 78 metabolites (6 acylcarnitines, 13 amino acids, hexose, 42 phosphatidylcholines, 2 lyso-phosphatidylcholines and 14 sphingolipids), while ICP-MS provided quantitative results for 33 metal ions in CSF. Literature analysis led to the identification of 57 more metabolites. In total, 476 compounds have now been confirmed to exist in human CSF. Conclusions The use of improved metabolomic and other analytical techniques has led to a 54% increase in the known size of the human CSF metabolome

  8. 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.

  9. Rapid and Quantitative Assay of Amyloid-Seeding Activity in Human Brains Affected with Prion Diseases

    PubMed Central

    Takatsuki, Hanae; Satoh, Katsuya; Sano, Kazunori; Fuse, Takayuki; Nakagaki, Takehiro; Mori, Tsuyoshi; Ishibashi, Daisuke; Mihara, Ban; Takao, Masaki; Iwasaki, Yasushi; Yoshida, Mari; Atarashi, Ryuichiro; Nishida, Noriyuki

    2015-01-01

    The infectious agents of the transmissible spongiform encephalopathies are composed of amyloidogenic prion protein, PrPSc. Real-time quaking-induced conversion can amplify very small amounts of PrPSc seeds in tissues/body fluids of patients or animals. Using this in vitro PrP-amyloid amplification assay, we quantitated the seeding activity of affected human brains. End-point assay using serially diluted brain homogenates of sporadic Creutzfeldt–Jakob disease patients demonstrated that 50% seeding dose (SD50) is reached approximately 1010/g brain (values varies 108.79–10.63/g). A genetic case (GSS-P102L) yielded a similar level of seeding activity in an autopsy brain sample. The range of PrPSc concentrations in the samples, determined by dot-blot assay, was 0.6–5.4 μg/g brain; therefore, we estimated that 1 SD50 unit was equivalent to 0.06–0.27 fg of PrPSc. The SD50 values of the affected brains dropped more than three orders of magnitude after autoclaving at 121°C. This new method for quantitation of human prion activity provides a new way to reduce the risk of iatrogenic prion transmission. PMID:26070208

  10. 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.

  11. Quantitation of fixative-induced morphologic and antigenic variation in mouse and human breast cancers.

    PubMed

    Cardiff, Robert D; Hubbard, Neil E; Engelberg, Jesse A; Munn, Robert J; Miller, Claramae H; Walls, Judith E; Chen, Jane Q; Velásquez-García, Héctor A; Galvez, Jose J; Bell, Katie J; Beckett, Laurel A; Li, Yue-Ju; Borowsky, Alexander D

    2013-04-01

    Quantitative Image Analysis (QIA) of digitized whole slide images for morphometric parameters and immunohistochemistry of breast cancer antigens was used to evaluate the technical reproducibility, biological variability, and intratumoral heterogeneity in three transplantable mouse mammary tumor models of human breast cancer. The relative preservation of structure and immunogenicity of the three mouse models and three human breast cancers was also compared when fixed with representatives of four distinct classes of fixatives. The three mouse mammary tumor cell models were an ER+/PR+ model (SSM2), a Her2+ model (NDL), and a triple negative model (MET1). The four breast cancer antigens were ER, PR, Her2, and Ki67. The fixatives included examples of (1) strong cross-linkers, (2) weak cross-linkers, (3) coagulants, and (4) combination fixatives. Each parameter was quantitatively analyzed using modified Aperio Technologies ImageScope algorithms. Careful pre-analytical adjustments to the algorithms were required to provide accurate results. The QIA permitted rigorous statistical analysis of results and grading by rank order. The analyses suggested excellent technical reproducibility and confirmed biological heterogeneity within each tumor. The strong cross-linker fixatives, such as formalin, consistently ranked higher than weak cross-linker, coagulant and combination fixatives in both the morphometric and immunohistochemical parameters.

  12. Development of Multiplexed Real-Time Quantitative PCR Assay for Detecting Human Adenoviruses

    PubMed Central

    Huang, Meei-Li; Nguy, Long; Ferrenberg, James; Boeckh, Michael; Cent, Anne; Corey, Lawrence

    2008-01-01

    Adenoviruses (AdV) have been associated with a wide variety of human disease and are increasingly recognized as viral pathogens that can cause significant morbidity and mortality in immunocompromised patients. Early detection of AdV DNA in plasma and sterile fluids has been shown to be useful for identifying patients at risk for invasive AdV disease. Due to the large number of existing Adv types, few real-time quantitative AdV PCR assays published effectively cover all AdV types. We designed a series of AdV PCR primers and probes and empirically multiplexed them into two separate real-time PCR assays to quantitatively detect all 49 serotypes of human AdV (Types 1-49) available from ATCC. We then subsequently multiplexed all the primers and probes into one reaction. The sensitivity of these assays was determined to be less than 10 copies per reaction (500 copies/ml plasma). In a retrospective evaluation we detected all 84 clinical AdV isolates isolated in cell culture from patients undergoing hematopoietic stem cell transplant (HSCT) between 1981 and 1987. Prospective analysis of 46 consecutive clinical samples submitted for adenovirus testing showed greater sensitivity and equal specificity of the AdV PCR than viral culture. This real time PCR assay allows rapid, sensitive and specific quantification of all currently defined adenoviruses into either two or one multiplex assay for clinical samples. PMID:18707838

  13. 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

  14. A rapid LC-MS/MS method for quantitation of eszopiclone in human plasma: application to a human pharmacokinetic study.

    PubMed

    Hotha, Kishore Kumar; Vijaya Bharathi, D; Jagadeesh, B; Ravindranath, L K; Jaya Veera, K N; Venkateswarulu, V

    2012-02-01

    A highly reproducible, specific and cost-effective LC-MS/MS method was developed for simultaneous estimation of eszopiclone (ESZ) with 50 μL of human plasma using paroxetine as an internal standard (IS). The API-4000 LC-MS/MS was operated under the multiple reaction-monitoring mode using the electrospray ionization technique. A simple liquid-liquid extraction process was used to extract ESZ and IS from human plasma. The total run time was 1.5 min and the elution of ESZ and IS occurred at 0.90 min; this was achieved with a mobile phase consisting of 0.1% formic acid-methanol (15:85, v/v) at a flow rate of 0.50 mL/min on a Discover C(18) (50 × 4.6 mm, 5 µm) column. The developed method was validated in human plasma with a lower limit of quantitation of 0.1 ng/mL for ESZ. A linear response function was established for the range of concentrations 0.10-120 ng/mL (r > 0.998) for ESZ. The intra- and inter-day precision values for ESZ were acceptable as per FDA guidelines. Eszopiclone was stable in the battery of stability studies, viz. bench-top, autosampler and freeze-thaw cycles. The developed assay method was applied to an oral bioequivalence study in humans.

  15. Exploiting for Breast Cancer Control a Proposed Unified Mechanism for Reduction of Human Breast Cancer Risk by the Hormones of Pregnancy

    DTIC Science & Technology

    2006-05-01

    Human Chorionic Gonadotropin (hCG) Twenty-one days after carcinogen administration, 30 female rats received an intra-peritoneal injection of...estriol (E3) or human chorionic gonadotropin (hCG), are at a reduced risk of developing carcinogen induced breast cancer (1-4). On the other hand...was decreased by 27% (p< 0.026). Figure 1b. Breast Cancer Incidence in Human Chorionic Gonadotropin (hCG) Treated Rats. Thirty Sprague-Dawley

  16. Mobile phone radiation interferes laboratory immunoenzymometric assays: Example chorionic gonadotropin assays.

    PubMed

    Shahbazi-Gahrouei, Daryoush; Mortazavi, Seyed Mohammad Javad; Nasri, Hamid; Baradaran, Azar; Baradaran-Ghahfarokhi, Milad; Baradaran-Ghahfarokhi, Hamid Reza

    2012-02-01

    The radiofrequency radiation is of concern in hospital laboratories as the microwaves have many health effects even on immune functions. The aim of this study was, however, to evaluate the effects of cell phone radiation on chorionic gonadotropin immunoassays of human serum. Two cell phones with 0.69 and 1.09W/kg (head SAR) emitting 900MHz radiation were used. Sixty wells with five human serum concentrations (0, 10, 100, 250, 500mIU/mL) were used in three batches. The well heads in each batch were exposed to 900MHz emitted from these phones, and the 0.69, 1.09W/kg exposed batches were compared with the unexposed controls. Radiation exposure from mobile phones altered the measured serum levels especially in the wells with 100, 250, 500mIU/mL hormone concentrations. Exposure at 1.09W/kg SAR caused a significant loss compared to 0.69W/kg SAR exposure. In conclusion, the microwave exposures may require attention in laboratories using immunoassays.

  17. 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.

  18. Immunological quantitation and localization of ACAT-1 and ACAT-2 in human liver and small intestine.

    PubMed

    Chang, C C; Sakashita, N; Ornvold, K; Lee, O; Chang, E T; Dong, R; Lin, S; Lee, C Y; Strom, S C; Kashyap, R; Fung, J J; Farese, R V; Patoiseau, J F; Delhon, A; Chang, T Y

    2000-09-08

    By using specific anti-ACAT-1 antibodies in immunodepletion studies, we previously found that ACAT-1, a 50-kDa protein, plays a major catalytic role in the adult human liver, adrenal glands, macrophages, and kidneys but not in the intestine. Acyl-coenzyme A:cholesterol acyltransferase (ACAT) activity in the intestine may be largely derived from a different ACAT protein. To test this hypothesis, we produced specific polyclonal anti-ACAT-2 antibodies that quantitatively immunodepleted human ACAT-2, a 46-kDa protein expressed in Chinese hamster ovary cells. In hepatocyte-like HepG2 cells, ACAT-1 comprises 85-90% of the total ACAT activity, with the remainder attributed to ACAT-2. In adult intestines, most of the ACAT activity can be immunodepleted by anti-ACAT-2. ACAT-1 and ACAT-2 do not form hetero-oligomeric complexes. In differentiating intestinal enterocyte-like Caco-2 cells, ACAT-2 protein content increases by 5-10-fold in 6 days, whereas ACAT-1 protein content remains relatively constant. In the small intestine, ACAT-2 is concentrated at the apices of the villi, whereas ACAT-1 is uniformly distributed along the villus-crypt axis. In the human liver, ACAT-1 is present in both fetal and adult hepatocytes. In contrast, ACAT-2 is evident in fetal but not adult hepatocytes. Our results collectively suggest that in humans, ACAT-2 performs significant catalytic roles in the fetal liver and in intestinal enterocytes.

  19. Quantitative analysis of human salivary gland-derived intact proteome using top-down mass spectrometry.

    PubMed

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

    2014-05-01

    There are several notable challenges inherent for fully characterizing the entirety of the human saliva proteome using bottom-up approaches, including polymorphic isoforms, PTMs, unique splice variants, deletions, and truncations. To address these challenges, we have developed a top-down based LC-MS/MS approach, which cataloged 20 major human salivary proteins with a total of 83 proteoforms, containing a broad range of PTMs. Among these proteins, several previously reported disease biomarker proteins were identified at the intact protein level, such as beta-2 microglobulin. In addition, intact glycosylated proteoforms of several saliva proteins were also characterized, including intact N-glycosylated protein prolactin inducible protein and O-glycosylated acidic protein rich protein. 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 and submandibular/sublingual gland secretion samples (2 μg of protein each) from six healthy individuals were compared using RPLC coupled with the 12T FT-ICR mass spectrometer. Significantly different proteoform profiles were resolved with high reproducibility between parotid secretion and submandibular/sublingual 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.

  20. Quantitative analysis of trihydroxybutyl mercapturic acid, a urinary metabolite of 1, 3-butadiene, in humans

    PubMed Central

    Kotapati, Srikanth; Matter, Brock A.; Grant, Amy L.; Tretyakova, Natalia Y.

    2011-01-01

    1,3-butadiene (BD)* is a known human carcinogen present in cigarette smoke and in automobile exhaust, leading to widespread exposure of human populations. BD requires cytochrome P450-mediated metabolic activation to electrophilic species, e.g. 3,4-epoxy-1-butene (EB), hydroxymethylvinylketone (HMVK), and 3,4-epoxy-1,2-diol (EBD), which form covalent adducts with DNA. EB, HMVK, and EBD can be conjugated with glutathione and ultimately excreted in urine as monohydroxybutenyl mercapturic acid (MHBMA), dihydroxybutyl mercapturic acid (DHBMA), and trihydroxybutyl mercapturic acid (THBMA), respectively, which can serve as biomarkers of BD exposure and metabolic processing. While MHBMA and DHBMA have been found in smokers and non-smokers, THBMA has not been previously detected in humans. In the present work, an isotope dilution HPLC-ESI−-MS/MS methodology was developed and employed to quantify THBMA in urine of known smokers and non-smokers (19–27 per group). The new method has excellent sensitivity (LOQ, 1 ng/mL urine) and achieves accurate quantitation using a small sample volume (100 µl). Mean urinary THBMA concentrations in smokers and non-smokers were found to be 21.6 and 13.7 ng/mg creatinine, respectively, suggesting that there are sources of THBMA other than exposure to tobacco smoke in humans, as is also the case for DHBMA. However, THBMA concentrations are significantly greater in urine of smokers than that of non-smokers (p < 0.01). Furthermore, THBMA amounts in human urine declined 25–50 % following smoking cessation, suggesting that smoking is an important source of this metabolite in humans. The HPLC-ESI−-MS/MS methodology developed in the present work will be useful for future epidemiological studies of BD exposure and metabolism. PMID:21749114

  1. Vasoactive intestinal polypeptide immunoreactivity in the human cerebellum: qualitative and quantitative analyses

    PubMed Central

    Benagiano, Vincenzo; Flace, Paolo; Lorusso, Loredana; Rizzi, Anna; Bosco, Lorenzo; Cagiano, Raffaele; Ambrosi, Glauco

    2009-01-01

    Although autoradiographic, reverse transcription-polymerase chain reaction and immunohistochemical studies have demonstrated receptors for vasoactive intestinal polypeptide (VIP) in the cerebellum of various species, immunohistochemistry has never shown immunoreactivity for VIP within cerebellar neuronal bodies and processes. The present study aimed to ascertain whether VIP immunoreactivity really does exist in the human cerebellum by making a systematic analysis of samples removed post-mortem from all of the cerebellar lobes. The study was carried out using light microscopy immunohistochemical techniques based on a set of four different antibodies (three polyclonal and one monoclonal) against VIP, carefully selected on the basis of control tests performed on human colon. All of the antibodies used showed VIP-immunoreactive neuronal bodies and processes distributed in the cerebellar cortex and subjacent white matter of all of the cerebellum lobes, having similar qualitative patterns of distribution. Immunoreactive neurons included subpopulations of the main neuron types of the cortex. Statistical analysis of the quantitative data on the VIP immunoreactivity revealed by the different antibodies in the different cerebellar lobes did not demonstrate any significant differences. In conclusion, using four different anti-VIP antibodies, the first evidence of VIP immunoreactivity is herein supplied in the human post-mortem cerebellum, with similar qualitative/quantitative patterns of distribution among the different cerebellum lobes. Owing to the function performed by VIP as a neurotransmitter/neuromodulator, it is a candidate for a role in intrinsic and extrinsic (projective) circuits of the cerebellum, in agreement with previous demonstrations of receptors for VIP in the cerebellar cortex and nuclei. As VIP signalling pathways are implicated in the regulation of cognitive and psychic functions, cerebral blood flow and metabolism, processes of histomorphogenesis

  2. A quantitative comparison of the similarity between genes and geography in worldwide human populations.

    PubMed

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

    2012-08-01

    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.

  3. 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.

  4. 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...

  5. 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 ...

  6. 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...

  7. Preimplantation diagnosis or chorionic villus biopsy? Women's attitudes and preferences.

    PubMed

    Miedzybrodzka, Z; Templeton, A; Dean, J; Haites, N; Mollison, J; Smith, N

    1993-12-01

    The objective of this work was to assess women's attitudes and preferences to two methods of prenatal diagnosis for genetic disease: embryo and chorionic villus biopsy (CVS). The design involved a structured postal questionnaire sent to women in the Grampian region with different reproductive experiences. The population sample included 46 women who had had genetic counselling because of a family history of a single gene disorder, 18 women who had had CVS for a single gene disorder, 158 women who had had CVS for other reasons, 200 women who had recently delivered a normal baby and 50 women who had experience of in-vitro fertilization (IVF). The main outcome measures were attitudes to family limitation, prenatal diagnosis, termination for fetal abnormality, embryo biopsy and CVS. Of the women surveyed, 38% preferred embryo diagnosis, whereas 42% favoured CVS and termination. Women with previous experience of CVS preferred this option whereas those with experience of IVF as infertility treatment were more likely to favour embryo diagnosis, as were women who had had genetic counselling for a single gene disorder. It was concluded that a substantial number of women find embryo diagnosis more acceptable than CVS when the pregnancy is at high risk. This is especially true amongst those with experience of IVF or who are at risk themselves. A demand for embryo diagnosis has been demonstrated.

  8. Quantitation of Human Immunodeficiency Virus Type 1 RNA in Different Biological Compartments

    PubMed Central

    Shepard, Robin N.; Schock, Jody; Robertson, Kevin; Shugars, Diane C.; Dyer, John; Vernazza, Pietro; Hall, Colin; Cohen, Myron S.; Fiscus, Susan A.

    2000-01-01

    Little information is available describing viral loads in body fluids other than blood. In addition, the suitability of commercially available assays for human immunodeficiency virus type 1 (HIV-1) RNA quantitation has not been evaluated in most nonblood fluids. We compared Organon Teknika's nucleic acid sequence-based amplification method (NASBA) and Roche's Amplicor HIV-1 Monitor (reverse transcriptase PCR [RT-PCR]) for quantitating HIV-1 RNA in cerebrospinal fluid (CSF), saliva, breast milk, seminal plasma, and cervical-vaginal lavage fluid (CVL). Saliva and breast milk frequently demonstrated some inhibition in the RT-PCR assay, similar to the inhibition previously described in seminal plasma. Inhibition of the RT-PCR assay was not observed with CSF or CVL, nor in any of the NASBA assays. When fluids from HIV-infected individuals were tested by RT-PCR and NASBA, 73 and 27% of CSF samples and 60 and 40% of breast milk specimens had detectable RNA, respectively. These differences were not statistically significant. In cross-sectional studies using RT-PCR to measure viral RNA in paired blood plasma and CSF samples, 71% of blood plasma samples and 42% of CSF samples were positive. A similar analysis using NASBA with paired blood plasma and CVL, saliva, or seminal plasma samples revealed 91% were blood plasma positive and 55% were CVL positive, 76% were blood plasma positive and 46% were saliva positive, and 83% were blood plasma positive and 63% were seminal plasma positive. NASBA worked fairly well to quantitate HIV-1 RNA from all fluids without apparent inhibition. RT-PCR performed well on CVL and CSF, frequently with greater sensitivity, although its use in other fluids appears limited due to the presence of inhibitors. These studies demonstrate that viral loads in nonblood fluids were generally lower than in blood. PMID:10747117

  9. Human judgment vs. quantitative models for the management of ecological resources.

    PubMed

    Holden, Matthew H; Ellner, Stephen P

    2016-07-01

    Despite major advances in quantitative approaches to natural resource management, there has been resistance to using these tools in the actual practice of managing ecological populations. Given a managed system and a set of assumptions, translated into a model, optimization methods can be used to solve for the most cost-effective management actions. However, when the underlying assumptions are not met, such methods can potentially lead to decisions that harm the environment and economy. Managers who develop decisions based on past experience and judgment, without the aid of mathematical models, can potentially learn about the system and develop flexible management strategies. However, these strategies are often based on subjective criteria and equally invalid and often unstated assumptions. Given the drawbacks of both methods, it is unclear whether simple quantitative models improve environmental decision making over expert opinion. In this study, we explore how well students, using their experience and judgment, manage simulated fishery populations in an online computer game and compare their management outcomes to the performance of model-based decisions. We consider harvest decisions generated using four different quantitative models: (1) the model used to produce the simulated population dynamics observed in the game, with the values of all parameters known (as a control), (2) the same model, but with unknown parameter values that must be estimated during the game from observed data, (3) models that are structurally different from those used to simulate the population dynamics, and (4) a model that ignores age structure. Humans on average performed much worse than the models in cases 1-3, but in a small minority of scenarios, models produced worse outcomes than those resulting from students making decisions based on experience and judgment. When the models ignored age structure, they generated poorly performing management decisions, but still outperformed

  10. Comparative quantitative analysis of osseous anatomy of the craniovertebral junction of tiger, horse, deer, and humans

    PubMed Central

    Goel, Atul; Shah, Abhidha; Kothari, Manu; Gaikwad, Santosh; Dhande, Prakash L.

    2011-01-01

    Aim: To compare the osseous anatomy of the craniovertebral junction of a horse, deer, and tiger with that of a human being. The variation in the structure of bones in these animals is analyzed. Materials and Methods: Various dimensions of the bones of the craniovertebral junction of the horse, deer, and tiger were quantitatively measured, and their differences with those of human bones were compared and analyzed. Results: Apart from the sizes and weights, there are a number of structural variations in the bones of these animals that depend on their functional needs. The more remarkable difference in joint morphology is noticed in the occipitoatlantal joint. The occipitoatlantal articulation is remarkably large and deep, resembling a ‘hinge joint’ in all the three animals studied. The odontoid process is ‘C shaped’ in the deer and horse and is ‘denslike’ in the tiger and humans. The transverse processes of the atlas are in the form of large wings in all the three animals. The arches of the atlas are large and flat, but the traverse of the vertebral artery resembles, to an extent, to that of human vertebral artery. The rotatory movements of the head at the craniovertebral junction are wider ranged in the horse and deer as compared with those of the tiger and humans. The bones of the craniovertebral junction of all the three animals are adapted to the remarkable thickness and strength of the extensor muscles of the nape of the neck. Conclusions: Despite the wide variations in the size of the bones, the basic patterns of structure, vascular and neural relationship, and joint alignments have remarkable similarities and a definite pattern of differences. PMID:22013373

  11. Age-related decline in prostacyclin synthesis by human aortic endothelial cells. Qualitative and quantitative analysis.

    PubMed Central

    Tokunaga, O.; Yamada, T.; Fan, J. L.; Watanabe, T.

    1991-01-01

    To investigate the functional alteration of human aortic endothelial cells with aging, prostacyclin synthesis was qualitatively and quantitatively examined. The endothelial cells of human aortas and umbilical veins or inferior vena cavae were immunohistochemically examined and found positive for prostacyclin, but the intensity of aortic endothelial cells from older subjects was low. In addition to the endothelial cells, smooth muscle cells in the thickened intima, not the media, of the aorta were also immunoreactive. Endothelial cells were successfully cultured from human aortas obtained from infants through aged subjects and were subdivided into three groups: young, middle, and old. Prostacyclin synthesis by endothelial cells from all types of blood vessels was extremely great at the primary culture, but decreased abruptly in the following subcultures. Among the aortic endothelial cells, the young group synthesized the largest amount of prostacyclin in a conventional culture condition, with synthesis progressively decreasing in the older groups. The in vitro prostacyclin biosynthesis was supported by the qualitative analysis on the tissue sections. These results indicate that prostacyclin synthesis of the aortic endothelial cells decreases with age, but intimal smooth muscle cells potentially have a back-up mechanism and substitute this synthesis to some extent. The decreased synthesis of prostacyclin with age may play an important role in the development and advancement of thrombosis and atherosclerosis. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:1707240

  12. Large-scale human skin lipidomics by quantitative, high-throughput shotgun mass spectrometry

    PubMed Central

    Sadowski, Tomasz; Klose, Christian; Gerl, Mathias J.; Wójcik-Maciejewicz, Anna; Herzog, Ronny; Simons, Kai; Reich, Adam; Surma, Michal A.

    2017-01-01

    The lipid composition of human skin is essential for its function; however the simultaneous quantification of a wide range of stratum corneum (SC) and sebaceous lipids is not trivial. We developed and validated a quantitative high-throughput shotgun mass spectrometry-based platform for lipid analysis of tape-stripped SC skin samples. It features coverage of 16 lipid classes; total quantification to the level of individual lipid molecules; high reproducibility and high-throughput capabilities. With this method we conducted a large lipidomic survey of 268 human SC samples, where we investigated the relationship between sampling depth and lipid composition, lipidome variability in samples from 14 different sampling sites on the human body and finally, we assessed the impact of age and sex on lipidome variability in 104 healthy subjects. We found sebaceous lipids to constitute an abundant component of the SC lipidome as they diffuse into the topmost SC layers forming a gradient. Lipidomic variability with respect to sampling depth, site and subject is considerable, and mainly accredited to sebaceous lipids, while stratum corneum lipids vary less. This stresses the importance of sampling design and the role of sebaceous lipids in skin studies. PMID:28266621

  13. 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.

  14. 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

  15. Quantitation and Identification of Thousands of Human Proteoforms below 30 kDa

    PubMed Central

    Durbin, Kenneth R.; Fornelli, Luca; Fellers, Ryan T.; Doubleday, Peter F.; Narita, Masashi; Kelleher, Neil L.

    2016-01-01

    Top-down proteomics is capable of identifying and quantitating unique proteoforms through the analysis of intact proteins. We extended the coverage of the label-free technique, achieving differential analysis of whole proteins <30 kDa from the proteomes of growing and senescent human fibroblasts. By integrating improved control software with more instrument time allocated for quantitation of intact ions, we were able to collect protein data between the two cell states, confidently comparing 1577 proteoform levels. To then identify and characterize proteoforms, our advanced acquisition software, named Autopilot, employed enhanced identification efficiency in identifying 1180 unique Swiss-Prot accession numbers at 1% false-discovery rate. This coverage of the low mass proteome is equivalent to the largest previously reported but was accomplished in 23% of the total acquisition time. By maximizing both the number of quantified proteoforms and their identification rate in an integrated software environment, this work significantly advances proteoform-resolved analyses of complex systems. PMID:26795204

  16. 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.

  17. 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.

  18. Quantitation of the residual DNA from rice-derived recombinant human serum albumin.

    PubMed

    Chen, Zhen; Dai, Huixia; Liu, Zhenwei; Zhang, Liping; Pang, Jianlei; Ou, Jiquan; Yang, Daichang

    2014-04-01

    Residual DNA in recombinant protein pharmaceuticals can potentially cause safety issues in clinical applications; thus, maximum residual limit has been established by drug safety authorities. Assays for residual DNA in Escherichia coli, yeast, and Chinese hamster ovary (CHO) cell expression systems have been established, but no rice residual DNA assay for rice expression systems has been designed. To develop an assay for the quantification of residual DNA that is produced from rice seed, we established a sensitive assay using quantitative real-time polymerase chain reaction (qPCR) based on the 5S ribosomal RNA (rRNA) genes. We found that a 40-cycle qPCR exhibited a linear response when the template concentration was in the range of 2×10(4) to 0.2pg of DNA per reaction in TaqMan and SYBR Green I assays. The amplification efficiency was 103 to 104%, and the amount of residual DNA from recombinant human serum albumin from Oryza sativa (OsrHSA) was less than 3.8ng per dosage, which was lower than that recommended by the World Health Organization (WHO). Our results indicate that the current purification protocol could efficiently remove residual DNA during manufacturing and processing. Furthermore, this protocol could be viable in other cereal crop endosperm expression systems for developing a residual DNA quantitation assay using the highly conserved 5S rRNA gene of the crops.

  19. Standardisation of the quantitation of serum amyloid A protein (SAA) in human serum.

    PubMed

    Godenir, N L; Jeenah, M S; Coetzee, G A; Van der Westhuyzen, D R; Strachan, A F; De Beer, F C

    1985-11-07

    An adequate method for standardising the quantitation of serum amyloid A protein (SAA) in human serum was developed. Acute phase high density lipoprotein3 (HDL3) was used as a standard. The concentration of the SAA in the standard was determined by the use of purified SAA. After protein determination, various concentrations of purified SAA were run on SDS-polyacrylamide gel together with the HDL3 standard containing an unknown amount of SAA amongst the apolipoproteins. From the standard curve obtained by pyridine extraction (Coomassie blue colour yield at A605 nm) the concentration of SAA in the HDL3 standard was determined. An established immunoradiometric assay (IRMA) for SAA was standardised with the HDL3. SAA concentrations in normal and acute phase sera were determined.

  20. Biomarkers are used to predict quantitative metabolite concentration profiles in human red blood cells

    PubMed Central

    Palsson, Bernhard O.

    2017-01-01

    Deep-coverage metabolomic profiling has revealed a well-defined development of metabolic decay in human red blood cells (RBCs) under cold storage conditions. A set of extracellular biomarkers has been recently identified that reliably defines the qualitative state of the metabolic network throughout this metabolic decay process. Here, we extend the utility of these biomarkers by using them to quantitatively predict the concentrations of other metabolites in the red blood cell. We are able to accurately predict the concentration profile of 84 of the 91 (92%) measured metabolites (p < 0.05) in RBC metabolism using only measurements of these five biomarkers. The median of prediction errors (symmetric mean absolute percent error) across all metabolites was 13%. The ability to predict numerous metabolite concentrations from a simple set of biomarkers offers the potential for the development of a powerful workflow that could be used to evaluate the metabolic state of a biological system using a minimal set of measurements. PMID:28264007

  1. Quantitative Biomechanics of Healthy and Diseased Human Red Blood Cells using Dielectrophoresis in a Microfluidic System.

    PubMed

    Du, E; Dao, Ming; Suresh, Subra

    2014-12-01

    We present an experimental method to quantitatively characterize the mechanical properties of a large number of biological cells by introducing controlled deformation through dielectrophoresis in a microfluidic device. We demonstrate the capability of this technique by determining the force versus deformation characteristics of healthy human red blood cells (RBCs) and RBCs infected in vitro with Plasmodium falciparum malaria parasites. These experiments clearly distinguish uninfected and healthy RBCs from infected ones, and the mechanical signatures extracted from these tests are in agreement with data from other independent methods. The method developed here thus provides a potentially helpful tool to characterize quickly and effectively the isolated biomechanical response of cells in a large population, for probing the pathological states of cells, disease diagnostics, and drug efficacy assays.

  2. Quantitative analysis of the toxicity of human amniotic fluid to cultured rat spinal cord.

    PubMed

    Drewek, M J; Bruner, J P; Whetsell, W O; Tulipan, N

    1997-10-01

    It has been proposed that the myelodysplastic components of a myelomeningocele are secondarily damaged as the result of exposure to amniotic fluid, the so-called 'two-hit' hypothesis. The critical time at which this secondary insult might occur has not been clearly defined. The present study addresses this issue by quantitatively assessing the toxic effects of human amniotic fluid of various gestational ages upon organotypic cultures of rat spinal cord. Using an assay for lactate dehydrogenase efflux to evaluate toxicity in such spinal cord cultures, we found that the amniotic fluid became toxic at approximately 34 weeks' gestation. This toxic effect of amniotic fluid appears to emerge rather suddenly. Accordingly, it seems reasonable to suggest that prevention of exposure of vulnerable spinal cord tissue to this toxicity by surgical closure of a myelomeningocele defect prior to the emergence of toxicity in amniotic fluid may prevent injury to vulnerable myelodysplastic spinal cord tissue.

  3. 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.

  4. Targeted LC-MS/MS Method for the Quantitation of Plant Lignans and Enterolignans in Biofluids from Humans and Pigs.

    PubMed

    Nørskov, Natalja P; Olsen, Anja; Tjønneland, Anne; Bolvig, Anne Katrine; Lærke, Helle Nygaard; Knudsen, Knud Erik Bach

    2015-07-15

    Lignans have gained nutritional interest due to their promising role in the prevention of lifestyle diseases. However, epidemiological studies are in need of more evidence to link the intake of lignans to this promising role. In this context, it is necessary to study large population groups to obtain sufficient statistical power. Therefore, there is a demand for fast, sensitive, and accurate methods for quantitation with high throughput of samples. This paper presents a validated LC-MS/MS method for the quantitation of eight plant lignans (matairesinol, hydroxymatairesinol, secoisolariciresinol, lariciresinol, isolariciresinol, syringaresinol, medioresinol, and pinoresinol) and two enterolignans (enterodiol and enterolactone) in both human and pig plasma and urine. The method showed high selectivity and sensitivity allowing quantitation of lignans in the range of 0.024-100 ng/mL and with a run time of only 4.8 min per sample. The method was successfully applied to quantitate lignans in biofluids from ongoing studies with humans and pigs.

  5. 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.

  6. 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

    2016-01-04

    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

  7. 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.

  8. An Alu-based, MGB Eclipse real-time PCR method for quantitation of human DNA in forensic samples.

    PubMed

    Nicklas, Janice A; Buel, Eric

    2005-09-01

    The forensic community needs quick, reliable methods to quantitate human DNA in crime scene samples to replace the laborious and imprecise slot blot method. A real-time PCR based method has the possibility of allowing development of a faster and more quantitative assay. Alu sequences are primate-specific and are found in many copies in the human genome, making these sequences an excellent target or marker for human DNA. This paper describes the development of a real-time Alu sequence-based assay using MGB Eclipse primers and probes. The advantages of this assay are simplicity, speed, less hands-on-time and automated quantitation, as well as a large dynamic range (128 ng/microL to 0.5 pg/microL).

  9. 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

  10. Rat Genome Database: a unique resource for rat, human, and mouse quantitative trait locus data.

    PubMed

    Nigam, Rajni; Laulederkind, Stanley J F; Hayman, G Thomas; Smith, Jennifer R; Wang, Shur-Jen; Lowry, Timothy F; Petri, Victoria; De Pons, Jeff; Tutaj, Marek; Liu, Weisong; Jayaraman, Pushkala; Munzenmaier, Diane H; Worthey, Elizabeth A; Dwinell, Melinda R; Shimoyama, Mary; Jacob, Howard J

    2013-09-16

    The rat has been widely used as a disease model in a laboratory setting, resulting in an abundance of genetic and phenotype data from a wide variety of studies. These data can be found at the Rat Genome Database (RGD, http://rgd.mcw.edu/), which provides a platform for researchers interested in linking genomic variations to phenotypes. Quantitative trait loci (QTLs) form one of the earliest and core datasets, allowing researchers to identify loci harboring genes associated with disease. These QTLs are not only important for those using the rat to identify genes and regions associated with disease, but also for cross-organism analyses of syntenic regions on the mouse and the human genomes to identify potential regions for study in these organisms. Currently, RGD has data on >1,900 rat QTLs that include details about the methods and animals used to determine the respective QTL along with the genomic positions and markers that define the region. RGD also curates human QTLs (>1,900) and houses>4,000 mouse QTLs (imported from Mouse Genome Informatics). Multiple ontologies are used to standardize traits, phenotypes, diseases, and experimental methods to facilitate queries, analyses, and cross-organism comparisons. QTLs are visualized in tools such as GBrowse and GViewer, with additional tools for analysis of gene sets within QTL regions. The QTL data at RGD provide valuable information for the study of mapped phenotypes and identification of candidate genes for disease associations.

  11. Quantitative risk assessment of human salmonellosis in Canadian broiler chicken breast from retail to consumption.

    PubMed

    Smadi, Hanan; Sargeant, Jan M

    2013-02-01

    The current quantitative risk assessment model followed the framework proposed by the Codex Alimentarius to provide an estimate of the risk of human salmonellosis due to consumption of chicken breasts which were bought from Canadian retail stores and prepared in Canadian domestic kitchens. The model simulated the level of Salmonella contamination on chicken breasts throughout the retail-to-table pathway. The model used Canadian input parameter values, where available, to represent risk of salmonellosis. From retail until consumption, changes in the concentration of Salmonella on each chicken breast were modeled using equations for growth and inactivation. The model predicted an average of 318 cases of salmonellosis per 100,000 consumers per year. Potential reasons for this overestimation were discussed. A sensitivity analysis showed that concentration of Salmonella on chicken breasts at retail and food hygienic practices in private kitchens such as cross-contamination due to not washing cutting boards (or utensils) and hands after handling raw meat along with inadequate cooking contributed most significantly to the risk of human salmonellosis. The outcome from this model emphasizes that responsibility for protection from Salmonella hazard on chicken breasts is a shared responsibility. Data needed for a comprehensive Canadian Salmonella risk assessment were identified for future research.

  12. Laser-induced fluorescence: quantitative analysis of atherosclerotic plaque chemical content in human aorta

    NASA Astrophysics Data System (ADS)

    Dai, Erbin; Wishart, David; Khoury, Samir; Kay, Cyril M.; Jugdutt, Bodh I.; Tulip, John; Lucas, Alexandra

    1996-05-01

    We have been studying laser-induced fluorescence as a technique for identification of selected changes in the chemical composition of atherosclerotic plaque. Formulae for quantification of chemical changes have been developed based upon analysis of fluorescence emission spectra using multiple regression analysis and the principal of least squares. The intima of human aortic necropsy specimens was injected with chemical compounds present in atherosclerotic plaque. Spectra recorded after injection of selected chemical components found in plaque (collagen I, III, IV, elastin and cholesterol) at varying concentrations (0.01 - 1.0 mg) were compared with saline injection. A single fiber system was used for both fluorescence excitation (XeCl excimer laser, 308 nm, 1.5 - 2.0 mJ/ pulse, 5 Hz) and fluorescence emission detection. Average spectra for each chemical have been developed and the wavelengths of peak emission intensity identified. Curve fitting analysis as well as multiple regression analysis were used to develop formulae for assessment of chemical content. Distinctive identifying average curves were established for each chemical. Excellent correlations were identified for collagen I, III, and IV, elastin, and cholesterol (R2 equals 0.92 6- 0.997). Conclusions: (1) Fluorescence spectra of human aortas were significantly altered by collagen I, collagen III, elastin and cholesterol. (2) Fluorescence spectroscopic analysis may allow quantitative assessment of atherosclerotic plaque chemical content in situ.

  13. 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

    2015-05-20

    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.

  14. Quantitation of venom antigens from European vipers in human serum or urine by ELISA.

    PubMed

    Audebert, F; Grosselet, O; Sabouraud, A; Bon, C

    1993-01-01

    We describe an enzyme-linked immunosorbent assay (ELISA) to quantitate venom antigens in human serum and urine, and thus to help evaluate the severity of envenomation due to viper bites. This assay, which is performed with commercially available polyclonal Fab'2s in a double-sandwich method, is rapid, simple, and specific for antigens of European vipers (Vipera aspis, Vipera berus, and Vipera ammodytes). No cross-reactivity was observed with other snake venoms or human serum proteins. It showed a good linear response over a wide range of concentrations of venom antigens (from 1 to 100 ng/mL). It was very sensitive, with detection limits of 7 and 2 ng/mL for Vipera aspis venom in serum and urine, respectively. This ELISA is also easily reproducible; the coefficients of variation determined at different concentrations of venom (50, 25, and 5 ng/mL) did not exceed 10% in serum and 14% in urine samples collected from different donors. This test was applied to determine the concentrations of venom in the serum of patients bitten by a viper in France and to follow its elimination as a function of time. The method is adaptable to other venoms by using other specific immunoglobulins.

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

    PubMed Central

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

    2014-01-01

    To assess responses to low-dose ionizing radiation (LD-IR) exposures potentially encountered during medical diagnostic procedures, nuclear accidents or terrorist acts, a quantitative proteomic approach was used to identify changes in protein abundance in a reconstituted human skin tissue model treated with 0.1 Gy of ionizing radiation. To improve the dynamic range of the assay, subcellular fractionation was employed to remove highly abundant structural proteins and to provide insight into radiation-induced alterations in protein localization. Relative peptide quantification across cellular fractions, control and irradiated samples was performing using 8-plex iTRAQ labeling followed by online two-dimensional nano-scale liquid chromatography and high resolution MS/MS analysis. A total of 107 proteins were detected with statistically significant radiation-induced change in abundance (>1.5 fold) and/or subcellular localization compared to controls. The top biological pathways identified using bioinformatics include organ development, anatomical structure formation and the regulation of actin cytoskeleton. From the proteomic data, a change in proteolytic processing and subcellular localization of the skin barrier protein, filaggrin, was identified, and the results were confirmed by western blotting. This data indicate post-transcriptional regulation of protein abundance, localization and proteolytic processing playing an important role in regulating radiation response in human tissues. PMID:28250387

  16. The quantitative proteomes of human-induced pluripotent stem cells and embryonic stem cells.

    PubMed

    Munoz, Javier; Low, Teck Y; Kok, Yee J; Chin, Angela; Frese, Christian K; Ding, Vanessa; Choo, Andre; Heck, Albert J R

    2011-11-22

    Assessing relevant molecular differences between human-induced pluripotent stem cells (hiPSCs) and human embryonic stem cells (hESCs) is important, given that such differences may impact their potential therapeutic use. Controversy surrounds recent gene expression studies comparing hiPSCs and hESCs. Here, we present an in-depth quantitative mass spectrometry-based analysis of hESCs, two different hiPSCs and their precursor fibroblast cell lines. Our comparisons confirmed the high similarity of hESCs and hiPSCS at the proteome level as 97.8% of the proteins were found unchanged. Nevertheless, a small group of 58 proteins, mainly related to metabolism, antigen processing and cell adhesion, was found significantly differentially expressed between hiPSCs and hESCs. A comparison of the regulated proteins with previously published transcriptomic studies showed a low overlap, highlighting the emerging notion that differences between both pluripotent cell lines rather reflect experimental conditions than a recurrent molecular signature.

  17. Novel quantitative pigmentation phenotyping enhances genetic association, epistasis, and prediction of human eye colour

    PubMed Central

    Wollstein, Andreas; Walsh, Susan; Liu, Fan; Chakravarthy, Usha; Rahu, Mati; Seland, Johan H.; Soubrane, Gisèle; Tomazzoli, Laura; Topouzis, Fotis; Vingerling, Johannes R.; Vioque, Jesus; Böhringer, Stefan; Fletcher, Astrid E.; Kayser, Manfred

    2017-01-01

    Success of genetic association and the prediction of phenotypic traits from DNA are known to depend on the accuracy of phenotype characterization, amongst other parameters. To overcome limitations in the characterization of human iris pigmentation, we introduce a fully automated approach that specifies the areal proportions proposed to represent differing pigmentation types, such as pheomelanin, eumelanin, and non-pigmented areas within the iris. We demonstrate the utility of this approach using high-resolution digital eye imagery and genotype data from 12 selected SNPs from over 3000 European samples of seven populations that are part of the EUREYE study. In comparison to previous quantification approaches, (1) we achieved an overall improvement in eye colour phenotyping, which provides a better separation of manually defined eye colour categories. (2) Single nucleotide polymorphisms (SNPs) known to be involved in human eye colour variation showed stronger associations with our approach. (3) We found new and confirmed previously noted SNP-SNP interactions. (4) We increased SNP-based prediction accuracy of quantitative eye colour. Our findings exemplify that precise quantification using the perceived biological basis of pigmentation leads to enhanced genetic association and prediction of eye colour. We expect our approach to deliver new pigmentation genes when applied to genome-wide association testing. PMID:28240252

  18. QUANTITATIVE SCREENING OF SINGLE COPIES OF HUMAN PAPILLOMA VIRAL DNA WITHOUT AMPLIFICATION

    PubMed Central

    Li, Jiangwei; Lee, Ji-Young; Yeung, Edward S.

    2008-01-01

    We describe a novel quantitative viral screening method based on single-molecule detection that does not require amplification. DNA of human papilloma virus (HPV), the major etiological agent of cervical cancer, served as the screening target in this study. Eight 100-nucleotide (nt) single-stranded (ss)-DNA probes were designed complementary to the E6-E7 gene of HPV-16 DNA. The probes were covalently stained with Alexa Fluor 532 and hybridized to the target in solution. The individual hybridized molecules were imaged with an intensified charge-coupled device (ICCD) in two ways. In the single-color mode, target molecules were detected via fluorescence from hybridized probes only. This system could detect HPV-16 DNA in the presence of human genomic DNA down to 0.7 copy/cell, and had a linear dynamic range of over six orders of magnitude. In the dual-color mode, we employed fluorescence resonance energy transfer (FRET) and added YOYO-3 dye as the acceptor. The two colors from Alexa Fluor 532 and YOYO-3 were dispersed by a transmission grating located in front of the ICCD. With this reinforced criteria for identifying the hybridized molecules, zero false-positive count was achieved. We also showed that DNA extracts from Pap test specimens did not interfere with the measurements. PMID:16970325

  19. Novel quantitative pigmentation phenotyping enhances genetic association, epistasis, and prediction of human eye colour.

    PubMed

    Wollstein, Andreas; Walsh, Susan; Liu, Fan; Chakravarthy, Usha; Rahu, Mati; Seland, Johan H; Soubrane, Gisèle; Tomazzoli, Laura; Topouzis, Fotis; Vingerling, Johannes R; Vioque, Jesus; Böhringer, Stefan; Fletcher, Astrid E; Kayser, Manfred

    2017-02-27

    Success of genetic association and the prediction of phenotypic traits from DNA are known to depend on the accuracy of phenotype characterization, amongst other parameters. To overcome limitations in the characterization of human iris pigmentation, we introduce a fully automated approach that specifies the areal proportions proposed to represent differing pigmentation types, such as pheomelanin, eumelanin, and non-pigmented areas within the iris. We demonstrate the utility of this approach using high-resolution digital eye imagery and genotype data from 12 selected SNPs from over 3000 European samples of seven populations that are part of the EUREYE study. In comparison to previous quantification approaches, (1) we achieved an overall improvement in eye colour phenotyping, which provides a better separation of manually defined eye colour categories. (2) Single nucleotide polymorphisms (SNPs) known to be involved in human eye colour variation showed stronger associations with our approach. (3) We found new and confirmed previously noted SNP-SNP interactions. (4) We increased SNP-based prediction accuracy of quantitative eye colour. Our findings exemplify that precise quantification using the perceived biological basis of pigmentation leads to enhanced genetic association and prediction of eye colour. We expect our approach to deliver new pigmentation genes when applied to genome-wide association testing.

  20. Quantitative second-harmonic generation imaging to detect osteogenesis imperfecta in human skin samples

    NASA Astrophysics Data System (ADS)

    Adur, J.; Ferreira, A. E.; D'Souza-Li, L.; Pelegati, V. B.; de Thomaz, A. A.; Almeida, D. B.; Baratti, M. O.; Carvalho, H. F.; Cesar, C. L.

    2012-03-01

    Osteogenesis Imperfecta (OI) is a genetic disorder that leads to bone fractures due to mutations in the Col1A1 or Col1A2 genes that affect the primary structure of the collagen I chain with the ultimate outcome in collagen I fibrils that are either reduced in quantity or abnormally organized in the whole body. A quick test screening of the patients would largely reduce the sample number to be studied by the time consuming molecular genetics techniques. For this reason an assessment of the human skin collagen structure by Second Harmonic Generation (SHG) can be used as a screening technique to speed up the correlation of genetics/phenotype/OI types understanding. In the present work we have used quantitative second harmonic generation (SHG) imaging microscopy to investigate the collagen matrix organization of the OI human skin samples comparing with normal control patients. By comparing fibril collagen distribution and spatial organization, we calculated the anisotropy and texture patterns of this structural protein. The analysis of the anisotropy was performed by means of the two-dimensional Discrete Fourier Transform and image pattern analysis with Gray-Level Co-occurrence Matrix (GLCM). From these results, we show that statistically different results are obtained for the normal and disease states of OI.

  1. 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.

  2. 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

  3. Detection and differentiation of human parvovirus variants by commercial quantitative real-time PCR tests.

    PubMed

    Hokynar, Kati; Norja, Päivi; Laitinen, Harri; Palomäki, Pekka; Garbarg-Chenon, Antoine; Ranki, Annamari; Hedman, Klaus; Söderlund-Venermo, Maria

    2004-05-01

    Parvovirus B19 causes a variety of diseases in humans, with outcomes ranging from asymptomatic to severe, such as chronic anemia in immunocompromised patients or fetal hydrops and death after maternal infection during pregnancy. The virus may be transmitted via plasma-derived products. According to the results of solvent-detergent safety studies, an upper limit of B19 DNA in plasma pools was recently defined. To restrict the input of B19 virus into production pools, a quantitative nucleic acid test is a prerequisite. We examined the suitability of the two commercial quantitative B19 PCR tests, LightCycler-Parvovirus B19 quantification kit (Roche Diagnostics) and RealArt Parvo B19 LC PCR (Artus) for detection, quantification, and differentiation of the three known B19 genotypes, including the newly described erythrovirus variants (genotypes 2 and 3). The former kit was highly sensitive for genotype 1 but was not suitable for detection of genotype 2 or one of two genotype 3 strains. The latter kit detected and differentiated all three genotypes, albeit with lower sensitivity for one of the genotype-3 strains. We furthermore assessed the prevalence of the three B19 virus genotypes in blood donors, by screening pooled plasma samples derived from 140,160 Finnish blood-donor units. None of the pools contained detectable levels of B19 virus genotypes 2 or 3. The origin, mode of transmission, and clinical significance of these genotypes are unknown and deserve further study. The RealArt Parvo B19 LC PCR is suitable for detection, quantification, and differentiation of all three B19 virus genotypes in molecular and clinical research.

  4. 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.

  5. 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.

  6. Quantitation of midazolam in human plasma by automated chip-based infusion nanoelectrospray tandem mass spectrometry.

    PubMed

    Kapron, James T; Pace, Ellen; Van Pelt, Colleen K; Henion, Jack

    2003-01-01

    An automated chip-based infusion nanoelectrospray ionization (nanoESI) platform was used to demonstrate reproducible quantitation of drug molecules from biological matrices. Three sample preparation strategies were explored including protein precipitation of plasma with acetonitrile, de-salting of the plasma, and a combination of protein precipitation with subsequent de-salting of the dried and reconstituted extract. The best results were obtained when fortified human plasma samples containing midazolam were precipitated with acetonitrile containing alprazolam as the internal standard (IS). The supernatant was concentrated to dryness, reconstituted in aqueous acid, and de-salted by automated reversed-phase solid-phase extraction (SPE) prior to infusion nanoESI-MS/MS. Analyses employed a triple quadrupole mass spectrometer operated in selected reaction monitoring (SRM) mode. Each sample was infused for approximately 10 s and the resulting ion current profiles were integrated. Area ratios were used for regression analysis of standard samples (1.5-500 ng/mL). Quality control samples (3, 250, and 400 ng/mL) in five replicates from three different analysis days demonstrated intra-assay precision (< or =16%), inter-assay precision (< or =5%), and overall accuracy (+/-9% deviation). Infusion reproducibility of the assay was established by analyzing extracts after storage for 24 h at ambient temperature. Control plasma samples from six different sources probed the potential utility of this technique for the analysis of clinical samples. At the lower limit of quantitation (LLQ), variability and mean overall accuracy were < or =13% CV and +/-3% deviation, respectively, while at the upper limit of quantitation (ULQ) variability and mean overall accuracy were < or =9% CV and +/-9% deviation, respectively. Inter-chip variability was established by determining standard sample extracts across five different chips (< or =12% CV). Throughput for the assay was 55 s per sample

  7. A quantitative threshold for high/low extent of urinary excretion of compounds in humans.

    PubMed

    Dave, Rutwij A; Morris, Marilyn E

    2016-07-01

    In this study, a quantitative threshold was determined for the high/low extent of urinary excretion (UE) of compounds in humans, using a straightforward but robust statistical method known as receiver operating characteristic curve (ROC) analysis, and also 18 potential physicochemical determinants of UE were evaluated. Data on the percent of drug excreted unchanged into the urine, %Ae , were used to determine the threshold for high/low UE. Compounds can be divided into high/low UE groups using the threshold value of Ae  = 16.8%, namely those with Ae  > 16.8% are classified as high UE and those with Ae  ≤ 16.8% as low UE. The %Ae negatively correlated with cLogP (r = -0.56); however, cLogP could not quantitatively predict the value of %Ae (R(2) adj. = 0.32). Several determinants of the extent of UE, including cLogP, ACD labs cLogP and ACD labs cLogD(pH=7.4) , were successfully evaluated as a priori indicators of the extent of UE using two cut-off values for each parameter. Moreover, 87% of the 90 compounds in the external validation set were correctly classified using this approach. Analysis of the physicochemical spaces of compounds in these two groups showed significant overlap, which hinders the a priori determination of the extent of UE of compounds using a single threshold/cut-off value of simple physicochemical parameters. In conclusion, 16.8% is a quantitative threshold value to distinguish between high and low UE and new molecular entities with cLogP and ACD labs cLogP values of ≤0.7 and ≥1.0 and ACD labs cLogD(pH=7.4) values of ≤0.0 and ≥0.5 could be identified as exhibiting high and low UE, respectively. Copyright © 2016 John Wiley & Sons, Ltd.

  8. Development of a Quantitative Bead Capture Assay for Soluble IL-7 Receptor Alpha in Human Plasma

    PubMed Central

    Faucher, Sylvie; Crawley, Angela M.; Decker, Wendy; Sherring, Alice; Bogdanovic, Dragica; Ding, Tao; Bergeron, Michele; Angel, Jonathan B.; Sandstrom, Paul

    2009-01-01

    Background IL-7 is an essential cytokine in T-cell development and homeostasis. It binds to the IL-7R receptor, a complex of the IL-7Rα (CD127) and common γ (CD132) chains. There is significant interest in evaluating the expression of CD127 on human T-cells as it often decreased in medical conditions leading to lymphopenia. Previous reports showed the usefulness of CD127 as a prognostic marker in viral infections such as HIV, CMV, EBV and HCV. A soluble CD127 (sCD127) is released in plasma and may contribute to disease pathogenesis through its control on IL-7 activities. Measuring sCD127 is important to define its role and may complement existing markers used in lymphopenic disease management. We describe a new quantitative assay for the measurement of sCD127 in plasma and report sCD127 concentrations in healthy adults. Methodology/Principal Findings We developed a quantitative bead-based sCD127 capture assay. Polyclonal CD127-specific antibodies were chosen for capture and a biotinylated monoclonal anti-CD127 antibody was selected for detection. The assay can detect native sCD127 and recombinant sCD127 which served as the calibrator. The analytical performance of the assay was characterized and the concentration and stability of plasma sCD127 in healthy adults was determined. The assay's range was 3.2–1000 ng/mL. The concentration of plasma sCD127 was 164±104 ng/mL with over a log variation between subjects. Individual sCD127 concentrations remained stable when measured serially during a period of up to one year. Conclusions/Significance This is the first report on the quantification of plasma sCD127 in a population of healthy adults. Soluble CD127 plasma concentrations remained stable over time in a given individual and sCD127 immunoreactivity was resistant to repeated freeze-thaw cycles. This quantitative sCD127 assay is a valuable tool for defining the potential role of sCD127 in lymphopenic diseases. PMID:19690616

  9. Morphology and Ploidy of Smooth Muscle Cells in Chorionic Arteries under Different Hemodynamic Conditions.

    PubMed

    Gansburgskii, A N; Yal'tsev, A V

    2017-02-01

    Smooth muscle cells from the arterial wall of placental chorion were studied at 39-40-week gestation. The content of mono- and binuclear tetraploid myocytes was higher in sites of arterial branching and turns (27.3% vs. 4.4% straight parts of the arteries; DNA cytophotometry data). Mitoses were found only in these arterial regions (0.18%). Regional changes in the sizes of diploid and polyploid myocytes were detected, associated with the blood flow pattern in the chorion; myocyte hypertrophy was 17-fold more incident in sites of arterial turns and branching than in straight arteries. Possible causes of changes in the proliferative characteristics and subsequent growth of the chorionic arterial wall myocytes are discussed.

  10. Chorion type as a possible influence on the results and interpretation of twin study data.

    PubMed

    Prescott, C A; Johnson, R C; McArdle, J J

    1999-12-01

    The estimation of genetic effects from twin studies usually relies upon the equal environment assumption--that monozygous (MZ) and dizygous (DZ) twin pairs experience equal similarity of their environments from prenatal experiences through adulthood. However, the sharing of a chorion may make a subset of identical twins more similar, or in some cases, more different, than twins that do not share a chorion. Recent studies suggest monochorionic MZ twins resemble one another more than dichorionic MZ twins in cognitive abilities, personality, and risk for psychiatric disorder. To the extent that prenatal environment affects these characteristics, the traditional twin method will yield biased estimates of genetic and environmental influences. We develop models for quantifying this bias and estimating the influence of chorion type on estimates of heritability.

  11. GWAS implicates a role for quantitative immune traits and threshold effects in risk for human autoimmune disorders.

    PubMed

    Gregersen, Peter K; Diamond, Betty; Plenge, Robert M

    2012-10-01

    Genome wide association studies in human autoimmune disorders have provided a long list of alleles with rather modest degrees of risk. A large fraction of these associations are probably owing 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 in 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.

  12. [THE COMPARATIVE ANALYSIS OF RESULTS OF DETECTION OF CARCINOGENIC TYPES OF HUMAN PAPILLOMA VIRUS BY QUALITATIVE AND QUANTITATIVE TESTS].

    PubMed

    Kuzmenko, E T; Labigina, A V; Leshenko, O Ya; Rusanov, D N; Kuzmenko, V V; Fedko, L P; Pak, I P

    2015-05-01

    The analysis of results of screening (n = 3208; sexually active citizen aged from 18 to 59 years) was carried out to detect oncogene types of human papilloma virus in using qualitative (1150 females and 720 males) and quantitative (polymerase chain reaction in real-time (843 females and 115 males) techniques. The human papilloma virus of high oncogene type was detected in 65% and 68.4% of females and in 48.6% and 53% of males correspondingly. Among 12 types of human papilloma virus the most frequently diagnosed was human papilloma virus 16 independently of gender of examined and technique of analysis. In females, under application of qualitative tests rate of human papilloma virus 16 made up to 18.3% (n = 280) and under application of quantitative tests Rte of human papilloma virus made up to 14.9% (n = 126; p ≤ 0.05). Under examination of males using qualitative tests rate of human papilloma virus 16 made up to 8.3% (n = 60) and under application of qualitative tests made up to 12.2% (n = 14; p ≥ 0.05). Under application of qualitative tests rate of detection on the rest ofoncogene types of human papilloma virus varied in females from 3.4% to 8.4% and in males from 1.8% to 5.9%. Under application of qualitative tests to females rate of human papilloma virus with high viral load made up to 68.4%, with medium viral load - 2.85% (n = 24) and with low viral load -0.24% (n = 2). Under application of quantitative tests in males rate of detection of types of human papilloma virus made up to 53% and at that in all high viral load was established. In females, the most of oncogene types of human papilloma virus (except for 31, 39, 59) are detected significantly more often than in males.

  13. 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…

  14. 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...

  15. Isolation and Quantitative Immunocytochemical Characterization of Primary Myogenic Cells and Fibroblasts from Human Skeletal Muscle

    PubMed Central

    Agley, Chibeza C.; Rowlerson, Anthea M.; Velloso, Cristiana P.; Lazarus, Norman L.; Harridge, Stephen D. R.

    2015-01-01

    The repair and regeneration of skeletal muscle requires the action of satellite cells, which are the resident muscle stem cells. These can be isolated from human muscle biopsy samples using enzymatic digestion and their myogenic properties studied in culture. Quantitatively, the two main adherent cell types obtained from enzymatic digestion are: (i) the satellite cells (termed myogenic cells or muscle precursor cells), identified initially as CD56+ and later as CD56+/desmin+ cells and (ii) muscle-derived fibroblasts, identified as CD56– and TE-7+. Fibroblasts proliferate very efficiently in culture and in mixed cell populations these cells may overrun myogenic cells to dominate the culture. The isolation and purification of different cell types from human muscle is thus an important methodological consideration when trying to investigate the innate behavior of either cell type in culture. Here we describe a system of sorting based on the gentle enzymatic digestion of cells using collagenase and dispase followed by magnetic activated cell sorting (MACS) which gives both a high purity (>95% myogenic cells) and good yield (~2.8 x 106 ± 8.87 x 105 cells/g tissue after 7 days in vitro) for experiments in culture. This approach is based on incubating the mixed muscle-derived cell population with magnetic microbeads beads conjugated to an antibody against CD56 and then passing cells though a magnetic field. CD56+ cells bound to microbeads are retained by the field whereas CD56– cells pass unimpeded through the column. Cell suspensions from any stage of the sorting process can be plated and cultured. Following a given intervention, cell morphology, and the expression and localization of proteins including nuclear transcription factors can be quantified using immunofluorescent labeling with specific antibodies and an image processing and analysis package. PMID:25650991

  16. Brain regions involved in human movement perception: a quantitative voxel-based meta-analysis.

    PubMed

    Grosbras, Marie-Hélène; Beaton, Susan; Eickhoff, Simon B

    2012-02-01

    Face, hands, and body movements are powerful signals essential for social interactions. In the last 2 decades, a large number of brain imaging studies have explored the neural correlates of the perception of these signals. Formal synthesis is crucially needed, however, to extract the key circuits involved in human motion perception across the variety of paradigms and stimuli that have been used. Here, we used the activation likelihood estimation (ALE) meta-analysis approach with random effect analysis. We performed meta-analyses on three classes of biological motion: movement of the whole body, hands, and face. Additional analyses of studies of static faces or body stimuli and sub-analyses grouping experiments as a function of their control stimuli or task employed allowed us to identify main effects of movements and forms perception, as well as effects of task demand. In addition to specific features, all conditions showed convergence in occipito-temporal and fronto-parietal regions, but with different peak location and extent. The conjunction of the three ALE maps revealed convergence in all categories in a region of the right posterior superior temporal sulcus as well as in a bilateral region at the junction between middle temporal and lateral occipital gyri. Activation in these regions was not a function of attentional demand and was significant also when controlling for non-specific motion perception. This quantitative synthesis points towards a special role for posterior superior temporal sulcus for integrating human movement percept, and supports a specific representation for body parts in middle temporal, fusiform, precentral, and parietal areas.

  17. Amniocentesis and chorionic villus sampling for prenatal diagnosis

    PubMed Central

    Alfirevic, Zarko; Mujezinovic, Faris; Sundberg, Karin

    2014-01-01

    Background A major disadvantage of second trimester amniocentesis is that the results are available relatively late in pregnancy (after 16 weeks’ gestation). Chorionic villus sampling (CVS) and early amniocentesis can be done in the first trimester of pregnancy and offer an earlier alternative. Objectives To assess comparative safety and accuracy of second trimester amniocentesis, early amniocentesis, transcervical and transabdominal CVS. Search methods We searched the Cochrane Pregnancy and Childbirth Group’s Trials Register (January 2008). Selection criteria All randomised trials comparing amniocentesis and CVS by either transabdominal or transcervical route. Data collection and analysis Two review authors independently assessed eligibility and trial quality and performed data extraction. Main results We included a total of 16 randomised studies. One study in a low-risk population (N = 4606) with a background pregnancy loss of around 2% found that a second trimester amniocentesis will increase total pregnancy loss by another 1%. This difference did not reach statistical significance and the confidence intervals (CI) around this excess risk were relatively large (risk ratio (RR) 1.41; 95% CI 0.99 to 2.00). In the same study, compared with no intervention, the increase in spontaneous miscarriages following second trimester amniocentesis was statistically significant (2.1% versus 1.3%; RR 1.60; 95% CI 1.02 to 2.52). Early amniocentesis is not a safe early alternative to second trimester amniocentesis because of increased pregnancy loss (7.6% versus 5.9%; RR 1.29; 95% CI 1.03 to 1.61) and higher incidence of talipes compared to CVS (RR 4.61; 95% CI 1.82 to 11.66). Compared with a second trimester amniocentesis, transcervical CVS carries a higher risk of pregnancy loss, although the results are quite heterogeneous. One study compared transabdominal CVS with second trimester amniocentesis and found no significant difference in the total pregnancy loss between the

  18. Quantitative Circulatory Physiology: an integrative mathematical model of human physiology for medical education.

    PubMed

    Abram, Sean R; Hodnett, Benjamin L; Summers, Richard L; Coleman, Thomas G; Hester, Robert L

    2007-06-01

    We have developed Quantitative Circulatory Physiology (QCP), a mathematical model of integrative human physiology containing over 4,000 variables of biological interactions. This model provides a teaching environment that mimics clinical problems encountered in the practice of medicine. The model structure is based on documented physiological responses within peer-reviewed literature and serves as a dynamic compendium of physiological knowledge. The model is solved using a desktop, Windows-based program, allowing students to calculate time-dependent solutions and interactively alter over 750 parameters that modify physiological function. The model can be used to understand proposed mechanisms of physiological function and the interactions among physiological variables that may not be otherwise intuitively evident. In addition to open-ended or unstructured simulations, we have developed 30 physiological simulations, including heart failure, anemia, diabetes, and hemorrhage. Additional stimulations include 29 patients in which students are challenged to diagnose the pathophysiology based on their understanding of integrative physiology. In summary, QCP allows students to examine, integrate, and understand a host of physiological factors without causing harm to patients. This model is available as a free download for Windows computers at http://physiology.umc.edu/themodelingworkshop.

  19. Absolute Quantitation of Water and Metabolites in the Human Brain. II. Metabolite Concentrations

    NASA Astrophysics Data System (ADS)

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

    A method for determining absolute metabolite concentrations with in vivo1H magnetic resonance spectroscopy is presented. Using the compartmentation model introduced in the preceding paper of this series ( J. Magn. Reson. B102, 1, 1993), it is possible to express NMR results in terms of most commonly used concentration units. The proposed scheme, involving the measurement of an external standard as well as of the localized water signal, is verified on cerebral spectra obtained from 22 subjects. Besides concentrations, longitudinal and transverse relaxation times are determined for parietal white and occipital gray matter. The determination of these quantities crucially depends on the analysis of the T2 signal decay as a function of echo time. The in vivo concentrations of the four metabolites N-acetyl aspartate, creatine plus phosphocreatine, choline, and myo-inositol are in good agreement with biochemical determinations performed in vitro. Two clinical examples emphasize the relevance of absolute quantitation in the investigation of human neuropathology and normal development.

  20. Quantitative analysis of untreated hair samples for monitoring human exposure to heavy metals

    NASA Astrophysics Data System (ADS)

    Sera, K.; Futatsugawa, S.; Murao, S.

    2002-04-01

    The method of quantitative analysis for untreated hair samples, which we developed three years ago, has proved to be quite useful for investigating environments contaminated by certain toxic elements. In the present work, the experimental conditions are improved. Loss of certain elements owing to irradiation damage, which has remained as one of the experimental uncertainties, was examined. It was found that the concentration of sulfur decreases gradually throughout the irradiation, while for the other elements, including arsenic and mercury, no changes occur under our measuring conditions. Furthermore, the degree of alteration of elemental concentration depending on the position along the hair was investigated. As a result, concentrations of some elements at different positions on a 14-cm-length hair, which was taken from a small-scale miner in the Philippines, showed some dependence on the distance from the root reflecting her history as a miner, while mercury does not show large deviation from a main trend. It was also found that concentration of mercury in hairs taken from different parts of a body does not show large difference. These results demonstrate that mercury and arsenic concentration in hairs, obtained by the present method, become a good index for an estimation of human exposure to these toxic elements. Changes of concentration of some elements depending on the way of cleaning before irradiation are studied in detail and the optimum way of washing is established.

  1. Quantitative susceptibility mapping for investigating subtle susceptibility variations in the human brain.

    PubMed

    Schweser, Ferdinand; Sommer, Karsten; Deistung, Andreas; Reichenbach, Jürgen Rainer

    2012-09-01

    Quantitative susceptibility mapping (QSM) is a novel magnetic resonance-based technique that determines tissue magnetic susceptibility from measurements of the magnetic field perturbation. Due to the ill-posed nature of this problem, regularization strategies are generally required to reduce streaking artifacts on the computed maps. The present study introduces a new algorithm for calculating the susceptibility distribution utilizing a priori information on its regional homogeneity derived from gradient echo phase images and analyzes the impact of erroneous a priori information on susceptibility map fidelity. The algorithm, Homogeneity Enabled Incremental Dipole Inversion (HEIDI), was investigated with a special focus on the reconstruction of subtle susceptibility variations in a numerical model and in volunteer data and was compared with two recently published approaches, Thresholded K-space Division (TKD) and Morphology Enabled Dipole Inversion (MEDI). HEIDI resulted in susceptibility maps without streaking artifacts and excellent depiction of subtle susceptibility variations in most regions. By investigating HEIDI susceptibility maps acquired with the volunteers' heads in different orientations, it was demonstrated that the apparent magnetic susceptibility distribution of human brain tissue considerably depends on the direction of the main magnetic field.

  2. Scale-Dependent Variability and Quantitative Regimes in Graph-Theoretic Representations of Human Cortical Networks

    PubMed Central

    Irimia, Andrei

    2016-01-01

    Abstract Studying brain connectivity is important due to potential differences in brain circuitry between health and disease. One drawback of graph-theoretic approaches to this is that their results are dependent on the spatial scale at which brain circuitry is examined and explicitly on how vertices and edges are defined in network models. To investigate this, magnetic resonance and diffusion tensor images were acquired from 136 healthy adults, and each subject's cortex was parceled into as many as 50,000 regions. Regions were represented as nodes in a reconstructed network representation, and interregional connectivity was inferred via deterministic tractography. Network model behavior was explored as a function of nodal number and connectivity weighing. Three distinct regimes of quantitative behavior assumed by network models as a function of spatial scale are identified, and their existence may be modulated by the spatial folding scale of the cortex. The maximum number of network nodes used to model human brain circuitry in this study (∼50,000) is larger than in previous macroscale neuroimaging studies. Results suggest that network model properties vary appreciably as a function of vertex assignment convention and edge weighing scheme and that graph-theoretic analysis results should not be compared across spatial scales without appropriate understanding of how spatial scale and model topology modulate network model properties. These findings have implications for comparing macro- to mesoscale studies of brain network models and understanding how choosing network-theoretic parameters affects the interpretation of brain connectivity studies. PMID:26596775

  3. 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.).

  4. Quantitative assessment of sensing and sequestration of spherocytic erythrocytes by the human spleen

    PubMed Central

    Buffet, Pierre A.; Deplaine, Guillaume; Perrot, Sylvie; Brousse, Valentine; Ndour, Alioune; Nguyen, Marie; Mercereau-Puijalon, Odile; David, Peter H.; Milon, Geneviève; Mohandas, Narla

    2012-01-01

    Splenic sequestration of RBCs with reduced surface area and cellular deformability has long been recognized as contributing to pathogenesis of several RBC disorders, including hereditary spherocytosis. However, the quantitative relationship between the extent of surface area loss and splenic entrapment remains to be defined. To address this issue, in the present study, we perfused ex vivo normal human spleens with RBCs displaying various degrees of surface area loss and monitored the kinetics of their splenic retention. Treatment with increasing concentrations of lysophosphatidylcholine resulted in a dose-dependent reduction of RBC surface area at constant volume, increased osmotic fragility, and decreased deformability. The degree of splenic retention of treated RBCs increased with increasing surface area loss. RBCs with a > 18% average surface area loss (> 27% reduced surface area-to-volume ratio) were rapidly and completely entrapped in the spleen. Surface-deficient RBCs appeared to undergo volume loss after repeated passages through the spleen and escape from splenic retention. The results of the present study for the first time define the critical extent of surface area loss leading to splenic entrapment and identify an adaptive volume regulation mechanism that allows spherocytic RBCs to prolong their life span in circulation. These results have significant implications for understanding the clinical heterogeneity of RBC membrane disorders. PMID:22510876

  5. Non-destructive assessment of human ribs mechanical properties using quantitative ultrasound.

    PubMed

    Mitton, David; Minonzio, Jean-Gabriel; Talmant, Maryline; Ellouz, Rafaa; Rongieras, Frédéric; Laugier, Pascal; Bruyère-Garnier, Karine

    2014-04-11

    Advanced finite element models of the thorax have been developed to study, for example, the effects of car crashes. While there is a need for material properties to parameterize such models, specific properties are largely missing. Non-destructive techniques applicable in vivo would, therefore, be of interest to support further development of thorax models. The only non-destructive technique available today to derive rib bone properties would be based on quantitative computed tomography that measures bone mineral density. However, this approach is limited by the radiation dose. Bidirectional ultrasound axial transmission was developed on long bones ex vivo and used to assess in vivo health status of the radius. However, it is currently unknown if the ribs are good candidates for such a measurement. Therefore, the goal of this study is to evaluate the relationship between ex vivo ultrasonic measurements (axial transmission) and the mechanical properties of human ribs to determine if the mechanical properties of the ribs can be quantified non-destructively. The results show statistically significant relationships between the ultrasonic measurements and mechanical properties of the ribs. These results are promising with respect to a non-destructive and non-ionizing assessment of rib mechanical properties. This ex vivo study is a first step toward in vivo studies to derive subject-specific rib properties.

  6. Quantitative proteomics of extracellular vesicles derived from human primary and metastatic colorectal cancer cells.

    PubMed

    Choi, Dong-Sic; Choi, Do-Young; Hong, Bok Sil; Jang, Su Chul; Kim, Dae-Kyum; Lee, Jaewook; Kim, Yoon-Keun; Kim, Kwang Pyo; Gho, Yong Song

    2012-01-01

    Cancer cells actively release extracellular vesicles (EVs), including exosomes and microvesicles, into surrounding tissues. These EVs play pleiotropic roles in cancer progression and metastasis, including invasion, angiogenesis, and immune modulation. However, the proteomic differences between primary and metastatic cancer cell-derived EVs remain unclear. Here, we conducted comparative proteomic analysis between EVs derived from human primary colorectal cancer cells (SW480) and their metastatic derivatives (SW620). Using label-free quantitation, we identified 803 and 787 proteins in SW480 EVs and SW620 EVs, respectively. Based on comparison between the estimated abundance of EV proteins, we identified 368 SW480 EV-enriched and 359 SW620 EV-enriched proteins. SW480 EV-enriched proteins played a role in cell adhesion, but SW620 EV-enriched proteins were associated with cancer progression and functioned as diagnostic indicators of metastatic cancer; they were overexpressed in metastatic colorectal cancer and played roles in multidrug resistance. As the first proteomic analysis comparing primary and metastatic cancer-derived EVs, this study increases our understanding of the pathological function of EVs in the metastatic process and provides useful biomarkers for cancer metastasis.

  7. 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

  8. Quantitative Analysis of Age Specific Variation in the Abundance of Human Female Parotid Salivary Proteins

    PubMed Central

    Ambatipudi, Kiran S.; Lu, Bingwen; Hagen, Fred K; Melvin, James E.; Yates, John R.

    2010-01-01

    Summary Human saliva is a protein-rich, easily accessible source of potential local and systemic biomarkers to monitor changes that occur under pathological conditions; however little is known about the changes in abundance associated with normal aging. In this study, we performed a comprehensive proteomic profiling of pooled saliva collected from the parotid glands of healthy female subjects, divided into two age groups 1 and 2 (20–30 and 55–65 years old, respectively). Hydrophobic charge interaction chromatography was used to separate high from low abundant proteins prior to characterization of the parotid saliva using multidimensional protein identification technology (MudPIT). Collectively, 532 proteins were identified in the two age groups. Of these proteins, 266 were identified exclusively in one age group, while 266 proteins were common to both groups. The majority of the proteins identified in the two age groups belonged to the defense and immune response category. Of note, several defense related proteins (e.g. lysozyme, lactoferrin and histatin-1) were significantly more abundant in group 2 as determined by G-test. Selected representative mass spectrometric findings were validated by western blot analysis. Our study reports the first quantitative analysis of differentially regulated proteins in ductal saliva collected from young and older female subjects. This study supports the use of high-throughput proteomics as a robust discovery tool. Such results provide a foundation for future studies to identify specific salivary proteins which may be linked to age-related diseases specific to women. PMID:19764810

  9. Functional analysis and quantitative determination of the expression profile of human parvovirus B19.

    PubMed

    Bonvicini, Francesca; Filippone, Claudia; Manaresi, Elisabetta; Zerbini, Marialuisa; Musiani, Monica; Gallinella, Giorgio

    2008-11-25

    Comprehension of the pathogenetic potential of human parvovirus B19 requires the definition of the complete spectrum of cellular tropism and a functional analysis of the viral genome in infected cells. In this study, we carried out a systematic functional analysis of B19 virus genome in the course of infection of susceptible bone marrow mononuclear cells and myeloblastoid UT7/EpoS1 cells, in terms of dynamics of nucleic acid synthesis. A PCR array was designed and a comprehensive analysis was performed by quantitative PCR and RT-PCR, yielding extended information on the presence and abundance of the diverse classes of viral nucleic acids, on the temporal regulation of genome expression and on its relationship with the cell cycle. The analysis performed indicate that the synthesis of viral nucleic acids is correlated to the progression through the S phase of the cell cycle, that an extended pattern of transcriptional activity occurs throughout the course of infection, with a maximal rate of transcription preceding the onset of S-phase dependent replication of the viral genome, and that utilization of transcript processing signals is relatively constant throughout the course of infection. The information obtained led to the definition of a unified model of functional and expression profiling of parvovirus B19 genome.

  10. 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

  11. ELITE S2 - A Facility for Quantitative Human Movement Analysis on Board the ISS

    NASA Astrophysics Data System (ADS)

    Neri, Gianluca; Mascetti, Gabriele; Zolesi, Valfredo

    2014-11-01

    This paper describes the activities for utilization and control of ELITE S2 on board the International Space Station (ISS). ELITE S2 is a payload of the Italian Space Agency (ASI) for quantitative human movement analysis in weightlessness. Within the frame of a bilateral agreement with NASA, ASI has funded a number of facilities, enabling different scientific experiments on board the ISS. ELITE S2 has been developed by the ASI contractor Kayser Italia, delivered to the Kennedy Space Center in 2006 for pre-flight processing, launched in 2007 by the Space Shuttle Endeavour (STS-118), integrated in the U.S. lab and used during the Increments 16/17 (2008) and 33/34 (2012/2013). The ELITE S2 flight segment comprises equipment mounted into an Express Rack and a number of stowed items to be deployed for experiment performance (video cameras and accessories). The ground segment consists in a User Support Operations Center (based at Kayser Italia) enabling real-time payload control and a number of User Home Bases (located at the ASI and PIs premises), for the scientific assessment of the experiment performance. Two scientific protocols on reaching and cognitive processing have been successfully performed in eight sessions involving three ISS crewmembers: IMAGINE 2 and MOVE.

  12. Integrating quantitative PCR and Bayesian statistics in quantifying human adenoviruses in small volumes of source water.

    PubMed

    Wu, Jianyong; Gronewold, Andrew D; Rodriguez, Roberto A; Stewart, Jill R; Sobsey, Mark D

    2014-02-01

    Rapid quantification of viral pathogens in drinking and recreational water can help reduce waterborne disease risks. For this purpose, samples in small volume (e.g. 1L) are favored because of the convenience of collection, transportation and processing. However, the results of viral analysis are often subject to uncertainty. To overcome this limitation, we propose an approach that integrates Bayesian statistics, efficient concentration methods, and quantitative PCR (qPCR) to quantify viral pathogens in water. Using this approach, we quantified human adenoviruses (HAdVs) in eighteen samples of source water collected from six drinking water treatment plants. HAdVs were found in seven samples. In the other eleven samples, HAdVs were not detected by qPCR, but might have existed based on Bayesian inference. Our integrated approach that quantifies uncertainty provides a better understanding than conventional assessments of potential risks to public health, particularly in cases when pathogens may present a threat but cannot be detected by traditional methods.

  13. Quantitative comparison of a human cancer cell surface proteome between interphase and mitosis.

    PubMed

    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

    2015-01-13

    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.

  14. 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.

  15. Oligopeptides of Chorionic Gonadotropin β-Subunit in Induction of T Cell Differentiation into Treg and Th17.

    PubMed

    Zamorina, S A; Shirshev, S V

    2015-11-01

    The role of oligopeptides of chorionic gonadotropin β-subunit (LQGV, AQGV, and VLPALP) in induction of differentiation into T-regulatory lymphocytes (Treg) and IL-17-producing lymphocytes (Th17) was studied in an in vitro system. Chorionic gonadotropin and oligopeptides promoted CD4(+) cell differentiation into functionally active Treg (FOXP3(+)GITR(+) and FOXP3(+)CTLA-4(+)), while chorionic gonadotropin and AQGV additionally stimulated IL-10 production by these cells. In parallel, chorionic gonadotropin and oligopeptides prevented CD4(+) cell differentiation into Th17 lymphocytes (ROR-gt(+)IL-17A(+)) and suppressed IL-17A secretion. Hence, oligopeptides of chorionic gonadotropin β-subunit promoted differentiation of CD4(+) cells into Treg and, in parallel, suppress Th17 induction, thus virtually completely reproducing the effects of the hormone, which opens new vista for their use in clinical practice.

  16. Detection of human herpesvirus 8 by quantitative polymerase chain reaction: development and standardisation of methods

    PubMed Central

    2012-01-01

    Background Human herpesvirus 8 (HHV-8), the aetiological agent of Kaposi’s sarcoma (KS), multicentric Castleman’s disease (MCD), and primary effusion lymphoma (PEL) is rare in Australia, but endemic in Sub-Saharan Africa, parts of South-east Asia and Oceania. While the treatment of external KS lesions can be monitored by clinical observation, the internal lesions of KS, MCD and PEL require extensive and expensive internal imaging, or autopsy. In patients with MCD and PEL, if HHV-8 viraemia is not reduced quickly, ~50% die within 24 months. HHV-8 qPCR is a valuable tool for monitoring HHV-8 viraemia, but is not available in many parts of the world, including those with high prevalence of KS and HHV-8. Methods A new molecular facility with stringent three-phase workflow was established, adhering to NPAAC and CLSI guidelines. Three fully validated quantitative assays were developed: two for detection and quantification of HHV-8; one for GAPDH, necessary for normalisation of viral loads in tissue and peripheral blood. Results The HHV-8 ORF73 and ORF26 qPCR assays were 100% specific. All qPCR assays, displayed a broad dynamic range (102 to 1010 copies/μL TE Buffer) with a limit of detection of 4.85x103, 5.61x102, and 2.59x102 copies/μL TE Buffer and a limit of quantification of 4.85x103, 3.01x102, and 1.38x102 copies/μL TE Buffer for HHV-8 ORF73, HHV-8 ORF26, and GAPDH respectively. The assays were tested on a panel of 35 KS biopsies from Queensland. All were HHV-8 qPCR positive with average viral load of 2.96x105 HHV-8 copies/μL DNA extract (range: 4.37x103 to 1.47x106 copies/μL DNA extract): When normalised these equate to an average viral load of 2.44x104 HHV-8 copies/103 cells (range: 2.20x102 to 7.38x105 HHV-8 copies/103 cells). Conclusions These are the first fully optimised, validated and MIQE compliant HHV-8 qPCR assays established in Australia. They worked well for qualitative detection of HHV-8 in archival tissue, and are well-suited for

  17. Development of an Alu-based, QSY 7-labeled primer PCR method for quantitation of human DNA in forensic samples.

    PubMed

    Nicklas, Janice A; Buel, Eric

    2003-03-01

    Determining the amount of human DNA extracted from a crime scene sample is an important step in DNA profiling. The forensic community relies almost entirely upon a technique (slot blot) to quantitate human DNA that is imprecise, time consuming, and labor intensive. This paper describes the development of a new technique based on PCR amplification of a repetitive Alu sequence. Specific primers were used to amplify a 124-bp fragment of Alu sequence; amplification was detected by SYBR Green I staining in a fluorescent plate reader. To reduce background in the plate reader assay, QSY-7 labeled primers were utilized. The assay was tested on animal DNAs, human blood spots, mock crime samples, and degraded DNA in comparison with the slot blot technique. The QSY Alu assay has a dynamic range of 10 ng to 10 pg, and is sensitive, specific, fast, quantitative, and comparable in cost to the slot blot assay.

  18. Development of an Alu-based, real-time PCR method for quantitation of human DNA in forensic samples.

    PubMed

    Nicklas, Janice A; Buel, Eric

    2003-09-01

    Determining the amount of human DNA extracted from a crime scene sample is an important step in DNA profiling. The forensic community relies almost entirely upon a technique (slot blot) to quantitate human DNA that is imprecise, time consuming, and labor intensive. We have previously described a method for quantitation of human DNA based on PCR amplification of a repetitive Alu sequence that uses a fluorescence plate reader. This manuscript describes and validates a variation of this assay using real-time PCR and SYBR Green I for quantitation. The advantages of the real-time assay over the plate reader assay are: reduced hands-on time, lower assay cost, and a greater dynamic range. The main disadvantage is the cost of the real-time instrument. However, for those forensic laboratories with access to a real-time instrument, this Alu-based assay has a dynamic range of 16 ng to 1 pg, is sensitive, specific, fast, quantitative, and uses only 2 microL of sample.

  19. 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.

  20. 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.

  1. Could bone tissue be a target for luteinizing hormone/chorionic gonadotropin?

    PubMed

    Mansell, Jason P; Bailey, Allen J; Yarram, Sarah J

    2007-04-15

    Ovariectomy (OVX) and Zoladex administration to adult rats gave conflicting results with respect to the excretion of total urinary hydroxyproline (OH-Pro), a valuable indicator of bone collagen catabolism. Whereas OVX culminated in early (1 week) increases in OH-Pro, the use of Zoladex actually lowered OH-Pro and showed no sign of increasing over controls for a 2-month period. Since both OVX and Zoladex produce a state of estrogen deficiency we reasoned that the differential effects of the two procedures on OH-Pro were attributed to LH status. Receptors for luteinizing hormone (LH)/human chorionic gonadotropin (hCG) have been identified in many, non-gonadal, estrogen sensitive sites and although bone is receptive to estrogen what effects LH/hCG might have upon bone metabolism have received scant attention. Treatment of osteoblasts in culture with a urinary derived formulation of hCG resulted in increased alkaline phosphatase (ALP) activity, raised matrix mettaloproteinase-2 (MMP-2) levels and increased expression of type I collagen. Further studies, using murine calvaria, supported a bone-resorbing effect of hCG. Taken together our initial findings suggested that raised hCG and/or LH might lead to an overall increase in bone matrix turnover as reported for puberty, pregnancy and the menopause. However, when the urinary derived preparation of hCG was replaced with recombinant hormone no changes in osteoblast activity were found implying the presence of contaminating agents in the urine derived hCG. Herein we describe that epidermal growth factor (EGF) could account for the changes observed for urinary derived hCG in osteoblast cultures and that the effects of LH/hCG on bone tissue are probably indirect.

  2. 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.

  3. Quantitation of Murine Stroma and Selective Purification of the Human Tumor Component of Patient-Derived Xenografts for Genomic Analysis

    PubMed Central

    Schneeberger, Valentina E.; Allaj, Viola; Gardner, Eric E.; 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

  4. 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…

  5. [Identification and quantitative determination of baclofen in human blood by HPLC with mass spectrometry detection].

    PubMed

    Dukova, O A; Kotlovsky, M Yu; Pokrovsky, A A; Suvorova, E V; Shivrina, T G; Krasnov, E A; Efremov, A A

    2016-03-01

    A method of identification and quantitative determination of baclofen in blood by HPLC with mass spectrometry detection has been developed. It is characterized by high sensitivity, specificity, linearity, accuracy, reproducibility, and a low detection for quantitative determination. The method has been used for diagnostics of acute baclofen poisoning in patients.

  6. Quantitative assessment of human whole blood RNA as a potential biomarker for infectious disease.

    PubMed

    Smith, Claire L; Dickinson, Paul; Forster, Thorsten; Khondoker, Mizanur; Craigon, Marie; Ross, Alan; Storm, Petter; Burgess, Stewart; Lacaze, Paul; Stenson, Benjamin J; Ghazal, Peter

    2007-12-01

    Infection remains a significant cause of morbidity and mortality especially in newborn infants. Analytical methods for diagnosing infection are severely limited in terms of sensitivity and specificity and require relatively large samples. It is proposed that stringent regulation of the human transcriptome affords a new molecular diagnostic approach based on measuring a highly specific systemic inflammatory response to infection, detectable at the RNA level. This proposition raises a number of as yet poorly characterised technical and biological variation issues that urgently need to be addressed. Here we report a quantitative assessment of methodological approaches for processing and extraction of RNA from small samples of infant whole blood and applying analysis of variation from biochip measurements. On the basis of testing and selection from a battery of assays we show that sufficient high quality RNA for analysis using multiplex array technology can be obtained from small neonatal samples. These findings formed the basis of implementing a set of robust clinical and experimental standard operating procedures for whole blood RNA samples from 58 infants. Modelling and analysis of variation between samples revealed significant sources of variation from the point of sample collection to processing and signal generation. These experiments further permitted power calculations to be run indicating the tractability and requirements of using changes in RNA expression profiles to detect different states between patient groups. Overall the results of our investigation provide an essential first step toward facilitating an alternative way for diagnosing infection from very small neonatal blood samples, providing methods and requirements for future chip-based studies.

  7. 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.

  8. Extrapolating cetacean densities to quantitatively assess human impacts on populations in the high seas.

    PubMed

    Mannocci, Laura; Roberts, Jason J; Miller, David L; Halpin, Patrick N

    2016-10-24

    As human activities expand beyond national jurisdictions to the high seas, there is increasing need to consider anthropogenic impacts to species that inhabit these waters. The current scarcity of scientific observations of cetaceans in the high seas impedes the assessment of population-level impacts of these activities. This study is directed towards an important management need in the high seas-the development of plausible density estimates to facilitate a quantitative assessment of anthropogenic impacts on cetacean populations in these waters. Our study region extends from a well-surveyed region within the United States Exclusive Economic Zone into a large region of the western North Atlantic sparsely surveyed for cetaceans. We modeled densities of 15 cetacean taxa using available line transect survey data and habitat covariates and extrapolated predictions to sparsely surveyed regions. We formulated models carefully to reduce the extent of extrapolation beyond covariate ranges, and constrained them to model simple and generalizable relationships. To evaluate confidence in the predictions, we performed several qualitative assessments, such as mapping where predictions were made outside sampled covariate ranges, and comparing them with maps of sightings from a variety of sources that could not be integrated into our models. Our study revealed a range of confidence levels for the model results depending on the taxon and geographic area, and highlights the need for additional surveying in environmentally distinct areas. Combined with their explicit confidence levels and necessary caution, our density estimates can inform a variety of management needs in the high seas, such as the quantification of potential cetacean interactions with military training exercises, shipping, fisheries, deep-sea mining, as well as delineation of areas of special biological significance in international waters. Our approach is generally applicable to other marine taxa and geographic

  9. 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

  10. 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

  11. 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

  12. The use of antibodies labelled with dyes for fast and simple assay of some human proteins by immunofiltration technique.

    PubMed

    Szewczuk, A; Kuropatwa, M; Rapak, A

    1992-01-01

    Immunofiltration technique with polyclonal and monoclonal antibodies for semi-quantitative assays of human albumin, chorionic gonadotropin, immunoglobulin G and transferrin was elaborated. An amount of antibody was immobolized in the form of 6 radially located small bars on a dry test filter made of glass microfibre sheet. The other amount of antibody, used in solution, was labelled with some dyes like commercial disperse dyes, colloidal elements, formazans and polypyrrole. Number of colour bars appearing on the test filter showed ranged of analyte concentration. Good results were obtained using antibodies labelled with colloidal gold, Disperse Red 11 and formazan from MTT. Assays with monoclonal antibodies were more sensitive than with polyclonal antibodies.

  13. Influence of water hardening of the chorion on cadmium accumulation in medaka (Oryzias latipes) eggs.

    PubMed

    González-Doncel, Miguel; Larrea, Maite; Sánchez-Fortún, Sebastián; Hinton, David E

    2003-07-01

    This report describes a study in which in vitro fertilization methods were used to expose medaka (Oryzias latipes) eggs to cadmium (Cd(2+)). This approach was applied to address the differential sensitivity and cumulative potential of Cd(2+) when exposure was initiated early (before fertilization and water hardening of the chorion) versus later during embryo development (i.e., well after the chorion has undergone water hardening). Following range finding exposures (2.5, 10, 20, 40 or 80 mg/l) under artificially controlled experimental procedures, results from hatching success and embryo malformations showed the earlier exposure interval more sensitive than the assay involving only the embryonated egg. Subsequent accumulation studies have shown that the exposure initiated before fertilization apparently led to more Cd(2+) deposition in the chorion compared to the exposure during embryonated stages of the eggs. Similarly, values for total Cd(2+) indicated higher concentrations in those eggs exposed prior to--and during--water hardening. Results suggest an alteration of the properties of the zona radiata in the early-stage eggs, making it more permeable to the potential exit or entrance of waterborne agents even after water hardening. Ongoing studies must now address the development of more realistic exposure conditions of the gametes by using incubation media with osmolarities similar to surface waters, and by shortening duration for gamete exposure. Also, sensitive methods to localize Cd(2+) and to delineate the transfer from the chorion to the embryo are needed.

  14. 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...

  15. The TissueNet v.2 database: A quantitative view of protein-protein interactions across human tissues

    PubMed Central

    Basha, Omer; Barshir, Ruth; Sharon, Moran; Lerman, Eugene; Kirson, Binyamin F.; Hekselman, Idan; Yeger-Lotem, Esti

    2017-01-01

    Knowledge of the molecular interactions of human proteins within tissues is important for identifying their tissue-specific roles and for shedding light on tissue phenotypes. However, many protein–protein interactions (PPIs) have no tissue-contexts. The TissueNet database bridges this gap by associating experimentally-identified PPIs with human tissues that were shown to express both pair-mates. Users can select a protein and a tissue, and obtain a network view of the query protein and its tissue-associated PPIs. TissueNet v.2 is an updated version of the TissueNet database previously featured in NAR. It includes over 40 human tissues profiled via RNA-sequencing or protein-based assays. Users can select their preferred expression data source and interactively set the expression threshold for determining tissue-association. The output of TissueNet v.2 emphasizes qualitative and quantitative features of query proteins and their PPIs. The tissue-specificity view highlights tissue-specific and globally-expressed proteins, and the quantitative view highlights proteins that were differentially expressed in the selected tissue relative to all other tissues. Together, these views allow users to quickly assess the unique versus global functionality of query proteins. Thus, TissueNet v.2 offers an extensive, quantitative and user-friendly interface to study the roles of human proteins across tissues. TissueNet v.2 is available at http://netbio.bgu.ac.il/tissuenet. PMID:27899616

  16. Quantitative T1, T2, and T2* Mapping and Semi-Quantitative Neuromelanin-Sensitive Magnetic Resonance Imaging of the Human Midbrain

    PubMed Central

    2016-01-01

    Purpose Neuromelanin is a dark pigment granule present within certain catecholamine neurons of the human brain. Here, we aimed to clarify the relationship between contrast of neuromelanin-sensitive magnetic resonance imaging (MRI) and MR relaxation times using T1, T2, and T2* mapping of the lower midbrain. Methods The subjects were 14 healthy volunteers (11 men and 3 women, mean age 29.9 ± 6.9 years). Neuromelanin-sensitive MRI was acquired using an optimized T1-weighted two-dimensional (2D)-turbo spin-echo sequence. To quantitatively evaluate the relaxation time, 2D-image data for the T1, T2, and T2* maps were also acquired. The regions of interest (substantia nigra pars compacta [SNc], superior cerebellar peduncles [SCP], cerebral peduncles [CP], and midbrain tegmentum [MT]) were manually drawn on neuromelanin-sensitive MRI to measure the contrast ratio (CR) and on relaxation maps to measure the relaxation times. Results The CR in the SNc was significantly higher than the CRs in the SCP and CP. Compared to the SCP and CP, the SNc had significantly higher T1 relaxation times. Moreover, the SNc had significantly lower T2 and T2* relaxation times than the other three regions (SCP, CP, and MT). Correlation analyses showed no significant correlations between the CRs in the SNc, SCP, and CP and each relaxation time. Conclusions We demonstrated the relationship between the CR of neuromelanin-sensitive MRI and the relaxation times of quantitative maps of the human midbrain. PMID:27768782

  17. Identification of the origin and localization of chorion (egg envelope) proteins in an ancient fish, the white sturgeon, Acipenser transmontanus.

    PubMed

    Murata, Kenji; Conte, Fred S; McInnis, Elizabeth; Fong, Tak Hou; Cherr, Gary N

    2014-06-01

    In many modern teleost fish, chorion (egg envelope) glycoproteins are synthesized in the liver of females, and the expression of those genes is controlled by endogenous estrogen released from the ovary during maturation. However, among the classical teleosts, such as salmonid, carp, and zebrafish, the chorion glycoproteins are synthesized in the oocyte, as in higher vertebrates. Sturgeon, which are members of the subclass Chondrostei, represent an ancient lineage of ray-finned fishes that differ from other teleosts in that their sperm possess acrosomes, their eggs have numerous micropyles, and early embryo development is similar to that of amphibians. In order to understand the molecular mechanisms of chorion formation and the phylogenetic relationship between sturgeon and other teleosts, we used specific antibodies directed against the primary components of sturgeon chorion glycoproteins, using immunoblotting and immunocytochemistry approaches. The origin of each chorion glycoprotein was determined through analyses of both liver and ovary, and their localization during ovarian development was investigated. Our data indicate that the origin of the major chorion glycoproteins of sturgeon, ChG1, ChG2, and ChG4, derive not only from the oocyte itself but also from follicle cells in the ovary, as well as from hepatocytes. In the follicle cell layer, granulosa cells were found to be the primary source of ChGs during oogenesis in white sturgeon. The unique origins of chorion glycoproteins in sturgeon suggest that sturgeons are an intermediate form in the evolution of the teleost lineage.

  18. 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.

  19. Evaluation of the treatment of human Schistosoma mansoni infection by the quantitative oogram technique*

    PubMed Central

    Cançado, J. Romeu; da Cunha, A. Sales; de Carvalho, D. Garcia; Cambraia, J. N. Santos

    1965-01-01

    Egg output is the only measure available for quantitative assessment of the activity of chemotherapeutic agents in Schistosoma mansoni infection. In the light of eight years' experience in the preparations of oograms, the authors suggest a simplified classification of S. mansoni eggs and certain improvements in the oogram technique by which quantitative data are obtained for comparison before and after treatment. Ten cases, taken from clinical trials on a variety of schistosomicidal compounds, are presented to illustrate the use of the quantitative oogram and the types of result obtained with active, partially active and inactive drugs. ImagesFIG. 3FIG. 2FIG. 1 PMID:5323117

  20. 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.

  1. TeratoScore: Assessing the Differentiation Potential of Human Pluripotent Stem Cells by Quantitative Expression Analysis of Teratomas

    PubMed Central

    Avior, Yishai; Biancotti, Juan Carlos; Benvenisty, Nissim

    2015-01-01

    Summary 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

  2. 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

  3. 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

  4. 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

  5. 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

  6. 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

  7. Qualitative and quantitative detection of human pathogenic Yersinia enterocolitica in different food matrices at retail level in Bavaria.

    PubMed

    Messelhäusser, Ute; Kämpf, Peter; Colditz, Janine; Bauer, Hans; Schreiner, Hermann; Höller, Christiane; Busch, Ulrich

    2011-01-01

    Yersinia enterocolitica is a major foodborne pathogen and the third most important bacteriological cause of diarrhea in Germany. However, studies investigating the occurrence of human pathogenic Y. enterocolitica in food at the retail level are very rare. Most of the studies published so far show qualitative but not quantitative data concerning the prevalence of this human pathogen. In this study the qualitative and quantitative assessment of human pathogenic Y. enterocolitica in different food matrices was investigated. For the qualitative analysis we used an enrichment method according to the International Organisation of Standardization (ISO) standard in combination with a real-time polymerase chain reaction (PCR) method detecting the ail gene of Y. enterocolitica. After detecting Y. enterocolitica in a sample, a quantitative investigation on Cefsulodin-Irgasan-Novobiocin (CIN) Agar was done to get information about the contamination level of the different samples. During the years 2008 and 2009, 446 samples of pork and pork products, 51 samples of game meat, and 61 raw milk samples were investigated for the presence of human pathogenic Y. enterocolitica. The samples were collected at the retail level in Bavaria. From the pork samples investigated, 81 samples (18%) were positive for the ail gene by real-time PCR, but human pathogenic Y. enterocolitica O:3 were found only in 46 (10%) pork samples by culture; the concentration in the samples ranged between 0.04 cfu/g and 2.30 × 10(5) cfu/g. Three game meat samples were positive by real-time PCR, but not by the cultural detection. All raw milk samples were negative by real-time PCR and culture.

  8. 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.

  9. 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.

  10. 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.

  11. 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…

  12. Inhibitory Activity of Human Immunodeficiency Virus Aspartyl Protease Inhibitors against Encephalitozoon intestinalis Evaluated by Cell Culture-Quantitative PCR Assay

    PubMed Central

    Menotti, Jean; Santillana-Hayat, Maud; Cassinat, Bruno; Sarfati, Claudine; Derouin, Francis; Molina, Jean-Michel

    2005-01-01

    Immune reconstitution might not be the only factor contributing to the low prevalence of microsporidiosis in human immunodeficiency virus (HIV)-infected patients treated with protease inhibitors, as these drugs may exert a direct inhibitory effect against fungi and protozoa. In this study, we developed a cell culture-quantitative PCR assay to quantify Encephalitozoon intestinalis growth in U-373-MG human glioblastoma cells and used this assay to evaluate the activities of six HIV aspartyl protease inhibitors against E. intestinalis. A real-time quantitative PCR assay targeted the E. intestinalis small-subunit rRNA gene. HIV aspartyl protease inhibitors were tested over serial concentrations ranging from 0.2 to 10 mg/liter, with albendazole used as a control. Ritonavir, lopinavir, and saquinavir were able to inhibit E. intestinalis growth, with 50% inhibitory concentrations of 1.5, 2.2, and 4.6 mg/liter, respectively, whereas amprenavir, indinavir, and nelfinavir had no inhibitory effect. Pepstatin A, a reference aspartyl protease inhibitor, could also inhibit E. intestinalis growth, suggesting that HIV protease inhibitors may act through the inhibition of an E. intestinalis-encoded aspartyl protease. These results showed that some HIV protease inhibitors can inhibit E. intestinalis growth at concentrations that are achievable in vivo and that the real-time quantitative PCR assay that we used is a valuable tool for the in vitro assessment of the activities of drugs against E. intestinalis. PMID:15917534

  13. 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

  14. The first comprehensive and quantitative analysis of human platelet protein composition allows the comparative analysis of structural and functional pathways.

    PubMed

    Burkhart, Julia M; Vaudel, Marc; Gambaryan, Stepan; Radau, Sonja; Walter, Ulrich; Martens, Lennart; Geiger, Jörg; Sickmann, Albert; Zahedi, René P

    2012-10-11

    Antiplatelet treatment is of fundamental importance in combatting functions/dysfunction of platelets in the pathogenesis of cardiovascular and inflammatory diseases. Dysfunction of anucleate platelets is likely to be completely attributable to alterations in posttranslational modifications and protein expression. We therefore examined the proteome of platelets highly purified from fresh blood donations, using elaborate protocols to ensure negligible contamination by leukocytes, erythrocytes, and plasma. Using quantitative mass spectrometry, we created the first comprehensive and quantitative human platelet proteome, comprising almost 4000 unique proteins, estimated copy numbers for ∼ 3700 of those, and assessed intersubject (4 donors) as well as intrasubject (3 different blood samples from 1 donor) variations of the proteome. For the first time, our data allow for a systematic and weighted appraisal of protein networks and pathways in human platelets, and indicate the feasibility of differential and comprehensive proteome analyses from small blood donations. Because 85% of the platelet proteome shows no variation between healthy donors, this study represents the starting point for disease-oriented platelet proteomics. In the near future, comprehensive and quantitative comparisons between normal and well-defined dysfunctional platelets, or between platelets obtained from donors at various stages of chronic cardiovascular and inflammatory diseases will be feasible.

  15. [Quantitative image analysis in pulmonary pathology - digitalization of preneoplastic lesions in human bronchial epithelium (author's transl)].

    PubMed

    Steinbach, T; Müller, K M; Kämper, H

    1979-01-01

    The report concerns the first phase of a quantitative study of normal and abnormal bronchial epithelium with the objective of establishing the digitalization of histologic patterns. Preparative methods, data collecting and handling, and further mathematical analysis are described. In cluster and discriminatory analysis the digitalized histologic features can be used to separate and classify the individual cases into the respective diagnostic groups.

  16. Microscopic and biochemical analysis of the viability and permeability of guinea pig amnion and chorion leave in vitro.

    PubMed

    Goldhawk, D E; Carter, D; Hobkirk, R

    1996-08-01

    Tissue viability and permeability of guinea pig amnion and chorion leave were analyzed microscopically and biochemically. The vital dyes T1111 and fluorescein diacetate were used to locate and determine the integrity of cell plasma membranes in early and late tissue in vitro using confocal laser scanning microscopy and scanning electron microscopy. Early amnion and chorion laeve were each found to contain a single epithelial cell layer, composed of membrane-intact cells. In contrast, plasma membrane lesions were present throughout the epithelium of late amnion. Late chorion laeve contained both regions of intact and damaged epithelial cells on its maternal side. There was also a layer of membrane-intact squamous cells on the fetal side of late chorion laeve. ATP measurements confirmed that early fetal membranes were viable after incubation in isotonic salt solutions at physiological pH. Late amnion was depleted of ATP stores while late chorion laeve retained its capacity for generating energy. These viability markers indicate that late guinea pig amnion is not a viable tissue in vitro, while late chorion laeve is a viable but probably degenerating tissue. Confocal X-Z scans were used to trace the movement of T1111 through the tissue as an indication of permeability to free solutes. Whereas dye will permeate across the main thickness of early amnion and chorion leave, it did not pass between cells, but was blocked, presumably by a line of tight junctions. Late amnion was characterized by the complete permeability to T1111. Late chorion leave contained regions where solute migration was blocked, but overall was a permeable tissue. These results provide an important context for the interpretation of molecular movement across fetal membranes.

  17. Quantitation of berberine chloride in human urine by use of selected ion monitoring in the field desorption mode.

    PubMed

    Miyazaki, H; Shirai, E; Ishibashi, M; Hosoi, K; Shibata, S; Iwanaga, M

    1978-10-01

    A method is described for the microdetermination of berberine chloride in human urine by a field desorption mass spectrometry selected ion monitoring system using a deuterium labelled analogue of berberine chloride as an internal standard. Prior to the quantitation of berberine in human urine, the fundamental problems related to field desorption selected ion monitoring, such as quality of emitters, amounts of sample loading, and the programming rate of the emitter current, were statistically investigated in detail. Berberine chloride can be determined in a concentration of 10 ng ml-1 in human urine by the method described. The analytical results were compared with those from gas chromatography mass spectrometry selected ion monitoring in the chemical ionization mode suggesting that the reliability of field desorption selected ion monitoring may be almost equivalent to that of gas chromatography chemical ionization selected ion monitoring.

  18. Severe pre-eclampsia is associated with alterations in cytotrophoblasts of the smooth chorion

    PubMed Central

    Garrido-Gomez, Tamara; Ona, Katherine; Kapidzic, Mirhan; Gormley, Matthew; Simón, Carlos; Genbacev, Olga

    2017-01-01

    Pre-eclampsia (PE), which affects ∼8% of first pregnancies, is associated with faulty placentation. Extravillous cytotrophoblasts (CTBs) fail to differentiate properly, contributing to shallow uterine invasion and deficient spiral artery remodeling. We studied the effects of severe PE (sPE) on the smooth chorion portion of the fetal membranes. The results showed a significant expansion of the CTB layer. The cells displayed enhanced expression of stage-specific antigens that extravillous CTBs normally upregulate as they exit the placenta. Transcriptomics revealed the dysregulated expression of many genes (e.g. placental proteins, markers of oxidative stress). We confirmed an sPE-related increase in production of PAPPA1, which releases IGF1 from its binding protein. IGF1 enhanced proliferation of smooth chorion CTBs, a possible explanation for expansion of this layer, which may partially compensate for the placental deficits. PMID:28232601

  19. Quantitative determination of 25-hydroxyvitamin D3 3-sulphate in human plasma using high performance liquid chromatography.

    PubMed

    Shimada, K; Mitamura, K; Kitama, N

    1995-01-01

    The quantitative determination of 25-hydroxyvitamin D3 3-sulphate in human plasma was completed using reversed phase high performance liquid chromatography with UV detection (265 nm) and an internal standard method. The vitamin D sulphate fraction was obtained from a plasma specimen with the combined use of a Bond Elut C18 cartridge for solid-phase extraction and a piperidinohydroxypropyl Sephadex LH-20 column for lipophilic ion-exchange chromatography. Separation of the compounds was performed on a YMC-Pack ODS-AM column. The limit of quantitation was 5 ng/mL and the assay was linear from 5 to 50 ng/mL. The proposed method is satisfactory in its accuracy and precision.

  20. Fast chiral chromatographic method development and validation for the quantitation of eszopiclone in human plasma using LC/MS/MS.

    PubMed

    Meng, Min; Rohde, Lisa; Cápka, Vladimír; Carter, Spencer J; Bennett, Patrick K

    2010-12-01

    Traditional chiral chromatographic separation method development is time consuming even for an experienced chromatographer. This paper describes the application of computer software ACD Lab to facilitate the development of chiral separation for the quantitation of eszopiclone using LC-MS/MS technology. Assisted by ACD/Chrom Manager and LC Simulator software, the optimal chiral chromatographic development was completed within hours. The baseline chiral separation was achieved with a total cycle time of 3 min. For sample extraction method development, a Waters Oasis Sorbent Selection Plate containing four different sorbents was utilized. Optimal conditions were determined using a single plate under various load, wash and elution conditions. This was followed by a GLP validation which demonstrated excellent intra- and inter-day accuracy and precision for the quantitation of eszopiclone in human plasma at 1.00-100 ng/mL range using LC/MS/MS technology. This method was utilized to support multiple clinic bioequivalence studies.

  1. Quantitative analysis of modified proteins and their positional isomers by tandem mass spectrometry: human histone H4.

    PubMed

    Pesavento, James J; Mizzen, Craig A; Kelleher, Neil L

    2006-07-01

    Here we show that fragment ion abundances from dissociation of ions created from mixtures of multiply modified histone H4 (11 kDa) or of N-terminal synthetic peptides (2 kDa) correspond to their respective intact ion abundances measured by Fourier transform mass spectrometry. Isomeric mixtures of modified forms of the same protein are resolved and quantitated with a precision of human proteins to derive quantitative information on the highly related and often isomeric protein forms created by combinatorial arrays of posttranslational modifications.

  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. MicroRNA125b-mediated Hedgehog signaling influences liver regeneration by chorionic plate-derived mesenchymal stem cells.

    PubMed

    Hyun, Jeongeun; Wang, Sihyung; Kim, Jieun; Kim, Gi Jin; Jung, Youngmi

    2015-09-15

    Although chorionic plate-derived mesenchymal stem cells (CP-MSCs) were shown to promote liver regeneration, the mechanisms underlying the effect remain unclear. Hedgehog (Hh) signaling orchestrates tissue reconstruction in damaged liver. MSCs release microRNAs mediating various cellular responses. Hence, we hypothesized that microRNAs from CP-MSCs regulated Hh signaling, which influenced liver regeneration. Livers were obtained from carbon tetrachloride (CCl4)-treated rats transplanted with human CP-MSCs (Tx) or saline (non-Tx). Sonic Hh, one of Hh ligands, increased in CCl4-treated liver, whereas it decreased in CP-MSC-treated liver with CCl4. The expression of Hh-target genes was significantly downregulated in the Tx. Reduced expansion of progenitors and regressed fibrosis were observed in the liver of the Tx rats. CP-MSCs suppressed the expression of Hh and profibrotic genes in co-cultured LX2 (human hepatic stellate cell) with CP-MSCs. MicroRNA-125b targeting smo was retained in exosomes of CP-MSCs. CP-MSCs with microRNA-125b inhibitor failed to attenuate the expression of Hh signaling and profibrotic genes in the activated HSCs. Therefore, these results demonstrated that microRNA-125b from CP-MSCs suppressed the activation of Hh signaling, which promoted the reduced fibrosis, suggesting that microRNA-mediated regulation of Hh signaling contributed to liver regeneration by CP-MSCs.

  4. High-resolution multi-parametric quantitative magnetic resonance imaging of the human cervical spinal cord at 7T.

    PubMed

    Massire, Aurélien; Taso, Manuel; Besson, Pierre; Guye, Maxime; Ranjeva, Jean-Philippe; Callot, Virginie

    2016-12-01

    Quantitative MRI techniques have the potential to characterize spinal cord tissue impairments occurring in various pathologies, from both microstructural and functional perspectives. By enabling very high image resolution and enhanced tissue contrast, ultra-high field imaging may offer further opportunities for such characterization. In this study, a multi-parametric high-resolution quantitative MRI protocol is proposed to characterize in vivo the human cervical spinal cord at 7T. Multi-parametric quantitative MRI acquizitions including T1, T2(*) relaxometry mapping and axial diffusion MRI were performed on ten healthy volunteers with a whole-body 7T system using a commercial prototype coil-array dedicated to cervical spinal cord imaging. Automatic cord segmentation and multi-parametric data registration to spinal cord templates enabled robust regional studies within atlas-based WM tracts and GM horns at the C3 cervical level. T1 value, cross-sectional area and GM/WM ratio evolutions along the cervical cord were also reported. An original correction method for B1(+)-biased T1 mapping sequence was additionally proposed and validated on phantom. As a result, relaxometry and diffusion parameters derived from high-resolution quantitative MRI acquizitions were reported at 7T for the first time. Obtained images, with unmatched resolutions compared to lower field investigations, provided exquisite anatomical details and clear delineation of the spinal cord substructures within an acquisition time of 30min, compatible with clinical investigations. Regional statistically significant differences were highlighted between WM and GM based on T1 and T2* maps (p<10(-3)), as well as between sensory and motor tracts based on diffusion tensor imaging maps (p<0.05). The proposed protocol demonstrates that ultra-high field spinal cord high-resolution quantitative MRI is feasible and lays the groundwork for future clinical investigations of degenerative spinal cord pathologies.

  5. Quantitative real-time PCR (qPCR) assay for human-dog-cat species identification and nuclear DNA quantification.

    PubMed

    Kanthaswamy, S; Premasuthan, A; Ng, J; Satkoski, J; Goyal, V

    2012-03-01

    In the United States, human forensic evidence collected from crime scenes is usually comingled with biomaterial of canine and feline origins. Knowledge of the concentration of nuclear DNA extracted from a crime scene biological sample and the species from which the sample originated is essential for DNA profiling. The ability to accurately detect and quantify target DNA in mixed-species samples is crucial when target DNA may be overwhelmed by non-target DNA. We have designed and evaluated a species-specific (human, dog and cat) nuclear DNA identification assay based on the TaqMan(®) quantitative real-time PCR (qPCR) technology that can simultaneously detect and measure minute quantities of DNA specific to either humans, dogs and/or cats. The fluorogenic triplex assay employs primers and hydrolysis probes that target the human TH01 locus as well as the dog and cat Melanocortin 1 Receptor (MC1R) sequences in a species-specific manner. We also demonstrate that the assay is a highly sensitive, reliable and robust method for identifying and quantifying mixed-species templates of human-dog-cat origin with as little as 0.4 pg of human and cat nuclear DNA, respectively, and 4.0 pg of dog nuclear DNA.

  6. 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...

  7. Detection of Chromosome Aneuploidies in Chorionic Villus Samples by Multiplex Ligation-Dependent Probe Amplification

    PubMed Central

    Kooper, Angelique J.A.; Faas, Brigitte H.W.; Feuth, Ton; Creemers, Johan W.T.; Zondervan, Hans H.; Boekkooi, Peter F.; Quartero, Rik W.P.; Rijnders, Robbert J.P.; van der Burgt, Ineke; van Kessel, Ad Geurts; Smits, Arie P.T.

    2009-01-01

    The objective of this study was to examine the suitability of multiplex ligation-dependent probe amplification (MLPA) in chorionic villus samples as a replacement for traditional karyotyping for the detection of (an)euploidies of chromosomes 21, 18, 13, X, and Y. Chorionic villus samples were diagnosed by traditional karyotyping using short-term cultures (STC) and long-term cultures (LTC), and by MLPA using kit P095. DNA was extracted after digestion of whole villi with proteinase K and/or trypsin and collagenase. Different cell-dissociation procedures were tested to obtain MLPA results representative of the cytotrophoblast layer and the mesenchymal core. Over 95% of the MLPA results were in concordance with the traditional karyotyping of STC and LTC. Traditional karyotyping revealed seven mosaics. After digestion of whole villi with proteinase K, only abnormal cell lines confined to the STC gave rise to abnormal MLPA results. In one sample, the complete discrepancy between STC and LTC was resolved after enzymatic dissociation of cells from the cytotrophoblast layer and the mesenchymal core. MLPA in chorionic villus samples was found to be a reliable test for the detection of (an)euploidies of chromosomes 21, 18, 13, X, and Y. Whole villi digestion with proteinase K resulted in the over-representation of cytotrophoblasts in the DNA pool. To obtain MLPA results representative for STC and LTC, enzymatic dissociation of cells from the cytotrophoblast layer and mesenchymal core is required. PMID:19074591

  8. Quantitative analysis of THC, 11-OH-THC, and THCCOOH in human hair by negative ion chemical ionization mass spectrometry.

    PubMed

    Wilkins, D; Haughey, H; Cone, E; Huestis, M; Foltz, R; Rollins, D

    1995-10-01

    A sensitive and specific method was developed for the quantitative analysis of delta9-tetrahydrocannabinol (THC), 11 -hydroxy-THC (11-OH-THC), and 11-nor-9-carboxy-THC (THCCOOH) in human hair. Deuterated internal standards were added to hair samples, and samples were digested overnight in 1 N NaOH at 37 degrees C. Digest solutions were extracted with a liquid-liquid extraction procedure, which was previously developed in our laboratory for the analysis of plasma and whole blood. Derivatized extracts were analyzed on a Finnigan 4500" mass spectrometer in negative ion chemical ionization mode using methane as the reagent gas, hydrogen as the carrier gas, and a Restek Rtx 200-15M-0.25-microm capillary column. The assay was linear up to 50 ng/mg hair (r, 0.99) for all three compounds and was capable of detecting 10 pg THC and THCCOOH and 100 pg 11-OH-THC on column. The intra-assay precision ranged from 2.1 to 11.2% for the three analytes; the interassay precision ranged from 4.4 to 13.0%. The method was used to detect and quantitate the presence of THC, 11-OH-THC, and THCCOOH in human hair obtained from eight regular users of cannabis. THC, but not 11-OH-THC or THCCOOH, was detectable in the hair shaft above the assay limit of quantitation. Four laboratory wash procedures were also evaluated for their effect on the measured concentration of THC in hair. in seven of eight subjects, a methylene chloride wash procedure substantially reduced the measured THC concentration by up to 50%. The gas chromatographic-mass spectrometric assay is currently being used to support pharmacokinetic studies of drug disposition into the hair of humans and animals.

  9. Bibliometric analysis of Human Factors (1970-2000): a quantitative description of scientific impact.

    PubMed

    Dee, John D; Cassano-Pinché, Andrea; Vicente, Kim J

    2005-01-01

    Bibliometric analyses use the citation history of scientific articles as data to measure scientific impact. This paper describes a bibliometric analysis of the 1682 papers and 2413 authors published in Human Factors from 1970 to 2000. The results show that Human Factors has substantial relative scientific influence, as measured by impact, immediacy, and half-life, exceeding the influence of comparable journals. Like other scientific disciplines, human factors research is a highly stratified activity. Most authors have published only one paper, and many papers are cited infrequently, if ever. A small number of authors account for a disproportionately large number of the papers published and citations received. However, the degree of stratification is not as extreme as in many other disciplines, possibly reflecting the diversity of the human factors discipline. A consistent trend of more authors per paper parallels a similar trend in other fields and may reflect the increasingly interdisciplinary nature of human factors research and a trend toward addressing human-technology interaction in more complex systems. Ten of the most influential papers from each of the last 3 decades illustrate trends in human factors research. Actual or potential applications of this research include considerations for the publication and distribution policy of Human Factors.

  10. 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...

  11. In vivo myogenic potential of human CD133+ muscle-derived stem cells: a quantitative study.

    PubMed

    Negroni, Elisa; Riederer, Ingo; Chaouch, Soraya; Belicchi, Marzia; Razini, Paola; Di Santo, James; Torrente, Yvan; Butler-Browne, Gillian S; Mouly, Vincent

    2009-10-01

    In recent years, numerous reports have identified in mouse different sources of myogenic cells distinct from satellite cells that exhibited a variable myogenic potential in vivo. Myogenic stem cells have also been described in humans, although their regenerative potential has rarely been quantified. In this study, we have investigated the myogenic potential of human muscle-derived cells based on the expression of the stem cell marker CD133 as compared to bona fide satellite cells already used in clinical trials. The efficiency of these cells to participate in muscle regeneration and contribute to the renewal of the satellite cell pool, when injected intramuscularly, has been evaluated in the Rag2(-/-) gammaC(-/-) C5(-/-) mouse in which muscle degeneration is induced by cryoinjury. We demonstrate that human muscle-derived CD133+ cells showed a much greater regenerative capacity when compared to human myoblasts. The number of fibers expressing human proteins and the number of human cells in a satellite cell position are all dramatically increased when compared to those observed after injection of human myoblasts. In addition, CD133+/CD34+ cells exhibited a better dispersion in the host muscle when compared to human myoblasts. We propose that muscle-derived CD133+ cells could be an attractive candidate for cellular therapy.

  12. In Vivo Myogenic Potential of Human CD133+ Muscle-derived Stem Cells: A Quantitative Study

    PubMed Central

    Negroni, Elisa; Riederer, Ingo; Chaouch, Soraya; Belicchi, Marzia; Razini, Paola; Di Santo, James; Torrente, Yvan; Butler-Browne, Gillian S; Mouly, Vincent

    2009-01-01

    In recent years, numerous reports have identified in mouse different sources of myogenic cells distinct from satellite cells that exhibited a variable myogenic potential in vivo. Myogenic stem cells have also been described in humans, although their regenerative potential has rarely been quantified. In this study, we have investigated the myogenic potential of human muscle–derived cells based on the expression of the stem cell marker CD133 as compared to bona fide satellite cells already used in clinical trials. The efficiency of these cells to participate in muscle regeneration and contribute to the renewal of the satellite cell pool, when injected intramuscularly, has been evaluated in the Rag2−/− γC−/− C5−/− mouse in which muscle degeneration is induced by cryoinjury. We demonstrate that human muscle–derived CD133+ cells showed a much greater regenerative capacity when compared to human myoblasts. The number of fibers expressing human proteins and the number of human cells in a satellite cell position are all dramatically increased when compared to those observed after injection of human myoblasts. In addition, CD133+/CD34+ cells exhibited a better dispersion in the host muscle when compared to human myoblasts. We propose that muscle-derived CD133+ cells could be an attractive candidate for cellular therapy. PMID:19623164

  13. Quantitative Evaluation of Dichloroacetic Acid Kinetics in Human -- A Physiologically-Based Pharmacokinetic Modeling Investigation

    DTIC Science & Technology

    2008-01-01

    use include mild liver dysfunction, transient central neuropathy , peripheral neuropathy and hypocalcemia. The clinical effects are generally...metabolite of trichloroethylene. DCA liver carcinogenicity has been demonstrated in rodents but epidemiological evidence in humans is not available. High...probable minor metabolite of trichloroethylene. DCA liver carcinogenicity has been demonstrated in rodents but epidemiological evidence in humans is not

  14. Human genome-specific real-time PCR method for sensitive detection and reproducible quantitation of human cells in mice.

    PubMed

    Song, Pengyue; Xie, Zhenhua; Guo, Ling; Wang, Chengmei; Xie, Weidong; Wu, Yaojiong

    2012-12-01

    Xenotransplantation of human cells into immunodeficiency mice has been frequently used to study stem cells in tissue repair and regeneration and cancer cell metastasis. However, a sensitive and reproducible method to quantify cell engraftment lacks. Here, we developed a Real-Time PCR-based method which facilitated consistent detection and quantification of small amounts of human cells distributed in mouse organs after infusion. The principle of the method was to directly detect a humans-specific sequence in the human-murine genomic DNA mixture. In a mouse myocardial infarction model, the Real-Time PCR-based method consistently determined the amounts of human mesenchymal stem cells (hMSCs) engrafted into the heart and other organs 7 days after infusion of as little as 2.5 × 10(5) cells, indicating a high sensitivity, and the amounts of hMSCs detected in mice highly correlated to the numbers of hMSCs transplanted. Importantly, different from previous PCR-based methods, our method produced highly consistent and reproducible results. The reliability of the method was further proven by parallel analyses of DiI-labeled hMSCs in tissue sections and in single cell suspensions of mice. Our data show that the present human genomic DNA-specific primers-based Real-Time PCR method is sensitive and highly reproducible in determining the amount of xenotransplanted human cells in murine tissues.

  15. 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.

  16. 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.

  17. A Rapid, Quantitative Method to Characterize The Human Lymphocyte Concentration for Automated High-Throughput Radiation Biodosimetry

    PubMed Central

    Xu, Yanping; Turner, Helen C.; Garty, Guy; Brenner, David

    2013-01-01

    We have developed a Quantitative Light Absorption Analysis (QLAA) method to rapidly estimate human lymphocyte concentrations isolated from small volumes of whole blood. Measurements of the light absorption analysis were calibrated for lymphocyte concentration levels using a hemocytometer. To validate the QLAA system, blood samples were collected from 17 healthy donors and lymphocyte absorption measurements were directly compared with the manual microscope counting. The results showed that lymphocyte measurements obtained using the QLAA system were comparable with the manually scored lymphocyte counts but with measurements taken in seconds. PMID:23781493

  18. Quantitative EEG parameters correlate with the progression of human prion diseases

    PubMed Central

    Wehner, Tim; Lowe, Jessica; Porter, Marie-Claire; Kenny, Joanna; Thompson, Andrew; Rudge, Peter; Collinge, John; Mead, Simon

    2016-01-01

    Background Prion diseases are universally fatal and often rapidly progressive neurodegenerative diseases. EEG has long been used in the diagnosis of sporadic Creutzfeldt-Jakob disease; however, the characteristic waveforms do not occur in all types of prion diseases. Here, we re-evaluate the utility of EEG by focusing on the development of biomarkers. We test whether abnormal quantitative EEG parameters can be used to measure disease progression in prion diseases or predict disease onset in healthy individuals at risk of disease. Methods In the National Prion Monitoring Cohort study, we did quantitative encephalography on 301 occasions in 29 healthy controls and 67 patients with prion disease. The patients had either inherited prion disease or sporadic Creutzfeldt-Jakob disease. We computed the main background frequency, the α and θ power and the α/θ power ratio, then averaged these within 5 electrode groups. These measurements were then compared among participant groups and correlated with functional and cognitive scores cross-sectionally and longitudinally. Results We found lower main background frequency, α power and α/θ power ratio and higher θ power in patients compared to control participants. The main background frequency, the power in the α band and the α/θ power ratio also differed in a consistent way among the patient groups. Moreover, the main background frequency and the α/θ power ratio correlated significantly with functional and cognitive scores. Longitudinally, change in these parameters also showed significant correlation with the change in clinical and cognitive scores. Conclusions Our findings support the use of quantitative EEG to follow the progression of prion disease, with potential to help evaluate the treatment effects in future clinical-trials. PMID:27413165

  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. 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...

  1. Quantitative bioanalysis of strontium in human serum by inductively coupled plasma-mass spectrometry

    PubMed Central

    Somarouthu, Srikanth; Ohh, Jayoung; Shaked, Jonathan; Cunico, Robert L; Yakatan, Gerald; Corritori, Suzana; Tami, Joe; Foehr, Erik D

    2015-01-01

    Aim: A bioanalytical method using inductively-coupled plasma-mass spectrometry to measure endogenous levels of strontium in human serum was developed and validated. Results & methodology: This article details the experimental procedures used for the method development and validation thus demonstrating the application of the inductively-coupled plasma-mass spectrometry method for quantification of strontium in human serum samples. The assay was validated for specificity, linearity, accuracy, precision, recovery and stability. Significant endogenous levels of strontium are present in human serum samples ranging from 19 to 96 ng/ml with a mean of 34.6 ± 15.2 ng/ml (SD). Discussion & conclusion: Calibration procedures and sample pretreatment were simplified for high throughput analysis. The validation demonstrates that the method was sensitive, selective for quantification of strontium (88Sr) and is suitable for routine clinical testing of strontium in human serum samples. PMID:28031925

  2. The primary transcription unit of the human alpha 2 globin gene defined by quantitative RT/PCR.

    PubMed Central

    Owczarek, C M; Enriquez-Harris, P; Proudfoot, N J

    1992-01-01

    We have set up an experimental system to map the primary transcription unit of the human alpha 2 globin gene. The duplicated human alpha globin genes (alpha 2-alpha 1) were linked to the alpha globin locus Positive Regulatory Element (PRE) and stably transfected into murine erythroleukaemia cells. We then developed a quantitative reverse transcriptase, polymerase chain reaction assay to map alpha 2 primary transcripts using primer pairs derived from different parts of the alpha 2 globin gene and its 3' flanking region. This approach has revealed the presence of steady state nuclear RNA past the poly(A) site of the alpha 2 globin gene at approximately 40% of the level of unspliced intron transcript. Furthermore, these 3' flanking transcripts diminish 500 bp into the 3' flanking region, identifying this part of the alpha 2 globin gene as the principal region of termination of transcription. Images PMID:1371868

  3. Quantitative keratinocyte assay detects two biological activities of human papillomavirus DNA and identifies viral types associated with cervical carcinoma.

    PubMed Central

    Schlegel, R; Phelps, W C; Zhang, Y L; Barbosa, M

    1988-01-01

    Keratinocytes electroporated with human papillomavirus (HPV) DNA (HPV-6, 11, 16 and 18) exhibited an increased cellular proliferation which was quantitated as microcolony and macrocolony formation. However, only macrocolonies induced by HPV-16 or HPV-18 DNA (the two viral types most commonly found in human cervical carcinomas) gave rise to proliferating, poorly-stratified colonies when grown in the presence of serum and calcium. Hydrocortisone increased the frequency of these differentiation-resistant colonies, and studies showed that they were immortalized, contained one copy of viral DNA per cell, expressed three discrete species of viral RNA and synthesized the viral E7 protein. HPV-induced cellular proliferation and altered differentiation are therefore separable events and may represent the activity of different viral genes. Images PMID:2460337

  4. 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.

  5. Multiplexed Quantitative Proteomics for High-Throughput Comprehensive Proteome Comparisons of Human Cell Lines.

    PubMed

    Edwards, Amanda; Haas, Wilhelm

    2016-01-01

    The proteome is the functional entity of the cell, and perturbations of a cellular system almost always cause changes in the proteome. These changes are a molecular fingerprint, allowing characterization and a greater understanding of the effect of the perturbation on the cell as a whole. Monitoring these changes has therefore given great insight into cellular responses to stress and disease states, and analytical platforms to comprehensively analyze the proteome are thus extremely important tools in biological research. Mass spectrometry has evolved as the most relevant technology to characterize proteomes in a comprehensive way. However, due to a lack of throughput capacity of mass spectrometry-based proteomics, researchers frequently use measurement of mRNA levels to approximate proteome changes. Growing evidence of substantial differences between mRNA and protein levels as well as recent improvements in mass spectrometry-based proteomics are heralding an increased use of mass spectrometry for comprehensive proteome mapping. Here we describe the use of multiplexed quantitative proteomics using isobaric labeling with tandem mass tags (TMT) for the simultaneous quantitative analysis of five cancer cell proteomes in biological duplicates in one mass spectrometry experiment.

  6. Quantitative risk assessment for human salmonellosis through the consumption of pork sausage in Porto Alegre, Brazil.

    PubMed

    Mürmann, Lisandra; Corbellini, Luis Gustavo; Collor, Alexandre Ávila; Cardoso, Marisa

    2011-04-01

    A quantitative microbiology risk assessment was conducted to evaluate the risk of Salmonella infection to consumers of fresh pork sausages prepared at barbecues in Porto Alegre, Brazil. For the analysis, a prevalence of 24.4% positive pork sausages with a level of contamination between 0.03 and 460 CFU g(-1) was assumed. Data related to frequency and habits of consumption were obtained by a questionnaire survey given to 424 people. A second-order Monte Carlo simulation separating the uncertain parameter of cooking time from the variable parameters was run. Of the people interviewed, 87.5% consumed pork sausage, and 85.4% ate it at barbecues. The average risk of salmonellosis per barbecue at a minimum cooking time of 15.6 min (worst-case scenario) was 6.24 × 10(-4), and the risk assessed per month was 1.61 × 10(-3). Cooking for 19 min would fully inactivate Salmonella in 99.9% of the cases. At this cooking time, the sausage reached a mean internal temperature of 75.7°C. The results of the quantitative microbiology risk assessment revealed that the consumption of fresh pork sausage is safe when cooking time is approximately 19 min, whereas undercooked pork sausage may represent a nonnegligible health risk for consumers.

  7. Quantitative podocyte parameters predict human native kidney and allograft half-lives

    PubMed Central

    Naik, Abhijit S.; Afshinnia, Farsad; Cibrik, Diane; Hodgin, Jeffrey B.; 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

  8. A comparative study of three approaches to the routine quantitation of human serum proteins.

    PubMed

    Bruver, R M; Salkie, M L

    1978-06-01

    The Laser Nephelometer PDQTM (Hyland Division, Travenol Laboratories Inc.) and the Abbott Bichromatic Analyser 100 (Abbott Laboratories) were compared with a radial immunodiffusion method. Seventy-eight serum specimens collected during routine blood testing were aliquoted and quantitated by the three procedures described. The nephelometric system was used as described by Hyland in their instruction accompanying the LAS-R Nephelometric test kits. The ABA-100 was used with a filter unit transmitting at 340 and 650 nm and at a water bath temperature of 30 degrees C. The Laser Nephelometer correlated well with the RID system giving a correlation coefficient varying from 0.94 for IgA to 0.79 for C3. It was not possible to quantitate IgA by the turbidometric method using the ABA-100. Results obtained for the other proteins were satisfactory and the correlation coefficient with the RID method varied from 0.88 for IgG to 0.90 for C3. The RID procedure in routine use takes 5 hours technologist time and a 16-hour incubation period to produce results for IgG, IgA and IgM on nine patient specimens. Using the Laser Nephelometer to obtain the same results on 20 patient specimens took two to three technologist hours. Nephelometry, therefore, appears to be a satisfactory alternative to RID with a comparable precision and a great saving in technologist time.

  9. 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.

  10. A quantitative model of human DNA base excision repair. I. Mechanistic insights.

    PubMed

    Sokhansanj, Bahrad A; Rodrigue, Garry R; Fitch, J Patrick; Wilson, David M

    2002-04-15

    Base excision repair (BER) is a multistep process involving the sequential activity of several proteins that cope with spontaneous and environmentally induced mutagenic and cytotoxic DNA damage. Quantitative kinetic data on single proteins of BER have been used here to develop a mathematical model of the BER pathway. This model was then employed to evaluate mechanistic issues and to determine the sensitivity of pathway throughput to altered enzyme kinetics. Notably, the model predicts considerably less pathway throughput than observed in experimental in vitro assays. This finding, in combination with the effects of pathway cooperativity on model throughput, supports the hypothesis of cooperation during abasic site repair and between the apurinic/apyrimidinic (AP) endonuclease, Ape1, and the 8-oxoguanine DNA glycosylase, Ogg1. The quantitative model also predicts that for 8-oxoguanine and hydrolytic AP site damage, short-patch Polbeta-mediated BER dominates, with minimal switching to the long-patch subpathway. Sensitivity analysis of the model indicates that the Polbeta-catalyzed reactions have the most control over pathway throughput, although other BER reactions contribute to pathway efficiency as well. The studies within represent a first step in a developing effort to create a predictive model for BER cellular capacity.

  11. A quantitative model of human DNA base excision repair. I. mechanistic insights

    PubMed Central

    Sokhansanj, Bahrad A.; Rodrigue, Garry R.; Fitch, J. Patrick; Wilson, David M.

    2002-01-01

    Base excision repair (BER) is a multistep process involving the sequential activity of several proteins that cope with spontaneous and environmentally induced mutagenic and cytotoxic DNA damage. Quantitative kinetic data on single proteins of BER have been used here to develop a mathematical model of the BER pathway. This model was then employed to evaluate mechanistic issues and to determine the sensitivity of pathway throughput to altered enzyme kinetics. Notably, the model predicts considerably less pathway throughput than observed in experimental in vitro assays. This finding, in combination with the effects of pathway cooperativity on model throughput, supports the hypothesis of cooperation during abasic site repair and between the apurinic/apyrimidinic (AP) endonuclease, Ape1, and the 8-oxoguanine DNA glycosylase, Ogg1. The quantitative model also predicts that for 8-oxoguanine and hydrolytic AP site damage, short-patch Polβ-mediated BER dominates, with minimal switching to the long-patch subpathway. Sensitivity analysis of the model indicates that the Polβ-catalyzed reactions have the most control over pathway throughput, although other BER reactions contribute to pathway efficiency as well. The studies within represent a first step in a developing effort to create a predictive model for BER cellular capacity. PMID:11937636

  12. 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

  13. Reduced expression of 15-hydroxy prostaglandin dehydrogenase in chorion during labor is associated with decreased PRB and increased PRA and GR expression.

    PubMed

    Li, Yuan; He, Ping; Sun, Qianqian; Liu, Jie; Gao, Lu; You, Xingji; Gu, Hang; Ni, Xin

    2013-05-01

    The chorion laeve controls the levels of active prostaglandins within the uterus by NAD-dependent 15-hydroxy prostaglandin dehydrogenase (PGDH). The expression of PGDH in chorion is modulated by glucocorticoids and progesterone. In this study, we investigated glucocorticoid receptor (GR) and progesterone receptor A and B (PRA and PRB) in the regulation of PGDH expression in chorion, and we determined whether reduced PGDH expression in chorion during labor is associated with the changes in GR and PR expression by real-time RT-PCR and Western blot analysis. Dexamethasone (DEX) inhibited PGDH expression whereas progesterone stimulated PGDH expression in chorionic trophoblasts. DEX suppressed PGDH expression in GR overexpression and PR knockdown cells. The inhibitory effect of DEX did not occur in GR knockdown cells. Progesterone inhibited PGDH in GR overexpression and PR knockdown cells and it stimulated PGDH in PRB overexpression cells whereas it suppressed PGDH in PRA overexpression cells. Knockdown of c-Jun resulted in a loss of progesterone- and DEX-induced effects. PGDH was down-regulated in chorion tissues during labor. PRB was decreased whereas PRA and GR were increased in chorion during labor. Glucocorticoids inhibit PGDH expression via GR in chorionic trophoblasts. Progesterone enhances PGDH expression through PRB, whereas it inhibits PGDH expression via GR and PRA. Decreased PGDH expression is associated with increased GR and PRA, although decreased PRB, in chorion during labor.

  14. Double strand breaks induced by low doses of {gamma} rays or heavy ions: Quantitation in nonradioactive human DNA

    SciTech Connect

    Sutherland, B.M.; Bennett, P.V.; Sutherland, J.C.

    1996-07-15

    We have developed a method of quantitating low frequencies (0-30 sites/10{sup 9} base pairs) of double strand breaks in {approximately}1 {mu}g of nonradioactive human DNA. Unirradiated or irradiated DNA is digested with the restriction endonuclease NotI, producing cleavage fragments that include a major group centered at {approximately}1.2-1.3 Mbp. The DNA molecules are separated as a function of size by transverse alternating field electrophoresis. The frequency of double strand breaks is computed directly from the decrease in number average molecular length induced in the 1.2 to 1.3-Mbp cleavage fragment group by {sup 137}Cs {gamma} or Fe{sup 26+} (1.1 GeV/nucleon) irradiation vs the corresponding unirradiated DNA samples. The double strand break frequency can be quantitated easily in the dose range of 0-10 cGy of {gamma} rays. The frequency of breaks per unit dose calculated for {gamma} irradiation of DNA in human cells ({approximately}4.6 double strand breaks/10{sup 9} bp/Gy) who used methods requiring higher doses. 55 refs., 4 figs.

  15. Quantitation of isobaric phosphatidylcholine species in human plasma using a hybrid quadrupole linear ion-trap mass spectrometer.

    PubMed

    Zacek, Petr; Bukowski, Michael; Rosenberger, Thad A; Picklo, Matthew

    2016-12-01

    Phosphatidylcholine (PC) species in human plasma are used as biomarkers of disease. PC biomarkers are often limited by the inability to separate isobaric PCs. In this work, we developed a targeted shotgun approach for analysis of isobaric and isomeric PCs. This approach is comprised of two MS methods: a precursor ion scanning (PIS) of mass m/z 184 in positive mode (PIS m/z +184) and MS(3) fragmentation in negative mode, both performed on the same instrument, a hybrid triple quadrupole ion-trap mass spectrometer. The MS(3) experiment identified the FA composition and the relative abundance of isobaric and sn-1, sn-2 positional isomeric PC species, which were subsequently combined with absolute quantitative data obtained by PIS m/z +184 scan. This approach was applied to the analysis of a National Institute of Standards and Technology human blood plasma standard reference material (SRM 1950). We quantified more than 70 PCs and confirmed that a majority are present in isobaric and isomeric mixtures. The FA content determined by this method was comparable to that obtained using GC with flame ionization detection, supporting the quantitative nature of this MS method. This methodology will provide more in-depth biomarker information for clinical and mechanistic studies.

  16. Quantitative GSL-glycome analysis of human whole serum based on an EGCase digestion and glycoblotting method[S

    PubMed Central

    Furukawa, Jun-ichi; Sakai, Shota; Yokota, Ikuko; Okada, Kazue; Hanamatsu, Hisatoshi; Kobayashi, Takashi; Yoshida, Yasunobu; Higashino, Kenichi; Tamura, Tomohiro; Igarashi, Yasuyuki; Shinohara, Yasuro

    2015-01-01

    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

  17. Advanced forensic validation for human spermatozoa identification using SPERM HY-LITER™ Express with quantitative image analysis.

    PubMed

    Takamura, Ayari; Watanabe, Ken; Akutsu, Tomoko

    2017-01-19

    Identification of human semen is indispensable for the investigation of sexual assaults. Fluorescence staining methods using commercial kits, such as the series of SPERM HY-LITER™ kits, have been useful to detect human sperm via strong fluorescence. These kits have been examined from various forensic aspects. However, because of a lack of evaluation methods, these studies did not provide objective, or quantitative, descriptions of the results nor clear criteria for the decisions reached. In addition, the variety of validations was considerably limited. In this study, we conducted more advanced validations of SPERM HY-LITER™ Express using our established image analysis method. Use of this method enabled objective and specific identification of fluorescent sperm's spots and quantitative comparisons of the sperm detection performance under complex experimental conditions. For body fluid mixtures, we examined interference with the fluorescence staining from other body fluid components. Effects of sample decomposition were simulated in high humidity and high temperature conditions. Semen with quite low sperm concentrations, such as azoospermia and oligospermia samples, represented the most challenging cases in application of the kit. Finally, the tolerance of the kit against various acidic and basic environments was analyzed. The validations herein provide useful information for the practical applications of the SPERM HY-LITER™ Express kit, which were previously unobtainable. Moreover, the versatility of our image analysis method toward various complex cases was demonstrated.

  18. 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.

  19. Quantitation of isobaric phosphatidylcholine species in human plasma using a hybrid quadrupole linear ion-trap mass spectrometer[S

    PubMed Central

    Zacek, Petr; Bukowski, Michael; Rosenberger, Thad A.; Picklo, Matthew

    2016-01-01

    Phosphatidylcholine (PC) species in human plasma are used as biomarkers of disease. PC biomarkers are often limited by the inability to separate isobaric PCs. In this work, we developed a targeted shotgun approach for analysis of isobaric and isomeric PCs. This approach is comprised of two MS methods: a precursor ion scanning (PIS) of mass m/z 184 in positive mode (PIS m/z +184) and MS3 fragmentation in negative mode, both performed on the same instrument, a hybrid triple quadrupole ion-trap mass spectrometer. The MS3 experiment identified the FA composition and the relative abundance of isobaric and sn-1, sn-2 positional isomeric PC species, which were subsequently combined with absolute quantitative data obtained by PIS m/z +184 scan. This approach was applied to the analysis of a National Institute of Standards and Technology human blood plasma standard reference material (SRM 1950). We quantified more than 70 PCs and confirmed that a majority are present in isobaric and isomeric mixtures. The FA content determined by this method was comparable to that obtained using GC with flame ionization detection, supporting the quantitative nature of this MS method. This methodology will provide more in-depth biomarker information for clinical and mechanistic studies. PMID:27688258

  20. 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.

  1. Automated quantitation of cold-inducible human brown adipose tissue with FDG PET/CT with application to fibromyalgia.

    PubMed

    Pardo, José V; Lee, Joel T; Larson, Robert C; Thuras, Paul; Larson, Alice A

    2017-01-01

    Increasing recognition of the importance of brown adipose tissue (BAT) motivates the development of reproducible and quantitative methods for measuring it. Positron emission tomography (PET)/computerized tomography (CT) with (18)F-fluorodeoxyglucose (FDG) has become the principal method to non-invasively detect brown adipose tissue (BAT) in humans. Improvements in quantitation and standardization will drive further clinical application. One disorder hypothesized to involve dysregulation in thermoregulation and the processing of pain involving BAT is fibromyalgia syndrome (FMS). This report describes an approach with additional technical standardization to measure cold-inducible, BAT activity (ci-BAT) semi-quantitatively and reliably with minimal operator intervention with the FDG PET/CT technique. Ci-BAT was measured to test whether FMS patients have decreased BAT activation compared to normal controls. Threshold parameters to optimally separate ci-BAT from non-ci-BAT were developed based on the distribution of the pixel-wise parametric data from each merged PET/CT scan for each study session occurring on different days. BAT activity was the same under warm conditions in both control and FMS subjects attesting to reproducibility and reliability. However, considerable variability arose between groups at cool temperatures consistent with other literature. Increases in ci-BAT activity were significantly less in FMS patients than in controls, as hypothesized. Ci-BAT recruitment can be quantified non-invasively using FDG PET/CT using semi-automated techniques in human subjects across different diagnostic groups or within groups undergoing manipulations of interest.

  2. Automated quantitation of cold-inducible human brown adipose tissue with FDG PET/CT with application to fibromyalgia

    PubMed Central

    Pardo, José V; Lee, Joel T; Larson, Robert C; Thuras, Paul; Larson, Alice A

    2017-01-01

    Increasing recognition of the importance of brown adipose tissue (BAT) motivates the development of reproducible and quantitative methods for measuring it. Positron emission tomography (PET)/computerized tomography (CT) with 18F-fluorodeoxyglucose (FDG) has become the principal method to non-invasively detect brown adipose tissue (BAT) in humans. Improvements in quantitation and standardization will drive further clinical application. One disorder hypothesized to involve dysregulation in thermoregulation and the processing of pain involving BAT is fibromyalgia syndrome (FMS). This report describes an approach with additional technical standardization to measure cold-inducible, BAT activity (ci-BAT) semi-quantitatively and reliably with minimal operator intervention with the FDG PET/CT technique. Ci-BAT was measured to test whether FMS patients have decreased BAT activation compared to normal controls. Threshold parameters to optimally separate ci-BAT from non-ci-BAT were developed based on the distribution of the pixel-wise parametric data from each merged PET/CT scan for each study session occurring on different days. BAT activity was the same under warm conditions in both control and FMS subjects attesting to reproducibility and reliability. However, considerable variability arose between groups at cool temperatures consistent with other literature. Increases in ci-BAT activity were significantly less in FMS patients than in controls, as hypothesized. Ci-BAT recruitment can be quantified non-invasively using FDG PET/CT using semi-automated techniques in human subjects across different diagnostic groups or within groups undergoing manipulations of interest. PMID:28123865

  3. 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.

  4. Expression Quantitative Trait Loci Analysis Identifies Associations Between Genotype and Gene Expression in Human Intestine

    PubMed Central

    KABAKCHIEV, BOYKO; SILVERBERG, MARK S.

    2013-01-01

    BACKGROUND & AIMS Genome-wide association studies have greatly increased our understanding of intestinal disease. However, little is known about how genetic variations result in phenotypic changes. Some polymorphisms have been shown to modulate quantifiable phenotypic traits; these are called quantitative trait loci. Quantitative trait loci that affect levels of gene expression are called expression quantitative trait loci (eQTL), which can provide insight into the biological relevance of data from genome-wide association studies. We performed a comprehensive eQTL scan of intestinal tissue. METHODS Total RNA was extracted from ileal biopsy specimens and genomic DNA was obtained from whole-blood samples from the same cohort of individuals. Cis- and trans-eQTL analyses were performed using a custom software pipeline for samples from 173 subjects. The analyses determined the expression levels of 19,047 unique autosomal genes listed in the US National Center for Biotechnology Information database and more than 580,000 variants from the Single Nucleotide Polymorphism database. RESULTS The presence of more than 15,000 cis- and trans-eQTL was detected with statistical significance. eQTL associated with the same expression trait were in high linkage disequilibrium. Comparative analysis with previous eQTL studies showed that 30% to 40% of genes identified as eQTL in monocytes, liver tissue, lymphoblastoid cell lines, T cells, and fibroblasts are also eQTL in ileal tissue. Conversely, most of the significant eQTL have not been previously identified and could be tissue specific. These are involved in many cell functions, including division and antigen processing and presentation. Our analysis confirmed that previously published cis-eQTL are single nucleotide polymorphisms associated with inflammatory bowel disease: rs2298428/UBE2L3, rs1050152/SLC22A4, and SLC22A5. We identified many new associations between inflammatory bowel disease susceptibility loci and gene expression

  5. Quantitation of Circulating Neuropilin-1 in Human, Monkey, Mouse, and Rat Sera by ELISA.

    PubMed

    Lu, Yanmei; Meng, Y Gloria

    2015-01-01

    Neuropilin-1 (NRP1) is a single spanning transmembrane glycoprotein that acts as a co-receptor for class 3 semaphorins and vascular endothelial growth factors. Naturally occurring soluble NRP1 isoforms containing partial extracellular domain (ECD) have been reported. In addition to soluble NRP1, full-length NRP1 ECD has also been identified in human and animal sera. Here, we describe primate and rodent NRP1 ELISAs that measure total circulating NRP1 including soluble NPR1 and NRP1 ECD in human, monkey, mouse, and rat sera.

  6. In vitro quantitative analysis of Salmonella typhimurium preference for amino acids secreted by human breast tumor

    NASA Astrophysics Data System (ADS)

    Choi, Eunpyo; Maeng, Bohee; Lee, Jae-hun; Chang, Hyung-kwan; Park, Jungyul

    2016-12-01

    Bacterial therapies have been paid significant attentions by their ability to penetrate deep into the solid tumor tissue and its propensity to naturally accumulate in tumors of living animals. Understanding the actual mechanism for bacteria to target the tumor is therapeutically crucial but is poorly understood. We hypothesized that amino acids released from the specific tumors induced bacteria to those tumors and the experiments for chemotactic response of bacteria toward the cancer secreting amino acids was then performed by using the diffusion based multiple chemical gradient generator constructed by in situ self-assembly of microspheres. The quantitative analysis was carried out by comparison of intensity using green fluorescent protein (GFP) tagged Salmonella typhimurium ( S. typhimurium) in the gradient generator, which showed the clear preference to the released amino acids, especially from breast cancer patients. The understanding chemotaxis toward the cancer secreting amino acids is essential for controlling S. typhimurium targeting in tumors and will allow for the development of bacterial therapies.

  7. 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.

  8. 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

    2016-02-25

    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.

  9. 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

  10. 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...