Pregnancy-associated plasma protein-A2 levels are increased in early-pregnancy gestational diabetes: a novel biomarker for early risk estimation.

AIM: To determine whether pregnancy-associated plasma protein-A2 levels are increased in early pregnancies complicated by gestational diabetes and whether gestation age influences levels. The possible use of pregnancy-associated plasma protein-A2 as a pre-screening biomarker to reduce the need for performing oral glucose tolerance tests in pregnant women was also investigated. METHODS: Pregnant women were diagnosed with gestational diabetes in early pregnancy after a 2-hour 75 g oral glucose tolerance test in the catchment area of Skåne University Hospital, Lund, Sweden during 2011-2015 (n = 99). Age- and BMI-matched pregnant women without diabetes were recruited at similar gestational ages from maternal healthcare centres in the same geographical area during 2014-2015 to act as controls (n = 100). Circulating pregnancy-associated plasma protein-A2 was analysed in participant serum using commercially available enzyme-linked immunosorbent assay kits. RESULTS: Circulating pregnancy-associated plasma protein-A2 was increased in women diagnosed with gestational diabetes [13.5 (9.58-18.8) ng/ml] compared with controls [8.11 (5.74-11.3) ng/ml; P < 0.001]. Pregnancy-associated plasma protein-A2 was associated with gestational diabetes independent of age, BMI, C-peptide and adiponectin (P < 0.001). Pregnancy-associated plasma protein-A2 as a pre-screening biomarker to identify women at a decreased risk of gestational diabetes resulted in a negative predictive value of 99.7%, with a sensitivity of 96% and a specificity of 30% at a cut-off level of 6 ng/ml. CONCLUSIONS: This is the first study to show increased pregnancy-associated plasma protein-A2 levels in gestational diabetes. Pregnancy-associated plasma protein-A2 also shows promise as a pre-screening biomarker with the potential to reduce the need for performing oral glucose tolerance tests in early pregnancy. Future prospective cohort studies in a larger group of both high- and low-risk women are, however, needed to further confirm this observation.

Expression of PAPPA2 in human fetomaternal interface and involvement in trophoblast invasion and migration.

Pregnancy-associated plasma protein-A 2 (PAPPA2) is a placental-enriched gene that is important for normal human placentation and defects in the gene can cause complications in pregnancy. Yet the exact expression pattern and role of PAPPA2 in the human fetomaternal interface are not clear. In this study, in situ hybridization (ISH) and immunohistochemistry (IHC) were employed to examine the spatial and temporal expression of PAPPA2 in the human fetomaternal interface. IHC results exhibited wide expression of PAPPA2 in the fetomaternal interface, with placental syncytiatrophoblast (STB) and extravillous trophoblast (EVT) showing strong expression and the cytotrophoblast (CTB) showing weak expression of PAPPA2. These results were confirmed by ISH. Quantitative reverse transcription-polymerase chain reaction and western blot showed the elevation of PAPPA2 in first trimester EVT differentiation and term CTB spontaneous syncytialization. PAPPA2-siRNA transfection significantly depressed the invasion and migration ability of a trophoblast cell line (HTR8/SVneo) in a transwell migration and Matrigel invasion model compared to a negative control siRNA (P < 0.05), also revealing that matrix metalloproteinase 9 (MMP9) secretion is downregulated. This was confirmed using a human first trimester placental villi explant culture model. Our results reveal the spatial and temporal expression of PAPPA2 in the human fetomaternal interface and show the positive regulatory role of PAPPA2 in human trophoblast invasion and migration through the secretion of MMP9.

IGFBP-4 and -5 are expressed in first-trimester villi and differentially regulate the migration of HTR-8/SVneo cells.

BACKGROUND: Adverse gestational outcomes such as preeclampsia (PE) and intrauterine growth restriction (IUGR) are associated with placental insufficiency. Normal placental development relies on the insulin-like growth factors -I and -II (IGF-I and -II), in part to stimulate trophoblast proliferation and extravillous trophoblast (EVT) migration. The insulin-like growth factor binding proteins (IGFBPs) modulate the bioavailability of IGFs in various ways, including sequestration, potentiation, and/or increase in half-life. The roles of IGFBP-4 and -5 in the placenta are unknown, despite consistent associations between pregnancy complications and the levels of two IGFBP-4 and/or -5 proteases, pregnancy-associated plasma protein -A and -A2 (PAPP-A and PAPP-A2). The primary objective of this study was to elucidate the effects of IGFBP-4 and -5 on IGF-I and IGF-II in a model of EVT migration. A related objective was to determine the timing and location of IGFBP-4 and -5 expression in the placental villi. METHODS: We used wound healing assays to examine the effects of IGFBP-4 and -5 on the migration of HTR-8/SVneo cells following 4 hours of serum starvation and 24 hours of treatment. Localization of IGFBP-4, -5 and PAPP-A2 was assessed by immunohistochemical staining of first trimester placental sections. RESULTS: 2 nM IGF-I and -II each increased HTR-8/SVneo cell migration with IGF-I increasing migration significantly more than IGF-II. IGFBP-4 and -5 showed different levels of inhibition against IGF-I. 20 nM IGFBP-4 completely blocked the effects of 2 nM IGF-I, while 20 nM IGFBP-5 significantly reduced the effects of 2 nM IGF-I, but not to control levels. Either 20 nM IGFBP-4 or 20 nM IGFBP-5 completely blocked the effects of 2 nM IGF-II. Immunohistochemistry revealed co-localization of IGFBP-4, IGFBP-5 and PAPP-A2 in the syncytiotrophoblast layer of first trimester placental villi as early as 5 weeks of gestational age. CONCLUSIONS: IGFBP-4 and -5 show different levels of inhibition on the migration-stimulating effects of IGF-I and IGF-II, suggesting different roles for PAPP-A and PAPP-A2. Moreover, co-localization of the pappalysins and their substrates within placental villi suggests undescribed roles of these molecules in early placental development.

miRNA-214 is related to invasiveness of human non-small cell lung cancer and directly regulates alpha protein kinase 2 expression.

The prognosis of non-small cell lung cancer (NSCLC) is poor, since it has often metastasized to distant organs by the time of diagnosis. Therefore, biomarkers predicting metastasis are crucial. miRNAs play important roles in the regulation of different tumor cell processes, including metastasis. We recently showed that miRNA-214 is linked to a radioresistant phenotype of NSCLC. miRNA-214 has been linked to metastasis in other tumor types. Therefore, we examined the role of miRNA-214 in the metastatic potential of NSCLC. We showed that downregulation of miRNA-214 increased invasive potential, and conversely, overexpression of miRNA-214 decreased invasiveness of NSCLC cells in vitro. Gene expression and bioinformatic analyses of NSCLC cells with ablated miRNA-214, identified a number of metastasis-related target genes, including pregnancy-associated plasma protein A (PAPP-A), alpha protein kinase 2 (ALPK2), cyclin-dependent kinase 6 (CDK6) and tumor necrosis-factor alpha-induced protein 3 (TNFAIP3). These were validated on mRNA and protein level to be regulated by miRNA-214. Through immunoprecipitation we showed that only ALPK2 is directly regulated by miRNA-214. We also examined the protein expression of these four genes in NSCLC tumors with respect to metastatic potential. These results showed that NSCLC tumors express these proteins at moderate-high levels in the nucleus, cytoplasm and/or plasma membrane although with no significant correlation to the overall survival or the metastatic potential of the patients. However, we also showed that the membrane-localized PAPP-A had a higher expression level compared to the cytoplasm-localized. In conclusion, we show that low miRNA-214 expression is linked to a higher invasive potential of NSCLC cells.

[Expression and activity of proteases in metastasis of ovarian cancer].

The total chymotrypsin-like activity of proteasornes, the activity of 20S- and 26S-proteasome pools and calpains, and the expression of metalloproteinase PAPP-A in primary tumors and metastasized tissues were studied in 13 patients with epithelial ovarian cancer. It was shown that initiation of the process of tumor dissemination occurs against the background of active proteolytic processes; A decrease in activity of 26S-proteasomes and total calpain activity and increased expression ofmetalloproteinase PAPP-A in the primary tu mors were found in patients with ascites as compared with patients without ascites. The disease progression after treatment and achieved stabilization were found in patients with decreased activity of intracellular proteases and a high content of PAPP-A in the primary tumors.

Is serum pregnancy-associated plasma protein A really a potential marker of atherosclerotic carotid plaque stability?

OBJECTIVES: The search for markers predicting risk of plaque rupture in carotid atherosclerosis is still ongoing. Previous findings showed that pregnancy-associated plasma protein-A (PAPP-A) levels correlate with an adverse plaque morphology. However, the role of PAPP-A in plaque destabilisation is still uncertain. MATERIAL AND METHODS: Patients with carotid artery stenosis involved in the study were asymptomatic (n=29) and symptomatic (n=37). Carotid plaques were characterised by histology (n=33). Immunohistochemistry (n=17) was used to determine expression of PAPP-A and CD68 within the plaques. Serum levels of PAPP-A were measured by the enzyme-linked immunosorbent assay (ELISA). RESULTS: Circulating PAPP-A levels were significantly higher in patients with unstable versus stable plaques (0.10+/-0.06 vs. 0.07+/-0.04 microg ml(-1), p=0.047) and interestingly, in asymptomatic versus symptomatic patients (0.11+/-0.05 vs. 0.069+/-0.09 microg ml(-1), p=0.025). These differences remained statistically significant after adjustment for age, gender and degree of stenosis (p=0.050). PAPP-A expression in plaques correlated significantly with CD68 positive macrophages, cap-thickness and its serological values (r=+0.291, p<0.001, r=-0.639, p<0.001 and r=0.618, p<0.008, respectively). Furthermore, PAPP-A serum values demonstrated a significant positive predictive value of 68.8% for unstable plaques. CONCLUSION: Our present data confirmed the close relationship between expression of PAPP-A and plaque instability and furthermore correlated significantly with cap thickness. However, the question whether PAPP-A is a useful predictive marker of plaque instability remains unresolved.

Chronic Venous Disease Patients Showed Altered Expression of IGF-1/PAPP-A/STC-2 Axis in the Vein Wall.

Chronic venous disease (CVeD) has a remarkable prevalence, with an estimated annual incidence of 2%. It has been demonstrated how the loss of homeostatic mechanisms in the vein wall can take part in the physiopathology of CVeD. In this regard, it has been described how different axis, such as IGF-1/PAPP-A/STC-2 axis, may play an essential role in tissue homeostasis. The aim of this research is to study both genetic and protein expressions of the IGF-1/PAPP-A/STC-2 axis in CVeD patients. It is a cross-sectional study in which genetic (RT-qPCR) and protein (immunohistochemistry) expression analysis techniques were accomplished in saphenous veins from CVeD patients (n = 35) in comparison to individuals without vascular pathology (HV). Results show a significant increase in both genetic and protein expressions of PAPP-A and IGF-1, and a decrement STC-2 expression at the same time in CVeD patients. Our study is a pioneer for demonstrating that the expression of the different components of the IGF-1/PAPP-A/STC-2 axis is altered in CVeD patients. This fact can be a part of the loss of homeostatic mechanisms of the venous tissue. Further research should be destined to deepen into alterations of this axis, as well as to evaluate the usage of these components as therapeutic targets for CVeD.

Expression of a protease-resistant insulin-like growth factor-binding protein-4 inhibits tumour growth in a murine model of breast cancer.

BACKGROUND: Insulin-like growth factor 1 (IGF1) promotes breast cancer and disease progression. Bioavailability of IGF1 is modulated by IGF-binding proteins (IGFBPs). IGFBP4 inhibits IGF1 activity but cleavage by pregnancy-associated plasma protein-A (PAPP-A) protease releases active IGF1. METHODS: Expression of IGF pathway components and PAPP-A was assessed by western blot or RT-PCR. IGFBP4 (dBP4) resistant to PAPP-A cleavage, but retaining IGF-binding capacity, was used to block IGF activity in vivo. 4T1.2 mouse mammary adenocarcinoma cells transfected with empty vector, vector expressing wild-type IGFBP4 or vector expressing dBP4 were implanted in the mammary fat pad of BALB/c mice and tumour growth was assessed. Tumour angiogenesis and endothelial cell apoptosis were assessed by immunohistochemistry. RESULTS: 4T1.2 cells expressed the IGF1R receptor and IGFBP4. PAPP-A was expressed within mammary tumours but not by 4T1.2 cells. Proliferation and vascular endothelial growth factor (VEGF) production by 4T1.2 cells was increased by IGF1(E3R) (recombinant IGF1 resistant to binding by IGFBPs) but not by wild-type IGF1. IGF1-stimulated microvascular endothelial cell proliferation was blocked by recombinant IGFBP4. 4T1.2 tumours expressing dBP4 grew significantly more slowly than controls or tumours expressing wild-type IGFBP4. Inhibition of tumour growth by dBP4 was accompanied by the increased endothelial cell apoptosis. CONCLUSION: Protease-resistant IGFBP4 blocks IGF activity, tumour growth and angiogenesis.

Adjustment of serum markers in first trimester screening.

First trimester combined screening can be performed using maternal serum pregnancy-associated plasma protein A, total human chorionic gonadotropin (hCG) and ultrasound measurement of nuchal translucency at 11-13 weeks of pregnancy. Our objective was to explore the effects of covariates on total hCG in the first trimester. First trimester total hCG levels were significantly increased in twins (median = 1.87 MoM), mildly increased in pregnancies achieved by in vitro fertilization (1.04 MoM) and decreased in smokers (0.80 MoM).

Levels of pregnancy-associated plasma protein A in patients with coronary artery disease.

PURPOSE: To investigate the levels of pregnancy-associated plasma protein A (PAPP-A) or insulin-like growth factor -1 (IGF-1) in patients with acute coronary syndrome. METHODS: Serum PAPP-A and IGF-1 was measured with biotin-tyramide-amplified enzyme immunoassay and Enzyme Linked Immuoserbent Assay, respectively, in patients with ST elevation acute myocardial infarction (STEMI, n=12), unstable angina (UAP, n=15), and stable angina (n=15). PAPP-A and IGF-1 was also measured in 16 healthy subjects (control group). RESULTS: The serum levels of PAPP-A in the STEMI (16.9+/-10.3 mIU/L) and UAP group (15.2+/-10.5 mIU/L) were higher than in the stable angina (8.5+/-3.1 mIU/L) or control group (8.4+/-2.0 mIU/L, P < 0.01). The serum levels of IGF-1 in the STEMI (132.3+/-40.9 microg/L) and UAP group (127.3+/-36.0 microg/L) were also higher than in the stable angina (44.9+/-18.5 microg/L) or control group (67.7+/-24.5microg/L, P < 0.01). There were no differences in serum levels of PAPP-A or IGF-1 among the single, double and three vessel lesion groups. The serum levels of PAPP-A (19.9+/-10.1 mIU/L) and IGF-1 (153.2+/-52.4 microg/L) after PCI were higher than those before PCI (15.1+/-10.0 mIU/L and 91.4+/-51.0 microg/L, respectively, P < 0.01). A positive correlation was found between PAPP-A and IGF-1 levels in the STEMI and UAP group before PCI (r=0.48P < 0.01). CONCLUSION: PAPP-A and IGF-1 are elevated in patients with acute coronary syndrome. They may be used as biomarkers for vulnerable plaques in patients with coronary artery disease. Whether post-PCI elevation of IGF-1 can be used to predict restenosis of coronary arteries remains to be seen.

Placental expression of PAPPA, PAPPA-2 and PLAC-1 in pregnacies is associated with FGR.

Fetal growth restriction (FGR) is a gynecological disorder of varying etiology. In the present study, an expression analysis of pregnancy-associated plasma protein A (PAPPA), pregnancy-associated plasma protein A2 (PAPPA2) and placenta-specific-1 (PLAC-1) was conducted in pregnancies with FGR and control pregnancies. Placental tissues were collected from pregnancies with FGR (n=16) and control pregnancies (n=16) and the expression of the genes of interest was examined by qPCR. The mean expression levels of PAPPA and PAPPA2 were significantly lower (P<0.001) in placental tissues from FGR pregnancies compared with tissues from healthy subjects, whereas the opposite pattern was observed for PLAC-1 (P<0.001). PAPPA and PLAC-1 expression in FGR and control subjects correlated with birth weight (P<0.001). The findings suggest a possible pathophysiological link between the development of FGR and the expression of PAPPA, PAPPA2 and PLAC-1.

Pregnancy associated plasma protein-A links pregnancy and melanoma progression by promoting cellular migration and invasion.

Melanoma is the most common cancer diagnosed in pregnant women and an aggressive course with poorer outcomes is commonly described during pregnancy or shortly after childbirth. The underlying mechanisms for this are not understood. Here, we report that melanoma migration, invasiveness and progression are promoted by Pregnancy-Associated Plasma Protein-A (PAPPA), a pregnancy-associated metalloproteinase produced by the placenta that increases the bioavailability of IGF1 by cleaving it from a circulating complex formed with IGFBP4. We show that PAPPA is widely expressed by metastatic melanoma tumors and is elevated in melanoma cells exhibiting mesenchymal, invasive and label-retaining phenotypes. Notably, inhibition of PAPPA significantly reduced invasion and migration of melanoma cells in vitro and in vivo within the embryonic chicken neural tube. PAPPA-enriched pregnancy serum treatment enhanced melanoma motility in vitro. Furthermore, we report that IGF1 can induce the phenotypic and functional effects of epithelial-to-mesenchymal transition (EMT) in melanoma cells. In this study, we establish a clear relationship between a pregnancy-associated protein PAPPA, melanoma and functional effects mediated through IGF1 that provides a plausible mechanism for accelerated melanoma progression during pregnancy. This opens the possibility of targeting the PAPPA/IGF1 axis therapeutically.

Pregnancy-associated plasma protein-a production in rat granulosa cells: stimulation by follicle-stimulating hormone and inhibition by the oocyte-derived bone morphogenetic protein-15.

Pregnancy-associated plasma protein-A (PAPP-A) is the major IGF binding protein-4 (IGFBP-4) protease in follicular fluid, consistent with its proposed role in folliculogenesis. Despite growing interest, almost nothing is known about how PAPP-A expression is regulated in any tissue. Here we show that FSH and oocytes regulate PAPP-A expression in granulosa cells (GCs). By in situ hybridization, ovary PAPP-A mRNA was markedly increased by pregnant mare serum gonadotropin treatment, and the message was localized to the membrana GCs but not cumulus GCs (CGCs) of dominant follicles. To explore the mechanism, we used primary cultures of rat GCs. Control (untreated) cells produced little or no PAPP-A spontaneously. Conversely, FSH markedly stimulated PAPP-A mRNA and protein in a dose- and time-dependent fashion. Interestingly, PAPP-A expression in isolated CGCs was also strongly induced by FSH, and the induction was inhibited by added oocytes. To investigate the nature of the inhibition, we tested the effect of oocyte-derived bone morphogenetic protein-15 (BMP-15). BMP-15 alone had no effect on basal levels of PAPP-A expression by cultures of membrana GCs or CGCs. However, BMP-15 markedly inhibited the FSH stimulation of PAPP-A production in a dose-dependent manner. The cleavage of IGFBP-4 by conditioned media from FSH-treated GCs was completely inhibited by anti-PAPP-A antibody, indicating the IGFBP-4 protease secreted by GCs is PAPP-A. These results demonstrate stimulatory and inhibitory roles for FSH and BMP-15, respectively, in regulating PAPP-A production by GCs. We propose that FSH and oocyte-derived BMP-15 form a controlling network that ensures the spatiotemporal pattern of GC PAPP-A expression in the dominant follicle.

The regulated expression of the pregnancy-associated plasma protein-A in the rodent ovary: a proposed role in the development of dominant follicles and of corpora lutea.

Compelling evidence exists displaying that the intrafollicular IGF-I system constitutes an obligatory mediator of FSH action in the murine ovary. Within this system, the ovarian IGF binding protein-4-directed protease (IGFBP-4ase) may have a critical role. Human IGFBP-4ase has been proved identical to the previously well-characterized pregnancy-associated plasma protein-A (PAPP-A). This communication reports the cloning and sequencing of the mouse PAPP-A cDNA as well as its expression and cellular localization in the mouse ovary. PAPP-A mRNA was undetectable in ovaries of untreated immature 25-d-old mice. Treatment with PMSG led to a marked time-dependent increase in PAPP-A expression in well-defined subsets of granulosa cells and follicles. Specifically, PAPP-A expression was detectable exclusively in centrifugally residing membrana granulosa cells of antral follicles during a 3- to 36-h period post PMSG. PAPP-A expression then fell to nondetectable levels in dominant preovulatory follicles at 48 h post PMSG. Treatment of PMSG-primed mice with human CG caused a rapid reinduction of PAPP-A expression in granulosa cells of dominant follicles and was sustained at relatively high levels throughout the ovulation and luteinization. These results suggest a role for gonadotropin-stimulated PAPP-A gene expression in the physiologic processes of dominant follicle development, ovulation, and luteogenesis in the mammalian ovary. The early onset and extended duration of gonadotropin-dependent PAPP-A expression in granulosa cells may serve to degrade the antigonadotropin IGFBP-4. Accordingly, successful antral follicle development, ovulation, and corpus luteum formation may be contingent on an IGFBP-4-deplete/PAPP-A-replete circumstance, hence resulting in an IGF-I-replete intrafollicular microenvironment.

Pregnancy-associated plasma protein-A gene expression in human ovaries is restricted to healthy follicles and corpora lutea.

Recently, Pregnancy-Associated Plasma Protein-A (PAPP-A) in human follicular fluid was identified as an insulin-like growth factor binding protein-4 protease (IGFBP-4ase). The ability of IGFBP-4ase to inactivate the FSH antagonist, IGFBP-4, has suggested a possible role for PAPP-A in regulating FSH action. Despite growing interest in this protease, the question of whether the PAPP-A gene is expressed in ovaries of normal cycling women is unknown. To fill this basic gap in our knowledge, we have identified the cellular sites of PAPP-A gene expression in normal human ovaries by in situ hybridization. PAPP-A mRNA was low or undetectable in preantral follicles, small (1-2 mm) healthy and atretic antral follicles, larger atretic antral follicles, surface epithelium, tunica albuginea and connective tissue cells. In contrast, an intense PAPP-A hybridization signal was evident in the healthy antral follicles examined from 5 mm to the preovulatory stage. In these follicles, the signal was restricted to the granulosa cells (GC). An intense signal for PAPP-A mRNA was also present in healthy corpora lutea (CL), being localized to a subset of large luteal cells. Collectively, these results provide the first evidence that the gene encoding PAPP-A is expressed in ovaries of normal cycling women and show that the gene is expressed almost exclusively in healthy GC and CL cells. The restricted pattern of PAPP-A expression in normal human ovaries suggests that PAPP-A may be a functional marker of the dominant follicle and its product, the CL. Although the physiological function of ovarian PAPP-A remains to be identified, we hypothesize it might play a role in controlling survival, growth, and/or differentiation of the dominant follicle and CL by inactivating the gonadotropin antagonist, IGFBP-4.

Localization and regulation of pregnancy-associated plasma protein a expression in healing human skin.

Pregnancy-associated plasma protein A (PAPP-A) is an IGF-binding protein-4 (IGFBP-4) metalloproteinase that cleaves inhibitory IGFBP-4 to amplify local IGF-I bioavailability in vitro. Thus it has functional implications in injury/repair responses. In this study we determined PAPP-A expression in healing human skin. Wounds were induced with a scalpel on the forearms of three normal subjects and were allowed to heal by first intention. Biopsies obtained on d 0, 2, 8, and 14 were processed for immunohistochemical detection of PAPP-A, IGF-I, and IGFBP-4. In uninjured skin (d 0), strong staining for PAPP-A was present in the epidermis, sweat and sebaceous gland epithelial cells, hair follicles, and blood vessels; no PAPP-A was detected in dermal fibroblasts or with mature collagen bundles. IGF-I localized strongly to epithelial cells of skin glands was weak to moderate in epidermis and blood vessels, and was absent in dermal cells. Weak focal staining for IGFBP-4 was found within uninjured epidermis. During wound healing, PAPP-A expression was induced in dermal granulation tissue within and adjacent to the injury. PAPP-A was present in dermis on d 2 and was increased in intensity and extent on d 8 and 14. PAPP-A expression also increased in the epidermis. PAPP-A expression in cells of granulation tissue colocalized with alpha-smooth actin staining of myofibroblasts and new blood vessels as well as with CD68 staining of macrophages and was associated with the compact, newly synthesized collagen of the healing wound. IGF-I staining was enhanced in the epidermis localized to the area of the incision and in granulation tissue associated with lymphoid cells. IGFBP-4 staining of the epidermis remained unchanged during wound healing, but was induced in the fibroblastic cells of granulation tissue over time. These data demonstrate localized and regulated expression of PAPP-A in human skin and suggest that PAPP-A may play an important role in an integrated IGF system in wound healing and tissue remodeling in vivo.

Evaluation of first trimester maternal serum and ultrasound screening for Down's syndrome in Eastern and Northern Finland.

The current trend in prenatal diagnosis is that trisomy screening is being moved to the first trimester and ultrasonographic nuchal translucency measurement is included in risk calculation. It is likely that biochemical screening in the second trimester will gradually be given up. In Eastern and Northern Finland, during the year 1999 we offered first-trimester ultrasonographic and serum screening for trisomy 21, with measurements of maternal serum PAPP-A and beta-hCG. A total of 2515 pregnant women participated in the screening, yielding the detection of eight foetuses with Down's syndrome. Six affected foetuses (75%) were detected by means of first-trimester serum screening. Since we were in the phase of collecting data for the Finnish medians for PAPP-A and beta-hCG, the women were not given the estimates of risk for trisomy 21. Only 1602 of the 2515 enrolled women had the combination of first-trimester ultrasonographic and serum screening performed, and in that group there were five foetuses with Down's syndrome. The combination ultrasonographic and serum approach yielded a Down's syndrome detection rate of 80% (four out of five) with a 5% false positive rate, whereas in nuchal translucency based-screening the detection rate was 60%, with a 5% false positive rate.

Regulation of pregnancy-associated plasma protein-A expression in cultured human osteoblasts.

Pregnancy-associated plasma protein-A (PAPP-A) is a metalloproteinase secreted by cultured human osteoblasts that has been implicated in the regulation of local insulin-like growth factor (IGF) bioavailability during bone growth and remodeling. However, very little is known about the regulation of PAPP-A expression in bone. In this study, we determined the effect of systemic and local osteoregulatory factors on PAPP-A mRNA and protein expression in normal human osteoblasts (hOB cells). Treatment of hOB cells with particular peptide growth factors (basic fibroblast growth factor, epidermal growth factor), steroid hormones (dexamethasone, 1,25-dihydroxyvitamin D(3)), and cytokines [interleukin-6 (IL-6), IL-13, oncostatin M] with known involvement in bone cell physiology had no significant effect on PAPP-A expression. Agents that increase intracellular cyclic AMP (forskolin, prostaglandin E(2)) increased PAPP-A mRNA and protein expression approximately 3-fold. Tumor necrosis factor alpha (TNFalpha), IL-1beta, and IL-4 also increased PAPP-A expression 3- to 4-fold. Transforming growth factor beta (TGFbeta) was previously shown to stimulate PAPP-A expression in hOB cells. The effects of TGFbeta, TNFalpha, and IL-1beta were additive, whereas the effects of TGFbeta and IL-4 were synergistic. In summary, TNFalpha, IL-1beta, and IL-4 were identified as potent stimulators of PAPP-A expression in primary cultures of human osteoblasts. These findings suggest a mechanism whereby cytokines present in bone and bone marrow could augment IGF bioavailability during skeletal growth and remodeling.

The influence of different sample collection types on the levels of markers used for Down's syndrome screening as measured by the Kryptor Immunosassay system.

BACKGROUND: In a rapid point-of-care screening programme for chromosomal anomalies, analysis of biochemical markers in maternal blood can now be accomplished in a rapid time frame (less than 20 min). The need to leave whole blood samples some 10 min for coagulation and a further 5 min for centrifugation adds additional processing time. METHODS: The possibilities for reducing this processing time were investigated using various anticoagulated blood collection systems and the Kryptor analytical platform. Plasma levels of alpha-fetoprotein (AFP), pregnancy-associated plasma protein-A (PAPP-A) and free human chronic gonadotrophin beta-subunit (beta-hCG) were compared with those in maternal serum. RESULTS: From the mean results from ten patients it was shown that use of heparin plasma resulted in a statistically significant reduction in levels of PAPP-A and that EDTA plasma reduced the levels of PAPP-A dramatically. For AFP, levels in citrated plasma and EDTA plasma were also significantly reduced, whereas levels of free beta-hCG were not affected. CONCLUSION: Use of alternative sample types for PAPP-A is not possible. The sample of choice for first trimester screening using the Kryptor platform is maternal serum.

First-trimester levels of pregnancy-associated plasma protein A2 (PAPP-A2) in the maternal circulation are elevated in pregnancies that subsequently develop preeclampsia.

Recent studies have consistently found pregnancy-associated plasma protein A2 (PAPP-A2) to be upregulated in preeclamptic placentae at term. We tested whether first-trimester circulating PAPP-A2 levels differed between complicated and uncomplicated pregnancies. We measured maternal PAPP-A2 levels at 10 to 14 weeks of gestational age in 17 pregnancies resulting in small-for-gestational-age (SGA) infants, 6 which developed preeclampsia (PE), 1 which developed PE and resulted in an SGA infant, and 37 gestational age-matched controls. The concentration of the PAPP-A2 isoform corresponding to the full-length protein was significantly higher in pregnancies that developed PE (35 ng/mL) compared with those that did not (23 ng/mL; P < .044). In contrast, we found no difference in PAPP-A2 levels between pregnancies that did or did not result in an SGA infant. The upregulation of PAPP-A2 that has previously been observed in PE at term appears to begin early in pregnancy, well before the symptoms develop.