Ectodysplasin has a dual role in ectodermal organogenesis: inhibition of Bmp activity and induction of Shh expression.

Ectodermal organogenesis is regulated by inductive and reciprocal signalling cascades that involve multiple signal molecules in several conserved families. Ectodysplasin-A (Eda), a tumour necrosis factor-like signalling molecule, and its receptor Edar are required for the development of a number of ectodermal organs in vertebrates. In mice, lack of Eda leads to failure in primary hair placode formation and missing or abnormally shaped teeth, whereas mice overexpressing Eda are characterized by enlarged hair placodes and supernumerary teeth and mammary glands. Here, we report two signalling outcomes of the Eda pathway: suppression of bone morphogenetic protein (Bmp) activity and upregulation of sonic hedgehog (Shh) signalling. Recombinant Eda counteracted Bmp4 activity in developing teeth and, importantly, inhibition of BMP activity by exogenous noggin partially restored primary hair placode formation in Eda-deficient skin in vitro, indicating that suppression of Bmp activity was compromised in the absence of Eda. The downstream effects of the Eda pathway are likely to be mediated by transcription factor nuclear factor-kappaB (NF-kappaB), but the transcriptional targets of Edar have remained unknown. Using a quantitative approach, we show in cultured embryonic skin that Eda induced the expression of two Bmp inhibitors, Ccn2/Ctgf (CCN family protein 2/connective tissue growth factor) and follistatin. Moreover, our data indicate that Shh is a likely transcriptional target of Edar, but, unlike noggin, recombinant Shh was unable to rescue primary hair placode formation in Eda-deficient skin explants.

Characterization of pip5k3 fleck corneal dystrophy-linked gene in zebrafish.

PIKfyve is a kinase encoded by pip5k3 involved in phosphatidylinositols (PdtIns) pathways. These lipids building cell membranes have structural functions and are involved in complex intracellular regulations. Mutations in human PIP5K3 are associated with François-Neetens mouchetée fleck corneal dystrophy [Li, S., Tiab, L., Jiao, X., Munier, F.L., Zografos, L., Frueh, B.E., Sergeev, Y., Smith, J., Rubin, B., Meallet, M.A., Forster, R.K., Hejtmancik, J.F., Schorderet, D.F., 2005. Mutations in PIP5K3 are associated with François-Neetens mouchetee fleck corneal dystrophy. Am. J. Hum. Genet. 77, 54-63]. We cloned the zebrafish pip5k3 and report its molecular characterization and expression pattern in adult fish as well as during development. The zebrafish PIKfyve was 70% similar to the human homologue. The gene encompassed 42 exons and presented four alternatively spliced variants. It had a widespread expression in the adult organs and was localized in specific cell types in the eye as the cornea, lens, ganglion cell layer, inner nuclear layer and outer limiting membrane. Pip5k3 transcripts were detected in early cleavage stage embryos. Then it was uniformly expressed at 10 somites, 18 somites and 24 hpf. Its expression was then restricted to the head region at 48 hpf, 72 hpf and 5 dpf and partial expression was found in somites at 72 hpf and 5 dpf. In situ on eye sections at 3 dpf showed a staining mainly in lens, outer limiting membrane, inner nuclear layer and ganglion cell layer. A similar expression pattern was found in the eye at 5 dpf. A temporal regulation of the spliced variants was observed at 1, 3 and 5 dpf and they were also found in the adult eye.

The current molecular phylogeny of Eutherian mammals challenges previous interpretations of placental evolution.

Based on histology, the placentae of eutherians are currently grouped in epitheliochorial, endotheliochorial and haemochorial placentae. In a haeckelian sense, the epitheliochorial contact with marked histiotrophic feeding by uterine milk is generally considered as primitive, especially since similar contacts exist in Marsupials. In contrast, the more intimate endotheliochorial and haemochorial contact, facilitating haemotrophic nutrition, is interpreted as a derived state. A cladistic analysis based on the phylogenetic relationships established by molecular analyses reveals that the basic clades are all characterized by an endotheliochorial or haemochorial placenta, and that the epitheliochorial placenta evolved at least three times in a convergent manner. This evolution may be explained by the fact that the epitheliochorial placenta in eutherians is more efficient in nutritional transfer (flow rate by exchange surface). Moreover, this arrangement may confer an advantage to the mother who can probably reduce the degree of manipulation by a genetically imprinted embryo.

Introducció experimental a la biotecnologia amb clavellina: cultiu in vitro

Treball de recerca realitzat per una alumna d'ensenyament secundari i guardonat amb un Premi CIRIT per fomentar l'esperit científic del Jovent l'any 2009. Es tracta d’una recerca experimental en la que s’han assajat vuit tècniques de cultiu in vitro amb clavellina. El material vegetal s’ha esterilitzat per immersió en una solució diluïda de lleixiu i s’ha manipulat de manera estèril. En tots els casos el medi de cultiu utilitzat ha estat el MS amb una concentració de sacarosa i reguladors de creixement variable segons l’experiment. La incubació dels cultius s’han dut a terme en una cambra amb control de fotoperíode durant 4 setmanes. Els diferents reguladors de creixement han mostrat un clar efecte sobre les seccions de tija. Els explants cultivats en medi lliure d’hormones han crescut menys que els exposats a diverses concentracions de NAA i BA. Aquests tractaments hormonals han originat símptomes de creixement anòmals (engruiximents a la base i vitrificació). La presencia de 2,4-D ha afavorit la formació de cal•lus i d’arrels per organogènesi adventícia indirecta. L’obtenció de plàntules per germinació in vitro de llavors ha permès reduir notablement les pèrdues per contaminació, mentre que el subcultiu d’aquestes ha donat unes tases de micropropagació de 7.2 seccions/plàntula. Ha estat possible aclimatar aquestes vitroplantes per tal d’adaptar-les a les condicions de camp. No hem pogut obtenir organogènesis adventícia ni embriogènesi somàtica a partir d anteres ni hem pogut iniciar un cultiu de cèl•lules a partir dels cal•lus. Tot i la complexitat d’aquestes tècniques, és possible dur-les a terme en un laboratori escolar.

PGC1alpha expression is controlled in skeletal muscles by PPARbeta, whose ablation results in fiber-type switching, obesity, and type 2 diabetes.

Mice in which peroxisome proliferator-activated receptor beta (PPARbeta) is selectively ablated in skeletal muscle myocytes were generated to elucidate the role played by PPARbeta signaling in these myocytes. These somatic mutant mice exhibited a muscle fiber-type switching toward lower oxidative capacity that preceded the development of obesity and diabetes, thus demonstrating that PPARbeta is instrumental in myocytes to the maintenance of oxidative fibers and that fiber-type switching is likely to be the cause and not the consequence of these metabolic disorders. We also show that PPARbeta stimulates in myocytes the expression of PGC1alpha, a coactivator of various transcription factors, known to play an important role in slow muscle fiber formation. Moreover, as the PGC1alpha promoter contains a PPAR response element, the effect of PPARbeta on the formation and/or maintenance of slow muscle fibers can be ascribed, at least in part, to a stimulation of PGC1alpha expression at the transcriptional level.

Vitellogenesis in insects and other groups: a review

The eggs from oviparous organisms contain large amounts of vitellus, or yolk, wich are utilized by the growing embryo. Vitellogenesis is the process of vitellus accumulation and involves massive heterosynthetic synthesis of the protein vitellogenin (Vg) and its deposition in the oocyte. This work summarizes data on Vg structure, synthesis, uptake by oocytes and its fate during embryogenesis. The hormonal control of vitellogenesis and its tissue, sex and temporal regulation are also discussed. Where it is available, data on structure and expression of Vg-coding genes are reviewed. Insect vitellogenesis is priorized although other oviparous animal groups outside insects are also treated.

The role of Pten in the Hematopoietic System and the Hematopoietic Stem Cells

Résumé Identification, localisation et activation des cellules souches hématopoiétiques dormantes in vivo Les cellules souches somatiques sont présentes dans la majorité des tissus régénératifs comme la peau, l'épithélium intestinal et le système hématopoiétique. A partir d'une seule cellule, elles ont les capacités de produire d'autres cellules souches du même type (auto-renouvellement) et d'engendrer un ensemble défini de cellules progénitrices différenciées qui vont maintenir ou réparer leur tissu hôte. Les cellules souches adultes les mieux caractérisées sont les cellules souches hématopoiétiques (HSC), localisées dans la moelle osseuse. Un des buts de mon travail de doctorat était de caractériser plus en profondeur la localisation des HSCs endogènes in vivo. Pour ce faire, la technique "label retaining assay", se basant sur la division peu fréquentes et sur la dormance des cellules souches, a été utilisée. Après un marquage des souris avec du BrdU (analogue à l'ADN) suivi d'une longue période sans BrdU, les cellules ayant incorporés le marquage ("label retaining cells" LCRs) ont pu être identifiées dans la moelle osseuse. Ces cellules LCRs étaient enrichies 300 fois en cellules de phenotype HSC et, en utilisant de la cytofluorométrie, il a pu être montré qu'environ 15% de toutes les HSCs d'une souris restent dormantes durant plusieures semaines. Ces HSCs dormantes à long terme ne sont probablement pas impliquées dans la maintenance de 'hématopoièse. Par contre, on assiste à l'activation rapide de ces HSCs dormantes lors d'une blessure, comme une ablation myéloide. Elles re-entrent alors en cycle cellulaire et sont essentielles pour une génération rapide des cellules progénitrices et matures qui vont remplacer les cellules perdues. De plus, la détection des LCRs, combinée avec l'utilisation du marqueur de HSCs c-kit, peut être utilisée pour la localisation des HSCs dormantes présentes dans la paroi endostéale de la cavité osseuse. De manière surprenante, les LCRs c-kit+ ont surtout étés trouvées isolées en cellule unique, suggérant que le micro-environement spécifique entourant et maintenant les HSCs, appelé niche, pourrait être très réduit et abriter une seule HSC par niche. Rôles complexes du gène supresseur de tumeur Pten dans le système hématopoiétique La phosphatase PTEN disparaît dans certains cancers héréditaires ou sporadiques humains, comme les gliomes, les cancers de l'utérus ou du sein. Pten inhibe la voie de signalisation de la PI3-kinase et joue un rôle clé dans l'apoptose, la croissance, la prolifération et la migration cellulaire. Notre but était d'étudier le rôle de Pten dans les HSC normale et durant la formation de leucémies. Pour ce faire, nous avons généré un modèle murin dans lequel le gène Pten peut être supprimé dans les cellules hématopoiétiques, incluant les HSCs. Ceci a été possible en croissant l'allèle conditionnelle ptenflox soit avec le transgène MxCre inductible par l'interféron α soit avec le transgène Scl-CreERt inductible par le tamoxifen. Ceci permet la conversion de l'allèle ptenflox en l'allèle nul PtenΔ dans les HSCs et les autres types cellulaires hématopoiétiques. Les souris mutantes Pten développent une splénomégalie massive causée par une expansion dramatiques de toutes les cellules myéloides. De manière interessante, alors que le nombre de HSCs dans la moelle osseuse diminue progressivement, le nombre des HSCs dans la rate augmente de manière proportionnelle. Etrangement, les analyses de cycle cellulaire ont montrés que Pten n'avait que peu ou pas d'effet sur la dormance des HSCs ou sur leur autorenouvellement. En revanche, une augmentation massive du niveau de la cytokine de mobilisation G-CSF a été détéctée dans le serum sanguin, suggérant que la suppression de Pten stimulerait la mobilisation et la migration des HSC de la moelle osseuse vers la rate. Finallement, la transplantation de moelle osseuse délétée en Pten dans des souris immuno-déficientes montre que Pten fonctionnerait comme un suppresseur de tumeur dans le système hématopoiétique car son absence entraîne la formation rapide de leucémies lymphocytaires. Summary Identification, localization and activation of dormant hematopoietic stun cells in vivo Somatic stem cells are present in most self-renewing tissues including the skin, the intestinal epithelium and the hematopoietic system. On a single cell basis they have the capacity to produce more stem cells of the same phenotype (self-renewal) and to give rise to a defined set of mature differentiated progeny, responsible for the maintenance or repair of the host tissue. The best characterized adult stem cell is the hematopoietic stem cell (HSC) located in the bone marrow. One goal of my thesis work was to further characterize the location of endogenous HSCs in vivo. To do this, a technique called "label retaining assay» was used which takes advantage of the fact that stem cells (including HSCs) divide very infrequently and can be dormant for months. After labeling mice with the DNA analogue BrdU followed by a long BrdU free "chase", BrdU "label retaining cells" (CRCs) could be identified in the bone marrow. These CRCs were 300-fold enriched for phenotypic HSCs and by using flow cytometry analysis it could be shown that about 15% of all HSCs in the mouse are dormant for many weeks. Our results suggest that these long-term dormant HSCs are unlikely to be involved in homeostatic maintenance. However they are rapidly activated and reenter the cell cycle in response to injury signals such as myeloid ablation. In addition, detection of LRCs in combination with the HSC marker c-Kit could be used to locate engrafted dormant HSCs close to the endosteal lining of the bone marrow cavities. Most surprisingly, c-Kit+LRCs were found predominantly as single cells suggesting that the specific stem cell maintaining microenvironment, called niche, has limited space and may house only single HSCs. Complex roles of the tumor suppressor gene Pten in the hematopoietic system. The phosphatase PTEN is lost in hereditary and sporadic forms of human cancers, including gliomas, endometrial and breast cancers. Pten inhibits the PI3-kina.se pathway and plays a key role in apoptosis, cell growth, proliferation and migration. Our aim was to study the role of Pten in normal HSCs and during leukemia formation. To do this, we generated a mouse model in which the Pten gene can be deleted in hematopoietic cells including HSCs. This was achieved by crossing the conditional ptenflox allele with either the interferona inducible MxCre or the tamoxifen inducible Scl-CreERT transgene. This allowed the conversion of the ptenflox allele into a pterr' null allele in HSCs and other hematopoietic cell types. As a result Pten mutant mice developed massive splenomegaly due to a dramatic expansion of all myeloid cells. Interestingly, while the number of bone marrow HSCs progressively decreased, the number of HSCs in the spleen increased to a similar extent. Unexpectedly, extensive cell cycle analysis showed that Pten had little or no effect on HSC dormancy or HSC self-renewal. Instead, dramatically increased levels of the mobilizing cytokine G-CSF were detected in the blood serum suggesting that loss-of Pten stimulates mobilization and migration of HSC from the BM to the spleen. Finally, transplantation of Pten deficient BM cells into immuno-compromised mice showed that Pten can function as a tumor suppressor in the hematopoietic system and that its absence leads to the rapid formation of T cell leukemia.

Effect of temperature on different stages of Romanomermis iyengari, a mermithid nematode parasite of mosquitoes

The effect of temperature (20 degrees-35 degrees C) on different stages of Romanomermis iyengari was studied. In embryonic development, the single-cell stage eggs developed into mature eggs in 4.5-6.5 days at 25-35 degrees C but, required 9.5 days at 20 degrees C. Complete hatching occurred in 7 and 9 days after egg-laying at 35 and 30 degrees C, respectively. At 25 and 20 degrees C, 85-96 of the eggs did not hatch even by 30th day. Loss of infectivity and death of the preparasites occurred faster at higher temperatures. The 50 survival durations of preparasites at 20 and 35 degrees C were 105.8 and 10.6 hr respectively. They retained 50 infectivity up to 69.7 and 30.3 hr. The duration of the parasitic phase increased as temperature decreased. Low temperature favoured production of a higher proportion of females which were also larger in size. The maximum time taken for the juveniles to become adults was 14 days at 20 degrees C and the minimum was 9 days at 35 degrees C. Oviposition began earlier at higher temperature than at lower temperature. However, its fecundic period was shorter at 20 degrees C than at 35 degrees C indicating enhanced rate of oviposition at 20 degrees C. Fecundity was adversely affected at 20 degrees C and 35 degrees C. It is shown that the temperature range of 25 degrees-30 degrees C favours optimum development of R. iyengari.

The L-Type Ca+ and KATP channels may contribute to pacing-induced protection against anoxia-reoxygenation in the embryonic heart model.

L-Type Ca(2+) and K(ATP) Channels in Pacing-Induced Cardioprotection. AIMS: The L-type Ca(2+) channel, the sarcolemmal (sarcK(ATP)), and mitochondrial K(ATP) (mitoK(ATP)) channels are involved in myocardial preconditioning. We aimed at determining to what extent these channels can also participate in pacing-induced cardioprotection. METHODS: Hearts of 4-day-old chick embryos were paced in ovo during 12 hour using asynchronous intermittent ventricular stimulation at 110% of the intrinsic rate. Sham operated and paced hearts were then submitted in vitro to anoxia (30 minutes) and reoxygenation (60 minutes). These hearts were exposed to L-type Ca(2+) channel agonist Bay-K-8644 (BAY-K) or blocker verapamil, nonselective K(ATP) channel antagonist glibenclamide (GLIB), mitoK(ATP) channel agonist diazoxide (DIAZO), or antagonist 5-hydroxydecanoate. Electrocardiogram, electromechanical delay (EMD) reflecting excitation-contraction (E-C) coupling, and contractility were determined. RESULTS: Under normoxia, heart rate, QT duration, conduction, EMD, and ventricular shortening were similar in sham and paced hearts. During reoxygenation, arrhythmias ceased earlier and ventricular EMD recovered faster in paced hearts than in sham hearts. In sham hearts, BAY-K (but not verapamil), DIAZO (but not 5-hydroxydecanoate) or GLIB accelerated recovery of ventricular EMD, reproducing the pacing-induced protection. By contrast, none of these agents further ameliorated recovery of the paced hearts. CONCLUSION: The protective effect of chronic asynchronous pacing at near physiological rate on ventricular E-C coupling appears to be associated with subtle activation of L-type Ca(2+) channel, inhibition of sarcK(ATP) channel, and/or opening of mitoK(ATP) channel.

In vivo measurement of tissue oxygenation by time-resolved luminescence spectroscopy: advantageous properties of dichlorotris(1, 10-phenanthroline)-ruthenium(II) hydrate.

Measuring tissue oxygenation in vivo is of interest in fundamental biological as well as medical applications. One minimally invasive approach to assess the oxygen partial pressure in tissue (pO2) is to measure the oxygen-dependent luminescence lifetime of molecular probes. The relation between tissue pO2 and the probes' luminescence lifetime is governed by the Stern-Volmer equation. Unfortunately, virtually all oxygen-sensitive probes based on this principle induce some degree of phototoxicity. For that reason, we studied the oxygen sensitivity and phototoxicity of dichlorotris(1, 10-phenanthroline)-ruthenium(II) hydrate [Ru(Phen)] using a dedicated optical fiber-based, time-resolved spectrometer in the chicken embryo chorioallantoic membrane. We demonstrated that, after intravenous injection, Ru(Phen)'s luminescence lifetime presents an easily detectable pO2 dependence at a low drug dose (1 mg∕kg) and low fluence (120 mJ∕cm2 at 470 nm). The phototoxic threshold was found to be at 10 J∕cm2 with the same wavelength and drug dose, i.e., about two orders of magnitude larger than the fluence necessary to perform a pO2 measurement. Finally, an illustrative application of this pO2 measurement approach in a hypoxic tumor environment is presented.

Análisis comparativo de tres técnicas de derivación de células madre

Las células madre embrionarias (Embryonic Stem Cells; ESC) son células pluripotentes que presentan la capacidad de dividirse indefinidamente a la vez que mantienen la habilidad para diferenciarse a cualquier tipo celular. Aunque de manera rutinaria se derivan a partir de la masa celular interna de embriones en estadio de blastocisto, también pueden derivarse a partir de embriones en estadios precompactacionales y de embriones reconstruidos por procesos de transferencia nuclear. Debido a que durante el desarrollo embrionario temprano, momento en el que se derivan las ESC, tienen lugar profundos cambios de metilación en el genoma, tanto la derivación como el cultivo se consagran como técnicas que pueden alterar los patrones de metilación en genes regulados por impronta genómica. Con el objetivo de analizar la estabilidad epigenética de embriones preimplantacionales y ESC murinas, en este trabajo se ha optimizado un protocolo de anàlisis de los niveles de metilación mediante pirosecuenciación. Para ello se han seleccionado tres genes regulados por impronta genómica (H19/Igf2, Snrpn and Peg3), dos genes relacionados con el mantenimiento de pluripotencia en ESC (Oct4, Nanog y Sox2) y dos genes marcadores de diferenciación temprana (Cdx2 y Gata6). Nuestros resultados muestran que algunos grupos de embriones preimplantacionales presentan una hipo e hipermetilación en las regiones diferencialmente metiladas (Differentially Methylated Regions, DMRs) de los genes Snrpn y Peg3. Además, la línea de ESC analizada presentó anomalías en los tres genes regulados por impronta genómica. No obstante, el hecho de que esta línea fuera inestable a nivel cariotípico no permite establecer una relación entre el cultivo in vitro o la técnica de derivación y la inestabilidad epigenética demostrada. Por todo esto, parece pertinente analizar tanto la integridad epigenética como la estabilidad cromosómica de ESC antes de proceder a realizar ensayos clínicos en humanos.

Expression of substance P and preprotachykinin mRNA by primary sensory neurons in culture: regulation by factors present in peripheral and central target tissues.

Primary sensory neurons display various neuronal phenotypes which may be influenced by factors present in central or peripheral targets. In the case of DRG cells expressing substance P (SP), the influence of peripheral or central targets was tested on the neuronal expression of this neuropeptide. DRG cells were cultured from chick embryo at E6 or E10 (before or after establishment of functional connections with targets). Preprotachykinin mRNA was visualized in DRG cell cultures by either Northern blot or in situ hybridization using an antisense labeled riboprobe, while the neuropeptide SP was detected by immunostaining with a monoclonal antibody. In DRG cell cultures from E10, only 60% of neurons expressed SP. In contrast, DRG cell cultures performed at E6 showed a significant hybridization signal and SP-like immunoreactivity in virtually all the neurons (98%). The addition of extracts from muscle, skin, brain or spinal cord to DRG cells cultured at E6 reduced by 20% the percentage of neurons which express preprotachykinin mRNA and SP-like immunoreactivity. Our results indicate that factors issued from targets inhibit SP-expression by a subset of primary sensory neurons and act on the transcriptional control of preprotachykinin gene.

Biological characterization of clones derived from the edmonston strain of measles virus in comparison with schwarz and CAM-70 vaccine strains

Four virus clones were derived from the Edmonston strain of measles virus by repeated plaque purification. These clones were compared with the vaccine strains Schwarz and CAM-70 in terms of biological activities including plaque formation, hemagglutination, hemolysis and replication in Vero cells and chick embryo fibroblasts (CEF). Two clones of intermediate plaque yielded mixed plaque populations on subcultivation whereas the other two, showing small and large plaque sizes, showed stable plaque phenotypes. The vaccine strains showed consistent homogeneous plaque populations. All the Edmonston clones showed agglutination of monkey erythrocytes in isotonic solution while both vaccine strains hemagglutinated only in the presence of high salt concentrations. Variation in the hemolytic activity was observed among the four clones but no hemolytic activity was detected for the vaccine virus strains. Vaccine strains replicated efficiently both in Vero cells and CEF. All four clones showed efficient replication in Vero cells but different replication profiles in CEF. Two of them replicated efficiently, one was of intermediate efficiency and the other showed no replication in CEF. Two of the clones showed characteristics similar to vaccine strains. One in terms of size and homogeneity of plaques, the other for a low hemolytic activity and both for the efficiency of propagation in CEF.

Serum-Mediane von Placentaprotein-Markern: Relevanz der Unterschiede zwischen spontanen und nach-in-vitro-Fertilisation eingetretenen Schwangerschaften für das fetale Trisomie-Screening.

Trisomy-21 (Down syndrome) is the most frequent chromosomal abnorm- ality but only one third of cases would be detected by amniocentesis based on maternal age alone. Serum screening tests in the early second trimester increase the detec- tion rate to 60-65%, and more recently it was found that such screening was also possible in the first trimester by quantifying a diffe- rent panel of markers. The concen- trations of these placental proteins are strongly dependent on gestatio- nal age; thus control medians must be established and precise dating is essential. Serum chorionic gonado- trophin (HCG) levels were recently found to be increased in IVF preg- nancies compared to spontaneous gestations, leading to a falsely ele- vated trisomy screening risk. The aim of this preliminary study was to find out whether, in the first-trime- ster screening, the markers similarly differed between IVF and spontane- ous pregnancies which would call for the establishment of separate normal medians for IVF patients. We compared 24 pregnancies ob- tained after ovarian stimulation and IVF with six women after thawed embryo transfer (unstimulated cycle) and 63 gestation- and maternal-age matched spontaneously pregnant controls. A single serum was ob- tained between 6 and 16 weeks of gestation and various placental protein levels determined by im- munometric assays. Serum levels of pregnancy-associated plasma protein A (PAPP-A), which is the major marker in the first-trimes- ter screening test, were reduced in IVF pregnancies: after 9 weeks of gestation, multiples of median (MoMs) ranged between 0.23 and 3.58 (logarithmic mean 0.743). For the frozen/thawed transfers, this value was 1.08. In the 9-12 week group containing 6 cases of IVF, three thawed transfers and 25 con- trols, PAPP-A was significantly redu- ced in the stimulated compared to the nonstimulated cycles. In the late first and early second trimester the difference was not significant in our small group but the trend persisted. Pregnancies after IVF will thus show an increased incidence of false positive results in fetal trisomy-21 screening, and special medians should be established for these pati- ents.