GRP78 as a marker of pre-eclampsia: an exploratory study

Although the exact mechanisms that lead to shallow invasion or defective trophoblastic differentiation in pre-eclampsia are still unknown, it is widely admitted that the etiology of pre-eclampsia is a defect in trophoblast invasion of the uterine spiral arteries. We have previously observed that the status of a chaperone protein, glucose regulated protein 78 (GRP78) is associated with the invasive properties of cytotrophoblastic cells; we therefore hypothesized that circulating GRP78 could serve as a diagnostic tool in pre-eclampsia. In a prospective case-control study, we quantified GRP78 autoantibodies, complexes of GRP78 with autoantibodies and GRP78 (C-term fragment, N-term fragment and full-length GRP78) by ELISA. Plasma from women diagnosed with pre-eclampsia (n = 16), from women during the first trimester of pregnancy who subsequently developed pre-eclampsia (n = 10) and from healthy pregnant women (controls, n = 58 at term, n = 26 at first trimester) were analysed and compared. We observed no significant difference between pre-eclamptic and healthy pregnant women for autoantibodies-GRP78 complexes or total GRP78 at both first trimester and at delivery. In contrast, the ratio of C-terminal GRP78 over full length GRP78 was significantly different in plasma of pre-eclamptic patients as compared with controls both during first trimester (P < 0.004) and at term (P < 0.0001). Our findings suggest that circulating C-terminal GRP78 reflect the invasive properties of cells, and could be used as a predictive marker for pre-eclampsia early in pregnanc

The NA48 LKr calorimeter digitizer electronics chain

The 13 500 channels of the NA48 liquid-krypton electromagnetic calorimeter readout electronics were put into operation in 1997. The digitizer electronics employs a new gain switching technique that expands the dynamic range of a standard 10-bit ADC to 14 bits at 40 MHz sampling rate employing a custom-developed integrated circuit (KRYPTON). The KRYPTON has been fabricated in 1.2 μm BiCMOS technology and was successfully developed together with industry on a short timescale. The performance and the experience from the first year of the operation of the liquid-krypton calorimeter electronics will also be briefly discussed

The NA48 LKr calorimeter readout electronics

The NA48 experiment at the CERN SPS accelerator is making a measurement of the direct CP violation parameter $\epsilon'/\epsilon$ by comparing the four rates of decay of $K_{S}$ and $K_{L}$ into $2\pi^{0}$ and $\pi^{+}\pi^{-}$. To reconstruct the decays into $2\pi^{0}$ the information from the almost 13500 channels of a quasi-homogeneous liquid krypton electromagnetic calorimeter is used. The readout electronics of the calorimeter has been designed to provide a dynamic range from a few MeV to about 50 GeV energy deposition per cell, and to sustain a high rate of incident particles. The system is made by cold charge preamplifiers (working at 120 degrees K), low-noise fast shapers followed by digitizer electronics at 40 MHz sampling rate that employs a gain switching technique to expand the dynamic range, where the gain can be selected for each sample individually (i.e. every 25 ns). To reduce the amount of data collected the system contains a zero suppression circuit based on halo expansion

Putative Role of the Aldo-Keto Reductase from Trypanosoma cruzi in Benznidazole Metabolism

Fil: Garavaglia, Patricia Andrea. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Parasitología; Argentina.Fil: Cannata, Joaquín J B. Instituto de Investigaciones Biotecnológicas, Universidad Nacional de General San Martín-CONICET; Argentina.Fil: Laverriere, Marc. Instituto de Investigaciones Biotecnológicas, Universidad Nacional de General San Martín-CONICET; Argentina.Fil: Garcia, Gabriela Andrea. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Parasitología; Argentina.Benznidazole (Bz), the drug used for treatment of Chagas' disease (caused by the protozoan Trypanosoma cruzi), is activated by a parasitic NADH-dependent type I nitroreductase (NTR I). However, several studies have shown that other enzymes are involved. The aim of this study was to evaluate whether the aldo-keto reductase from T. cruzi (TcAKR), a NADPH-dependent oxido-reductase previously described by our group, uses Bz as the substrate. We demonstrated that both recombinant and native TcAKR enzymes reduce Bz by using NADPH, but not NADH, as a cofactor. TcAKR-overexpressing epimastigotes showed higher NADPH-dependent Bz reductase activity and a 50% inhibitory concentration (IC50) value for Bz 1.8-fold higher than that of the controls, suggesting that TcAKR is involved in Bz detoxification instead of activation. To understand the role of TcAKR in Bz metabolism, we studied TcAKR expression and NADPH/NADH-dependent Bz reductase activities in two T. cruzi strains with differential susceptibility to Bz: CL Brener and Nicaragua. Taking into account the results obtained with TcAKR-overexpressing epimastigotes, we expected the more resistant strain, Nicaragua, to have higher TcAKR levels than CL Brener. However, the results were the opposite. CL Brener showed 2-fold higher TcAKR expression and 5.7-fold higher NADPH-Bz reduction than the Nicaragua strain. In addition, NADH-dependent Bz reductase activity, characteristic of NTR I, was also higher in CL Brener than in Nicaragua. We conclude that although TcAKR uses Bz as the substrate, TcAKR activity is not a determinant of Bz resistance in wild-type strains and may be overcome by other enzymes involved in Bz activation, such as NADPH- and NADH-dependent reductases

Trypanosoma cruzi: death phenotypes induced by ortho-naphthoquinone substrates of the aldo-keto reductase (TcAKR). Role of this enzyme in the mechanism of action of β-lapachone

Fil: Garavaglia, Patricia A ANLIS Dr.C.G.Malbrán. Instituto Nacional de Parasitología; Argentina.Fil: Rubio, María Fernanda. Laboratorio de Biología Molecular y Apoptosis,Instituto de Investigaciones Médicas Alfredo Lanari (IDIM-CONICET),Universidad de Buenos Aires,Ciudad de Buenos Aires (1427); Argentina.Fil: Laverrière, Marc. Instituto de Investigaciones Biotecnológicas (IIB-INTECH),Universidad Nacional de General San Martín-CONICET,San Martín (1650),Prov. Buenos Aires; Argentina.Fil: Tasso, Laura Mónica. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Parasitología; Argentina.Fil: Fichera, Laura E. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Parasitología; Argentina.Fil: Cannata, Joaquín J B. Instituto de Investigaciones Biotecnológicas (IIB-INTECH),Universidad Nacional de General San Martín-CONICET,San Martín (1650),Prov. Buenos Aires; Argentina.Fil: García, Gabriela Andrea. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Parasitología; Argentina.Several ortho-naphthoquinones (o-NQs) have trypanocidal activity against Trypanosoma cruzi, the aetiological agent of Chagas disease. Previously, we demonstrated that the aldo-keto reductase from this parasite (TcAKR) reduces o-NQs, such as β-lapachone (β-Lap) and 9,10-phenanthrenequinone (9,10-PQ), with concomitant reactive oxygen species (ROS) production. Recent characterization of TcAKR activity and expression in two T. cruzi strains, CL Brener and Nicaragua, showed that TcAKR expression is 2.2-fold higher in CL Brener than in Nicaragua. Here, we studied the trypanocidal effect and induction of several death phenotypes by β-Lap and 9,10-PQ in epimastigotes of these two strains. The CL Brener strain was more resistant to both o-NQs than Nicaragua, indicating that greater TcAKR activity is unlikely to be a major influence on o-NQ toxicity. Evaluation of changes in ROS production, mitochondrial membrane potential, phosphatidylserine exposure and monodansylcadaverine labelling evidenced that β-Lap and 9,10-PQ induce different death phenotypes depending on the combination of drug and T. cruzi strain analysed. To study whether TcAKR participates in o-NQ activation in intact parasites, β-Lap and 9,10-PQ trypanocidal effect was next evaluated in TcAKR-overexpressing parasites. Only β-Lap was more effective and induced greater ROS production in TcAKR-overexpressing epimastigotes than in controls, suggesting that TcAKR may participate in β-Lap activation

High rate large dynamic range analog circuitry and digitizers for fast calorimeter

This paper describes the issues encountered in the design of a large dynamic range digitizer for calorimetry. A gain switching technology developed for the liquid krypton calorimeter of the NA48 CP-violation experiment at CERN is described in detail. 14 bit dynamic range and better than 300 ps time resolution were achieved in beam tests with an 8 channel prototype. A mixed analog/digital ASIC has been developed. A 13,500 channel system is being produced which will be put into operation in 1996

Supplementary Material for: Identification and Analysis of Two Novel Sites of Rat GnRH Receptor Gene Promoter Activity: The Pineal Gland and Retina

<b><i>Background and Aims:</i></b> In mammals, activation of pituitary GnRH receptor (GnRHR) by hypothalamic GnRH increases the synthesis and secretion of LH and FSH, which, in turn, regulate gonadal functions. However, GnRHR gene <i>(Gnrhr)</i> expression is not restricted to the pituitary. <b><i>Methods:</i></b> To gain insight into the extrapituitary expression of <i>Gnrhr</i>, a transgenic mouse model that expresses the human placental alkaline phosphatase reporter gene driven by the rat <i>Gnrhr</i> promoter was created. <b><i>Results:</i></b> This study shows that the rat <i>Gnrhr</i> promoter is operative in two functionally related organs, the pineal gland, as early as embryonic day (E) 13.5, and the retina where activity was only detected at E17.5. Accordingly, <i>Gnrhr</i> mRNA were present in both tissues. Transcription factors known to regulate <i>Gnrhr</i> promoter activity such as the LIM homeodomain factors LHX3 and ISL1 were also detected in the retina. Furthermore, transient transfection studies in CHO and gonadotrope cells revealed that OTX2, a major transcription factor in both pineal and retina cell differentiation, is able to activate the <i>Gnrhr</i> promoter together with either CREB or PROP1, depending on the cell context. <b><i>Conclusion:</i></b> Rather than using alternate promoters, <i>Gnrhr</i> expression is directed to diverse cell lineages through specific associations of transcription factors acting on distinct response elements along the same promoter. These data open new avenues regarding GnRH-mediated control of seasonal and circadian rhythms in reproductive physiology