Humanization of a highly stable single-chain antibody by structure-based antigen-binding site grafting.

The murine single-chain variable fragment F8 (scFv(F8)) is endowed with high intrinsic thermodynamic stability and can be functionally expressed in the reducing environment of both prokaryotic and eukaryotic cytoplasm. The stability and intracellular functionality of this molecule can be ascribed mostly to its framework regions and are essentially independent of the specific sequence and structure of the supported antigen-binding site. Therefore, the scFv(F8) represents a suitable scaffold to construct stable scFv chimeric molecules against different antigens by in vitro evolution or antigen-binding site grafting.

Induction of parturition with daily low-dose oxytocin injections in pregnant mares at term: clinical applications and limitations.

The aim of the present study was to evaluate the clinical applications and limitations of daily low-dose oxytocin injections for the induction of parturition in pregnant at term mares, the attention was focussed on the efficacy of the treatment and on its possible negative effects on mare and foal. Three-hundred and fifty pregnant full term Standardbred mares were used: 176 were allowed to foal spontaneously, 174 were treated daily with 3.5 IU of oxytocin i.

The effect of selecting oocytes for insemination and transferring all resultant embryos without selection on outcomes of assisted reproduction.

Objective: To evaluate the effect of selecting three best oocytes for insemination and transfer of all resultant embryos on outcomes of assisted reproductive technologies (ART).

Study Design: Comparative study.

Setting: The Center of Reproductive Medicine, Arcispedale Santa Maria Nuova, Reggio Emilia, Italy.

Atf2 transcription factor binds to the APP1 promoter in Cryptococcus neoformans: stimulatory effect of diacylglycerol.

The fungus Cryptococcus neoformans is an environmental human pathogen which enters the lung via the respiratory tract and produces a unique protein, called antiphagocytic protein 1 (App1), that protects it from phagocytosis by macrophages. In previous studies, we proposed genetic evidences that transcription of APP1 is controlled by the enzymatic reaction catalyzed by inositol phosphorylceramide synthase 1 (Ipc1) via the production of diacylglycerol through the activating transcription factor 2 (Atf2). We investigated here the mechanism by which Atf2 binds to the APP1 promoter in vitro and in vivo.

Pharmacokinetics of oral fenretinide in neuroblastoma patients: indications for optimal dose and dosing schedule also with respect to the active metabolite 4-oxo-fenretinide.

Purpose: Pharmacokinetic data on fenretinide (4-HPR) are scant, thus limiting the rational use of the drug. We investigated the pharmacokinetics of 4-HPR and its active metabolite 4-oxo-fenretinide (4-oxo-4-HPR).

Experimental Design: Pharmacokinetics were assessed in 18 children (3 for each dose) with neuroblastoma who received oral 4-HPR once daily for 28 days at the doses of 100, 300, 400, 600, 1,700 and 4,000 mg/m(2)/day.

Validation of a stable recombinant antibodies repertoire for the direct selection of functional intracellular reagents.

We have previously generated a semi-synthetic single-chain variable fragments (scFv) phage display library built on a thermodynamically stable single-framework scaffold. All scFv antibodies selected from this repertoire showed high thermodynamic stability and were expressed as soluble molecules in bacterial cytoplasm. In this work, two complementary methodologies have been adopted to assess the functionality of library-derived scFvs as intracellular antibodies and to verify the possibility to directly use this repertoire for the selection of antibodies able to function in a reducing environment.

The effect of legislation on outcomes of assisted reproduction technology: lessons from the 2004 Italian law.

Objective: To evaluate the effect of the 2004 Italian regulations (insemination of
Design: Case-control study.

Setting: The Center of Reproductive Medicine, Arcispedale Santa Maria Nuova, Reggio Emilia, Italy.

Influence of age and foaling on plasma protein electrophoresis and serum amyloid A and their possible role as markers of equine neonatal septicaemia.

A field study was undertaken to investigate plasma protein electrophoresis (PPE) and serum amyloid A (SAA) concentrations at 1, 3 and 7 days of age in foals born by oxytocin-induced (group A, n =17) or spontaneous (group B, n =12) parturition. The putative diagnostic role of PPE and SAA in four septicaemic foals was also examined. At birth, beta-gamma-globulins were higher in group B, and then increased in both groups, probably due to colostrum intake.

Why a simple model of genetic regulatory networks describes the distribution of avalanches in gene expression data.

In a previous study it was shown that a simple random Boolean network model, with two input connections per node, can describe with a good approximation (with the exception of the smallest avalanches) the distribution of perturbations in gene expression levels induced by the knock-out of single genes in Saccharomyces cerevisiae. Here we address the reason why such a simple model actually works: we present a theoretical study of the distribution of avalanches and show that, in the case of a Poissonian distribution of outgoing links, their distribution is determined by the value of the Derrida exponent. This explains why the simulations based on the simple model have been effective, in spite of the unrealistic hypothesis about the number of input connections per node.

Dynamics of HIV viral load in blood and semen of patients under HAART: impact of therapy in assisted reproduction procedures.

We examined the efficacy and effect of HAART in HIV-1-infected men confronted with assisted fertilization procedures. We showed that HAART did not always reduce the HIV-1-RNA level in blood and semen compartments, and that a significant upward shift in mitochondrial DNA was observed in spermatozoa from a HAART-treated patient group compared with spermatozoa from HAART-untreated or HIV-1-uninfected groups (P < 0.001).

Analysis of gene expression identifies PLAB as a mediator of the apoptotic activity of fenretinide in human ovarian cancer cells.

Fenretinide (4-HPR) is a synthetic retinoid with antitumor activity, which induces apoptosis in cancer cell lines of different histotypes. To identify genes contributing to its apoptotic activity in ovarian cancer cells, we monitored, by cDNA arrays, gene expression changes after 4-HPR exposure in A2780, a human ovarian carcinoma cell line sensitive to the retinoid. Among the differentially expressed transcripts, PLAcental Bone morphogenetic protein (PLAB), a proapoptotic gene, was the most highly induced.

Effects of mating on plasma concentrations of testosterone, cortisol, oestrone sulphate and 15-ketodihydro-PGF2alpha in stallions.

Very little information is available regarding the physiological mechanisms involved in the normal sexual activity in the stallion and, in particular, the endocrine control of reproduction is still not clearly understood. This experiment was designed to determine the short-term effect of sexual stimulation on plasma concentrations of testosterone, cortisol, oestrone sulphate and 15-ketodihydro-PGF(2alpha) in stallions. Semen samples were collected from 10 lighthorse stallions of proven fertility using a Missouri model artificial vagina.

Outcome of 518 salvage oocyte-cryopreservation cycles performed as a routine procedure in an in vitro fertilization program.

Objective: To document outcomes of oocyte cryopreservation performed as a routine procedure in an IVF program.

Design: Describing the rate of oocyte survival, embryo transfer (ET), implantation, and live births of IVF-intracytoplasmic sperm injection performed on thawed oocytes.

Setting: Reproductive medicine center in Italy.

New insights on the use of desipramine as an inhibitor for acid ceramidase.

Treatment of different cancer cell lines with desipramine induced a time- and dose-dependent downregulation of acid ceramidase. Desipramine's effect on acid ceramidase appeared specific for amphiphilic agents (desipramine, chlorpromazine, and chloroquine) but not other lysomotropic agents such as ammonium chloride and bafilomycin A1, and was not transcriptionally regulated. The cathepsin B/L inhibitor, CA074ME, but not the cathepsin D inhibitor, pepstatin A, blocked desipramine's effect on acid ceramidase.

Sex pheromone of the cranberry root grub Lichnanthe vulpina.

The cranberry root grub Lichnanthe vulpina (Hentz) (Coleoptera: Glaphyridae) is a pest of cranberries in Massachusetts, reducing yield and vine density. (Z)-7-Hexadecenol and (Z)-7-hexadecenal were identified from the female effluvia collection by gas chromatographic-electroantennographic detection and gas chromatography-mass spectrometry. The double-bond position was confirmed by dimethyl disulfide derivatization.

Pregnancy loss and assisted reproduction: preliminary results after the law 40/2004 in Italy.

The new Italian law on assisted reproduction technology rules that no more than three oocytes can be fertilized at one time, and that all embryos obtained must be transferred to the maternal uterus simultaneously. The aim of the present study was to investigate the influence of the new law on spontaneous embryonic/fetal losses through comparison of data collected over an identical period of time: the first year of application of the new law compared with the same time period 1 year before (March 10, 2004 to March 9, 2005 versus March 10, 2003 to March 9, 2004). A total of 271 clinical pregnancies were analysed.

Effect of the mode of assisted reproductive technology conception on obstetric outcomes for survivors of the vanishing twin syndrome.

Survivors of the "vanishing" twin syndrome, which occurred in dichorionic twins only, and singletons that began as singletons, had similar mean gestational duration and birth weights, as well as similar frequencies of maternal and neonatal complications. This similarity persisted when conventional IVF and IVF plus ICSI cases were separately evaluated.

4-oxo-fenretinide, a recently identified fenretinide metabolite, induces marked G2-M cell cycle arrest and apoptosis in fenretinide-sensitive and fenretinide-resistant cell lines.

4-oxo-N-(4-hydroxyphenyl)retinamide (4-oxo-4-HPR) is a recently identified metabolite of fenretinide (4-HPR). We explored the effectiveness of 4-oxo-4-HPR in inducing cell growth inhibition in ovarian, breast, and neuroblastoma tumor cell lines; moreover, we investigated the molecular events mediating this effect in two ovarian carcinoma cell lines, one sensitive (A2780) and one resistant (A2780/HPR) to 4-HPR. 4-oxo-4-HPR was two to four times more effective than 4-HPR in most cell lines, was effective in both 4-HPR-sensitive and 4-HPR-resistant cells, and, in combination with 4-HPR, caused a synergistic effect.

Trapping Phyllophaga spp. (Coleoptera: Scarabaeidae: Melolonthinae) in the United States and Canada using sex attractants.

The sex pheromone of the scarab beetle, Phyllophaga anxia, is a blend of the methyl esters of two amino acids, L-valine and L-isoleucine. A field trapping study was conducted, deploying different blends of the two compounds at 59 locations in the United States and Canada. More than 57,000 males of 61 Phyllophaga species (Coleoptera: Scarabaeidae: Melolonthinae) were captured and identified.

Spontaneous embryonic loss rates in twin and singleton pregnancies after transfer of top- versus intermediate-quality embryos.

Objective: To determine whether embryo quality is associated with early spontaneous loss rates in twin and singleton pregnancies after IVF/intracytoplasmic sperm injection (ICSI).

Design: Retrospective, single center analysis.

Setting: The Center of Reproductive Medicine, Arcispedale Santa Maria Nuova, Reggio Emilia, Italy.

Plasma concentrations of 15-ketodihydro-PGF(2alpha), cortisol and progesterone during manual twin reduction in thoroughbred mares.

The aim of the present study was to highlight the effect of two different techniques of one embryo crushing on some hormonal changes. Ten twinning mares were submitted to the mobile or fixed manual crushing of one blastocyst within day 19 after the last mating. Blood sample was collected from 20 min before to 90 min, 24 and 72 h after the procedure was performed to analyse 15-ketodihydro-PGF(2alpha), cortisol and progesterone plasma concentrations.

Lower embryonic loss rates among twin gestations following assisted reproduction.

Purpose: To determine whether maternal age and number of transferred embryos influence early pregnancy losses in twin pregnancies compared to singletons following IVF/ICSI.

Methods: We compared the pregnancy loss rates in singleton (n = 549) and twin (n = 252) gestations, stratified by maternal age (< or = 35 and > 35 years) and the number of transferred embryos (1-3 and 4-9).

Results: Loss rates of singleton pregnancies were significantly higher than that in twins (OR 3.

Immunomodulation of cucumber mosaic virus infection by intrabodies selected in vitro from a stable single-framework phage display library.

Immunomodulation by the ectopic expression of intracellular antibodies ('intrabodies') has a great potential for interfering with physiological or pathological functions in vivo in a highly specific manner. One of the major obstacles of this technology is the inability of most antibodies to properly fold and function in the reducing environment of the cytoplasm, which prevents the formation of essential disulfide bonds. We wished to assess the intracellular performance of antibodies derived from a semi-synthetic single-chain variable fragment (scFv) phage display library ('F8 library') built on a thermodynamically stable single-framework scaffold.

Uncertainty in rate loss corrections based on counting a periodic pulser.

Tomographic gamma scanning of waste produces three-dimensional transmission and emission images. These are used to derive item-specific attenuation correction factors that improve the accuracy of non-destructive waste assay. For each vertical layer, data grabs of short duration are acquired as the waste item is rotated and translated.