Uterine fluid proteome changes during diapause and resumption of embryo development in roe deer (Capreolus capreolus).

The uterine microenvironment during pre-implantation presents a pro-survival milieu and is essential for embryo elongation in ruminants. The European roe deer (Careolus capreolus) pre-implantation embryo development is characterised by a 4-month period of reduced development, embryonic diapause, after which the embryo rapidly elongates and implants. We investigated the uterine fluid proteome by label-free liquid chromatography tandem mass spectrometry at four defined stages covering the phase of reduced developmental pace (early diapause, mid-diapause and late diapause) and embryo elongation. We hypothesised that embryo development during diapause is halted by the lack of signals that support progression past the blastocyst stage. Three clusters of differentially abundant proteins were identified by a self-organising tree algorithm: (1) gradual reduction over development; (2) stable abundance during diapause, followed by a sharp rise at elongation; and (3) gradual increase over development. Proteins in the different clusters were subjected to gene ontology analysis. 'Cellular detoxification' in cluster 1 was represented by alcohol dehydrogenase, glutathione S-transferase and peroxiredoxin-2. ATP-citrate synthase, nucleolin, lamin A/C, and purine phosphorylase as cell proliferation regulators were found in cluster 2 and 'cortical cytoskeleton', 'regulation of substrate adhesion-dependent cell spreading' and 'melanosome' were present in cluster 3. Cell cycle promoters were higher abundant at elongation than during diapause, and polyamines presence indicates their role in diapause regulation. This study provides a comprehensive overview of proteins in the roe deer uterine fluid during diapause and forms a basis for studies aiming at understanding the impact of the lack of cell cycle promoters during diapause.

More than just mothers: The neurobiological and neuroendocrine underpinnings of allomaternal caregiving.

In a minority of mammalian species, mothers depend on others to help raise their offspring. New research is investigating the neuroendocrine mechanisms supporting this allomaternal behavior. Several hormones have been implicated in allomaternal caregiving; however, the role of specific hormones is variable across species, perhaps because allomothering independently evolved multiple times. Brain regions involved in maternal behavior in non-human animals, such as the medial preoptic area, are also critically involved in allomaternal behavior. Allomaternal experience modulates hormonal systems, neural plasticity, and behavioral reactivity. In humans, fatherhood-induced decreases in testosterone and increases in oxytocin may support sensitive caregiving. Fathers and mothers activate similar neural systems when exposed to child stimuli, and this can be considered a global "parental caregiving" network. Finally, early work on caregiving by non-kin (e.g., foster parents) suggests reliance on similar mechanisms as biologically-related parents. This article is part of the 'Parental Brain and Behavior' Special Issue.

Early deprivation, atypical brain development, and internalizing symptoms in late childhood.

Children exposed to extreme early-life neglect such as in institutional rearing are at heightened risk for developing depression and anxiety disorders, and internalizing problems more broadly. These outcomes are believed to be due to alterations in the development of neural circuitry that supports emotion regulation. The specific neurodevelopmental changes that contribute to these difficulties are largely unknown. This study examined whether microstructural alterations in white matter pathways predicted long-term risk for internalizing problems in institutionally reared children. Data from 69 children were drawn from the Bucharest Early Intervention Project, a randomized clinical trial of foster care for institutionally reared children. White matter was assessed using diffusion tensor imaging (DTI) when children were between 8 and 10years of age. Internalizing symptoms were assessed at the time of the MRI scan, and once children reached 12-14years of age. Results indicated that neglect-associated alterations in the external capsule and corpus callosum partially explained links between institutional rearing status and internalizing symptoms in middle childhood and early adolescence. Findings shed light on neural mechanisms contributing to increased risk for emotional difficulties among children reared in adverse conditions and have implications for prevention and intervention.

Back table outflow graft anastomosis technique for HeartWare HVAD implantation.

The management of concomitant aortic and aortic valve disease with left ventricular assist device (LVAD) implantation for patients with severe cardiomyopathy is challenging, and has not been established given the complexity of LVAD surgery with concomitant aortic interventions. A 45-year-old patient presented to our institution with end-stage heart failure symptoms and non-ischemic cardiomyopathy. The patient was found to have a bicuspid aortic valve, severe native aortic regurgitation, a significant ascending aortic aneurysm, and severely depressed left ventricular (LV) function requiring two inotropes. He underwent a successful hemiarch repair of the ascending aortic aneurysm using a back table outflow graft anastomosis technique, and subsequent placement of a HeartWare Ventricular Assist Device (HVAD) with concomitant aortic valve closure with a modified Park's stitch. The patient did well postoperatively and is currently listed for heart transplantation.

Cryptococcal meningitis and SLE: a diagnostic and therapeutic challenge.

Cryptococcal meningitis is a rare occurrence in systemic lupus erythematosus (SLE). The risk factors of developing this infection are duration of SLE, intensity of glucocorticoid use, and SLE-related intrinsic immune abnormalities. Early recognition and prompt initiation of antifungals can prevent complications and improve survival. There is a dearth of evidence with regards to optimal treatment of cryptococcosis in non-HIV infected and non-transplant patients. The general consensus is to follow treatment guidelines for HIV-positive patients with cryptococcal meningitis. We describe a girl with active SLE and cryptococcal meningitis, and discuss the diagnostic and therapeutic challenges faced in this case.

Magnetic field dependent small-angle neutron scattering on a Co nanorod array: evidence for intraparticle spin misalignment.

The structural and magnetic properties of a cobalt nanorod array have been studied by means of magnetic field dependent small-angle neutron scattering (SANS). Measurement of the unpolarized SANS cross section dΣ/dΩ of the saturated sample in the two scattering geometries where the applied magnetic field H is either perpendicular or parallel to the wavevector ki of the incoming neutron beam allows one to separate nuclear from magnetic SANS, without employing the usual sector-averaging procedure. The analysis of the SANS data in the saturated state provides structural parameters (rod radius and centre-to-centre distance) that are in good agreement with results from electron microscopy. Between saturation and the coercive field, a strong field dependence of dΣ/dΩ is observed (in both geometries), which cannot be explained using the conventional expression of the magnetic SANS cross section of magnetic nanoparticles in a homogeneous nonmagnetic matrix. The origin of the strong field dependence of dΣ/dΩ is believed to be related to intradomain spin misalignment, due to magnetocrystalline and magnetoelastic anisotropies and magnetostatic stray fields.

First-trimester placental protein 13, PAPP-A, uterine artery Doppler and maternal characteristics in the prediction of pre-eclampsia.

OBJECTIVE: To test the hypothesis that a combination of PP13, PAPP-A and first-trimester uterine artery Doppler would improve the prediction of pre-eclampsia. METHODS: This is a prospective cohort study of pregnant women followed from the first-trimester to delivery. PP13 and PAPP-A were determined by immunoassay of maternal serum at 11-14 weeks', when uterine artery Doppler measurements were assessed. Cases identified with any form of pre-eclampsia were compared with a control group without pre-eclampsia. The sensitivity of each marker or their combinations in predicting pre-eclampsia for different fixed false positive rates was calculated from the ROC curves. RESULTS: Forty two women were diagnosed with pre-eclampsia and 410 women with pregnancies not complicated by pre-eclampsia were used as controls. For a fixed false positive rate (FPR) of 20%, PP13, PAPP-A and mean uterine artery pulsatility index identified 49%, 58% and 62% respectively, of women who developed any form of pre-eclampsia. PP13 was best in predicting early onset pre-eclampsia with a sensitivity of 79% at a 20% FPR. Combinations of the three first-trimester assessments did not improve the prediction of pre-eclampsia in later pregnancy. CONCLUSION: First-trimester PP13, PAPP-A and uterine artery PI are reasonable, individual predictors of women at risk to develop pre-eclampsia. Combinations of these assessments do not further improve the prediction of pre-eclampsia.

Regulation of the plant-type 5'-adenylyl sulfate reductase by oxidative stress.

5'-Adenylyl sulfate (APS) reductase (EC 1.8.4.9) catalyzes a key reaction in the plant sulfate assimilation pathway leading to the synthesis of cysteine and the antioxidant glutathione. In Arabidopsis thaliana APS reductase is encoded by a family of three genes. In vitro biochemical studies revealed that the enzyme product derived from one of them (APR1) is activated by oxidation, probably through the formation of a disulfide bond. The APR1 enzyme is 45-fold more active when expressed in a trxB strain of Escherichia coli than in a trxB(+) wild type. The enzyme is inactivated in vitro by treatment with disulfide reductants and is reactivated with thiol oxidants. Redox titrations show that the regulation site has a midpoint potential of -330 mV at pH 8.5 and involves a two-electron redox reaction. Exposure of a variety of plants to ozone induces a rapid increase in APS reductase activity that correlates with the oxidation of the glutathione pool and is followed by an increase in free cysteine and total glutathione. During the response to ozone, the level of immunodetectable APS reductase enzyme does not increase. Treatment of A. thaliana seedlings with oxidized glutathione or paraquat induces APS reductase activity even when transcription or translation is blocked with inhibitors. The results suggest that a posttranslational mechanism controls APS reductase. A model is proposed whereby redox regulation of APS reductase provides a rapidly responding, self-regulating mechanism to control the glutathione synthesis necessary to combat oxidative stress.

HIV clinical trials in correctional settings: right or retrogression?

The right of incarcerated prison and jail inmates to health care is protected by the 8th and the 14th amendments of the Constitution, respectively. Does the right to health care include access to clinical trials? At the time of this writing, clinical trials have become part of the fabric of HIV/AIDS care, allowing patients to participate in studies of new and often lifesaving treatments. Participation in trials can also be dangerous, as illustrated by the recent death of a subject in a gene therapy trial. This danger is compounded by ethical dilemmas that can arise from the large amount of financial support for clinical trials (greater than 75%) that is derived from for-profit corporations. Indeed, clinical trials are the subject of grave concern on the part of the United States Government, which has recently taken steps to shore up human subject safeguards. Following a conference on the conduct of clinical trials in correctional settings, the Office for Human Research Protections suspended prison research conducted by 4 prestigious academic institutions.

Identification of a new class of 5'-adenylylsulfate (APS) reductases from sulfate-assimilating bacteria.

A gene was cloned from Burkholderia cepacia DBO1 that is homologous with Escherichia coli cysH encoding 3'-phosphoadenylylsulfate (PAPS) reductase. The B. cepacia gene is the most recent addition to a growing list of cysH homologs from a diverse group of sulfate-assimilating bacteria whose products show greater homology to plant 5'-adenylylsulfate (APS) reductase than they do to E. coli CysH. The evidence reported here shows that the cysH from one of the species, Pseudomonas aeruginosa, encodes APS reductase. It is able to complement an E. coli cysH mutant and a cysC mutant, indicating that the enzyme is able to bypass PAPS, synthesized by the cysC product. Insertional knockout mutation of P. aeruginosa cysH produced cysteine auxotrophy, indicating its role in sulfate assimilation. Purified P. aeruginosa CysH expressed as a His-tagged recombinant protein is able to reduce APS, but not PAPS. The enzyme has a specific activity of 5.8 micromol. min(-1). mg of protein(-1) at pH 8.5 and 30 degrees C with thioredoxin supplied as an electron donor. APS reductase activity was detected in several bacterial species from which the novel type of cysH has been cloned, indicating that this enzyme may be widespread. Although an APS reductase from dissimilatory sulfate-reducing bacteria is known, it shows no structural or sequence homology with the assimilatory-type APS reductase reported here. The results suggest that the dissimilatory and assimilatory APS reductases evolved convergently.

Cloning of a cDNA coding for an amino acid carrier from Ricinus communis (RcAAP1) by functional complementation in yeast: kinetic analysis, inhibitor sensitivity and substrate specificity.

A cDNA for the amino acid permease gene RcAAP1 has been isolated from Ricinus communis by yeast complementation and subjected to a detailed kinetic analysis. RcAAP1 cDNA is 1.5 kb with an open reading frame that codes for a protein with 486 amino acids and a calculated molecular mass of 53.1 kDa. RcAAP1-mediated histidine uptake was pH dependent with highest transport rates at acidic pH; it was sensitive to protonophores and uncouplers and the Km for histidine uptake was 96 microM. The substrate specificity was investigated by measuring the levels of inhibition of histidine uptake by a range of amino acids. The basic amino acids (histidine, lysine and arginine) showed strongest inhibition of uptake whereas acidic amino acids competed less effectively. Alanine was the most efficient competitor of the neutral amino acids. Glutamine, serine, asparagine, methionine and cysteine showed moderate inhibition whereas threonine, isoleucine, leucine, phenylalanine, tyrosine and tryptophan showed only low levels of inhibition. Glycine, proline and citrulline caused slight stimulation. More detailed competition kinetics indicated that both lysine and arginine showed simple competitive inhibition of histidine uptake. When direct uptake measurements were carried out, both lysine and arginine were found to be effective substrates for RcAAP1.

Expression analysis of a sucrose carrier in the germinating seedling of Ricinus communis.

This study describes the expression of a sucrose carrier at various developmental stages in Ricinus communis. A partial-length cDNA clone, RcSUT1, was isolated by RT-PCR from Ricinus seedling RNA. This is almost identical to a sucrose carrier cDNA, Rscr1, which has previously been isolated by library screening. However, we have observed a very different expression pattern in the seedling to that previously reported. Northern analysis, with RcSUT1 as a probe, revealed high expression of a 2 kb transcript in the cotyledons of the germinating seedling; transcript levels were similar in cotyledons harvested 3-6 days after germination. A much lower level of this transcript was detected in the root, hypocotyl and endosperm RNA of the seedling and very low levels were also present in the sink and source leaves of the mature plant. This pattern of expression was also reflected at the protein level with an antipeptide antibody raised to part of the RcSUT1 deduced amino acid sequence. Tissue print hybridisation analysis of the hypocotyl revealed that the sucrose carrier transcripts were localised to the phloem cells of the vascular bundles. A more detailed analysis of sucrose carrier gene expression in the cotyledons of the germinating seedling was carried out by in situ hybridisation; the strongest signals were observed from the lower epidermal layer and the phloem, consistent with an active loading role for these cells. An ultrastructural study of the cells in the lower epidermis showed that they have wall ingrowths which are characteristic of transfer cells. The results are discussed in relation to the physiological role of the sucrose carrier in the Ricinus seedling and to the pathways of sucrose movement from endosperm to the sieve elements in the cotyledons.

Glutaredoxin function for the carboxyl-terminal domain of the plant-type 5'-adenylylsulfate reductase.

5'-Adenylylsulfate (APS) reductase (EC 1.8.99.-) catalyzes the reduction of activated sulfate to sulfite in plants. The evidence presented here shows that a domain of the enzyme is a glutathione (GSH)-dependent reductase that functions similarly to the redox cofactor glutaredoxin. The APR1 cDNA encoding APS reductase from Arabidopsis thaliana is able to complement the cysteine auxotrophy of an Escherichia coli cysH [3'-phosphoadenosine-5'-phosphosulfate (PAPS) reductase] mutant, only if the E. coli strain produces glutathione. The purified recombinant enzyme (APR1p) can use GSH efficiently as a hydrogen donor in vitro, showing aKm[GSH] approximately of 0.6 mM. Gene dissection was used to express separately the regions of APR1p from amino acids 73-327 (the R domain), homologous with microbial PAPS reductase, and from amino acids 328-465 (the C domain), homologous with thioredoxin. The R and C domains alone are inactive in APS reduction, but the activity is partially restored by mixing the two domains. The C domain shows a number of activities that are typical of E. coli glutaredoxin rather than thioredoxin. Both the C domain and APR1p are highly active in GSH-dependent reduction of hydroxyethyldisulfide, cystine, and dehydroascorbate, showing a Km[GSH] in these assays of approximately 1 mM. The R domain does not show these activities. The C domain is active in GSH-dependent reduction of insulin disulfides and ribonucleotide reductase, whereas APR1p and R domain are inactive. The C domain can substitute for glutaredoxin in vivo as demonstrated by complementation of an E. coli mutant, underscoring the functional similarity between the two enzymes.

Amino acid carriers of Ricinus communis expressed during seedling development: molecular cloning and expression analysis of two putative amino acid transporters, RcAAP1 and RcAAP2.

This study reports on the isolation of two putative amino acid carrier cDNAs, RcAAP1 and RcAAP2, from Ricinus communis. Northern analysis shows that RcAAP1 and RcAAP2 are expressed abundantly in the cotyledon and root tissues of developing seedlings and at lower levels in the endosperm and hypocotyl. In the mature plant low expression was observed in the source and sink leaves. We have further characterized the expression of RcAAP1 in Ricinus roots by in situ hybridization. The transcripts are localized in many cell types of the root tip region, including the epidermal and cortical cells, but the highest expression was observed in the cells of the stele situated adjacent to the xylem poles. This is the first report describing the cellular expression of an amino acid transporter in roots, and the results are discussed in relation to the physiological role of this transporter.

Plant sulfur metabolism--the reduction of sulfate to sulfite.

Until recently the pathway by which plants reduce activated sulfate to sulfite was unresolved. Recent findings on two enzymes termed 5'-adenylylsulfate (APS) sulfotransferase and APS reductase have provided new information on this topic. On the basis of their similarities it is now proposed that these proteins are the same enzyme. These discoveries confirm that the sulfate assimilation pathway in plants differs from that in other sulfate assimilating organisms.

Biochemical and molecular characterization of sucrose and amino acid carriers in Ricinus communis.

The use of energized plasma membrane vesicle preparations from cotyledons and roots of Ricinus communis seedlings is described, and evidence is presented for the existence of plasma membrane H(+)/sucrose and H(+)/amino acid symporters. Using fractions isolated from roots, there is evidence for at least two carriers which can transport neutral amino acids and one which can also transport basic amino acids. A method for the solubilization and reconstitution of glutamine transport activity is described. Preliminary results on the molecular characterization of two putative amino acid carriers and a putative sucrose transporter from Ricinus are presented.

AIDS: Part II.

Great strides have been made in the therapy of human immunodeficiency virus (HIV) infection. Currently approved drugs include zidovudine and didanosine. A third drug, dideoxycytidine (zalcitibine), has recently been filed for approval with the Food and Drug Administration. All these drugs work through inhibition of the reverse transcriptase enzyme. Zidovudine is the only drug that has shown clinical efficacy against HIV. Treatment of patients with advanced HIV disease (i.e., acquired immune deficiency syndrome [AIDS] or symptomatic infection with < 200 CD4+ lymphocytes per mm3), results in a prolongation and improved quality of life. Zidovudine is the only antiretroviral agent approved for the treatment of asymptomatic patients. Early intervention with zidovudine has been shown to delay progression to AIDS when patients' CD4+ lymphocyte counts decline to less than 500/mm3, irrespective of clinical signs or symptoms of HIV infection. Didanosine is currently indicated for the treatment of patients with advanced HIV disease who are intolerant to or failing zidovudine therapy. The major toxicity of zidovudine is bone marrow suppression with anemia and granulocytopenia (which occurs in from 1% to 45% of patients, depending on the clinical stage of disease and the dose of the drug). Didanosine and zalcitibine have both been associated with a severe peripheral neuropathy, which is generally reversible on cessation of the drug. In addition, didanosine has been implicated as a cause of pancreatitis that has been fatal in a small percentage of cases. The toxicities of didanosine and zalcitibine range from 1% to 10%, depending on dose, duration of therapy, and the presence of underlying HIV-related peripheral neuropathy or a previous history of pancreatitis. The clinical hallmark of HIV infection is the development of opportunistic infections and malignancies, which are a consequence of the profound immunodeficiency. The risk of an opportunistic infection increases significantly as the T-helper lymphocyte count declines to less than 20%, or 200 to 250/mm3. The spectrum of opportunistic infections ranges from viruses to protozoa. Patients with advanced HIV disease are also at increased risk of infection with nonopportunistic, community-acquired pathogens. Primary and secondary prophylaxis against the most common AIDS-defining opportunistic infection, Pneumocystis carinii pneumonia, is now recommended. Studies are currently underway to determine the efficacy of prophylaxis against other opportunistic pathogens. Treatment of opportunistic infections associated with AIDS has improved significantly over the past 5 years as new drugs and combination regimens of antimicrobials have been developed.(ABSTRACT TRUNCATED AT 400 WORDS)