HLA-A*02:687, a novel allele identified by sequence-based typing in cord blood from a Korean woman.

A*02:687 showed one nucleotide difference with A*02:01:01:01 resulting in an amino acid change.

Improvement of radiocephalic fistula maturation: rationale and design of the Liposomal Prednisolone to Improve Hemodialysis Fistula Maturation (LIPMAT) study - a randomized controlled trial.

BACKGROUND: Non-maturation is a frequent complication of radiocephalic arteriovenous fistulas (RCAVF). In an animal model, liposomal prednisolone improved maturation of experimental fistulas. The Liposomal Prednisolone to Improve Hemodialysis Fistula Maturation (LIPMAT) study investigates if liposomal prednisolone improves RCAVF maturation. METHODS AND RESULTS: The LIPMAT study is an investigator-initiated, multicenter, double-blinded, placebo-controlled randomized controlled trial with 1:1 randomization to liposomal prednisolone or placebo. Eighty patients receiving an RCAVF will be included. The primary outcome is the cephalic vein diameter six weeks after surgery, measured by ultrasound. The LIPMAT study started in May 2016. Enrollment is expected to be completed by the end of 2017. CONCLUSIONS: The LIPMAT study is the first to evaluate the efficacy of liposomal prednisolone to enhance RCAVF maturation.

The effects of formoterol on the serum, peritoneal VEGF, MDA, and VEGF levels in the ovaries and endometrium of rats with OHSS.

PURPOSE: To investigate the effects of formoterol (a beta2-adrenoreceptor agonist) on serum and peritoneal fluid VEGF, malondialdehyde (MDA) levels, and on VEGF-stained cell counts in the ovaries and endometrium of rats with ovarian hyperstimulation syndrome (OHSS) within the framework of immunohistochemical analysis. MATERIALS AND METHODS: A total of 28 immature female Wistar rats were randomly divided into four groups. Three groups were given ten IU pregnant mare serum gonadotropin/day on days 22-25 of life. They were administered 30 IU hCG on day 26 of life to mimic OHSS. On days 26 and 27 of life, 24 mcg/kg/day formoterol in group 3 and 48 mcg/kg formoterol in group 4 were administered intraperitoneally per animal. RESULTS: Although, there were no statistically significant differences between the groups in terms of serum and peritoneal fluid VEGF or MDA levels (serum VEGF: p = 0.28 1, peritoneal VEGF: p = 0.674, serum MDA: p = 0.543, peritoneal MDA: p = 0.506), there was a significant difference between the control and the OHSS placebo groups (p = 0.013) regarding the VEGF in the ovarian cortex. There was a significant difference between the control and the other groups in terms of ovarian stroma (p = 0.001), and there was also a statistically significant difference between the OHSS placebo and the other groups regarding VEGF in the endometrium (OHSS placebo vs. control group p = 0.002, OHSS placebo vs. the formoterol 24 mcg/kg group, p = 0.008, and OHSS-placebo vs. the formoterol 48 mcg/kg group, p = 0.001). CONCLUSIONS: Formoterol represents a potential novel strategy for the management of OHSS. Further studies, including those examining the dosage of formoterol, are warranted.

HLA-B*59:09, a novel allele identified by sequence-based typing in a cord blood donated by a Korean woman.

The new allele B*59:09 showed two nucleotide differences with B*59:01:01:01 in exon 3.

Ubiquitin-proteasome genes as targets for modulation of cisplatin sensitivity in fission yeast.

BACKGROUND: The ubiquitin(Ub)-proteasome pathway is implicated in the regulation of a variety of cellular functions and plays a major role in stress response in eukaryotic cells, by targeting misfolded and damaged proteins for degradation. In addition, in the presence of DNA damage, the Ub-proteasome system regulates proteins involved in sensing, repairing, and/or tolerating the damage. Antitumor agents such as cisplatin can activate the pathway, but the role of specific pathway components in cell sensitivity/response to the drug is not known. Since platinum compounds represent clinically relevant antitumor agents and a major limitation to their use is the development of drug resistance, there is an urgent need for identifying targets for improving their efficacy. RESULTS: In the present study, we performed a genome-wide screening for sensitivity to cisplatin using non-essential haploid deletion mutants of the fission yeast Schizosaccharomyces pombe, belonging to a collection of haploid strains constructed through homologous recombination. Using this approach, we identified three Ub-proteasome mutants exhibiting hypersensitivity to cisplatin (ubp16, ubc13 and pmt3) and ten mutants (including ufd2, beta7 20S, rpt6/let1) resistant to the drug. In addition, the importance of lub1 gene emerged from the comparison between the present screening and gene expression profile data previously obtained in fission yeast. CONCLUSIONS: The factors identified in the present study allowed us to highlight most finely the close relationship between the Ub-proteasome system and DNA damage response mechanisms, thus establishing a comprehensive framework of regulators likely relevant also in higher eukaryotes. Our results provide the proof of principle of the involvement of specific genes modulated by cisplatin treatment in cell response to the drug, suggesting their potential role as targets for modulating cisplatin sensitivity. In this regard, the prospective identification of novel targets for modulation of cisplatin sensitivity in an eukaryotic model organism appears particularly intriguing towards the discovery of strategies to overcome cisplatin resistance in human tumors.

Extragastrointestinal stromal tumor of the retroperitoneum mimicking kidney tumor.

A case of extragastrointestinal tumor of the retroperitoneum in a 48-year-old woman complaining right sided flank pain without hematuria is reported. The mass was excised from the kidney without a positive margin. The histopathological examination revealed an extragastrointestinal stromal tumor of the retroperitoneum. These tumors usually originate from the small intestine or stomach. On the other hand, they are rarely located in the retroperitoneum. These tumors typically exhibit CD117 immunoreactivity, whereas they may reveal CD34, neuron specific enolase, smooth muscle actin, desmin and S-100 protein. In our case the specimen of the patient was positive for CD117, actin and desmin.

Concomitant vagal neurofibroma and aplasia of the internal carotid artery in neurofibromatosis type 1.

We report the case of a patient with neurofibromatosis type 1 who had both aplasia of an internal carotid artery (ICA) and a vagal neurofibroma. To our knowledge, this is the first report in the literature of the simultaneous presence of these two rare disorders in a single patient. We believe that this is also the first report of an absence of an ICA in a patient with neurofibromatosis type 1. The patient was a 19-year-old woman who complained of a slowly growing neck mass. The mass occupied the right parapharyngeal space and upper cervical region. The patient had no other masses on physical examination, but widespread café au lait spots were evident. This led us to suspect the presence of a vagal neurofibroma. The tumor was removed, and pathology confirmed the diagnosis. No intracranial aneurysms were detected on cerebral angiography.

Involvement of p38 mitogen-activated protein kinase and apoptosis signal-regulating kinase-1 in nitric oxide-induced cell death in PC12 cells.

Although nitric oxide (NO) plays key signaling roles in the nervous systems, excess NO leads to cell death. In this study, the involvement of p38 mitogen-activated protein kinase (p38 MAPK) and apoptosis signal-regulating kinase-1 (ASK1) in NO-induced cell death was investigated in PC12 cells. NO donor transiently activated p38 MAPK in the wild type parental PC12 cells, whereas the p38 MAPK activation was abolished in NO-resistant PC12 cells (PC 12-NO-R). p38 MAPK inhibitors protected the cells against NO-induced death, whereas the inhibitors were not significantly protective against the cytotoxicity of reactive oxygen species. Stable transfection with dominant negative p38 MAPK mutant reduced NO-induced cell death. Stable transfection with dominant negative mutant of ASK1 attenuated NO-stimulated activation of p38 MAPK and decreased NO-induced cell death. These results suggest that p38 MAPK and its upstream regulator ASK1 are involved in NO-induced PC12 cell death.

Molecular cloning and cultivar specific expression of MAP kinases from Capsicum annuum.

Two MAP kinases, MK1 and MK2, were cloned from Capsicum annuum (pepper) cv. Subicho using a parsley MAP kinase gene as a heterologous probe. MK1 and MK2 encode stress-inducible protein kinases that can contribute to the response to wounding, UV-C, and cold. MK1 has a 92% amino acid identity with WIPK of tobacco. It was transcriptionally induced in response to wounding. In contrast, no detectable MK1 transcript was found in unwounded leaves of pepper. MK2 has a high level of amino acid homology to MAP kinases, such as NTF4 and SIPK and was constitutively expressed in all tissues. Both MK transcripts were downregulated by UV-C treatment. Each MK protein activation was independently wound-inducible in a cultivar dependent manner. MK1 is phosphorylated in cv. Pungchon but not cv. Subicho; whereas, the MK2 protein activation by wounding is restricted to cv. Subicho. In addition, de novo synthesis of the MK1 protein and tyrosine phosphorylation was rapidly and transiently induced in cv. Pungchon by wounding. In contrast, it is highly unlikely that the MK1 protein is produced in cv. Subicho, even though there is an abundant expression of MK1 mRNA after wounding in this cultivar. In Escherichia coli, which overexpresses MK1, autophosphorylation is observed at conserved threonine and tyrosine phosphorylation sites.

Transgenic rice plants expressing a Bacillus subtilis protoporphyrinogen oxidase gene are resistant to diphenyl ether herbicide oxyfluorfen.

Protoporphyrinogen oxidase (Protox), the penultimate step enzyme of the branch point for the biosynthetic pathway of Chl and hemes, is the target site of action of diphenyl ether (DPE) herbicides. However, Bacillus subtilis Protox is known to be resistant to the herbicides. In order to develop the herbicide-resistant plants, the transgenic rice plants were generated via expression of B. subtilis Protox gene under ubiquitin promoter targeted to the cytoplasm or to the plastid using Agrobacterium-mediated gene transformation. The integration and expression of the transgene were investigated at T0 generation by DNA and RNA blots. Most transgenic rice plants revealed one copy transgene insertion into the rice genome, but some with 3 copies. The expression levels of B. subtilis Protox mRNA appeared to correlate with the copy number. Furthermore, the plastidal transgenic lines exhibited much higher expression of the Protox mRNA than the cytoplasmic transgenic lines. The transgenic plants expressing the B. subtilis Protox gene at T0 generation were found to be resistant to oxyfluorfen when judged by cellular damage with respect to cellular leakage, Chl loss, and lipid peroxidation. The transgenic rice plants targeted to the plastid exhibited higher resistance to the herbicide than the transgenic plants targeted to the cytoplasm. In addition, possible resistance mechanisms in the transgenic plants to DPE herbicides are discussed.

Reciprocal regulation of HFE and NNamp2 gene expression by iron in human intestinal cells.

The newly identified hemochromatosis gene, HFE, and a candidate iron transporter gene, Nramp2, have been proposed as key factors responsible for the regulation of intestinal iron absorption. Although the exact functions of these proteins in intestinal iron absorption are unknown, HFE may be required for the down-regulation of iron absorption that occurs with increasing iron status, and Nramp2 may up-regulate iron absorption when iron status is low. Thus, we examined whether the expression of the HFE and Nramp2 genes are regulated by iron status in the human intestinal cell line Caco-2. HFE mRNA and HFE protein were increased and Nramp2 mRNA was decreased by increasing cellular iron status in Caco-2 cells. This iron-mediated modulation of mRNA levels was specific to iron. Moreover, super-induction of HFE mRNA in the presence of cycloheximide suggests that HFE gene expression may be controlled by a short-lived repressor protein. HFE and Nramp2 mRNA levels also changed in opposite directions during cellular differentiation. This reciprocal modification of the HFE and Nramp2 gene expression during both iron treatment and cell differentiation in Caco-2 cells is consistent with an opposing role for these proteins in homeostatic regulation of human intestinal iron absorption.

Recently identified molecular aspects of intestinal iron absorption.

Gene mapping techniques to identify gene mutations in humans and animals with phenotypic abnormalities in iron metabolism are providing important insights into the probable molecular mediators of intestinal iron absorption. Positional gene cloning in humans with hereditary hemochromatosis has identified a mutation in a novel major histocompatibility complex (MHC) gene called HFE that is likely to be involved in regulating intestinal iron absorption. In addition, recent observations based on positional cloning strategies in the mk/mk mouse and the Belgrade (b/b) rat rodent models of hypochromic, microcytic anemia have shown that the phenotypic abnormality in iron metabolism is associated with a mutation in the Nramp2 gene. Functional cloning studies in Xenopus oocytes have characterized DCT1 (Nramp2) as an iron-regulated proton-coupled divalent cation transporter. Nramp2 is likely to be the membrane transporter that functions in controlling iron entry across the apical membrane and in the export of iron out of endosomal vesicles. The observation that the expression of both HFE and Nramp2 mRNAs are reciprocally regulated by cellular iron status in Caco-2 cells, a human intestinal cell line, lends additional credence to the notion that these proteins may work in concert to regulate intestinal iron absorption.

Strain induces Caco-2 intestinal epithelial proliferation and differentiation via PKC and tyrosine kinase signals.

Although the intestinal epithelium undergoes complex deformations during normal function, nutrient absorption, fasting, lactation, and disease, the effects of deformation on intestinal mucosal biology are poorly understood. We previously demonstrated that 24 h of cyclic deformation at an average 10% deformation every 6 s stimulates proliferation and modulates brush-border enzyme activity in human intestinal Caco-2 cell monolayers. In the present study we sought potential mechanisms for these effects. Protein kinase C (PKC) activity increased within 1 min after initiation of cyclic deformation, and the PKC-alpha and -zeta isoforms translocated from the soluble to the particulate fraction. Cyclic deformation also rapidly increased tyrosine kinase activity. Tyrosine phosphorylation of several proteins was increased in the soluble fraction but decreased in the particulate fraction by cyclic deformation for 30 min. Inhibition of PKC and tyrosine kinase signals by calphostin C, G-06967, and erbstatin attenuated or blocked cyclic deformation-mediated modulation of Caco-2 DNA synthesis and differentiation. These results suggest that cyclic deformation may modulate intestinal epithelial proliferation and brush-border enzyme activity by regulating PKC and tyrosine kinase signals.

Generation of Resistance to the Diphenyl Ether Herbicide, Oxyfluorfen, via Expression of the Bacillus subtilis Protoporphyrinogen Oxidase Gene in Transgenic Tobacco Plants.

In an effort to develop transgenic plants resistant to diphenyl ether herbicides, we introduced the protoporphyrinogen oxidase (EC 1.3.3.4) gene of Bacillus subtilis into tobacco plants. The results from a Northern analysis and leaf disc assay indicate that the expression of the B. subtilis protoporphyrinogen oxidase gene under the cauliflower mosaic virus 35S promoter generated resistance to the diphenyl ether herbicide, oxyfluorfen, in transgenic tobacco plants.

Ascorbate offsets the inhibitory effect of inositol phosphates on iron uptake and transport by Caco-2 cells.

Differentiated monolayer cultures of Caco-2 human intestinal cells were used as a model to examine interactions between various dietary factors related to the intestinal uptake and absorption of nonheme Fe. Caco-2 cells accumulated 91-98 pmol Fe/mg protein from uptake buffer containing 12 nmol of Fe(III)-nitrilotriacetate during a 1-hr incubation at 37 degrees C. Addition of a 10-fold molar excess of inositol hexaphosphate (IP6) and its lesser phosphorylated derivatives (IP3, IP4, and IP5) decreased cellular uptake and transport of Fe from the lumenal compartment. Addition of ascorbic acid (AA) to the solution containing IPs stimulated Fe uptake and transport in a manner dependent upon the ratio of AA to IP and inversely proportional to the degree of phosphorylation of inositol (i.e., IP3 > IP4 > IP5 > IP6). A mixture of essential amino acids had minimal impact on Fe uptake in either the absence or presence of IPs. Cellular acquisition of Fe from solutions containing IPs was further enhanced by simultaneous addition of essential amino acids and AA. The stimulatory influence of ascorbic acid on Fe uptake from solutions containing IP6 was associated with an increase in the level of ferrous ion. These data further support the usefulness of Caco-2 cells as a model for investigating the effects of various dietary factors on mineral bioavailability.

Reduction of Fe(III) is required for uptake of nonheme iron by Caco-2 cells.

Differentiated cultures of Caco-2 human colonic cells were used to examine the importance of reduction of nonheme ferric iron, Fe(III), for transport across the brush border surface. Cultures accumulated approximately 100 pmol Fe/(h.mg protein) when 10 mumol Fe(III) as the nitrilotriacetic acid complex (1Fe:2NTA) was added to the apical compartment. Ascorbic acid enhanced cellular acquisition of iron in a dose-dependent manner, with a concentration as low as 8 mumol/L ascorbate increasing iron uptake by 50%. Similarly, the rate of iron transport from the apical to the basolateral compartment increased 5.6- and 30-fold when 100 and 1000 mumol/L ascorbic acid, respectively, were present in the apical chamber. Ascorbate-mediated stimulation of iron uptake was temperature dependent and required the reduction of Fe(III) to Fe(II), because it was inhibited by ascorbate oxidase and chelators of Fe(II). Moreover, Caco-2 cells recycled dehydroascorbic acid to ascorbic acid. Ferricyanide and Fe(II) chelators also partially inhibited iron uptake from a medium devoid of ascorbic acid. Intact Caco-2 cells exhibited a ferrireductase activity on the apical surface that accounted for the majority of iron accumulated by cells incubated in the absence of exogenous reductant. These data suggest that reduction of Fe(III) within the lumen or at the cell surface is required for transfer of this essential micronutrient across the intestinal brush border surface.

Inositol phosphates inhibit uptake and transport of iron and zinc by a human intestinal cell line.

To examine the influence of inositol phosphates on the uptake and absorption of Fe and Zn, Caco-2 cells were grown on either plastic (uptake studies) or porous membranes in bicameral chambers (transport/absorption studies). Caco-2, a human colon adenocarcinoma cell line, was selected as the test cell because it spontaneously differentiates into polarized enterocyte-like cells at confluency. Uptake of Fe (added as Fe-nitrilotriacetate complex) from a calcium-free solution by fully differentiated cells was 37 pmol/cm2. Addition of 10-fold molar excess of individual inositol phosphates (IP3, IP4, IP5 or IP6) decreased Fe solubility by 13 to 25% and reduced Fe uptake by 50 to 65%. The rate of transport of Fe from the apical solution into the basolateral chamber [1.4 +/- 0.1 pmol/(h.cm2)] decreased (34-96%) in proportion to the degree of phosphorylation of the inositol derivative in the apical compartment. Uptake and transepithelial transport of Zn were 246 +/- 5 pmol/cm2 and 23 +/- 1 pmol/(h.cm2), respectively. The solubility, uptake and rate of transport of Zn also decreased in proportion to the degree of phosphorylation of inositol. These results demonstrate the inhibitory influence of IP3-IP6 on the uptake and transport of Fe and Zn and support the usefulness of the Caco-2 human cell line as an appropriate model for evaluating the effects of specific dietary factors on trace metal bioavailability.

Mechanistic studies of the biosynthesis of 3,6-dideoxyhexoses in Yersinia pseudotuberculosis. Purification and characterization of CDP-6-deoxy-delta 3,4-glucoseen reductase based on its NADH:dichlorophenolindolphenol oxidoreductase activity.

CDP-6-deoxy-delta 3,4-glucoseen reductase, the key enzyme catalyzing the biosynthetic formation of CDP-ascarylose (CDP-3,6-dideoxy-L-arabino-hexose), was purified from Yersinia pseudotuberculosis by monitoring its NADH:dichlorophenolindolphenol oxidoreductase activity. A protocol consisting of DEAE-cellulose, phenyl-Sepharose, and Sephadex G-100 column chromatography yielded a mixture of two proteins. The low molecular weight protein contaminant was removed by limited tryptic digestion leaving a purified enzyme consisting of a single polypeptide with a molecular weight of 41,000. A weak, featureless uv spectrum above 300 nm suggested no common chromophoric cofactor contributes to enzyme activity and no protein-associated metals were detected. The stereospecificity of nicotinamide oxidation was determined to be pro-R stereospecific. Reduction of ferricyanide during NADH oxidation and confirmation of the intermediacy of O2- in the reaction flux suggested that enzyme-catalyzed H2O2 formation is not a direct two-electron reduction of molecular oxygen, but is rather the consequence of an enzymatic 2e-/1e- switch. The sugar deoxygenation reaction may therefore proceed through a radical mechanism.

Empleo de un nuevo antiinflamatorio no esteroideo en el tratamiento de lesiones deportivas. / Use of a new non-steroidal antiinflammatory for the treatment of athletic lesions

Cuarenta y nueve pacientes de diferentes equipos deportivos (lucha, karate, basketbol, futbol y otros) de la Universidad Nacional Autonoma de Mexico; 24 del sexo femenino y 25 del masculino, con edades entre 17 y 37 anos (media 24.9) que tenian lesiones de tejidos blandos o de articulaciones, fueron tratados con piroxicam, capsulas de 20 mg cada 12 horas por 2 dias y despues una capsula diaria hasta que desaparecieran los sintomas. Los resultados fueron como sigue: el dolor en reposo desaparecio en un tiempo promedio de 3.8 dias (N=29); dolor con movimientos activos en 5.4 dias (N=46); inflamacion en 4.4 dias (N= 42); limitacion de movimientos en 5.3 dias (N=26); debilidad muscular en 5.3 dias(N=29); dolor con movimientos pasivos en 4.4 dias (N= 43). En la autoevaluacion hecha por los pacientes, el dolor desaparecio en 4.8 dias (N= 49). Al final de los 12 dias de tratamiento, 46 pacientes estaban totalmente curados y 3 tenian mejoria clinica importante. Los efectos secundarios fueron minimos. Solo 2 pacientes se quejaron de dolor epigastrico ligero y no fue necesario suspender el tratamiento