bluuebunny/crossref_metadata_2025 huggingface dataset:
Conformational changes upon binding of a receptor loop to lipid structures: possible role in signal transduction
Authors: Pertinhez Thelma A. , Nakaie Clóvis R. , Carvalho Regina S.H. , Paiva Antonio C.M. , Tabak Marcel , Toma Flavio , Schreier Shirley
Published: July 2002

The mas oncogene codes for a seven transmembrane helix protein. The amino acid sequence 253–266, from the third extracellular loop and beginning of helix 7, was synthesized either blocked or carrying an amino acid spin label at the N‐terminus. Peptide binding to bilayers and micelles was monitored by ESR, fluorescence and circular dichroism. Binding induced tighter lipid packing, and caused an increase of peptide secondary structure. While binding to bilayers occurred only when peptide and phospholipid bore opposite charges, in micelles the interaction took place irrespective of charge. The results suggest that changes in lipid packing could modulate conformational changes in receptor loops related to the triggering of signal transduction.
Gastrin induces tyrosine phosphorylation of Shc proteins and their association with the Grb2/Sos complex
Authors: Seva Catherine , Kowalski-Chauvel Aline , Blanchet Jean-Sébastien , Vaysse Nicole , Pradayrol Lucien
Published: July 2002

Gastrin/CCKB G protein‐coupled receptors have been shown to mediate proliferative effects of their endogenous ligands. In the present study, we examined the signal transduction mechanisms linked to the G/CCKB receptor occupancy. We report here that gastrin stimulates MAP kinase activation in a dose‐ and time‐dependent manner, a pathway known to play a key role in cell proliferation. We also characterized the molecular events, upstream of p21‐Ras, that may link the MAP kinase pathway to G/CCKB receptors. Gastrin induced a rapid and transient increase in tyrosine phosphorylation of several proteins including the 2 isoforms (46 and 52 kDa) of the adaptor protein She. Phosphorylated Shc subsequently associated with a complex that includes Grb2 and the p21‐Ras activator, Sos. Our results also indicate that Sos becomes phosphorylated in response to gastrin as shown by a reduction in electrophoretic mobility of the protein. Tyrosine phosphorylation of Shc and subsequent complex formation with Grb2 and Sos appear to be a common mechanism by which tyrosine kinase receptors and the G/CCKB G protein‐coupled receptor stimulate the Ras‐dependent MAP kinase pathway.
Role of arachidonic acid metabolism in thymocyte apoptosis after irradiation
Authors: Korystov Yuri N. , Dobrovinskaya Oksana R. , Shaposhnikova Vera V. , Eidus Lazar Kh.
Published: July 2002

The present study demonstrates that DNA fragmentation, nuclear pycnosis and trypan blue staining of irradiated thymocytes is prevented by inhibition of the lipoxygenase pathway of arachidonic acid metabolism and is not affected by cyclooxygenase inhibition. Exposed to irradiation [3H]arachidonic acid‐labeled thymocytes release radioactive products to the external medium. The process is blocked by the lipoxygenase inhibitor, nordihydroguaiaretic acid. Thus, it can be concluded that irradiation activates arachidonic acid metabolism and that lipoxygenase metabolites play an important role in thymocyte apoptosis.
Ethanol neuronotoxicity in the embryonic chick brain in ovo and in culture: Interaction of the neural cell adhesion molecule (NCAM)
Authors: Kentroti S. , Rahman H. , Grove J. , Vernadakis A.
Published: July 2002

The present study was undertaken to investigate the involvement of NCAM in the neuroteratogenic effects of ethanol demonstrated by us and others. In the first experiment we examined the effect of in‐ovo ethanol exposure on expression of NCAM in various regions of the embryonic CNS throughout development. Chick embryos received ethanol (10 mg/50 μl/day) or saline (control) at days 1–3 of development (E1–E3), were sacrificed at various embryonic ages and whole brain (WB), cerebral hemispheres (CH) and cerebellum (CE) processed for SDS‐polyacrylamide gel electrophoresis. The normal developmental profile of NCAM in the chick brain exhibited the same dynamics as previously reported by others. When compared to age‐matched control brains, an increase was observed in expression of high molecular weight forms of NCAM in cerebral hemispheres between E8 and E10. These bands represented highly sialated (> 180 kDa) forms of NCAM. In fact, the NCAM band from ethanol‐treated embryos at E8 migrated at a higher molecular weight than did its control counterpart, indicating an increase in sialic acid content. In contrast, no clear change was observed in NCAM expression in cerebellum from E10 through E20 as a result of ethanol exposure. In the second experiment, we examined the involvement of NCAM in the alterations in neuronal growth patterns observed in ethanol‐exposed cultures. Neuroblast‐enriched cultures derived from three‐day‐old whole chick embryos (E3WE) were maintained on poly‐l‐lysine pre‐coated Petri dishes in DMEM + 5% fetal bovine serum with or without 50 mM ethanol. Cultures were fixed at 3, 6 or 9 DIV and co‐stained for NCAM and neurofilament (160 kDa). E3WE cultures exhibited intense NCAM immunoreactivity at 3 and 6 DIV decreasing by 9 DIV. NCAM positive structures included all neuronal perikarya, neuritic processes and growth cones. Addition of 50 mM ethanol to the medium resulted in profound alterations in growth patterns of developing neurons which continued to exhibit intense NCAM staining. Ethanol‐induced changes in the developmental profile of NCAM expression (i.e. increased sialation) in cerebral hemispheres correspond temporally with the shift in neuronal phenotype from cholinergic to catecholaminergic and GABAergic which we have reported previously. Changes in the normal pattern of cellular contact and interaction as a result of altered NCAM expression may influence establishment of neurotransmitter phenotype. Findings from this study support the view that NCAM may be involved both directly and indirectly in shaping of the CNS during development and we speculate that ethanol neuroembryotoxicity uncouples this relationship.
Definition of therapy-resistant schizophrenia and its assessment
Authors: Brenner HD , Merlo MCG
Published: July 2002

The treatment of nonresponsive schizophrenia poses perennial problems. The situation is made yet more complex when the great variety of patient types, their social background, family circumstances and psychosocial requirements are taken into account. The comparative lack of response to neuroleptic therapy seen in some patients, especially those with minimal structural brain changes, further complicates the problem, provoking—as such therapy may—more EPS than are normally seen in patients without more obvious physical changes in brain structure. Until recently, the treatment of nonresponsive schizophrenia has tended to involve switching from one neuroleptic (usually a “highpotent” agent) to another of different chemical structure. However, as these “typical” neuroleptics occupy D2 receptors, the rationale for using a less conventional antipsychotic, such as clozapine, is increasingly appreciated in the management of the treatment-refractory schizophrenic.
The effect of reactive oxygen species generated from the mitochondrial electron transport chain on the cytochrome c oxidase activity and on the cardiolipin content in bovine heart submitochondrial particles
Authors: Paradies Giuseppe , Petrosillo Giuseppe , Pistolese Marilva , Ruggiero Francesca Maria
Published: July 2002

The effect of reactive oxygen species (ROS), produced by the mitochondrial respiratory chain, on the activity of cytochrome c oxidase and on the cardiolipin content in bovine heart submitochondrial particles (SMP) was studied. ROS were produced by treatment of succinate‐respiring SMP with antimycin A. This treatment resulted in a large production of superoxide anion, measured by epinephrine method, which was blocked by superoxide dismutase (SOD). Exposure of SMP to mitochondrial mediated ROS generation, led to a marked loss of cytochrome c oxidase activity and to a parallel loss of cardiolipin content. Both these effects were completely abolished by SOD+catalase. Added cardiolipin was able to almost completely restore the ROS‐induced loss of cytochrome c oxidase activity. No restoration was obtained with peroxidized cardiolipin. These results demonstrate that mitochondrial mediated ROS generation affects the activity of cytochrome c oxidase via peroxidation of cardiolipin which is needed for the optimal functioning of this enzyme complex. These results may prove useful in probing molecular mechanism of ROS‐induced peroxidative damage to mitochondria which have been proposed to contribute to aging, ischemia/reperfusion and chronic degenerative diseases.
Characterization of Arabidopsis AtAMT2, a novel ammonium transporter in plants
Authors: Sohlenkamp Christian , Shelden Megan , Howitt Susan , Udvardi Michael
Published: July 2002

We have cloned and characterized the first member of a novel family of ammonium transporters in plants: AtAMT2 from Arabidopsis thaliana. AtAMT2 is more closely related to bacterial ammonium transporters than to plant transporters of the AMT1 family. The protein was expressed and functionally characterized in yeast. AtAMT2 transported ammonium in an energy‐dependent manner. In contrast to transporters of the AMT1 family, however, AtAMT2 did not transport the ammonium analogue, methylammonium. AtAMT2 was expressed more highly in shoots than roots and was subject to nitrogen regulation.
Effect of the sugar chain of soluble recombinant CD59 on complement inhibitory activity
Authors: Suzuki Hiroshi , Yamaji Noboru , Egashira Akira , Yasunaga Kunio , Sugita Yuji , Masuho Yasuhiko
Published: July 2002

A soluble recombinant CD59#77 (rCD59#77), consisting of 77 amino acids starting from the N terminus of membrane‐bound CD59, was prepared using a gene expression system in CHO cells. The rCD59#77 preparation was composed of glycosylated and non‐glycosylated forms (G and NG forms). Unexpectedly, NG form was 7 times more potent than G form in complement inhibitory activity. Postulating that sialic acids on G‐form molecules make it difficult for rCD59#77 to access nascent membrane attack complexes on the cell surface, the sialic acids were removed by neuraminidase treatment. However, the inhibitory activity was not changed. Next, one of two putative N‐glycosylation sites was mutated by substituting Gln18 for Asn18. The mutant, designated rCD59#77(N/Q), had no sugar moiety and was as active as the NG form of rCD59#77. These results suggest that the bulky sugar moiety at Asn18 is not necessary for the complement‐inhibitory activity of rCD59 and actually hampers that function.
Specific binding of [3H]GppNHp to extracellular membrane receptors in chick cerebellum: possible involvement of kainic acid receptors
Authors: Ramos Milagros , Souza Diogo O , Ramı́rez Galo
Published: July 2002

Guanine nucleotides (GNs), including GMP, displace [3H]kainic acid binding to chick cerebellar lysed and vesiculated membranes. Saturation studies of [3H]GppNHp binding, under conditions that prevent the occupation of the nucleotide binding sites in G‐proteins, demonstrate the existence of extracellular membrane receptors specific for guanine nucleotides. Affinity‐labeling of a vesicle preparation with [α‐32P]GTP gives one single labeled band, upon electrophoresis, with an apparent molecular mass of 50 kDa. Additional experiments with partially purified kainate receptors suggest that the GN extracellular sites may overlap, at least partially, the kainic acid binding sites, being then responsible for the displacement of [3H]kainic acid by GNs. The physiological significance of these findings remains unclear.
In vitro expressed dystrophin fragments do not associate with each other
Authors: Chan Yiu-mo , Kunkel Louis M.
Published: July 2002

Dystrophin, a component of the muscle membrane cytoskeleton, is the protein altered in Duchenne Muscular Dystrophy (DMD) and Becker Muscular Dystrophy (BMD). Dystrophin shares significant homology with other cytoskeletal proteins, such as α‐actinin and spectrin. On the basis of its sequence similarity with α‐actinin and spectrin, dystrophin has been proposed to function as dimer. However, the existence of both dimers and monomers have been observed by electron microscopy. To address this apparent discrepancy, we expressed dystrophin fragments composed of different domains in an in vitro translation system. The expressed fragments were tested for their ability to interact with each other and full‐length dystrophin by both immunoprecipitation and blot overlay assays. These assays were successfully used to demonstrate the dimerization of α‐actinin and spectrin, yet failed to detect any interaction between dystrophin fragments. Although these in vitro results do not prove that dystrophin is not a dimer in vivo, they do indicate that this interaction is not like that of the α‐actinin and spectrin.
Escherichia coliccm in‐frame deletion mutants can produce periplasmic cytochrome b but not cytochrome c
Authors: Throne-Holst Mimmi , Thöny-Meyer Linda , Hederstedt Lars
Published: July 2002

Escherichia coli CcmA, CcmB and CcmC polypeptides are required for cytochrome c synthesis and are thought to constitute the subunits of an ABC‐type transporter as judged from sequence data. Using a periplasmic reporter system based on Bacillus subtilis cytochrome c‐550 and E. coli cytochrome b‐562 we show that the synthesis of the b‐type cytochrome in the periplasm is normal in E. coli ccmA and ccmC in‐frame deletion mutants. Mutants deleted for ccmF or ccmG encoding a component of a putative cytochrome c‐heme lyase and a membrane bound thioredoxin‐like protein, respectively, have the same phenotype. The ccm mutants produce cytochrome c‐550 polypeptide, but not holocytochrome c. Taken together the results demonstrate that heme can be transported to the periplasm by a ccm‐independent mechanism.
Thermodynamic and spectral properties of DNA miniduplexes with the terminal G·A mispairs and 3′ or 5′ dangling bases
Authors: Lokhov S.G , Pyshnyi D.V
Published: July 2002

The tetradeoxyribonucleotide pAGCG in 1 M NaCl forms duplexes with terminal non‐canonical pA·G pairs with stability significantly exceeding that for the duplex (pAGCT)2 and lower than that for the duplex (pCGCG)2. The deoxyriboduplex (pAGCG)2 is considerably stabilized by 3′‐ and slightly by 5′‐ dangling bases. Therefore, the stability of duplexes with 3′ dangling bases decreases in the order (pAGCG)2>(pCGCG)2>(pAGCT)2. The sum of the independent stabilizing effect of the of 5′‐p and 3′‐ dangling bases on the (pAGCG)2 core duplex is higher than that of the additional terminal pG·A pair in pG‐A‐/‐G‐A tandem of the duplex (pAGCG)2.
Carboxy‐terminal truncation of long‐tailed amyloid β‐peptide is inhibited by serine protease inhibitor and peptide aldehyde
Authors: Hamazaki Hideaki
Published: July 2002

The 42/43‐residue amyloid β‐peptide (Aβ) is widely believed to play a major role in Alzheimer's disease. The present study shows that the rat brain contains a carboxypeptidase that efficiently deletes three amino acids from Aβ1–43. The carboxypeptidase activity in the brain was completely inhibited by 1 mM phenylmethylsulfonyl fluoride, suggesting the protease is a serine carboxypeptidase. The carboxy‐terminal truncation of Aβ1–43 was moderately inhibited by carbobenzoxy‐Leu‐leucinal, carbobenzoxy‐Leu‐Leu‐leucinal, and carbobenzoxy‐Leu‐Leu‐norvalinal, and weakly by antipain. The present data suggest that the serine carboxypeptidase contributes to the generation of short‐tailed Aβ peptides and is important in the intracellular clearance of Aβ1–42/43 in brains.
Subunit exchange of lens α‐crystallin: a fluorescence energy transfer study with the fluorescent labeled αA‐crystallin mutant W9F as a probe
Authors: Sun Tian-Xiao , Akhtar Nila J , Liang Jack J.-N
Published: July 2002

A Trp‐free αA‐crystallin mutant (W9F) was prepared by site‐directed mutation. This mutant appears to be identical to the wild‐type in terms of conformation (secondary and tertiary structures). W9F was labeled with a sulfhydryl‐specific fluorescent probe, 2‐(4′‐maleimidylanilino) naphthalene‐6‐sulfonate (MIANS), and used in a subunit exchange between αA‐ and αA‐crystallins as well as between αA‐ and αB‐crystallins, studied by measurement of fluorescence resonance energy transfer. Energy transfer was observed between Trp (donor, with emission maximum at 336 nm) of wild‐type αA‐ or αB‐crystallin and MIANS (acceptor, with absorption maximum at 313 nm) of labeled W9F when subunit exchange occurred. Time‐dependent decrease of Trp and increase of MIANS fluorescence were recorded. The exchange was faster at 37°C than at 25°C. The energy transfer efficiency was greater between homogeneous subunits (αA‐αA) than between heterogeneous subunits (αA‐αB). A previous exchange study with isoelectric focusing indicated a complete but slow exchange between αA and αB subunits. The present study showed that the exchange was a fast process, and the different energy transfer efficiencies between αA‐αA and αA‐αB indicated that αA‐ and αB‐crystallins were not necessarily structurally equivalent.
Glycerol diversifies phage repertoire selections and lowers non‐specific phage absorption
Authors: Kjær Svend , Stausbøl-Grøn Brian , Wind Troels , Ravn Peter , Hobolt Jensen Kristian , Kahns Liselotte , Clark Brian F.C
Published: July 2002

Using a semi‐synthetic phage displayed antibody repertoire, isoform‐specific and cross‐reactive phage‐antibodies to eukaryotic elongation factor 1A (eEF1A) have been selected. Enrichment of specific antibodies was found to depend on the presence of glycerol. Further selections against lactate dehydrogenase (LDH) revealed that the dominance of a phage‐antibody clone to LDH was inhibited by glycerol, a notable feature for selection strategies where a broad variety of binding clones is desired. The impact of glycerol in distinct steps of the selection protocol was examined and glycerol found to affect certain antibody‐antigen interactions. Furthermore, the non‐specific phage binding was lowered by three orders of magnitude at a 20% (v/v) glycerol concentration.
Activation of nitric oxide synthase is involved in tamoxifen‐induced apoptosis of human erythroleukemia K562 cells
Authors: Maccarrone Mauro , Fantini Cristina , Ranalli Marco , Melino Gerry , Finazzi Agrò Alessandro
Published: July 2002

Tamoxifen induces apoptosis (programmed cell death) in human erythroleukemia K562 cells. Nitric oxide synthase activity and expression increased in apoptotic cells by 315% and 280%, respectively, compared to controls. The specific inhibitor of nitric oxide synthase, l‐NAME, protected K562 cells from tamoxifen‐induced apoptosis, whereas the nitric oxide donor, sodium nitroprusside (SNP), potentiated the apoptotic effect of the drug. In addition, 5‐lipoxygenase was activated by tamoxifen and the specific enzyme inhibitor, MK886, protected K562 cells against the drug. Conversely, the 5‐lipoxygenase product, 5‐hydroperoxyeicosatetraenoic acid, enhanced the tamoxifen‐induced apoptosis. Finally, tamoxifen altered also membrane properties of K562 cells.
7‐Deazaxanthine, a novel prototype inhibitor of thymidine phosphorylase
Authors: Balzarini Jan , Gamboa Antonio Esteban , Esnouf Robert , Liekens Sandra , Neyts Johan , De Clercq Erik , Camarasa Maria-José , Pérez-Pérez Maria-Jesus
Published: July 2002

7‐Deazaxanthine (7DX) was identified as a novel inhibitor of thymidine (dThd) phosphorylase (TPase). It inhibited the TPase reaction in a concentration‐dependent manner. At 1 mM, it almost completely prevented the TPase‐catalysed hydrolysis of dThd to thymine. The 50% inhibitory concentration (IC50) of 7DX was 40 μM in the presence of 100 μM of the natural substrate dThd. 7DX is also endowed with a marked inhibitory effect on angiogenesis. It significantly prevents neovascularisation in the chicken chorioallantoic membrane during development. 7DX is the first purine derivative shown to be a potent inhibitor of purified TPase and angiogenesis.
Production and characterization of monoclonal antibodies directed against native epitopes of NhaA, the Na+/H+ antiporter of Escherichia coli
Authors: Padan Etana , Venturi Miro , Michel Hartmut , Hunte Carola
Published: July 2002

Monoclonal antibodies (mAbs) recognizing native epitopes are an important tool for functional and structural studies of proteins, yet they have rarely been used with transport proteins. In an attempt to raise monoclonal antibodies against the NhaA Na+/H+ antiporter of Escherichia coli we encountered difficulties in the screening procedure, which is based on the standard enzyme‐linked immunosorbent assay (ELISA). Here we report a rapid and efficient method of screening for anti‐NhaA mAbs which recognize native epitopes of the antiporter. The method is based on the use of His‐tagged protein, Ni2+‐nitrilotriacetic acid coated plates and non‐denaturing conditions in the assay. With this procedure four mAbs were obtained, three of which recognize the NhaA in its native conformation and one preferentially recognizes the denatured form. The latter mAb is Western blot positive, the others are Western blot negative and bind the detergent solubilized NhaA as assayed by gel filtration chromatography. Competition experiments show that the native epitopes are common to both the His‐tagged and the wild‐type protein. We suggest that in the standard ELISA the NhaA protein is not presented to the antibody in the native conformation whereas the His tag based protocol favors this presentation. Indeed, we could remarkably improve the low reactivity of the standard ELISA by coating the plates with anti‐NhaA mAb and use it to present NhaA (‘sandwich' ELISA or two antibodies assay). Remarkably, two of the mAbs (5H4, 2C5) which bind native NhaA inhibit drastically the ΔpH driven 22Na uptake mediated by His‐tagged NhaA reconstituted in proteoliposomes. Hence, these mAbs afford a new tool to study the structure/function relationship of the antiporter.
Immunohistochemical localization of 5‐oxo‐l‐prolinase, an enzyme of the γ‐glutamyl cycle, in porcine brain microvessels
Authors: Jäger Martina , Weber Peter , Wolf Sabine
Published: July 2002

The immunohistochemical analysis of the distribution of 5‐oxo‐l‐prolinase in porcine brain at the light microscopic level was performed with an antibody raised against the enzyme purified from pig kidney. The present study reveals the specific expression of 5‐oxo‐l‐prolinase in brain capillaries with an average diameter of 4.1±0.9 μm, while larger blood vessels remain unstained. Porcine kidney and skeletal muscle show no endothelial‐specific staining with the antibody. In some cases, the asymmetrical staining pattern in cross and longitudinal sections of brain microvessels indicate endothelial‐ but also pericyte‐specific expression.
Pax6 and Cdx2/3 form a functional complex on the rat glucagon gene promoter G1‐element
Authors: Andersen Frank Gellert , Heller R.Scott , Petersen Helle Vestergård , Jensen Jan , Madsen Ole Dragsbæk , Serup Palle
Published: July 2002

α‐cell specific transcription of the glucagon gene is mainly conferred by the glucagon promoter G1‐element, while additional elements G2, G3, and G4 have broad islet cell specificity. Transcription of the glucagon gene has been shown to be stimulated by Pax6 through binding to the glucagon gene promoter G3‐element. In this report, we show that Pax6 additionally binds the glucagon gene promoter G1‐element and forms a transcriptionally active complex with another homeodomain protein, Cdx2/3. Two distinct mutations in the G1‐element, that both reduce promoter activity by 85–90%, is shown to eliminate binding of either Pax6 or Cdx2/3. Additionally, Pax6 enhanced Cdx2/3 mediated activation of a glucagon reporter in heterologous cells. We discuss how Pax6 may contribute to cell‐type specific transcription in the pancreatic islets by complex formation with different transcription factors.
Surface display of functional fibronectin‐binding domains on Staphylococcus carnosus
Authors: Liljeqvist Sissela , Cano François , Nguyen Thien Ngoc , Uhlén Mathias , Robert Alain , Ståhl Stefan
Published: July 2002

The surface expression in Staphylococcus carnosus of three different fibronectin binding domains (FNBDs), derived from fibronectin binding proteins of Streptococcus dysgalactiae and Staphylococcus aureus, has been investigated. Surface localization of the chimeric proteins containing the FNBDs was demonstrated. All three surface‐displayed FNBDs were demonstrated to bind fibronectin in whole‐cell enzyme‐linked binding assays. Furthermore, for one of the constructs, intranasal immunizations with the recombinant bacteria resulted in improved antibody responses to a model immunogen present within the chimeric surface proteins. The implications of the results for the design of live bacterial vaccine delivery systems are discussed.
The biological significance of non‐enzymatic reaction of menadione with plasma thiols: enhancement of menadione‐induced cytotoxicity to platelets by the presence of blood plasma
Authors: Chung Sun-Hwa , Chung Seung-Min , Lee Joo-Young , Kim Se-Ryun , Park Ki-Sook , Chung Jin-Ho
Published: July 2002

To test the hypothesis that the non‐enzymatic reaction of quinones with thiols in plasma can generate reactive oxygens (ROS), thereby leading to potentiated cellular toxicity, we have studied the effect of a representative quinone compound, menadione, on plasma isolated from rats. The experimental results are as follows: (1) menadione generated ROS via non‐enzymatic reaction with protein thiols in plasma; (2) the presence of plasma increased menadione‐induced cytotoxicity to platelets; (3) pretreatment of plasma with a thiol‐depleting agent significantly suppressed menadione‐induced ROS and cytotoxicity. These results suggest that the non‐enzymatic reaction of menadione with plasma thiols could be an important process in quinone‐induced cellular toxicity.
Neuronal nAChR stereoselectivity to non‐natural epibatidine derivatives
Authors: Bertrand Sonia , Patt Jörg T , Spang Jörg E , Westera Gerrit , Schubiger P.August , Bertrand Daniel
Published: July 2002

The frog toxin epibatidine is one of the most powerful ligands of the neuronal nicotinic receptors and derivatives show promising possibilities for labeling in positron emission tomography studies. In an attempt to reduce epibatidine toxicity, new methyl derivatives were synthesized, tested in positron emission tomography imaging and in electrophysiology. labeling as well as physiological experiments highlighted the differences in sensitivity of the neuronal nicotinic acetylcholine receptors between two methyl enantiomers and the reduction in sensitivity caused by introducing the methyl group. At present, epibatidine derivatives seem the most promising compounds for in vivo labeling of neuronal nicotinic acetylcholine receptors.
A cognate dopamine transporter‐like activity endogenously expressed in a COS‐7 kidney‐derived cell line1
Authors: Sugamori Kim S. , Lee Frank J.S. , Pristupa Zdenek B. , Niznik Hyman B.
Published: July 2002

The activity of the dopamine transporter is an important mechanism for the maintenance of normal dopaminergic homeostasis by rapidly removing dopamine from the synaptic cleft. In kidney‐derived COS‐7, COS‐1 and HEK‐293 but not in other mammalian cell lines (CHO, Y1, Ltk−), we have characterized a putative functional dopamine transporter displaying a high affinity (K
m∼250 nM) and a low capacity (∼0.1 pmol/105 cells/min) for [3H]dopamine uptake. Uptake displayed a pharmacological profile clearly indicative of the neuronal dopamine transporter. Estimated K
i values of numerous substrates and inhibitors for the COS‐dopamine transporter and the cloned human neuronal transporter (human dopamine transporter) correlate well with the exception of a few notable compounds, including the endogenous neurotransmitter dopamine, the dopamine transporter inhibitor GBR 12,909 and the dopaminergic agonist apomorphine. As with native neuronal and cloned dopamine transporters, the uptake velocity was sodium‐sensitive and reduced by phorbol ester pre‐treatment. Two mRNA species of 3.8 and 4.0 kb in COS‐7 cells were revealed by Northern blot analysis similar in size to that seen in native neuronal tissue. A reverse‐transcribed PCR analysis confirmed the existence of a processed dopamine transporter. However, no immunoreactive proteins of expected dopamine transporter molecular size or [3H]WIN 35,428 binding activity were detected. A partial cDNA of ∼1.3 kb, isolated from a COS‐1 cDNA library and encoding transmembrane domains 1–6, displayed a deduced amino acid sequence homology of ∼96% to the human dopamine transporter. Taken together, the data suggest the existence of a non‐neuronal endogenous high affinity dopamine uptake system sharing strong functional and molecular homology to that of the cloned neuronal dopamine transporter.
The novel transmembrane Escherichia coli proteins involved in the amino acid efflux
Authors: Zakataeva Natalia P. , Aleshin Vladimir V. , Tokmakova Irina L. , Troshin Petr V. , Livshits Vitaliy A.
Published: July 2002

A novel gene of Escherichia coli, rhtB, has been characterized. Amplification of this gene provides resistance to homoserine and homoserine lactone. Another E. coli gene, rhtC, provides resistance to threonine. The homologues of RhtB are widely distributed among various eubacteria and archaea, from one to 12 copies of family members that differ in their primary structure were found in the genomes. Most of them are genes that encode hypothetical transmembrane proteins. Experimental data that indicate participation of the rhtB product in the excretion of homoserine have been obtained.
Aluminum fluoride inhibits phospholipase D activation by a GTP‐binding protein‐independent mechanism
Authors: Li Liang , Fleming Norman
Published: July 2002

Aluminum fluoride (AlF4
−) inhibited guanine nucleotide‐activated phospholipase D (PLD) in rat submandibular gland cell‐free lysates in a concentration‐dependent response. This effect was consistent in permeabilized cells with endogenous phospholipid PLD substrates. Inhibition was not caused by either fluoride or aluminum alone and was reversed by aluminum chelation. Inhibition of PLD by aluminum fluoride was not mediated by cAMP, phosphatases 1, 2A or 2B, or phosphatidate phosphohydrolase. AlF4
− had a similar inhibitory effect on rArf‐stimulated PLD, but did not block the translocation of Arf from cytosol to membranes, indicating a post‐GTP‐binding‐protein site of action. Oleate‐sensitive PLD, which is not guanine nucleotide‐dependent, was also inhibited by AlF4
−, supporting a G protein‐independent mechanism of action. A submandibular Golgi‐enriched membrane preparation had high PLD activity which was also potently inhibited by AlF4
−, leading to speculation that the known fluoride inhibition of Golgi vesicle transport may be PLD‐mediated. It is proposed that aluminum fluoride inhibits different forms of PLD by a mechanism that is independent of GTP‐binding proteins and that acts via a membrane‐associated target which may be the enzyme itself.
Differentiation of mucous neck cells into parietal cells: a new concept of mitochondrial biogenesis
Authors: Pradal Gilbert , Berreur Martine , Pouyet Brigitte , Riet Anne
Published: July 2002

Parietal cells of the gastric fundic mucosa are small and contain only a few tiny mitochondria when they begin to differentiate from mucous neck cells. The canalicular ATPase activity characteristic of mature parietal cells is discrete in these young cells, whereas areas of very high activity are apparent in the Golgi complex, reticulum, nuclear envelope, mitochondrial wall, and plasma membrane. Close relations and contacts occur between mitochondria and these organelles, and the size and number of mitochondria increase progressively. These relations, as well as mitochondrial ATPase activity (a true differentiation marker), cease once the mitochondria become as numerous and large as those of a mature parietal cell. Our observations suggest that a secondary form of mitochondrial biogenesis, involving the massive participation of other organelles and independent of the classical mechanisms inherent in mitosis, occurs in parietal cells at the beginning of G1 phase during the 6 days of their maturation.
Morphological brain asymmetries in male mice withcallosal defects due to prenatal gamma irradiation
Authors: Caparelli‐Dáquer Egas M. , Schmidt Sergio L.
Published: July 2002

We have previously suggested a relationship between the development of the corpuscallosum and the direction of cerebral asymmetries (Schmidt and Caparelli‐Dáquer, 1989 ; Schmidt et al., 1991). Here we report a study on morphological brainasymmetries using a distinct experimental approach. At embryonic day 16, mice were exposed toa gamma source, receiving a total dose of 2 Gy. At adulthood 32 irradiated and 28 normal maleSwiss mice were evaluated for individual and populational interhemispheric asymmetries ofweight, dorsal area, and neocortical volume. All irradiated mice showed callosal defects rangingfrom total absence to a partial reduction of the midsagittal callosal area. The normal and irradiatedgroups displayed a pronounced individual asymmetry in all measurements. In contrast,populational asymmetry could be identified only in the normal group. These results are consistentwith our previous data in acallosal mutant mice and support the hypothesis that the corpuscallosum may play a role in directing interhemispheric morphological brain asymmetries.
Need for support and participation in treatment differences among subgroups of relatives to compulsorily and voluntarily admitted mentally ill individuals
Authors: Östman Margareta , Hansson Lars
Published: July 2002

The need for support and participation in treatment of relatives to voluntarily and compulsorily admitted patients was addressed in a study of the quality of mental health services in two Swedish county councils. The aims of the study were to investigate differences in the above aspects between subgroups of relatives, the differences between two years of investigation, 1986 and 1991, and the differences between relatives of voluntarily and compulsorily admitted patients. The relatives investigated consisted of 79 spouses, 118 parents and 31 grown-up children. The results showed that there were only minor differences between the subgroups concerning their participation in care. Grown-up children experienced significantly less need of support and received less help for this need. In 1991, relatives participated more in the care situation, were more interested in support with regard to their own life situation, and also showed more positive attitudes towards the psychiatric services than in 1986. The relatives of the voluntarily admitted patients felt more involved in the patient's treatment, whereas the relatives of those compulsorily admitted felt less involved and perceived obstacles to admission.
Feasibility and accuracy of transthoracic Doppler echocardiographic estimation of pulmonary capillary wedge pressure applying different methods
Authors: Poelzl Gerhard , Gattermeier Martin , Kratzer Horst , Zeindlhofer Eduard , Kuehn Peter
Published: July 2002

Background:

Pulmonary wedge pressure (PWP) is an established index of cardiac function and an essential component in the management of patients with congestive heart failure and in critically ill patients.

Aim:

To evaluate feasibility and accuracy of non‐invasive prediction of PWP by Doppler echocardiography in daily clinical practice.

Methods:

Agreement was assessed between values predicted by Doppler vs. invasively measured PWP. Forty‐five consecutive patients [mean (S.D.) age 62 (10) years] with CAD (44%), DCMP (40%) and without structural heart disease (16%) were studied (EF ≤ 40% in 58% of the patients). Doppler transmitral and pulmonary venous flow velocity profiles were recorded. For binary and quantitative prediction of PWP, four different methods and five different linear equations, suggested previously in the literature, were evaluated.

Results:

Predictive values to identify elevated PWP were highest for pulmonary venous flow reversal exceeding the duration of forward mitral flow during atrial systole (PPV 1 and NPV 0.96). Likewise, agreement with measured PWP was highest for equations comprising both transmitral and pulmonary venous flow variables (relative mean difference 0.11, S.D. ± 4.01 mmHg for the most accurate equation). Feasibility was slightly, but not statistically, lower when pulmonary venous flow was considered vs. transmitral flow parameters alone for binary prediction (87 vs. 93%) as well as for quantitative assessment (82 vs. 93%).

Conclusion:

Semiquantitative prediction of elevated PWP by Doppler echocardiography is feasible as well as accurate in daily clinical practice. However, accuracy of numeric estimates is limited. Hence, invasive measurement of PWP is still necessary in certain clinical settings.
THE IMPACT OF LANGUAGE BARRIERS ON THE HEALTH CARE OF LATINOS IN THE UNITED STATES: A REVIEW OF THE LITERATURE AND GUIDELINES FOR PRACTICE
Authors: Timmins Caraway L.
Published: July 2002

Although Spanish was the primary language of an estimated 11% of adults in the year 2000, the U.S. health care system is largely geared toward serving English speakers. A systematic review was conducted of studies published in biomedical journals from 1990 to 2000 examining language barriers in health care for Latino populations. Aspects of the problem examined were access to health care, quality of care, and health status/health outcomes. Five (55%) of the nine studies examining access to care found a significant adverse effect of language; three (33%) found mixed or weak evidence that language affected access. Six (86%) of the seven studies evaluating quality of care found a significant detrimental effect of language barriers. Two of the three studies examining health status or outcomes found language to be a risk factor for adverse outcomes. Evidence was mixed as to the level of importance attributable to language when access to care is considered; however, non‐English‐speaking status was a marker of a population at risk for decreased access. Solid evidence showed that language barriers can adversely affect quality of care. Health care practitioners are recommended to devise an effective strategy to bridge language barriers in their setting. National laws and policies are discussed; practical guidelines and resources for providing language access in health care are provided. This article is intended to supply basic knowledge for providers and institutions in devising effective strategies for bridging the language barrier.
CERTIFIED NURSE‐MIDWIVES' KNOWLEDGE OF REIMBURSEMENT ISSUES
Authors: Ament Lynette
Published: July 2002

A recent survey examined the impact of reimbursement policies and admitting privileges on the ability of certified nurse‐midwives (CNMs) to practice and maintain an income. Many CNMs reported deficits in their knowledge about reimbursement issues. The responses to the survey indicated that service directors either do not view this information as important to their leadership role, have delegated their authority to someone else, or do not have authority over this information. Education program directors may need to examine their curricular content to determine if students are being provided adequate information to better value and understand this data.
Crystal structure of l‐rhamnose 1‐dehydrogenase involved in the nonphosphorylative pathway of l‐rhamnose metabolism in bacteria
Authors: Yoshiwara Kentaroh , Watanabe Seiya , Watanabe Yasunori
Published: January 2021

Several microorganisms can utilize l‐rhamnose as a carbon and energy source through the nonphosphorylative metabolic pathway, in which l‐rhamnose 1‐dehydrogenase (RhaDH) catalyzes the NAD(P)+‐dependent oxidization of l‐rhamnose to l‐rhamnono‐1,4‐lactone. We herein investigated the crystal structures of RhaDH from Azotobacter vinelandii in ligand‐free, NAD+‐bound, NADP+‐bound, and l‐rhamnose‐ and NAD+‐bound forms at 1.9, 2.1, 2.4, and 1.6 Å resolution, respectively. The significant interactions with the 2′‐phosphate group of NADP+, but not the 2′‐hydroxyl group of NAD+, were consistent with a preference for NADP+ over NAD+. The C5‐OH and C6‐methyl groups of l‐rhamnose were recognized by specific residues of RhaDH through hydrogen bonds and hydrophobic contact, respectively, which contribute to the different substrate specificities from other aldose 1‐dehydrogenases in the short‐chain dehydrogenase/reductase superfamily.
Adoptive transfer of immature myeloid cells lacking NF‐κB p50 (p50‐IMC) impedes the growth of MHC‐matched high‐risk neuroblastoma
Authors: Cui Cheng , Barberi Theresa , Suresh Rahul , Friedman Alan D.
Published: January 2021

High‐risk neuroblastomas harbor abundant myeloid cells that suppress antitumor immunity and support tumor growth. Macrophages lacking the inhibitory NF‐κB p50 subunit adopt a pro‐inflammatory phenotype. We now report that murine 9464D neuroblastoma cells, which express high levels of exogenous MYCN, grow slower in syngeneic p50(f/f);Lys‐Cre mice that lack p50 in macrophages and neutrophils, compared with p50(f/f) littermates. Tumors in p50(f/f);Lys‐Cre mice possess increased numbers of total and activated CD4+ and CD8+ T cells, and depletion of both of these T‐cell populations accelerates tumor growth. Anti‐PD‐1 T‐cell checkpoint blockade, or DNA methyltransferase and histone deacetylase inhibition, further slows tumor growth. In addition, adoptive transfer of immature myeloid cells lacking NF‐κB p50 (p50‐IMC), generated either from the bone marrow of p50−/− mice or via nucleofection of a p50 sgRNA:Cas9 complex into wild‐type hematopoietic progenitors, also slowed growth of MHC‐matched 9464D tumors but not of MHC‐mismatched Neuro2A tumors. These findings further validate the utility of targeting myeloid NF‐κB p50 as a strategy for cancer therapy and demonstrate activity of p50‐IMC generated by gene editing of syngeneic marrow cells, a cell product relevant to clinical translation.
Another trial falls short in search for stimulant use disorder treatment
Authors: Enos Gary
Published: January 2021

Numerous challenges in identifying treatment targets have left the addiction treatment field without an approved medication option for treating stimulant use disorder, making the response to the growing stimulant crisis look markedly different from opioid crisis response. A great deal of hope has been directed toward identifying existing medications or medication combinations that can combat stimulant addiction, but a much‐anticipated study released this month likely will dampen the enthusiasm.
New acting leadership at SAMHSA and ONDCP under Biden
Authors: Knopf Alison
Published: January 2021

Last week, the Substance Abuse and Mental Health Services Administration (SAMHSA) got a new acting head — Tom Coderre, who was chief of staff at the agency during the Obama administration. He was then senior advisor to Rhode Island Governor Gina Raimondo, a strong proponent of substance use disorder treatment who President Biden nominated to be commerce secretary. Coderre is beloved in the recovery community, and SAMHSA staff will be relieved to have him in charge while waiting for an official nomination for permanent assistant secretary for mental health and substance use with the Department of Health and Human Services.
Stretch, fold, and break: Intensification of emulsification of high viscosity ratio systems by fractal mixers
Authors: Hofmann Martin , Bayles Alexandra V. , Vermant Jan
Published: January 2021

During emulsification process design, the bulk and interfacial rheology of the target formulation must be carefully considered. Formulations with high viscosity ratios and/or finite interfacial elasticity are particularly challenging to emulsify, as conventional drop‐breakup methods consume significant energy and provide limited control over polydispersity. Here, we develop a two‐stage process that produces monodisperse emulsions from high viscosity ratio constituents. In the first stage, a custom static mixer generates co‐flowing layers of alternate phases, and progressively thins layers until they rupture, thus forming a high‐internal phase emulsion. The interfacial properties and flow conditions that promote stable fractal multiplication are discussed. In the second stage, extensional flow elements refine the polydispersity. We demonstrate the utility of this novel process by producing remarkably monodisperse polyisobutylene‐in‐water emulsions with an energy efficiency that is orders of magnitude higher than classical emulsification methods. The moderate throughputs achieved show promise for upscaling and intensification in industrial applications.
Philipp Jolly and his 1913 “the heredity of psychosis”: Homogeneity versus heterogeneity of familial transmission and an early look at Mendelian models for manic‐depressive illness and dementia praecox
Authors: Kendler Kenneth S. , Klee Astrid
Published: January 2021

Philipp Jolly's 1913 extensive monograph “The Heredity of Psychoses” provides, in both his detailed literature review and new pedigree study, an extensive assessment of a key issue in the psychiatric genetics of his day: the degree to which the familial transmission of psychiatric disorders was specific (or homogeneous) versus nonspecific (or heterogeneous). Contrary to a number of earlier observations, Jolly concludes that heterogeneous transmission is rare. Multiple psychiatric disorders can occur in one family because members of a family may have elevated predispositions to more than one disorder rather than one predisposition which increases risk for a wide range of conditions. A notable exception to this, he notes, is within manic‐depressive illness (MDI), where different forms of illness run together within families. Jolly is also among the earliest investigators to evaluate simple Mendelian models for MDI and dementia praecox (DP). While his methods were primitive and contained only “eye‐ball” examinations rather than statistical modeling, Jolly demonstrated considerable conceptual sophistication in the early application of such models. He concludes that DP is likely a recessive disorder. MDI, he states, is either a recessive condition or demonstrates “gender‐based dominant heredity” where heterozygote females are affected, and heterozygote males are not.
Computational Modeling of a Low‐Cost Fluidic Oscillator for Use in an Educational Respiratory Simulator
Authors: Dillon Tom , Ozturk Caglar , Mendez Keegan , Rosalia Luca , Gollob Samuel Dutra , Kempf Katharina , Roche Ellen Tunney
Published: January 2021

Herein, the computational modeling of a fluidic oscillator for use in an educational respiratory simulator apparatus is presented. The design provides realistic visualization and tuning of respiratory biomechanics using a part that is (i) inexpensive, (ii) easily manufactured without the need for specialized equipment, (iii) simple to assemble and maintain, (iv) does not require any electronics, and (v) has no moving components that could be prone to failure. A computational fluid dynamics (CFD) model is used to assess flow characteristics of the system, and a prototype is developed and tested with a commercial benchtop respiratory simulator. The simulations show clinically relevant periodic oscillation with outlet pressures in the range of 8–20 cmH2O and end‐user‐tunable frequencies in the range of 3–6 s (respiratory rate [RR] of 10–20 breaths per minute). The fluidic oscillator presented here functions at physiologically relevant pressures and frequencies, demonstrating potential as a low cost, hands‐on, and pedagogical tool. The model will serve as a realistic model for educating Science, Technology, Engineering, and Mathematics (STEM) students on the relationship between flow, pressure, compliance, and volume in respiratory biomechanics while simultaneously exposing them to basic manufacturing techniques.