bluuebunny/crossref_metadata_2025 huggingface dataset:
Adenosine evokes potassium currents by protein kinase C activated via a novel signaling pathway in superior colliculus neurons
Authors: Nishizaki Tomoyuki , Ikeuchi Youji
Published: July 2002

Adenosine evoked whole‐cell potassium currents and enhanced intracellular free Ca2+ concentration ([Ca2+]i) in superior colliculus neurons through a P2Y purinoceptor linked to a pertussis toxin‐insensitive G‐protein, possibly Gq‐protein, which is involved in a protein kinase C(PKC) activation pathway. The [Ca2+]i increase was inhibited by a phospholipase C(PLC) inhibitor, whereas the evoked currents were not affected by a PLC inhibitor or a phospholipase A2(PLA2) inhibitor. Adenosine elicited single channel currents via PKC activation in cell‐attached patches and furthermore, those currents with conductances of the same slope were induced even in excised patches, suggesting that PKC can be activated only by cell membrane factors without intracellular components. These results thus indicate that the P2Y purinoceptor‐coupled potassium channel is regulated via a novel PKC activation pathway independent of PLC or PLA2.
Flow cytometric investigation of neutrophil activation pathways by n‐formyl‐Met‐Leu‐Phe and phorbol myristate acetate
Authors: Zhang Jing , Kaupke Charles J , Yousefi Shookooh , Cesario Thomas C , Vaziri Nick D
Published: July 2002

Summary— Recent evidence suggests that multiple pathways exist in PMN activation and that specific leukocyte response may be due to the activation of a particular signaling pathway. Using flow cytometry, PMN activation pathways were studied through the parallel comparison of n‐formyl‐Met‐Leu‐Phe (fMLP)‐ and phorbol‐12‐myristate‐13‐acetate (PMA)‐induced stimulation and by simultaneous assays for CD11b expression and morphology. The maximal CD11b expression was higher with PMA than with fMLP, suggesting different activation pathways. Under these experimental conditions, a morphological response to fMLP was not observed. However, significant shape change was detected in PMA treated samples and was suppressed by either the removal of extracellular calcium or staurosporine at the concentrations above 14.5 μM. Calcium ionophore induced a similar light scattering pattern to that by PMA and enhanced CD11b expression, both of which were not inhibitable by staurosporine. These observations, for the first time, indicated that Ca2+ was a mediator in activation processes and that the treatment of PMN with PMA resulted in Ca2+ influx.
Cell‐cycle dependent biosynthesis and localization of p53 protein in untransformed human cells
Authors: Katsumoto Tetsuo , Higaki Katsumi , Ohno Kousaku , Onodera Kazukiyo
Published: July 2002

Summary— Localization of p53 in human cultured lymphocytes and in cultured skin fibroblasts was studied by immuno‐fluorescent microscopy and post‐embedded immunoelectron microscopy using Lowicryl K4M. In quiescent lymphocytes, p53 was found in small amounts in both the cytoplasm and the nucleus. p53 in the nucleus was found associated with the non‐chromatin structure. At 24 h or 72 h of PHA stimulation, p53 increased markedly just beneath the plasma membrane and in the nucleus, which stained diffusely with anti‐p53. In resting fibroblasts, small amounts of p53 were present in both the cytoplasm and the nucleus. After 16 h of stimulation of confluent‐resting fibroblasts by trypsinization and replating, a phase just prior to the initiation of DNA synthesis, p53 slightly increased in both the cytoplasm and the nucleus. Afterwards, p53 was present predominantly in the cytoplasm, closely associated with the cytoskeletal actin filaments. In mitotic cells, p53 was distributed throughout the cytoplasm. When fibroblasts were extracted with saponin, p53 was still associated with the actin filaments, as well as mitochondrial membranes and granular structures of the nuclear matrix. Our data suggest that the initial increase of p53 in cells that enter the cell cycle through G1 first bind to the actin cytoskeleton, and that some of the p53 then move into the nucleus to initiate gene activation and DNA synthesis for cell proliferation. This implies that there is some functionally significant interaction between p53 and actin in the cells.
Visible light‐induced destabilization of endocytosed liposomes
Authors: Miller Christina R , Clapp Paula J , O'Brien David F
Published: July 2002

The potential biomedical utility of the photoinduced destabilization of liposomes depends in part on the use of green to near infrared light with its inherent therapeutic advantages. The polymerization of bilayers can be sensitized to green light by associating selected amphiphilic cyanine dyes, i.e. the cationic 1,1′‐dioctadecyl‐3,3,3′,3′‐tetramethylindocarbocyanine (DiI), or the corresponding anionic disulfonated DiI (DiI‐DS), with the lipid bilayer. The DiI sensitization of the polymerization of 1,2‐dioleoyl‐sn‐glycero‐3‐phosphoethanolamine/1,2‐bis[10‐(2′,4′‐hexadienoyloxy)‐decanoyl]‐sn‐glycero‐3‐phosphocholine liposomes caused liposome destabilization with release of encapsulated aqueous markers. In separate experiments, similar photosensitive liposomes were endocytosed by cultured HeLa cells. Exposure of the cells and liposomes to 550 nm light caused a net movement of the liposome‐encapsulated 8‐hydroxypyrene‐1,3,6‐trisulfonic acid (HPTS) from low pH compartment(s) to higher pH compartment(s). This suggests that photolysis of DiI‐labelled liposomes results in delivery of the contents of the endocytosed liposomes to the cytoplasm. The release of HPTS into the cytoplasm appears to require the photoactivated fusion of the labelled liposomes with the endosomal membrane. These studies aid in the design of visible light sensitive liposomes for the delivery of liposome‐encapsulated reagents to the cytoplasm.
Efficient phage display of polypeptides fused to the carboxy‐terminus of the M13 gene‐3 minor coat protein
Authors: Fuh Germaine , Sidhu Sachdev S.
Published: July 2002

We report that, contrary to common belief, polypeptides fused to the carboxy‐terminus of the M13 gene‐3 minor coat protein are functionally displayed on the phage surface. In a phagemid display system, carboxy‐terminal fusion through optimized linker sequences resulted in display levels comparable to those achieved with conventional amino‐terminal fusions. These findings are of considerable importance to phage display technology because they enable investigations not suited to amino‐terminal display, including the study of protein–protein interactions requiring free carboxy‐termini, functional cDNA cloning efforts, and the display of intracellular proteins.
Genomic Exploration of the Hemiascomycetous Yeasts: 11. Kluyveromyces lactis
Authors: Bolotin-Fukuhara Monique , Toffano-Nioche Claire , Artiguenave François , Duchateau-Nguyen Guillemette , Lemaire Marc , Marmeisse Roland , Montrocher Robert , Robert Catherine , Termier Michel , Wincker Patrick , Wésolowski-Louvel Micheline
Published: July 2002

Random sequencing of the Kluyveromyces lactis genome allowed the identification of 2235–2601 open reading frames (ORFs) homologous to S. cerevisiae ORFs, 51 ORFs which were homologous to genes from other species, 64 tRNAs, the complete rDNA repeat, and a few Ty1‐ and Ty2‐like sequences. In addition, the complete sequence of plasmid pKD1 and a large coverage of the mitochondrial genome were obtained. The global distribution into general functional categories found in Saccharomyces cerevisiae and as defined by MIPS is well conserved in K. lactis. However, detailed examination of certain subcategories revealed a small excess of genes involved in amino acid metabolism in K. lactis. The sequences are deposited at EMBL under the accession numbers AL424881–AL430960.
Corticosterone‐induced rapid phosphorylation of p38 and JNK mitogen‐activated protein kinases in PC12 cells
Authors: Li Xiaoyu , Qiu Jian , Wang Jianwen , Zhong Yongping , Zhu Jianqin , Chen Yizhang
Published: July 2002

The present study showed that corticosterone (B) could induce a rapid activation of p38 and c‐Jun NH2‐terminal protein kinase (JNK) in PC12 cells. The dose–response and time–response curves were bell‐shaped with maximal activation at 10−9 M and at 15 min. RU38486 had no effect, and bovine serum albumin‐coupled B could induce the activation. Genistein failed to block the phosphorylation, suggesting the pathway was not involved in tyrosine kinase activity. Phorbol 12‐myristate 13‐acetate could mimic, while Gö6976 could abolish the actions. These results demonstrated that B might act via a putative membrane receptor to activate p38 and JNK rapidly through a protein kinase C‐dependent pathway.
Aluminum interaction with plasma membrane lipids and enzyme metal binding sites and its potential role in Al cytotoxicity
Authors: Jones David L , Kochian Leon V
Published: July 2002

The trivalent cation aluminum can cause chronic cytotoxicity in plants, animals and microorganisms. It has been suggested that Al interaction with cell membranes and enzyme metal binding sites may be involved in Al cytotoxicity. In this study, the binding of Al to microsomes and liposomes was found to be lipid dependent with the signal transduction element phosphatidylinositol‐4,5‐bisphosphate having the highest affinity for Al with an Al:lipid stoichiometry of 1:1. Al binding was only reduced in the presence of high concentrations of Ca2+ (>1 mM). Both citrate and, to a lesser extent, malate were capable of preventing Al lipid binding, which is consistent with the involvement of these organic acids in a recently described Al detoxification mechanism in plants. The effects of AlCl3, Al‐citrate and ZnSO4 on metal‐dependent enzyme activities (enolase, pyruvate kinase, H+‐ATPase, myosin, Calpain, proteinase K, phospholipase A2 and arginase) was assayed in vitro. While Zn2+ was capable of inhibiting all the enzymes except the H+‐ATPase, AlCl3 and Al‐citrate had minimal effects except for with phospholipase A2 where an interaction with AlCl3 occurred. However, this could be negated by the addition of citrate. The results indicate that, contrary to current hypotheses, the toxic mode of Al is not through an interaction with enzymatic catalytic metal binding sites but may be through the interaction with specific membrane lipids.
Hepatocyte growth factor/scatter factor (HGF/SF) signals via the STAT3/APRF transcription factor in human hepatoma cells and hepatocytes
Authors: Schaper Fred , Siewert Elmar , Gómez-Lechón Maria José , Gatsios Petros , Sachs Martin , Birchmeier Walter , Heinrich Peter C , Castell José
Published: July 2002

© 1997 Federation of European Biochemical Societies.
Human osteosarcoma cell susceptibility to natural killer cell lysis depends on CD54 and increases after TNFα incubation
Authors: Mariani Erminia , Tarozzi Andrea , Meneghetti Alessandra , Cattini Luca , Facchini Andrea
Published: July 2002

Osteosarcoma cell lines vary widely in their susceptibility to natural killer (NK) cell lysis in vitro although it is still unclear why this occurs. In this study we investigated the expression of some cell adhesion molecules on osteosarcomas to determine which of these can modify the susceptibility to NK lysis and we also attempted to modulate the cytolytic susceptibility of these targets with TNFα. We found that osteosarcoma lysis induced by NK cells correlates with different expression of the CD54 adhesion molecule on osteosarcomas and the increased susceptibility after TNFα treatment mostly depends on the expression of CD54 molecules on target cells.
Mg2+ modulates membrane lipids in vascular smooth muscle: a link to atherogenesis
Authors: Morrill Gene A , Gupta Raj K , Kostellow Adele B , Ma Gui-Ying , Zhang Amin , Altura Bella T , Altura Burton M
Published: July 2002

Epidemiological studies associate low dietary magnesium intake with an increased incidence of ischemic heart disease and sudden cardiac death. We have used proton‐magnetic resonance (1H‐NMR) techniques and Mg2+‐selective electrodes to monitor changes in lipid extracts of aortic and cerebrovascular smooth muscle as extracellular ionized magnesium ion concentration ([Mg2+]o) is lowered. We have found that, within the pathophysiological range of Mg2+ concentrations, fatty acid chain length and double bond content are progressively reduced as [Mg2+]o is lowered. In contrast, the plasmalogen content is progressively increased. A concomitant decrease in fatty acid chain length and double bonds indicates oxidation of double bonds resulting in truncation of the fatty acids. A decrease in lipid oxidation in the presence of elevated Mg2+ could contribute to the apparent protective role of increased Mg2+ intake on vascular function in humans.
Addition of G418 and other aminoglycoside antibiotics to mammalian cells results in the release of GPI‐anchored proteins
Authors: Küng Matthias , Stadelmann Barbara , Brodbeck Urs , Bütikofer Peter
Published: July 2002

Resistance to the neomycin analogue G418 forms the basis of a dominant marker selection system for mammalian cells transfected with the bacterial neomycin gene. We found that COS‐1 cells stably transfected with the neomycin resistance gene had a greater than 50% reduction in cell‐associated glycosylphosphatidylinositol (GPI)‐anchored alkaline phosphatase (AP). A similarly reduced amount of AP was also observed in wild‐type COS‐1 cells incubated in the presence of G418 or other aminoglycoside antibiotics. The AP was released from cells into the culture supernatant in its GPI‐anchored form. Our data suggest that the G418‐induced reduction of AP involves a vesiculation process of COS‐1 cells.
Large‐scale preparation of biologically active recombinant ovine obese protein (leptin)
Authors: Gertler Arieh , Simmons Julie , Keisler Duane H
Published: July 2002

Prokaryotic expression vector pMON3401 encoding full size A(‐1) ovine leptin was prepared by polymerase chain reaction (PCR) of previously described cDNA. E. coli cells transformed with this vector overexpressed large amounts of ovine leptin upon induction with nalidixic acid. The expressed protein found in the inclusion bodies was refolded and purified to homogeneity on Q‐Sepharose and SP‐Sepharose columns, yielding two electrophoretically pure fractions (leptin‐Q and leptin‐SP), composed respectively of 90 and 95% of monomeric protein of the expected molecular mass of 16 kDa. The purified protein was capable of interacting with antibodies raised against GST‐ovine leptin and to bind specifically to ventromedial hypothalamus of ewes. The biological activity of both fractions resulting from proper renaturation was further evidenced by their ability to stimulate DNA synthesis in leptin‐sensitive BAF/3 cells transfected with a long form of human leptin receptor construct.
Amyloidogenic determinant as a substrate recognition motif of insulin‐degrading enzyme
Authors: Kurochkin Igor V
Published: July 2002

Insulin‐degrading enzyme (IDE) is an evolutionarily conserved neutral thiol metalloprotease expressed in all mammalian tissues whose biological role is not well established. IDE has highly selective substrate specificity. It degrades insulin, glucagon, atrial natriuretic peptide, transforming growth factor α but does not act on related hormones and growth factors. The structural properties determining whether a peptide is an IDE substrate are essentially unknown. The reported cleavage sites are not consistent with simple peptide‐bond recognition and it was proposed that IDE recognizes in its substrates some elements of tertiary structure. We noticed that although IDE substrates are functionally unrelated, the majority of them share a specific property, an ability to form under certain conditions amyloid fibrils. Utilizing the residue pattern recognition procedure, this study reveals a common motif in the sequences of IDE substrates, HNHHHPSH, where H is wholly or partly hydrophobic character, N is small and neutral, P is polar, and S is polar and/or small amino acid residue. It is proposed that this sequence motif predetermines a structure recognized by IDE. The identified motif appears to be essentially the same as the proposed earlier consensus sequence for amyloid‐forming peptides [Turnell and Finch, J. Mol. Biol. 227 (1992) 1205–1223]. The study suggests that IDE may play a role in elimination of potentially toxic amyloidogenic peptides.
The proregion of papaya proteinase IV inhibits Colorado potato beetle digestive cysteine proteinases
Authors: Visal Savita , Taylor Mark A.J. , Michaud Dominique
Published: July 2002

Three distinct digestive protease systems were induced in larvae of the herbivorous pest, Colorado potato beetle (CPB; Leptinotarsa decemlineata Say), and used as a model to assess the ability of the proregion of papaya proteinase IV (PPIV; glycyl endopeptidase, EC 3.4.22.25) to act as an inhibitor of insect digestive cysteine proteinases. As shown by gelatin/PAGE and complementary inhibition assays, a recombinant form of the proregion produced in Escherichia coli inhibited a fraction of the insect proteases also inhibited by the well‐characterized inhibitor of cysteine proteinases, oryzacystatin I (OCI). In contrast with OCI, the inhibitory potency of the proregion was affected by an increase of the temperature, suggesting a certain alteration of its structural integrity by the insect non‐target proteases. This apparent susceptibility to proteolysis was confirmed by SDS‐PAGE, after challenging the proregion with the different insect extracts. As seen on gel, selective inhibition of the insect aspartate proteinase, cathepsin D, with the inhibitor pepstatin A preserved the activity of the proregion against cysteine proteinases by preventing its hydrolysis. Taken together, these observations suggest the potential of plant protease proregions as regulators of cysteine proteinases in biotechnological systems, and show the ability of protease inhibitors to preserve the integrity of ‘companion’ defense‐related proteins from the action of insensitive proteases in target pests.
Interleukin‐1β activates a short STAT‐3 isoform in clonal insulin‐secreting cells
Authors: Morton Nicholas M. , de Groot Rolf P. , Cawthorne Michael A. , Emilsson Valur
Published: July 2002

Interleukin‐1β (IL‐1β) is a potent inflammatory cytokine involved in type 1 diabetes and acts through defined IL‐1β signaling pathways. In the present work we describe induction of DNA binding activity to signal transducer and activator of transcription (STAT) in response to IL‐1β in clonal insulin‐secreting cells. Moreover, IL‐1β activates a short isoform of STAT‐3 that potently stimulates transcription. Immunoprecipitation studies reveal an interaction between the activated STAT‐3 and the IL‐1 receptor accessory protein indicating an association between the two signaling pathways. This may be a novel point of transduction cross talk and an additional mechanism utilised by IL‐1β in the pancreatic β‐cell during the process of type 1 diabetes.
High‐level expression of uniformly 15N‐labeled hen lysozyme in Pichia pastoris and identification of the site in hen lysozyme where phosphate ion binds using NMR measurements
Authors: Mine Shouhei , Ueda Tadashi , Hashimoto Yoshio , Tanaka Yoshitsugu , Imoto Taiji
Published: July 2002

The non‐enzymatic deamidation of Asn to Asp is known to occur in proteins and peptides and is accelerated by phosphate buffer [Tyler‐Cross, R. and Schirch, V. (1991) J. Biol. Chem. 25, 22549–22556]. We attempted to identify the site in lysozyme where a phosphate ion binds by means of 1H‐15N HSQC measurements of 15N‐labeled lysozyme, which was successfully obtained using Pichia pastoris. As a result, we found that the phosphate ion was preferentially bound to Asn‐103 in hen lysozyme. The method presented here may be useful for identifying the binding site of a protein with low molecular weight substances.
Mistletoe lectin dissociates into catalytic and binding subunits before translocation across the membrane to the cytoplasm
Authors: Agapov Igor I. , Tonevitsky Alexander G. , Moysenovich Mikhail M. , Maluchenko Natalya V. , Weyhenmeyer Roland , Kirpichnikov Mikhail P.
Published: July 2002

Hybridomas producing monoclonal antibodies (mAbs) against the mistletoe lectin A‐chain (MLA) were obtained to investigate the intracellular routing and translocation of ribosome‐inactivating proteins. Anti‐MLA mAb MNA5 did not bind the holotoxin but interacted with isolated MLA. This epitope was not recognized upon MLA denaturation or conjugation of MLA with the ricin binding subunit (RTB). Furthermore, the mAbs did not appreciably react with a panel of MLA synthetic octapeptides linked to the surface of polyethylene pins. A study of the cytotoxicity of mistletoe lectin, ricin, and chimeric toxin MLA/RTB for the hybridomas revealed that interchain disulfide bond reduction and subunit dissociation are required for cytotoxic activity of mistletoe lectin.
Examination of the signal transduction pathways leading to activation of extracellular signal‐regulated kinase by formyl‐methionyl‐leucyl‐phenylalanine in rat neutrophils
Authors: Chang Ling-Chu , Wang Jih-Pyang
Published: July 2002

The signaling pathways leading to extracellular signal‐regulated kinase (ERK) activation in formyl‐methionyl‐leucyl‐phenylalanine (fMLP)‐stimulated rat neutrophils were examined. fMLP‐stimulated ERK activation based on immunoblot analysis with antibodies against the phosphorylation form of ERK was attenuated by the pretreatment of cells with pertussis toxin but not with a dual cyclo‐oxygenase/lipoxygenase inhibitor BW755C. Exposure of cells to the tyrosine kinase inhibitor genistein, phosphatidylinositol 3‐kinase (PI3K) inhibitors wortmannin and LY294002, or protein kinase C (PKC) inhibitors Gö6976, Gö6983, and GF109203X inhibited fMLP‐stimulated ERK phosphorylation in a concentration‐dependent manner. In addition, both the phospholipase C (PLC) inhibitor U73122 and the Ca2+ chelator BAPTA attenuated ERK activation. These results indicate that Gi/o protein, tyrosine kinase, PI3K, PKC, and PLC/Ca2+, but not arachidonate metabolites, act upstream of fMLP‐stimulated ERK activation.
Short‐term block of Na+/K+‐ATPase in neuro‐glial cell cultures of cerebellum induces glutamate dependent damage of granule cells
Authors: Stelmashook E.V. , Weih M. , Zorov D. , Victorov I. , Dirnagl U. , Isaev N.
Published: July 2002

Granule cells in a dissociated neuro‐glial cell culture of cerebellum when exposed to ouabain (10−3 M) for 25 min apparently swell, increase their [Ca2+]i with obvious depolarization of the mitochondrial membrane. In 3 h after ouabain was omitted from the solution, 62±3% of granule cells had pycnotic nuclei. The supplement of a solution with competitive specific antagonist of NMDA receptors, L‐2‐amino‐7‐phosphonoheptanoate (10−4 M, APH) together with ouabain prevented cells from swelling, mitochondrial deenergization, neuronal death and increase of [Ca2+]i. These data suggest that cellular Na+/K+‐ATPase inactivation in neuro‐glial cell cultures of cerebellum leads to glutamate (Glu) accumulation, hyperstimulation of glutamate receptors, higher Ca2+ and Na+ influxes into the cells through the channels activated by Glu. This process leads to cell swelling, mitochondrial deenergization and death of granule cells. Possibly, the decrease of Na+/K+‐ATPase activity in brain cells can lead to the onset of at least some chronic neurological disorders.
Association of the rat heterogeneous nuclear RNA‐ribonucleoprotein F with TATA‐binding protein
Authors: Yoshida Tatsushi , Makino Yasutaka , Tamura Taka-aki
Published: July 2002

Heterogeneous nuclear ribonucleoprotein F (hnRNP‐F) has been shown to be a pre‐mRNA splicing factor. Recent studies have uncovered the coordination of synthesis of pre‐mRNA and its processing, including post‐transcriptional modification and splicing. Here, we present evidence for an association between a splicing factor, hnRNP‐F, and TATA‐binding protein (TBP), which is an essential factor needed for transcription initiation. An affinity detection experiment revealed hnRNP‐F in the preparation of TBP‐interacting proteins. HnRNP‐F was associated with TBP in nuclear extracts and was capable of direct binding to TBP in vitro. These results suggest that hnRNP‐F is associated with TBP in the cell. HnRNP‐F was observed in abundance in the thymus, spleen and testis, and its distribution pattern was similar to that of TBP, implying a functional coordination of transcription and splicing. We assume that the splicing machinery is associated with the transcription apparatus as a prerequisite prior to transcriptional elongation.
Binding properties of a highly potent and selective iodinated aminopeptidase N inhibitor appropriate for radioautography
Authors: Noble Florence , Luciani Nathalie , Da Nascimento Sophie , Laï-Kuen René , Bischoff Laurent , Chen Huixiong , Fournié-Zaluski Marie-Claude , Roques Bernard P.
Published: July 2002

Aminopeptidase N (APN) is a zinc metallopeptidase involved in the inactivation of biologically active peptides. The knowledge of its precise distribution is crucial to investigate its physiological role. This requires the use of appropriate probes such as the recently developed highly potent and selective radiolabeled APN inhibitor 2(S)‐benzyl‐3‐[hydroxy(1′(R)‐aminoethyl)phosphinyl]propanoyl‐L‐3‐[125I]iodotyrosine ([125I]RB 129). Its binding properties were investigated using rat brain homogenates (K
d=3.4 nM) or APN expressed in COS‐7 cells (K
d=0.9 nM). The specific binding was 95% at [K
d], and preliminary autoradiography in intestine is promising. The decreased affinity of [125I]RB 129 (=10−6 M) for the E350D APN mutant, supports the critical role of E350 in the amino‐exopeptidase action of APN.
Mistletoe lectin A‐chain unfolds during the intracellular transport
Authors: Agapov Igor I , Tonevitsky Alexander G , Maluchenko Natalya V , Moisenovich Mikhail M , Bulah Yuliya S , Kirpichnikov Mikhail P
Published: July 2002

Protein conformation during intracellular routing and translocation of the ribosome‐inactivating proteins was investigated on hybridomas producing monoclonal antibodies (monAbs) against mistletoe lectin (ML). Decrease in the toxin activity towards these hybridomas is accounted for by the intracellular interaction of monAbs and the toxin resulting in the interruption of enzymatic subunit translocation into the cytosol. Obtained monAbs interacted with denatured ML A‐chain (MLA) and a panel of MLA synthetic octapeptides linked to the surface of polyethylene pins. Enzyme‐linked immunosorbent assay (ELISA) shows that monAbs recognize five epitopes in denatured MLA. Treatment of MLA by 3 M of guanidine hydrochloride leads to appearance of the epitopes. Hybridoma TA7 has been shown to be insensitive to cytotoxic action of ML. TA7 monAb as we have shown recognizes epitope 101–105, FTGTT, and inhibits the liposome aggregation induced by MLA. A study of the cytotoxicity of ML and ricin for the hybridomas revealed that the unfolding of A‐chain is probably required for intracellular transport and cytotoxic activity of ML.
Conservative treatment of low grade squamous intraepithelial lesions (LSIL) of the cervix
Authors: Lee S.S.N. , Collins R.J. , Pun T.C. , Cheng D.K.L , Ngan H.Y.S.
Published: July 2002

Objective: To assess the accuracy of the cervical smear and colposcopy in diagnosing low grade squamous intraepithelial lesions (LSIL), and the natural history of LSIL. Method: The cytological, colposcopic and histological findings of 145 patients with a smear diagnosis of LSIL were compared, and the final outcome studied. Results: The diagnosis of LSIL either by cytology alone or in combination with colposcopy was associated with a rate of overdiagnosis of 11.7% and 6.9%, respectively, and a rate of underdiagnosis of 31.0% and 26.2%, respectively. Spontaneous regression of lesions occurred in 81.1% of patients with proven LSIL, with regression within 24 months in 4/5 of cases. Conclusions: The degree of dysplasia shown in the cervical smear correlated poorly with histology. All patients with cervical smear showing LSIL should have colposcopy and colposcopic‐directed biopsy to exclude the presence of more advanced lesions. In the absence of some serious lesions, it is then acceptable to observe the patient for 24 months before adopting definite treatment, as spontaneous regression is common.
Extending the treatment boundaries: Zoladex and add‐back
Authors: Schlaff W.D
Published: July 2002

Objective: To review the evidence that add‐back hormone replacement therapy (HRT) can ameliorate the metabolic consequences of gonadotropin‐releasing hormone (GnRH) agonist treatment in women with symptomatic endometriosis. Methods: A review of relevant literature. Results: Early studies suggested that add‐back HRT maintained bone mineral density (BMD) without reducing the symptomatic benefit of GnRH treatment. Both high‐dose progestogen and low dose progestogen plus cyclical etidronate are effective in maintaining BMD. Standard and low dose HRT add‐back may be more effective in relieving the hypo‐estrogenic side‐effects of GnRH agonist therapy. Randomized controlled studies have shown that both low‐dose and standard‐dose add‐back HRT reduce the side‐effects of GnRH agonist therapy, and that this benefit extends to 12 months of treatment. Conclusions: GnRH agonist treatment with add‐back HRT seems to offer the hope of improved treatment for women with endometriosis, but the optimum treatment duration and time to start HRT have yet to be defined.
GABAergic system in the developing mammalian retina: dual sources of GABA at early stages of postnatal development
Authors: Yamasaki Edna N. , Barbosa Valéria D. , de Mello Fernando G. , Hokoç Jan Nora
Published: July 2002

In the present work, we have characterized the maturation of the GABAergic system in mammalian retina. Immunoreactivity for GABA, GAD (glutamic acid decarboxylase, EC 4.1.1.15) ‐65 and ‐67 in the adult rat retina was localized in cells in the inner nuclear and ganglion cell layers. This pattern was established around postnatal day 8 and included transient GABA and GAD‐67 expression in horizontal cells. GAD activity was very low at P1 and P4, increasing after P8, reaching maximal activity by P21 and decreasing to attain adult values by P30. GABA content was approximately constant from P1 to P13, increasing thereafter to reach adult levels. GAD protein content increased progressively with postnatal development and the two isoforms could be distinguished at P8.

The disparity between retinal GABA content vs presence and activity of the synthesizing enzyme, led us to investigate the alternative pathway for GABA synthesis that utilizes putrescine as a substrate. Highest levels of ornithine decarboxylase activity (the limiting step for putrescine synthesis) were found between P1 and P4, decreasing to very low levels after P13. The same pattern was observed for putrescine content in the retina. Highest amounts were found at P1, that decreased and remained constant after P13. Additionally, approximately 40% of tritiated putrescine incorporated by P1, P4 and adult retinas was converted into GABA.

Our results suggest the existence of two different sources of GABA in mammalian retina, one that uses glutamate as a precursor and predominates in the mature nervous system and another that utilizes putrescine and is present transiently at early developmental stages.
Pharmacotherapeutic profile of venlafaxine
Authors: Preskorn S
Published: July 2002

When selecting an antidepressant, a number of factors must be considered. These considerations are summarized under the mnemonic STEPS: Safety, Tolerability, Efficacy, Payment (eg, cost-effectiveness), and Simplicity of use. Venlafaxine is the first of a new class of antidepressants that selectively blocks the serotonin and noradrenaline uptake pumps without blocking muscarinic, histaminergic and adrenergic receptors or inhibiting sodium fast channels. Because venlafaxine avoids these mechanisms of action, it has a wide therapeutic index, an improved tolerability profile and a reduced risk of causing pharmacodynamically mediated drug-drug interactions when compared to tricyclic antidepressants (TCAs). In contrast to some other new antidepressants, venlafaxine also avoids effects on cytochrome P450 which are likely to cause clinically meaningful, pharmacokinetically mediated drug-drug interactions. The effects on the uptake pumps of both serotonin and noradrenaline appear to be responsible for some of venlafaxine's unique features in terms of antidepressant efficacy, including its ascending antidepressant dose-response curve and its apparent rapid onset of antidepressant action at the upper end of its clinically relevant dosing range. Venlafaxine is effective in a broad spectrum of patients, including outpatients and inpatients, those with and without melancholia, patients with symptoms of anxiety or agitation or retardation and patients with first time or recurrent episodes of major depression. An important factor when selecting an antidepressant is the simplicity of the dosing regimen and the ability to rapidly and confidently achieve the optimal dose for the patient. In this regard, venlafaxine can be initiated at a clinically effective dose from the beginning. If the patient fails to respond to this dose, there is evidence that increased antidepressant efficacy can be achieved by increasing the dose rather than having to resort to an augmentation strategy or switch to another class of antidepressants. In the immediate release form, venlafaxine has proven antidepressant efficacy when using a twice-or three-times-a-day schedule. A sustained release formulation is expected to be marketed soon and will permit once-a-day-dosing.
Intravenous magnesium sulfate for bronchial hyperreactivity: A randomized, controlled, double‐blind study
Authors: Schenk Peter , Vonbank Karin , Schnack Beate , Haber Paul , Lehr Stephan , Smetana Ronald
Published: July 2002

Background

Magnesium has been shown to be helpful in the treatment of acute exacerbations of asthma. Conflicting data exist concerning the effect of magnesium on bronchial hyperreactivity.

Methods

We performed a randomized, double‐blind, placebo‐controlled study to investigate the effect of intravenous magnesium sulfate on bronchial reactivity to metacholine in 30 subjects with bronchial hyperreactivity. Two days after baseline metacholine provocation, 20 subjects received 0.3 mmol/kg/h of intravenous magnesium sulfate and 10 subjects received normal saline solution. Metacholine provocation was repeated 30 minutes after the initiation of the magnesium or placebo infusion.

Results

The difference of the postinterventional minus the baseline provocative dose of metacholine required to decrease the forced expiratory volume in 1 second by 20% (PC20) was significantly higher in the magnesium group compared with the placebo group (0.48 ± 0.46 mg/mL versus 0.05 ± 0.73 mg/mL, P = .028). In the magnesium group, the PC20 significantly increased (from 0.83 ± 0.54 mg/mL to 1.31 ± 0.66 mg/mL, P = .0001), whereas there was no change in the placebo group (0.86 ± 0.52 mg/mL to 0.91 ± 0.54 mg/mL, P = .83).

Conclusions

In the magnesium group, 30% of the subjects reached a normal PC20 compared with 10% in the placebo group. We conclude that intravenous magnesium sulfate significantly improved bronchial hyperreactivity and may serve as an adjunct to standard treatment.

Clinical Pharmacology & Therapeutics (2001) 69, 365–371; doi: 10.1067/mcp.2001.114926
A Guide for Businesses and Employers Responding to Novel Coronavirus disease (COVID-19): 4th edition
Authors:   Members of Occupational Health Committee on JSTH (Japanese Society of Travel and Health), Suzuki Hidetaka , Miyamoto Toshiaki , Hamada Atsuo , Nakano Akiyasu , Okoshi Hirofumi , Yamasawa Fumihiro
Published: April 2021

The Japanese Society of Travel and Health (JSTH) and the Japanese Society for Occupational Health (JSOH) have compiled “Novel Coronavirus Information” together as a joint document, which has been shared with the public on their respective websites since February 2020. In May 11, 2020, this document was to be published as “A Guide for Businesses and Employers Responding to Novel Coronavirus Disease (COVID-19)” (hereinafter referred to as “this Guide”). This Guide was prepared for persons in charge of COVID-19 control measures in their workplace. It should be used at the discretion of each business operator according to their workplace environment. The examples of infection control measures shown in this Guide are not guaranteed to work for all situations, and they do not limit or bind actual measures being put in place. When selecting actual measures, it is necessary to obtain new information and thoroughly understand individual cases and situations. This Guide was prepared based on findings and reports about the virus and response measures taken by the relevant ministries (Ministry of Health, Labour and Welfare, Ministry of Foreign Affairs, etc.) that could be confirmed as of December 15, 2020. Therefore, the contents of this Guide may need to be modified in the future, depending on changes in the situations mentioned above. In the preparation of this Guide, every effort has been made to ensure the accuracy of currently obtainable information. However, neither JSTH or JSOH shall be held liable for any unfavorable circumstance, such as loss and damage (including lost profits and various expenses), harmful rumors, etc. experienced by a business operator, his/her employees, and any other persons concerned as a result of various measures considered/implemented using this Guide by persons responsible for infection control in the workplace.
KRAS mutant rectal cancer cells interact with surrounding fibroblasts to deplete the extracellular matrix
Authors: Kim Jin K. , Marco Michael R. , Choi Seo‐Hyun , Qu Xuan , Chen Chin‐Tung , Elkabets Moshe , Fairchild Lauren , Chow Oliver , Barriga Francisco M. , Dow Lukas E. , O’Rourke Kevin , Szeglin Bryan , Yarilin Dmitry , Fujisawa Sho , Manova‐Todorova Katia , Paty Philip B. , Shia Jinru , Leslie Christina , Smith J. Joshua , Lowe Scott , Pelossof Raphael , Sanchez‐Vega Francisco , Garcia‐Aguilar Julio
Published: April 2021

Somatic mutations in the KRAS oncogene are associated with poor outcomes in locally advanced rectal cancer but the underlying biologic mechanisms are not fully understood. We profiled mRNA in 76 locally advanced rectal adenocarcinomas from patients that were enrolled in a prospective clinical trial and investigated differences in gene expression between KRAS mutant (KRAS‐mt) and KRAS‐wild‐type (KRAS‐wt) patients. We found that KRAS‐mt tumors display lower expression of genes related to the tumor stroma and remodeling of the extracellular matrix. We validated our findings using samples from The Cancer Genome Atlas (TCGA) and also by performing immunohistochemistry (IHC) and immunofluorescence (IF) in orthogonal cohorts. Using in vitro and in vivo models, we show that oncogenic KRAS signaling within the epithelial cancer cells modulates the activity of the surrounding fibroblasts in the tumor microenvironment.
Dosimetric feasibility of stereotactic ablative radiotherapy in pulmonary vein isolation for atrial fibrillation using intensity‐modulated proton therapy
Authors: Ren Xue‐Ying , He Peng‐Kang , Gao Xian‐Shu , Zhao Zhi‐Lei , Zhao Bo , Bai Yun , Liu Si‐Wei , Li Kang , Qin Shang‐Bin , Ma Ming‐Wei , Zhou Jing , Rong Yi
Published: April 2021

Purpose

To evaluate dosimetric properties of intensity‐modulated proton therapy (IMPT) for simulated treatment planning in patients with atrial fibrillation (AF) targeting left atrial‐pulmonary vein junction (LA‐PVJ), in comparison with volumetric‐modulated arc therapy (VMAT) and helical tomotherapy (TOMO).

Methods

Ten thoracic 4D‐CT scans with respiratory motion and one with cardiac motion were used for the study. Ten respiratory 4D‐CTs were planned with VMAT, TOMO, and IMPT for simulated AF. Targets at the LA‐PVJ were defined as wide‐area circumferential ablation line. A single fraction of 25 Gy was prescribed to all plans. The interplay effects from cardiac motion were evaluated based on the cardiac 4D‐CT scan. Dose‐volume histograms (DVHs) of the ITV and normal tissues were compared. Statistical analysis was evaluated via one‐way Repeated‐Measures ANOVA and Friedman’s test with Bonferroni’s multiple comparisons test.

Results

The median volume of ITV was 8.72cc. All plans had adequate target coverage (V23.75Gy ≥ 99%). Compared with VMAT and TOMO, IMPT resulted in significantly lower dose of most normal tissues. For VMAT, TOMO, and IMPT plans, Dmean of the whole heart was 5.52 ± 0.90 Gy, 5.89 ± 0.78 Gy, and 3.01 ± 0.57 Gy (P < 0.001), mean dose of pericardium was 4.74 ± 0.76 Gy, 4.98 ± 0.62 Gy, and 2.59 ± 0.44 Gy (P < 0.001), and D0.03cc of left circumflex artery (LCX) was 13.96 ± 5.45 Gy, 14.34 ± 5.91 Gy, and 8.43 ± 7.24 Gy (P < 0.001), respectively. However, no significant advantage for one technique over the others was observed when examining the D0.03cc of esophagus and main bronchi.

Conclusions

IMPT targeting LA‐PVJ for patients with AF has high potential to reduce dose to surrounding tissues compared to VMAT or TOMO. Motion mitigation techniques are critical for a particle‐therapy approach.
Neurofilament light chain is a cerebrospinal fluid biomarker in hereditary spastic paraplegia
Authors: Kessler Christoph , Serna‐Higuita Lina M. , Rattay Tim W. , Maetzler Walter , Wurster Isabel , Hayer Stefanie , Wilke Carlo , Hengel Holger , Reichbauer Jennifer , Armbruster Marcel , Schöls Ludger , Martus Peter , Schüle Rebecca
Published: April 2021

Objective

Despite the need for diagnostics and research, data on fluid biomarkers in hereditary spastic paraplegia (HSP) are scarce. We, therefore, explore Neurofilament light chain (NfL) levels in cerebrospinal fluid (CSF) of patients with hereditary spastic paraplegia and provide information on the influence of demographic factors.

Methods

The study recruited 59 HSP cases (33 genetically confirmed) and 59 controls matched in age and sex. Neurofilament light chain levels were assessed by enzyme‐linked immunosorbent assay. The statistical analysis included the effects of age, sex, and genetic status (confirmed vs. not confirmed).

Results

Levels of CSF NfL were significantly increased in patients with hereditary spastic paraplegia compared to controls (median 741 pg/mL vs. 387 pg/mL, p < 0.001). Age (1.4% annual increase) and male sex (81% increase) impacted CSF NfL levels in patients. The age‐dependent increase of CSF NfL levels was steeper in controls (2.6% annual increase). Thus, the CSF NfL ratio of patients and matched controls—expressing patients’ fold increases in CSF NfL—declined considerably with age.

Interpretation

CSF NfL is a reliable cross‐sectional biomarker in hereditary spastic paraplegia. Sex is a relevant factor to consider, as male patients have remarkably higher CSF NfL levels. While levels also increase with age, the gap between patients and controls is narrowing in older subjects. This indicates distinct temporal dynamics of CSF NfL in patients with hereditary spastic paraplegia, with a rise around phenotypic conversion and comparatively static levels afterward.
Effect of TU‐100 on Peyer's patches in a bacterial translocation rat model
Authors: Takasu Chie , Miyazaki Katsuki , Yoshikawa Kozo , Nishi Masaaki , Tokunaga Takuya , Kashihara Hideya , Yoshimoto Toshiaki , Ogawa Hirohisa , Morine Yuji , Shimada Mitsuo
Published: April 2021

Background

Daikenchuto (TU‐100), a Japanese herbal medicine, is widely used for various gastrointestinal diseases. We have previously reported that TU‐100 suppresses CPT‐11‐induced bacterial translocation (BT) by maintaining the diversity of the microbiome. In this study we show that TU‐100 modulates the immune response during BT by inducing PD‐1 expression in Peyer's patches.

Methods

Eighteen male Wistar rats were divided into four groups: a control group; a control + TU‐100 group, given TU‐100 1000 mg/kg orally for 5 d; a BT group, given CPT‐11 250 mg/kg intra‐peritoneal for 2 d; and a TU‐100 group, given TU‐100 1000 mg/kg orally for 5 d with CPT‐11 250 mg/kg intra‐peritoneal on days 4 and 5.

Results

The size of Peyer's patch was significantly bigger in the BT group compared to the control group (9.0 × 104 µm2 vs 29.4 × 104 µm2, P < .05), but improved in the TU‐100 group (15.4 × 104 µm2, P < .005). TU‐100 significantly induced PD‐1 expression in Peyer's patch compared to the control group and the BT group (control vs BT vs TU‐100 = 4.3 ± 4.9 vs 5.1 ± 10.3 vs 17.9 ± 17.8). The CD4+ cells were increased in the BT group (P < .05) compared to the control group but decreased in the TU‐100 group. The Foxp3+ cells were increased in the BT group compared to the control group (P < .05), and further increased in the TU‐100 group compared to the BT group. CPT‐11 significantly increased TLR4, NF‐κβ, TNF‐α mRNA expressions in the BT group. TU‐100 cotreatment significantly reversed these mRNA expressions.

Conclusion

TU‐100 may have a protective effect against BT through PD‐1 expression in Peyer's patch.
Communicating complex marine science: Does media format matter?
Authors: Kolandai‐Matchett Komathi , Armoudian Maria , Thrush Simon
Published: April 2021

Considering the deteriorated state of the ocean, maximizing the impact of scientists' media communication is essential for gaining public interest in, and support for, marine conservation initiatives. Unlike conventional news, that involves journalists' interpretation, the media Q&A offers scientists a way to convey insights on complex marine ecosystem issues in their own words. However, the Q&A approach for communicating marine science and conservation issues has not been empirically examined for format‐related differences in impact.

The modality theory in multimedia learning suggests that information is more effective for accelerating learning and retention when visuals are combined with narrated words, than when combined with written text. This study compared the effects of identically framed information presented in two different media Q&A formats: narrated multimedia (a video on YouTube) and written text with photos (designed to resemble an online mainstream news article). In both formats, a marine scientist explained the complex phenomenon of marine ecosystem tipping points.

In an online experiment, 116 respondents were randomly assigned to either view the video or read the article. Effects on knowledge, awareness, and motivation were tested for format‐related differences. Data related to time spent on the interventions were used to test the effects of attention retention on awareness and motivation gains. Additionally, the study gauged participants' assessment about the effectiveness of the communication features and framings used and suggestions for enhancing communication effectiveness.

Findings showed positive effects on knowledge, awareness, and motivation irrespective of media format, suggesting that other communication aspects such as the messengers, the scientist's communication style, educative content, and message frames probably contributed to impact. Findings also suggest that multimedia formats, which tend to be more costly, may not necessarily be more effective.