Enhanced heparan sulfate proteoglycan-mediated uptake of cell-penetrating peptide-modified liposomes

Protein transduction domains (PTDs) are used to enhance cellular uptake of drugs, proteins, polynucleotides or liposomes. In this study, functionalized Antennapedia (Antp, aa 43–-58) and HIV Tat (aa 47–57) peptides were coupled to small unilamellar liposomes via thiol-maleimide linkage. Modified liposomes showed higher uptake into a panel of cell lines including tumor and dendritic cells than unmodified control liposomes. Liposome uptake was time and concentration dependent as analyzed by flow cytometry and live-cell microscopy. At least 100 PTD molecules per small unilamellar liposome (100 ± 30 nm) were necessary for efficient translocation into cells. Cellular uptake of PTD-modified liposomes was 15- to 25-fold increased compared to unmodified liposomes and was inhibited by preincubation of liposomes with heparin. Glycosaminoglycan-deficient CHO cells showed dramatically reduced cell association of PTD-modified liposomes, confirming the important role of heparan sulfate proteoglycans in PTD-mediated uptake. Antp-liposomes used as carriers of the cytotoxic drug N⁴-octadecyl-1--D-arabinofuranosylcytosine-(5- 5)-3-C-ethinylcytidine showed a reduction of the IC₅₀ by 70% on B16F1 melanoma cells compared with unmodified liposomes. PTD-functionalized liposomes, particularly Antp-liposomes, represent an interesting novel carrier system for enhanced cell-specific delivery of a large variety of liposome-entrapped molecules.Received 16 April 2004; received after revision 13 May 2004; accepted 25 May 2004     

Effects of calcium-EGTA buffers on active calcium transport in inside-out red cell membrane vesicles

Summary In inside-out red cell membrane vesicles, the free calcium concentration half-maximally stimulating active calcium uptake is about 2 orders of magnitude smaller in a calcium-EGTA buffer than in media containing unbuffered calcium. In calcium-EGTA buffer, the maximum rate of calcium uptake is determined by the total calcium concentration present. A possible model for explaining these findings is presented.Acknowledgments. This work was supported by the Scientific Research Council, Ministry of Health, Hungary (6-03-0306-01-1/Gá). We thank for the valuable comments of Dr Ilma Szász and for the skilful technical assistance of Mrs M. Sarkadi.     

Localization of concanavalin-A binding sites inTetrahymena by scanning electron microscopy

Summary Binding sites of concanavalin-A were detected inTetrahymena on the body ciliature, while the cell membrane itself and the oral ciliature failed to bind the lectine.Supported by the Scientific Research Council, Ministry of Health, Hungary 1-01-0302-02-1/Cs.     

Time-qualified reference intervals — chronodesms

Summary Methods are presented for qualifying clinical reference intervals, for individuals as well as peer groups, according to circadian and other rhythms, using chronobiologically-defined single samples or time series.Supported by the Hoechst Italia Foundation, US Environmental Protection Agency (R804512-01-0), US Public Health Service (5-K6-GM-13, 981), National Cancer Institute (1R01-CA-14446-01), Cancer Supplement (CA-14445-01S1), National Institute of Aging (A6-00158) and National Institute of Occupational Safety and Health (OH 00631).     

Degenerative atrophy of central terminals of primary sensory neurons induced by blockade of axoplasmic transport in peripheral nerves

Summary Perineural colchicine- or vinblastine-treatment results in disappearance of non-lysosomal acid phosphatase activity, characterizing axon terminals in the Rolando substance of the normal spinal cord. Ultrastructural alterations are identical with those seen in the course of degenerative atrophy.Supported by research grant No. 4-01-0303-01-1 from the Hungarian Ministry of Health.     

Circadian variation in urinary melatonin in clinically healthy women in Japan and the United States of America

Summary Urinary melatonin excretion is lower in East-Asian (Japanese) than in North-American (whites of mixed ethnic origin) women. Moreover, a statistically significant circadian rhythm is demonstrated by population-mean cosinor in the data pool from both groups of women. Furthermore, statistical significance characterizes interactions of effects from geographic differences (between ethnic groups) with temporal factors. Such spatio-temporal interactions await further scrutiny with a view inter alia of carcinogenesis as it is influenced by a spectrum of intermodulating rhythms.Supported by U.S. Public Health Service (5-K6-GM-13981-17), National Cancer Institute (1R01-CA-14445-05 and N01-CP-5-5702), National Institute of Occupational Safety and Health (OH-00631-01), National Institute of Aging (AG-00158), Environmental Protection Agency (R804513-01-0), Swedish Medical Research Council grant 3371 and the St Paul Ramsey Medical Research and Education Foundation. R.B. Sothern and Jung-Keun Lee, Scientists, Chronobiology Laboratories, University of Minnesota provided valuable help.     

Synthesis and pharmacological properties of [1-N4-dimethyl-asparagine]-angiotensin II

Summary [1-N⁴-Dimethyl-asparagine]-angiotensin II was synthesized by Merrifield's solid-phase procedure. The analogue gave rat blood pressure about 70% relative potency to Hypertensin (Ciba). Rabbit aorta strips gave intrinsic activity _E=1, a PD₂ of 6.92±0.09 and an affinity relative to [Asn¹]-angiotensin II of 6.5%.The editors do not hold themselves responsible for the opinions expressed in the authors' brief reports. — Les auteurs sont seuls responsables des opinions exprimées dans ces brèves communications. — Für die Kurzmitteilungen ist ausschliesslich der Autor verantwortlich. — Per le brevi comunicazioni è responsabile solo l'autore. — . — Solo los autores son responsables de las opiniones expresadas en estas comunicationes breves.Acknowledgment. The authors wish to express their appreciation to Dr Emanuel Escher, School of Medicine, Centre Hospitalier Universitaire, Sherbrooke, Quebec, for his help in providing the biological data and critical comments.     

FGFR1、FGFR3和Pokemon在膀胱癌中的表达及相关性研究

目的通过观察FGFR1、FGFR3和Pokemon在膀胱癌组织的表达情况,探讨它们在膀胱癌发生、发展中的作用。方法采用免疫组化检测296例膀胱癌和48例正常膀胱组织中FGFR1、FGFR3和Pokemon蛋白的表达情况。结果 1)FGFR1在膀胱癌不同病理分期、分级及初发复发的表达率为:Ta-T1期7.5%,T2-T4期18.1%(p0.01);低级别2.8%,低-高级别11.9%(p0.05),高级别14.3%的(p0.01);初发表达6.8%,复发表达率17.1%(p0.01)。2)FGFR3在膀胱癌不同病理分期、分级的表达率为:Ta-T1期68.1%,T2-T4期35.0%(p0.01));低级别75.0%,低-高级别60.3%(p0.05),高级别44.9%(p0.01)。3)Pokemon在膀胱癌不同病理分级及初发复发的表达率为:低级别47.2%,低-高级别66.7%(p0.01),高级别80.6%(p0.01);初发59.2%,复发80.0%(p0.01)。结论 FGFR1在膀胱癌中的表达与病理分级、分期成正相关,复发表达明显高于初发;Pokemon在膀胱尿路上皮癌中的表达与病理分级成正相关,复发表达也明显高于初发,表明它们可能主要参与膀胱癌发展及转移等过程。FGFR3在膀胱癌中的表达与病理分级、分期成负相关,表明它可能在膀胱癌发展早期及低级别肿瘤发展中发挥着重要作用。     

Myelination in chronically-alcoholic mice

Summary Chronically-administered ethanol, not impaired nutrition, exerts a specific effect to decrease myelination as measured by the activity of the marker enzyme for myelin, 2, 3-cyclic nucleotide 3-phosphohydrolase.Acknowledgment. This work was supported in part by N.I.H. Biomedical Research Support Grant Program, Grant No. 1-507-RR07155-01. To whom reprint requests should be addressed.     

Effect of 6-hydroxydopamine pretreatment on spontaneous convulsions induced by barbital withdrawal

Summary Follwing withdrawal from chronic barbital administration, 6-hydroxydopamine pretreated rats show a greater number and an earlier onset of spontaneous convulsive seizures than do rats pretreated with the saline-ascorbic acid vehicle.Supported by NIMH Grant No. 5 RO1 DA00755-02.Recipient of Research Scientist Development Award, No. 1 KO2 MH00028-01.     

Effectiveness of phosphocitrate and N-sulpho-2-amino tricarballylate,a new analogue of phosphocitrate,in blocking hydroxyapatite induced crystal growth and calcium accumulation by matrix vesicles

Summary Phosphocitrate and its analogue N-sulpho-2-amino tricarballylate were compared with ethane-1-hydroxy-1, 1-diphosphonate for inhibition of calcium phosphate crystallization in hydroxyapatite induced crystal growth and⁴⁵Ca uptake by matrix vesicles. Phosphocitrate (1 M) was the most potent inhibitor followed by ethane-1-hydroxy-1, 1-diphosphonate and N-sulpho-2-amino tricarballylate, the latter requiring a high concentration (100 M) to be equally effective as an inhibitor.Acknowledgments. We wish to thank Mr J. Jordan and Miss L.C. Ward for the excellent technical assistance and Mr R.J. Tennant for the transmission electron microscopy. We also thank the Golden Poultry Farming Industries Ltd., Hobart, Tasmania for their generous supply of broiler strain chickens.     

Photoreceptor properties of an ectopic eye in the fleshfly,Sarcophaga bullata

Summary Ectopic eyes were produced on the fleshfly,Sarcophaga bullata by transplantation of imaginal eye discs. Electrophysiological and histological observations of these supernumerary eyes indicate the absence of synaptic connections between retinular cells and higher order neurons.Supported by an University of New Brunswick research grant to P.S. and NSF grant BNS-76-11921 and NIH grant 1-R01-EY-02487-01A1 to W.S.S.We would like to thank Dr I.A. Meinertzhagen for suggestions and Mary Johnson for technical assistance.     

Changes in body temperature produced by injecting prostaglandin E1, EGTA and bacterial endotoxins into the PO/AH region and the medulla oblongata of the rat

Résumé La prostaglandine E₁ a causé de l'hyperthermie si on l'injectait dans la région préoptique/hypothalamique antérieure/des rats et de l'hypothermie au niveau de la région médullaire oblongue. L'EGTA, le Piromen et le vaccin typhoïdes ont rarement causé des changements parallèles dans la température rectale si on les injectait dans les deux régions cérébrales.

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This research was supported by PHS Research Grant No. 1-R01-NS-10046-01 from the National Institute of Neurological Diseases and Stroke. We thank Dr.J. Pike of Upjohn Laboratories for supplying the PGE₁.

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Medical student fellow in Psychiatry. Supported by the Hogg Foundation and PHS Grant No. 5 TO2 MH 5928 from the National Institute of Mental Health.     

Free radicals produced in a nitrosofluorene-unsaturated lipid reaction

Summary We report here the first demonstration that the carcinogen 2-nitrosofluorene reacts directly with lipid molecules containing carbon-carbon double bonds to yield free radicals which appear to be the nitroxyl free radical of the carcinogen covalently bound to the lipid.This investigation was supported in part by the U.S. Public Health Service Grant 5 RO1 CA18591-01 from the National Institutes of Health.     

Protohemin in bile during primate development

Summary Protohemin is excreted in biles of primate fetuses and premature newborns. It also was found in relatively large quantities in the lower ileum and mesenteric lymph-nodes of newborn primates.This work was supported by National Institute of Child Health and Human Development, grants 1RO1 HD 07331-02 and 1F22 HD 00191-01.     

Bovine microvascular endothelial cells of separate morphology differ in growth and response to the action of interferon-γ

Five cell types recently isolated from the bovine corpus luteum differed in their epithelioid morphology and their cytoskeleton, but shared common criteria of microvascular endothelial cells^1,2. To give strong evidence for the separate entity, the growth rate of the 5 phenotypically different cells was studied. They were seeded at low density on day 0. Most of these cells were treated with 200 to 1000 U recombinant bovine interferon- (IFN-) for 3 days. The untreated remainder served as controls. Cell counts were made for all cultures on days 4, 7, 10 and 13. morphology: 13 d after treatment with IFN- senescent cells as well as intact cells occurred in cultures of cell types 1 to 4. Cultures of cell type 5 were apparently unchanged and resembled their untreated counterparts. Desminpositive cells in cultures of cell type 2 developed cell processes. Growth rate: In the absence of IFN-, the growth rate was high for cell types 3 and 4, moderate for cell type 1, and low for cell types 2 and 5. The presence of IFN- caused anti-proliferative effects. These were higher for cell types 3 and 4 than for cell types 1 and 2. IFN- could be cytotoxic on cell type 3. In contrast, the cytokine tended to support the cell growth of cell type 5. These findings substantiate the postulate that endothelial cells exhibiting separate morphology in culture also function differently.     

Synthesis of 5′-deoxypyridoxal derivatives

Summary A convenient synthesis of 5-deoxypyridoxal derivatives is described. The method involves catalytic hydrogenolysis of the corresponding 5-phosphorylated derivatives; products are obtained in high yields.Supported by PHS Grant No. 5429-16-9.     

A novel cannabinoid containing a 1,8-cineol moiety

Summary The synthesis of a novel cannabinoid containing a 1,8-cineol moiety (1) is described. It is much less biologically active than ¹-tetrahydrocannabinol.Acknowledgment. This work was supported by NIDA (grant No. DA-00574-01).     

(−)-Δ8-tetrahydrocannabinol: Two novel in vitro metabolites

Zusammenfassung (–)-⁸-Tetrahydrocannabinol ist einer der Aktivstoffe in Marihuana (Cannabis sativa L.). Inkubation dieser Verbindung mit der überstehenden Zentrifugationsfraktion (9000g) aus männlicher Hundeleber ergab zwei Hauptmetaboliten, welche durch Massenspektrometrie und Kernresonanzspektroskopie identifiziert wurden. Den beiden Verbindungen werden die Strukturen des 1-Hydroxy-⁸-tetrahydrocannabinols und des 3-Hydroxy-⁸-tetrahydrocannabinols zugeordnet.     

Interaction of extrinic fluoresence probes with E. coli glutamine synthetase

Summary Binding of 2-p-toluidinylnaphthalene-6-sulfonate (TNS) to adenylylated glutamine synthetase is cooperative and time-dependent, with 3 dye sites per subunit. In fluorescence polarization experiments TNS and pyrene butyrate give normalized Perrin plots that indicate a symmetrical arrangement of dye excited state dipoles, relative to the rotational axis of the oblate ellipsoid of the dodecameric native enzyme.Acknowledgment. Drs G. Weber and L. Brand are thanked for helpful comments on this work. — Supported in part by grants from the National Science Foundation (BMS 72-02165 and GU-3182). Portions of this work were presented in preliminary form at the 9th International Congress of Biochemistry, Stockholm, Sweden, July 1973.