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Biomedical subjects

R Jordan

Publications and source records attributed to R Jordan.

At least 19 recordsLinked to original sources
PubMed1992

Development of a monoclonal antibody to the conserved region of p34cdc2 protein kinase.

Mice and rabbits were injected with various forms of a 16 amino acid synthetic peptide representing PSTAIR, the evolutionarily conserved region of the protein kinase p34cdc2, for polyclonal antisera and hybridoma-monoclonal antibody production. Antisera from mice injected with an unconjugated monomeric form of the peptide showed no reaction to the peptide. Of four animals injected with the monomeric form of the peptide conjugated to keyhole limpet hemocyanin via m-maleimidobenzoyl-N-hydroxysulfosuccinimide (MBS), antisera from only one mouse had a very low titer to the peptide, and all four animals produced antibody to the MBS bridge. Both mice injected with an octameric multiple antigen peptide (MAP) of PSTAIR produced antisera reactive to the octameric MAP form of the peptide in ELISA and also to the cdc2 protein expressed in bacteria in an immunoblotting assay. Splenocytes from one mouse injected with the octameric MAP form of the peptide were successfully used for hybridoma-monoclonal antibody production. A monoclonal antibody was produced that reacted with octamer, monomer and cdc2-expressed protein and specifically with the carboxyl terminus of the 16 amino acid peptide.

Amino Acid Sequence↗
PubMed1992

Synthesis and biological activity of trans(+-)-N-methyl-2-(3-pyridyl)-2-tetrahydrothiopyrancarbothioamide 1-oxide (RP 49356) and analogues: a new class of potassium channel opener.

The synthesis and biological activity of trans-(+-)-N-methyl-2-(3-pyridyl)-2-tetrahydrothiopyrancarbothioamid+ ++ e 1-oxide (8a, RP 49356) and analogues is reported. These compounds constitute a new structural class of K(+)-channel opener. The effects of changes in pyridyl group, thioamide, and thiane ring on in vitro K(+)-channel opening reactivity are discussed. A 3-pyridyl or 3-quinolyl group, a small N-alkyl thioamide function, and a thiane oxide ring, in which the sulfoxide is in a trans relationship to the thioamide, are preferred for activity. Selected compounds were tested intravenously in the normotensive anaesthetized rat for hypotensive effects, and the activities reflect their in vitro K(+)-channel opening activity. This led to further evaluation of compound 8a and the selection of the (-)-enantiomer 8b (RP 52891) for development as an antihypertensive and antianginal agent.

Animals↗
PubMed1992

Different conformations for the same polypeptide bound to chaperones DnaK and GroEL.

The proteins DnaK (hsp70) and GroEL (cpn60) from Escherichia coli are prototypes of two classes of molecular chaperones conserved throughout evolution. The analysis of transferred nuclear Overhauser effects in two-dimensional NMR spectra is ideally suited to determine chaperone-bound conformations of peptides. The peptide vsv-C (amino-acid sequence KLIGVLSSLFRPK) stimulates the ATPase of BiP and Hsc70 (ref. 3) and the intrinsic ATPase of DnaK. The affinity of the vsv-C peptide for DnaK is greatly reduced in the presence of ATP. Here we analyse transferred nuclear Overhauser effects and show that the peptide is in an extended conformation while bound to DnaK but is helical when bound to GroEL. NMR also indicates that the mobility of the peptide backbone is reduced more by binding to DnaK than by binding to GroEL, whereas the side chains are less mobile when bound to GroEL.

Amino Acid Sequence↗
PubMed1992

Vein grafts in reconstructive microsurgery of the lower extremity.

This paper describes surgical techniques and results of lower extremity microsurgical reconstruction using vein grafts. During a 5 year period, free tissue transfers were used to reconstruct 118 lower extremity defects in 109 patients. In 28 of these cases (24%), saphenous vein grafts were used when reconstruction would otherwise have been difficult or impossible. A preliminary arteriovenous fistula was established between the proximal arterial source and the final venous drainage in the majority (26) of cases. Venous drainage was left intact at the point of entry into the deep venous system in 23 cases. Three complications included anastomotic clotting in 2 patients and postoperative blow-out of the vein graft in another; all were treated without flap loss. Flap survival was 86% in vein grafted patients and 94% in those cases in which vein grafts were not used. Flap loss was related in each case to the nature of the pre-existing condition, rather than to vein grafting per se.

Arteriovenous Shunt, Surgical↗
PubMed1992

Potyviruses, monoclonal antibodies, and antigenic sites.

Virus-specific and cross-reactive monoclonal antibodies have been produced to at least 19 different aphid-transmitted potyviruses. This report summarizes the development of these monoclonal antibodies as well as presents information on the delineation of the virus-specific and group-common epitopes defined by these monoclonal antibodies. Virus-specific and group-common antigenic determinants were mapped by a variety of techniques, including analysis of antigen: antibody reactivity patterns, determination of N-terminal vs. trypsin-resistant core peptide-specificity, immunoanalysis of overlapping synthetic peptides, and immunoanalysis of bacterially expressed coat-protein gene products. Of those monoclonal antibodies that have been examined, monoclonal antibody-defined virus-specific epitopes are virion surface-located within the 30+ amino acid amino terminus, whereas the group-common epitopes are found in the trypsin-resistant core protein not usually located on the virion surface, as has been shown previously with certain polyclonal antibodies. New information is presented on the analysis of bean yellow mosaic virus amino terminal epitopes as well as on the identification of amino terminal antigenic determinants shared between strains of bean yellow mosaic virus and pepper mottle virus. A recommendation on the evaluation and use of a panel of potyvirus broad-spectrum reacting monoclonals as reference monoclonal antibodies for the detection and classification of aphid-transmitted potyviruses is also presented.

Amino Acid Sequence↗
PubMed1992

Evidence that pepper mottle virus and potato virus Y are distinct viruses: analyses of the coat protein and 3' untranslated sequence of a California isolate of pepper mottle virus.

Pepper mottle virus (PepMoV) is a member of the large and complex genus Potyvirus, and is classically distinguished from other members of the genus by differential host range and cytopathology as well as serology of the coat protein and cytoplasmic inclusion body proteins. Here we report the deduced amino acid sequence of the coat protein of a California potyvirus identified by a variety of classical methods as PepMoV (PepMoV C). Comparison of the 3' untranslated nucleic acid sequence and the deduced coat-protein amino acid sequence of the PepMoV C isolate with those of PVY and other potyviruses indicates that PepMoV C is sufficiently diverged to be considered a distinct virus species. Thus, comparative sequence analyses of the PepMoV C isolate support earlier serological and biological evidence that PepMoV and PVY are distinct viruses.

Amino Acid Sequence↗
PubMed1991

Inhibition of transient gene expression with plasmids encoding herpes simplex virus type 1 UL55 and alpha genes.

Herpes simplex virus type 1 (HSV-1) subgenomic sequences from 0.743 to 0.782 map units have been molecularly cloned as plasmid AT1 and shown to inhibit stable DNA-mediated gene transformation of Ltk- cells with the HSV-1 thymidine kinase (tk) gene. Here it is shown that AT1 also inhibits transient gene expression. Expression from the chloramphenicol acetyltransferase (CAT) gene under the control of either the HSV-1 tk gene or the Rous sarcoma virus (RSV) promoter was inhibited when cotransfected into Ltk- and CV-1 cells with equimolar amounts of AT1. AT1 was subcloned as three overlapping plasmids called AT1a, alpha 27 and AT1b. The alpha 27 plasmid encodes the HSV-1 immediate early gene, alpha 27; AT1a possesses sequences that specify an open reading frame in HSV-1 strain KOS used in these studies, although the significance of this open reading frame is unknown; AT1b possesses the sequences for UL55 and UL56, also genes for which no function has been reported. No single subclone or pair of subclones demonstrated significant inhibition of transient gene expression. Cotransfection of all three subclones did result in inhibition of RSV-CAT gene expression, suggesting that information from each subclone is necessary. One of the three subclones, alpha 27, contains the HSV-1 immediate early gene, alpha 27, so the possibility that other immediate early genes could substitute for alpha 27 was tested. Inhibition of RSV-CAT gene expression was also achieved by cotransfection of AT1a and AT1b with either an alpha 0- or alpha 4-containing plasmid, suggesting that the role of the alpha 27-containing plasmid can be replaced by other alpha genes with trans-regulating capability. Finally, AT1a and AT1b linker insertion mutants have been constructed and used to study the role these plasmids play in mediating inhibition. These results suggest that AT1 contains HSV-1 functions in addition to that of alpha 27 that interfere with gene expression.

Animals↗
PubMed1991

Comparison and differentiation of potyvirus isolates and identification of strain-, virus-, subgroup-specific and potyvirus group-common epitopes using monoclonal antibodies.

A panel of monoclonal antibodies (MAbs) generated against an admixture of 12 potyvirus isolates was used to compare and differentiate diverse potyviruses. Both native and denatured virions of strains of bean yellow mosaic (BYMV), potato virus Y, tobacco etch, pea seed-borne mosaic, iris severe mosaic, iris mild mosaic and asparagus virus-1 potyviruses were used as immunogen and as antigen for screening of the hybridoma cell lines. Thirty cell lines secreting potyvirus-specific antibodies reactive in indirect antigen-coated plate (ACP-) ELISA were selected for detailed analysis. All 30 MAbs reacted with at least one strain of BYMV; 11 MAbs reacted with between one and eight of the nine BYMV strains and an additional three MAbs reacted only with isolates within the BYMV subgroup (BYMV, pea mosaic virus and clover yellow vein virus). The remaining 16 MAbs reacted with a BYMV isolate and with at least one of the other 43 potyvirus isolates tested. MAb PTY 1 reacted with all 55 potyvirus isolates tested (representing at least 33 different and distinct aphid-transmissible potyviruses). The potyvirus cross-reactive MAbs generally gave higher reactivity values in ACP-ELISA with dissociated virus than with polyclonal antibody-trapped intact virions in triple antibody sandwich ELISA (i.e. were cryptotope-specific). The BYMV strain- and virus-specific MAbs reacted strongly with both types of antigens (i.e. were metatope-specific). At least 25 distinct epitopes (12 cryptotopes and 13 metatopes) could be identified from the MAb-antigen reactivity patterns. The distribution of these epitopes between virus isolates can be used to detect and differentiate potyviruses in infected plant extracts and to examine virus architectures. Some of these epitopes are shared by potyvirus isolates not previously shown to be serologically related. The broad spectrum-reacting MAb PTY 1 recognizes a cryptotope conserved on all of the aphid-transmissible potyviruses examined and should be a valuable tool for the detection and assay of these potyviruses.

Antibodies, Monoclonal↗
PubMed1990

Pharmacodynamic study of F(ab')2 fragments of murine monoclonal antibody 7E3 directed against human platelet glycoprotein IIb/IIIa in patients with unstable angina pectoris.

The pharmacodynamics of intravenous bolus injections of 0.05, 0.10, 0.15, and 0.20 mg/kg of F(ab')2 fragments of the murine monoclonal antibody 7E3, 7E3-F(ab')2, directed against the glycoprotein IIb/IIIa (GPIIb/IIIa) receptor of human platelets, were studied in groups of four patients with unstable angina pectoris. With 0.20 mg/kg, the template bleeding time prolonged from 6.3 +/- 1.9 (mean +/- SD) to greater than 30 min; it subsequently decreased to 13 +/- 7.8 min after 12 h and to 8.3 +/- 1.5 min after 24 h. The number of unblocked GPIIb/IIIa receptors (preinfusion value, 32,000 +/- 3,000 per platelet) decreased to 13 +/- 7% of the preinfusion value 1 h after infusion, and then increased to 33 +/- 10% at 12 h, 44 +/- 8% at 24 h and 67 +/- 7% at 72 h. The logarithm of the bleeding time was inversely proportional with the residual GPIIb/IIIa receptors (r = 0.73, P less than 0.0001). ADP-induced platelet aggregation (measured by changes in light transmittance in percent) decreased from 60 +/- 5% before infusion to 1.5 +/- 3% 1 h after infusion; it then increased to 29 +/- 3% after 24 h and 39 +/- 6% after 72 h. Platelet counts decreased by 16% at 1 h and returned to control values within 24 h. Proportionally smaller effects were seen at lower doses of 7E3-F(ab')2. Antibody injection did not induce spontaneous bleeding. Angina was not observed during the first 12 h when the bleeding time was significantly prolonged, but occurred in 6 of the 16 patients within the next 3 d. 2 of the 16 patients developed low titers of IgG antibodies specific for 7E3-F(ab')2. Thus 7E3-F(ab')2 induces dose-related inhibition of platelet function; at a dose of 0.20 mg/kg, it causes profound inhibition of platelet aggregation and prolongation of the bleeding time, but no spontaneous bleeding.

Adult↗
PubMed1989

Role of selective cyclic GMP phosphodiesterase inhibition in the myorelaxant actions of M&B 22,948, MY-5445, vinpocetine and 1-methyl-3-isobutyl-8-(methylamino)xanthine.

1. The mechanism by which M&B 22,948, MY-5445, vinpocetine and 1-methyl-3-isobutyl-8-(methylamino)xanthine (MIMAX), which have been described as selective cyclic GMP phosphodiesterase (PDE) inhibitors, relax rat aorta was investigated. 2. Three cyclic nucleotide PDEs were identified in the soluble fraction of rat aorta; a Ca2+-insensitive form exhibiting substrate selectivity for cyclic GMP (cGMP PDE), a Ca2+/calmodulin-stimulated form which also preferentially hydrolyzed cyclic GMP (Ca2+ PDE), and a form demonstrating substrate selectivity for cyclic AMP (cAMP PDE). 3. M&B 22,948 and MIMAX inhibited cGMP PDE (Ki = 0.16 microM and 0.43 microM, respectively) and Ca2+ PDE (Ki = 9.9 microM and 0.55 microM, respectively), but exhibited weak activity against cAMP PDE (Ki = 249 microM and 42 microM, respectively). MY-5445 selectivity inhibited cGMP PDE (Ki = 1.3 microM) and vinpocetine selectively inhibited Ca2+ PDE (Ki = 14 microM). 4. M&B 22,948 and MIMAX induced dose-dependent increases in the accumulation of cyclic GMP, but not cyclic AMP, in rat aorta pieces. These effects were greatly reduced by endothelial denudation and by methylene blue (5 microM) which blocks the actions of endothelium-derived relaxant factor. MY-5445 and vinpocetine had no effect on rat aorta cyclic GMP or cyclic AMP accumulation. 5. All four compounds caused dose-related relaxation of 5-hydroxytryptamine (10 microM) contracted, endothelium-intact rat aorta, the effects of M&B 22,948 and MIMAX being greatly reduced by methylene blue (5 microM). Methylene blue also caused 10 fold and 100 fold rightward shifts in the dose-response curves of MY-5445 and vinpocetine, respectively. 6. The results are consistent with the smooth muscle relaxant actions of M&B 22,948 and MIMAX, but not vinpocetine and MY-5445, being mediated through a mechanism involving inhibition of cyclic GMP hydrolysis.

1-Methyl-3-isobutylxanthine↗
PubMed1989

Abolition of in vivo platelet thrombus formation in primates with monoclonal antibodies to the platelet GPIIb/IIIa receptor. Correlation with bleeding time, platelet aggregation, and blockade of GPIIb/IIIa receptors.

We studied the dose-response effects of the F(ab')2 fragments of murine monoclonal antibodies to the platelet GPIIb/IIIa receptor (7E3 and 10E5) on in vivo platelet thrombus formation in a well-characterized monkey model in which the carotid artery is stenosed and thrombus formation is provoked and augmented by intimal damage and the infusion of subaggregating doses of epinephrine. Both antibodies abolished thrombus formation with a mean dose of -0.2 mg/kg. Ex vivo platelet aggregation was not always abolished at doses that abolished thrombus formation; similarly, bleeding times were only moderately prolonged (9.1 +/- 1.4 minutes) at these doses. Increasing the dose above that required to abolish thrombus formation consistently produced abolition of ex vivo platelet aggregation, marked prolongation of the bleeding time (14.2 +/- 1.5 minutes), and nearly quantitative blockade of GPIIb/IIIa receptors. We conclude that in a significant percentage of animals, the extent of GPIIb/IIIa blockade required to prevent vasoocclusive thrombus formation in this model is less than that required for abolition of platelet aggregation, and that the preservation of only a minority of functional GPIIb/IIIa receptors might be adequate to maintain a nearly normal bleeding time.

Animals↗
PubMed1988

Herpes simplex virus type 1 alpha gene containing plasmids can inhibit expression regulated from an alpha promoter in CV-1 but not HeLa cells.

Transfection of plasmids containing the herpes simplex virus type 1 (HSV-1) alpha gene 27 has been observed to inhibit gene expression from a virus alpha promoter in monkey (CV-1) but not human (HeLa) cells in a transient gene expression system. DNA mediated gene transfer to CV-1 and HeLa cells of the bacterial gene for chloramphenicol acetyl transferase (CAT) linked to the promoter regulatory domain from the HSV-1 alpha 4 gene results in production of substantial levels of CAT enzyme. Cotransfection of equal mole amounts of an alpha 27 containing plasmid with an alpha 4-CAT construct to CV-1 cells results in a greater than 85% inhibition of CAT activity. No significant inhibition of CAT activity was observed when transfection was done in HeLa cells, with the same concentrations tested. Intact alpha 27 structural genes were necessary to achieve inhibition since subgenomic fragments and restriction enzyme digested alpha 27 genes were not effective inhibitors. Cotransfection of alpha 27 genes to CV-1 cells also prevented alpha 0 as well as alpha 4 from mediating their trans-stimulation of the HSV thymidine kinase (tk) regulated CAT gene, B-CAT. This suggests that the alpha 27 gene product may down-regulate gene expression from alpha promoters.

Animals↗
PubMed1988

Osteoblasts increase their rate of division and align in response to cyclic, mechanical tension in vitro.

Bone adapts to physical deformation in vivo, yet the mechanism of the adaptive process remains unknown. One reason for this perplexity has been the difficulty in examining the effects of a well-defined deformation regimen on individual bone cells. With the utilization of novel, flexible-bottomed cell culture plates, one can study the effects of cyclic strain on the morphologic and biochemical adaptations of individual osteoblasts in vitro. Avian, calvarial osteoblast-like cells, from passes 2-5, responded to cyclic strain, by increasing their rates of DNA synthesis and cell division during the first 72 h after initiation of a continuous deformation regimen comprised of 3 cycles per min of 0-24% elongation. In addition, within hours after initiation of the deformation regimen, cells oriented 90 degrees to the applied strain field at the periphery of the culture plate in the region of maximum strain and elongation.

Animals↗
PubMed1988

Antithrombin-III prevents the lethal effects of Escherichia coli infusion in baboons.

Infusion of Escherichia coli (LD100) was followed by coagulopathic and cell injury responses, cardiovascular collapse, and death in 18 to 32 hr in four out of four baboons. Infusion of AT-III in sufficient amounts to achieve AT-III levels of more than 4 units/ml of plasma before and during the infusion of E. coli reduced the intensity of the coagulopathic and cell injury response and prevented vascular collapse and death in four out of four baboons. Failure to achieve AT-III levels of more than six units/ml at T +60 min during the infusion of E. coli resulted in failure to prevent its lethal effects in three out of three baboons even though levels as high as 10 units/ml were achieved later in the course of the experiment. These studies suggest that thrombin and/or its products can contribute to the inflammatory response to E. coli and that AT-III is of potential value as a prophylactic but not as a therapeutic agent in the treatment of patients at high risk of developing gram negative sepsis.

Animals↗
PubMed1987

Beta-adrenergic activities of octopamine and synephrine stereoisomers on guinea-pig atria and trachea.

The activities of the (-)- and (+)-forms of m- and p-octopamine and m- and p-synephrine on beta 1- and beta 2-adrenoceptors in guinea-pig atria and trachea have been compared with that of noradrenaline. The rank order of potency of the (-)-forms on beta 1-adrenoceptors was noradrenaline greater than m-synephrine greater than m-octopamine = p-octopamine greater than p-synephrine. m-Synephrine was 100-fold, m- and p-octopamine about 6000-fold, and p-synephrine about 40,000-fold less active than noradrenaline. The (+)-forms were 1-2 orders of magnitude less active than their (-)-counterparts. The four (-)-compounds were more than four orders of magnitude less active than noradrenaline on beta 2-adrenoceptors, and the (+)-forms had no detectable activity in concentrations as high as 10(-4) M. If m- and p-octopamine are co-released with noradrenaline in amounts proportional to their concentration, their activities at these structures are too low to be physiologically significant.

Adrenergic beta-Agonists↗