Biomedical subjects

J M Li

Publications and source records attributed to J M Li.

At least 19 recordsLinked to original sources

PubMed1996

Functional fetal nigral grafts in a patient with Parkinson's disease: chemoanatomic, ultrastructural, and metabolic studies.

A patient with Parkinson's disease received bilateral fetal human nigral implants from six donors aged 6.5 to 9 weeks post-conception. Eighteen months following a post-operative clinical course characterized by marked improvement in clinical function, this patient died from events unrelated to the grafting procedure. Post-mortem histological analyses revealed the presence of viable grafts in all 12 implant sites, each containing a heterogeneous population of neurons and glia. Approximately 210,146 implanted tyrosine hydroxylase-immunoreactive (TH-ir) neurons were found. A greater number of TH-ir grafted neurons were observed in the right (128,162) than the left (81,905) putamen. Grafted TH-ir neurons were organized in an organotypic fashion. These cells provided extensive TH-ir and dopamine transporter-ir innervation to the host striatum which occurred in a patch-matrix fashion. Quantitative evaluations revealed that fetal nigral grafts reinnervated 53% and 28% of the post-commissural putamen on the right and left side, respectively. Grafts on the left side innervated a lesser area of the striatum, but optical density measurements were similar on both sides. There was no evidence that the implants induced sprouting of host TH-ir systems. Electron microscopic analyses revealed axo-dendritic and occasional axo-axonic synapses between graft and host. In contrast, axo-somatic synapses were not observed. In situ hybridization for TH mRNA revealed intensely hybridized grafted neurons which far exceeded TH mRNA expression within residual host nigral cells. In addition, gamma-amino butyric acid (GABA)-ergic neurons were observed within the graft that formed a dense local neuropil which was confined to the implant site. Serotonergic neurons were not observed within the graft. Cytochrome oxidase activity was increased bilaterally within the grafted post-commissural putamen, suggesting increased metabolic activity. In this regard, a doubling of cytochrome oxidase activity was observed within the grafted post-commissural putamen bilaterally relative to the non-grafted anterior putamen. The grafts were hypovascular relative to the surrounding striatum and host substantia nigra. Blood vessels within the graft stained intensely for GLUT-1, suggesting that this marker of blood--brain barrier function is present within human nigral allografts. Taken together, these data indicate that fetal nigral neurons can survive transplantation, functionally reinnervate the host putamen, establish synaptic contacts with host neurons, and sustain many of the morphological and functional characteristics of normal nigral neurons following grafting into a patient with PD.

Carrier Proteins↗

PubMed1996

A universal tag for recombinant proteins.

Incorporation of tags into recombinant proteins can facilitate their identification and purification. In addition, these tags can also be used to monitor the trafficking or localization of the recombinant proteins inside the cells. Several such tags have been developed. However, the lengths of these tags make it cumber-some to incorporate them into the desired proteins. Typically, one must subclone the desired cDNA into a plasmid containing the tag sequence at a suitable restriction site or ligate a synthetic oligonucleotide containing the tag sequence at a suitable restriction site in the cDNA of the desired protein. These manipulations can be avoided, if one uses a short peptide tag that can be incorporated by PCR. We show here that a short peptide tag, RYIRS can be easily incorporated at the C termini of recombinant proteins by PCR. We also showed that by using a mAb specific for this peptide sequence, the tagged proteins could be easily detected in Western blot analysis, immunofluorescence staining, and immunoprecipitation. The interactions between this tag sequence and the mAb have been well characterized. One can take advantage of this information and control the reactivities between the tagged proteins and the mAb by varying the lengths of the peptide tags. Furthermore, we showed that this tag can be used to monitor whether a recombinant protein is properly translated and terminated because the interactions between this tag sequence and the mAb requires that the tag be at the C-terminus of the protein.

Amino Acid Sequence↗

PubMed1996

Placental and fetal cardiac laminin are targets for cross-reacting autoantibodies from mothers of children with congenital heart block.

The association of congenital heart block (CHB) with maternal autoantibodies to the Ro and La ribonucleoprotein antigens may be due to cross-reactions between maternal anti-La antibodies and fetal cardiac specific antigens. One of the major components of cardiac myocytes, laminin, is accessible for binding by maternal autoantibodies and we have previously reported cross-reactivity of mouse laminin with anti-La antibodies affinity purified from the sera of patients with primary Sjögren's syndrome. Affinity purified anti-La antibodies from ten women who had at some time given birth to a child with CHB were examined for cross-reactivity with human placental laminin, which shares structural similarities with cardiac laminin. All ten anti-La antibodies bound to the surface of cryosections of normal full term placental trophoblasts. Binding could be inhibited by pre-incubation of antibodies with either La or placental laminin. Eight anti-La antibodies also reacted with placental laminin by ELISA and La inhibited up to 82% of binding to laminin while laminin inhibited up to 85% of binding to La in a dose dependent manner. Eight anti-La antibodies also bound to the surface of fetal cardiac myocytes at 10.3 weeks of gestation and five showed lower levels of reactivity with the surface of fetal cardiac myocytes at 16.5 weeks of gestation. None showed any surface staining of normal adult heart. These data confirm the cross-reactivity of anti-La antibodies with laminin and may support a placental role in preventing the majority of potentially pathogenic antibodies from reaching the fetal circulation.

Autoantibodies↗

PubMed1996

Adhesion of activated platelets to venous endothelial cells is mediated via GPIIb/IIIa.

Normal circulating platelets do not adhere to intact, undisturbed endothelium. Studies have shown, however, that platelets will adhere to virally infected or thrombin-stimulated human umbilical vein endothelial cells. Using a novel platelet/endothelial cell adhesion assay we studied the interaction of thrombin-activated platelets to human saphenous vein endothelial cells (HSVEC), and its mechanism(s). Biotinylated platelets were exposed to Hepes-Tyrode buffer, 10E5 or PAC-1 [monoclonal antibodies (Mabs) blocking GPIIb-IIIa], AK4 (Mab blocking P-selectin, 6D1 (Mab blocking vWf binding to GPIb), RGDS (small peptide blocking the fibrinogen binding site), or EDTA (dissociates GPIIb-IIIa complex) and then activated with thrombin. The platelets were subsequently exposed to thrombin-stimulated monolayer HSVEC. Phycoerythrin-streptavidin was added to the wells to fluorescently label the platelets, followed by formaldehyde fixation and washing to remove nonadherent platelets. Adhesion of platelets to HSVEC was assessed using a fluorescent multiwell plate reader. Antibodies which blocked the GPIIb-IIIa receptor and agents which competitively bound the receptor all significantly inhibited activated platelet adhesion to the activated HSVEC. We have found that thrombin significantly increases platelet/HSVEC adhesion, and this event is mediated via the integrin GPIIb-IIIa (fibrinogen receptor). These GPIIb-IIIa receptor blocking Mabs and RGDS may be useful adjuncts for improving patency following angiographic intervention and/or vein grafting in patients with high risk of thrombosis. The assay we have developed is a valuable and relatively simple method for assessing platelet/endothelial cell adhesion and activation.

Amino Acid Sequence↗

PubMed1996

Reduction of bleeding after heart operations through the prophylactic use of epsilon-aminocaproic acid.

UNLABELLED: Excessive postoperative bleeding after heart operations continues to be a source of morbidity. This prospective double-blind study evaluated epsilon-aminocaproic acid as an agent to reduce postoperative bleeding and investigated its mode of action. One hundred three patients were randomly assigned to receive either 30 gm epsilon-aminocaproic acid (51 patients) or an equivalent volume of placebo (52 patients). In a subset of these patients (14 epsilon-aminocaproic acid, 12 placebo), tests of platelet function and fibrinolysis were performed. RESULTS: By multivariate analysis, three factors were associated with decreased blood loss in the first 24 hours after operation: epsilon-aminocaproic acid versus placebo (647 ml versus 839 ml, p = 0.004), surgeon 1 versus all other surgeons (582 ml versus 978 ml, p = 0.002), and no intraaortic balloon versus intraaortic balloon pump use (664 ml versus 1410 ml, p = 0.02). No significant differences in platelet function could be demonstrated between the two groups. Inhibited fibrinolysis, as reflected by less depression of the euglobulin clot lysis and no rise in D-dimer levels, was significant in the epsilon-aminocaproic acid group compared with the placebo group. CONCLUSION: The intraoperative use of epsilon-aminocaproic acid reduces postoperative cardiac surgical bleeding.

Aminocaproic Acid↗

PubMed1996

Common factors direct transcription through the proximal sequence elements (PSEs) of the embryonic sea urchin U1, U2, and U6 genes despite minimal similarity among the PSEs.

The proximal sequence element (PSE) for the sea urchin U6 small nuclear RNA gene has been defined. The most critical nucleotides for expression, located 61 to 64 nucleotides (nt) from the transcription start site, are 4 nt, AACT, at the 5' end of the PSE. Two nucleotide mutations in this region abolish transcription of the sea urchin U6 gene in vitro. The same two nucleotide mutations greatly reduce the binding of specific factors detected by an electrophoretic mobility shift assay. There is also a conserved AC dinucleotide 57 nt from the start site of the sea urchin U1 and U2 PSEs. The sea urchin U1 and U2 PSEs were substituted for the sea urchin U6 PSE, with the conserved AC sequences aligned with those of the U6 PSE. Both of these genes were expressed at levels higher than those observed with the wild-type U6 gene. Similar complexes are formed on the U1 and U2 PSEs, and formation of the complexes is inhibited efficiently by the U6 PSE. In addition, the E-box sequence present upstream of the PSE enhances U6 transcription from both the U1 and U2 PSEs. Finally, depletion of a nuclear extract with a DNA affinity column containing the U6 PSE sequence reduces expression of the U6 genes driven by the U6, U1, or U2 PSE but does not affect expression of the 5S rRNA gene. These data support the possibility that the same factor(s) interacts with the PSE sequences of the U1, U2, and U6 small nuclear RNA genes expressed in early sea urchin embryogenesis.

Animals↗

PubMed1995

Transforming growth factor beta activates the promoter of cyclin-dependent kinase inhibitor p15INK4B through an Sp1 consensus site.

Transforming growth factor beta (TGF-beta) causes growth arrest in the G1 phase in many cell types. One probable pathway for this growth inhibition is through the TGF-beta-mediated up-regulation of the cyclin-dependent kinase (CDK) inhibitor p15INK4B, which specifically inhibits the enzymatic activities of CDK4 and CDK6. An active cyclin D-CDK4/6 complex is required for pRb phosphorylation to allow the cell cycle to progress from G1 to S phase. To study the molecular mechanism of the p15INK4B induction by TGF-beta, we isolated a 780-base pair promoter sequence of the human p15 gene and inserted this fragment upstream of a luciferase reporter gene. When this construct was transiently transfected into HaCaT cells, luciferase activity was induced more than 10-fold upon TGF-beta treatment, indicating that the induction of p15INK4B expression by TGF-beta is partly exerted at the transcription level. Promoter deletion analysis revealed that the sequence from -110 to -40 relative to the transcription start site is capable of conferring the 10-fold induction by TGF-beta. Within this region there are three Sp1 consensus sites. Mutation of one of these sites, GGGGCGGAG, substantially reduced both the induction by TGF-beta and the basal promoter activity, whereas mutations in the other two Sp1 sites and the spacer sequences had little effect. In addition, gel mobility shift assay indicates that the transcription factors Sp1 and Sp3 bind to this Sp1 site. Taken together, these data suggest that a specific Sp1 consensus site is involved in the mediation of TGF-beta induction as well as the basal promoter activity of the p15 gene and that Sp1 and Sp3 transcription factors might be involved in this regulation.

Base Sequence↗

PubMed1995

Thrombocytopoietic effect of heparin given in chronic immune thrombocytopenic purpura.

Our previous in-vitro and in-vivo studies showed that heparin enhanced murine and human megakaryocytopoiesis. 20 patients with chronic immune thrombocytopenic purpura were randomly divided into two groups and given 10 mg per day of prednisone for 30 days, for haemostatic purposes. One group received in addition heparin (1250 IU twice a day subcutaneously for 30 days). From day 10, a significant increase in platelet count was observed in eight of the ten patients treated with heparin (p < 0.05), with return to the initial value after heparin cessation in six of the responders. These data demonstrate the effectiveness of heparin and suggest its use or that of other related compounds for therapy of chronic immune thrombocytopenic purpura.

Adult↗

PubMed1995

Anti-La (SS-B) but not anti-Ro52 (SS-A) antibodies cross-react with laminin--a role in the pathogenesis of congenital heart block?

Cross-reactions between maternally derived autoantibodies and fetal cardiac antigens have been postulated to play a role in the pathogenesis of congenital heart block (CHB). We have explored the cross-reactivity of autoantibodies to the small ribonuclear autoantigens, La/SS-B and Ro/SS-A, with laminin, the major component of cardiac sarcolemmal membrane using affinity-purified antibodies from patients with Sjögren's syndrome (SS). Anti-La antibodies purified from eight of 10 patients cross-reacted significantly with mouse laminin by ELISA. In contrast, purified antibodies to Ro52 from the same 10 patients showed little or no binding to laminin. Laminin inhibited up to 70% binding of anti-La antibodies to La antigen, and La inhibited up to 65% binding of anti-La antibodies to laminin. The cross-reaction was further examined on cryosections of 10 human fetal hearts aged from 8.7 to 14.9 weeks of gestation, two normal adult hearts, and one pathological adult heart with a diagnosis of dilated cardiomyopathy. Anti-Ro52 antibodies did not bind to the surface of cardiac cells. However, anti-La antibodies from seven of 10 patients tested bound to the surface of fetal myocytes from hearts aged 9.4 to 14.9 weeks of gestation, and also to the myocytes from the pathological adult heart but not to normal adult hearts. Preincubation with La antigen abolished the binding of anti-La antibodies to the surface of adult heart myocytes with dilated cardiomyopathy, and pre-incubation with mouse laminin could partially block this binding. These results suggest that molecular mimicry between laminin and La, but not Ro52, may act as a target for specific maternal autoantibodies, and contribute to the pathogenesis of CHB at a critical stage during fetal cardiac development.

Adult↗

PubMed1995

Adenosine enhances nitric oxide production by vascular endothelial cells.

Adenosine per se is a potent vasodilator of vascular smooth muscle. Endothelial cells modulate vascular tone via the release of nitric oxide (NO), which also elicits vasodilation. This study was undertaken to determine whether adenosine could directly stimulate endothelial cells to enhance NO production, which could subsequently reduce vascular tone. NO production was evaluated in porcine carotid artery endothelial cells (PCAEC) and human saphenous vein endothelial cells (HSVEC) seeded on multiwell plates, grown to confluence, and treated with adenosine for 1 h. The bathing medium was collected, and the NO production was determined as reflected by the formation of NO2- and NO3-. NO production by PCAEC was significantly increased by adenosine in a dose-dependent manner, whereas there was only an insignificant tendency for an increase by HSVEC. The addition of the NO synthase competitive inhibitor, NG-monomethyl-L-arginine (NMMA), or the adenosine receptor antagonist, theophylline, prevented the increase in NO production by adenosine. The results suggest that adenosine stimulates, by a receptor-mediated mechanism, the production of NO by arterial, but not by venous, endothelial cells.

Adenosine↗

PubMed1995

Contact-dependent stimulation of monocytic cells and neutrophils by stimulated human T-cell clones.

By means of direct cell-cell contact, fixed, stimulated T lymphocytes trigger the production of interleukin-1 beta (IL-1 beta) and tumour necrosis factor by monocytes and prime polymorphonuclear leucocytes (PMN) for the respiratory burst induced by N-formyl-methionyl-leucyl-phenylalanine (FMLP). In order to assess whether this activity is displayed by a particular subpopulation of T lymphocytes, 88 T-cell clones (TCC) were generated from healthy human peripheral blood. The clones were stimulated by Phaseolus vulgaris agglutinin (PHA) and phorbol 12-myristate acetate monocytic cell line THP-1. All fixed, stimulated TCC induced IL-1 beta production by THP-1 cells, although to varied degrees. The activity of TCC on THP-1 cells correlated with their activity on PMN (r2 = 0.84, P < 0.001), suggesting that this pro-inflammatory activity of TCC may similarly affect both types of infiltrating cells. The ability of TCC to modulate target cells did not correlate with either their phenotype (CD4 or CD8) or their cytokine production profile (interferon-gamma, IL-4 and IL-6), although it tended to correlate inversely with their capacity to produce IL-6 (P < 0.02). These observations suggest that a large proportion, if not all, of human peripheral blood T lymphocytes may potentially affect monocytes and PMN by direct cell-cell contact. This activity may be relevant to the maintenance of chronic inflammation.

Antigens, CD↗

PubMed1995

Genes of intracellular calcium metabolism and blood pressure control in primary hypertension.

In the last two decades, major progress has been made in understanding the role of calcium (Ca) metabolism in blood pressure (BP) control. This article discusses the intracellular Ca handling systems that could potentially be involved in the pathogenesis of primary hypertension. We begin by reviewing our current knowledge of intracellular Ca handling, alterations of intracellular Ca metabolism in primary hypertension, and possible mechanisms of BP control. We have analyzed data on the structure and alternative splicing of major genes controlling intracellular free Ca concentration, ie, genes encoding intracellular Ca-binding protein (calmodulin, calbindin, plasma membrane Ca2+ pump, Na+/Ca2+ exchanger, and voltage-dependent Ca channels). Data are presented on the relationship of gene polymorphism and alternative splicing with membrane architecture and the function of gene products. Numerous observations on abnormalities of these gene products in primary hypertension are summarized. Studies on the polymorphism of genes regulating intracellular Ca in hypertension are only now being performed.

Animals↗

PubMed1995

Effects of Panax notoginseng saponin Rg1 on cardiac electrophysiological properties and ventricular fibrillation threshold in dogs.

AIM: To study the effects of Rg1 isolated from saponins of Panax notoginseng on cardiac electrophysiological properties and ventricular fibrillation threshold (VFT). METHODS: Seventeen open-chest dogs were randomly allocated into a Rg1 group (20 mg kg-1, iv) and a control group. The electrophysiological variables and VFT were evaluated by standard electric stimuli and monophasic action potential (MAP) recording. RESULTS: Rg1 prolonged sinus node recovery time (SNRT) by 19.1%, AV conduction Wenckebach cycle length (AVWCL) by 7.1%, and ventricular effective refractory period (VERP) by 7.9%. It prolonged ventricular MAPD30, MAPD50, and MAPD90 by 25.5%, 24.2%, and 13.5%, respectively. VFT was increased by 19.2%. CONCLUSION: Rg1 prolonged ventricular refractoriness and repolarization, and increased VFT. It was indicated that cardiac electrophysiological effects of Rg1 were similar to those of amiodarone.

Action Potentials↗

PubMed1994

The activation-induced decrease in the platelet surface expression of the glycoprotein Ib-IX complex is reversible.

Thrombin decreases the platelet surface expression of the glycoprotein (GP) Ib-IX complex. To determine whether this effect is reversible, flow cytometric studies were performed with GPIb-IX-specific monoclonal antibodies. In both whole blood and washed platelet systems, incubation of platelets with thrombin or a combination of adenosine diphosphate and epinephrine resulted in a maximal decrease of the platelet surface expression of GPIb-IX within 5 minutes, after which there was a time-dependent return of the platelet surface GPIb-IX complex, which was maximal by 60 minutes. Exposure of the same platelets to additional exogenous thrombin resulted in a second decrease in platelet surface GPIb-IX, followed by a second reconstitution of platelet surface GPIb-IX. Throughout these experiments there was no measurable release from the platelets of glycocalicin (a proteolytic fragment of GPIb). Experiments in which platelets were preincubated with a biotinylated GPIb-specific MoAb showed that the GPIb molecules that returned to the platelet surface were the same molecules that had been translocated to the intraplatelet pool. The GPIb molecules that returned to the platelet surface were functionally competent to bind von Willebrand factor, as determined by ristocetin-induced platelet agglutination and ristocetin-induced binding of exogenous von Willebrand factor. Inhibitors of protein kinase C and myosin light-chain kinase enhanced the reexpression of platelet surface GPIb. In summary, the activation-induced decrease in the platelet surface expression of the GPIb-IX complex is reversible. Inactivation of protein kinase C and myosin light-chain kinase are important mechanisms in the reexpression of the platelet surface GPIb-IX complex.

Adult↗

PubMed1994

Transcription of the sea urchin U6 gene in vitro requires a TATA-like box, a proximal sequence element, and sea urchin USF, which binds an essential E box.

The tandemly repeated gene set encoding the sea urchin U6 gene has been cloned from the sea urchin Strongylocentrotus purpuratus. The U6 gene is transcribed by RNA polymerase III in a sea urchin nuclear extract. Like that of the vertebrate U6 genes, transcription of the sea urchin U6 gene does not require any internal sequences or 3' sequences but requires only 5' flanking sequences. Only 88 nucleotides of 5' flanking sequence are required for maximal expression in vitro. Mutagenesis experiments demonstrated the requirement for three elements, a CACGTG element at -80, a proximal sequence element at about -55, and the TATA-like box at -25. The major protein in sea urchin extracts that interacts with the CACGTG element is sea urchin USF, and immunodepletion of sea urchin USF greatly reduces transcription. The USF binding site in the U6 gene is highly homologous (11 of 13 nucleotides) with the USF binding sites found in the promoter of the S. purpuratus spec genes.

Animals↗

PubMed1994

Effects of 3'-angeloyloxy-4'-acetoxy-3',4'-dihydroseselin on cardiohemodynamics in anesthetized dogs.

Cardiohemodynamic effects of 3'-angeloyloxy-4'-acetoxy-3',4'-dihydroseselin (Pd-Ia) isolated from the Chinese medicinal plant Peucedanum praeruptorum Dunn were compared with those of diltiazem (Dil) in anesthetized open-chest dogs. Pd-Ia 3 mg.kg-1 increased coronary blood flow from 25 +/- 11 to 58 +/- 16 ml.min-1 (n = 7, P < 0.01) and decreased mean aortic pressure (MAP) from 13 +/- 2 to 8 +/- 1 kPa (P < 0.01), rate pressure product (RPP) from 1.9 +/- 0.5 to 1.3 +/- 0.3 MPa.bpm (P < 0.05), +dp/dtmax from 246 +/- 56 to 160 +/- 36 kPa.s-1 (P < 0.01) and systemic vascular resistance from 19 +/- 4 to 12 +/- 7 Pa.ml-1.min-1 (P < 0.05), together with an increase in HR. Dil showed effects similar to those of Pd-Ia except for a marked decrease in HR. The effects of Pd-Ia on MAP and RPP were approximately one-tenth as potent as those of Dil. The results demonstrated that Pd-Ia was a Ca2+ channel blocker.

Animals↗

PubMed1994

Effect of dl-praeruptorin A on calcium current in ventricular cells of guinea pig.

With patch clamp technic (whole cell recording), the effect of dl-praeruptorin A (Pra), an ingredient of Peucedanum praeruptorum Dunn on calcium current (ICa) in the single ventricular cells of guinea pig was studied. Results showed that under Cs/Cs condition, when the holding potential was -40 mV and in the presence of Pra (1, 10, 100 mumol.L-1), ICa was decreased dose-dependently from 2.02 +/- 0.24, 2.00 +/- 0.12, 2.12 +/- 0.33 nA (control) to 1.60 +/- 0.24, 1.32 +/- 0.08, 1.16 +/- 0.43 nA, respectively, and their inhibitory rates were 21%, 33.5%, 45%, respectively. The current-voltage relation curve showed that the reversal potential of ICa was +60 mV; the potential producing peak value of ICa was about 0 mV. The results indicated that Pra had a Ca2+ channel blocking effect.

Animals↗

PubMed1994

Effects of cyproheptadine on plasma superoxide dismutase activity and malondialdehyde content in rabbits with hemorrhagic shock.

Profound hemorrhagic shock was produced in thirty rabbits by exsanguination via the carotid artery until blood pressure (BP) reached 5.3 kPa (40 mmHg) and was sustained for a period of 90 minutes. The rabbits were equally divided into cyproheptadine (Cyp) treated group and control group. Blood samples 30 minutes after liquid and blood infusion and administration of Cyp (10 mg/kg) were collected from the carotid artery, and the plasma superoxide dismutase (SOD) activity and malondialdehyde (MDA) content measured. The results showed that Cyp remarkably enhanced the plasma SOD activity (2462 +/- 338 vs 1955 +/- 596, P < 0.01) and reduced MDA content (2.68 +/- 0.24 vs 3.20 +/- 0.49, P < 0.01). We believe that the increase of O2 production plays an important role in the development of shock, the single blood and liquid infusion can not significantly improve the shock conditions. Scavenging oxygen free radicals and alleviating cellular damage and multiple organ failure are the possible mechanisms of cyproheptadine anti-shock effect.

Animals↗