Biomedical subjects

J J Liu

Publications and source records attributed to J J Liu.

At least 73 records · Page 4Linked to original sources

PubMed1997

A novel method for quantitative analysis of apoptosis.

In the present study, we describe a method of quantifying DNA fragmentation. This assay is based on saturation labeling 3'-ends of DNA fragments with alpha(32)PdCTP in the presence of 2',3'-dideoxy-cytidine-5'-triphosphate (ddCTP) by terminal deoxynucleotidyl transferase (TdT). The saturation labeling of 3'-ends of DNA fragments was performed by adding different concentrations of alpha(32)PdCTP to a DNA sample, from which a maximal labeling (Lmax) and a kinetic parameter (Km) of the TdT reaction are calculated. The saturated labeling gives true quantitation that makes it possible to accurately compare quantities of DNA fragments among different samples. This method requires as little as 5 ng of DNA and increases the sensitivity of apoptotic DNA detection by at least 200-fold relative to the widely used ethidium staining method. The application of this method in an apoptosis study showed that (a) a time- and dose-dependent increase in the number of DNA strand breaks in apoptotic lymphocytes was induced by dexamethasone, and (b) age-dependent apoptosis occurred in the cardiac tissues of spontaneously hypertensive rats. Results of this assay were confirmed by the DNA ladder pattern exhibited after electrophoresis as well as the morphologic changes of apoptosis observed under electron microscopy and were very consistent with results obtained in quantifying apoptotic cells by flow cytometric analysis (r = 0.98, p = 0.002). Thus, this assay is quantitative, simple, sensitive, and useful for assessing apoptosis.

Animals↗

PubMed1996

Identification, molecular cloning, and transcription analysis of the Choristoneura fumiferana nuclear polyhedrosis virus spindle-like protein gene.

The Choristoneura fumiferana nuclear polyhedrosis virus spindle-like protein (slp) gene has been identified and localized immediately downstream and in the same orientation as the CfMNPV DNA polymerase gene. The slp gene is 1101 bp long, predicted to code for a 366 amino acid (42.1 kDa) polypeptide. Transcriptional analysis revealed that the CfMNPV slp gene is expressed at late times postinfection, beginning at 24 hr postinfection and is most abundantly expressed after 36 hr. Transcription initiates within a single baculovirus consensus late start site sequence (GTAAG) at position -18 relative to the translation start codon. Based on amino acid comparisons, the CfMNPV gene is closely related to other similar baculovirus genes and distantly but recognizably related to the fusolin proteins of two entomopoxviruses. The conservation of amino acid sequence, glycosylation signals and specific domains throughout the protein suggest that this gene product may play an important role in insect DNA virus replication.

Amino Acid Sequence↗

PubMed1996

Support for the prion hypothesis for inheritance of a phenotypic trait in yeast.

A cytoplasmically inherited genetic element in yeast, [PSI+], was confirmed to be a prionlike aggregate of the cellular protein Sup35 by differential centrifugation analysis and microscopic localization of a Sup35-green fluorescent protein fusion. Aggregation depended on the intracellular concentration and functional state of the chaperone protein Hsp104 in the same manner as did [PSI+] inheritance. The amino-terminal and carboxy-terminal domains of Sup35 contributed to the unusual behavior of [PSI+]. [PSI+] altered the conformational state of newly synthesized prion proteins, inducing them to aggregate as well, thus fulfilling a major tenet of the prion hypothesis.

Amino Acid Sequence↗

PubMed1996

Studies of Choristoneura fumiferana nuclear polyhedrosis virus gene expression in insect cells.

To investigate the mechanisms regulating baculovirus virulence and host range we have begun to study Choristoneura fumiferana nuclear polyhedrosis virus (CfMNPV) and its gene expression in permissive and nonpermissive cells. We have identified and mapped three genes on the CfMNPV genome. The polyhedrin gene is located from 0.0 to 2.0 m.u. and two other genes, dnapol and p143, both of which are essential for baculovirus DNA replication, are located from 35.3 to 40.9 m.u. and 55.5 to 63.4 m.u., respectively. To gain insight into the expression of CfMNPV genes in permissive C. fumiferana and nonpermissive Spodoptera frugiperda cells, we constructed three expression plasmids in which the promoter region of the dnapol, the p143, and polyhedrin genes were placed in front of a chloramphenicol acetyltransferase reporter gene. All three CfMNPV promoters were active in nonpermissive cells in the presence of Autographa californica nuclear polyhedrosis virus (AcMNPV) DNA, but no activity was detected in permissive cells either in the presence of CfMNPV DNA or AcMNPV DNA. This lack of promoter activity was not due to failure of viral or plasmid DNA to enter the cell nucleus. It was possible that the reporter plasmids were inefficient templates for transcriptional transactivation so we developed a CfMNPV transfer vector and generated a recombinant virus in which the polyhedrin promoter driving CAT gene cassette was integrated into the CfMNPV genome. In this case, the CfMNPV polyhedrin promoter was highly active in the permissive cells.

Animals↗

PubMed1996

Enhanced denaturation of the alpha (II) chains of type-II collagen in normal adult human intervertebral discs compared with femoral articular cartilage.

The mechanical strength of connective tissues is dependent on the integrity of their fibrillar collagen frameworks. The objective of the present study was to assess type-II collagen damage (denaturation) in the adult human intervertebral disc compared with articular cartilage, in order to determine whether damage to this molecule may vary in different anatomical sites in the same person. A new immunochemical assay was used to measure the amounts of denatured and total type-II collagen in the annulus fibrosus and nucleus pulposus of the L5-S1 disc and in cartilage from the femoral condyles of the same individuals (n = 7). Denaturation of type-II collagen was significantly higher in both the annulus fibrosus and the nucleus pulposus than in articular cartilage. Such increased damage to type-II collagen in the adult disc may have relevance to the more pronounced degenerative changes observed in this tissue compared with articular cartilage.

Adult↗

PubMed1996

Individual serum bile acids in apprentice spray painters in association with solvent exposure.

The effects of exposure to solvents on serum bile acids were investigated by comparing a group of apprentice vehicle spray painters (exposed group) with one of apprentice electricians. Apprentice spray painters from the study were subdivided into high- and low-solvent-exposure groups. Concentrations of individual serum bile acids (SBA) were measured and compared with conventional liver function tests (LFTs). Total, free, glycine- and taurine-conjugated SBA were consistently found to be present at higher levels in the spray painters than in the electricians, even at the beginning of the apprenticeship. Total SBA tended to increase in spray painters with increasing years of exposure during the apprenticeship, but this was significant at only one time point. No rises were observed over the sampling period in electricians. The mean values of individual and total SBA concentrations were all found to be higher in the high-exposure group than in the low-exposure group, with some differences reaching statistical significance. None of the routine liver biochemistry parameters was different between spray painters and electricians. gamma-Glutamyl transferase (GGT) was the only enzyme found to be significantly different between the high- and low-exposure groups, but all values were within the normal range. This study suggests that occupational exposure even to low levels of solvent mixtures results in increases in SBA. The increased SBA may be indicative of a subclinical liver dysfunction. Alternatively, they may reflect solvent exposure only, with the raised levels having no pathologic implication or consequence.

Adolescent↗

PubMed1996

Unique response of human arteries to endothelin B receptor agonist and antagonist.

1. The role of the ET(B) receptor in human arteries has not been well studied because of the lack of specific ET(B) receptor antagonists. In the present studies the specific ET(B) receptor antagonist BQ-788 and the specific ET(B) agonist IRL-1620 were used to characterize the function of the ET(B) receptor in human radial arteries and internal mammary arteries. 2. The results showed that the ET(B) antagonist BQ-788 significantly inhibited endothelin-1-induced contraction in internal mammary arteries, but not in radial arteries. In internal mammary arteries, BQ-788 at a concentration of 10 micromol/l shifted the endothelin-1-induced concentration-dependent curve to the right by one order. By comparison, the ET(A) receptor antagonist BQ-610 at 1 micromol/l produced a much more potent inhibitory effect (three-order shifting) on endothelin-1-induced contraction in internal mammary arteries, and also potently inhibited the contraction in radial arteries. 3. The ET(B) agonist IRL-1620 caused a contraction in internal mammary arteries, but not in radial arteries, although the response of radial arteries to endothelin-1 was very strong. The contraction induced by IRL-1620 was weaker than that induced by endothelin-1; however, the maximal contraction to IRL-1620 was obtained at 3 nmol/l, which was lower than that with endothelin-1 (maximal contraction at 10 nmol/l). 4. In internal mammary arteries the contraction to endothelin-1 and IRL-1620 gradually changed to relaxation with high concentrations of endothelin-1 (from 30 nmol/l) and IRL-1620 (from 3 nmol/l), whereas it did not in radial arteries; suggesting that the ETB receptor on human arterial smooth muscle cells may mediate contraction at low agonist concentrations and relaxation at high agonist concentrations. 5. The ET(B) agonist IRL-1620, endothelin-1 and endothelin-3 did not cause endothelium-dependent relaxation in either precontracted radial arteries or internal mammary arteries, although endothelium-dependent relaxation was fully induced by acetylcholine in these two arterial preparations. 6. In conclusion, the present studies demonstrate that the responses of internal mammary arteries and radial arteries to an ET(B) antagonist and an ET(B) agonist are significantly different from those of animal vascular vessels, and also from each other. The ET(B) receptor may play only a minor role in endothelium-dependent relaxation of these human arteries. Endothelin-1-induced contraction is mediated by both the ET(A) (major) and the ET(B) (minor) receptors in internal mammary arteries, but only by the ET(A) receptor in radial arteries. These studies may help to determine therapeutic strategy.

Aged↗

PubMed1996

The radial artery as a bypass graft.

The radial artery is being reintroduced into clinical practice to increase the number of arterial grafts for patients undergoing coronary artery bypass surgery. The radial artery is readily available from one or both forearms and removal is safe in patients who have a normal collateral circulation to the hand. Harvesting the radial artery with the adjacent veins using a minimal touch technique and vasodilators will prevent vasospasm and possibly early occlusion. A concern is that subclinical atheroma is present in many patients. The radial artery can be used as a single graft, anastomosed in a "Y" fashion with the internal mammary artery, or used as a sequential graft. The early mortality and complications are low. There are potential cost savings because the need for an incision in the leg is avoided, so that patients may be discharged early. The early results of radial artery grafting are encouraging. Further follow-up is required to determine the late patency and effects on survival of using the radial artery graft.

Aged↗

PubMed1995

Identification, localization, transcription, and sequence analysis of the Choristoneura fumiferana nuclear polyhedrosis virus DNA polymerase gene.

The location of the Choristoneura fumiferana baculovirus DNA polymerase gene was determined by hybridization analysis using a probe prepared from the previously identified polymerase gene from the Autographa californica multiple nuclear polyhedrosis virus. DNA sequence analysis revealed that the Choristoneura fumiferana baculovirus DNA polymerase gene consists of 2970 base pairs encoding 990 amino acids (114.2 kDa). Transcriptional analysis demonstrated that overlapping transcripts of 3.2 and 4.6 kb, first detected at 6 hr postinfection, potentially coded for the DNA polymerase gene. The major transcription starts sites, identified at 6 hr postinfection, mapped to baculovirus consensus early start sites CGTGCTCA and CAGT. The relatively low level and late initiation of the DNA polymerase gene coupled with our previous data on the temporal control of DNA replication and late gene synthesis (Liu and Carstens, 1993) suggests that the low virulence of the spruce budworm baculovirus may be related to the regulation of its gene expression at the transcriptional level.

Amino Acid Sequence↗

PubMed1995

Induction of Ca2+ signaling and possible exocytosis in endothelial cells by a stable leukocyte-derived factor.

Previous studies have shown that polymorphonuclear leukocytes (PMNs) release a stable factor that inhibits endothelium-dependent relaxation. In the present studies, the effects of the factor on Ca2+ signaling and on ultrastructure of endothelial cells were investigated. In the cultured endothelial cells, the PMN-derived factor induced an increase in [Ca2+]i in a pattern of oscillations. The frequency of the Ca2+ oscillations was less than 3 spikes/10 minutes. Removal of extracellular Ca2+ by perfusion with Ca(2+)-free Krebs' solution abolished the spikes. The results of electron microscopy showed that this factor induced an increase in vesicle formation on the luminal surface of the rat aortic endothelium. The increased vesicle formation may represent exocytosis. The structure of the smooth muscle cells was not changed. In conclusion, the PMN-derived factor induces a Ca2+ influx and In conclusion, the PMN-derived factor induces a Ca2+ influx and possible exocytosis, suggesting that the factor may have other biological functions besides the inhibition of the vascular relaxation.

Animals↗

PubMed1995

Potent inhibitory effect of SR 49059, an orally active non-peptide vasopressin VIa receptor antagonist, on human arterial coronary bypass graft.

1. The effect of vasopressin receptor antagonists varies between analogues (peptide, non-peptide) and across species. In this study the effect of the novel non-peptide vasopressin V1a receptor antagonist SR 49059 on human internal mammary arteries was investigated. 2. SR 49059 produced a potent, concentration-dependent, inhibitory effect on vasopressin-induced contraction of human coronary bypass graft internal mammary arteries. Both SR 49059 (1 mumol/l) and a peptide selective V1a antagonist ([d(CH2)5sarcosine7]arginine vasopressin) (1 mumol/l) abolished vasopressin-induced contraction. The non-peptide V1a receptor antagonist OPC-21268 (1 mumol/l) had no effect on vasopressin-induced contraction. 3. The effect of SR 49059 was specific to vascular vasopressin receptors as noradrenaline-induced contraction was not influenced by SR 49059. 4. The results of this study in vitro indicate that the non-peptide SR 49059 is a potent, specific vasopressin V1a receptor antagonist in the human internal mammary artery and suggest that it may be a useful tool for studying the pathophysiological role of vasopressin in man.

Antidiuretic Hormone Receptor Antagonists↗

PubMed1995

Galactinol synthase from kidney bean cotyledon and zucchini leaf. Purification and N-terminal sequences.

Galactinol synthase (GS) was purified 1591-fold with a 3.9% recovery from the cotyledon of kidney bean (Phaseolus vulgaris) by a novel scheme consisting of ammonium sulfate fractionation followed by diethylaminoethyl, Affi-Gel Blue, and UDP-hexanolamine affinity chromatography. The purified enzyme had a specific activity of 8.75 mumol mg-1 min-1, a pH optimum of 7.0, and requirements for manganese ion and DTT. The enzyme exhibited a Km = 0.4 mM for UDP-galactose and a Km = 4.5 mM for myo-inositol. It was identified as a 38-kD peptide that co-purified with a 41- and a 43-kD peptide as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Purification to homogeneity was achieved by isolating the 38-kD peptide from the SDS-PAGE gel. To clarify conflicting reports in the literature about the relative molecular mass of purified GS from zucchini leaf (Cucurbita pepo), a similar scheme with modified eluting conditions was used to purify GS from this source. Zucchini leaf GS was purified to homogeneity and identified as a 36-kD peptide on SDS-PAGE. Partial N-terminal sequences of the 38-kD peptide from kidney bean cotyledon and the 36-kD peptide from zucchini leaf were obtained. To facilitate identification of GS during the purification, an assay utilizing thin-layer chromatography and an isotopic analytic imaging scanner was developed.

Amino Acid Sequence↗

PubMed1995

Ras transformation results in an elevated level of cyclin D1 and acceleration of G1 progression in NIH 3T3 cells.

Ectopic overexpression of v-H-Ras protein in NIH 3T3 cells resulted in cellular transformation and an acceleration of G1 progression of these cells. A shortened G1 phase was found to be associated with an increased level of cyclin D1 but not cyclin E protein. Using an antisense blocking method, reduced synthesis of cyclin D1 in v-H-Ras transformants resulted in a slower G1 progression rate of these cells. Although constitutive overexpression of cyclin D1 in NIH 3T3 cells accelerated G1 progression, cells remained untransformed. Furthermore, inhibition of cyclin D1 synthesis greatly impaired the soft-agar cloning efficiency of v-H-Ras transformants. These results suggest that increased expression of cyclin D1 is necessary but not sufficient for the transforming activity of v-H-Ras. Similar effect on cell cycle progression was also observed in Raf-transformed cells. In addition to cyclin D1, cyclin E protein was found to be elevated in Src transformants. This may account for the further shortening of the G1 phase of these cells. Activation of an additional Ras-independent pathway was suggested to be responsible for the further acceleration of the G1 phase in Src transformants.

3T3 Cells↗

PubMed1995

Various cells release a stable small molecule that inhibits endothelium-dependent relaxation.

Previous studies have shown that neutrophils release a stable factor that inhibits endothelium-dependent relaxation. In the present studies, the effects of supernatants derived from various cells on endothelium-dependent relaxation were studied. Cells were obtained from seven sources: human hematopoietic cells including mononuclear leukocytes (MONO), polymorphonuclear leukocytes (PMNs), and chronic lymphocytic leukemia (CLL) cells; cells of the cardiovascular system including human endothelial cell line ECV304, human smooth muscle cells, and rat myocardial cells; and the tumor cell line HPB. These isolated or cultured cells were incubated for 1 h in Krebs solution to release the factor. The results showed that the supernatants from 10(5) cells/ml of all cells except the tumor cell line HPB produced a potent inhibitory effect on endothelium-dependent relaxation of rat aortic rings in response to acetylcholine and Ca2+ ionophores A23187 and ionomycin but not on endothelium-independent relaxation to nitroprusside and glyceryl trinitrate. When the concentration increased to 10(6) cell/ml, the supernatants from the tumor cell line HPB also slightly but significantly inhibited endothelium-dependent relaxation. The potency order was PMNs = MONO = CLL cells > cardiac cells > smooth muscle cells > the endothelial cell line ECV304 > the tumor cell line HPB. It seems that the hematopoietic cells and the cardiac cells are more active in release of the factor. The effect of this factor was rapid in onset and hard to wash out. A cyclooxygenase inhibitor or a thromboxane A2-prostaglandin H2 receptor antagonist partially but significantly reduced the effect of the factor.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Role of thrombosis in the pathogenesis of congestive hepatic fibrosis (cardiac cirrhosis).

The pathogenesis of congestive cirrhosis is generally thought to be a reaction of the hepatic stroma to hypoxia, pressure, or necrosis. This does not explain the poor correlation between symptoms and severity of fibrosis and the irregular distribution of fibrosis within the liver. We have observed healed hepatic vein (HV) thrombosis in patients with congestive heart failure (CHF). The purposes of this study were to document hepatic vascular lesions in autopsy livers of patients with chronic CHF, to determine the role of these lesions in the pathogenesis of congestive cirrhosis, and to refine the definition of congestive cirrhosis. Twenty-five livers were studied, 13 with multiple large blocks 4 x 5 cm. Sections were graded for parenchymal fibrous septa, sinusoidal fibrosis, and intimal fibrosis of portal veins (PVs) and HVs. Fibrous septa were found in livers of 7 of 13 patients with CHF and in none of 12 controls without CHF (P = .007). Parenchymal fibrosis was highly variable in distribution, often with severe septation in some areas and nearly normal morphology in others. Intimal fibrosis and obstruction of small- and medium-HVs were found only in livers of patients with CHF. The vascular lesions were confined to regions with fibrous septation and had morphology suggestive of organized thrombosis. Acute thrombi in sinusoids were noted in livers of 4 patients with CHF and in livers of 2 patients without CHF. These findings support the hypothesis that congestive cirrhosis is a response to intrahepatic thrombosis. The pattern of disease suggests that thrombus begins in sinusoids, occasionally propagates to HVs, and causes secondary local PV thrombosis, ischemia, parenchymal extinction, and fibrosis.

Aged↗

PubMed1995

A method of stimulating an immune response with a hapten immobilized on a nitrocellulose membrane.

Hapten refers to a chemical compound of small molar mass (typically less than 1000 daltons) that can bind with an antibody, but cannot initiate an immune response by itself unless it is conjugated to a protein carrier of larger molar mass. A novel method to prepare a hapten to generate anti-hapten immunity without covalent conjugation to a carrier was developed. Coating both water-soluble and -insoluble haptens onto a nitrocellulose membrane effectively presented haptens to the system and caused the generation of specific anti-hapten B lymphocytes and antibodies by immunization both in vitro and in vivo. This method has a potential to substitute for conventional hapten carrier conjugation to generate anti-hapten immunity.

Adsorption↗

PubMed1994

A stable leukocyte-derived factor inhibits platelet aggregation.

The effect of human polymorphonuclear leukocyte (PMN)-derived supernatants on platelet aggregation was studied. The results showed that the supernatants produced a concentration-dependent inhibition of the platelet aggregation in response to thrombin. The effect was mainly on the second phase of the aggregation, suggesting that the secretory mechanisms of the platelets are disturbed. The activity of the supernatants was not reduced after treatment with heat (95 degrees C for 30 minutes), extreme pH (pH 12 or pH 2) or protease. In conclusion, human PMNs release a stable non-protein factor that inhibits platelet aggregation.

Biological Factors↗

PubMed1994

Purification and properties of GTP:GTP guanylyltransferase from encysted embryos of the brine shrimp Artemia.

GTP:GTP guanylyltransferase has been purified from the yolk platelets of Artemia cysts. The 480-kDa enzyme catalyzes the reversible reaction 2GTP<==>Gp4G + PPi and contains immunologically related polypeptides of 142, 88, and 45 kDa and a distinct 80-kDa component. The 88 and 45 kDa species can be covalently labeled with [alpha-32P]GTP. Even in crude extracts, the enzyme appears to be partially proteolyzed, suggesting that it is a nonfunctional residue of the pre-encystment stages of development. A native alpha 2 beta 2 structure comprising 2 mol each of the 142- and 80-kDa polypeptides is proposed. The reaction follows ping-pong kinetics with a covalent enzyme-guanylate intermediate containing a phosphoramidate linkage, probably phospholysine. The enzyme has two GTP-binding sites: a "donor" site in which the enzyme-guanylate is formed and which is highly specific for guanine nucleotides (GTP, p4G, dGTP, and GppNHp) and an "acceptor" site which additionally binds XTP, ITP, GDP, and ADP. Thus, the enzyme will form the homodinucleotides Gp4G, Gp5G, Gp3G, dGp4dG, and GppNHppG and the heterodinucleotides Gp4X, Gp4I, and Gp3A, but not Xp4X, Ip4I, or Gp4A. The Km for GTP was 6.7 mM and kcat was 1.6 s-1. XTP was a fully uncompetitive inhibitor of Gp4G synthesis while ITP was a partially uncompetitive inhibitor. In the reverse reaction, certain pyrophosphate analogs could substitute for PPi. The structure and mechanism of this enzyme suggest an evolutionary relationship to mRNA capping enzymes.

Animals↗