Biomedical subjects

F Chen

Publications and source records attributed to F Chen.

At least 559 records · Page 31Linked to original sources

PubMed1993

Clonability and tumorigenicity of human epithelial cells expressing the EBV encoded membrane protein LMP1.

Two isolates of the EBV-LMP1 gene were compared for their ability to induce phenotypic changes in a non-tumorigenic human keratinocyte line, Rhek-1, immortalized with an adenovirus 12-SV40 hybrid virus. One isolate, designated B-LMP1, was derived from B95-8, a B-cell line of marmoset origin, that carries a viral strain from a mononucleosis patient. The other, designated C-LMP1, originated from a nude mouse passaged Chinese NPC tumor, CAO. Both types of transfectants were less serum dependent than the non-transfected and the vector-transfected controls. The ability to grow on low serum increased with increasing LMP1 expression. All transfectants were more highly clonable than the non-transfected or vector-transfected controls. Clonability in soft agarose increased with increasing LMP1 expression. Nine of 24 C-LMP1 transfectants produced tumors in SCID mice. Seven of them grew invasively into the surrounding tissue. Only one of 12 B-LMP1 transfected Rhek-1 clones was tumorigenic. It did not grow invasively. All tumorigenic transfectants expressed LMP1 at high or moderate levels. All tumors were found to express LMP1. Transfectants with low LMP1 expression did not produce tumors. The untransfected Rhek-1 cells and six vector control clones failed to produce tumors.

Adenoviridae↗

PubMed1992

Two human liver cDNAs encode UDP-glucuronosyltransferases with 2 log differences in activity toward parallel substrates including hyodeoxycholic acid and certain estrogen derivatives.

Two human liver UDP-glucuronosyltransferase cDNA clones, HLUG25 [Jackson, M. R., et al. (1987) Biochem. J. 242, 581-588] and UDPGTh-2 [Ritter, J. K., et al. (1990) J. Biol. Chem. 266, 7900-7906] have previously been shown to encode isozymes active in the glucuronidation of hyodeoxycholic acid (HDCA) and certain estrogen derivatives (estriols and 3,4-catechol estrogens), respectively. Here we report that the UDPGTh-2-encoded isoform (udpgth-2) and the HLUG25-encoded isoform (udpgth-1) have parallel aglycon specificities. Following expression in COS-1 cells, each isoform metabolized three types of dihydroxy- or trihydroxy-substituted ring structures, including the 3,4-catechol estrogen (4-hydroxyestrone), estriol and 17-epiestriol, and HDCA, but the udpgth-2 isozyme is 100-fold more efficient than udpgth-1. udpgth-1 and udpgth-2 are 86% identical overall (76 differences out of 528 amino acids), including 55 differences in the first 300 amino acids of the amino terminus, a domain which confers isoform substrate specificity. The data indicate that a high level of conservation in the amino terminus is not required for the preservation of substrate selectivity. Analysis of glucuronidation activity encoded by UDPGTh-1/UDPGTh-2 chimeric cDNAs constructed at their common restriction sites, SacI (codon 297), NcoI (codon 385), and HhaI (codon 469), showed that nine amino acids between residues 385 and 469 are important for catalytic efficiency, suggesting that this region represents a domain which is critical for catalysis but distinct from that responsible for aglycon selection.(ABSTRACT TRUNCATED AT 250 WORDS)

Binding Sites↗

PubMed1992

Availability of the B beta(15-21) epitope on cross-linked human fibrin and its plasmic degradation products.

The binding of radiolabeled monoclonal antifibrin antibody 59D8 (specific for fibrin but not fibrinogen) to a series of degraded fibrin clots showed that the availability of the B beta(15-21) epitope (against which 59D8 had been raised) was inversely proportional to the extent of clot lysis. Examination of digest supernatants revealed that the B beta(15-21) epitope was released from clots as a high molecular weight degradation product in the presence of calcium ions but that the generation of low molecular weight peptides occurred in the absence of calcium ions. To address the question of epitope accessibility, we compared levels of fibrin clot binding among four radioactively labeled antibodies: antifibrin monoclonal antibody 59D8, two antifibrinogen monoclonal antibodies that cross-reacted with fibrin, and an affinity-purified polyclonal antifibrinogen antibody. We expected that the antifibrinogen antibodies would show enhanced binding to clots in comparison with the antifibrin antibody. However, the epitope accessibility experiments showed that all four antibody preparations bound fibrin clots at comparable levels. Taken together, these studies demonstrated that one fibrin-specific epitope, B beta(15-21), remains available on clots as they undergo degradation by plasmin and, importantly, that the epitope is not solubilized at a rate faster than the rate at which the clot is itself solubilized. The availability of the B beta(15-21) epitope during the course of plasminolysis assures the potential utility of antifibrin antibodies such as 59D8 for detecting thrombi and targeting plasminogen activators.

Antibodies, Monoclonal↗

PubMed1992

A novel complex locus UGT1 encodes human bilirubin, phenol, and other UDP-glucuronosyltransferase isozymes with identical carboxyl termini.

Two human liver UDP-glucuronosyltransferase (transferase) cDNAs, HUG-Br1 and HUG-Br2, were previously isolated (Ritter, J. K., Crawford, J. M., and Owens, I. S. (1991) J. Biol. Chem. 266, 1043-1047), and each was shown to encode a bilirubin transferase isozyme which catalyzes the formation of all physiological conjugates of bilirubin IX alpha following expression in COS-1 cells. Sequence data showed that the cDNAs contained identical 3' ends (1469 base pairs in length) to each other and to that of the human phenol transferase cDNA, HLUG P1 (Harding, D., Fournel-Gigleux, S., Jackson, M. R., and Burchell, B. (1988) Proc. Natl. Acad. Sci. U.S.A. 85, 8381-8385). Here we report that the two corresponding bilirubin transferases and the phenol transferase are encoded by a novel locus, UGT1, which is also predicted to encode three other bilirubin transferase-like isozymes all having identical carboxyl termini. The transcriptional arrangement utilizes six nested promoter elements, each of which is positioned upstream of a unique exon 1. Each exon 1 encodes the NH2-terminal domain (286 amino acids) and confers the substrate specificity of the isoform. The 3' end of the locus contains 4 common exons which encode the identical carboxyl termini (246 amino acids). It is predicted that six nested primary transcripts are synthesized and that each exon 1 is differentially spliced to the 4 common exons to produce six unique, mature mRNAs. Although the gene organization is present as a single copy, it provides the flexibility of independent regulation of each isoform which is known to occur in the case of bilirubin and phenol transferase activities. With an understanding of the gene structure, lethal, as well as the nonlethal defects, associated with bilirubin transferase activity can now be determined.

Amino Acid Sequence↗

PubMed1992

A sensitive fluorometric assay for reducing sugars.

A simple and rapid fluorometric assay for reducing sugars that is sensitive to the nanomolar range has been developed. The assay involves the derivatization of a given sugar with hydrazine at pH 3 to form a hydrazone, which is reacted with fluorescamine following adjustment of pH to first 9.4 and then 7.4. The amount of sugar in a sample is quantitated by measuring the fluorescence intensity at an excitation wavelength of 400 nm and an emission wavelength of 490 nm. The assay is precise and reproducible, as indicated by intra- and inter-run variations of at most 3% and 4%, respectively. In addition to reducing sugars, the assay can also be used to measure aliphatic and aromatic aldehydes, but not acetone. Compared with an existing fluorometric sugar assay, the assay reported here does not require chromatographic separation of the fluorescent derivative from unreacted fluorescamine. The assay can, however, be potentially adapted for postcolumn detection of aldehydes, reducing sugars, and hydrazones in HPLC.

Aldehydes↗

PubMed1992

The effect of lipid peroxides and superoxide dismutase on systemic lupus erythematosus: a preliminary study.

Serum lipid peroxide (LPO) levels, superoxide dismutase (SOD) activity in peripheral blood erythrocytes, serum ANA, anti-dsDNA, and C3 were measured in 83 patients with systemic lupus erythematosus (SLE) and in 29 healthy controls. Compared to the controls, LPO levels were significantly increased in active SLE patients while SOD activity was markedly decreased. From the active phase to the inactive phase, a gradual decline in LPO levels and an increase in SOD activity were observed. There was a close correlation between LPO levels and disease activity or the parameters including ANA titers, anti-DNA titers, and C3 levels; a significant negative correlation was found between SOD activity and LPO levels or these parameters. Further, a marked difference in SOD activity was found in untreated active cases with and without nephritis. As the SLE patients improved, LPO levels gradually declined and SOD activity increased. The present study indicates that free radicals and resultantly formed lipid peroxide levels are higher in patients with SLE than those in normal persons, probably contributing to the production of autoantibodies, nephritis, and vasculitis of other organs, and that excessively generated free radicals may play an important role in the pathogenesis of SLE, which is possibly due to diminished SOD activity.

Adolescent↗

PubMed1992

Dietary trace elements and esophageal cancer mortality in Shanxi, China.

To explore the relation between esophageal cancer and dietary trace elements in humans, we estimated the average daily intake of zinc, copper, iron, selenium, molybdenum, silicon, cadmium, and nickel in 21 Chinese communes, where the annual mortality rate from esophageal cancer among the population 30 years of age and over ranged from 0 to 495/100,000 person-years. We also estimated the relative level of calcium consumption. Zinc and copper intake were inversely related to esophageal cancer mortality, and calcium intake levels was positively related to esophageal cancer mortality. The predicted esophageal cancer mortality among a vegetarian population with a high level of dietary calcium and a low level of dietary zinc was 5.3 times as high as that in a vegetarian population with a low level of dietary calcium and a high level of dietary zinc. The influence of a high level of dietary calcium in a vegetarian population may be explained by a reduction in the absorption of dietary zinc.

Adult↗

PubMed1992

Mechanisms and rates of decay of marine viruses in seawater.

Loss rates and loss processes for viruses in coastal seawater from the Gulf of Mexico were estimated with three different marine bacteriophages. Decay rates in the absence of sunlight ranged from 0.009 to 0.028 h, with different viruses decaying at different rates. In part, decay was attributed to adsorption by heat-labile particles, since viruses did not decay or decayed very slowly in seawater filtered through a 0.2-mum-pore-size filter (0.2-mum-filtered seawater) and in autoclaved or ultracentrifuged seawater but continued to decay in cyanide-treated seawater. Cyanide did cause decay rates to decrease, however, indicating that biological processes were also involved. The observations that decay rates were often greatly reduced in 0.8- or 1.0-mum-filtered seawater, whereas bacterial numbers were not, suggested that most bacteria were not responsible for the decay. Decay rates were also reduced in 3-mum-filtered or cycloheximide-treated seawater but not in 8-mum-filtered seawater, implying that flagellates consumed viruses. Viruses added to flagellate cultures decayed at 0.15 h, corresponding to 3.3 viruses ingested flagellate h. Infectivity was very sensitive to solar radiation and, in full sunlight, decay rates were 0.4 to 0.8 h. Even when UV-B radiation was blocked, rates were as high as 0.17 h. Calculations suggest that in clear oceanic waters exposed to full sunlight, most of the virus decay, averaged over a depth of 200 m, would be attributable to solar radiation. When decay rates were averaged over 24 h for a 10-m coastal water column, loss rates of infectivity attributable to sunlight were similar to those resulting from all other processes combined. Consequently, there should be a strong diel signal in the concentration of infectious viruses. In addition, since sunlight destroys infectivity more quickly than virus particles, a large proportion of the viruses in seawater is probably not infective.

Journal Article↗

PubMed1992

Antigen-specific suppressor cells in experimental autoimmune uveitis.

Anti-I-A antibodies, administered in vivo at the time of S-antigen injection, suppress development of experimental autoimmune uveitis (EAU) in Lewis rats. While the effects of anti-I-A are profound, the exact mechanism for this suppression is unknown. We attempted adoptive transfer of this form of suppression by injecting lymphocytes from anti-I-A-treated animals into syngeneic recipients which were later injected with S-antigen. Histologically, globes of 75% of the anti-I-A-treated animals showed no inflammation while 25% of these animals developed mild uveitis. In the group of animals which were injected with S-antigen and also received spleen cells from anti-I-A-treated rats, only 1 showed mild uveitis while the remaining 7 had no inflammation. The animals undergoing adoptive transfer of spleen cells and which were primed with an irrelevant antigen, readily developed uveitis. Suppression of S-antigen-induced EAU was abrogated by pretreatment of donor animals with cyclophosphamide. In vitro studies revealed that spleen cells of S-antigen-primed, anti-I-A-treated donors specifically suppressed lymphocyte responses to S-antigen. These in vivo and in vitro results suggest that generation of antigen-specific suppressor cells play a role in the anti-I-A immunotherapy of EAU.

Animals↗

PubMed1992

Acute effects of amiodarone on sodium currents in isolated neonatal ventricular myocytes: comparison with procainamide.

Recent studies suggest that amiodarone's acute clinical effects in infants and children are related predominantly to its class I antiarrhythmic activity. However, the effects of amiodarone on Na+ currents have not been investigated directly in immature cardiac cells. Accordingly, the tight seal whole cell voltage clamp technique was used to measure time- and voltage-dependent Na+ currents in acutely isolated neonatal ventricular myocytes from 2- to 5-day-old rabbits, before and after addition of amiodarone (0.1-10 microM). To evaluate the class I antiarrhythmic activity of amiodarone in this age group, the effects of amiodarone on Na+ currents were compared with those of procainamide. Similar to procainamide, amiodarone significantly decreased peak inward Na+ current in neonatal ventricular myocytes. Moreover, both amiodarone and procainamide shifted the steady-state inactivation curve to more negative membrane potentials and delayed recovery of the Na+ current from inactivation. Thus, the effects of amiodarone on the Na+ current in immature myocardium are qualitatively similar to those of procainamide, suggesting that amiodarone may act acutely as a class I antiarrhythmic agent in the newborn heart.

Amiodarone↗

PubMed1992

Developmental changes in membrane Ca2+ and K+ currents in fetal, neonatal, and adult rabbit ventricular myocytes.

Whole-cell calcium current (ICa) and inwardly rectifying potassium current (IK1) were studied in 21-day fetal, 28-day fetal (total gestation, 31 days), 2-5-day neonatal, and adult rabbit ventricular myocytes isolated by enzymatic dissociation. Whole-cell peak ICa and IK1 at -100 mV increased significantly after birth. Cell size approximated from cell membrane capacitance also increased with age, with the most significant increase occurring after birth. When normalized to cell surface area, peak ICa density increased from day 21 of gestation to the neonate and then increased again from neonate to adult. In all age groups, peak ICa occurred at a test potential of +10 mV, and the shape of the Ca2+ current-voltage relation did not change with age. These findings suggest that there are no significant developmental changes in the voltage dependence of ICa. Therefore, the measured age-related increase in Ca2+ current density may result from increased channel expression. IK1 also exhibited a pattern of increasing current density with age. For IK1, the increase in current density was most rapid between day 21 and the perinatal period and much slower after birth. These results demonstrate that ICa and IK1 undergo significant changes during late fetal and postnatal development.

Action Potentials↗

PubMed1992

ATP-sensitive potassium channels in neonatal and adult rabbit ventricular myocytes.

The properties of the ATP-sensitive potassium (KATP) current were studied in freshly isolated rabbit ventricular myocytes using the patch clamp technique. Removing ATP from the bath (intracellular) solution activated a large K+ conductance in patches from neonatal cells with properties similar to those of KATP channels in other preparations. In membrane patches from neonatal ventricular myocytes, the density of KATP channels was higher than the density of inwardly rectifying K+ channels and the mean patch KATP current was approximately 10 times that of the inwardly rectifying K+ current, at a patch membrane potential of -60 mV. Glibenclamide (10 microM) in the bath solution decreased the number of functional KATP channels, the open-state probability, and the mean patch membrane current. The single-channel conductance of the KATP channel was dependent on the external K+ concentration, and the relationship between channel conductance and external K+ concentration was fit by an exponential equation. In addition, the voltage dependence, channel density, and open-state probability of this channel were compared between neonatal and adult isolated ventricular myocytes. The single-channel conductance and channel density of the KATP channel in neonatal myocytes were significantly smaller than in adult cells. These results suggest that age-related changes occur in the properties of KATP channels.

Action Potentials↗

PubMed1992

The carboxy-terminal region of the glycoprotein hormone alpha-subunit: contributions to receptor binding and signaling in human chorionic gonadotropin.

The glycoprotein hormones are heterodimeric and contain a common alpha-subunit, which is noncovalently associated with a hormone-specific beta-subunit. The alpha-subunit has been highly conserved throughout evolution; for example, the five amino acid residues of the carboxy-terminus, Tyr-Tyr-His-Lys-Ser-COOH, are identical in nine of the 10 available amino acid sequences. It has been shown that enzymatic removal of these five amino acid residues, while not affecting holoprotein formation, results in a heterodimer that exhibits very little, if any, binding to the CG/LH receptor. Using site-directed mutagenesis on the human alpha-subunit, we have prepared two deletion mutants, Des-(88-92)alpha and Des-(89-92)alpha, and two point mutants, where each of the two tyrosines, 88 and 89, was replaced with phenylalanine, in order to delineate more specifically the contributions of these aromatic side-chains to receptor binding. The cDNAs for wild-type hCG alpha and mutants were introduced into a pcDNAINEO expression vector, and the cDNA for hCG beta was inserted into a pRSV plasmid; both were transiently cotransfected into DUXB-11 cells. The media were collected, and RIAs showed that all mutants formed heterodimers; moreover, there was no discernable difference in subunit assembly between wild-type hCG alpha and the various mutant alpha-subunits. The gonadotropin mutants were assayed in vitro using a competitive binding assay with [125I]hCG and stimulation of progesterone production in the transformed murine Leydig cell line MA-10.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1992

A single amino acid residue replacement in the beta subunit of human chorionic gonadotrophin results in the loss of biological activity.

The heterodimer, human chorionic gonadotrophin (hCG), contains an alpha subunit that is common to the glycoprotein hormones and a hormone-specific beta subunit. A comparison of all known beta amino acid sequences shows that an aspartic acid at position 99 (with the numbering scheme for hCG-beta) is one of the seven non-Cys invariant residues. Using site-directed mutagenesis we have replaced hCG-beta Asp99 with Arg. Chinese hamster ovary cells, containing a stably integrated gene for bovine alpha subunit, were transiently transfected with plasmids containing wild-type and mutant hCG-beta cDNAs. The Arg99 beta mutant associated with the alpha subunit, but the resulting heterodimer failed to enhance intracellular cyclic AMP production in a gonadotrophin-responsive transformed murine Leydig cell line. Thus, a single amino acid residue replacement in this glycosylated heterodimer containing 237 amino acid residues is sufficient to abolish activity.

Amino Acid Sequence↗

PubMed1992

Lipid peroxidation in experimental uveitis: sequential studies.

Previously we have detected the occurrence of retinal lipid peroxidation initiated by phagocyte-derived oxygen radicals in experimental autoimmune uveitis (EAU). In the current studies, the confirmation of inflammation-mediated lipid peroxidation was proceeded further to include measurement of multiple parameters, including conjugated dienes, ketodienes, thiobarbituric acid reactive substances and fluorescent chromolipids. The assay for myeloperoxidase, a measure for the number of polymorphonuclear leukocytes in the inflammatory sites was also carried out. The levels of all these parameters were followed through the course of EAU development. The sequential evaluation of histologic changes using both light and electron microscopy was also carried out and the results were correlated with lipid peroxidation indices. These data suggest that the retinal lipid peroxidation plays a causative role in the subsequent retinal degeneration.

Animals↗

PubMed1992

A double (exchange transfusion-carbon clearance) model for testing post-resuscitation reticuloendothelial function.

A double exchange transfusion-double carbon clearance method was evaluated for assessing reticuloendothelial (RE) function following exchange transfusion with hemoglobin solutions. Fifty percent of estimated blood volume (3% body weight) was withdrawn from anesthetized Sprague-Dawley rats and isovolumically replaced with shed blood (SB, control), lysed shed blood (LB, pos. control), human stroma-free hemoglobin solution (SFH), or polyhemoglobin solution (PHS). Thirty minutes after the exchange transfusion, colloidal carbon was injected intravenously and its vascular clearance followed for 1 hour. Then, the 50% exchange transfusion was repeated and the second carbon clearance measured. The intravascular carbon clearance constants, K, and clearance half-times, T1/2, were calculated and compared. No apparent differences in RE function were seen among the groups after the initial exchange transfusion. However, following the second exchange transfusion significant (P less than 0.05) slowing of carbon clearance was observed in lysed blood treated animals. The RE function of SFH or PHS treated animals were not different (P less than 0.05) from that of SB animals. A double exchange transfusion-double carbon clearance method seems to reveal changes in RE function that are not apparent after a single exchange transfusion and clearance test.

Animals↗

PubMed1992

Hepatic reticuloendothelial function following resuscitation with hemoglobin solutions.

Red cell substitutes could lead to depressed reticuloendothelial (RE) particulate clearance function. This hypothesis was tested using an animal model of hypovolemia-resuscitation. Anesthetized male Sprague-Dawley rats were subjected to 50% blood volume hemorrhage followed by isovolumic replacement with stroma-free hemoglobin (SFH, 7 gHb/dl), polyhemoglobin (PHS, 14 gHb/dl), or shed blood (SB). At 30 min post-transfusion, the liver was isolated and perfused with colloidal carbon. Hepatic RE function was assessed from the carbon clearance kinetics. In separate experiments, the hepatic Kupffer cells were isolated and cultured from rats that were previously hemorrhaged and transfused with normal saline solution. The cultured Kupffer cells were incubated with SFH or bovine albumin (ALB) and their phagocytic function assessed in-vitro. The hepatic carbon clearance following exchange transfusion with hemoglobin solutions was not significantly altered as compared to shed blood controls (P greater than 0.05). Similarly, phagocytic function of hemoglobin treated Kupffer cells was not significantly different (P greater than 0.05) from that of ALB treated cells.

Animals↗

PubMed1992

[Studies on the chemical constituents from the skin of Bufo bufo gargarizans Cantor].

Four bufotoxins were isolated by means of preparative reversed phase HPLC from the water soluble fraction of the skin of Bufo bufo gargarizans Cantor. On the basis of their spectral data (UV, IR, 1HNMR, 13CNMR, EIMS and FDMS), amino acid analyses and chemical properties, they were identified as bufotalin 3-succinoylarginine ester(I), bufalin 3-succinoylarginine ester(II), cinobufagin 3-succinoylarginine ester(III), and resibufogenin 3-succinoylarginine ester(IV). Compound I is a new bufotoxin.

Animals↗