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H Lee

Publications and source records attributed to H Lee.

At least 613 records · Page 34Linked to original sources
PubMed1994

Survival of lux-lac-marked biosurfactant-producing Pseudomonas aeruginosa UG2L in soil monitored by nonselective plating and PCR.

Two reporter systems, lacZY and luxAB, were stably integrated into the chromosome of Pseudomonas aeruginosa UG2, a biosurfactant-producing strain. Growth and rhamnolipid production of the UG2 wild-type and reporter gene-bearing UG2L strains were similar in liquid culture. A spontaneous rifampin-resistant detecting UG2Lr, allowed antibiotic selection. Phenotypic characteristics were compared for usefulness in detecting UG2Lr colonies: morphology, fluorescent pigment production, light emission (lux), X-Gal (5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside) cleavage (lac), and rifampin resistance. Survival patterns of UG2, UG2L, and UG2Lr strains were similar in sandy loam soil microcosms over 12 12 weeks. The lac marker was not suitable for monitoring P. aeruginosa UG2Lr in soil since 20 to 42% of cultured, aerobic, heterotrophic soil microorganisms formed blue, lactose-positive colonies. The lux genes provided a stable and unequivocal reporter system, as effective as conventional antibiotic plating, for tracking microorganisms nonselectively at 10(3) CFU/g of soil. Three months after inoculation into oil-contaminated and uncontaminated soil microcosms, UG2Lr cells were recovered at 10(7) and 10(4) cells per g (dry weight) of soil, respectively. Detection by PCR amplification of part of the luxA gene confirmed a decrease in UG2Lr cell numbers in uncontaminated soil. In combination, antibiotic resistance, bioluminescence, and PCR analyses provided sensitive detection and quantitative enumeration of P. aeruginosa UG2Lr in soil.

Base Sequence↗
PubMed1994

Bioluminescent Most-Probable-Number Method To Enumerate lux-Marked Pseudomonas aeruginosa UG2Lr in Soil.

Bioluminescence measured with a luminometer and charge-coupled device was an effective marker in most-probable-number assays for luxAB-marked Pseudomonas aeruginosa UG2Lr in soil. Most-probable-number assays with microtiter plate wells and luminometer tubes gave estimates for UG2Lr that were similar to viable colony counts. Both methods detected five cells per g of soil.

Journal Article↗
PubMed1994

Diagnosis of Chlamydia trachomatis infections in men and women by testing first-void urine by ligase chain reaction.

From April to September 1993, 305 men and 447 women in Hamilton, Canada, consented to the collection of a urethral or cervical swab, respectively, for culture and 20 ml of first-void urine (FVU) for testing by the enzyme immunoassay Chlamydiazyme and by ligase chain reaction (LCR) in the form of a kit from Abbott Laboratories called LCx Chlamydia trachomatis. Evaluation of test performance with each specimen was calculated on the basis of an expanded "gold standard" of a patient found to be positive by culture or by a confirmed nonculture test. By using this expanded standard, the prevalence of infection was determined to be 6% (27/447) for the women and 18.4% (56/305) for the men. LCR testing of FVU in both studies was the most sensitive approach (96%). The performance of Chlamydiazyme was as follows: cervical swab, 78.3% sensitivity; female FVU, 37% sensitivity; and male FVU, 67.9% sensitivity. Culture was the least sensitive approach to diagnosis: female cervix, 55.6%; and male urethra, 37.5%. LCR testing of FVU from men or women diagnosed the greatest number of genitourinary tract infections with no false positives.

Bacteriuria↗
PubMed1994

The middle hepatitis B virus envelope protein is not necessary for infectivity of hepatitis delta virus.

The hepatitis delta virus (HDV) envelope contains the large (L), middle (M), and small (S) surface proteins encoded by coinfecting hepatitis B virus. Although HDV-like particles can be assembled with only the S protein in the envelope, the L protein is essential for infectivity in vitro (C. Sureau, B. Guerra, and R. Lanford, J. Virol. 67:366-372, 1993). Here, we demonstrate that the M protein, previously described as carrying a site for binding to polymerized human albumin, is not necessary for infectivity. HDV-like particles coated with the S plus L or the S plus M plus L proteins are infectious in primary cultures of chimpanzee hepatocytes. We conclude that the S and L proteins serve two essential functions in the HDV replication cycle; the S protein ensures the export of the HDV genome from an infected cell by forming a particle, and the L protein ensures its import into a human hepatocyte.

Animals↗
PubMed1994

Intracellular association of the protein product of the c-myc oncogene with the TATA-binding protein.

The c-myc proto-oncogene encodes nuclear phosphoproteins that bind DNA in a sequence-specific fashion and appear to function as transcriptional activators. Here we demonstrate that a 40-kDa nuclear protein coimmunoprecipitated with c-Myc specifically when nuclear proteins, extracted from nuclei of exponentially growing murine B-lymphoma WEHI 231 cells by using procedures for preparation of trans-acting factors, were reacted with anti-c-Myc antibodies made against different regions of the c-Myc protein. In contrast, preparation of nuclear lysates under denaturing conditions significantly reduced this coprecipitation. Upon incubation of WEHI 231 cells with the reversible chemical cross-linking agent dithiobis(succinimidyl propionate), the 40-kDa protein could be cross-linked to c-Myc protein intracellularly. Identification of the 40-kDa protein as the TATA-binding protein (TBP) of the TFIID transcription initiation complex was made by comigration and V-8 protease mapping, which yielded identical peptide fragments upon digestion of the 40-kDa protein and material immunoprecipitated with an anti-TBP specific antibody. Furthermore, in vitro-translated TBP bound to the amino-terminal portion of c-Myc. Column chromatography of cross-linked nuclear proteins showed TBP to be in a large-molecular-weight complex with c-Myc, consistent with a transcription initiation complex. These results indicate that intracellularly, c-Myc interacts with TBP, suggesting a mechanism of interaction of this oncoprotein with the basal transcription machinery.

Animals↗
PubMed1994

Radiation effects on DNA synthesis in a defined chromosomal replicon.

It has recently been shown that the tumor suppressor p53 mediates a signal transduction pathway that responds to DNA damage by arresting cells in the late G1 period of the cell cycle. However, the operation of this pathway alone cannot explain the 50% reduction in the rate of DNA synthesis that occurs within 30 min of irradiation of an asynchronous cell population. We are using the amplified dihydrofolate reductase (DHFR) domain in the methotrexate-resistant CHO cell line, CHOC 400, as a model replicon in which to study this acute radiation effect. We first show that the CHOC 400 cell line retains the classical acute-phase response but does not display the late G1 arrest that characterizes the p53-mediated checkpoint. Using a two-dimensional gel replicon-mapping method, we then show that when asynchronous cultures are irradiated with 900 cGy, initiation in the DHFR locus is completely inhibited within 30 min and does not resume for 3 to 4 h. Since initiation in this locus occurs throughout the first 2 h of the S period, this result implies the existence of a p53-independent S-phase damage-sensing pathway that functions at the level of individual origins. Results obtained with the replication inhibitor mimosine define a position near the G1/S boundary beyond which cells are unable to prevent initiation at early-firing origins in response to irradiation. This is the first direct demonstration at a defined chromosomal origin that radiation quantitatively down-regulates initiation.

Animals↗
PubMed1994

Bronchodilator aerosol administered by metered dose inhaler and spacer in subacute neonatal respiratory distress syndrome.

There is increasing evidence that bronchodilators are effective in ventilator dependent preterm infants. The effects of single doses of salbutamol (400 micrograms), ipratropium bromide (72 micrograms), and placebo (four puffs) given by metered dose inhaler and spacer (MDIS) were examined in 10 ventilated preterm infants, with a mean birth weight of 800 g at a postnatal age of 1 week, who were suffering from respiratory distress syndrome. The agents were each given in an open, random design. Blood gases were measured and ventilatory efficiency index (VEI) and arterial/alveolar oxygen tension ratio (PaO2/PAO2) were calculated five minutes before and 30 minutes after administration. Heart rate and mean arterial blood pressure were noted. The mean PaO2 improved by 0.61 kPa and 0.69 kPa after salbutamol and ipratropium bromide, respectively and these changes were significantly greater than the 0.5 kPa fall seen with placebo. The mean arterial carbon dioxide tension fell by 0.98 kPa after salbutamol and 0.59 kPa after ipratropium bromide. After both salbutamol and ipratropium bromide, VEI improved significantly (by 23% and 20% respectively) but there was no significant change in the PaO2/PAO2, suggesting that respiratory mechanics and not ventilation/perfusion balance had improved after a single dose of bronchodilator. We conclude that both salbutamol and ipratropium bromide given by MDIS have useful short term effects in ventilator dependent neonates with respiratory distress syndrome. Precise dose regimens and long term effects remain to be worked out.

Aerosols↗
PubMed1994

Survival and respiratory activity of genetically engineered Pseudomonas spp. exposed to antimicrobial agents in broth and soil.

The effectiveness of seven chemical disinfectants were tested against genetically engineered Pseudomonas spp. under optimal growth conditions. Each chemical was tested to determine how quickly 10(8) cells/ml Pseudomonas fluorescens C5t (containing Bacillus thuringiensis endotoxin gene) were killed in King's B broth at 30 degrees C. The minimal bactericidal concentrations (MBC) for calcium hypochlorite, benzalkonium chloride, Germiphene and Spectrum Clear Bath were 0.06% (w/v), 0.01% (w/v), 0.08% (v/v) and 0.044 (v/v), respectively. Virocidin X and CanLab Neutral did not kill P. fluorescens C5t at concentrations up to 20% (v/v). Baxter Bacdown was ineffective as a killing agent at concentrations up to 10% (v/v). These agents were also tested on P. aureofaciens RNL11 (lacZY), P. putida strains PaW8 and PaW340 (containing the plasmid pLV1013 encoding the xylE gene) and P. aeruginosa UG2L (lacZY and luxAB). All strains were killed by similar disinfectant concentrations with the exception of P. aeruginosa UG2L. The MBCs for this strain were double the MBCs for P. fluorescens C5t with all disinfectants used except for Spectrum Clear Bath, which was similar. Oxygen consumption and carbon dioxide evolution were used to assess the activity of P. fluorescens C5t inoculated in non-sterile soil in the presence of antimicrobial agents. Concentrations chosen were 0.1 and 1.0% (w/v) calcium hypochlorite, 0.01 and 0.1% (w/v) benzalkonium chloride, 0.1 and 1.0% (v/v) Germiphene and 0.1 and 1.0% (v/v) Spectrum Clear Bath. Over a 15-day period, the antimicrobial agents did not reduce respiratory activity of P. fluorescens C5t and indigenous soil micro-organisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Containment of Biohazards↗
PubMed1994

Keratinocyte growth factor causes cystic dilation of the mammary glands of mice. Interactions of keratinocyte growth factor, estrogen, and progesterone in vivo.

Keratinocyte growth factor (KGF) is a paracrine mediator of epithelial cell proliferation that has been reported to induce marked proliferation of mammary epithelium in rats. In this study, systemic administration of KGF into naive and oophorectomized mice causes mammary gland proliferation, as evidenced histologically by the appearance of cysts lined by a single layer of epithelium and by hyperplastic epithelium. Whole mount preparations of the mammary glands reveal that the histologically noted cysts are actually ducts that are dilated along much of their length. The histology of the mammary glands of KGF-treated mice is similar to the histology of fibrocystic disease in the human female breast. The response in mice differs significantly from the appearance of the mammary glands in KGF-treated rats in which ductal epithelial proliferation is most prominent. Estrogen and progesterone when administered in combination but not alone cause the development of numerous endbuds in the mouse mammary gland. KGF in estrogen- and progesterone-pretreated mice causes the growth of dilated ducts, hyperplastic epithelium within ducts and endbuds, and a fibrous metamorphosis of periductal adipose tissue. The mammary epithelial hyperplasia caused by KGF is rapidly reversible in both mice and rats after cessation of KGF treatment. The spectrum of KGF-, estrogen-, and progesterone-induced mammary histopathology in mice provides a model for the study of fibrocystic and hyperplastic breast disease.

Animals↗
PubMed1994

SILASTIC ring vertical banded gastric bypass for the treatment of obesity: two years of follow-up in 84 patients [corrected].

The SILASTIC ring vertical gastric bypass (SRVGBP) has evolved as the rational operation to control obesity. The operation consists of a proximal vertical gastric pouch < 30 cc in size. The pouch is banded with a 5.5-cm SILASTIC ring, and this functions as the stoma which does not stretch and is large enough to allow patients to eat all varieties of food, including vegetables and meats, with minimal incidence of postprandial emesis. The continuity of the gastrointestinal tract is formed with a Roux-en-Y gastroenterostomy with each limb about 60 cm long. The bypass of the gastroduodenal axis causes decreased digestion and thus decreased absorption of fats and carbohydrates, resulting in comparably more weight loss than seen in the standard restrictive gastroplasty. The dumping experienced in this operation, which prevents patients from becoming sweet eaters and thus provides long-term weight maintenance, is not as severe as in the regular gastric bypass with a dilatable stoma. In trained hands, the morbidity and mortality from this operation is comparable to that seen in the simple restrictive gastroplasty. The complications due to this operation include staple line breakdown, marginal ulcers, stenosis, incisional hernia, dumping, and iron, vitamins A, B12, D, and E deficiencies. These deficiencies are correctable by oral or parenteral supplements as necessary. This operation yields a 90% or higher success rate (> 40% excess weight loss) in the treatment of morbid obesity [corrected].

Adult↗
PubMed1993

Engineered topographic determinants with alpha beta, beta alpha beta, and beta alpha beta alpha topologies show high affinity binding to native protein antigen (lactate dehydrogenase-C4).

The use of peptides has attracted much interest in the development of synthetic vaccines. Although our current understanding of peptide antigens as immunogens has been greatly advanced recently, there still remain many obstacles. The B cell response elicited by a peptide antigen is governed by a number of poorly understood events such as epitope structure, T cell dependency and major histocompatibility complex restriction, adjuvancy, route of immunization, and immunogen stability. In this paper, we extend our previous studies on the problem of the topographical nature of antigenic sites on native protein antigens, in terms of how much molecular mimicry must be maintained in an antigenic determinant for the induction of high affinity antibodies specific for native protein. We show here that an antigenic epitope from the model contraceptive vaccine candidate lactate dehydrogenase (LDH-C4) can be rationally engineered into a highly structured conformation that mimics the corresponding site in the native three-dimensional protein. Our strategy is based on the selection of an antigenic segment which exhibits certain secondary structural properties and by design principles is fixed in three dimensions by appropriate grafting onto a supersecondary structural motif such as alpha beta, beta alpha beta, or beta alpha beta alpha. The biophysical data are consistent with the proposed secondary structures, and antibodies raised to the various construct show high affinity for the native protein. These studies lend further credence to the conformational nature of peptide epitopes and provide a basis for the rational design of peptide vaccines.

Amino Acid Sequence↗
PubMed1993

Solution conformation of the (+)-trans-anti-[BPh]dA adduct opposite dT in a DNA duplex: intercalation of the covalently attached benzo[c]phenanthrene to the 5'-side of the adduct site without disruption of the modified base pair.

Benzo[c]phenanthrene diol epoxide can covalently bind to the exocyclic amino group of deoxyadenosine to generate [BPh]dA adducts where the polycyclic aromatic hydrocarbon is attached to the major groove edge of DNA. This paper reports on NMR-energy minimization structural studies of the (+)-trans-anti-[BPh]dA adduct positioned opposite dT in the sequence context d(C5-[BPh]A6-C7).d-(G16-T17-G18) at the 11-mer duplex level. The exchangeable and nonexchangeable protons of the benzo[c]phenanthrenyl moiety and the nucleic acid were assigned following analysis of two-dimensional NMR data sets in H2O and D2O solution. The solution structure of the (+)-trans-anti-[BPh]dA.dT 11-mer duplex has been determined by incorporating intramolecular and intermolecular proton-proton distances defined by upper and lower bounds deduced from NOESY data sets as restraints in energy minimization computations. The covalently attached benzo[c]phenanthrene ring intercalates to the 5'-side of the [BPh]-dA6 lesion site without disruption of the flanking Watson-Crick dC5.dG18 and [BPh]dA6.dT17 base pairs. The observed buckling of the intercalation cavity reflects the selective overlap of the intercalated phenanthrenyl ring with dT17 and dG18 bases on the unmodified strand. The structure provides new insights into how a polycyclic aromatic hydrocarbon covalently attached to the major groove edge of deoxyadenosine can still unidirectionally intercalate into the helix without disruption of the modified base pair. Our study establishes that among the contributing factors are a propeller-twisted [BPh]dA6.dT17 base pair, displacement of the carcinogen-DNA linkage bond from the plane of the dA6 base, the specific pucker adopted by the benzylic ring, and the propeller-like nonplanar geometry for the aromatic phenanthrenyl ring system. Our combined experimental-computational studies to date have now identified three structural motifs adopted by covalent polycyclic aromatic hydrocarbon-DNA adducts with their distribution determined by the chiral characteristics of individual stereoisomers and by whether the covalent adducts are generated at the minor or the major groove edge of the helix.

Base Sequence↗
PubMed1993

Physicochemical characterization of a ouabain isomer isolated from bovine hypothalamus.

Recent reports have shown the presence of a ouabain-like inhibitor of Na+/K(+)-ATPase in humans. We have purified a bovine hypothalamic Na+/K(+)-ATPase inhibitory factor (HIF) by using affinity chromatography combined with HPLC. This inhibitor has a molecular weight of 584 as determined by ion-spray mass spectrometry, making it isobaric with ouabain. Glycosidase treatment or acid hydrolysis of HIF released only L-rhamnose, the hexose isomer found in ouabain, as detected by chiral GC/MS. Additionally, enzymatically generated desrhamnosyl HIF was found to have a molecular weight of 438, as does ouabagenin, the aglycone of ouabain. HIF and its aglycone were indistinguishable from ouabain and ouabagenin, respectively, by reversed-phase HPLC retention times. However, derivatization with naphthoylimidazole followed by HPLC revealed different retention times for naphthoylation products of HIF and ouabain. Subsequent CD spectroscopy on isolated naphthoylation products of HIF and ouabain confirmed that they were different. This study provides chromatographic and spectroscopic evidence that ouabain and HIF are isomeric cardenolides. The structural difference is presumed to account for the significant differences in biological properties observed for HIF and ouabain.

Animals↗
PubMed1993

Effect on lymphocyte subsets of clotting factor therapy in human immunodeficiency virus-1-negative congenital clotting disorders. The Transfusion Safety Study Group.

Patients with hemophilia A without human immunodeficiency virus type 1 (HIV-1) infection have lower CD4+ counts and CD4+/CD8+ ratios than controls. This is usually interpreted as a therapy-induced immunodeficiency. Our data re-examine the effect of therapy on peripheral blood mononuclear cell immunophenotypic subpopulations in all congenital clotting disorders. Since late 1985 we have prospectively observed HIV-1 uninfected persons with all types and severity of disorder. Controls were household members without clotting disorders or HIV-1 infection. Analyses of immunophenotype and treatment included a longitudinal random effects model. Compared with controls, age-adjusted CD4+ counts were significantly lower in treated patients (P < .0001) and in patients with all types of clotting disorders who were seldom or never treated (P = .0005). Significantly lower values among both treated and untreated clotting disorder subjects (P < .05) were likewise found for total lymphocytes, several other T-cell subsets, and the CD4+/CD8+ ratio. For most indexes, including the CD4+ count and CD4+/CD8+ ratio, the type of clotting deficiency was not a significant variable. Comparing persons who had no or minimal therapy with those having the most showed increases in CD8+ (P = .0017) and CD20+ CD21- counts (P = .0255), and a lower CD20+ CD21+/CD20+ ratio (P = .0106) in the latter. Controls and persons with clotting disorders differ in CD4+ count. Among those with clotting factor disorders, there is no difference attributable to type of clotting disorder or factor therapy. Large amounts of treatment increased CD8+ and CD20+ CD21- counts, but were not associated with a change in CD4+ count.

Antigens, CD↗
PubMed1993

Signal transduction through extracellular signal-regulated kinase-like pp57 blocked in differentiated colon carcinoma cells having low levels of c-src kinase.

Basic fibroblast growth factor (FGF) induced a rapid increase in tyrosine phosphorylation of a 57-kDa cytoplasmic protein with functional myelin basic protein kinase activity and antigenic properties common to mitogen-activated kinases or extracellular signal-regulated kinases. Basic and acidic FGFs and the diacylglycerol diolein used the same signal transduction pathway to activate pp57. These FGFs, like diolein (Lee, H., Ghose-Dastidar, J., Winawer, S., and Friedman, E. (1993) J. Biol. Chem., 268, 5255-5263), increased the cellular concentration of long-chain diacylglycerols within the same short time period as they increased pp57 tyrosine phosphorylation. Both FGF and diolein increased phosphorylation of pp57 on the same V8 protease-generated fragments, suggesting a common pathway for the phosphorylation of pp57. FGF-induced signal transduction through pp57 mitogen-activated kinase led to cell growth in two undifferentiated colon carcinoma cell lines. In contrast, basic FGF neither increased tyrosine phosphorylation of pp57 nor increased cell growth in two colon goblet cell differentiated lines derived from the same parental line as the undifferentiated cells. Both goblet cell lines exhibited levels of protein-tyrosine kinase activity about one-fifth that of the undifferentiated lines. The decrease in tyrosine kinase activity was not due to down-regulation of FGF receptors or their tyrosine kinase activities. c-src kinase-specific activity was decreased 4-5-fold in both goblet cell lines, suggesting a role for c-src in pp57-mediated signal transduction.

CSK Tyrosine-Protein Kinase↗
PubMed1993

Release of the p53-induced repression on thymidine kinase promoter by single p53-binding sequence.

Herpes simplex thymidine kinase (TK) promoter was shown to be repressed by the wild-type p53. Using a model system that the p53-binding site was linked to the thymidine kinase promoter, we demonstrated that single p53-specific binding site was sufficient to abolish the repression. On the contrary, the mutant p53 had the opposite effects on the HSV-TK promoter in BHK cells. The results suggest that the p53-binding site may act as an enhancer to regulate the gene expression in a novel way in vivo.

Animals↗