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Biomedical subjects

B L Lee

Publications and source records attributed to B L Lee.

At least 109 records · Page 6Linked to original sources
PubMed1994

Purification and molecular cloning of cDNA for an inducible antibacterial protein from larvae of the coleopteran, Tenebrio molitor.

Antibacterial activity was induced in the hemolymph of larvae of the coleopteran Tenebrio molitor by injection of Escherichia coli. An antibacterial protein, named tenecin 1, was purified to homogeneity from the larval hemolymph and characterized. A cDNA clone for tenecin 1 was isolated and its complete sequence was determined. This protein was found to inhibit the growth of Gram-positive bacteria and to consist of 43-amino acid residues including six cysteine residues. The disulfide structure of tenecin 1 was determined by sequencing cysteine containing peptides obtained by digesting tenecin 1 with endopeptidase Lys-C, trypsin, and thermolysin. The amino acid sequence and its disulfide bonds were similar to those of sapecin and sapecin C, antibacterial proteins of Sarcophaga peregrina.

Animals↗
PubMed1993

Simultaneous determination of hydroquinone, catechol and phenol in urine using high-performance liquid chromatography with fluorimetric detection.

A method was developed for simultaneous determination of urinary hydroquinone, catechol and phenol using high-performance liquid chromatography (HPLC) with variable-wavelength fluorimetric detection. Urine samples, after acid hydrolysis, were saturated with sodium sulphate and extracted by diethyl ether. The two buffers used for gradient elution were (A) 10 mM sodium acetate containing 0.5% (v/v) acetic acid and (B) the same as buffer A but containing an additional 20% (v/v) acetonitrile. Hydroquinone, catechol and phenol were separated in a C18 column and detected at 2.9, 6.8 and 13.6 min, respectively. The recovery and reproducibility were generally over 90%. Over 300 extracted samples were analysed and no change in column efficiency was noted. Comparisons were also made with HPLC using ultraviolet (UV) detection and with gas chromatography (GC). The proposed method appears to be more sensitive and reliable than other existing methods. This new method was also validated with urine samples collected from cigarette smokers and from refinery workers exposed to low concentrations of benzene.

Animals↗
PubMed1993

Determination of plasma ascorbic acid by high-performance liquid chromatography with ultraviolet and electrochemical detection.

A convenient and reliable reversed-phase liquid chromatographic method for the routine determination of ascorbic acid with ultraviolet detection is described. This system avoids the use of modifier and ion-pairing reagent. The mobile phase consists of 20 mM ammonium dihydrogenphosphate with 0.015% metaphosphoric acid. This method enables the detection of plasma ascorbic acid at a concentration of 120 ng/ml within 5 min. The recovery and reproducibility were above 95%. A comparative study was also performed using ultraviolet and electrochemical detectors. Excellent agreement was observed between the two detection modes, with a correlation coefficient of 0.99. In addition, the storage conditions and stability of ascorbic acid in plasma and whole blood were investigated. The results showed that ascorbic acid was more stable in whole blood when stored below 4 degrees C.

Ascorbic Acid↗
PubMed1993

Altered patterns of drug metabolism in patients with acquired immunodeficiency syndrome.

Caffeine was used to assess acetylation status and indexes of oxidative drug metabolism (demethylation, xanthine oxidation, and 8-hydroxylation) in a control group and in three groups of patients infected with human immunodeficiency virus (HIV): patients with acquired immunodeficiency syndrome (AIDS) who had acute illnesses, stable patients with AIDS, and asymptomatic patients infected with HIV. The prevalence of apparent slow acetylation was greater in AIDS patients with acute illnesses compared with control subjects (27 of 29 [93%] versus 18 of 29 [62%]). Indexes of demethylation were decreased and 8-hydroxylation increased in these patients. Xanthine oxidation was the same as that in the control subjects. In the stable AIDS patients, oxidative pathways were altered in a manner similar to that observed in the AIDS patients with acute illnesses, but acetylation was the same as that in the control subjects. In HIV-infected asymptomatic patients, drug metabolism was the same as that in the control subjects. The increased prevalence of apparent slow acetylation and the altered activity of the oxidative pathways in AIDS patients with acute illnesses may partly explain the increased incidence of adverse drug reactions in these patients.

Acetylation↗
PubMed1993

Expression of follistatin mRNA by somatotropes and mammotropes early in the rat estrous cycle.

We previously found follistatin (FS) mRNA in gonadotropes [predominantly in cells with luteinizing hormone (LH) antigens] and folliculostellate cells (with S100 antigens) in diestrus rats pituitaries. However, earlier in the cycle, when percentages of gonadotropes are lowest, percentages of cells expressing FS are 1.5-2-fold higher than in diestrus. This study was designed to detect FS mRNA and other pituitary antigens to identify the additional cells with dual in situ hybridization and immunolabeling protocols. The mRNA was detected with biotinylated complementary oligonucleotide probes and avidin-biotin-peroxidase complexes. Significant labeling for FS mRNA was found in cells with the following antigens: growth hormone (GH) (7% of pituitary cells); prolactin (PRL) (5%); S100 protein (5%); follicle-stimulating hormone (FSH beta) (4%); LH beta (3%); and thyroid-stimulating hormone (TSH beta) (3%). Optimal conditions for detection included: overnight plating of > 50,000 cells/well (24-well tray) in media containing 10% fetal bovine serum; hybridization at 37 degrees C; and fixation in 2% glutaraldehyde. Whereas FS is expressed predominantly by LH gonadotropes at midcycle, FS mRNA can be expressed by all types of antigen-bearing cells earlier in the cycle. Its function in the pituitary may relate to its role in binding activin, which would result in inhibition of FSH release. However, since activin inhibits secretion of GH, PRL, and adrenocorticotropin (ACTH), FS may also control activin's effects on these cells. The FS-expressing cells may therefore be paracrine or autocrine regulators.

Animals↗
PubMed1993

Structural studies of telomeric DNA, RNA and DNA-RNA hybrid oligonucleotides by non-denaturing polyacrylamide gel electrophoresis.

Five kinds of DNA, RNA and DNA-RNA hybrid oligomers(T1-T5) containing Tetrahymena and human telomeric sequences were synthesized. T1 is d(T2G4). T2, T3, T4 and T5 are d(T2AG3), h(U2dG4), h(T2rG4) and r(U2G4) in sequences, respectively. Their structural changes were monitored by gel mobility shift on non-denaturing PAGE. The susceptibility of dG in telomeric structure was tested by methylation reaction by DMS. Our results show that telomeric DNA structure is dependent on dG to form intramolecular hydrogen bonding in the hairpin structure.

Animals↗
PubMed1993

Combination drug testing of Mycobacterium chelonae.

PURPOSE: Medical therapy of Mycobacterium chelonae keratitis is difficult because there are so few effective antimicrobial agents and single agent therapy frequently fails clinically. To identify more effective medical treatment regimens, the in vitro antimicrobial efficacy of amikacin, the most frequently used single agent, was investigated in combination with four antibiotics previously reported to have activity against M. chelonae: erythromycin, imipenem, ciprofloxacin, and vancomycin. METHODS: The drug combinations were tested by the checkerboard method against seven corneal isolates of M. chelonae. RESULTS: The combination of amikacin with erythromycin or vancomycin consistently led to synergistic or additive effect, however the minimum inhibitory concentrations for vancomycin were very high. The combination of amikacin with imipenem or ciprofloxacin led to results ranging from antagonism to additive effects. CONCLUSIONS: Of the antibiotics tested, erythromycin showed the most activity against M. chelonae in combination with amikacin. In vitro combination drug testing of M. chelonae by the checkerboard method should be further evaluated for clinical relevance in microbial keratitis.

Amikacin↗
PubMed1993

Urinary trans,trans-muconic acid determined by liquid chromatography: application in biological monitoring of benzene exposure.

We describe a sensitive and specific high-performance liquid-chromatographic method for determining the benzene metabolite, trans,trans-muconic acid (ttMA) in urine by measuring ultraviolet absorbance at 265 nm. We mix 1 mL of urine sample with 2 mL of Tris buffer containing vanillic acid as internal standard (IS) and percolate this through a preconditioned ion-exchange column. After rinsing the column with phosphoric acid solution, acetate buffer, and deionized water, we elute the analytes with 2 mL of an equivolume solution of 1.5 mol/L sodium chloride and methanol. Of this, 5 microL is injected into the HPLC column. The mobile phase used consists of, per liter, 10 mL of acetic acid, 100 mL of methanol, and the rest 5 mmol/L sodium acetate. The flow rate was started at 1 mL/min and increased to 1.5 mL/min after 6 min. ttMA and IS were detected at 5.2 and 10.2 min, respectively. The lowest detection limit is 125 pg. Analytical recovery and reproducibility generally exceeded 90%. We validated the method with urine samples collected from normal persons and from refinery workers exposed to benzene concentrations < 1 microL/L. The results show that urinary ttMA is a promising biological marker for risk assessment of low-concentration benzene exposure.

Benzene↗
PubMed1992

High-performance liquid chromatographic method for routine determination of vitamins A and E and beta-carotene in plasma.

A simple and reliable reversed-phase high-performance liquid chromatographic (HPLC) method for the routine determination of vitamins A and E and beta-carotene in plasma (or serum) with wavelength-programmed ultraviolet-visible absorbance detection is described. A 200-microliters aliquot of serum or plasma sample, after deproteinization with ethanol, and containing tocopherol acetate as internal standard, was extracted with butanol-ethyl acetate. Sodium sulphate was added for dehydration. Analytes of extracted samples were found to be stable for at least four days. A 10-microliters aliquot of this organic extract was used for HPLC analysis. The mobile phase was methanol-butanol-water (89.5:5:5.5, v/v) and the flow-rate was set at 1.5 ml/min. The analytes of interest were well separated from other plasma constituents within 22 min at 45 degrees C. The lowest detection limits of vitamins A and E and beta-carotene were 0.02, 0.5 and 0.1 microgram/ml, respectively. The recovery and reproducibility of the present method were around 90%. The method is sensitive, specific and can be used for epidemiological studies and for routine determination of vitamin deficiency. Several important factors that may affect the analysis are also discussed in this paper.

Carotenoids↗
PubMed1992

The solubility of antibiotic and corticosteroid combinations.

The use of collagen shields soaked in various combinations of medications has been advocated to enhance drug delivery to the cornea. Recently, severe corneal toxicity associated with aggregate formation in mixtures of gentamicin and methylprednisolone prompted our study of the effect of drug concentration, pH, and temperature on the solubility of several antibiotic and corticosteroid formulations commonly used to treat ocular disease. Selected combinations of cefazolin, vancomycin, tobramycin, gentamicin, methylprednisolone, and dexamethasone were evaluated. Mixtures of tobramycin and vancomycin produced no precipitates, but many spheroid aggregates were seen when methylprednisolone and gentamicin were combined. Although the effects of precipitate formation on drug bioavailability and toxicity have not been fully determined, until such information is available, the use of combinations of drugs that remain in solution during administration is recommended.

Anti-Bacterial Agents↗
PubMed1992

Bile pigments in gallbladder and freshly-secreted hepatic duct bile from fed and fasted rainbow trout, Oncorhynchus mykiss.

1. Chromatographic analyses of bile pigments in rainbow trout reveal the presence of primarily unconjugated biliverdin (BV) and bilirubin (BR) glycosyl conjugates. Only trace amounts of unconjugated BR are present in hepatic duct (HD) bile: no beta-glucuronidase activity is detectable. 2. The per cent of BV and BR in HD and gallbladder biles is similar in fasted trout; however, the per cent of BV is significantly increased in HD bile from fed fish. 3. Fasting decreases the rate of choleresis but does not alter the excretory rate of endogenous BV or BR. 4. Erythrocyte life span is estimated to be approximately 500 days.

Animals↗
PubMed1992

Interactions and toxicities of drugs used in patients with AIDS.

As clinicians have come to realize, patients who have AIDS are uniquely predisposed to drug-associated toxic effects and allergic reactions. In San Francisco General Hospital's first clinical trial involving patients with pneumonia due to Pneumocystis carinii, only one-third of all patients who were treated initially with either trimethoprim-sulfamethoxazole or pentamidine were able to complete the 3-week course. Because of drug-related toxic effects, most patients required an alteration in therapy. In addition, with the multi-drug approach to management of infection due to human immunodeficiency virus (HIV) and its complications, drug-drug interactions commonly complicate the pharmacokinetics and toxicities of the therapeutic agents used. Both factors make caring for HIV-infected patients a complicated and perplexing process for the clinician. We've invited Drs. Belle Lee and Sharon Safrin of the University of California, San Francisco School of Medicine to contribute the AIDS Commentary this month. Drs. Lee and Safrin are active investigators in the field of pharmacokinetics and conduct clinical trials of agents used in the treatment of patients who have AIDS. In this commentary, they review what is known about the complexities involved in treating patients with AIDS and offer some practical suggestions for management.

Acquired Immunodeficiency Syndrome↗
PubMed1992

Heightened secretion by small and medium-sized luteinizing hormone (LH) gonadotropes late in the cycle suggests contributions to the LH surge or possible paracrine interactions.

LH secretion from pituitary cell fractions separated by centrifugal elutriation was compared to learn whether any contributed to the LH surge. After plating (1 h) and stimulation with 0-10 nM [D-Lys6]GnRH (4 h), some fractions secreted levels that were out of proportion to their enrichment. Large cells from proestrous morning rats (3-fold enriched) secreted 4.3 times more LH, and medium-sized fractions (1.5-fold enriched) secreted 2-3.4 times more LH than unseparated cells during estrus or metestrus. Normalized data (nanograms per 20,000 cells) showed that basal levels reflected the enrichment in the fractions. Data were also normalized to nanograms of LH per 1,000 LH gonadotropes to focus on LH cell activity. Basal secretion in unseparated cultures (4-6 ng/ml/1,000 LH cells) was lower than that in small or large LH cells during all stages except proestrus, when small gonadotropes became as active as those in unseparated cultures, and large gonadotropes secreted 2-3 times more LH. Basal secretion from medium-sized gonadotropes was comparable to that in unseparated cultures. [D-Lys6]GnRH-mediated secretion from unseparated, small, and large LH cells was comparable during most stages. However, during proestrus, large gonadotropes secreted 2.2 times more LH than unseparated counterparts. Stimulated medium-sized LH cells were 1.3-2.3 times more active in most stages than those in unseparated cultures and 1.75-2.8 times more active than those in large cell fractions (from proestrous PM to metestrus). Whereas this enhanced secretion late in proestrus suggests that medium-sized LH cells may support the LH surge, it also may reflect the removal of regulatory factors from cells in other fractions. Studies of autocrine or paracrine interactions with gonadotropes are needed to test this hypothesis.

Animals↗
PubMed1992

Follistatin gene expression in the pituitary: localization in gonadotropes and folliculostellate cells in diestrous rats.

Follistatin, a glycosylated single chain protein that was originally isolated from ovarian follicular fluid, can specifically inhibit the biosynthesis and secretion of FSH by the pituitary. Follistatin has also been isolated from bovine pituitary and shown to have activin-binding activity. We wished to determine whether the follistatin gene is expressed in the rat pituitary and, if so, to identify the specific cell types. A 337-basepair fragment of the follistatin cDNA was amplified by polymerase chain reaction from a rat ovarian cDNA library and subcloned into pGEM3. Low levels of follistatin mRNA from rat pituitary poly(A)+RNA were detected by ribonuclease protection analysis using a specific follistatin riboprobe generated from the cDNA clone. The presence of follistatin mRNA in the pituitary was confirmed using polymerase chain reaction to amplify the follistatin cDNA generated by reverse transcription from total rat pituitary RNA. Furthermore, in situ hybridization studies combined with immunostaining for pituitary hormones were used to localize follistatin mRNA within the rat pituitary. When a biotinylated oligonucleotide complementary to follistatin mRNA was used with dispersed pituitary cells from rats in diestrus II, labeling was found in 5-7% of the cells. The in situ hybridization protocol was then combined with immunolabeling protocols for LH beta, FSH beta, or S-100 protein (a marker for folliculostellate cells). Follistatin mRNA was detected in 70 +/- 5% of LH beta cells, 44 +/- 11% of FSH beta cells, and 35 +/- 2% of folliculostellate cells. These results suggest that follistatin is expressed in pituitary gonadotropes and folliculostellate cells during diestrus II, where it may have a role in the local autocrine or paracrine regulation of FSH biosynthesis and secretion, possibly by binding to and modulating the effects of activin in the pituitary.

Animals↗
PubMed1992

Maturation of follicle-stimulating hormone gonadotropes during the rat estrous cycle.

FSH mRNA is transcribed after the onset of high FSH secretion during proestrus and estrus. Pituitary cell fractions separated by size and density were studied to determine if expression of FSH mRNA activity was predominantly in one subset during the estrous cycle and to determine the source and significance of "silent FSH" cells that secrete FSH, but store too little for detection. Pituitary cells were separated by centrifugal elutriation, plated, and then exposed to 0.1-1 nM [D-Lys6]GnRH for 3 h. Media were assayed for FSH by RIA, and the cells were fixed for immunocytochemistry or in situ hybridization. The percentages of immunoreactive FSH cells in unseparated populations increased from 8% at metestrus to 12% during proestrus. Percentages of cells with FSH beta mRNA showed the same rise; however, peak levels were higher (17%) during proestrus and estrus. Small cells with FSH beta mRNA were more frequent than those with antigens early in the cycle. The largest cell fractions contained 38-44% immunoreactive cells. Only 8-21% of these cells had FSH beta mRNA, except during the morning of proestrus (33%). The distribution analyses showed that the increment in immunoreactive FSH cells during diestrus initially stemmed from smaller subsets; however, over half of immunoreactive FSH cells were large by the evening of proestrus. During the time of active transcription of FSH mRNA, more than half of the cells with FSH beta mRNA were small or medium-sized. Thus, early in the cycle, FSH beta mRNA is transcribed in the smaller cells, which may be the source of the silent FSH cells reported in previous studies. During proestrus, smaller FSH cells also secreted as well if not better than those in the unseparated population or large fractions. When they secreted more than expected from their percentages of FSH cells, this response was interpreted to be due to either the presence of cells that are immunoreactively silent or the possible removal of autocrine or paracrine regulatory factors.

Animals↗
PubMed1992

Oral SCH 39304 as primary, salvage, and maintenance therapy for cryptococcal meningitis in AIDS.

To determine the efficacy and toxicity of SCH 39304 in the treatment and suppression of cryptococcal meningitis, we conducted a prospective, noncomparative study in three groups of patients: patients with acute cryptococcal meningitis, patients with acute cryptococcal meningitis in whom other therapies have failed (salvage), and patients who required maintenance therapy. As primary therapy, the patients received up to 14 days or 1 g of amphotericin B followed by SCH 39304 200 mg once daily for 12 weeks. As maintenance therapy, the patients received SCH 39304 600 mg once weekly for 12 months. Of five salvage patients, none completed the study. Two patients died, two patients clinically deteriorated, and one patient was noncompliant. Two of three patients with acute cryptococcal meningitis completed the 12-week primary therapy, and one patient was discontinued from therapy because of a skin rash (95% confidence interval, 14-100%). All four patients who were receiving weekly maintenance therapy followed up to 27 weeks were clinically stable with no change in their serum cryptococcal antigen titer from baseline when the study was prematurely terminated. Elevation of liver function test results developed in three patients and skin rash developed in one patient. The unique pharmacologic and pharmacokinetic properties of SCH 39304 (low incidence of toxicity, long serum half-life, and good penetration into the cerebrospinal fluid) lend promise to pursue other triazole antifungals at higher doses as primary therapy and less frequent dosing for maintenance therapy.

Acquired Immunodeficiency Syndrome↗