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Biomedical subjects

C P Schaffner

Publications and source records attributed to C P Schaffner.

At least 19 recordsLinked to original sources
PubMed1995

Cloning and characterization of a 3-N-aminoglycoside acetyltransferase gene, aac(3)-Ib, from Pseudomonas aeruginosa.

A novel gene encoding an aminoglycoside 3-N-acetyltransferase, which confers resistance to gentamicin, astromicin, and sisomicin, was cloned from Pseudomonas aeruginosa Stone 130. Its sequence was determined and found to show considerable similarity to an aac(3)-I gene previously cloned from R plasmids from Enterobacter, Pseudomonas, and Serratia spp. We have designated the genes from the R plasmids and this work aac(3)-Ia and aac(3)-Ib, respectively. The two aac(3)-I genes share 74% nucleotide identity, and their deduced protein products are 88% similar. These data suggest that the genes derive from a common ancestor. Homology between the flanking sequences of both aac(3)-I genes and other resistance determinants known to reside in integron environments was also observed. Intragenic probes specific for either aac(3)-Ia or aac(3)-Ib were used in hybridization studies with a series of gentamicin-, astromicin-, and sisomicin-resistant clinical isolates. Of 59 clinical isolates tested, no isolates hybridized with both probes, 30 (51%) hybridized with the aac(3)-Ia probe, 12 (20%) hybridized with the aac(3)-Ib probe, and 17 (29%) did not hybridize with either probe. These data suggest the existence of at least one other aac(3)-I gene.

Acetyltransferases↗
PubMed1995

Cellular sterol accumulation stimulated by cholesterol 5 beta,6 beta-epoxide in J774 macrophages.

A significant accumulation of cellular free cholesterol and steryl esters is observed in J774 macrophages when cells are exposed to low-density lipoproteins (LDL) containing cholesterol 5 beta,6 beta-epoxide. This cellular sterol accumulation is mainly due to the formation of esterified cholesterol and desmosterol. Cellular steryl esters increased to 39.4 and 22.4 micrograms/mg cell protein with 0.8 microM of cholesterol 5 beta,6 beta-epoxide and 3,5-cholestadien-7-one, respectively, whereas hardly detectable levels were observed with the absence of oxysterols. The total cellular sterols increased 45% above the value of control with cholesterol 5 beta,6 beta-epoxide. The uptake of [3H] cholesteryl oleate-LDL was also enhanced by cholesterol 5 beta,6 beta-epoxide. The rapid displacement of desmosterol with cholesterol was observed when cells were treated with cholesterol 5 beta,6 beta-epoxide or 3,5-cholestadien-7-one in the presence of LDL. Cholesterol 5 beta,6 beta-epoxide became associated with LDL in the culture conditions, and its uptake into J774 cells and the cytotoxicity were reduced significantly by the association with LDL. The comparison of selected oxysterols for their ability to stimulate cellular sterol accumulation indicated that cholesterol 5 beta,6 beta-epoxide is the most potent. Cholesterol esterification was enhanced significantly by cholesterol 5 beta,6 beta-epoxide whereas cholesterol 5 alpha,6 alpha-epoxide and 3,5-cholestadien-7-one produced a modest response. In contrast, although cholestantriol, the metabolic hydrolysis product of cholesterol epoxides, also associated with LDL, it showed no stimulating effect on both cellular sterol content and sterol esterification. These results indicate that some oxysterols, such as cholesterol 5 beta,6 beta-epoxide and possibly 3,5-cholestadien-7-one, stimulate cellular sterol accumulation in J774 macrophages and may play an important role in atherogenesis.

Animals↗
PubMed1993

Comparative neurotoxicities of amphotericin B and its mono-methyl ester derivative in rats.

The intracisternal administration of amphotericin B (AmB) and its mono-methyl ester derivative (AME), via direct intraventricular injection (0.01 to 5 mg/ml, 6 microliters) in adult female Wistar rats, revealed that AmB was significantly more toxic than AME, as measured by weight loss, lethargy, death, and central nervous system histopathology. Light and electron microscopy confirmed a greater neurotoxicity for AmB, manifested as edema and modest gliosis extending along and beyond the injection tract. Neuronal degeneration and myelin damage were present in AmB-treated (1 mg/ml) animals but were present only modestly in animals treated with AME at a fivefold greater concentration. Intravenous administration of AmB to adult female Wistar rats as five daily doses of 5 mg/kg of body weight resulted in significant weight loss and some deaths. Histopathologic examination of the brains, spinal cords, and sural nerves of surviving animals revealed neurotoxicity manifested by neuronal degeneration, gliosis, and myelin edema. In sharp contrast, similar treatment with AME at a 10-fold greater dose resulted in neither death nor significant neurotoxicity. The administration of five daily doses of a mixture of AME-AmB (9:1; wt/wt) at 50 mg/kg of body weight resulted in neurotoxicity. These results indicate that AmB exhibits significantly greater in vivo neurotoxicity than AME.

Amphotericin B↗
PubMed1992

Human prostatic aldehyde dehydrogenase of healthy controls and diseased prostates.

Aldehyde dehydrogenase (ALDH, EC 1.2.1.3) of the human prostate was the subject of investigation in this study. The possible physiological role of aldehyde dehydrogenase in the human prostate might be to detoxify aldehydes arising from the oxidation of the polyamines via monoamine or diamine oxidases. The specific activity of the enzyme with 1 mM propionaldehyde as substrate and 0.5 mM NAD at pH 7.4 in the control normal prostates and prostates afflicted with the disease, benign prostatic hyperplasia (BPH), was 26.06 +/- 2.96 and 5.17 +/- 0.48 nmol/g prostate per min, respectively. When 100 microM gamma-aminobutyraldehyde was used as a substrate, the specific activity in the normal controls and prostates with benign prostatic hyperplasia was 19.80 +/- 1.33 and 2.95 +/- 2.46 nmol/g prostate per min, respectively. Upon isoelectric focusing of the extracts of the control prostates when the gels were developed for aldehyde dehydrogenase activity, there were three aldehyde dehydrogenase activity bands visible, pI 4.9 (mitochondrial), 5.4 (cytosolic) and about 6.0-6.5, on the IEF gels developed with gamma-aminobutyraldehyde as a substrate. With the extracts of prostates with benign prostatic hyperplasia the pI 4.9 band was significantly reduced, the pI 5.4 band enhanced and the approx. pI 6.0 band was not detectable on the IEF gels with propionaldehyde as a substrate. There was no detectable aldehyde dehydrogenase activity in the extract of the prostate with cancer on IEF gels nor in the activity assays with propionaldehyde or gamma-aminobutyraldehyde as substrates.

Aged↗
PubMed1991

Effect of cholesta-3,5-dien-7-one on human liver aldehyde dehydrogenase.

A recently isolated cholesterol oxidation product, cholesta-3,5-dien-7-one, which was present at high concentrations in fatty/cirrhotic alcoholic liver was identified as a potent endogenous inhibitor of the cytosolic, E1, isozyme of aldehyde dehydrogenase (EC 1.2.1.3). The oxysterol was a less potent inhibitor of mitochondrial, E2, isozyme. The inhibition of the E1 isozyme was irreversible on the IEF gels, upon dilution and with 33 microM 2-mercaptoethanol during activity assay. The calculated 1-50% values from the inhibition curves for the E1 isozyme were 5-10 microM and approx. 180 microM for the E2 isozyme. The E3 isozyme was not sensitive to the oxysterol. Judging from the Lineweaver-Burk plot, the inhibition of the E1 isozyme with a constant concentration of cholesta-3,5-dien-7-one (52 microM) appeared to be noncompetitive.

Adolescent↗
PubMed1990

Applications of thin layer chromatography, high performance liquid chromatography and mass spectrometry in the fermentation and isolation of the antibiotic nybomycin.

Thin layer chromatography (TLC), high performance liquid chromatography (HPLC) and mass spectrometry (MS) methods have been developed for the analysis of the antibiotic nybomycin, its derivatives deoxynybomycin and nybomycin acetate, during the fermentation and isolation of nybomycin. Using a quantitative HPLC based assay, the time course of nybomycin production (nybomycin titers) in 1000 liter fermentations was determined. Desorption chemical ionization mass spectrometry (DCI/MS) of standard nybomycin samples, fermentation broth samples and purified fractions suggested the co-production of deoxynybomycin which was not reported previously from this organism. TLC and HPLC were used to confirm the presence of deoxynybomycin in the crude extracts of fermentation broths.

Anti-Bacterial Agents↗
PubMed1990

Oxysterols and alcoholic liver disease.

A new theory is presented implicating oxidative cholesterol metabolism and oxysterols as possible factors in the development of alcoholic liver disease. Our present studies have revealed the accumulation of cholesta-3,5-dien-7-one, 13.05 +/- 2.75 micrograms/g (n = 8), and cholesta-4,6-dien-3-one, 2.26 +/- 0.88 micrograms/g (n = 8) in fatty alcoholic liver, as compared with controls, 0.21 +/- 0.12 microgram/g (n = 7) and 0.3 +/- 0.33 microgram/g (n = 7), respectively. Acetaldehyde at 1 to 6 micromolar concentration in the blood and tissues of alcoholics cannot account for the extent of tissue damage, nor can it adequately explain liver steatosis characterized by accumulation of cholesterol and fatty acids and their esters in the liver of alcoholics known for their poor dietary habits. Oxysterols may be the primary cause for the development of alcoholic liver diseases and damage to accessory tissues. Significantly lower levels of 7-ketocholesterol in fatty liver, 6.8 +/- 3.5 micrograms/g (n = 8), as compared with control, 36.85 +/- 22.25 micrograms/g (n = 7), may be responsible for the increased cholesterol content of the alcoholic liver due to the inhibitory properties of this sterol on HMG-CoA reductase in cholesterol biosynthesis.

Cholesterol↗
PubMed1990

Comparative toxicities of amphotericin B and its monomethyl ester derivative on glial cells in culture.

Amphotericin B (AmB) is a potent antifungal polyene macrolide antibiotic and is the drug of choice for the treatment of deep-seated mycotic infections. Its use is limited, owing to its nephrotoxicity, and it must be dispersed in deoxycholate for parenteral administration. In contrast, AME (the monomethyl derivative of AmB) is water dispersible, is appreciably less cytotoxic than AmB toward a variety of cell types, and is reportedly active against the acquired immunodeficiency syndrome virus (human immunodeficiency virus type 1). The latter activity has generated interest in AME as an antiviral drug. However, AME is perceived to be neurotoxic, based on the outcome of a human clinical trial of AME as an antifungal drug. AmB is not regarded as neurotoxic, presumably because any neurotoxicity in vivo is precluded by its nephrotoxicity. It was important, therefore, to determine the potential for neurotoxicity of the two agents in comparative tests, assessing the effects of their direct action against neural cells in culture. Rat cortical cells, comprising astrocytes and oligodendrocytes, were used. AME was at least 10 times less toxic than AmB and equally less toxic against several other nonneural cell types also included in these tests. Equally important, AmB disrupted the myelin sheath in these cultures, and it inhibited its generation. AME did not, even at a concentration 10 times greater than the toxic concentration of AmB. AmB is, therefore, potentially more neurotoxic than AME, contrary to current perception. AME is effective as an antifungal and antiviral drug at a concentration far below its toxic concentration for neural cells. Also, AME does not cross the blood-brain barrier appreciably, so that a therapeutic level in blood can be expected without encountering neurotoxicity.

Amphotericin B↗
PubMed1989

Inhibition of HIV replication by liposomal encapsulated amphotericin B.

This report shows the potential of using a liposomal encapsulated preparation of amphotericin B (a polyene macrolide antibiotic) for the in vitro inhibition of HIV. There was no significant difference between the effective doses of the free form of drug when compared to the liposomal encapsulated preparation in inhibiting the growth of HIV. Virus expression was suppressed at a concentration of 5-10 micrograms/ml of the drugs. The liposomal preparation showed greatly reduced cytotoxicity in experiments using cultures of murine leukocytes. These results show the potential usefulness of liposomal encapsulated drugs in the treatment of patients with AIDS or AIDS related complex.

Acquired Immunodeficiency Syndrome↗
PubMed1988

Treatment of fungal infections with semisynthetic derivatives of amphotericin B alpha.

AME appeared to be as effective as AmB in the treatment of mycoses in humans. AME was much less nephrotoxic than AmB, and was better tolerated in terms of rapid onset and reversible adverse reactions. AME may be more ototoxic than AmB. AME, even as AmB and OAME, may cause neurotoxicity and leukoencephalopathy, particularly when high doses are given for long periods.

Adult↗
PubMed1988

Crisamicin C, a new isochromanequinone antibiotic. Isolation, structure determination, and biosynthesis.

Micromonospora purpureochromogenes subsp. halotolerans was found to produce crisamicin C, a novel antibiotic, together with crisamicin A. Crisamicin C was purified by silica gel column chromatography and its physico-chemical properties, structure and biosynthesis were studied. Crisamicin C, mp 260 degrees C (dec), showed UV maxima at 392 (epsilon 9,497), 261 (epsilon 32,959) and 232 nm (epsilon 24,623) in CH3CN, and gave an IR spectrum with absorbances at 1782 (lactone), 1705 and 1655 (quinone) cm-1. Crisamicin C plasma desorption mass spectrometry (PD-MS) m/z 615.9 [M + H)+, hydroquinone) was 16 amu higher than crisamicin A PD-MS m/z 600 [M + H)+, hydroquinone) suggesting that the two antibiotics differ by one additional oxygen in crisamicin C. Analysis of 1H and 13C NMR spectra, in comparison with those of crisamicin A, indicated that crisamicin C was the 4'a, 10'a epoxide derivative of crisamicin A. Carbon-thirteen labeled acetate feeding experiments were used to confirm the positions of the epoxide and other structural features. Crisamicin C was a more potent antibiotic than crisamicin A, but shared the same spectrum of antimicrobial activity (Gram-positive only).

Anti-Bacterial Agents↗
PubMed1988

Studies on the biosynthesis of the antibiotic crisamicin A and carbon-13 magnetic resonance assignments.

The biosynthesis of crisamicin A, a novel dimeric isochromanequinone antibiotic from Micromonospora purpureochromogenes subsp. halotolerans has been investigated by [1-13C] and [2-13C] labeled acetate precursor feeding experiments. Analysis of the proton noise decoupled and off resonance 13C NMR spectra of 13C enriched and unenriched crisamicin A and their acetate derivatives indicated the biosynthesis via the polyketide pathway, as expected. Further analysis of the enriched spectra allowed the complete assignment of the carbon signals. Of particular interest was the establishment of the linkage between the two monomeric halves of the molecule and determination of the location of the phenolic hydroxyls.

Anti-Bacterial Agents↗
PubMed1986

Crisamicin A, a new antibiotic from Micromonospora. I. Taxonomy of the producing strain, fermentation, isolation, physico-chemical characterization and antimicrobial properties.

A microorganism, designated as RV-79-9-101 and now identified as Micromonospora purpureochromogenes subsp. halotolerans, isolated from a mud sample in the Philippines, has been shown to produce a complex of antibiotics called crisamicins. Thin-layer chromatography and bioautography, employing solvent extracts of whole fermentation broths, revealed a minimum of five antimicrobial components. The major biologically-active component of the antibiotic complex, crisamicin A, was obtained in pure form after preparative silica gel column chromatography followed by crystallization. Based on physico-chemical data crisamicin A has been identified as a novel member of the isochromanequinone group of antibiotics. It exhibits excellent in vitro activity against Gram-positive bacteria but little or no activity towards Gram-negative bacteria or fungi.

Anti-Bacterial Agents↗
PubMed1984

Comparative analysis of hexaene antibiotics.

A study of nine hexaene antibiotics resulted in their assignment to three subgroups on the basis of their bioactivities. Separation of individual components of the nine antibiotic complexes was accomplished by thin-layer chromatography. Similarities and differences among members of the subgroups were established by thin-layer chromatography, spectrophotometry and high performance liquid chromatography. Two antibiotics (endomycin and hexafungin) were found to be similar.

Antifungal Agents↗
PubMed1983

Activity of two polyene and two imidazole antimicrobics on Candida albicans in human fibrin clots.

The vegetations of infective fungal endocarditis were simulated by preparing human fibrin clots containing 10(5) CFU of Candida albicans per clot. Clots were incubated in human serum without drugs (controls) or with equimolar concentrations (2.5 to 20 nmol/ml) of four antifungal antimicrobics: AMB and miconazole (both colloidal suspensions) and DAB and KET (both molecularly dispersed). Over a period of 4 days of incubation, clots were resuspended in freshly prepared serum-antimicrobic every 12 hr. Cohorts of control and drug-exposed clots were cultured quantitatively at 24 hr intervals. Neither imidazole at any concentration tested prevented growth of C. albicans to an extent that was significantly different from growth in the control clots. Both polyenes reduced the CFU/clot, with AMB significantly more active than DAB.

Amphotericin B↗
PubMed1982

Epoxycholesterols in secretions and tissues of normal, benign, and cancerous human prostate glands.

Extracts of prostatic secretions taken from 25 control subjects, twenty-three to seventy-five years of age, were analyzed by gas-liquid chromatography (GLC) for cholesterol and possible epoxycholesterol content. Only subjects fifty-three years or older exhibited any signs of prostatic enlargement. Whereas cholesterol (893 +/- 133 micrograms/ml) was found in all prostatic secretions, the carcinogenic epoxycholesterols (22.6 +/- 3.3 micrograms/ml) were detected only in one third of the specimens examined. Prostatic secretion collected from 29 surgical patients prior to primary (20) and secondary (9) prostatectomies revealed cholesterol contents of 1,182 +/- 148 and 1,134 +/- 157 micrograms/ml, respectively. The epoxycholesterols were essentially present at concentrations of 33.7 +/- 7.9 and 24.0 +/- 10.3 micrograms/ml, respectively. Tissue extracts of the different prostatic lobes taken from 40 surgical BPH patients at the time of primary (26) and secondary (14) prostatectomy were examined by capillary GLC. Of these patients 8 had prostatic carcinoma. The cholesterol content of all tissues was essentially double that of normal prostates. The epoxycholesterols were present in all tissues. The mean cholesterol 5 alpha, 6 alpha-epoxide and cholesterol 5 beta, 6 beta-epoxide contents of all tissue were 120 +/- 23.4 and 9.9 +/- 5.2 micrograms/Gm dry tissue, respectively. There were no apparent differences beteeen the lobes nor between BPH and cancers. No epoxycholesterol could be detected in 8 seminal vesicles obtained at autopsy.

Adult↗
PubMed1981

Effect of candicidin on cholesterol and bile acid metabolism in the rat.

Sterol metabolism studies were carried out in rats maintained on a diet containing a polyene antibiotic, candicidin, (30 mg/kg/day) for 2-1/2 months. Compared to the controls, the candicidin-treated animals had a smaller food intake and weight gain during this period. There was no difference between the 2 groups in serum cholesterol levels, biliary cholesterol or bile acid concentrations. However, in the experimental group, liver cholesterol content decreased by 27% and hepatic HMG-CoA reductase increased by 36%. Candicidin administration produced an 84% increase in neutral sterol output without change in bile acid output. Cholesterol absorption was reduced 80% by candicidin feeding. The weight of ventral prostate was reduced 33% by candicidin administration. Prostatic HMG-CoA reductase levels were 3 times higher than those of the liver, but enzyme activity was unchanged by candicidin treatment.

Animals↗