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Biomedical subjects

J Kiefer

Publications and source records attributed to J Kiefer.

At least 73 records · Page 4Linked to original sources
PubMed1993

Analysis of the inversion effect in pulsed field gel electrophoresis by a two-dimensional contour-clamped homogeneous electric field system.

A two-dimensional contour-clamped homogeneous electric field electrophoresis was used for the separation of the yeast Saccharomyces cerevisiae chromosomes. The role of different pulse times and field strengths was examined. The resulting complex migration curves were similar to those predicted by the bag model. They are characterized by a straight line in the region of lower molecular weight where the migration is inversely proportional to the molecular weight and a mobility inversion in the region of higher molecular weight both depending on the conditions chosen for the two directions. The quantitative analysis of the data was compared with the predictions of the bag model which is shown to be a good approximation. Using this experimental approach the separation pattern of a commercially available S. cerevisiae standard was examined under standard electrophoresis conditions, and chromosomes IV and XII were found to be inverted.

Chromosomes, Fungal↗
PubMed1993

Problems, pitfalls, perspectives and potentials of quantitative theoretical models for cellular radiation action.

The problem of quantitative mathematical models in cellular radiation biology is discussed in a general way. It is emphasized that there are a number of stages, starting from the spatial pattern of energy deposition and ending with repair/misrepair processes which all need to be incorporated. Since different types of radiation commonly yield very similar dose-response curves a model which is only valid for one special case cannot claim general applicability. Interaction experiments with ultraviolet and ionizing radiation are discussed in this context. Also the role of different experimental systems (microorganisms versus mammalian cells) has to be taken into account. A number of current model approaches are discussed within this context, and it is shown that most of them do not satisfy the criterion of universal applicability and can therefore not claim to give a 'true' picture of biological reality. Shouldered survival curves are taken as an example to illustrate these points in a more specific way.

Animals↗
PubMed1993

The significance of DNA double-strand breaks in the UV inactivation of yeast cells.

The radiation-sensitive mutant rad 54-3 of Saccharomyces cerevisiae temperature-conditional for the repair of DNA double-strand breaks was exposed to 254 nm ultraviolet radiation and incubated at the restrictive and permissive temperatures. A large difference in survival was seen indicating the involvement of double-strand breaks in cellular inactivation at least in this strain. Pulsed-field gel electrophoresis of DNA showed that double-strand breaks are not directly induced but develop upon incubation under growth conditions. Their number is highest after about 4 h, after 8 h repair is complete in wild-type cells. With the aid of the excision-deficient double mutant rad3rad54 it could be demonstrated that strand break formation proceeds independent of excision repair.

DNA Damage↗
PubMed1993

DNA double-strand break induction in yeast by X-rays and alpha-particles measured by pulsed-field gel electrophoresis.

Pulsed-field gel electrophoresis was used to separate the chromosomes of the diploid yeast Saccharomyces cerevisiae 211*B after irradiation with X-rays and alpha-particles. After electrophoresis, gels were stained with ethidium bromide, placed on a UV-transilluminator and photographed with a digitizing camera connected to a personal computer. The pictures obtained were processed with the help of specially developed software which allows for the correction of the camera's shading effect and background fluorescence. Linearity between DNA amount and fluorescence was demonstrated. Fluorescence intensity for the band with the lowest electrophoretic mobility was found to decrease exponentially with dose. Based on the known size of the native DNA molecules, double-strand break yields could be calculated. These were found to be (8.2 +/- 0.4) and (14.8 +/- 0.5)10(-12) (g/mol)-1 Gy-1 for 80 keV X-rays and 3.5 MeV 241Am alpha-particles respectively which gives a relative biological effectiveness of 1.8 +/- 0.1.

Alpha Particles↗
PubMed1993

A novel technetium-99m labeled monoclonal antibody (174H.64) for staging head and neck cancer by immuno-SPECT.

A novel murine monoclonal antibody (MAb 174H.64) was labeled with 99mTc by a direct method. MAb 174H.64 detects a cytokeratin-associated antigen which is expressed by over 90% of all squamous cell carcinomas. Panendoscopy, sonography and computerized tomography scan were performed in all cases as well as magnetic resonance imaging (in selected patients). Pre-operative immunoscintigraphy was performed in 21 patients with histologically proven primary carcinomas (18 with remaining primary tumors and 3 with lymph node recurrences). Scintigraphic images were obtained 4-6 h after injection of 1.1 GBq of the 99mTc-labeled antibody (2 mg). Late images were acquired 18 to 24 h after injection. Single-Photon-Emission-Computed Tomography (SPECT) of the head and thorax was performed in all patients. The primary tumors were immunoscintigraphically visualized in all 18 patients with remaining primary tumor. Fifteen of 18 loco-regional lymph node metastases were visualized by immunoscintigraphy (the smallest lesions had a diameter of < 1 cm), in one patient lymph node metastases were detected by immunoscan only. Two metastatically involved lymph nodes were identified by histology only (micrometastases). Distant metastases were present in 3 patients, of which two were identified by immunoscintigraphy. Immuno-SPECT according to this method was a sensitive and specific imaging modality for preoperative staging of patients with squamous cell carcinoma of the head and neck and detected lymph node metastases with higher accuracy than conventional clinical and imaging modalities.

Animals↗
PubMed1992

Inactivation and mutation induction in Saccharomyces cerevisiae exposed to simulated sunlight: evaluation of action spectra.

The effectiveness of polychromatic light irradiation was investigated for haploid yeast cells. Inactivation and mutation induction were measured in both a RAD-wildtype strain and an excision-repair defective strain. The behaviour of vegetative "wet" cells was compared to that of dehydrated cells. The aim of the study was to assess the interaction of UVC with other wavelengths in cells of different states of humidity. The irradiation procedure was therefore carried out using a solar simulator either with full spectrum or with a UVC-blocking filter (modified sunlight) added. The results were analysed on the basis of separately determined action spectra. The summation of the efficiency of individual wavelengths was compared to the values obtained from polychromatic irradiation. It is shown that the effects caused by the whole-spectrum irradiation in wet cells can be predicted sufficiently from the calculation, while dried wildtype cells exhibit higher mutation rates. Thus it can be assumed that drying-specific damage leads to lethal and mutagenic lesions which are processed in different ways, causing a synergistic behaviour in mutation induction. Irradiation of vegetative cells with modified sunlight (UVC-) results in less inactivation and lower mutation rates than were calculated. From these results it can be concluded that this antagonistic behaviour is caused by the interaction of near-UV photoproducts.

Dose-Response Relationship, Radiation↗
PubMed1992

Heavy ion effects on cells: chromosomal aberrations, mutations and neoplastic transformations.

The quantification of chromosomal aberrations, mutations and neoplastic transformations induced by heavy charged particles meets with considerable experimental and conceptual difficulties which are related to the specific pattern of energy deposition. These problems are discussed on the background of current ideas on track structure, and some new experimental results are analysed.

Animals↗
PubMed1992

Mutation induction in mammalian cells by very heavy ions.

V79 Chinese hamster cells were exposed to heavy ions (O to U) and assayed for mutants at the HGPRT-locus by incubation in selective medium containing 6-thioguanine. The LET ranged from 300 to 18000 keV/micrometer. Mutants could be recovered from all particle radiation but the effectivity per deposited energy decreased with atomic numbers greater than 8. The results are discussed with regard to fundamental processes of cell reactions to very heavy ions and with respect to possible implications for hazard estimations.

Animals↗
PubMed1992

Heavy ion-induced DNA double-strand breaks in yeast.

DNA double-strand break (dsb) induction in diploid yeast was measured by neutral sucrose sedimentation after exposure to very heavy ions with values of linear energy transfer (LET) ranging from about 300 to 11500 ke V/microns. Linear fluence dependencies were found in all cases from which dsb production cross-sections (sigma dsb) could be calculated. Corresponding cross-sections for cell killing (sigma i) were derived from final slopes of survival curves measured in parallel and for the same fluence range. A close correlation was found between sigma i and sigma dsb. It is calculated that over the entire LET range, including 30 MeV electron irradiation, about 22 dsb are induced per lethal event when high exposures are considered.

Acceleration↗
PubMed1992

Protein synthesis in irradiated cells. I. Ultraviolet radiation.

Excision-deficient haploid yeast cells (Saccharomyces cerevisiae) were exposed to 254-nm UV radiation and protein synthesis inhibition was measured for a large number of different proteins resolved by two-dimensional gel electrophoresis. The derived UV-radiation sensitivities exhibited an overall increase with protein molar mass. Quantitatively, this behavior is compatible with a well known mechanism of transcription inactivation--termination of RNA chains at UV-radiation-induced pyrimidine dimers--if the respective target sizes are inferred from protein molar mass. The observed deviations from the predicted response suggest that (i) UV-radiation damage may also interfere with recognition/binding of RNA polymerase to regulatory sequences and (ii) the frequency of photolesions for a specific protein encoding gene may differ markedly from the mean induction rate for the total yeast genome.

Fungal Proteins↗
PubMed1991

Changes in cell cycle distribution of V79 Chinese hamster fibroblasts after irradiation at different wavelengths.

Changes in cell cycle distribution of V79 Chinese hamster fibroblasts were investigated at different wavelengths between 254 and 313 nm. The fluences applied led to surviving fractions of 0.61. In all cases, the S fraction was temporarily increased within 8-12 h after irradiation, whereas the G1 fraction was decreased. The maximum deviations from the initial values did not significantly depend on the wavelength.

Animals↗
PubMed1991

Heavy ion effects on cells: survival of a temperature-conditional repair mutant of yeast.

The temperature-conditional double-strand break repair mutant rad 54-3 of the yeast Saccharomyces cerevisiae was exposed to Ar, Ti and U ions with LET values between 900 and 15,000 keV/micron. Survival was assessed after incubation at the permissive (23 degrees C) and the restrictive temperature (36 degrees C) in aliquots of the same sample. Repair could be demonstrated in all instances, although to a somewhat reduced extent with the heavy ions as compared to X-rays. The results suggest that very densely ionizing radiations do not produce lesions which are irreparable per se, but that their ultimate fate depends on the particular repair system which may be different in different cell types.

Argon↗
PubMed1991

Inhibitory effects of furocoumarins in Salmonella typhimurium TA98 on the mutagenicity of dictamnine and rutacridone, promutagens from Ruta graveolens L.

Eight furocoumarins differing in their basic structure and substitution pattern (angular, linear, dihydrofuran type) were tested for their ability to reduce the mutagenic potency of dictamnine and rutacridone, two alkaloids present in extracts from Ruta graveolens L. Both compounds need metabolic activation by S9 mix in order to exhibit mutagenicity in Salmonella typhimurium strain TA98. The furocoumarins used in this study did not show any mutagenicity either with or without S9 mix within the dose range tested. However, all the furocoumarins were able to inhibit the mutagenicity induced by dictamnine as well as by rutacridone in a dose-dependent manner. Imperatorin turned out to be the most efficient inhibitor. The inhibitory effect is probably due to the inactivation of the cytochrome P450 enzyme complex which prevents the activation of the promutagens. This is indicative of the desmutagenic character of the furocoumarins. However, there is also some evidence that the reduction of the mutagenicity induced by dictamnine might be caused to a small extent by a mechanism which possibly depends on the competition with furocoumarins for the same sites in the DNA molecule.

Alkaloids↗
PubMed1991

Phenotype-specific phosphorylation of simian virus 40 tsA mutant large T antigens in tsA N-type and A-type transformants.

To identify molecular differences between simian virus 40 (SV40) tsA58 mutant large tumor antigen (large T) in cells of tsA58 N-type transformants [FR(tsA58)A cells], which revert to the normal phenotype after the cells are shifted to the nonpermissive growth temperature, and mutant large T in tsA58 A-type transformants [FR(tsA58)57 cells], which maintain their transformed phenotype after the temperature shift, we asked whether the biological activity of these mutant large T antigens at the nonpermissive growth temperature might correlate with phosphorylation at specific sites. At the permissive growth temperature, the phosphorylation patterns of the mutant large T proteins in FR(tsA58)A (N-type) cells and in FR(tsA58)57 (A-type) cells were largely indistinguishable from that of wild-type large T in FR(wt648) cells. After a shift to the nonpermissive growth temperature, no significant changes in the phosphorylation patterns of wild-type large T in FR(wt648) or of mutant large T in FR(tsA58)57 (A-type) cells were observed. In contrast, the phosphorylation pattern of mutant large T in FR(tsA58)A (N-type) cells changed in a characteristic manner, leading to an apparent underphosphorylation at specific sites. Phosphorylation of the cellular protein p53 was analyzed in parallel. Characteristic differences in the phosphorylation pattern of p53 were observed when cells of N-type and A-type transformants were kept at 39 degrees C as opposed to 32 degrees C. However, these differences did not relate to the different phenotypes of FR(tsA58)A (N-type) and FR(tsA58)57 (A-type) cells at the nonpermissive growth temperature. Our results, therefore, suggest that phosphorylation of large T at specific sites correlates with the transforming activity of tsA mutant large T in SV40 N-type and A-type transformants. This conclusion was substantiated by demonstrating that the biological properties as well as the phosphorylation patterns of SV40 tsA28 mutant large T in cells of SV40 tsA28 N-type and A-type transformants were similar to those in FR(tsA58)A (N-type) and in FR(tsA58)57 (A-type) cells, respectively. The phenotype-specific phosphorylation of tsA mutant large T in tsA A-type transformants probably is a cellular process induced during establishment of SV40 tsA A-type transformants, since tsA28 A-type transformant cells could be obtained by a large-T-dependent in vitro progression of cells of the tsA28 N-type transformant tsA28.3 (M. Osborn and K. Weber, J. Virol. 15:636-644, 1975).

Animals↗