PubMed Health⌕ Search

Biomedical subjects

A Stary

Publications and source records attributed to A Stary.

At least 19 recordsLinked to original sources
PubMed1992

Molecular analysis of DNA junctions produced by illegitimate recombination in human cells.

In a human HeLa derived-cell line carrying permanently a single integrated copy of an SV40 shuttle vector, the transient expression of the SV40 T-antigen led to the production of heterogeneous populations of circular DNA molecules which retained both integrated vector and its surrounding cellular sequences. Comparison between the integrated copy and the linear maps of 80 different plasmids rescued in bacteria suggested that the formation of circular DNA was the result of bidirectional replication from the SV40 origin of replication followed by a single intramolecular joining leading to the cyclization of the replicated molecules. Sequence analysis of 45 recombinational junctions demonstrated that the cyclization occurred via illegitimate recombination process which did not require preferential nucleotide sequence at the joining sites. However, extensive characterization of recombination junctions revealed that the sequences involved in the recombination at each side of the SV40 origin of replication were not randomly distributed, suggesting the presence of regions which were more prone to be involved in the illegitimate recombination process in human cells. Search of common features usually implied in illegitimate recombination in mammalian cells revealed some association of these regions with palindromes, A + T-rich DNA segments, alternating purine/pyrimidine sequences and Alu family repeats.

Antigens, Polyomavirus Transforming↗
PubMed1992

Description of a new amplifiable shuttle vector for mutagenesis studies in human cells: application to N-methyl-N'-nitro-N-nitrosoguanidine-induced mutation spectrum.

In order to analyze the mechanisms of mutagenesis in human cells, we have established a human 293 cell-derived line containing a permanent mutagenesis target, the bacterial lacZ' gene, on an episomal EBV/SV40-based shuttle vector. This plasmid was maintained at a low copy number per cell which rendered it closer to an endogenous gene as compared to the usual transient shuttle vectors. Transient amplification of vectors, inside the host cell due to expression of the SV40 T-antigen, allowed the recovery of a large number of bacterial colonies transformed by plasmids extracted from human cells. Mutations produced in human host cells on the lacZ' locus were easily and rapidly scored and identified in bacteria using the blue/white color assay. Over a 6-month period in culture, we have shown that the lacZ' gene exhibited a low background frequency of point mutations (< 4.8 x 10(-6)). The efficiency of our system for detecting genotoxic-induced mutations was investigated by treating cells with a potent mutagen, the direct alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). A significant increase (< 230-fold) in the frequency of single-base substitutions was observed after MNNG treatment. In total, 63 MNNG-induced independent mutations were characterized. All substitutions but one involved G:C base pairs with 89% being G:C to A:T transitions which is consistent with the MNNG mutagenic specificity already reported in bacteria and mammalian cells. Mutations were distributed along the two strands of the lacZ' gene and there was no obvious influence of either the 5' or the 3' flanking base near the G:C to A:T transition sites. The low spontaneous point mutation frequency on the mutagenesis locus and the ability to detect induced point mutations indicate that this system could be readily used in human mutagenesis studies at the molecular level.

Base Sequence↗
PubMed1992

Coincidence of hepatitis B-virus markers and other sexually transmitted diseases in different STD-risk groups.

To evaluate the prevalence of hepatitis-B-virus (HBV)-markers in STD patients and the significance of promiscuous heterosexual activity as a risk factor for the transmission of HBV, a serological screening was performed in 499 patients, in addition to the routine STD diagnostic programme. Two groups of patients were evaluated: group 1 (120 patients) was drawn from the STD clinic of the Public Health Office (PHO), group 2 (379 patients) from a private STD outpatient clinic. Promiscuous activity was reported significantly more often by persons of group 1 than by those attending the private clinic (59.3% vs. 5.1%). The infection rate of gonorrhea, syphilis and Chlamydia trachomatis was high in patients of the PHO (46.7%, 35.3%, 27.5%) whereas most of the STDs were seldom ascertained in patients of the private clinic (1.1%, 0%, 5.6%). Similar to other STDs, the prevalence of HBV markers differed significantly between patients of the PHO and those of the private clinic (33.3% vs. 6.3%; p = .0000). Comparison of HBV and other STDs showed the highest coincidence of HBV markers in patients with serological evidence of syphilis (44.2%), and in one third of patients with Neisseria gonorrhoeae as well as HIV infection. The data obtained in the present study demonstrate that also in Austria, in addition to homosexual preference and drug abuse, promiscuous heterosexual activity must be considered a substantial risk factor for the transmission of HBV.

Adolescent↗
PubMed1992

Simian virus 40 (SV40) large T antigen-dependent amplification of an Epstein-Barr virus-SV40 hybrid shuttle vector integrated into the human HeLa cell genome.

We analysed the DNA rearrangements that occurred during the integration and amplification of an Epstein-Barr virus (EBV)-simian virus 40 (SV40) hybrid shuttle vector in human cells. The human HeLa cell line was episomally transformed with the EBV-SV40 p205-GTI plasmid. After a 2 month culture in a selective medium, a HeLa cell-derived population (H-G1 cells) was obtained in which the p205-GTI vector was integrated as a single intact copy deleted in the EBV latent origin of replication (OriP). Sequencing data showed that the endpoints of the plasmid sequences, at the plasmid-cell DNA junctions, are located within the two essential elements of EBV OriP, which may form several secondary structures. This result suggests that a specific DNA sequence (OriP) or palindromic structures could play a role in this integration process. This represents the first fully characterized site of integration of an EBV vector in human cells. The transient expression of the SV40 large T antigen in H-G1 cells leads to the appearance of episomal molecules with an extremely heterogeneous size pattern. Individual analysis of these episomes after rescue in bacteria indicated that they retained sequences of both the p205-GTI plasmid and cellular DNA. Comparison of the structure of these circular DNAs with those of the integrated p205-GTI copy indicated that large T antigen expression in human cells leads to the amplification of the integrated shuttle vector according to the 'onion skin' model developed for transformed rodent cells. Indeed, amplified sequences were colinear with the integrated p205-GTI copy and its surrounding cellular sequences, distributed almost equally around the SV40 replication origin, and circularized by illegitimate recombination which did not involve specific nucleotide sequences. This system is of interest in that it enables easy recovery of individual recombined molecules in host bacteria. Each isolated clone contains a unique recombination junction which is easily and rapidly characterized and sequenced.

Antigens, Polyomavirus Transforming↗
PubMed1992

Rapid diagnosis of Chlamydia trachomatis in male patients by antigen detection in urine samples.

To investigate the diagnostic value of testing urine samples as a rapid method for the detection of chlamydial antigen in males, first-catch urine (FCU) and urethral swab samples were obtained from 668 male patients and examined by an enzyme immunosorbent assay (EIA). Positive results were further analyzed by direct fluorescence antibody tests of the EIA sediment. Antigen detection was possible in 12.7% out of the urethra, in 10.8% out of FCU and in a total of 14.5% of the tested persons. Testing only FCU would have missed chlamydia detection in 25 (25.8%) out of a total of 97 chlamydia-positive males. Testing only genital samples would have missed 12 positive cases (12.4%). The sensitivity and specificity of the EIA test of FCU as compared with urethral swabs were 70.6 and 97.9%, respectively, and differed between urine collected before (sensitivity: 84%; specificity: 98.6%) and after (sensitivity: 65%; specificity: 97.7%; p = 0.1254) urethral sampling. The quantitative evaluation of the EIA results demonstrates that the mean value of the extinction rates was highest in specimens corresponding to a positive result from both sampling sites. This study indicates that the chlamydial detection rate was lower in FCU than in urethral samples. FCU testing may be suitable when urethral sampling is not possible; due to its high rate of unconfirmed borderline extinction, positive results should be confirmed with another chlamydial antigen detection test such as direct immunofluorescence.

Adolescent↗
PubMed1992

Quinolones as an alternative treatment of chlamydial, mycoplasma and gonococcal urogenital infections.

We report the results of several trials aimed at evaluating the quinolones in urogenital infections. In Chlamydia trachomatis infections, ofloxacin (200 mg b.i.d. for 10 days) gave a cure rate of 98% (n = 66), and fleroxacin (400 mg s.i.d. for 7 days) provided a cure rate of 89% (n = 19). A double-blind study comparing fleroxacin (600 mg s.i.d.) to doxycycline (100 mg b.i.d.) for 7 days showed similar high cure rates for both regimens (100%; n = 23). In Mycoplasma hominis infections, ofloxacin (200 mg b.i.d. for 10 days) yielded a cure rate of 86% (n = 50) for M. hominis and 55% (n = 43) for Ureaplasma urealyticum. Gonococcal infections (n = 122) were all cured by a single dose of 200 mg ofloxacin. Both ofloxacin and fleroxacin were well tolerated and may be recommended for patients with chlamydial or uncomplicated gonococcal infections, although 600 mg fleroxacin showed a higher incidence of adverse events compared to doxycycline.

4-Quinolones↗
PubMed1992

[DNA repair and related diseases].

DNA-repair is a complex enzymatic process which enables all living cells to withstand the deleterious and mutagenic effects of most genotoxic agents. Defective DNA-repair is caused by mutations involving genes which encode the enzymes responsible for recognition and excision of DNA lesions. Some of these genes have been identified in humans. Several severe human diseases are caused by defective DNA repair affecting the entire genome (e.g. xeroderma pigmentosum and trichotiodystrophy) or only actively transcribed genes (Cockayne's syndrome). Some of these conditions are associated with extremely high rate of cancer.

Cockayne Syndrome↗
PubMed1991

Amplification of Epstein-Barr virus-based shuttle vectors by ultraviolet light in human cells.

In order to approach the mechanism of gene amplification, we have developed a model system in human cells based on the use of episomally-replicating shuttle vectors. Shuttle vectors carrying the replication origin of the Epstein-Barr virus can be stably maintained in human cells. These vectors replicate as an episome with a low copy number. We also constructed hybrid plasmids containing both the EBV and the SV40 replication origins. These molecules are able to replicate episomally either like an EBV vector or like SV40 if the SV40 large T antigen is provided at the same time. UV irradiation of both human adenovirus transformed 293 or SV40-transformed MRC5 host cells leads to vector amplification whatever the type of replication origin used for the episomal maintenance. Our result clearly shows that the EBV latent replication origin (OriP), in the presence of the Epstein-Barr nuclear antigen-1 (EBNA-1) and the SV40 large T antigen, is sensitive to over-replication in UV-irradiated human cells. Since the UV doses were small enough to induce very little damage, if any, on the plasmid sequences, this amplification should be mediated through a cellular factor acting in trans. The interest in using shuttle vectors for this kind of study lays in the easy analysis of the amplified vectors in rescued bacterial colonies. The accuracy of the amplification process can be monitored by studying restriction maps of individual plasmid molecules or more precisely the integrity of a target gene, such as the lacZ' sequence, carried by our vectors.

Cells, Cultured↗
PubMed1991

[Results of virostatic treatment of varicella with various severity].

Chickenpox is the result of primary infection with the varicella zoster virus (VZV), which--like other herpes-viruses--has the ability to remain latent within the nervous system; reactivation then sometimes causes shingles many decades later. While chickenpox is benign in normal children, infection in the immunocompromised patient is characterized by a period of prolonged viral replication, delayed healing and a high frequency of extracutaneous manifestations, such as pneumonitis and involvement of the nervous system. Therefore, current therapeutic research efforts have focused on both the prevention of VZV infections through the development of a live, attenuated vaccine and improved therapeutic modalities. The existing antiviral drug acyclovir has been shown to be effective in reducing severe complications in risk groups. It has been generally accepted that the varicella vaccine is useful in immunocompromised children with acute leukaemia or solid malignant tumours. Healthy seronegative siblings will also benefit from the varicella vaccine. In addition, zoster hyperimmunoglobulin has also been shown to provide protection against primary VCV infection in the incubation period. Preventive and therapeutic efforts should mean that varicella will soon no larger be a medical problem even for immunocompromised patients.

Acyclovir↗
PubMed1991

Detection of Chlamydia trachomatis in first-void urine collected from men and women attending a venereal clinic.

Cervical, urethral and first-void urine (FVU) specimens from 196 men and 245 women attending a venereal outpatient clinic were studied by culture and a commercial enzyme immunoassay (EIA) (CHLAMYDIAZYME). Confirmatory chlamydial testing by a direct fluorescence assay (DFA) (MICROTRAK) was performed on the sediments of the positive EIA samples from culture-negative patients. Chlamydia trachomatis was isolated from 11% of the men and 12% of the women. Of the women, 67% were positive in both sampling sites and 33% in the cervix only. No further cases were found when a female urethral swab was cultured. All the chlamydia-positive urine samples were obtained from women who were positive in the urethra. The denominator used to calculate sensitivities was the combination of patients with culture- and EIA-positive results which could be confirmed by DFA. The sensitivity of our culture method was 85% for men and 77% for women. In men, the sensitivity of EIA was greater on urine than on urethral specimens (77% vs 62%; p less than 0.1). In women, the sensitivity of EIA on urine was significantly poorer than that on cervical specimens (54% vs 85%, p less than 0.001). The specificity of EIA ranged between 94 and 100%. Our study suggests that it may be worth using FVU in a trial for the diagnosis of genital chlamydial infections in symptomatic men, but not in symptomatic women.

Adult↗
PubMed1990

Mechanisms and consequences of mutation induction in mammalian cells.

Mutations have been studied for several decades in order to understand biological processes of great significance and the selection of better-adapted species. Our knowledge both of mutation spectra induced by genotoxic agents and the mechanisms involved in DNA damage processing is more advanced in bacteria than in animal cells. However, the use of new technologies such as shuttle vectors or the polymerase chain reaction will undoubtedly allow rapid progress in the next few years. Shuttle vectors consist of target sequences for monitoring mutagenic activity and additional sequences permitting DNA replication and selection, both in bacteria and in mammalian cells. These plasmids are very efficient in allowing the production of mutation spectra of a particular genotoxin in animal cells. In most cases, base substitutions occur predominantly at the sites of base damage and the type of substitution depends on the kind of damage. This has been well characterized using ultraviolet (UV) light as a mutagen. UV-induced mutations are targeted opposite pyrimidine-pyrimidine sites, where the two major UV lesions are produced. The direct relationships existing between mutation and cancer are exemplified by some hereditary diseases where deficiency in an enzymatic repair system is linked to a high incidence of tumours. Similarly, activation of some cellular proto-oncogenes occurs via specific point mutations. A correlation does exist between the mutation spectra found in model systems and the specific mutation found in the activated oncogene in tumours induced by a given genotoxin. This is particularly well illustrated in the DNA repair deficiency syndrome, xeroderma pigmentosum. The specific mutations found in activated ras oncogenes isolated from UV-stimulated skin tumours correlate well with the mutagenic properties of unrepaired UV-induced DNA lesions.

Animals↗
PubMed1990

[Relevance of routine diagnosis of Chlamydia and HIV infections in Vienna prostitutes].

At the Control Center for Veneral Diseases of the Vienna Health Office, 5000 examinations for Chlamydia infection were performed on registered and non-registered prostitutes between 1980 and 1988. During this period, we observed a continuous decrease of the rate of infections from 20.4% to 4% among registered prostitutes and from 31.4% to 12.4% among non-registered prostitutes. Between 1985 and 1988, the cumulative number of HIV infections only rose from 7 to 8 cases among registered prostitutes and from 2 to 3 cases among non-registered prostitutes. All these HIV carriers were associated with i.v. drug abuse. The correlations between these two infections as well as the implications regarding health politics are discussed.

Antibodies, Bacterial↗
PubMed1989

[Eczema diseases and erythemato-squamous dermatoses of aging skin--studies of the effectiveness and tolerance of Prednicarbate in various vehicles].

In an open multi-center study on 1,286 patients aged 60 years and more, we tested the efficacy and compatibility of prednicarbate (Dermatop) in various vehicles. The eczemas and erythematosquamous dermatoses of 857 patients (66.6%) disappeared within three weeks treatment. In 142 patients (11%), it took more than 3 weeks (62 day at the most) to achieve a complete healing. A very good or good tolerance was observed in 97.1% of the patients. Only in 1.5% the study had to be broken off because of a change for the worse. Prednicarbate, therefore, is especially suitable in the treatment of eczemas and chronic erythematosquamous dermatoses of aged skin.

Aged↗
PubMed1989

[New knowledge regarding herpes zoster].

Zoster is the clinical manifestation of the endogenous reactivation of the varicella-zoster virus. Current observations of viral reactivation emphasize the role of cellular immunity and show an inverse correlation between the specific cellular immune response of the host and the incidence of zoster. Thus, immunocompromised persons like patients with immune deficiency syndrome, lymphoproliferative cancer, or immunosuppressive therapy are at a high risk for the development of disseminated zoster, which may either involve the skin only, or affect more than one organ. During the last few years zoster has been proved a prognostic marker for HIV-positive persons. The incidence of zoster and post-zoster neuralgia increases with advancing age. In young children, immunosuppressive therapy and varicella in utero or during the first year of life are the only risk factors for zoster infection. Prevention of dissemination has been one of the major goals in antiviral chemotherapy of zoster in immunocompromised patients. Among the antiviral drugs available at present, aciclovir has proved especially useful, acting as an inhibitor of viral DNA polymerase. It is well-tolerated and can be applied together with corticoids, analgetics, and retrovir. It is most effective in reducing complications of zoster.

Adult↗
PubMed1989

High recombination rate of an Epstein-Barr virus-simian virus 40 hybrid shuttle vector in human cells.

The stability of an Epstein-Barr virus (EBV)-simian virus 40 (SV40) hybrid shuttle vector, the p205-GTI plasmid, was analyzed in human cells during EBV- or SV40-type replication mode. When the p205-GTI plasmid was maintained as an episomal EBV vector in the human 293 cell line, no rearrangement was detected. To induce the SV40 replication mode, cells containing the episomal p205-GTI plasmid were either transfected with vectors carrying the T antigen gene or infected with SV40. Surprisingly, we observed both production and amplification of different classes of recombinant molecules. Particular types of modifications were found in most of the recombinants. The most striking rearrangement was a duplication of the promoter and enhancer regions of SV40 which was inserted in the thymidine kinase (TK) promoter. This recombination process involved a few bases of homology, and one of the recombination junctions implicated the GC boxes which constitute the essential components of the TK and SV40 early promoters. Our results suggest that a combination of a low level of base homology and a specific DNA sequence function (promoter and enhancer sites) leads to a very high level of recombinational activity during T-antigen-dependent plasmid replication.

Base Sequence↗