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M Toth

Publications and source records attributed to M Toth.

At least 55 records · Page 3Linked to original sources
PubMed1994

Antagonist-mediated down-regulation of 5-hydroxytryptamine type 2 receptor gene expression: modulation of transcription.

Prolonged exposure to an agonist results in a progressive loss of most G protein-coupled receptors, whereas exposure to an antagonist causes increased receptor response. The 5-hydroxytryptamine (5-HT)2 receptor is down-regulated by agonists but, paradoxically, antagonists can also elicit a decrease in receptor density. Here we show that long term treatment with serotonin or mianserin, an antagonist and antidepressant, results in reduced levels of both the 5-HT2 receptor and its RNA. Antagonist-induced down-regulation requires the presence of the 5-HT2 receptor, it occurs at the level of transcription, and it is mediated by a drug response sequence in the 5' flanking region of the 5-HT2 receptor gene. The effect of mianserin might result, at least in part, from its ability to modulate transcription.

Animals↗
PubMed1993

Glial cell-specific expression of the serotonin 2 receptor gene: selective reactivation of a repressed promoter.

The 5' flanking region of the 5-HT2 receptor gene has been cloned, sequenced and its transcriptional regulatory functions analyzed. The promoter lacks an identifiable TATA motif, and utilizes at least 11 clustered start sites. Promoter function was analyzed by transient assays in rat C6 glioma cells, which were shown to express the endogenous 5-HT2 receptor gene, as well as in rat CREF and human HeLa cells which do not express the endogenous gene. The basal promoter functioned equally well in all three cell lines; and a repression domain, located upstream of the basal promoter, inhibited activity of the promoter in all three cell lines. A far upstream cell specific activator domain restored promoter activity in C6 glioma cells, but did not reactivate the silenced promoter in CREF or HeLa cells. The upstream activator domain, repressor domain and basal promoter functioned in concert to achieve cell type specific expression. The activator domain did not direct C6 glioma cell specific expression in the absence of the repressor domain or in constructs carrying a heterologous basal promoter. These results indicate that glial cell expression of the 5-HT2 receptor gene is achieved through a cell type specific reactivation of a repressed promoter.

Amino Acid Sequence↗
PubMed1993

Detection of Chlamydia trachomatis by the polymerase chain reaction in the cervices of women with acute salpingitis.

OBJECTIVE: Our objective was to determine whether an increased prevalence of Chlamydia trachomatis could be detected by the polymerase chain reaction as opposed to culture in the cervices of women with acute salpingitis. STUDY DESIGN: Endocervical samples from 15 women with laparoscopy-verified acute salpingitis and 20 women seeking medical help for conditions other than pelvic pain were tested for Chlamydia trachomatis with the polymerase chain reaction. The oligonucleotide primer pairs used were specific for a 144 bp region of the major outer membrane protein that contained a single EcoRI endonuclease cleavage site. The detection of a 144 bp band that was cleaved by EcoRI to a 103 bp band denoted a Chlamydia trachomatis-positive sample. Cervical samples were cultured for Chlamydia trachomatis with the use of McCoy cells. The lymphocyte proliferative response to Chlamydia trachomatis elementary bodies was also determined. RESULTS: Nine of the 15 women (60%) with salpingitis had positive results when tested with the polymerase chain reaction for cervical Chlamydia trachomatis. Only two of these women (13%), both of whom had positive results when tested with the polymerase chain reaction, had cultures that were positive for Chlamydia trachomatis (p < 0.02). Among the 20 other women, only two patients with cervicitis had positive cultures for Chlamydia. Those women plus two women with unexplained recurrent abortions had positive polymerase chain reaction test results for Chlamydia trachomatis. A lymphocyte proliferative response to Chlamydia trachomatis was detected in five of eight women with salpingitis, as well as three of the other four patients, all of whom had positive polymerase chain reaction test results; lymphocytes from the remaining women were unresponsive. Follow-up cervical samples were obtained 4 to 6 months after treatment from six of the patients with salpingitis who had positive polymerase chain reaction test results; at that time five had negative polymerase chain reaction test results for Chlamydia trachomatis. CONCLUSION: The polymerase chain reaction appeared to be more sensitive and more rapid than culture in detecting Chlamydia trachomatis in the cervices of women with acute salpingitis. This assay may be of value for the early diagnosis of chlamydial infections.

Acute Disease↗
PubMed1993

Cell-mediated immune response to the recombinant 57-kDa heat-shock protein of Chlamydia trachomatis in women with salpingitis.

Peripheral blood mononuclear cells (PBMC) from 9 of 18 women with laparoscopy-verified salpingitis proliferated in response to recombinant Chlamydia trachomatis 57-kDa heat shock protein (hsp). In contrast, PBMC from 0 of 10 women with cervicitis, 1 of 5 women with recurrent abortions, and 3 (7.1%) of 42 healthy reproductive-age women were responsive to hsp (P < .001). After passage of the hsp through an endotoxin-removing column, PBMC from 6 of 14 additional women with salpingitis were responsive to hsp, while those from 10 controls, including the 3 previously positive women, were negative. PBMC from all patients responsive to the chlamydial hsp were unresponsive to Mycobacterium bovis 65-kDa hsp. PBMC from 6 of the 15 women with a positive hsp-induced lymphocyte response were unresponsive to C. trachomatis elementary bodies. Induction of a cell-mediated immune response to the chlamydial 57-kDa hsp is a common feature of an upper genital tract infection but does not appear to be limited to women with apparent chlamydial infections.

Adult↗
PubMed1993

The role of free radicals in the pathogenesis of amiodarone toxicity.

INTRODUCTION: In vitro and in vivo studies were performed to elucidate the pathogenesis of amiodarone toxicity. METHODS AND RESULTS: Rats were treated with amiodarone alone (500 mg/kg body weight per day) or together with antioxidants (silibinin or MTDQ-DA: 50 mg/kg body weight per day) or with either antioxidant alone. They received amiodarone for 30 days and antioxidant for 33 days (3 days pretreatment). In vitro, amiodarone induced a dose-dependent chemiluminescence signal, which was inhibited by the two dihydroquinolin-type antioxidants (MTDQ-DA, CH 402). Chemiluminometric results from liver homogenate demonstrated that simultaneous treatment with silibinin partially prevented the liver homogenate superoxide anion radical scavenger capacity decreasing effect of amiodarone. Amiodarone treatment caused a significant increase of NADPH and Fe3+ induced lipid peroxidation in the liver microsomal fraction, which antioxidants (silibinin, MTDQ-DA) were unable to prevent. Light microscopy of the lung tissue in amiodarone-treated rats showed accumulation of foamy macrophages with thickening of the interalveolar septa, pneumonitis, and variable interstitial fibrosis. Antioxidant treatment did not prevent these changes. Electron micrographs of lung from amiodarone-treated rats showed lysosomal phospholipoidosis, intralysosomal electron dense deposits, and increased lysosome number and size. In contrast to rats treated with amiodarone alone, those treated with both amiodarone and silibinin had significantly fewer lysosomes (P < 0.01); the lysosome size, shape, and internal characteristics remained the same. Simultaneous treatment with silibinin and amiodarone decreased lysosomal phospholipoidosis compared to amiodarone treatment alone. Simultaneous treatment with MTDQ-DA and amiodarone did not show any beneficial effect. Pulse radiolysis and cobalt 60-gamma (60Co-gamma) radiolysis studies showed that the main free radical product in a reducing environment was a very reactive aryl radical formed after the partial deiodination of the amiodarone molecule. The radiosensitizing effect of amiodarone was also verified in rat liver microsomal preparations using in vivo amiodarone with or without MTDQ-DA pretreatment and 60Co-gamma irradiation with or without the in vitro addition of antioxidants (CH 402, MTDQ-DA). In vivo, the MTDQ-DA treatment also had a radiosensitizing effect; however, the in vitro addition of both antioxidants resulted in a radioprotective effect. The aryl radical also may emerge in vivo during the metabolism of amiodarone. CONCLUSION: These observations suggest that amiodarone in vitro and in vivo generates free radicals that may play a role in the pathogenesis of amiodarone toxicity beside other well-established mechanisms, and antioxidants may have a partial protective effect against amiodarone toxicity.

Amiodarone↗
PubMed1992

Adenovirus DNA replication facilitates binding of the MLTF/USF transcription factor to the viral major late promoter within infected cells.

The activity of the adenovirus major late promoter is substantially increased as the infection proceeds from the early to late phase. To gain insight into the regulation of this promoter, we analyzed protein-DNA interactions by in vivo DMS and DNasel footprinting during the course of adenovirus infection. Little or no protein interaction at promoter sequences was detected early (5 hr) after infection but strong interactions at the major late transcription factor (MLTF/USF) binding site and at the TATA box were evident late (12 hr) after infection. Comparison of in vivo and in vitro footprints revealed that the in vivo interaction late after infection results from binding of the cellular transcription factor MLTF/USF. Nuclear extracts prepared from uninfected cells as well as cells harvested at 5 and 12 hr after infection contained similar levels of MLTF/USF footprint activity, therefore the lack of a detectable interaction early after infection is not due to reduced levels of the factor early in the viral growth cycle. Viral DNA replication was required for MLTF/USF binding at the major late promoter. These results indicate that DNA replication participates in the regulation of adenovirus late gene expression by facilitating the binding of a transcription factor to the major late promoter.

Adenoviruses, Human↗
PubMed1992

Prognostic value of DNA flow cytometry in the locally recurrent, conservatively treated breast cancer patient.

PURPOSE: This study attempted to determine the prognostic value of DNA flow cytometry in the treatment of patients with locally recurrent, conservatively treated breast cancer. METHODS AND MATERIALS: Of 433 patients with clinical stage I and II breast cancer treated with conservative surgery and radiotherapy at Yale-New Haven Hospital before January 1985, 50 patients experienced an ipsilateral breast relapse as a first site of treatment failure. Using standard flow-cytometric techniques, DNA ploidy, DNA index, and S-phase fraction (SPF) were measured for 38 of the 50 (76%) paraffin-embedded specimens available for analysis. RESULTS: At a median postrecurrence follow-up of 5.8 years, the 5-year and disease-free survival rates following ipsilateral breast treatment failure were 48% and 54%, respectively. Sixty-three percent of the recurrent tumors were DNA diploid and 37% were aneuploid. Both DNA ploidy and SPF were statistically significant prognostic indicators for 5-year survival and disease-free survival after local recurrence. The 5-year survival rate of the DNA diploid population was 64%, compared with 15% in the aneuploid population (P < .02). Patients with low SPF (< 12%) experienced an 83% 5-year survival rate, compared with a 24% 5-year survival rate in patients with high SPF (> or = 12%) (P < .03). Ploidy and SPF were combined to define the categories of favorable (diploid, low SPF) and unfavorable (diploid, high SPF or any aneuploid subgroups). Patients in the favorable category experienced an 89% 5-year postrecurrence survival rate and a 100% disease-free survival rate, whereas patients in the unfavorable category had a 24% 5-year survival rate and a 32% disease-free survival rate (P < .01). The flow cytometry as a factor correlated with other clinical parameters previously shown to be of prognostic significance in this patient population. In a multivariate analysis, flow cytometry was a statistically significant and independent prognostic factor for disease-free survival following local recurrence. CONCLUSIONS: DNA ploidy and SPF as measured by currently available flow-cytometric techniques show promise as a tool in determining prognosis for the patient with locally recurrent breast cancer. Implications of these findings with respect to issues of adjuvant systemic therapy at the time of local recurrence are discussed.

Adult↗
PubMed1992

[Atypical fractionation in advanced squamous-cell carcinomas of the head-neck area].

From May 1990 to May 1991, 23 patients with advanced, inoperable squamous cell cancers, clinically staged as III or IV, were treated by unconventional fractionation radiotherapy. Treatment consisted of a continuous hyperfractionated accelerated radiotherapy, delivering a total dose of 55.3 Gy within 17 consecutive days. In ten patients radiation therapy was combined with chemotherapy; 20 mg mitomycin C/m2, administered by intravenous bolus injection on day 5 of treatment. Apart from a confluent mucositis, treatment tolerance was good. Haematological toxicity from mitomycin C was minor and did not require any specific therapy. The mucosal reaction lasted six weeks (median duration) and was not thought to be increased by additional chemotherapy. In twelve of 23 patients a complete remission of the primary tumour was seen, in patients with lymph node metastases there was a complete response in 14 out of 20 patients. After a median follow-up of 18 months, ten of 23 patients have survived (8/23 without evidence of disease). Eleven patients have died due to local tumour progression and one patient died with distant metastases, being without evidence of local tumour. The advantage of this unconventional fractionation, which takes the described short potential tumour doubling time for head and neck cancers into account, is discussed.

Adult↗
PubMed1992

In vivo tumor necrosis factor production in women with salpingitis.

Immune mediated mechanisms might contribute to damage of the fallopian tube in instances of salpingitis. Using a filter paper technique to obtain samples during the surgical procedure, we examined fluids from the reproductive tract organs of seven women with salpingitis and five controls, for evidence of tumor necrosis factor (TNF). TNF, produced principally by macrophages, is a substantial mediator of inflammatory responses. In three women culture-positive for Chlamydia trachomatis, TNF was identified only in those fallopian tubes with visual evidence of disease. Fluids obtained from morphologically normal tubes, as well as from the ovaries and uterus, were negative. In three women with negative fallopian tube cultures but visual evidence of salpingitis, TNF was also identified in fluids from damaged, but not from normal, tubes. Ovarian and uterine fluids of the women were also TNF positive. The last patient, also culture-negative, had TNF only in one affected tube. All five patients in the control group had negative findings at all genital tract sites. Only one patient had TNF in her serum. Thus, localized cell-mediated immune system activation, identified by TNF production, appears to be a typical component of salpingitis.

Bacteria↗
PubMed1991

Increased inducibility of inflammatory mediators from peripheral blood mononuclear cells of women with salpingitis.

To investigate whether immune system activation may contribute to the tissue damage observed in salpingitis, we isolated peripheral blood mononuclear cells and quantitated production of the monocyte activation products tumor necrosis factor-alpha, interleukin-1, and interleukin-6. Unstimulated cells from 7 of 20 women with salpingitis spontaneously released tumor necrosis factor at a concentration greater than 2 SD above the mean value produced by cells from 29 healthy donors. Interferon gamma (200 U/ml) further induced production of tumor necrosis factor from mononuclear cells of 11 women with salpingitis. In contrast, production of tumor necrosis factor by each of 23 other patients who lacked laparoscopic or clinical evidence of salpingitis was similar to that of the controls. In a subset of women whose cells were tested for production of other monokines, three of nine women with salpingitis spontaneously released interleukin-1 but none of the others did so. Four of nine patients with salpingitis also produced interleukin-6, but none of the others did so. None of the monokines were detected in serum from any subject. The results suggest that monocytes from women with salpingitis are primed in vivo and produce inflammatory mediators under conditions where monocytes from other women are poorly responsive. This increased monokine inducibility may contribute to the tubal damage that is the hallmark of salpingitis.

Cells, Cultured↗
PubMed1990

Establishment of de novo DNA methylation patterns. Transcription factor binding and deoxycytidine methylation at CpG and non-CpG sequences in an integrated adenovirus promoter.

The establishment of de novo patterns of DNA methylation in mammalian genomes is characterized by the gradual spreading of methylation, which has been documented to occur across an entire integrated adenovirus genome as well as at the nucleotide level in the integrated late E2A promoter of adenovirus type 2. By applying the techniques of genomic sequencing and dimethylsulfate or DNase I genomic footprinting in vivo, we have now demonstrated that the spreading of methylation in cell lines that carry the late E2A promoter with three in vitro pre-methylated 5'-CCGG-3' sequences initially involves a DNA domain of this promoter that is devoid of bound proteins. Subsequently, methylation further spreads to neighboring regions, and the patterns of complexed transcription factors are altered. Evidence has been adduced that DNA methylation at sequences homologous to the AP-1 and octamer binding factor sites interferes with protein binding. In contrast, the methylation of sequences in the vicinity of but not involving sequences homologous to an AP-2 site still permits the binding of proteins to these sites. It is significant that during the spreading of methylation a few 5'-CG-3' sequences can remain hemimethylated for several cell generations, before they also become methylated in both complements. Moreover, in cell line HE2, the integrated, heavily methylated late E2A promoter has been shown by the genomic sequencing technique to contain 5-methyldeoxycytidine residues, not only in all 5'-CG-3' dinucleotides but also in a 5'CA-3' and a 5'-CT-3' dinucleotide sequence. Hence, 5-methyldeoxycytidine occurs in a silenced mammalian DNA sequence also in dinucleotides other than 5'-CG-3'. This finding raises the question of whether 5-methyldeoxycytidine in non-5'-CG-3' dinucleotides can be maintained in the methylated state during continuous cell propagation.

Adenoviridae↗
PubMed1990

Eukaryotic DNA methylation: facts and problems.

Patterns of DNA methylation in complex genomes like those of mammalian cells have been viewed as indicators of different levels of genetic activities. It is as yet unknown how these complicated patterns are generated and maintained during cell replication. There is evidence from many different biological systems that the sequence-specific methylation of promoters in higher eukaryotes is one of the important factors in controlling gene activity at a long-term level. In general, the fifth nucleotide 5-methyldeoxycytidine can be considered as a modulator of protein-DNA interactions. The degree and direction of this modulation has to be assessed experimentally in each individual instance. The establishment of de novo patterns of DNA methylation is characterized by the gradual non-random spreading of DNA methylation by an essentially unknown mechanism. In this review, some of the general concepts of DNA methylation in mammalian systems are presented, and research currently performed in the authors' laboratory has been summarized.

Animals↗
PubMed1990

Interindividual concordance of methylation profiles in human genes for tumor necrosis factors alpha and beta.

The DNA in mammalian genomes is characterized by complex patterns of DNA methylation that reflect the states of all genetic activities of that genome. The modified nucleotide 5-methyldeoxycytidine (5mdC) can affect the interactions of specific proteins with DNA sequence motifs. The most extensively studied effect of sequence-specific methylations is that of the long-term silencing of eukaryotic (mammalian) promoters. We have initiated studies on the methylation status of parts of the human genome to view patterns of DNA methylation as indicators for genetic activities. In this report, analyses using both restriction enzyme--Southern blotting and the very precise genomic sequencing technique have been done. The genes for tumor necrosis factors (TNF) alpha and beta--in particular, their 5'-upstream and promoter regions--have been investigated in DNA isolated from human lymphocytes, granulocytes, and sperm. The results are characterized by a remarkable interindividual concordance of DNA methylation in specific human cell types. The patterns are identical in the DNA from one cell type for different individuals even of different genetic origins but different in the DNA from different cell types. As an example, in the DNA from human granulocytes of 15 different individuals (ages 20-48 yr, both sexes), 5mdC residues have been localized by the genomic sequencing technique in three identical sequence positions in the 5'-upstream region and in one downstream position of the gene encoding TNF-alpha. The promoter of this gene is free of 5mdC, and TNF-alpha is expressed in human granulocytes. The TNF-beta promoter is methylated in granulocytes from 9 different individuals, and TNF-beta is not expressed. In human lymphocytes, the main source of TNF-beta, the TNF-beta promoter is free of 5mdC residues. All 5'-CG-3' sites studied in the TNF-alpha and -beta genes are methylated in DNA from human sperm. In human cell lines HL-60, Jurkat, and RPMI 1788, the extent of DNA methylation in TNF-alpha and -beta genes has also been studied.

5-Methylcytosine↗
PubMed1990

[The nursing audit: its introduction and development in the University Hospital of the University of São Paulo].

The study reports an experience carried out by the Departamento de Assistência de Enfermagem do Hospital Universitário da USP, the implantation and development of the retrospective auditory process for the evaluation of their activities as an instrument for control and maintenance of the quality of nursing care given the community. It describes the various steps, instruments and reports used, as well as the use of the auditory's findings for reviewing the assistential and educational programs developed by the department.

Brazil↗
PubMed1989

Prostaglandins E and F in cervical mucus of pregnant women.

Cervical mucus concentrations of prostaglandins E (PGE) and F (PGF) were measured by radioimmunoassay in samples collected from 30 pregnant women. The amount of mucus that could be aspirated did not vary significantly at various stages of gestation and was 67 mg on the average. Concentrations of both prostaglandins were lowest in the first trimester and increased significantly during the second trimester. No significant further increase was observed during the third trimester. Serial samples taken from five women at approximately weekly intervals from the 35th to the 36th week onward did not show any significant increase toward term, although several individual samples had very high concentrations, suggesting that the prostaglandins were secreted into the cervical lumen in pulses. During the first and second trimester, the concentrations of PGE and PGF were similar, but during the third trimester PGF levels were significantly higher. The mean concentrations of PGE and PGF in cervical mucus during the second half of gestation were 10 to 20 times higher than their reported levels in amniotic fluid at the same stage, and in the range reported by others to be effective in modulating cervical tissue responses in vitro. Two to 4 hours after intercourse, prostaglandin concentrations in the cervical mucus were found to be about 10- to 50-fold higher than normal. The prostaglandins in cervical mucus may well participate in the gradual changes in the extracellular matrix of the cervix that result in the extensive remodeling of the cervix during pregnancy.

Cervix Mucus↗
PubMed1989

Genomic sequencing reveals a 5-methylcytosine-free domain in active promoters and the spreading of preimposed methylation patterns.

Previous work demonstrated an inverse correlation between methylation at the three 5'-CCGG-3' sequences in positions +24, +6, and -215 relative to the cap site of the late E2A promoter of adenovirus type 2 (Ad2) DNA and its activity. In the study presented here, we used the genomic sequencing method to detect 5-methyl-2'-deoxycytidine (m5dC) residues in 5'-CG-3' sequences other than the 5'-CCGG-3' (Hpa II) sites. The patterns of methylation in all 5'-CG-3' sequences over a region of about 180 base pairs required for gene activity in the late E2A promoter of integrated Ad2 DNA were determined in cell lines that carry this promoter in an active or inactive state. In cell lines HE1 and uc2, the late E2A promoter is active and all thirteen 5'-CG-3' sequences between positions +24 and -160 are unmethylated. In cell line HE2, the same promoter is permanently shut off and all 5'-CG-3' sequences are methylated in both strands. Thus, the inverse correlation is perfect in these cell lines over a region of about 180 base pairs in the late E2A promoter. The same promoter segment was analyzed in cell lines mc23 and mc40, in which a late E2A promoter-chloramphenicol acetyltransferase (CAT) gene construct had been genomically fixed after in vitro 5'-CCGG-3' methylation and subsequent transfection. In cell line mc23, the preimposed methylation pattern was stable and the CAT gene was inactive. Genomic sequencing confirmed the presence of m5dC in the 5'-CCGG-3' sequences and revealed the spreading of methylation to neighboring 5'-CG-3' sequences along the entire promoter. Some of these sites were hemimethylated. In cell line mc40, several of the 120 integrated copies became demethylated in positions +24 and +6, but the promoter was methylated in some of the copies upstream of position -50. Cell line mc40 expressed the CAT gene.

5-Methylcytosine↗
PubMed1989

Aspects of molecular biology and biochemistry of the cardiac renin-angiotensin system.

1. Recent evidence for the existence of extrarenal tissue renin-angiotensin systems has raised the question of whether such a system also exists in the heart. 2. Evidence is presented for a cardiac renin-angiotensin system based on molecular biological and biochemical data. In addition, the question of whether the components of this system interact as a locally integrated, biologically functioning unit is addressed. 3. Using radio-labelled cRNA probes prepared from specific cDNA fragments, renin and angiotensin gene expression in atria and ventricles of the rat heart have been documented by Northern blot and liquid hybridization analysis. Relative signal strength for both mRNAs was highest in the atria, followed by the right and left ventricle. 4. Using specific, h.p.l.c.-controlled RIAs for angiotensin peptides, the presence of both angiotensin I and angiotensin II in all anatomical regions of the monkey and rat heart have been demonstrated; similarly, presence of converting enzyme activity was also ascertained by direct in vitro determinations. 5. Additional experiments evaluating the spontaneous release of angiotensin from rat isolated, perfused hearts revealed a bimodal pattern of high, but rapidly declining rates during the first hour (perhaps representing washout of a pool sequestrated from plasma), followed by a prolonged period of steady, low level release, consistent with the secretion of locally synthesized protein. 6. In separate experiments aimed at examining the possible local integration of the components of the cardiac renin-angiotensin system, angiotensin II concentrations were measured in the coronary sinus effluent of rat isolated, perfused hearts.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗