Biomedical subjects

G Stein

Publications and source records attributed to G Stein.

At least 253 records · Page 14Linked to original sources

PubMed1988

Native genomic blotting: a novel approach to mapping DNase I hypersensitive sites and protein-DNA interactions at high resolution.

We have developed a new high resolution method for screening 400-600 base pairs of DNA in chromatin for DNase I hypersensitive sites and protein-DNA interactions. By separating the DNA isolated from nuclease-digested nuclei in small, native polyacrylamide gels prior to electroblotting onto nylon membranes, we increased the resolution by greater than 3-fold as compared with the traditional approach whereby the nuclease-digested DNA is fractionated electrophoretically in agarose gels (11). In addition, our native genomic blotting method has the advantage of combining the ability of the traditional agarose approach to detect DNase I hypersensitive sites, with the genomic sequencing method (2), where individual protein-DNA contacts can be observed. Native genomic blotting therefore permits for the first time the display of DNase I hypersensitive sites and protein-DNA interactions at high resolution on the same autoradiograph. This method allows us to investigate a new level of chromatin structure and to therefore obtain better insight into levels of gene structure, organization and gene regulation.

Autoradiography↗

PubMed1988

[Drug-induced hepatitis after dihydralazine treatment with fatal consequences].

Adverse drug side effects of the antihypertensive substance dihydralazine have been increasingly observed in recent years. Damage is concentrated on the liver and is characterized by drug-induced hepatitis with confluent necrosis. Three cases (60 year old woman, 79 year old woman, 61 year old man) with lethal course after antihypertensive treatment, using dihydralazine are reported in this paper.

Aged↗

PubMed1987

Differing patterns of human protooncogene expression in peripheral blood and bone marrow acute leukemia cells.

The levels of protooncogene RNA in matched bone marrow and peripheral blood cells obtained from patients with newly diagnosed acute myelogenous leukemia were compared. While the absolute amounts of c-myc RNA in the matched specimens are similar, the levels are not correlated. In contrast, while the levels of c-fos RNA in the matched bone marrow and peripheral blood cells are correlated, the absolute levels of c-fos RNA differ substantially. The level of histone H3 RNA is higher in bone marrow cells than in peripheral blood cells. These substantial differences in protooncogene RNA levels between leukemic cells found in the bone marrow and in the peripheral blood make it impossible to accurately "characterize" gene expression in leukemic cells if studies are restricted to the cells in either compartment. Additionally, there appears to be a significant relationship between the levels of c-fos RNA and triose phosphate isomerase RNA and the height of the white blood cell count and between the level of c-fos RNA in marrow cells and the proportion of monocytic cells present.

Bone Marrow Cells↗

PubMed1987

Protein-DNA interactions in vivo upstream of a cell cycle-regulated human H4 histone gene.

Cell cycle-dependent histone genes are transcribed at a basal level throughout the cell cycle, with a three- to fivefold increase during early S phase. Protein-DNA interactions in the 5' promoter region of a cell cycle-regulated human H4 histone gene have been analyzed at single-nucleotide resolution in vivo. This region contains two sites, with four potential protein-binding domains, at which the DNA is protected from reaction with dimethyl sulfate in cells and from digestion with deoxyribonuclease I in nuclei. These protein-DNA interactions persist during all phases of the cell cycle and dissociate with 0.16 to 0.2M sodium chloride.

Base Sequence↗

PubMed1987

Coordinate turnover of nuclear and cytoplasmic histone messenger RNA following inhibition of DNA replication in HeLa S3 cells.

We have examined the metabolism of human H4 histone mRNA in the nucleus and cytoplasm of HeLa S3 cells following inhibition of DNA synthesis to address the extent to which histone mRNA stability in these cellular compartments is coupled to DNA replication. The nuclear and cytoplasmic levels of histone mRNAs encoded by the pF0108A human H4 histone gene were determined by S1 nuclease analysis using a 32P-labeled probe that could distinguish pF0108A transcripts from those of other members of the H4 histone multigene family. Hydroxyurea treatment resulted within 15 min in a 75% reduction in the level of histone H4 mRNA in the nucleus, which corresponds to the 85% decrease observed for H4 histone mRNA in the cytoplasm. The kinetics of nuclear and cytoplasmic H4 mRNA turnover following hydroxyurea treatment were also similar. Northern blot analysis using a 32P-labeled mitochondrial cytochrome b probe indicated that the association of cytoplasmic RNA with the nuclear fraction was less than 0.5%. Treatment of cells with a protein synthesis inhibitor resulted in a 1.3-fold increase in nuclear H4 histone mRNA levels and a 1.5-fold increase of H4 mRNA in the cytoplasm after 45 min. Together, these results indicate that nuclear and cytoplasmic H4 histone mRNAs respond similarly to metabolic perturbations that influence message stability and that mechanisms operative in the turnover of histone mRNAs in the nucleus and cytoplasm may be similar.

Cell Cycle↗

PubMed1987

Aluminium induced damage of the lysosomes in the liver, spleen and kidneys of rats.

The influence of repeated aluminium (Al) administration (0.05 or 0.5 mg 100 g-1 b.w.t. i.p. 5 times weekly for 12 weeks) on the lysosomal enzymes N-acetyl-beta-D-glucosaminidase (beta-NAG) and beta-glucuronidase (beta-Gluc) in serum, liver, spleen and kidneys of adult female rats with intact kidneys, (NR), or following partial nephrectomy (5/6 NX) was investigated. After A1 loading, at the high dose only, the beta-NAG in serum and the free beta-NAG in liver, spleen and kidneys increased. Latent beta-NAG levels decreased in all three organs the effect being dose related. Following A1 loading no elevation in total enzyme activity was observed, with one exception. Depending on A1 doses the spleen of the non-operated animals, the liver of both groups of animals and the serum showed a decrease in beta-Gluc activity. No effect on beta-Gluc activity was observed in the spleen of 5/6 NX animals or in the kidneys of either group of animals. The results confirm that high doses of Al induce toxic effects and damage the lysosomes in the liver, the spleen and the kidneys. The results indicate that the extent of lysosomal damage correlates with dose and duration of Al loading. Repeated administration of Al also interferes selectively with enzyme synthesis.

Acetylglucosamine↗

PubMed1987

Phosphocreatine and adenine nucleotides in postasphyxial hearts with normal basal function and normal oxygen demand.

We investigated whether there is a relationship between the prolonged dysfunction after myocardial ischaemia and the postischaemic phosphocreatine overshoot phenomenon. In 16 open-chest rats 3 periods of 4 minutes of oxygen deficiency were performed and basal haemodynamic variables and the myocardial oxygen demand were determined during the recovery period. At the end of the 20 minutes recovery period, left ventricular pressure, dp/dtmax, ejection fraction, and myocardial oxygen demand were completely recovered. High energy phosphate levels, however, were still altered. The sum of adeninenucleotides was decreased to 78 +/- 4% of control (mean +/- SEM, p less than 0.05). The level of phosphocreatine was markedly elevated to 162 +/- 14 (mean +/- SEM). The persistence of the phosphocreatine overshoot phenomenon, while basal function was already normalized, indicates that a reduced function and thus a reduced energy demand of the contractile apparatus are not the cause of the phosphocreatine overshoot. We found no close relationship between high energy levels and basal function or oxygen demand in myocardium after mild oxygen deficiency.

Adenine Nucleotides↗

PubMed1987

A rat histone H4 gene closely associated with the testis-specific H1t gene.

A rat histone H4 gene closely associated with the testis-specific H1t gene was isolated by screening the Sargent-Bonner rat genomic library using cloned human histone genes as probes. Both the H4 gene and the H1t gene are located on a 7-kb EcoRI genomic DNA fragment. Although the deduced amino acid sequence of the rat H4 histone is identical to that of the sequence of human histone H4, the nucleotide sequence of the coding region differs significantly from the coding region of the human H4 gene. Moreover, the relative spacing between the 5'-consensus sequence elements is unique for an H4 gene. S1-nuclease protection analyses reveal that both the H4 and H1t mRNA species are present in a fraction of rat testis cells highly enriched in pachytene spermatocytes, while only the H4 mRNA species is present in a rat myeloma cell line (Y3-Ag1.2.3). During a 1-h hydroxyurea treatment of the Y3 cells, which produces a 99% inhibition of DNA synthesis, the level of this H4 mRNA drops by only 50%, indicating that the stability of this mRNA is only partially coupled with DNA synthesis.

Amino Acid Sequence↗

PubMed1987

Histone gene expression remains coupled to DNA synthesis during in vitro cellular senescence.

Despite a decrease in the extent to which confluent monolayers of late compared to early passage CF3 human diploid fibroblasts can be stimulated to proliferate, the time course of DNA synthesis onset is similar regardless of the in vitro age of the cells. A parallel and stoichiometric relationship is maintained between the rate of DNA synthesis and the cellular levels of histone mRNA independent of the age of the cell cultures. Furthermore, DNA synthesis and cellular histone mRNA levels decline in a coordinate manner after inhibition of DNA replication by hydroxyurea treatment. These results indicate that while the proliferative activity of human diploid fibroblasts decreases with passage in culture, those cells that retain the ability to proliferate continue to exhibit a tight coupling of DNA replication and histone gene expression.

Cell Survival↗

PubMed1987

Proximal and distal regulatory elements that influence in vivo expression of a cell cycle-dependent human H4 histone gene.

We have examined the sequences required in vivo to promote transcription of a cell cycle-regulated human H4 histone gene. Deletion mutants of the 5' flanking region were assayed in mouse cells or fused with the chloramphenicol acetyltransferase (CAT) gene for assay in HeLa cells. The functional limits of the regulatory sequences were shown to extend at least 6.5 kilobases (kb) upstream. Sequences sufficient for correctly initiated transcription were found in the 70 base pairs (bp) immediately 5' to the cap site. A proximal element located 200-400 bp upstream increased the level of transcription several times above the basal level, although not to maximal levels. Maximal levels of expression were achieved with 6.5 kb of 5' flanking sequence adjacent to the proximal promoter sequences or when a distal enhancer element with both position- and orientation-independent function was moved proximal to the promoter. Our results indicate that a series of 5' cis-acting sequences are functionally related to the fidelity and level of expression of this human H4 histone gene.

Animals↗

PubMed1987

Cloning and expression of cDNA for human poly(ADP-ribose) polymerase.

cDNAs encoding poly(ADP-ribose) polymerase from a human hepatoma lambda gt11 cDNA library were isolated by immunological screening. One insert of 1.3 kilobases (kb) consistently hybridized on RNA gel blots to an mRNA species of 3.6-3.7 kb, which is consistent with the size of RNA necessary to code for the polymerase protein (116 kDa). This insert was subsequently used in both in vitro hybrid selection and hybrid-arrested translation studies. An mRNA species from HeLa cells of 3.6-3.7 kb was selected that was translated into a 116-kDa protein, which was selectively immunoprecipitated with anti-poly (ADP-ribose) polymerase. To confirm that the 1.3-kb insert from lambda gt11 encodes for poly(ADP-ribose) polymerase, the insert was used to screen a 3- to 4-kb subset of a transformed human fibroblast cDNA library in the Okayama-Berg vector. One of these vectors [pcD-p(ADPR)P; 3.6 kb] was tested in transient transfection experiments in COS cells. This cDNA insert contained the complete coding sequence for polymerase as indicated by the following criteria: A 3-fold increase in in vitro activity was noted in extracts from transfected cells compared to mock or pSV2-CAT transfected cells. A 6-fold increase in polymerase activity in pcD-p(ADPR)P transfected cell extracts compared to controls was observed by "activity gel" analysis on gels of electrophoretically separated proteins at 116 kDa. A 10- to 15-fold increase in newly synthesized polymerase was detected by immunoprecipitation of labeled transfected cell extracts. Using pcD-p(ADPR)P as probe, it was observed that the level of poly(ADP-ribose) polymerase mRNA was elevated at 5 and 7 hr of S phase of the HeLa cell cycle, but was unaltered when artificial DNA strand breaks are introduced in HeLa cells by alkylating agents.

Carcinoma, Hepatocellular↗

PubMed1987

Targeting of a chimeric human histone fusion mRNA to membrane-bound polysomes in HeLa cells.

The subcellular location of histone mRNA-containing polysomes may play a key role in the posttranscriptional events that mediate histone mRNA turnover following inhibition of DNA synthesis. Previously, it has been shown that histone mRNA is found primarily on free polysomes that are associated with the cytoskeleton. We report here the construction of an Escherichia coli pBR322 beta-lactamase signal peptide-human H3 histone fusion gene. The fusion transcript is targeted to membrane-bound polysomes and remains stable following interruption of DNA replication. Relocating mRNA within the cell may provide a procedure for studying the posttranscriptional regulation of gene expression.

Cell Membrane↗

PubMed1987

Comparison of enalapril and thiazide diuretics in the elderly hypertensive patient.

One hundred seventy-four patients, 65 years of age or older, entered a double-blind, seven-center, 16-week, controlled study to compare the effects of enalapril and hydrochlorothiazide (HCTZ) in an elderly hypertensive population. Sixty-eight percent of the patients were men, 32% were women. Thirty-two percent of the patients had isolated systolic hypertension. Approximately 80% of the patients were white. After a 4-week placebo run-in period, patients with sitting diastolic BP (DBP) of 90-120 mm Hg or systolic BP (SBP) greater than or equal to 160 mg Hg and DBP less than 90 mm Hg (isolated systolic hypertension) were randomized to receive 10 mg of enalapril or 12.5 mg of HCTZ once daily. If after 4 weeks their BPs were not controlled (i.e., DBP greater than 85 mm Hg or SBP greater than 140 mm Hg), the dose of the drug was doubled. If after successive 4-week intervals, their BPs were still not controlled, the other drug was added to their regimen and this was then doubled. The initial mean BPs were 167/94 mm Hg in both groups. By the end of the monotherapy phase at 8 weeks, the mean BPs had fallen significantly (p less than or equal to 0.01) to 148/85 mm Hg in both groups. By 16 weeks, the mean BPs had again fallen similarly: in the enalapril group to 144/83 mm Hg, and in the HCTZ group to 145/83 mm Hg. Seventy-nine percent of the enalapril group and 85% of the HCTZ group had controlled BPs at this time (DBP less than or equal to 85 or SBP less than or equal to 140 mm Hg). White and nonwhite patients in both drug groups had similar falls in SBP and DBP both at the end of the monotherapy period and the overall study. The white patients experienced more rapid falls in BP with enalapril, the nonwhite patients with HCTZ. Three serious adverse experiences occurred in the enalapril group, none of which were considered likely to be due to the drug therapy. Overall, 49% of the enalapril group and 61% of the HCTZ group reported an adverse effect during the study (not significant). Laboratory adverse effects occurred 10% more frequently in the HCTZ group: enalapril, 22%; HCTZ, 32% (not significant); none was serious. Both drugs therefore appeared to be equally efficacious antihypertensive agents in these elderly patients.

Aged↗

PubMed1987

[Serum protein determinations in patients with chronic glomerulonephritis and their mathematical-statistical analysis in relation to suitability for renal immunogram].

Different immunological parameters (immunoglobulins, complement, lysozyme, and others) were tested in the sera of 65 patients with two forms of chronic glomerulonephritis (mesangiopathic and membranoproliferative GN). The results were compared with values received from 30 healthy human beings. All the data were analyzed carefully by different statistical methods to find out parameters available for a renal immunogram. Out of 14 parameters 4 were indispensable to the GN-groups: alpha 2-macroglobulin, beta-lipoprotein, alpha 1-acid-glycoprotein, lysozyme.

Acute-Phase Proteins↗

PubMed1987

The influence of selected urinary constituents on the adhesion process of Escherichia coli to human uroepithelial cells.

In the pathogenesis of infectious diseases, great importance is attached to the problem of adhesion of bacteria to cells. In 100 urine specimens from normal test persons, patients with infections of the lower urinary tract, with chronic pyelonephritis and glomerulonephritis were studied. The adhesion of strains of Escherichia coli to human uroepithelial cells depended on the concentration of single urinary factors. While increased concentrations of urea and creatinine favored the adhesion process, a statistically significant negative influence was found with regard to potassium, immunoglobulins and pH value. Considering the multifactorial effects of the urinary constituents, we found in a multivariate comparison that none of the studied physiological features alone exerts a fundamental influence upon adhesion, but in their entirety they determine the environmental conditions for the adhesion of bacteria to cells in the urine.

Adult↗

PubMed1987

Pharmacokinetics and pharmacodynamics of high dose furosemide in patients with chronic renal failure or nephrotic syndrome.

The pharmacokinetics and pharmacodynamics of repeated oral administration of furosemide were studied in patients with chronic renal failure or nephrotic syndrome using three different dosage regimens (4 x 40, 4 x 80 and 4 x 250 mg/d). Concentrations of the unchanged drug in serum and urine were measured fluorometrically. The elimination kinetics of furosemide was linear over the dose range studied. In contrast, the bioavailability of this diuretic decreased with increasing doses. The diuretic effects of all three dosage regimens were distinct only on the first day of therapy, thereafter decreasing rapidly. It can be concluded that the hormonal regulation of water and electrolyte balance are responsible for this behavior rather than pharmacokinetics. Creatinine and urea nitrogen excretion were not influenced by any furosemide dosage regimen.

Adult↗

PubMed1987

[Significance of parathyroid hormone (PTH) within the scope of central nervous system disorders in hemodialysis patients].

The role of PTH as possible uraemic toxin within the scope of disturbances of the central nervous system (progressive dialysis encephalopathy, PDE) was investigated in 88 patients undergoing haemodialysis. A radioimmunoassay covering the C-terminal PTH fragment was used. Patients undergoing haemodialysis with a PDE showed the highest values with 2,015.4 +/- 457.9 pg/ml, and also in the preclinical stage of a PDE the PTH values with 1,845.7 +/- 663.1 pg/ml lay significantly above those ones of the patients undergoing haemodialysis without PDE (794.8 +/- 364.7 pg/ml). The findings speak for the importance of PTH in the development of complications of the central nervous system within the scope of the uraemia syndrome.

Adult↗

PubMed1986

Histone gene stability during cellular senescence.

The extent to which human histone gene organization is conserved during the in vitro lifespan of human diploid fibroblast-like cells was determined by comparing the restriction patterns of a human H4 and an H3 histone gene from cells of various in vitro ages. No age related change in the organization of these two genes was detected.

Aging↗