[Need for rehabilitation from the viewpoint of the stoma patient. Experiences of the German ILCO].
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Aftercare↗
Biomedical subjects
H Schröter
Publications and source records attributed to H Schröter.
Macrogolgin--a new 376 kD Golgi complex outer membrane protein as target of antibodies in patients with rheumatic diseases and HIV infections.
Antibodies against the Golgi complex (GC) were found by indirect immunofluorescence microscopy in the serum of two patients with sclerodermia and Sjögren syndrome. Serum from one patient was used to screen clones from an oligo (dT) primed HeLa cDNA expression library. Four overlapping cross hybridizing clones (G1, G12, G13, G14) were found. One additional 5' clone (G15) was retrieved from a random primed lambda gt11 human thyroid cDNA library by nucleic acid hybridization, exploiting sequence information of clone G12. Additional clones for both the 5' and 3' ends were generated by RF-PCR from HeLa cell mRNA. Alignment of the overlapping clones resulted in a consensus cDNA of 10,300 bp encoding a protein of 376 kD. A corresponding mRNA of about 10 kb was found in Northern blots of RNA from various cultured cells. The most distinct features of the protein were the extraordinarily high fraction of alpha-helical domains containing heptad repeats with the probability of forming coiled-coils and the structure similarities with the myosin family and the yeast intracellular transport protein USO1. Five overlapping cDNA fragments covering the entire open reading frame were used to synthesize recombinant proteins for affinity purification of the antibodies in the two patients' sera. By use of these affinity purified antibodies, staining of the GC of various cultured cell lines was reproduced. The antibody target was dissociated within 15 min after brefeldin A exposure of cultured cells, a phenomenon, which was fully reversible within 30 min after withdrawal of the drug. Sucrose step gradient separation of GC enriched microsomal fractions from rat liver showed a natural antigen of about 380 kD co-fractionating with the GC marker galactosyltransferase. KCl extraction, Triton X-114 partition, as well as trypsin digestion of microsomal fractions revealed that the hydrophilic protein has to be located on the cytoplasmatic surface of GC vesicles. Using the five blotted recombinant protein fragments, anti-GC antibodies were found in 18 of 164 (11%) HIV positive patients but in none of the 64 healthy controls. HIV patients as well as the two original patients showed a diverse antibody spectrum recognizing different epitopes of the recombinant proteins. The protein characterized herein, for which we propose the provisional name macrogolgin, constitutes the largest protein known so far associated with the GC.
Amino Acid Sequence↗
The ambiguous effect of ascorbic acid on chromate induced proteinuria in rats.
The influence of ascorbic acid (AA, 5 g/kg body weight) on chromate (Cr, 10 mg/kg) induced proteinuria, which is a sensitive parameter of its nephrotoxicity, was investigated in adult female Wistar rats. The concentrations of Cr and ascorbic acid (AA) were determined in renal tissue. Cr nephrotoxicity is related to its intracellular reduction from Cr(VI) to Cr(III). Proteinuria was completely prevented by enhancement of extracellular reduction of Cr(VI) to Cr(III) followed by rapid renal excretion when Cr and AA were given concomitantly. With an interval up to 1 h between Cr and AA, proteinuria was decreased probably by the radical scavenging function of AA. At an interval of 3 h AA enhanced Cr toxicity by increased intracellular Cr reduction. If the interval was increased to 5 h or if Cr was given 24 h after AA, no influence of AA could be detected. Our results confirm that AA is a very effective reductant of Cr which can influence Cr nephrotoxicity in very high concentrations. It depends on the interval between Cr and AA administration whether or not there is a beneficial effect of AA in Cr nephrotoxicity.
Animals↗
Molecular genetic analyses of a 376-kilodalton Golgi complex membrane protein (giantin)
Molecular genetic analyses of a 376-kDa Golgi complex (GC) membrane protein (giantin) are described. The immunoglobulin G fraction of a human serum containing antibodies against GC antigens as revealed by indirect immunofluorescence microscopy with Hep-2 cells was used to screen a HeLa cDNA expression library, yielding four overlapping cross-hybridizing clones. Additional cDNA clones were retrieved from a lambda gt11 human thyroid cDNA library or generated by reverse transcriptase-mediated PCR from HeLa cell mRNA. Alignment of the clones resulted in a consensus cDNA of 10,300 bp encoding a protein of 376 kDa. The corresponding mRNA with a size of about 10 kb was detected by Northern (RNA) blotting of HeLa, Hep-G2, and Jurkat cell RNA. Sequence analyses of the protein revealed an extraordinarily high content of heptad repeats with the probability of forming coiled coils similar to the proteins of the myosin family. Five overlapping recombinant proteins covering the entire sequence were synthesized and used for antibody production in rabbits and for affinity purification of human and rabbit antibodies. Indirect immunofluorescence experiments also done with brefeldin A-treated Hep-2 and Pt K1 cells revealed an identical GC staining of both the affinity-purified human and rabbit antibodies. Double labeling experiments with antibodies against the GC marker mannosidase II as well as immunoelectron microscopic studies confirmed the localization of the protein within the GC. A corresponding endogenous large-molecular-mass protein of about 390 kDa was found in [35S]methionine-labeled Hep-2 cell lysates as well as in GC-enriched subcellular fractions from rat liver. The protein as well as the recently described proteins golgin-95 and golgin-160 (M. J. Fritzler, J. C. Hamel, R. L. Ochs, and E. K. L. Chan, J. Exp. Med. 178:49-62, 1993) may belong to a new group of Golgi proteins with a high content of heptad repeats which may exert functions in scaffold formation or vesicle transport. As far as can be concluded from immunological and personally communicated partial cDNA sequence data, the protein seems to be identical with a 400-kDa Golgi protein (giantin) recently described (A. D. Linstedt and H. P. Hauri, Mol. Biol. Cell 4:679-693, 1993). Therefore, we agreed to adopt the name giantin.
Amino Acid Sequence↗
[Antibodies against topoisomerase I (Scl-70)--comparative studies on detection with recombinant protein antigens and natural enzyme].
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Antibody Specificity↗
Autoantibodies against topoisomerase I detected with the natural enzyme and overlapping recombinant peptides.
Antibodies against topoisomerase I (anti-topo I, anti-Scl-70) are regarded as a marker of systemic sclerosis. The various frequencies of anti-topo I detected in those patients depends at least in part on the test design and the kind of the antigen used. We therefore analyzed three overlapping recombinant topo I fragments (N-terminal, center and C-terminal part of the molecule) covering the full length of the enzyme for substitution of highly purified natural antigens (n-topo I) in ELISA for antibody screening. 49 of 50 sera reacting with n-topo I in ELISA also recognized the recombinant C-terminal topo I fragment under identical test conditions, 37 sera recognized the recombinant center and two sera the recombinant N-terminal peptide. All sera reactive with the N-terminal and center peptide reacted with the recombinant C-terminus which therefore may substitute for the natural antigen. In immunoblot assays 92% (46/50) of the sera reacted with n-topo I and 86% (43/50) with the recombinant C-terminal peptide. Immunoblots therefore seem to be less sensitive for detecting anti-topo I antibodies than ELISA regardless the source of the antigen used. In a screening of 696 sera submitted for routine antibody tests the recombinant peptide ELISA on the other hand detected two sera which did not react with n-topo I in ELISA. Because of the high rate of agreement within the results obtained with the two antigens, n-topo I can be substituted by the recombinant peptide ELISA allowing better standardization and interlaboratory comparison of results.
Animals↗
Anti-LKM-1 antibodies determined by use of recombinant P450 2D6 in ELISA and western blot and their association with anti-HCV and HCV-RNA.
Several subtypes of anti-liver-kidney microsome antibodies (LKM) are known. LKM-1 antibodies associated with autoimmune chronic active hepatitis recognize P450 2D6, a cytochrome P450 mono-oxygenase. The frequent association of anti-LKM-1 antibodies and hepatitis C virus (HCV) infections and the probable existence of an infectious and autoimmune form of anti-LKM-1-associated hepatitis, requiring different therapeutical strategies, necessitates the exact determination of anti-LKM-1 specificities. Therefore, we compared various antibody tests (immunofluorescence, ELISA with recombinant P450 2D6, and Western blot with recombinant and natural antigens and agargel double diffusion) with sera of 27 anti-LKM-1-positive chronic active hepatitis (CAH) patients, with 61 sera harbouring anti-mitochondrial antibodies, 100 sera each from HCV-RNA-positive and HCV-RNA-negative patients, and 50 sera of healthy persons. Western blot techniques using recombinant MS2-polymerase P450 2D6 fusion protein were found to be the most sensitive and specific method for anti-LKM-1 antibody determination in routine laboratory. The recently recognized association of anti-LKM-1 antibody and HCV infection was confirmed by the results of this study. In anti-HCV and HCV-RNA-positive patients with anti-LKM-1 antibodies there was a preponderance of males with higher mean age and lower antibody titres. The results support the hypothesis of the existence of an autoimmune as well as an infectious (HCV triggered) subgroup of anti-LKM-1-positive hepatitis.
Adolescent↗
Murine cyclophilin-S1: a variant peptidyl-prolyl isomerase with a putative signal sequence expressed in differentiating F9 cells.
Fractionation of differentiating murine teratocarcinoma F9 cells and extraction of the nuclear/microsomal pellets with ethidium bromide led to the purification and microsequencing of the protein mCyP-S1, a novel cyclosporin A-sensitive peptidyl-prolyl cis-trans isomerase (PPIase). mCyP-S1 is a new member of the cyclophilin class of proteins. Cloning and sequencing of the mCyP-S1 cDNA revealed extended coding capacity for a putative N-terminal signal sequence, suggesting processing of mCyP-S1 during intracellular translocation across the membrane of the endoplasmic reticulum. mCyP-S1 is abundantly expressed in a variety of mouse organ tissues and its mRNA levels increase during F9 cell differentiation. Specific subcellular localization of PPIases is postulated to contribute to functional specificities of this class of enzymes.
Amino Acid Isomerases↗
[Neurologic and psychiatric manifestations of lead poisoning in adults (case report and review of the literature)].
A 59-year-old potter presented with lead polyneuropathy after 37 years of occupational exposure. There was a 25-year history of normochromic normocytic anaemia with moderate basophilic stippling, mild renal failure, hyperuricaemia and abnormal porphyrins. Since 1964 three short psychotic episodes were noted. Cranial computed tomography showed extensive bilateral symmetrical calcification in the cerebellar hemispheres and slight calcification in the subcortical area of the cerebral hemispheres and basal ganglia. T2-weighted magnetic resonance imaging disclosed high signal intensities in the periventricular white matter, basal ganglia, insula, posterior thalamus and pons. Differential diagnostic aspects are discussed with special regard to CT and MRI findings. A review of the literature on neurological and psychiatric manifestations of lead intoxication in adults is given.
Brain↗
Synergism in ternary complex formation between the dimeric glycoprotein p67SRF, polypeptide p62TCF and the c-fos serum response element.
Transcriptional regulation of the c-fos proto-oncogene requires the serum response element (SRE) which is complexed by a multi-protein assembly observed both in vitro and in vivo. Two protein factors, p67SRF and p62TCF (previously called p62), are required to interact with the SRE for efficient induction of c-fos by serum. By quantitative band shift electrophoresis we measure at least a 50-fold increase in SRE affinity for p67SRF/p62TCF over p67SRF alone. Stoichiometrically we determine that the ternary complex with p62TCF involves p67SRF in dimeric form. We demonstrate that p67SRF is a glycosylated nuclear transcription factor carrying terminal N-acetylglucosamine (GlcNAc) as a post-translational modification. A proteolytic limit digestion product, approximately 13 kd in size, was generated from the p67SRF-SRE complex. This p67SRF-core domain binds SRE, can dimerize with p67SRF and is still able to form a ternary complex with p62TCF. Therefore, three functional activities can be ascribed to this small p67SRF-core domain: specific DNA binding, dimerization and interaction with p62TCF. We demonstrate that these functions map within the p67SRF core fragment containing the region between amino acids 93 and 222.
Base Sequence↗
Triiodothyronine (T3) increases cisplatin nephrotoxicity in young and adult rats.
The influence of pretreatment with triiodothyronine (T3) on cisplatin (CP)-induced nephrotoxicity was investigated in 10- and 55-day-old rats. Triiodothyronine pretreatment enhanced CP proteinuria in young and adult rats, and increased blood urea nitrogen concentration in 55-day-old rats. As T3 decreased Pt concentrations in renal tissue, the enhanced nephrotoxicity of CP by T3 must have another mechanism. Enhanced CP nephrotoxicity is discussed in connection with an increase of glutathione concentration obtained in renal tissue as a consequence of T3 pretreatment.
Aging↗
Silver-stained protein gels allow recovery of transcription factors still active in specific DNA-binding.
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DNA↗
The ability of a ternary complex to form over the serum response element correlates with serum inducibility of the human c-fos promoter.
Rapid induction of c-fos transcription by serum and phorbol esters requires the serum response element (SRE). The SRE contains a 20 bp element of interrupted dyad symmetry (DSE) that is bound by p67/SRF. We have identified a hitherto unrecognized protein with an apparent size of 62 kd. This novel component, p62, is shown to be an integral but physically separable part of a ternary complex formed with p67/SRF and the SRE. Alone, p62 fails to bind the SRE but requires DSE-bound p67/SRF and sequences both within and outside the DSE for its interaction with DNA. In vivo, the response of the c-fos promoter to serum is severely impaired by mutations that abolish ternary complex formation in vitro.
Base Sequence↗
Pharmacokinetics of platinum cytostatics after cisplatin-related equitoxic dosing to rats.
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Animals↗
Purification of intercalator-released p67, a polypeptide that interacts specifically with the c-fos serum response element.
Incubation of intact nuclei in buffers containing the DNA intercalating drug chloroquine leads to release of proteins that interact with DNA. We demonstrate here that a protein which binds to a motif within the human c-fos promoter, identified as the serum response element (SRE), is quantitatively released from HeLa nuclei, whereas nuclear factor 1 (NF 1) is not. Purification of the SRE binding protein by affinity chromatography to greater than 95% homogeneity allowed us to identify it as a polypeptide of approximately 67,000 daltons. The DNA contacts made by p67, as identified by methylation interference experiments, are indistinguishable from those of the serum response factor described previously.
Binding Sites↗
Dose-response studies on the effects of alpha-, beta-, and gamma-hexachlorocyclohexane on putative preneoplastic foci, monooxygenases, and growth in rat liver.
Alpha-, beta-, and gamma-hexachlorocyclohexane (alpha-, beta-, gamma-HCH) isomers are widespread environmental pollutants; alpha-HCH can cause liver tumors in rats and mice. In the present study we have checked first the tumor-initiating activity of HCH using the appearance of phenotypically altered foci in female rat liver as an end point. Foci were identified by means of the gamma-glutamyltransferase (GGT) reaction and by morphological alterations. No evidence of initiating activity was found. Secondly, we have attempted to determine quantitatively the ability of HCH isomers to promote tumor development. For this purpose growth and phenotypic changes of foci were used as an end point. Rats received a single dose of N-nitrosomorpholine. Then five different doses of each HCH isomer or phenobarbital (PB) (as a positive control) were administered continuously via the diet for 4, 15, and 20 weeks. Both number and size of altered foci were enhanced by doses of 2 to 3 mg/kg or more of the three isomers; in addition, foci phenotypes showed a pronounced shift towards strong expression of GGT and sharp demarcation from the surrounding liver. Based on daily doses the three HCH isomers were approximately equipotent; based on concentrations in liver or adipose tissue, gamma-HCH was severalfold more effective than alpha- and beta-HCH. Thirdly, size and DNA and monooxygenase activities of the liver were determined. All three parameters were enhanced by HCH isomers and PB. However, no strict correlations were found. Rather, at the highest doses tested PB was the most effective inducer of monooxygenases, alpha-HCH was the most potent inducer of liver growth, and all three HCHs were more potent than PB as inducers of focal expansion. Thus, induction of liver growth appears to be associated with foci expansion (tumor promotion); however, neither liver growth nor monooxygenase induction can be used for quantitative predictions of foci expansion by chemical compounds. Finally, no-observed-effect levels were estimated for the parameters studied and are discussed in relation to human exposure.
Adipose Tissue↗
[Desferal treatment of aluminum poisoning; effect on serum concentrations of aluminum (Al), iron (Fe) and copper (Cu)].
In 2 patients with a progressive dialysis encephalopathy and increased serum aluminium concentrations a Desferal-treatment with 500 mg per week was performed and in these cases the kinetics of Al, Fe, Cu was investigated. After the application of Desferal an increase of the serum levels, the ultrafiltrable proportion, the dialysance and the eliminated quantity of aluminium developed. In the course of the treatment in the two patients the aluminium level could be reduced to approximately normal values. Nevertheless one patient died of a severe course of a progressive dialysis encephalopathy. An increase of the copper concentration independent of Desferal-treatment after the dialysator passage remained without influence on the course of concentration of the serum copper in the examination period. A remarkable effect of the Desferal-therapy on the iron metabolism could not be established. Since in the dosage used there did not appear any side effects the Desferal-application is to characterized as a distinct and effective technique for the decrease of the serum-aluminium-concentration in patients undergoing dialysis.
Adult↗
DNA intercalators induce specific release of HMG 14, HMG 17 and other DNA-binding proteins from chicken erythrocyte chromatin.
Chicken erythrocyte nuclei were incubated with DNA intercalating agents in order to isolate from chromatin specific DNA-binding proteins whose binding specificity may be determined by DNA secondary and/or tertiary structure. The intercalating agents ethidium bromide (EtBr) and propidium iodide induce the specific release of high mobility group proteins HMG 14 and HMG 17 under low ionic strength conditions. Chloroquine (CQ) intercalation also results in the selective liberation of HMG 14 and HMG 17, but, in addition, selectively releases other nuclear proteins (including histone H1A) in a pH- and ionic strength-dependent fashion. The use of this new 'elutive intercalation' technique for the isolation and purification of 'sequence-specific' and 'helix-specific' DNA-binding proteins is suggested.
Animals↗