Biomedical subjects
Publications and source records attributed to G Brunner.
Enzyme-based artificial organs are being developed as metabolic assist devices. These are required when normal metabolism is impaired, or when the body is overloaded by undesired metabolites or toxins. The implementations of this approach for treating a genetic disease, and for metabolic support in liver failure are envisaged. The kinetic aspects and mass transfer characteristics of bioreactors for these systems are considered in detail.
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A lipophilic hollow-fiber technique was used in the enzymatic sulfation of lipophilic toxins. Endogenous and exogenous toxins, for which glucuronidation reactions already were demonstrated with this technique, were submitted to sulfate transferase reaction as an alternative phase-II detoxification route. Native enzyme was circulated on the external side of a lipophilic hollow-fiber membrane while the toxin-containing media (serum or aqueous solution) were circulated inside the hollow fiber. Sulfation reactions were catalyzed by rabbit liver cytosol, with a specific sulfate transferase activity of 626 nmol/min-mg protein. Clearance of the hollow-fiber module for phenol, p-cresol, paracetamol, 2-aminophenol, and 5-hydroxyindol were determined to be 38.1, 47.2, 2.7, 5.3, and 3.2 pmol/mg protein/h/cm2 hollow-fiber surface, respectively. This technique allows sulfation reactions with crude enzyme preparations over long periods without loss of activity from covalent immobilization and without immunological hazards.
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Many techniques have been developed to support liver function in patients with hepatic failure. Only two of these methods are suited to be used in adequately equipped hospitals under appropriate safety conditions. Exchange of toxin-rich patient plasma against fresh frozen plasma will decrease plasma toxin levels and substitute coagulation factors and proteins. This procedure however should be used in intervals of at least 3-4 days because more frequent application will lead to lung or cerebral complications. Dialysis and hemofiltration will remove water soluble toxins from patient blood as well as excess amino acids which are considered to be precursors of toxic lipophilic metabolites. Addition of certain amino acids to dialysis fluid in physiological concentrations will help to avoid loss of essential amino acids (with approx. normal plasma concentration). In a very few special centers treatment by liver perfusion of human or pavian livers, or by liver transplantation can be performed.
In order to find optimal long term cofactor supply for continuous enzymatic detoxification processes, different resins of varying surface, dipole moment, pore size, and chemical structure were investigated for their adsorptive capacity as well as their desorption behaviour towards various nucleotide based coenzymes. UDPGA, NADPGH, NADH, and SAM were gently shaken with agarose coated resins XAD-12, XAD-8, XAD-7, XAD-4, XAD-2, Dowex 1 X 2 (50-100; 200-400), Dowex 1 X 4 (20-50; 200-400), Dowex 2 X 8, and charcoal until all nucleotide was adsorbed or a saturation of the resins was achieved. High adsorption capacity was not always found to correlate with a steady release of cofactor in desorption experiments. Under this premise the optimal resin-cofactor combinations for long term cofactor supply were found to be XAD-12 for UDPGA, Dowex 2 X 8 for NADPH, Dowex 1 X 4 (20-50) for NADH, and XAD-7 for SAM.
Adsorbents for high-performance affinity chromatography were prepared by bonding proteins and reactive Procion triazine dyes to 3-isothiocyanatopropyl- and 3-aminopropylsilicas. The materials prepared were used successfully in the separation of hydrophobic plasma membrane proteins of cultured human fibroblasts. The data obtained show that the reaction of 3-isothiocyanatopropyltriethoxysilane (ITCPS) with the surface hydroxyl groups of silica yields a new and convenient route to preparing an "activated carrier" that is capable of coupling with potential affinity ligands containing amino functional groups. The reaction and bonding procedures of 3-isothiocyanatopropyltriethoxysilane and 3-aminopropyltriethoxysilane with silica were optimized with regard to a controlled and reproducible ligand density by adjusting the type of solvent and organic base as reaction catalyst. The ligand content of reactive triazine dyes directly coupled to 3-aminopropylsilica was significantly higher than that of the 6-aminohexyl derivatives coupled to 3-isothiocyanatopropylsilica. The stability of Procion Blue MX-R bonded to 3-aminopropylsilica and 3-isothiocyanatopropylsilica in phosphate-buffered aqueous solution at pH 5.0 and 8.0 was examined.
In a multicentre single-blind study, ranitidine was compared to cimetidine as prophylactic treatment against stress-induced upper gastrointestinal bleeding in seriously ill patients in the intensive care unit (ICU). 380 patients entered the study. 192 patients were treated with ranitidine 50 mg q.i.d. as i.v. bolus followed by 150 mg orally twice daily. 188 patients received cimetidine 400 mg q.i.d. intravenously and 1,000 mg daily orally in divided doses. Five patients in the ranitidine group (2.6%) and 12 in the cimetidine group (6.4%) developed gastrointestinal bleeding definitely or possibly due to stress lesions. This difference was not significant. The incidence of stress erosions or ulcerations developing during the study was 11.8% for the ranitidine group and 18.3% for the cimetidine group (non-significant difference). Adverse events in the ranitidine group were nausea, tachycardia or vomiting in 4 patients. 5 cimetidine-treated patients developed cholestasis, and 5 additional central nervous system problems. The high degree of efficacy of both drugs compared very favourably with the high incidence of stress ulceration and hemorrhage in similar untreated populations.
A lipophilic hollow-fiber membrane preparation was used in the enzymatic detoxification of lipophilic toxins. Native enzymes were circulated on one side of the lipophilic membrane, while the toxin-containing media (blood, serum, waste fluids, etc.) were circulated inside the hollow fiber. Lipophilic substances that accumulate in and penetrate the lipophilic membrane were converted by the corresponding enzymes. Phase II detoxifying enzymes converted the lipophilic toxins to hydrophylic compounds that made a rediffusion of these molecules back to the blood impossible. Glucuronidation reactions were catalyzed by a crude preparation of solubilized uridine-5'-diphosphate-glucuronyltransferase (EC 2.1.1.9) with a specific activity of 10.0 nmol/min/mg protein in the presence of uridine-5'- diphosphoglucuronic acid (acceptor: phenol as reference substrate). Clearance rates of phenol, 1- naphthol , 2- naphthol , and p-cresol were measured, respectively, to be 141, 195, 300, and 251 pmol/h/mg protein/cm2 hollow fiber. The advantage of this technique lies in the possibility of using highly active, soluble enzyme preparations and a fully utilizable cofactor supply. The lipid membrane ensures the absence of immunological hazards, while mass transfer of the toxin is not impaired.
The value of the electrophysiological investigations can be summarized as follows: Differentiation of different types of polyneuropathies. Detection of subclinical neuropathies. Approach to etiopathogenical aspects. Verification of therapeutic results. Prognosis. The single questions are discussed.
Incidence of peptic ulcers in hospitalized patients with arterial occlusive disease without history or clinical symptoms of ulcer disease was 32% and four times higher than in patients with acute phlebothrombosis. Possible etiological factors could be: decreased blood perfusion of gastric or duodenal mucosa because of generalized arteriosclerosis, intake of ulcerogenic drugs, chronic abuse of nicotine, and stress due to the underlying arterial occlusive disease. Patients with inflammatory vascular disease are affected as often as are patients with arteriosclerotic disease. Routine endoscopic examination of the upper GI-tract should be performed in such patients in order to exclude potential bleeding sources before treatment with fibrinogen lowering drugs is started.
Recently lipophilic fluid membrane hollow fiber modules have been developed, which allow continuous extracorporal enzymatic detoxification by elimination of endogenous toxins out of the blood in vivo without loss of enzymatic activity and without immunological risks. The clearance for phenol glucuronisation is 6.8 nMol/min/mg enzyme protein/cm2 hollow fiber surface the hydroxylation rate for dodecanic acid using the cytochrome - P-450 system is 0.5 nMol/nMol P450/min/cm2 hollow fiber surface. Detoxification procedures using methyl, sulfat and glutathion transferase are still in an experimental phase. This new enzyme fluid membrane technology in conjunction with modified established techniques like aminoacid dialysis and plasma exchange will make possible the development of partial extracorporal substitution of liver function in the near future.
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N-Acetylneuraminic acid-bearing glycoproteins and lectins isolated from the plasma membrane of GH3 cells are coupled covalently to glass dishes and are used as substrates for cell culture. Under serum-free, hormone-supplemented culture conditions these coupled molecules inhibit the growth rate of GH3 cells. This inhibition is non-toxic and is concentration-and time-dependent. For an inhibition of 50% 2 X 10(6) coupled molecules contacting one cell are needed. The data suggest that specific plasma membrane-located glycoproteins and lectins are involved in the 'density-dependent growth regulation'.
S-methyltransferase was solubilized from pig liver microsomes by treatment with N-dodecyl-N,N-dimethyl-3-ammonio-1-sulfonate (Zwittergent). The soluble enzyme was immobilized by covalent binding to agarose and by copolymerization with acrylamide. The specific activity for the agarose-bound enzyme towards the substrate ethane thiol was 0.87 nmol/min/mg and for the acrylamide-bound enzyme 0.55 nmol/min/mg. The specific activity of the soluble enzyme was found to vary with increasing chain length of the substrate molecules from 0.5 nmol/min/mg for methane thiol (C1) to 6.3 nmol/min/mg for n-heptane thiol (C7). After binding of the enzyme to agarose beads, the increase in specific activity towards substrates with increasing chain length was no longer detectable. Instead, a relatively constant specific activity of 1.1 nmol/min/mg was observed for the whole range of substrates tested from C1 to C7. The stability of the agarose immobilized enzyme at -20 degrees C is twice as good as the soluble enzyme. The acrylamide immobilized enzyme is less stable than the soluble enzyme.
S-methyltransferase was solubilized from pig liver microsomes by five different types of detergents: Triton X-100, zetyltrimethyl-ammonium bromide, sodium cholate, Zwittergent, and sodium dodecylsulfate. As regards enzyme activity, stability, and critical detergent concentration, Zwittergent proved superior to the four other detergents utilized. A striking difference was found in the catalytic activity relative to the chain length of homologous substrates between the microsomal and the solubilized enzyme. The microsomal enzyme preparation showed highest activity towards C1 substrate (methane thiol) while the solubilized enzyme had its maximal activity for C7 substrate. This observation accorded with the active site affinity for the corresponding substrates. From the reaction of the enzyme to the different detergents and the distribution coefficient of substrates in membranes and cytosol, it can be deduced that the enzyme is located on the surface of microsomal membranes with the active site directed towards the cytoplasm.
We entered 174 patients with healed duodenal ulcer and 77 with healed gastric ulcer into a double-blind, placebo-controlled, 6-month trial to investigate the efficacy of 1 g sucralfate twice daily in preventing ulcer recurrence. Endoscopy was performed after 6 months or earlier for symptoms compatible with ulcer disease. The relapse rate in the 126 patients with duodenal ulcer who could be evaluated for efficacy was 14/66 (21.2%) under sucralfate and 30/60 (50%) under placebo treatment (p less than 0.01). No significant difference in relapse rate was found in the 55 gastric ulcer patients who could be evaluated; 11 of 30 (37%) relapsed on sucralfate and 11 of 25 (44%) relapsed on placebo. Among the duodenal ulcer patients in the placebo group, those who had been treated initially with H2-receptor blockers for acute ulcer had significantly more relapses than patients who had been treated with other drugs. The recurrence rate was independent of prior treatment in the sucralfate group. Duodenal ulcer patients with a history of multiple episodes of active ulcer disease had a significantly higher rate of relapse than patients with only a few previous episodes. Both treatments were well tolerated. Two patients in each treatment group complained of nausea and epigastric pain immediately after drug intake. No other drug-related symptoms were observed. We conclude that sucralfate is effective in the prophylaxis of duodenal ulcer. No significant effect was found in the prevention of gastric ulcer recurrence.(ABSTRACT TRUNCATED AT 250 WORDS)