Wilhelm Ehrlich

Inhibition of the induction of collagenase by interleukin-1 β in cultured rabbit synovial fibroblasts after treatment with the poly(ADP-ribose)-polymerase inhibitor 3-aminobenzamide

Abstract3-Aminobenzamide is an inhibitor of poly-(ADP-ribosyl)ation. In concentrations from 3 to 10 mM it reduced the collagenase activity in culture supernatants of interleukin-1β-stimulated rabbit synovial fibroblasts. 3-Aminobenzoate, not an inhibiter of poly(ADP-ribosyl)ation, had no effect on collagenase activity at a concentration of 10 mM. We concluded that poly(ADP-ribosyl)ation plays a role in the induction of the expression of collagenase and that 3-aminobenzamide can inhibit this process.

Protection from acetaminophen-induced liver damage by the synergistic action of low doses of the poly(ADP-ribose) polymerase-inhibitor nicotinamide and the antioxidant N-acetylcysteine or the amino acid l-methionine

1. An array of therapeutically used analgetic and antirheumatic drugs cause severe liver damage. The present study investigates the hepatoprotective effects of inhibitors of NAD-dependent adenoribosylation reactions and of antioxidants in analgesic-induced hepatic injury. 2. Male NMRI mice were treated PO with 500 mg/kg of acetaminophen, and the activities of both glutamate-oxaloacetate transaminase (GOT) and glutamate-pyruvate transaminase (GPT) were determined in serum. 3. The acetaminophen-induced release of both GOT and GPT from injured liver cells could be inhibited in a dose-dependent manner, when mice were injected additionally either with increasing amounts (from (25 mg/kg to 100 mg/kg i.p.) of the PARP-inhibitor nicotinamide, with increasing amounts (from 25 mg/kg to 100 mg/kg i.p.) of the antioxidant N-acetylcysteine, or with increasing amounts (from 50 mg/kg to 300 mg/kg i.p.) of the amino acid L-methionine. 4. A combination of both nicotinamide and N-acetylcysteine (at the low dose of 12.5 mg/kg i.p. each) results in a complete protection from acetaminophen-induced release of GOT and GPT from injured liver cells. 5. A combination of both L-methionine and N-acetylcysteine or nicotinamide (at the low dose of 12.5 mg/kg IP each) resulted also in complete protection from acetaminophen-induced release of GOT and GPT.

Intracellular metabolism of 4-hydroxynonenal in primary cultures of rabbit synovial fibroblasts.

The intracellular metabolism of 4-hydroxynonenal (HNE), a secondary product of lipid peroxidation and mediator of inflammation, which was found in the joints of patients with rheumatoid arthritis, was investigated in primary cultures of rabbit synovial fibroblasts. A consumption rate of 27.3 nmol/min x 10(6) cells was measured for the cultivated fibroblasts. It could be shown, that 4-hydroxynonenal enters the synovial fibroblasts and is metabolized mainly oxidatively to 4-hydroxynonenoic acid, intermediates of the tricarboxylic acid cycle and water and by formation of the glutathione-HNE adduct. The share of protein-bound HNE was about up to 8% of the total added HNE after 10 min of incubation. All metabolites accumulates intracellularly within the incubation time except of 4-hydroxynonenal itself. An increase of 4-hydroxynonenoic acid could be detected also extracellularly during the intracellular metabolism of 4-hydroxynonenal. Therefore, an involvement of synovial fibroblasts in the secondary antioxidant defense system of the joints during conditions of higher HNE concentrations like rheumatoid arthritis is suggested.

The effect of tryptophan plus methionine, 5-azacytidine, and methotrexate on adjuvant arthritis of rat.

Within the wider framework of our studies on the genesis of rheumatoid arthritis we have investigated the two signal processes in arthritis: adenoribosylation of proteins and DNA methylation. Arthritis can be induced when Freunds complete adjuvant is applied to rats. This form of arthritis can then be reduced or even totally suppressed through the application of several different substances. In the present article we have investigated if the effect of two of these substances, 5-azacytidine and methotrexate can be influenced by the application of tryptophan plus methionine. When applied singly, these latter two substances are known to reduce the formation of arthritis. This effect is intensified by a combination of tryptophan plus methionine. Application of tryptophan plus methionine without methotrexate or 5-azacytidine causes an enhanced development of an adjuvant induced arthritis.

Influence of diet free of NAD-precursors on acetaminophen hepatotoxicity in mice

Recently, we demonstrated the hepatoprotective effects of nicotinic acid amide, a selective inhibitor of poly(ADP-ribose) polymerase (PARP; EC 2.4.2.30) on mice suffering from acetaminophen (AAP)-hepatitis, suggesting that the AAP-induced liver injury involves a step which depends on adenoribosylation. The present study investigates the effects of a diet free of precursors of NAD, the substrate on which PARP acts, in female NMRI mice with AAP hepatitis and evaluates the influence of simultaneous ethanol consumption in these animals. Liver injuries were quantified as serum activities of glutamate-oxaloacetate transaminase (GOT) and glutamate-pyruvate transaminase (GPT). While AAP caused a 117-fold elevation of serum transaminase activities in mice kept on a standard laboratory diet, which was significantly exacerbated by ethanol and inhibited by nicotinic acid amide (NAA), adverse effects were noted in animals fed a diet free of precursors of NAD. In these animals, only minor increases of serum transaminase activities were measured in the presence of AAP, and unlike the exacerbation caused by ethanol in mice on a standard diet, the liver damage was inhibited by 50% by ethanol. A further 64% reduction of hepatitis was observed, when NAA was given to ethanol/AAP-mice. Our results provide evidence that the AAP-induced hepatitis and its exacerbation by ethanol can either be reduced by end-product inhibition of PARP by NAA or by dietary depletion of the enzymes substrate NAD. We see the main application of NAA as for the combinational use in pharmaceutical preparations of acetaminophen in order to avoid hepatic damage in patients treated with this widely used analgesic.

Induction and activation of procollagenase in rabbit synovial fibroblasts after treatment with active oxygen released by xanthine/xanthine oxidase.

Treatment of rabbit synovial fibroblasts with active oxygen (AO) released by xanthine/xanthine oxidase resulted in an induction of procollagenase in these cells in concentrations ranging from 12.5 μg/ml xanthine plus 0.0025 U/ml xanthine oxidase to 50 μg/ml xanthine plus 0.01 U/ml xanthine oxidase. Preceding this there was an accumulation of poly(ADP-ribose) for the same concentration range of xanthine/xanthine oxidase. Furthermore, it was found that AO caused activation of the latent procollagenase to the active enzyme in concentrations ranging from 0.1 μg/ml xanthine plus 0.00002 U/ml xanthine oxidase to 1 μg/ml xanthine plus 0.0002 U/ml xanthine oxidase. It is suggested that poly(ADP-ribosyl)ation participates in the induction of procollagenase by relaxing chromatin. Furthermore, it is proposed that AO activates latent procollagenase under physiological conditions.

Enhancing the Inhibitory Effect of Nicotinamide Upon Collagen II Induced Arthritis in Mice Using N-Acetylcysteine

We could show that both nicotinamide and N-acetylcysteine inhibit collagen induced arthritis in mice. In the present paper, using lower doses of each, we applied combinations of these two substances. We were able to confirm potentiating effects of these combinations. These results may allow new perspectives for the therapy of arthritis to emerge.

The therapeutic effect of a combination of cofpropamine, a caffeine derivative, and cyclophosphamide on the development of adjuvant arthritis of rats and collagen arthritis of mice.

1. Cofpropamine (Cofa), a caffeine derivative that inhibits polyadenoribosylation, enhances the therapeutic effect of cyclophosphamide (CPA) in two animal models of arthritis. 2. The development of adjuvant arthritis of rats is reduced by treatment with 2 x 50 mg/kg IP CPA and 2 x 50 mg/kg IP Cofa. 3. The development of collagen arthritis in mice is prevented by treatment with 12.5 mg/kg IP CPA and 150 mg/kg IP Cofa three times per week.

Synthesis of195mPt radiolabeled cisdiammine (glycolato) platinum (II) of high radionuclidic purity

An experimental method is described for the synthesis of195mPt-radiolabeled cis-diammine (glycolato) platinum (II) (254-S). Ten mg of 95% enriched194Pt was irradiated for 75 h in the hydraulic conveyer of KUR at a thermal neutron flux of ca. 8.15×1013 n·cm−2·s−1, and the195mPt-radiolabeled 254-S was synthesized and purified using HPLC (column: Dichrosorb Diol, elution: 80% CH3CN). The chemical yield was approximately 45%, with chemical purity greater than 98.4%. The radionuclidic purity was nearly 100% and the specific activity, 7.2 MBq·mg−1 254-S.

Synthesis of195mPt radiolabeled (−)-(R)-2-aminomethyl-pyrrolidine (1,1-cyclobutanedicarboxylato)-2-platinum(II) monohydrate using high performance liquid chromatography

An experimental method is described for the synthesis of195mPt-radiolabeled (−)-(R)-2-aminomethylpyrrolidine (1,1-cyclobutanedicarboxylato)-2-platinum (II) (DWA2114R). An amount of 10 mg of 95% enriched194Pt was irradiated for 75 h in the hydraulic conveyer of the KUR at a thermal neutron flux of approximately 8.1×1013 n.cm−2.sec−1 and the195mPt-radiolabeled DWA2114R was purified using HPLC. The chemical yield is higher than 65%, its chemical purity is greater than 99.2%, the radiochemical purity is nearly 100%, and the specific activity is 6.7 MBq. mg−1 DWA2114R.