methylnitronitrosoguanidine and Hemolysis

Although N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and methylmethanesulfonate (MMS) cause injury and malondialdehyde formation in rat hepatocytes, MNNG toxicity is much more sensitive to inhibition by antioxidants. In order to quantify the relationship between toxicity and antioxidation potential, we compared 14 antioxidants that protected against MNNG and MMS toxicity. Chemoprotection was quantified as the concentration that delayed by 1 h the decline in trypan blue exclusion to less than or equal to 50%. While chemoprotection against MNNG and antioxidant efficacy were directly related (R = 0.86), chemoprotection against MMS and antioxidant efficacy were unrelated (R = 0.37). Since we hypothesized that protection against MMS involved stabilization of membranes, the capacity of the 14 compounds to stabilize membranes in an unrelated system (i.e. prevention of erythrocyte osmotic rupture) was assayed. Chemoprotection against both MNNG and MMS correlated with reduced RBC fragility (R = 0.97 and 0.70, respectively). One of the better protecting compounds, 4b,5,9b,10-tetrahydroindeno[1,2-b]indole, was also protective against hepatocellular toxicity mediated by acetaminophen, carbon tetrachloride and tert-butyl hydroperoxide, suggesting a fundamental basis in the mechanism of chemoprotection. We propose that methylating agents and perhaps other chemical toxicants destabilize cellular membranes resulting in hepatocellular injury. For MNNG, radical mediated events may result in membrane destabilization; for MMS, membranes are destabilized without concurrent radical events. The current studies provide a basis for future work to determine structure-activity relationships of chemoprotective agents, examine protection mechanisms, and develop better protective compounds.

Topics: Animals; Antioxidants; Cell Survival; Cells, Cultured; Erythrocyte Membrane; Hemolysis; In Vitro Techniques; Indoles; Lipid Peroxides; Liver; Methyl Methanesulfonate; Methylnitronitrosoguanidine; Rats

Seven beta-hemolytic Streptococcus salivarius isolates produced bacteriocin-like inhibitory activity in deferred antagonism tests using a set of nine indicator bacteria (I1-I9). Five of these S. salivarius strains (KWF, TOVE-R, K17, K21, and K26) were inhibitory to indicators I2, I5, I6, and I7. Mutated non-hemolytic derivatives showed concomitant loss of inhibitory activity against I2, I5, and I6, but retained activity against I7. Inhibitory activity against I2, I5, and I6 was restored in beta-hemolytic revertants of such mutants. Strain 3638 was inhibitory to all of the indicator organisms except I3, and this pattern of inhibitory activity was retained by non-hemolytic derivatives. It appeared that strain 3638 produced an additional broadly-active inhibitory agent, since a mutant (strain 3638A), which was apparently defective in the production of this inhibitor, retained both the beta-hemolytic and I2-, I5-, I6-, and I7-inhibitory activities. Non-hemolytic derivatives of strain 3638A were inhibitory only to I7. Strain 3638, therefore, appeared to produce at least three inhibitory agents: one active only on I7; another acting on I2, I5, and I6 (and associated with beta-hemolytic activity); and a third apparently active on all of the indicators other than I3. S. salivarius strain JH inhibited all nine indicator strains and possessed a beta-hemolytic character which differed from that of the other strains in being readily eliminated on treatment with the plasmid-curing agent novobiocin. Non-hemolytic derivatives of JH retained inhibitory activity against the complete set of indicators.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Bacteriocins; Culture Media; Dialysis; Hemolysis; Methylnitronitrosoguanidine; Novobiocin; Streptococcus

Hemolytic mutants of Lancefield strain SS-95 and ATCC 19615 Streptococcus pyogenes were produced by treatment with N-methyl-N'-nitro-N-nitrosoguanidine. These mutants contained the same levels of streptolysin O, nicotinamide adenine dinucleotidase, deoxyribonuclease, and hyaluronidase. The mutants were deficient in streptolysin S, as was the naturally occurring nonhemolytic Lowry strain. The mutants retained their pathogenicity for mice and, when reisolated from the dead animals, produced the mutant hemolytic pattern.

Topics: Animals; Hemolysis; Methylnitronitrosoguanidine; Mice; Mutagens; Mutation; Streptococcus pyogenes

A fully virulent classical type A strain of Clostridium perfringens was treated during its logarithmic growth phase with 100 mug/ml of N-méthyl-N'-nitro-N-nitrosoguanidine, the bacteria being exposed to the mutagen for 30 min at 37 degrees C in a phosphate buffer adjusted to pH 6.2; after treatment the suspension was streaked on sheep blood agar plates, and colonies that showed an alteration in the theta-hemolysis pattern were selected for isolation. The virulence of two mutants, thus altered in their theta-hemolysis, was studied. One, designated LNG 5, was still capable of killing most of the inoculated guinea pigs in less than 24 h with all the clinical, macroscopic, and bacteriological signs of gas gangrene; however, histological sections showed that tissue damage was not as marked as with the wild strain. On the contrary, the second mutant, labelled LNG 11, was completely avirulent as far as gas gangrene was concerned; indeed, the injection of fluid cultures containing 1 times 10(8) - 10(9)/ml viable bacteria, was not followed by any clinical, bacteriological, or histological signs of gas gangrene. However, strain LNG 11 did give rise to a firm swelling of the inoculated thigh with a corresponding acute inflammatory response of the connective tissue, although the muscle fiber was unaltered. Eventually, this local reaction was followed by necrosis of the skin accompanied by an acute or subacute inflammation with fibroblastic proliferation. These superficial lesions healed spontaneously. They could not be reproduced with crude filtrate alone or with washed bacilli. Strain LNG 11 was therefore considered to be soletly an attenuated strain since, although avirulent as far as gas gangrene is concerned. it is still capable of producing low levels of toxic material. This appears to be the first time that such a strain of C. perfringens type A has been obtained by nitrosoguanidine treatment.

Topics: Animals; Clostridium perfringens; Gas Gangrene; Guinea Pigs; Hemolysis; Male; Methylnitronitrosoguanidine; Muscles; Mutagens; Mutation; Virulence