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Oxygen free radicals induced release of lysosomal enzymes in vitro

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Summary

The effect of oxygen free radicals, generated by xanthine and xanthine oxidase, was studied on the release of lysosomal hydrolase from rat liver lysosomes in vitro. A lysosomal enriched subcellular fraction was prepared, using differential centrifugation technique, from the homogenate of rat liver. The biochemical purity of the lysosomal fraction was established by using the markers of different cellular organelles. Oxygen free radicals were generated in vitro by the addition of xanthine and xanthine oxidase. The release of lysosomal hydrolase (β-glucuronidase) from the lysosomal fraction was measured. There was a 3 to 4 fold increase in the release of β-glucuronidase activity in the presence of xanthine and xanthine oxidase when compared to that in the absence of xanthine and xanthine oxidase. In the presence of superoxide dismutase (SOD), a scavenger of oxygen free radicals, the xanthine and xanthine oxidase system was unable to induce the release of β-glucuronidase activity from the lysosomes. Sonication (2 bursts for 15 sec each) and Lubrol (2 mg/10 mg lysosomal protein) treatment, which are known to cause membrane disruption, also induced the release of β-glucuronidase from lysosomal fraction. This release of β-glucuronidase by sonication and lubrol treatment was not prevented by SOD. These data indicate that lysosomal disruption is a consequence of oxygen free radicals, generated by xanthine and xanthine oxidase.

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Abbreviations

HEPES:

N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid

EGTA:

Ethylene Glycol Bis-(β-aminoethyl ether)N,N,-N′,N′-tetracetic acid

Tris:

Tris (hydroxymethyl) aminomethane

SOD:

Superoxide Dismutase

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Authors and Affiliations

  1. Department of Pathology, College of Medicine, University of Saskatchewan, S7N 0W0, Saskatoon, Saskatchewan, Canada

    Jawahar Kalra, Dave Lautner, K. Lorne Massey & Kailash Prasad

  2. Department of Physiology, College of Medicine, University Hospital, S7N 0W0, Saskatoon, Saskatchewan, Canada

    Jawahar Kalra, Dave Lautner, K. Lorne Massey & Kailash Prasad

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  1. Jawahar Kalra
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  2. Dave Lautner
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  3. K. Lorne Massey
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  4. Kailash Prasad
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Kalra, J., Lautner, D., Massey, K.L. et al. Oxygen free radicals induced release of lysosomal enzymes in vitro . Mol Cell Biochem 84, 233–238 (1988). https://doi.org/10.1007/BF00421058

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