Immunotargeting of glucose oxidase: Intracellular production of H2O2 and endothelial oxidative stress

Andrew J. Gow, Frank Branco, Melpo Christofidou-Solomidou, Linda Black-Schultz, Steven M. Albelda, Vladimir R. Muzykantov

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

Extracellular and intracellular reactive oxygen species attack different targets and may, therefore, result in different forms of oxidative stress. To specifically study an oxidative stress induced by a regulated intracellular flux of a defined reactive oxygen species in endothelium, we used immunotargeting of the H2O2-generating enzyme glucose oxidase (GOX) conjugated with an antibody to platelet-endothelial cell adhesion molecule (PECAM)-1, an endothelial surface antigen. Anti-PECAM-125I-GOX conjugates specifically bind to both endothelial and PECAM-transfected cells. Approximately 70% of cell-bound anti-PECAM-125I-GOX was internalized. The cell-bound conjugate was enzymatically active and generated H2O2 from glucose. Use of the fluorescent dye dihydrorhodamine 123 revealed that 70% of H2O2 was generated intracellularly, whereas 30% of H2O2 was detected in the cell medium. Catalase added to the cells eliminated H2O2 in the medium but had little effect on the intracellular generation of H2O2 by anti- PECAM-GOX. Both H2O2 added exogenously to the cell medium (extracellular H2O2) and that generated by anti-PECAM-GOX caused oxidative stress manifested by time-and dose-dependent irreversible plasma membrane damage. Inactivation of cellular catalase by aminotriazole treatment augmented damage caused by either extracellular H2O2 or anti-PECAM-GOX. Catalase added to the medium protected either normal or aminotriazole-treated cells against extracellular H2O2, yet failed to protect cells against injury induced by anti-PECAM-GOX. Therefore, treatment of PECAM-positive cells with anti-PECAM- GOX leads to conjugate internalization, predominantly intracellular H2O2 generation and intracellular oxidative stress. These results indicate that anti-PECAM-GOX 1) provides cell-specific intracellular delivery of an active enzyme and 2) causes intracellular oxidative stress in PECAM-positive cells.

Original languageEnglish
JournalAmerican Journal of Physiology - Lung Cellular and Molecular Physiology
Volume277
Issue number2 21-2
Publication statusPublished - Aug 1999
Externally publishedYes

Fingerprint

Glucose Oxidase
Cell Adhesion Molecules
Oxidative Stress
Blood Platelets
Endothelial Cells
Amitrole
Catalase
Reactive Oxygen Species
CD31 Antigens
Enzymes
Surface Antigens
Fluorescent Dyes
Endothelium
Cell Membrane

Keywords

  • Bioconjugation
  • CD31
  • Drug delivery
  • Hydrogen peroxide
  • Platelet-endothelial cell adhesion molecule-1

ASJC Scopus subject areas

  • Pulmonary and Respiratory Medicine
  • Cell Biology
  • Physiology
  • Physiology (medical)

Cite this

Gow, A. J., Branco, F., Christofidou-Solomidou, M., Black-Schultz, L., Albelda, S. M., & Muzykantov, V. R. (1999). Immunotargeting of glucose oxidase: Intracellular production of H2O2 and endothelial oxidative stress. American Journal of Physiology - Lung Cellular and Molecular Physiology, 277(2 21-2).

Immunotargeting of glucose oxidase : Intracellular production of H2O2 and endothelial oxidative stress. / Gow, Andrew J.; Branco, Frank; Christofidou-Solomidou, Melpo; Black-Schultz, Linda; Albelda, Steven M.; Muzykantov, Vladimir R.

In: American Journal of Physiology - Lung Cellular and Molecular Physiology, Vol. 277, No. 2 21-2, 08.1999.

Research output: Contribution to journalArticle

Gow, AJ, Branco, F, Christofidou-Solomidou, M, Black-Schultz, L, Albelda, SM & Muzykantov, VR 1999, 'Immunotargeting of glucose oxidase: Intracellular production of H2O2 and endothelial oxidative stress', American Journal of Physiology - Lung Cellular and Molecular Physiology, vol. 277, no. 2 21-2.
Gow, Andrew J. ; Branco, Frank ; Christofidou-Solomidou, Melpo ; Black-Schultz, Linda ; Albelda, Steven M. ; Muzykantov, Vladimir R. / Immunotargeting of glucose oxidase : Intracellular production of H2O2 and endothelial oxidative stress. In: American Journal of Physiology - Lung Cellular and Molecular Physiology. 1999 ; Vol. 277, No. 2 21-2.
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