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Respiratory burst facilitates the digestion of Escherichia coli killed by polymorphonuclear leukocytes
Abstract:
We examined factors that may limit degradation of bacterial protein of Escherichia coli S15 killed by polymorphonuclear leukocytes (PMN). Both human and rabbit PMN degraded up to 40% of [14C]amino acid-labeled protein of ingested and killed E. coli in 2 h as determined by loss of acid-precipitable radioactivity. In contrast, equally bactericidal broken-PMN preparations or isolated granules degraded only about 10% of bacterial protein regardless of pH. To determine whether activation of the respiratory burst contributes to digestion, we compared degradation by intact PMN in room air and under N2. Depletion of O2 by N2 flushing had no effect on the bactericidal activity of either human or rabbit PMN but reduced degradation by approximately 50%. Protein degradation during phagocytosis was also reduced in the presence of cyanide or azide, inhibitors of myeloperoxidase (MPO). PMN of two patients with chronic granulomatous disease ingested and killed E. coli S15 as well as did normal PMN but degraded bacterial protein as did normal PMN incubated under N2. The low degradative activity of PMN disrupted by sonication could be raised to nearly the level of intact PMN incubated in room air by preincubation of the PMN with 10(-7) M formyl-methionyl-leucyl-phenylalanine (fMLP) before sonication and by pretreatment of E. coli with MPO. Depletion of O2 or chloride during these preincubations with formyl-methionyl-leucyl-phenylalanine respectively, virtually abolished and markedly diminished stimulation of bacterial protein degradation. We conclude that enhanced MPO-mediated O2 metabolism of intact PMN plays a role in the digestion of killed E. coli.
Insights
Polymorphonuclear leukocytes (PMN) degrade bacterial protein from Escherichia coli. Myeloperoxidase (MPO)-mediated oxygen metabolism by intact PMN enhances this protein digestion, particularly during phagocytosis.
Area of Science:
- Immunology
- Microbiology
- Cell Biology
Background:
- Polymorphonuclear leukocytes (PMN) are key immune cells involved in clearing bacterial infections.
- The degradation of bacterial components by PMN is crucial for effective pathogen clearance.
- Factors influencing the efficiency of bacterial protein degradation by PMN require further investigation.
Purpose of the Study:
- To identify factors limiting the degradation of Escherichia coli protein by human and rabbit PMN.
- To elucidate the role of the respiratory burst and myeloperoxidase (MPO) in bacterial protein digestion.
- To compare the degradative capacity of intact PMN versus disrupted PMN preparations.
Main Methods:
- Quantification of [14C]amino acid-labeled E. coli protein degradation by intact and disrupted PMN.
- Assessment of protein degradation under varying oxygen conditions (room air vs. N2) and in the presence of enzyme inhibitors (cyanide, azide).
- Evaluation of PMN from patients with chronic granulomatous disease and stimulation assays using formyl-methionyl-leucyl-phenylalanine (fMLP) and MPO.
Main Results:
- Intact human and rabbit PMN degraded up to 40% of labeled E. coli protein, while broken PMN preparations degraded only ~10%.
- Oxygen depletion (N2 flushing) reduced protein degradation by approximately 50%, indicating a role for the respiratory burst.
- Inhibition of MPO activity and impaired respiratory burst function (chronic granulomatous disease) significantly reduced bacterial protein degradation.
Conclusions:
- Enhanced myeloperoxidase (MPO)-mediated oxygen metabolism by intact PMN plays a significant role in the digestion of killed E. coli.
- The respiratory burst and MPO activity are critical for efficient bacterial protein degradation during phagocytosis.
- Intact PMN possess a superior capacity for bacterial protein digestion compared to disrupted preparations.