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A flow cytometric method for measurement of hydrogen peroxide generation by pediatric polymorphonuclear leukocytes
T Ihara1, K Yamawaki, T Fujiwara
1Department of Pediatrics, Mie National Hospital, Japan.
Insights
Infants have a reduced ability to generate hydrogen peroxide (H2O2) when exposed to Staphylococcus aureus and Escherichia coli. This impaired immune response in infants may increase their susceptibility to infections.
Area of Science:
- Immunology
- Microbiology
- Pediatrics
Background:
- Reactive oxygen species, including hydrogen peroxide (H2O2), are crucial for immune responses.
- Polymorphonuclear leukocytes (PMN) are key immune cells involved in combating bacterial infections.
- Infant immune systems are still developing, potentially impacting their ability to fight pathogens like Staphylococcus aureus and Escherichia coli.
Purpose of the Study:
- To investigate the capacity of infant polymorphonuclear leukocytes (PMN) to generate hydrogen peroxide (H2O2) in response to bacterial stimulation.
- To compare H2O2 generation in infants of different age groups with adult levels.
- To explore the role of serum factors in modulating PMN H2O2 production in infants.
Main Methods:
- Cytofluorography was used to measure H2O2 generation by PMN.
- Heparinized whole blood from infants and adults was incubated with Staphylococcus aureus and Escherichia coli.
- H2O2 production was assessed after 60 minutes of incubation.
- Experiments involved adding infant serum to adult PMN and adult serum to infant PMN.
Main Results:
- Infants under 1 year showed significantly reduced H2O2 generation by PMN when stimulated by S. aureus and E. coli.
- H2O2 generation increased with age, reaching adult levels in children aged 10-15 years.
- Infant serum decreased H2O2 generation in adult PMN, while adult serum enhanced it in infant PMN.
Conclusions:
- Infants exhibit a lower capacity for H2O2 generation by PMN against S. aureus and E. coli.
- This reduced immune function in infants may contribute to their increased susceptibility to these bacterial infections.
- Serum factors may play a role in the age-related differences in PMN function.
Abstract:
The generation of hydrogen peroxide (H2O2), one of the reactive oxygen species, by polymorphonuclear leukocytes (PMN) stimulated by Staphylococcus aureus and Escherichia coli was studied in infants by cytofluorography. After heparinized whole blood was incubated with bacteria for 60 min, generated H2O2 was measured. The positive rate of H2O2 generation of PMN and mean fluorescent intensity of positive PMN stimulated by S. aureus and E. coli were significantly reduced in infants aged < 1 year and H2O2 generation increased with advancing age. In 10-15 year old children, the level of generated H2O2 reached adult levels. When sera from 1 year old children were added to separated PMN from healthy adults, H2O2 generation was reduced. In contrast, H2O2 generation by PMN from 1 year old children was increased by the addition of adult sera. These results suggest that the ability to generate H2O2 in response to S. aureus and E. coli is lower in infants and that such reduced activity may be related to the susceptibility of such infants to S. aureus and E. coli infections.