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Inhibition of neutrophil oxidative burst and phagocytosis by meconium
1Division of Maternal-Fetal Medicine, University of Florida College of Medicine, Gainesville, USA.
Objective:
Meconium in amniotic fluid has been associated with an increased prevalence of chorioamnionitis. In an effort to delineate the mechanism of this association, we determined the effect of meconium on the neutrophil's capacity for phagocytosis and microbial killing by oxidative burst in vitro.
Study Design:
Sterile meconium samples were obtained from four fetuses at the time of breech delivery and were then pooled and lyophilized. Neutrophils were purified from whole blood of each of 13 pregnant nonlaboring patients. Phagocytosis and the oxidative burst of neutrophils in the presence and absence of meconium were assessed by single-cell analysis with flow cytometry. Phagocytosis was measured as the mean fluorescence intensity produced after 30 minutes of incubation with fluorescein-labeled Escherichia coli. Oxidative burst was measured as the mean fluorescence intensity resulting from the oxidation of internalized reduced dichlorodihydrofluorescein after 15 minutes of stimulation with phorbol myristate acetate. Oxidative burst was expressed as the neutrophil oxidative index and the net fluorescence intensity. Neutrophil oxidative index was equivalent to the quotient of the mean fluorescence intensity for phorbol myristate acetate-stimulated and unstimulated cells. Net fluorescence intensity was equivalent to the absolute difference between stimulated and unstimulated cells.
Results:
Exposure of neutrophils to light and very light meconium each resulted in significantly lower mean neutrophil oxidative index compared with unexposed controls (3.2 +/- 4.9 and 4.2 +/- 5.9 vs 16.2 +/- 7.5, p = 0.00002 and p = 0.0007, respectively) and significantly lower mean net fluorescence intensity than that of control cells (112 +/- 220 and 188 +/- 294 vs 613 +/- 328, p = 0.0001 and p = 0.005, respectively). Phagocytosis was significantly impaired in the presence of moderate meconium compared with control cells (2239 +/- 393 vs 4645 +/- 2071, p = 0.0001). Light meconium did not significantly affect phagocytosis.
Conclusion:
Meconium has significant effects on neutrophil function in vitro. Both light and very light meconium inhibit the oxidative burst. Moderate meconium inhibits phagocytosis.
Insights
Meconium exposure impairs neutrophil function. Light meconium inhibits the oxidative burst, while moderate meconium impairs phagocytosis, potentially explaining links to chorioamnionitis.
Area of Science:
- Obstetrics and Gynecology
- Immunology
- Neonatal Health
Background:
- Meconium-stained amniotic fluid is linked to increased chorioamnionitis rates.
- Understanding meconium's impact on maternal immune cells is crucial for explaining this association.
Purpose of the Study:
- To investigate the in vitro effects of meconium on neutrophil phagocytosis and oxidative burst capacity.
- To elucidate the mechanisms underlying the association between meconium and chorioamnionitis.
Main Methods:
- Neutrophils were isolated from pregnant patients.
- In vitro assays assessed phagocytosis and oxidative burst using flow cytometry after exposure to varying concentrations of meconium.
- Escherichia coli and phorbol myristate acetate were used for stimulation and measurement.
Main Results:
- Light and very light meconium significantly inhibited neutrophil oxidative burst (p < 0.0007).
- Moderate meconium significantly impaired neutrophil phagocytosis (p = 0.0001).
- Light meconium did not significantly affect phagocytosis.
Conclusions:
- Meconium significantly impacts neutrophil immune functions in vitro.
- Inhibition of oxidative burst and phagocytosis by meconium may contribute to chorioamnionitis development.