Aggregation of human neutrophils by heparin
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Abstract:
Commercial heparin preparations induce the aggregation of human polymorphonuclear neutrophils (PMNs). The minimum aggregating concentration (MAC) of different heparin lots was measured under standardized conditions. It was found to vary between 0.3 and 15 units/ml. Gel filtration of heparin showed that the aggregating activity eluted in a sharp peak with a pattern different from heparin. When heparin was bound to antithrombin III-Sepharose, the aggregating activity eluted totally in the high-affinity fraction. When PMNs are partially aggregated with heparin, further aggregation by N-formyl-methionyl-leucyl-phenylalanine (FMLP) or phorbol myristate-acetate (PMA) is not affected. Pretreatment of PMNs with a low-aggregating heparin fraction, inhibits further aggregation by a high-aggregating heparin fraction. These results suggest that the PMNs have binding sites for heparin different from those for PMA and FMLP. Binding of heparin is not enough for inducing aggregation. Certain structural requirements of the heparin molecule are probably essential for aggregation.
Insights
Commercial heparin preparations cause human polymorphonuclear neutrophils (PMNs) to aggregate. Specific structural features of heparin, not just binding, are essential for this PMN aggregation.
Area of Science:
- Immunology
- Biochemistry
- Pharmacology
Background:
- Commercial heparin preparations are known to induce the aggregation of human polymorphonuclear neutrophils (PMNs).
- The variability in heparin's aggregating activity necessitates understanding the underlying mechanisms.
Purpose of the Study:
- To investigate the specific properties of heparin responsible for inducing PMN aggregation.
- To explore the binding sites and structural requirements of heparin involved in PMN activation.
Main Methods:
- Measurement of the minimum aggregating concentration (MAC) of various heparin lots under standardized conditions.
- Gel filtration chromatography of heparin to isolate fractions with aggregating activity.
- Affinity chromatography using antithrombin III-Sepharose to study heparin-PMN interactions.
- Assessment of PMN aggregation responses to heparin fractions, N-formyl-methionyl-leucyl-phenylalanine (FMLP), and phorbol myristate-acetate (PMA).
Main Results:
- The MAC of different heparin lots varied significantly (0.3–15 units/ml).
- Heparin's aggregating activity eluted as a distinct peak during gel filtration, differing from unfractionated heparin.
- Aggregating activity was concentrated in the high-affinity fraction when heparin was bound to antithrombin III-Sepharose.
- Partial PMN aggregation by heparin did not affect subsequent aggregation induced by FMLP or PMA.
- Pre-incubation with low-aggregating heparin fractions inhibited aggregation by high-aggregating fractions, suggesting specific binding sites.
Conclusions:
- Human PMNs possess distinct binding sites for heparin, separate from those for FMLP and PMA.
- Heparin binding alone is insufficient for inducing PMN aggregation; specific structural characteristics of the heparin molecule are critical.
- These findings elucidate the mechanism of heparin-induced PMN aggregation and highlight the importance of heparin's structure.