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Pentoxifylline inhibits actin polymerization in human neutrophils after stimulation by chemoattractant factor
G Freyburger1, F Belloc, M R Boisseau
1Laboratoire d'Hémobiologie, Hôpital Cardiologique, Pessac-Bordeaux, France.
Abstract:
Pentoxifylline (PTX) has been recently reported to stimulate PMN chemotaxis under dense agarose. The present study was designed to characterize the effect of PTX on actin polymerization before and after stimulation by the chemotactic factor f-MLP- We used two different methods to determine the proportion of actin in the filamentous form: SDS-polyacrylamide gel electrophoresis to study the Triton X-100 insoluble cytoskeleton, and flow cytometry using fluorescent Rhodamine-Phalloidin to study actin conformation. PTX (10(-3) M) did not affect the amount of F-actin (polymerized G-actin) incorporated into the cytoskeleton, but reduced total F-actin in a dose-dependent manner, at all concentrations of f-MLP used. Moreover, this inhibitory effect appeared more clearly in PMN with the higher activation ratios. Thus F-actin is only partially incorporated into the cytoskeleton, and PTX-induced reduction of non-incorporated actin may reduce the stiffness of activated PMN. This could explain the increased chemotaxis of PMN across the small holes of dense agarose.
Insights
Pentoxifylline (PTX) reduces total filamentous actin (F-actin) in neutrophils (PMN) without affecting cytoskeletal incorporation. This reduction in non-incorporated actin may explain PTX
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- Pentoxifylline (PTX) has been shown to enhance polymorphonuclear neutrophil (PMN) chemotaxis.
- The precise mechanism by which PTX influences PMN function, particularly actin dynamics, remains to be fully elucidated.
Purpose of the Study:
- To investigate the effect of Pentoxifylline (PTX) on actin polymerization in neutrophils (PMN).
- To characterize the influence of PTX on actin dynamics before and after stimulation with formyl-methionyl-leucyl-phenylalanine (f-MLP).
Main Methods:
- Quantification of filamentous actin (F-actin) using SDS-polyacrylamide gel electrophoresis of the Triton X-100 insoluble cytoskeleton.
- Flow cytometry with Rhodamine-Phalloidin to assess actin conformation and polymerization status.
- Evaluation of PTX effects across various f-MLP concentrations and PMN activation levels.
Main Results:
- Pentoxifylline (PTX) at 10(-3) M did not alter the incorporation of F-actin into the cytoskeleton.
- PTX significantly reduced total F-actin in a dose-dependent manner across all tested f-MLP concentrations.
- The inhibitory effect of PTX on total F-actin was more pronounced in highly activated PMN.
Conclusions:
- F-actin is only partially incorporated into the neutrophil cytoskeleton.
- PTX-induced reduction of non-incorporated actin may decrease the stiffness of activated PMN.
- This modulation of actin dynamics by PTX could underlie the observed enhancement of PMN chemotaxis through restrictive environments.