Morphological polarization of human polymorphonuclear leucocytes in response to three different chemoattractants: an effector response independent of calcium rise and tyrosine kinases.
Chemoattractants such as interleukin-8, C5a and N-formylmethionyl-leucyl-phenylalanine induce a cytosolic calcium rise involved in triggering the secretory functions of human polymorphonuclear leucocytes. We studied the possible role of calcium rise in membrane ruffling, actin polymerization, filamentous actin distribution, and morphological polarization, which are all events contributing to chemotaxis. Membrane ruffling was assessed by right-angle light-scatter changes, the cellular content of polymerized actin by fluorescence of bodipy phallacidin, the intracellular distribution of filamentous actin by fluorescence microscopy and image digitization, and morphological polarization by scanning electron microscopy. Pretreatment of polymorphonuclear leucocytes with 50 microM BAPTA/AM, an intracellular calcium chelator, lowered the basal level in cell calcium and inhibited the transient calcium rise stimulated by 2 nM interleukin-8, 2 nM C5a, and 10 nM N-formylmethionyl-leucyl-phenylalanine. However, BAPTA pretreatment of polymorphonuclear leucocytes did not modify membrane ruffling, actin polymerization, filamentous actin distribution, and morphological polarization stimulated by chemoattractants. Downstream effectors may be protein tyrosine kinases. However, the tyrosine kinase inhibitor tyrphostin did not affect the cytoskeletal characteristics elicited by chemoattractants. Taken together, our results suggest that the transductional pathway leading to cytoskeleton organization and morphological polarization of polymorphonuclear leucocytes is different from that leading to secretion.
Lepidi
H
H
Unité INSERM U387, Hôpital de Sainte-Marguerite, Marseille, France.
Zaffran
Y
Y
Ansaldi
J L
JL
Mege
J L
JL
Capo
C
C
D000199:Actins / Q000097:blood / Q000737:chemistry / Q000187:drug effects; D000328:Adult; D002118:Calcium / Q000097:blood*; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D002614:Chelating Agents / Q000494:pharmacology; D002630:Chemotactic Factors / Q000494:pharmacology*; D015936:Complement C5a / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D016209:Interleukin-8 / Q000494:pharmacology; D007700:Kinetics; D008855:Microscopy, Electron, Scanning; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009504:Neutrophils / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000097:blood*; D013481:Superoxides / Q000097:blood
D000199:Actins; D002614:Chelating Agents; D002630:Chemotactic Factors; D016209:Interleukin-8; D013481:Superoxides; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D009240:N-Formylmethionine Leucyl-Phenylalanine; D015936:Complement C5a; D011505:Protein-Tyrosine Kinases; D002118:Calcium