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1.
J Exp Med ; 176(3): 819-28, 1992 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-1512545

RESUMO

We describe a quantitative assay of transendothelial migration (TEM) that allows us to selectively study the interaction of monocytes with confluent human endothelial cell (HEC) monolayers. The HEC are grown on hydrated collagen gels; the monocytes need not be purified. 100% of monocytes transmigrated the monolayer within 1 h at 37 degrees C and accumulated in the subendothelial collagen; TEM of lymphocytes was not detected within this time. Migration of neutrophils from the same donor was much slower and incomplete, with only 14% of PMN transmigrating in 2 h. This rapid TEM occurs in the absence of exogenous chemoattractants, and HEC in this system do not express cytokine-inducible leukocyte adhesion molecules. A slight modification of the TEM assay allowed us to separate binding to the apical HEC surface from TEM. We found that tight apical surface binding was the rate-limiting step for TEM. Two-thirds of this binding and TEM could be blocked by a monoclonal antibody against the leukocyte beta 2 integrin chain CD18. This assay will allow us to dissect the mechanisms of both the binding and transmigration stages of diapedesis.


Assuntos
Endotélio Vascular/citologia , Monócitos/citologia , Ciclo Celular , Divisão Celular , Movimento Celular , Células Cultivadas , Colágeno , Humanos , Solubilidade
2.
J Exp Med ; 178(2): 449-60, 1993 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-8340753

RESUMO

Platelet/endothelial cell adhesion molecule 1 (PECAM-1; CD31) is crucial to the process of leukocyte transmigration through intercellular junctions of vascular endothelial cells. A monoclonal antibody to PECAM, or recombinant soluble PECAM, blocks transendothelial migration of monocytes by 70-90%. Pretreating either the monocytes or the endothelial junctions with antibody blocks transmigration. If the endothelium is first activated by cytokines, anti-PECAM antibody or soluble recombinant PECAM again block transmigration of both monocytes and neutrophils. Anti-PECAM does not block chemotaxis of either cell type. Light and electron microscopy reveal that leukocytes blocked in transmigration remain tightly bound to the apical surface of the endothelial cell, precisely over the intercellular junction. Thus, the process of leukocyte emigration can be dissected into three successive stages: rolling, mediated by the selectin class of adhesion molecules; tight adhesion, mediated by the leukocyte integrins and their endothelial cell counter-receptors; and now transmigration, which, based on these studies, requires PECAM-1.


Assuntos
Antígenos de Diferenciação Mielomonocítica/fisiologia , Moléculas de Adesão Celular/fisiologia , Quimiotaxia de Leucócito/fisiologia , Endotélio Vascular/citologia , Anticorpos Monoclonais/imunologia , Adesão Celular , Células Cultivadas , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/ultraestrutura , Humanos , Interleucina-1/farmacologia , Microscopia Eletrônica de Varredura , Monócitos/citologia , Monócitos/ultraestrutura , Neutrófilos/citologia , Neutrófilos/ultraestrutura , Molécula-1 de Adesão Celular Endotelial a Plaquetas , Proteínas Recombinantes , Fator de Necrose Tumoral alfa/farmacologia
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