Motilidade do citoesqueleto de macrófagos e sua relação com o processo de fagocitose estudados através da microscopia de desfocalização

Abstract

Defocusing microscopy, a novel, recently developed, approach on quantitative analysis of bright field optical microscopy images, was used for real-time in vivo observation and quantification of both morphological and dynamical characteristics of the structures formed on the membrane surface of murine bone marrow macrophages. Two distinct types of membrane shape fluctuations were detected and studied: small, random fluctuations, uniformly distributed over all membrane surface at any time, and large, localized, propagating structures, commonly referred as membrane ruffles. Data collected from these fluctuations allowed determination of membrane bending modulus and cytoplasm viscosity of the macrophages. The effects of several drugs and treatments on the behavior of the structures studied were also investigated. The combined use of defocusing microscopy and optical tweezers allowed membrane surface dynamics to be followed closely during phagocytosis of parasites (Leishmania amazonensis). Analysis of the changes in ruffling activity in a large area of membrane around the forming phagosomes indicated that ruffles may be related to the acceleration of the engulfment of the parasites. Investigation of temperature decrease effects on the fluctuations observed and on the phagocytosis of the parasites provided evidence of how these phenomena share the same basic origin, directly related to cytoskeleton dynamics.