IFN-γ-induced increase in the mobility of MHC class II compartments in astrocytes depends on intermediate filaments

Nina Vardjan,Robert Zorec,Mateja Gabrijel,Yolanda De Pablo,Marko Kreft,Milos Pekny,Maja Potokar,Maryam Faiz,Urban Švajger,Matjaž Jeras

doi:10.1186/1742-2094-9-144

Abstract

BackgroundIn immune-mediated diseases of the central nervous system, astrocytes exposed to interferon-γ (IFN-γ) can express major histocompatibility complex (MHC) class II molecules and antigens on their surface. MHC class II molecules are thought to be delivered to the cell surface by membrane-bound vesicles. However, the characteristics and dynamics of this vesicular traffic are unclear, particularly in reactive astrocytes, which overexpress intermediate filament (IF) proteins that may affect trafficking. The aim of this study was to determine the mobility of MHC class II vesicles in wild-type (WT) astrocytes and in astrocytes devoid of IFs.MethodsThe identity of MHC class II compartments in WT and IF-deficient astrocytes 48 h after IFN-γ activation was determined immunocytochemically by using confocal microscopy. Time-lapse confocal imaging and Alexa Fluor546-dextran labeling of late endosomes/lysosomes in IFN-γ treated cells was used to characterize the motion of MHC class II vesicles. The mobility of vesicles was analyzed using ParticleTR software.ResultsConfocal imaging of primary cultures of WT and IF-deficient astrocytes revealed IFN-γ induced MHC class II expression in late endosomes/lysosomes, which were specifically labeled with Alexa Fluor546-conjugated dextran. Live imaging revealed faster movement of dextran-positive vesicles in IFN-γ-treated than in untreated astrocytes. Vesicle mobility was lower in IFN-γ-treated IF-deficient astrocytes than in WT astrocytes. Thus, the IFN-γ-induced increase in the mobility of MHC class II compartments is IF-dependent.ConclusionsSince reactivity of astrocytes is a hallmark of many CNS pathologies, it is likely that the up-regulation of IFs under such conditions allows a faster and therefore a more efficient delivery of MHC class II molecules to the cell surface. In vivo, such regulatory mechanisms may enable antigen-presenting reactive astrocytes to respond rapidly and in a controlled manner to CNS inflammation.

Highlights

In immune-mediated diseases of the central nervous system, astrocytes exposed to interferon-γ (IFN-γ) can express major histocompatibility complex (MHC) class II molecules and antigens on their surface
Fluorescent dextran labels MHC class II–positive late endosomes/lysosomes in WT and Glial fibrillary acidic protein (GFAP)-/-Vim-/- astrocytes Before reaching the cell surface of Antigen-presenting cells (APCs), MHC class II molecules are generally located in late endosomal/lysosomal compartments of APCs [1] with diameters above 200 nm [26,27], which can be labeled by long-term loading of cells with fluorescent dextrans [28]
We determined whether dextran labels late endosomes/lysosomes in WT and GFAP-/-Vim-/- astrocytes and whether these compartments express MHC class II molecules upon IFN-γ stimulation

Summary

Introduction

In immune-mediated diseases of the central nervous system, astrocytes exposed to interferon-γ (IFN-γ) can express major histocompatibility complex (MHC) class II molecules and antigens on their surface. The aim of this study was to determine the mobility of MHC class II vesicles in wild-type (WT) astrocytes and in astrocytes devoid of IFs. Antigen-presenting cells (APCs) are key players in the immune response. In primary astrocyte cultures without neurons, IFN-γ induces expression of surface MHC class II and some APC-specific co-stimulatory molecules [9,10]. Such cultures may be useful for studying antigen presentation at the subcellular level

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