Laminin and fibronectin treatment leads to generation of dendritic cells with superior endocytic capacity.
BACKGROUND
Sampling the microenvironment at sites of microbial exposure by dendritic cells (DC) and their subsequent interaction with T cells in the paracortical area of lymph nodes are key events for initiating immune responses. Most of our knowledge of such events in human is based on in vitro studies performed in the absence of extracellular matrix (ECM) proteins. ECM in basement membranes and interstitial spaces of different tissues, including lymphoid organs, plays an important role in controlling specific cellular functions such as migration, intracellular signalling and differentiation. The aim of this study was, therefore, to investigate the impact of two abundant ECM components, fibronectin and laminin, on the phenotypical and functional properties of DC and how that might influence DC induced T-cell differentiation.


RESULTS
Human monocyte derived DC were treated with laminin and fibronectin for up to 48 hours and their morphology and phenotype was analyzed using scanning electron microscopy, flow cytometry and real time PCR. The endocytic ability of DC was determined using flow cytometry. Furthermore, co-culture of DC and T cells were established and T cell proliferation and cytokine profile was measured using H(3)-thymidine incorporation and ELISA respectively. Finally, we assessed formation of DC-T cell conjugates using different cell trackers and flow cytometry. Our data show that in the presence of ECM, DC maintain a 'more immature' phenotype and express higher levels of key endocytic receptors, and as a result become significantly better endocytic cells, but still fully able to mature in response to stimulation as evidenced by their superior ability to induce antigen-specific T cell differentiation.


CONCLUSIONS
These studies underline the importance of including ECM components in in vitro studies investigating DC biology and DC-T cell interaction. Within the context of antigen specific DC induced T cell proliferation, inclusion of ECM proteins could lead to development of more sensitive assays.
García-Nieto
Samuel
S
Allergy Research Group, School of Molecular Medical Sciences and Respiratory Biomedical Research Unit, University of Nottingham, Nottingham, United Kingdom.
Johal
Ramneek K
RK
Shakesheff
Kevin M
KM
Emara
Mohamed
M
Royer
Pierre-Joseph
PJ
Chau
David Y S
DY
Shakib
Farouk
F
Ghaemmaghami
Amir M
AM
D002454:Cell Differentiation; D002478:Cells, Cultured; D018920:Coculture Techniques; D016207:Cytokines / Q000032:analysis; D003713:Dendritic Cells / Q000187:drug effects / Q000502:physiology*; D004705:Endocytosis; D005353:Fibronectins / Q000494:pharmacology*; D006801:Humans; D007797:Laminin / Q000494:pharmacology*; D010641:Phenotype; D037182:T-Cell Antigen Receptor Specificity; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology
D016207:Cytokines; D005353:Fibronectins; D007797:Laminin