* Corresponding author. Tel.: #33-1-46835627; fax: #33-1- 46619334. E-mail address: gillian.barratt@cep.u-psud.fr (G. Barratt). Biomaterials 22 (2001) 2967}2979 Relationship between complement activation, cellular uptake and surface physicochemical aspects of novel PEG-modi"ed nanocapsules Vanessa Carla Furtado Mosqueira, Philippe Legrand, Annette Gulik, Olivier Bourdon, Ruxandra Gref , Denis Labarre, Gillian Barratt* Laboratoire de Physico-Chimi, Faculte & de Pharmacie, Universite & de Paris XI Sud, UMR CNRS 8612, 5 rue Jean-Baptiste Cle & ment, Cha L tenay Malabry Cedex 92296, France Centre de Ge & ne & tique Mole & culaire*UPR A2420, Gif-sur-Yvette 91198, France LPBC, UPRESA CNRS 7033, Universite & Pierre et Marie Curie, 4 place Jussieu, Case 318, 75252 Paris Cedex 05, France Departamento de Farma & cia da Escola de Farma & cia, Universidade Federal de Ouro Preto, Rua Costa Sena, 171-Ouro Preto-35400000, Minas Gerais, Brazil Received 24 January 2000; accepted 24 January 2001 Abstract The aim of our work was to examine the relationship between modi"cations of the surface of nanocapsules (NC) by adsorption or covalent grafting of poly(ethylene oxide) (PEG), and changes in their phospholipid (PL) content on complement activation (C3 cleavage) and on uptake by macrophages. The physicochemical characterization of the NC included an investigation of their properties, such as surface charge, size, hydrophilicity, morphology and homogeneity. This is the "rst time that such properties have been correlated with biological interactions for NC, a novel carrier system with a structure more complex than nanospheres. C3 crossed immunoelectrophoresis revealed the reduced activation for NC with longer PEG chain and higher density, although all formulations induced C3 cleavage to a lesser or greater extent. NC bearing PEG covalently bound to the surface were weaker activators of complement than plain PLA [poly(D,L-lactide)] NC or nanospheres (NS). Furthermore, the #uorescent/confocal microscopy of J774A1 cells in contact with NC reveal a dramatically reduced interaction with PEG-bearing NC. However, the way in which PEG was attached (covalent or adsorbed) seemed to a!ect the mechanism of uptake. Taken together, these results suggest that the low level of protein binding to NC covered with a high density of 20 kDa PEG chains is likely to be due to the steric barriers surrounding these particles, which prevents protein adsorption and reduces their interaction with macrophages.  2001 Elsevier Science Ltd. All rights reserved. Keywords: Nanocapsules; Poly(D,L-lactide-co-ethylene oxide) copolymers; Complement activation; Cellular uptake; Physicochemical characterization; Surface properties 1. Introduction When administrated intravenously, conventional col- loidal drug carriers are rapidly cleared from the blood- stream by the mononuclear phagocyte system (MPS), mainly represented by the Kup!er cells of the liver and spleen macrophages [1]. This removal from the circula- tion generally occurs through speci"c recognition by cellular receptors speci"c for plasma proteins bound to the carriers rather than the carriers themselves. The pat- terns of protein absorption that determine colloid recog- nition by macrophages has been extensively studied and is a very useful tool to evaluate the ability of colloids that undergo delayed plasma protein adsorption [2}4] and thus reduced macrophage uptake. In particular, the complement system plays a major role in the immune system recognition of foreign particles [5]. The concept of surface modi"cation of particulate carriers, developed in the last 10 years in order to control the opsonization process, the speci"c and non-speci"c interaction of par- ticulate carriers with MPS and blood components, raises questions about the optimal surface properties of the carrier [1]. These properties have been modi"ed by adsorption or by covalent attachment of hydrophilic polymers at the colloid surface. Surface charge, size, hydrophilicity and the conformation of the polymer chains are other factors that in#uence their interactions with biological media [6}9]. 0142-9612/01/$-see front matter  2001 Elsevier Science Ltd. All rights reserved. PII:S0142-9612(01)00043-6