Effects of endocytosis inhibitors on internalization of human IgG by Caco-2 human intestinal epithelial cells

Sato Koya

Nagai Junya

Mitsui Naoko

Yumoto Ryoko

Takano Mikihisa

Life Sciences

Aims: The purpose of this study was to characterize the internalization mechanism of human IgG into the epithelial cells of human small intestine. employing human intestinal epithelial cell line Caco-2 as an in vitro model system.

Main methods: Real-time PCR analysis and uptake studies of fluorescein isothiocyanate-labeled IgG (FITC-IgG) from human serum were performed using Caco-2 cells.

Key findings: Real-time PCR analysis showed that mRNA level of the neonatal Fc receptor (FcRn) was increased during the differentiation process in Caco-2 cells. The binding of FITC-labeled human IgG to the membrane surface of Caco-2 cells increased with a decrease in pH of incubation buffer. The uptake of FITC-IgG was also stimulated at acidic pH and was time-dependent. The binding and uptake of FITC-IgG at pH 6.0 was partially, but significantly, decreased by human gamma-globulin in a concentration-dependent manner. A mixture of metabolic inhibitors (sodium azide and 2-deoxyglucose) significantly inhibited the uptake, but not the binding, of FITC-IgG. In addition, endosomal acidification inhibitors such as bafilomycin A, and chloroquine significantly increased the accumulation of FITC-IgG. Clathrin-dependent endocytosis inhibitors (phenylarsine oxide and chlorpromazine) and caveolin-dependent endocytosis inhibitors (nystatin and indomethacin) did not decrease the uptake of FITC-IgG at pH 6.0. In contrast, macropinocytosis inhibitors such as cytochalasin B and 5-(N-ethyl-N-isopropyl) amiloride significantly decreased the uptake of FITC-IgG at pH 6.0.

Significance: The internalization of human IgG in human intestine might be, at least in part, due to FcRn-mediated endocytosis. which could occur by a process other than clathrin- and caveolin-dependent mechanisms.

IgG

FcRn

Endocytosis

Caco-2 cells

Intestine

Medical sciences [ 490 ]

eng

Pergamon

Elsevier Science Ltd

Copyright (c) 2009 Elsevier Inc. All rights reserved.

[ISSN] 0024-3205

[DOI] 10.1016/j.lfs.2009.10.012

[NCID] AA00717011

[DOI] http://dx.doi.org/10.1016/j.lfs.2009.10.012