Transport of β-lactoglobulin and α-lactalbumin in enterocyte-like Caco-2 cells

Abstract

Summary ― The transport of [ 14 C]-radiolabelled p-lactoglobulin and a-iactaibumin through Caco-2 cell monolayers grown on permeable filters was studied in order to evaluate the different protein pathways through the intestinal epithelium. p-Lactoglobulin or a-lactalbumin (0.25-3 mg/ml) was introduced on the apical side of the monolayer and both the transport and the release of labelled material from the cells were measured following different incubation times. The labelled material was analysed by either trichloroacetic acid precipitation or by high pressure liquid chromatography. Despite some differences between the 2 proteins, the overall mechanism followed approximately the same pattern. Part of the intact internalized protein was either recycled (10-15%) or transported via transcytosis (about 5%). Another pathway corresponded to the intracellular degradation of the protein. The calculation of the different routes followed by the proteins indicated that the main part of the degraded fraction (about 70%) was recycled whereas approximately 30% was transported to the other side. Moreover, 5-10% of the endocytosed material was retained intracellularly. milk protein / protein absorption I intestine / Caco-2 cells Résumé ― Transport de la 5-lactoglobuline et de l'a-lactalbumine dans les cellules de type entérocytaire Caco-2. Afin de quantifier les différentes voies de transport des protéines à travers l'épithélium intestinal, nous avons étudié le transport de la p -lactoglobuline et de l â -lactalbumine, marquées au !4C, à travers des cellules Caco-2 cultivées en monocouches sur des filtres perméables. Après addition de la p-lactoglobuline ou de 1'(x-lactaibumine (0,25 à 3 mglml) dans le compartiment apical, le transport et le relargage des produits radiomarqués ont été mesurés à différents temps d'incubation. Le matériel marqué a été analysé par précipitation à l'acide trichloracétique et par chromatographie liquide haute performance. Malgré quelques différences entre les 2 protéines, leur mécanisme général de transport était comparable. Une partie des protéines intactes intemalisées a été soit recyclée (10-15%) soit «transcytosée» (environ 5%). Les protéines peuvent également être dégradées à l'intérieur des cellules. La quantification des différentes voies indique que les produits de dégrada-tion sont majoritairement recyclés (70%) alors que 30% environ sont transportés vers la face opposée. De plus, 5 à 10% du matériel internalisé est stocké dans les cellules.

β-Lactoglobulin-linoleate complexes: In vitro digestion and the role of protein in fatty acid uptake

“…As linoleate concentration was constant (50 µM linoleate), βlg concentration was three times smaller for a linoleate/βlg molar ratio of 3 compare to a molar ratio of 1. Native βlg has been shown to be transported into the cells (Caillard and Tome, 1995). This could explain why the kinetics of linoleate transport from the medium to the cells is affected by βlg concentration.…”

In vitro digestion methods for assessing the effect of food structure on allergen breakdown

“…It has also been demonstrated that b-Lg from milk is taken up by the duodenal epithelium, possibly by a paracellular route [8]. In vitro studies on confluent cultures of the enterocyte-like cell line Caco-2 have confirmed the endocytic nature of b-Lg uptake, and whilst the majority of the protein was degraded intracellularly, around a third was transported across the cells [9]. The nature of the transported proteins has not been clearly defined, but circumstantial evidence, such as serum IgE, indicates that intact and degraded proteins cross the mucosal barrier.…”

“…To date, no common features of protein molecules or protein molecule fragments have been identified as the major cause of allergy, although some resistance to pepsin digestion is a common property [9]. Resistance to pepsinolysis has been identified using conditions that are far from physiological, as part of the allergenicity risk assessment process for target transgenes.…”

Paracellular Transport of Avidin Saturated or not with Biotinylated Cobalamin through Caco-2 Cell Epithelium Monolayer

“…β-lactoglobulin epithelial transport has been studied using the Ussing chamber [4,23] and the Caco-2 cell line model [24]. β-lactoglobulin absorption undergoes two transcellular pathways.…”

“…The main route is a degradative one, with a lysosomal processing of the protein. The second route allows the transcytosis of the intact protein for less than 10 % of the total transport [24]. These transcellular pathways could explain why its Papp was 4-fold lower than that of saturated avidin.…”