Development of an Advanced Intestinal in Vitro Triple Culture Permeability Model To Study Transport of Nanoparticles

Abstract

Intestinal epithelial cell culture models, such as Caco-2 cells, are commonly used to assess absorption of drug molecules and transcytosis of nanoparticles across the intestinal mucosa. However, it is known that mucus strongly impacts nanoparticle mobility and that specialized M cells are involved in particulate uptake. Thus, to get a clear understanding of how nanoparticles interact with the intestinal mucosa, in vitro models are necessary that integrate the main cell types. This work aimed at developing an alternative in vitro permeability model based on a triple culture: Caco-2 cells, mucus-secreting goblet cells and M cells. Therefore, Caco-2 cells and mucus-secreting goblet cells were cocultured on Transwells and Raji B cells were added to stimulate differentiation of M cells. The in vitro triple culture model was characterized regarding confluence, integrity, differentiation/expression of M cells and cell surface architecture. Permeability of model drugs and of 50 and 200 nm polystyrene nanoparticles was studied. Data from the in vitro model were compared with ex vivo permeability results (Ussing chambers and porcine intestine) and correlated well. Nanoparticle uptake was size-dependent and strongly impacted by the mucus layer. Moreover, nanoparticle permeability studies clearly demonstrated that particles were capable of penetrating the intestinal barrier mainly via specialized M cells. It can be concluded that goblet cells and M cells strongly impact nanoparticle uptake in the intestine and should thus be integrated in an in vitro permeability model. The presented model will be an efficient tool to study intestinal transcellular uptake of particulate systems.

A 3D co-culture of three human cell lines to model the inflamed intestinal mucosa for safety testing of nanomaterials

“…Regarding the intestinal epithelium, i.e. after oral ingestion, most of the models focus on co-cultures to study absorption, particle celltranslocation or particle-mucus interaction (Schimpel et al, 2014). In our system, we focused on an intestinal model that could mimic either healthy or inflamed conditions.…”

Prediction of Oral Absorption of Nanoparticles from Biorelevant Matrices Using a Combination of Physiologically Relevant In Vitro and Ex Vivo Models

“…In addition, clathrin-and/ or caveolin-mediated endocytosis of NPs is also possible, but these processes are under-represented in Caco-2 cells [38]. Different co-or tri-cultures with mucus secreting goblet cells and/or M-cells have been used to get more representative results of these studies for the in vivo situation, but it remains difficult to accurately mimic the physiological complexity of the gastrointestinal tract in vivo with regard to anatomy and physiological processes [2,[8][9][10]. Alternatively, animal studies can be applied to determine the oral absorption of NPs in physiological environments and during physiological process.…”

“…Walczak et al [8], Schimpel et al [9], Lautenschlager et al [14], des Rieux et al [17]). However, it has been shown tht NPs can be digested or otherwise structurally altered (including processes such as degradation and/or agglomeration) in the gut lumen in vivo, which might influence their absorption [19,20].…”

“…However, single cell type models do not represent the whole complexity of the intestinal wall in vivo. Different co-or tri-cultures with mucus secreting goblet cells and/or M-cells have therefore been developed to mimic the in vivo situation, however the discontinuous mucus layer and relative presence of M cells in vitro do not match the situation in vivo [8][9][10].…”

Co-culture Caco-2/Immune Cells

“…Several methodological reports are describing co-culture conditions between Caco-2 and CD14 + monocyte derived DCs, THP-1 or PBMCs (Araujo and Sarmento 2013 ;Leonard et al 2010 ;Schimpel et al 2014 ;Antunes et al 2013 ;Kerneis et al 2000 ;Parlesak et al 2004 ). However, there are still few reports where these co-culture systems have been used to scrutinize communication between epithelial cells and immune cells.…”