In Vitro Tissue Models

While the performance of a new drug or delivery formulation is most critically evaluated using in vivo models, initial screening and development are more efficiently accomplished using a well-defined and controlled in vitro cell culture model. In vitro screening for transport of a drug across an epithelial barrier requires growth of a confluent cell monolayer on a suitable porous support membrane and differentiation into polarized epithelium with robust intercellular TJs. Ideally, these cultures mimic nasal epithelium in their composition of ciliated and nonciliated columnar cells, mucus-producing goblet cells, and basal cells. However, HTS efforts for formulation optimization may be associated with concerns of cost and reproducibility of the cell model.

Human epithelial cell lines are popular choices for oral, nasal, and respiratory drug delivery studies. Models such as Caco-2 are widely used as an oral (i.e., intestinal) delivery model in assays for drug metabolism and transport (89). Most nasal delivery studies utilize Calu-3 (90) or 16HBE14o- (91) cell lines, which readily differentiate into polarized epithelial cells on porous support membranes but are derived from respiratory epithelial cells and may not mimic nasal epithelium [for more in-depth review, see Merkle et al. (92)]. Recently, culture methods for the nasal epithelial cell line RPMI 2650 have been optimized for TJ formation in monolayers (93). When grown at an air-liquid interface, RPMI 2650 differentiates to form confluent monolayers with significantly higher transepithelial resistance (TEER) than cells cultured at a liquid interface. TEER is an important characteristic since its reduction can be used as a formulation-screening tool, that is, an indication of TJ opening. Formation of robust TJs was further indicated by the ability to resist permeation of paracellular flux markers such as mannitol. Additionally, RPMI 2650 cell line has been shown to be useful in drug metabolism studies (92).

Primary respiratory epithelial cell cultures can be differentiated to form pseudostratified, mucociliary cultures. While cost, tissue availability, and reproducibility are major barriers for HTS in primary cell cultures, there are well-established methods for the culture of primary human bronchial/tracheal epithelial cells, as well as companies that provide reproducible, ready-to-use, differentiated cultures in formats suitable for HTS (MatTek Corporation, Ashland, Massachusetts, U.S.A.; Epithelix Sarl, Plan-Les-Ouates, Geneve, Suisse). Further, the recent development of hTERT-immortalized primary nasal cells (94), which allows for extended lifespan while preserving karyotype, holds promise for a differentiated nasal cell model, which preserves the key biological characteristics of the nasal mucosa while allowing for HTS.

0 0

Post a comment