A HIGH-THROUGHPUT IN VITRO MODEL OF HUMAN TRACHEAL/ BRONCHIAL EPITHELIUM (EPIAIRWAY™) FOR PRE-CLINICAL SAFETY AND EFFICACY TESTING OF PHARMACEUTICALS.

This study by scientists at MatTek Corp. demonstrated that MatTek’s EpiAirway in vitro 3-D human tracheal/bronchial tissue equivalent can be produced in a 96-well high-throughput microwell plate format in a highly reproducible manner, enabling the use of this format for in vitro irritation/toxicity screening, high content cell and molecular biology assays, as well as drug permeation studies. A growing need exists for high throughput in vitro models which can provide rapid, reliable safety and efficacy screening in preclinical drug development applications. The current poster describes development of an in vitro model of human tracheal/bronchial epithelium (TBE) in a 96-well high-throughput screening (HTS) format compatible with robotic manipulation. The HTS model is derived from normal human cells cultured at the air/liquid interface in 96-well microporous membrane plates to produce three-dimensional organotypic cultures. Histology of the culture model was evaluated by H&E staining of formalin fixed paraffin sections. The 96-well TBE culture is pseudo-stratified, and displays a differentiated mucociliary phenotype with barrier properties similar to native tracheal/bronchial or nasal epithelium, including development of transepithelial electrical resistance (TEER), conferred by functional tight junctions. Over the course of several consecutive culture lots, the intra-plate and inter-lot barrier function reproducibility of the 96-HTS tracheal/bronchial epithelium cultures was determined by measurement of TEER. In initial studies to date, the average baseline TEER readings of all wells in a given 96-well plate (intra-plate average) ranged from 388.6 to 445.0 cm2. The average coefficient of variation between wells on the same plate (i.e. intra-plate variability) was 19.5 %, while the variability between plates (i.e. inter-lot variabil-ity) was 6.8 %. The utility of the HTS tracheal/bronchial epithelium model was demonstrated by evaluation of the effects of various cytokine treatments on 15-lipoxygenase gene expression and TARC secretion. PCR analysis of total RNA isolated from the cultures showed that TNF-plus IL-4 induced 15-lipoxygenase-2 expression. ELISA experiments revealed the same treatments also induced TARC secretion. The high-throughput screening (HTS) airway model should find utility for in vitro irritation/toxicity screening, high content cell and molecular biology assays, as well as drug permeation studies. This work was supported by NIEHS grant #5R44 ES010237-03.

15-lipoxygenase gene, 96-well, Bronchial, Cytokine treatments, Drug permeability studies, ELISA experiment, Efficacy screening, EpiAirway, Epithelium, HTS, High content cell biology assays, High content molecular biology assays, High-throughput screening, In vitro efficacy screening, In vitro irritation screening, Nasal epithelium, Pre-clinical efficacy, Pre-clinical toxicity, Safety and efficacy testing, TARC secretion, TEER, Tracheal, Tracheal/bronchial epithelium (TBE), Transepithelial electrical resistance

Cytokines, IL-13, IL-4, INF-a, TNF2