Screening for Chemical Toxicity Using Cryopreserved Precision Cut Lung Slices

Abstract

To assess chemical toxicity, current high throughput screening (HTS) assays rely primarily on in vitro measurements using cultured cells. Responses frequently differ from in vivo results due to the lack of physical and humoral interactions provided by the extracellular matrix, cell-cell interactions, and other molecular components of the native organ. To more accurately reproduce organ complexity in HTS, we developed an organotypic assay using the cryopreserved precision cut lung slice (PCLS) from rats and mice. Compared to the never-frozen PCLS, their frozen-thawed counterpart slices showed viability or metabolic activity that is decreased to an extent comparable to that observed in other cryopreserved cells and tissues, but shows no differences in further changes in cell viability, mitochondrial integrity, and glutathione activity in response to the model toxin zinc chloride (ZnCl2). Notably, these measurements were successfully miniaturized so as to establish HTS capacity in a 96-well plate format. Finally, PCLS responses correlated with common markers of lung injury measured in lavage fluid from rats intratracheally instilled with ZnCl2. In summary, we establish that the cryopreserved PCLS is a feasible approach for HTS investigations in predictive toxicology.

Cigarette smoke up‐regulates PDE3 and PDE4 to decrease cAMP in airway cells

“…Although FRET has previously been used to estimate cAMP levels in human airway epithelial cells (Schmid et al ., 2006, 2015), airway smooth muscle cells (Billington and Hall, 2011) and endothelial cells (Yañez‐Mó et al ., 2008), real‐time monitoring cAMP levels in the lung tissue has not been reported so far. Due to the fact that PCLS retain the complex micro‐composition and environment of the airways, they are recognized as a reliable model to study airway responsiveness and drug toxicity (Schlepütz et al ., 2012; Oenema et al ., 2013; Watson et al ., 2015). In the present study, we report that PCLS from Epac1‐camps FRET reporter mice represent a useful model to monitor airway cAMP levels in real‐time.…”

Organ-on-a-chip for assessing environmental toxicants

“…Conventional HTS relies on 2D cultured cells to evaluate the cytotoxicity to drugs or toxicants, whose responses differ from those obtained in vivo due to the lack of physical and humoral interactions provided by the ECM, cell-cell interactions, and other molecular components of the native organ [5]. Indeed animal models do reproduce organ complexity more accurately but deduction of toxicological responses may be ambiguous due to inter-species differences and thus irrelevant to human physiological responses.…”

“…Indeed animal models do reproduce organ complexity more accurately but deduction of toxicological responses may be ambiguous due to inter-species differences and thus irrelevant to human physiological responses. Also, the time consumption, cost, and ethical concerns of animal testing disfavors its use in toxicological research [5,6]. …”

“…On the other hand, precision-cut organ models, where thin tissue slices are used rather than 2D cultured cells, demonstrate the sheer advantage of direct interspecies comparison with respect to metabolic capacity and sensitivity for toxicants [6], and therefore has been identified as useful models for toxicological assessment [5,11,12]. However, obtained tissue slices are largely constrained by the limited viability for toxicological testing, which inhibits long-term toxicity studies [5].…”

“…However, obtained tissue slices are largely constrained by the limited viability for toxicological testing, which inhibits long-term toxicity studies [5]. …”