Non Animal Testing, Alternative Test Methods, In Vitro Toxicology, IIVS | Increased Throughput and Cryopreservation of Precision-cut Lung Slices Extend the Utility of Human-relevant, 3-Dimensional Pulmonary Test Systems
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Increased Throughput and Cryopreservation of Precision-cut Lung Slices Extend the Utility of Human-relevant, 3-Dimensional Pulmonary Test Systems

March 18, 2019

Human-relevant, in vitro/ex vivo assays are considered an ethical and economically viable manner by which to screen the thousands of chemicals requiring hazard assessment. Of the 3-dimensional models, human precision-cut lung slices (PCLS) are often considered the most physiologically relevant pulmonary test system, but lower throughput and difficulties in cryopreservation have hampered PCLS use.

We have modified a tissue slicer to accommodate 3 tissue cores for simultaneous slicing. Increased slice production was quantified using agarose and tissue cores in the slicer. To evaluate cryopreservation of PCLS, we have tested 5 cryopreservation formulations using PCLS (frozen on the day of slicing, or after overnight culture). Thawed slice viability in each of the groups was assessed with the WST-8 viability assay, prior to fixation and histological evaluation.

The slicer modification resulted in 2.8-fold and 2.4-fold more slices from agarose cores, and lung cores, respectively. Cryopreservation efforts indicated freezing after slicing yields better average viability (48-73% of fresh, non-frozen control) than culturing overnight and freezing (13-54% of control) when assessing health over 4 days, post-thaw. Cryopreservation buffers containing University of Wisconsin preservation solution preserved viability the best (54%-90% of non-frozen control). Histological findings concurred with WST-8 viability results and indicated the retention of healthy lung tissue features (>75% of normal), post-thaw.

The increased PCLS production indicates larger (or multiple) studies can be initiated from one donor lung. The promising cryopreservation results suggest slices can be banked and utilized at a later date, potentially even allowing the same donor’s tissue to be used repeatedly.