IN VITRO TOXICOLOGY--PRECISION TISSUE SLICE TECHNOLOGY

The goal is to develop precision-cut organ slice culture into in vitro
systems for the study of toxicology. Although organ slices have been used
for years in in vitro studies, they have been hampered by non-reproducible
slicing techniques and inadequate culturing systems. This study proposes
to solve the key problems hampering the use of slices in in vitro
toxicology studies. Kidney and liver tissue from rats and rabbits will be
utilized; however, other tissues and other species will be used in future
studies. The Specific Aims are to: 1.) DEVELOP TECHNIQUES FOR THE
PREPARATION OF PRECISION-CUT TISSUE SLICES. Basically, mechanical slicers
will be designed allowing reproducible, precision slices to be made from
various tissues. Slicers will be designed and fabricated for both
producing slices in a sterile environment and for slicing non-homogenous
tissue into positional slices. 2.) DEVELOP TECHNIQUES FOR THE CULTURING
OF PRECISION-CUT TISSUE SLICES. This is a key Aim since past organ culture
produced partially dead tissue. Various apparatuses for culturing slices
will be designed and fabricated. This will include both closed
(recirculatory) and open flow-through systems which will allow prolonged
culturing of the slices. 3.) DEVELOP TECHNIQUES FOR ASSESSING VIABILITY
(TOXICITY) OF CULTURED SLICES. Due to maintenance of tissue architecture,
the slices contain all the tissue's cell types. Thus viability assays that
reflect the complexity of the slice's cell distribution and their
susceptibility to intoxication by site-specific toxins will be developed.
These will include indicators of specific biochemical and functional
processes. 4.) EVALUATE CULTURED SLICES FOR THEIR RESPONSE TO
ORGAN-SPECIFIC TOXICANTS. Using the optimum slicing, culturing, and
toxicity indicators, organ-specific toxicants will be profiled. Initially
only renal and liver tissue will be examined, but these studies will expand
to other tissues as techniques are developed. Besides a considerable
savings in animals, this proposal will produce systems in which mechanistic
toxicity questions can be addressed that not easily answered in vivo.