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.
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