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Abstract

Grant Number:	2R44ES010887-02A1
PI Name:	CAMPBELL, LIA H.
PI Email:	lcampbell@organ-recovery.com
PI Title:
Project Title:	Development of Cryoplates for Cytotoxicity Testing

Abstract: DESCRIPTION (provided by applicant): Advancements in high throughput screening, and mandates all over the world to reduce animal testing, have driven the development of a variety of assays to evaluate potentially new drugs and their side effects. Environmental monitoring of potentially harmful chemicals and by-products has also driven the development of new assays. Convenient cell based systems for testing a wide variety of products with a variety of cell types has become a great need. In Phase I, we proposed the development of a system providing ready to use cells on microtiter plates for any desirable assay. We demonstrated that bovine corneal endothelial cells (BCE) attached to tissue culture plastic or an extracellular matrix can be cryopreserved and be viable after thawing. We have also shown that we are able to uniformly cool and warm microtiter plates and improve overall retention of cells to their substrate. The goal of the Phase II proposal, is to further define and optimize the parameters that will provide successful cryopreservation of BCE cells attached to microtiter plates. It was shown that cells attached to an extracellular matrix demonstrated better viability after cryopreservation than cells attached to tissue culture plastic. Further examination of extracellular matrix components will be performed to determine if adjustments to the composition of the matrix can improve viability and attachment. In addition other cryobiological variables will be evaluated including optimization of cooling, the choice of cryoprotectant vehicle solution and its influence on cell survival as well as long term storage of cells after cryopreservation on microtiter plates. All these experiments are designed to further optimize the system and provide cells on plates that are ready to use for a variety of applications. Validation of cryopreserved BCE cells will be provided by a battery of functional assays including, but not limited to, responses to toxicants, extracellular matrix formation, barrier function and nitric oxide production. It is anticipated that optimization of procedures for corneal endothelial cells will provide a technology platform that can be applied, with further development, to other cell types.

Thesaurus Terms:
corneal endothelium, cryopreservation, cytotoxicity, technology /technique development
apoptosis, cell proliferation, collagen, enzyme activity, extracellular matrix, fibronectin, high throughput technology, superoxide dismutase, transforming growth factor
tissue /cell culture

Institution:	ORGAN RECOVERY SYSTEMS, INC.
	701 E BAY ST, STE 433
	CHARLESTON, SC 29403
Fiscal Year:	2003
Department:
Project Start:	30-SEP-2000
Project End:	30-JUN-2005
ICD:	NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
IRG:	ZRG1

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