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Development of a high-throughput screening assay for chemical effects on proliferation and viability of immortalized human neural progenitor cells
Citation:
Breier, J. M., N. RADIO, W. R. MUNDY, AND T. J. SHAFER. Development of a high-throughput screening assay for chemical effects on proliferation and viability of immortalized human neural progenitor cells. TOXICOLOGICAL SCIENCES. Society of Toxicology, RESTON, VA, 105(1):119-133, (2008).
Impact/Purpose:
In vitro models may be useful for the rapid toxicological screening of large numbers of chemicals for their potential to produce toxicity. Such screening could facilitate prioritization of resources needed for in vivo toxicity testing towards those chemicals most likely to result in adverse health effects. Cell cultures derived from nervous system tissue have proven to be powerful tools for elucidating cellular and molecular mechanisms of nervous system development and function, and have been used to understand the mechanism of action of neurotoxic chemicals. This study focuses on the development and optimization of an automated high content screening assay to examine chemical effects on proliferation and viability in RenCell CX cells, an immortalized human neuroprogenitor cell line. Effects on proliferation and viability were examined following exposure to chemicals known to be inhibitors of proliferation in other cell types, and on a set of test chemicals consisting of toxic and neurotoxic compounds across a wide concentration range (1 nM – 100 µM). For all five compounds known to inhibit proliferation in other systems, significant inhibition of proliferation of RencellCX cells was observed at concentrations below those that resulted in significant cytotoxicity. For six of eight neurotoxic compounds, significant effects were observed on proliferation and/or viability: proliferation was more sensitive than viability for five of these compounds. Conversely, six of eight non-toxic compounds were without effect on either proliferation or viability. The results indicate the potential for high content analysis to rapidly quantify chemical effects on proliferation and viability of human neuroprogenitor cells in vitro. The manuscript demonstrates the application of a new approach for assessment of developmental neurotoxicity that can be used as part of a battery of tests for screening large numbers of chemicals.
Description:
There is considerable public concern that the majority of commercial chemicals have not been evaluated for their potential to cause developmental neurotoxicity. Although several chemicals are assessed annually under the current developmental neurotoxicity guidelines, time, resource, and animal constraints prevent testing of large numbers of chemicals using this approach. Thus, incentive is mounting to develop in vitro methods to screen chemicals for their potential to harm the developing human nervous system. As an initial step toward this end, the present studies evaluated an automated, high-throughput method for screening chemical effects on proliferation and viability using ReNcell CX cells, a human neural progenitor cell (hNPC) line. ReNcell CX cells doubled in approximately 36 h and expressed the neural progenitor markers nestin and SOX2. High-throughput assays for cell proliferation (5-bromo-2'-deoxyuridine incorporation) and viability (propidium iodide exclusion) were optimized and tested using known antiproliferative compounds. The utility of this in vitro screen was evaluated further using a set of compounds containing eight known to cause developmental neurotoxicity and eight presumably nontoxic compounds. Six out of eight developmental neurotoxicants significantly inhibited ReNcell CX cell proliferation and/or viability, whereas two out of eight nontoxic chemicals caused only minimal effects. These results demonstrate that chemical effects on cell proliferation and viability can be assessed via high-throughput methods using hNPCs. Further development of this approach as part of a strategy to screen compounds for potential effects on nervous system development is warranted.
