ToxSci Advance Access published online on January 19, 2005
Toxicological Sciences, doi:10.1093/toxsci/kfi085
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1 Dept. of Environmental and Occupational Health Sciences, University of Washington
* To whom correspondence should be addressed. The development of in vitro models for testicular toxicity may provide important tools for investigating specific mechanisms of toxicity in the testis. Although various systems have been reported, their application in toxicological studies have been limited due to the poor ability to replicate the complex biochemical, molecular and functional interactions observed in the testis. In the present study, we evaluated a significantly improved Sertoli cell/gonocyte co-culture (SGC) system that utilizes a 3-dimensional extracellular matrix Matrigel (ECM) applied as an overlay instead of a substratum. We explored the dose- and time-dependent effects of the addition of such an ECM overlay on cytoskeletal and morphological changes in the SGC system, and the resultant impacts on cellular integrity. Furthermore, we correlated the latter effects with the ECM-dependent modulation of stress and survival signaling pathways and most critically the expression levels of the spermatagonia-specific protein, c-Kit. Finally, we applied this co-culture system to investigate the dose and time-dependant effects on the morphology and induction of apoptosis of cadmium. We observed that the dose dependent addition of an ECM overlay led to an enhanced attachment of Sertoli cells and facilitated the establishment of SGC communication and cytoskeletal structure with a dramatic improvement in cell viability. The latter was consistent with the observed dose and time-dependent modulation of both stress (SAPK/JNK) and survival (ERK and AKT) signaling pathways in the presence of the ECM overlay. Furthermore, the dose-dependent stabilization of c-Kit protein expression confirmed the functional integrity of this co-culture system. We conclude that this modified SGC system will provide investigators with a simple, efficient, and highly reproducible alternative in the screen for testicular cell-specific cytotoxicity and the assessment of molecular mechanisms associated with both normal development as well as of reproductive toxicity induced by environmental toxicants.
Received October 19, 2004
Accepted January 4, 2005
Reproductive and Developmental Toxicology
Essential role of extracellular matrix (ECM) overlay in establishing the functional integrity of primary neonatal rat Sertoli cell/gonocyte co-cultures: an improved in vitro model for assessment of male reproductive toxicity
Elaine M Faustman, E-mail: faustman{at}u.washington.edu
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