PARENT SESSION

(25) THE EPIDIDYMAL JUNCTIONAL COMPLEX IS IMPAIRED BY TRIBUTYLTIN EXPOSURE.

Barthelemy, Johanna¹, Dufresne, Julie¹, Cyr, Daniel¹, ¹ INRS-Institut Armand-Frappier, Pointe-Claire, QC, Canada

ABSTRACT- Tributyltin (TBT) is widely distributed in the environment. TBT is used as a biocide and as an anti-fouling agent. We have previous observed that TBT exposure during in utero development can result in alterations to the male reproductive system which persist to adulthood. These effects include altered sperm motility, decreased ventral prostate weight and effects on junctional proteins. The development of the male reproductive system is complex and requires sex hormones as well as direct interactions between cells. These interactions permit the creation of specific micro-environments which are critical for proper development and maturation of spermatozoa. Our objectives were to determine the effects of the TBT on the epididymis of rats exposed in utero and to assess how TBT alters the expression of Cldn-1. Proteins from adherent junctions (E-Cadherin and -Catenin) and tight junctions (Claudin-1 and Zonula Occludens-1) were significantly decreased by TBT as was their localization. To determine the mechanism of TBT action, an immortalized epididymal epithelial cell line was used. Epididymal cells were exposed to either 500 nM or 1M of TBT for 24h. This resulted in a dose dependent decrease in both Cldn-1 and Cldn-3 mRNA levels as determined by RT-PCR. Important effects were noted on the junctional complex in the epididymis. In order to determine if TBT altered the transcription of Cldn-1, a luciferase expression assay was developed. To do this we cloned the rat Cldn-1 promoter into a promoterless luciferase expression vector. Cells were transfected with the full length Cldn-1 luciferase construct, and allowed us to we established that the promoter sequence contained sufficient information to drive the expression of Cldn-1. Using this construct, epididymal cells were then transfected with the construct and exposed to either vehicle or TBT (500nM or 1M), at doses which were previously determined to be non-lethal and which lowered Cldn-1 levels. Results indicated a dose-dependent decrease in luciferase activity in the cells treated with TBT. Based on these results, we propose a mechanism by which TBT affects tight junctions trough an inhibitory effect on the transcription factors regulating the expression of Cldn-1. Supported by NSERC and CIHR.

KEY WORDS: epididymis, Tributyltin, junctions