116. Contribution of claudin-11 to the inter-Sertoli cell tight junction, in vitro
M. J. McCabe A B and P. G. Stanton AA Male Reproductive Endocrinology, Prince Henry’s Institute of Medical Research, Clayton, VIC, Australia
B Applied Biology/Biotechnology, Royal Melbourne Institute of Technology, Bundoora, VIC, Australia
Reproduction, Fertility and Development 17(9) 72-72 https://doi.org/10.1071/SRB05Abs116
Submitted: 26 July 2005 Accepted: 26 July 2005 Published: 5 September 2005
Abstract
The inter-Sertoli cell tight junction (TJ) forms the blood testis barrier (BTB) between Sertoli cells and is composed of three major transmembrane proteins: claudin-11, occludin and junctional adhesion molecule. Formation of the BTB occurs during puberty associating with an increase in circulating gonadotrophins. Claudin-11 and occludin are hormonally regulated in vitro although their importance to the function of the TJ is unknown. The aim of this study was to investigate the contribution of claudin-11 to the inter-Sertoli cell TJ in vitro by blocking gene expression using RNA interference. Two claudin-11-specific siRNA fragments were designed for this purpose. Sertoli cells in primary culture formed stable TJs within 5 days as measured by transepithelial electrical resistance (TER). The addition of siRNA for 2 days resulted in a significant (P < 0.01) 55% (mean, SD, n = 4 cultures) decrease in TER along with a major reduction in claudin-11 localisation to the TJ as assessed by immunocytochemistry. The specificity of the siRNA was shown by the presence of extensive immunostaining of occludin and of the adherens junction protein β-catenin in the same treatments. Similarly, claudin-11 mRNA expression significantly (P < 0.01) decreased by 71% (mean, SD, n = 3 cultures) in response to both claudin-11 siRNA fragments. Occludin mRNA expression was not affected. It is concluded that claudin-11 contributes at least 55% to the function of the rat Sertoli cell TJ in vitro. It is hypothesised that the remaining 45% of TJ function can be attributed to other integral proteins, such as occludin and junctional adhesion molecule. It is expected that claudin-11 and other TJ proteins play a pivotal role in the function of the BTB in vivo with potential implications in fertility and contraception.
