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Transfection of Sertoli cells ex vivo; Opening the door to the in vitro study of junction turnover in the seminiferous epithelium
Author(s) -
Guttman Julian Andrew,
Rusnak Chris,
Wilkinson Darcy,
Deng Wanyin,
Roskelley Calvin,
Finlay Brett,
Takai Yoshimi,
Vogl Wayne
Publication year - 2006
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.20.5.a883-a
Subject(s) - adherens junction , sertoli cell , microbiology and biotechnology , transfection , internalization , cadherin , cell junction , tight junction , biology , gap junction , hela , nectin , cell–cell interaction , cell adhesion , cell culture , chemistry , intracellular , spermatogenesis , cell , endocrinology , genetics
Sertoli cells interact with adjacent Sertoli cells and with spermatids through elaborate intercellular adhesion junctions termed ectoplasmic specializations (ESs). Among adhesion molecules present at the sites is Nectin‐2. Tubulobulbar complexes have been proposed to play a role in junction internalization. Here we establish a culture model for exploring the role of tubulobulbar complexes during junction turnover. Cell clumps enriched for Sertoli cells were isolated from ten day‐old rat pups and cultured on Matrigel coated Transwells. These cells established a columnar morphology and formed identifiable basal ESs. Significantly, structures resembling tubulobulbar complexes developed at the junction sites. We generated eGFP‐tagged nectin‐2 constructs and transfected them first into Hela cells (cells that do not normally form adherens junctions) and then into our primary cultures of morphologically differentiated Sertoli cells. In Hela cells, eGFP‐Nectin‐2 localized to sites of intercellular contact between adjacent cells indicating that the protein was expressed and targeted to the plasma membrane where it appeared to establish adherens junctions. In Sertoli cells, the protein was localized to areas consistent with the position of ESs and to tubular structures associated with them. Primary Sertoli cell cultures may be a model for studying the role of tubulobulbar complexes in junction turnover. CIHR MOP 62728