Polarized Sertoli Cell Functions in a New Two‐compartment Culture System

Abstract

Sertoli cells in vivo are highly polarized and interact with the inner (tubular) and outer (interstitial) fluids. To simulate this condition in vitro we developed a two-compartment culture system in which confluent Sertoli cell monolayers were grown on permeable supports (Millipore filters) separating the inner and outer fluid compartments. Monolayer permeability to (3H)-inulin decreased by 90% after 5 to 7 days of culture, presumably due to formation of tight junctions. This process was influenced by cell plating density. The cells were highly polarized morphologically, resembling their appearance in vivo, and secreted transferrin bidirectionally into both fluid compartments. The amount of transferrin secreted was 166% to 250% of that secreted by the same number of Sertoli cells cultured in plastic dishes. Testosterone (5 X 10(-8) M) doubled and testosterone + FSH (0.1 microgram/ml) increased transferrin secretion 3.6-fold. These results demonstrate that under suitable culture conditions the Sertoli cells remain both morphologically and functionally polarized, reflecting a more physiologic state.

Intercellular adhesion molecule-1 is a regulator of blood–testis barrier function

“…Cells were seeded at a density of 0.5610 6 cells/cm 2 in 12-well plates for lysate preparation, RNA isolation and overexpression experiments, at a density of 1.2610 6 cells/cm 2 in Millicell inserts (Millipore Corp., Billerica, MA) for transepithelial electrical resistance (TER) measurements and overexpression experiments, or at a density of 0.05610 6 cells/ cm 2 on round glass coverslips (18 mm; Thomas Scientific, Swedesboro, NJ) for immunofluorescent staining. It is worth noting that functional junctions were assembled and maintained in control Sertoli cells cultured at all densities used in this study, a conclusion that is based on nearly two decades of research from this and other laboratories Chung et al, 1999;Dym and Papadopoulos, 1992;Florin et al, 2005;Hadley et al, 1987;Janecki et al, 1991;Janecki and Steinberger, 1986;Lie et al, 2011;Lui et al, 2001;Siu et al, 2003;Su et al, 2010b;Tung and Fritz, 1993). All substrata were coated with Matrigel TM basement membrane matrix as previously described (Xiao et al, 2011).…”

The Blood-Testis Barrier and Its Implications for Male Contraception

“…Other studies using Sertoli cells cultured in vitro (Byers et al, 1986;Janecki and Steinberger, 1986;Kaitu'uLino et al, 2007;Yan et al, 2008b) have also demonstrated the role of androgens in conferring BTB integrity, because the presence of androgens was shown to promote the assembly of the Sertoli cell TJ-permeability barrier and to lessen the cadmium-induced disruptive effects on the Sertoli cell TJ barrier function (Janecki et al, 1992;Chung and Cheng, 2001). Testosterone was also shown to stimulate the steady-state levels of occludin (Kaitu'u-Lino et al, 2007;Yan et al, 2008b) and claudin-11 (Kaitu'u-Lino et al, 2007) in the Sertoli cell epithelium and the proper localization of these proteins at the Sertoli-Sertoli cell interface.…”

Vitamin C and oxidative stress in the seminiferous epithelium

“…Sertoli cells in vivo are highly polarized and interact with the inner (adluminal) and outer (basal) fl uids (Janecki & Steinberger 1986). Because of the physical separation produced by tight junctions, germ cells have to move from one compartment to the other, thus defi ning the polarity of the epithelium (Maeda et al, 2007).…”