Adhesion, Shape, Proliferation, and Gene Expression of Mouse Leydig Cells are Influenced by Extracellular Matrix in Vitro1

Abstract

Interactions between Leydig cells and the extracellular matrix (ECM) within the interstitial compartment of the mammalian testis have not been characterized. We have examined the influence of ECM on adult mouse Leydig cells by culturing the cells on different ECM substrates. Leydig cells adhere weakly to hydrated gels of type I collagen (including those supplemented with collagen types IV, V, or VIII), or to air-dried films of collagen types I, V, or VIII. In contrast, the cells attach firmly to substrates of purified type IV collagen, fibronectin, or laminin. Leydig cells also attach rapidly and adhere strongly to gelled basement membrane matrix derived from the murine Englebreth-Holm-Swarm sarcoma (Matrigel). Leydig cells assume spherical shapes and form aggregates on thick (1.5-mm) layers of Matrigel; however, on thin (0.1-mm) layers, networks of cell clusters linked by cords of elongated cells are formed within 48 h. Similar networks are formed on thick layers of Matrigel that are supplemented with type I collagen. On substrates with high ratios of collagen I to Matrigel or on untreated tissue culture plastic, Leydig cells flatten and do not aggregate. On substrates that induce rounded shapes, proliferation is inhibited and the cells maintain the steroidogenic enzyme 3 beta-hydroxysteroid dehydrogenase for as long as 2 wk. Under conditions where Leydig cells are flattened, they divide and cease expressing the enzyme. Proliferating Leydig cells also exhibit elevated levels of mRNA for SPARC (Secreted Protein, Acidic and Rich in Cysteine), a Ca2(+)-binding glycoprotein associated with changes in cell shape that accompany morphogenesis and tissue remodeling. Our results indicate that the shape, association, proliferation, and expression of gene products by Leydig cells can be significantly affected in vitro by altering the composition of the extracellular substratum.