Involvement of vitronectin and of a natural extracellular matrix in prostatic cell to cell contact and interaction with the substratum in culture

Abstract

The kinetics and regulation of attachment, the spreading and initial growth of canine prostatic epithelial cells in primary monolayers, as well as the ability of the cells to produce a natural extracellular matrix (nECM) have been studied. Within 1 day of culture, attachment without spreading occurred in uncoated dishes while spreading was observed in nECM-coated dishes. The attachment was stimulated by dialyzed fetal bovine serum (dFBS) and/or by the nECM and was not affected by the presence of cycloheximide at 10(-6) M. This effect of dFBS on attachment was maximal at a concentration of 10% and increased with cell density and was mainly attributed to vitronectin as demonstrated by gel filtration, selective adsorption of serum proteins to the substratum, and indirect immunofluorescence of prostatic epithelial cells incubated with a rabbit antiserum against bovine vitronectin. Dog serum reduced attachment while bovine serum albumin, phenol red, and steroids were inactive. The maximal effect of the nECM on attachment was observed with matrices produced by canine prostatic epithelial cells cultured in the presence of dog serum for more than 10 days. As revealed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, several proteins of relative mass (Mr) of 210, 113, 61, 56, 52, 49, 45, and 42 kDa remained in the dish following nECM denudation and could be responsible for its effect on attachment. Their deposition as well as intercell adhesion and cell-to-substratum interactions required an active protein synthesis plus non-steroidal serum components. Once attached and spread in primary monolayers, prostatic epithelial cells divided by the 3rd to 4th day; their growth, stimulated by dFBS at an optimal concentration of 20%, was inhibited by cycloheximide while the nECM was inactive. Thus, vitronectin in serum and the nECM synthesized by canine prostatic epithelial cells are responsible for the attachment and spreading of homologous cells to an in vitro substratum; nECM proteins also contribute to cell-to-cell contact and cell-to-substratum interaction in the monolayers.

Serum and prostatic growth‐promoting factors for steroid‐independent epithelial cells of adult dog prostate

“…Unless otherwise indicated, epithelial cell cultures were initiated in 60 X 15 mm dishes with 2 X lo5 cells in 3 ml of Minimum Essential Medium D-valine (MEM D-Val) supplemented with 10% dialyzed fetal bovine serum (dFBS) and 1% antibiotic solution (10,000 units of penicillin and 10,000 pg of streptomycin/ ml). After a 3 day attachment period, mainly regulated by serum vitronectin [14], the number of epithelial cells in monolayers was determined on triplicate cultures and the medium of the remaining dishes was renewed and supplemented as described below to study their proliferation.…”

Techniques for Studies on Growth Characteristics of Human Prostatic Cancer Cells

“…One-third of all men over the age of 50 are affected by nonclinical, i.e., latent, prostate cancer. However, only a small fraction of the latent become Furthermore, proliferation and growth of even malignant epithelial prostate cells are regulated by growth factors of and by contact with healthy epithelial cells and stroma cells (Story et al, 1989; Thompson, 1990; Chang and Chung, 1989; Chevalier and Chapdelaine, 1991; Diakiew et al, 1991; Gleave et al, 1991; Orlowski and Clark, 1991).…”