Culture of endocrine pancreatic cells in protein-free, chemically defined media

Abstract

Cell suspensions prepared by collagenase digestion of pancreata obtained from 21.5-d-old rat fetuses were preincubated in RPMI medium containing 10% fetal bovine serum (FBS), to ensure cell adhesion. Twenty hours later, this medium was replaced by a chemically defined medium. Dulbecco's modified Eagle's (DME)-F12 was used alone or supplemented with various combinations of transferrin, sodium selenite, or Ultroser G. The evolution of the culture and the islet ultrastructure were similar in defined and serum-containing media. However, in the defined medium, the neoformed islets seemed less numerous, and the fibroblast layer less dense, when compared to the RPMI + 10% FBS control medium. At Day 7, in defined media, the total insulin content per dish was half that of control cultures. None of the tested additives improved the yield of the cultures. The fractional insulin release per day was elevated in defined media. In subsequent incubations, glucose and leucine stimulated insulin release in a way characteristic of these cells of fetal origin. The labeling index of islet cells cultured in DME-F12 reached 10.7%, which is not far from that observed in RPMI + 10% FBS. Such a defined medium is useful to study B cell physiology, avoiding the possible interaction of serum components with substances to be tested.

Development of embryonic chick insulin cells in culture: Beneficial effects of serum-free medium, raised nutrients, and biomatrix

“…Insulin, transferrin, and to a lesser extent selenium are recommended constituents of serum-free media (5,8,16). Although transferrin is essential for the satisfactory culture of a number of cell types (8), its effect on islet cell growth is not particularly marked (see 13,22,34). According to Kinard et al (22) and Fong et al (13) selenium is not 778 RAWDON AND ANDREW essential for satisfactory islet cell development.…”

“…bryonic chick insulin cells in culture may well differ from those found suitable for prenatal rat islets (22,28). In order to identify the most important components of a culture medium for chick cells, we have adopted use of serum-free medium: this has obvious advantages over serum-containing medium (13) for our purpose.…”

Morphogenesis and differentiation of the avian endocrine pancreas, with particular reference to experimental studies on the chick embryo

“…The latter three components are all recommended constituents of serum-free media (see for example, Barnes and Sato, 1980) but there are indications that for development of islet cells, selenium and transferrin are not as important as insulin (Fong et al, 1981;Kinard et al, 1990). In view of the known stimulatory effect of insulin on proliferation of rat islet (mainly insulin) cells in vitro (Hayek et al, 1989;Rabinovitch et al, 1982), we analysed the role of insulin in our culture system.…”