Serum-free growth of normal and transformed fibroblasts in milk: differential requirements for fibronectin.

Abstract

Bovine milk may be used as a supplement for the serum-free growth of certain fibroblastic cells in culture. The growth properties of three representative cell types in milksupplemented medium were examined ; fibroblastic cell strains, fibroblastic cell lines, and transformed fibroblasts . Transformed fibroblasts, which included RNA and DNA tumor virustransformed cells and carcinogen-transformed cells, grew in milk . Instead of growing attached to the culture dishes, as they normally do in serum, transformed fibroblasts grew in milk as large clusters in suspension . In contrast, nontransformed fibroblastic cell strains and cell lines did not grow in milk-supplemented medium . Fibroblasts transformed by a temperaturesensitive transformation mutant of Rous sarcoma virus were temperature-sensitive for growth in milk . The failure of cells to adhere to the substratum in milk-supplemented medium suggested that milk might be deficient in attachment factors for fibroblasts . When the attachment of fibroblastic cells in milk-supplemented medium was facilitated by pretreating culture dishes with fibronectin, (a) transformed cells grew attached rather than in suspension, (b) normal cell lines attached and grew to confluence, and (c) normal cell strains adhered and survived but did not exhibit appreciable cell proliferation.The physiological function of milk is to provide substances important to the nutrition, growth, and development of the newborn. In addition, mammalian milk contains viable cells (1, 2) . Both human and bovine milks contain up to 4 X 10 6 cells/ml, primarily macrophages, T lymphocytes, and B lymphocytes. In previous studies, we demonstrated that both human (3, 4) and bovine (5) milks contain polypeptide growth factors that stimulate DNA synthesis and cell division of confluent cultures of fibroblasts. The presence of mitogens and viable cells in milk prompted us to evaluate milk as a replacement for serum in cell culture. In an initial report, we demonstrated that milk could support the long-term growth of epithelial cells in culture (6). Only colostrum, a highly enriched milk produced by mammals in the first day or two after birth of the newborn, was effective in supporting epithelial cell proliferation. In addition, colostrum was selective, supporting the growth of epithelial cells but not fibroblasts. When plated in either colostrum or regular milk, fibroblasts adhered very poorly to the substratum, suggesting that milk lacks factors that are necessary for the attachment of fibroblasts. This ob-294 servation prompted us to investigate whether milk would be selective for the growth of cells that do not require attachment to the substratum to proliferate. In this report, we show that transformed fibroblasts grow readily in milk-supplemented medium as suspension cultures . On the other hand, normal fibroblasts do not grow in milk unless the attachment of cells is mediated by an adhesion-promoting factor, such as fibronectin . MATERIALS AND METHODS Cell CultureRat embryo cultures (LRI) prepared fro...

Mare's Colostrum Globules Stimulate Fibroblast GrowthIn Vitro: A Biochemical Study

“…Fibroblasts are a cell type that is representative of the skin and have been widely studied. [43][44][45] It was therefore decided to investigate them further, by comparing the activating effects of mare's colostrum and milk on fibroblast proliferation.…”

Peptides from milk protein hydrolysates to improve the growth of human keratinocytes in culture

“…Supplementation of cell culture media with milk, colostrum and whey extracts has been reported to improve the growth rate of many cell types, mostly epithelial cells from kidney and skin, but also fibroblasts, smooth muscle cells and chondrocytes from cartilage (Klagsbrun, 1980;Sereni & Baserga, 1981;Steimer & Klagsbrun, 1981;Belford et al, 1995;Francis, Regester, Webb, & Ballard, 1995). The activity of milk to promote cell growth has been related to many of its constituents, including nutrients such as minerals and vitamins (Alais, 1984) and amino acids such as glutamine (Rombeau, 1990), hormones (Nabet, Belleville-Nabet, & Linden, 1991) and growth factors including HMGF (Human Growth Factor)-I, -II and -III (Shing & Klagsbrun, 1984), IGF (Insulinlike Growth Factor) (Campbell & Baumrucker, 1989), TGF (Tumour Growth Factor) (Cox & Burk, 1991) and FGF (Fibroblast Growth Factor) .…”

Inhibition of mouse x mouse hybridoma growth by milk and colostrum

“…Propagation of non-hybridoma cell lines in colostrum or milk supplemented media has also been attempted by other workers (Klagsbrun, 1980;Steimer and Klagsbrun, 1981). Inhibition by colostrum was reported to be cell line de pende nt.…”

“…Colostrum did not support human and rat fibroblasts in long-term culture, whereas canine kidney epithelial cell (MOCK) grew exponentially (Klagsbrun, 1980). Steimer and Klagsbrun (1981) suggested that milk was deficient in attachment factor su ch as fibronectin. This may not be the only reason for inhibition of VI0103 hybridoma by colostrum since coating of 24-well plate with collagen did not improve binding and multiplication of hybridoma.…”