The method of isolation of primary cells and their subculture influences the expression of LDL receptors on pigeon and chicken embryo cells in culture.
Previous studies from this laboratory have shown that pigeon embryo fibroblasts in culture, derived from embryo explants, do not express LDL receptors. In contrast, pigeon peritoneal macrophages and pigeon hepatocytes in culture express LDL receptors. The presence of an active LDL receptor pathway in pigeons has been further confirmed by in vivo studies which showed that receptor-mediated mechanisms were responsible for greater than 80 percent of whole body LDL clearance. The purpose of the present study was to determine the factors responsible for the failure of explant-derived pigeon embryo cells to express LDL receptors. To address this problem, both chicken and pigeon embryo cells were studied. Primary pigeon and chicken embryo cells isolated by collagenase digestion expressed high levels of LDL receptor activity using 125I-labelled pigeon LDL as the ligand. Cells isolated from chicken embryo by enzyme digestion expressed an approximately 10-fold greater LDL receptor activity than cells derived from explants. Pigeon LDL and beta-VLDL, but not methyl-LDL, HDL or mammalian LDL, bound to a limited number of receptors on pigeon embryo cells with specificity and high affinity. Other properties of the pigeon LDL receptor were similar to the mammalian LDL receptor with the exception that the pigeon LDL receptor was resistant to the proteolytic enzyme, pronase. In both pigeon and chicken embryo cells LDL receptor activity was lost with subsequent passage in culture. As a result, the failure of explant derived pigeon cells to express LDL receptors appears to be the result of two factors, the use of explant rather than enzyme-isolated cells and the loss of any remaining LDL receptor activity as a result of passage in culture. Whether this difference between pigeon and mammalian cells with respect to LDL receptor activity is the result of differences in the molecular structure of the pigeon LDL receptor remains to be determined.
Sulistiyani
Department of Pathology, Bowman Gray School of Medicine, Wake Forest University, Winston-Salem, NC 27103.
St Clair
R W
RW
D000818:Animals; D001667:Binding, Competitive; D002642:Chick Embryo; D002738:Chloroquine / Q000494:pharmacology; D010856:Columbidae; D046508:Culture Techniques / Q000379:methods*; D004625:Embryo, Nonmammalian; D005347:Fibroblasts / Q000378:metabolism*; D007457:Iodine Radioisotopes; D008077:Lipoproteins, LDL / Q000378:metabolism; D003012:Microbial Collagenase; D011973:Receptors, LDL / Q000378:metabolism*
D007457:Iodine Radioisotopes; D008077:Lipoproteins, LDL; D011973:Receptors, LDL; D002738:Chloroquine; D003012:Microbial Collagenase