Regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and the esterification of cholesterol in human long term lymphoid cell lines

Abstract

The regulation of the rate-controlling enzyme in cholesterol biosynthesis and of the incorporation of [14C]oleate into cholesterol esters were studied in established lymphoid cell lines from normal subjects and compared with that of eight patients with genetic abnormalities of lipid metabolism. The activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase, the rate-controlling enzyme in cholesterol biosynthesis, increases in lymphoid cell lines derived from normal subjects after the culture medium is changed to a lipid deficient medium and reaches peak activity after 48 hr. The addition of whole serum and of low density lipoproteins to cell lines derived from normal subjects suppressed 3-hydroxy-3-methylglutaryl coenzyme A reductase activity by 50%, but failed (almost completely) to suppress the activity in the lymphoid cell lines derived from two patients with homozygous familial hypercholesterolemia. When 7-ketocholesterol was added, the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase was markedly suppressed in both normal and abnormal lymphoid cell lines. Lymphoid cell lines derived from patients presumably heterozygous for familial hypercholesterolemia were difficult to distinguish from normal cells in these studies. The incorporation of [14C]oleate into the fatty acid fraction of cholesteryl esters was stimulated by the addition of the low density lipoproteins to the culture media of the lymphoid cell lines derived from the normal human subjects. The lymphoid cell lines derived from the patients with homozygous familial hypercholesterolemia showed no increase in [14C]oleate incorporation into cholesteryl esters even when a fourfold amount of low density lipoprotein was added to the media; a modest increase in [14C]oleate incorporation was observed in lymphoid cell lines from patients with heterozygous familial hypercholesterolemia. The results of these studies in lymphocyte cell lines are compared with the findings in cultured human fibroblasts obtained from normal subjects and from patients with homozygous familial hypercholesterolemia. Studies of the regulation of cholesterol biosynthesis in the apparently permanent lymphoid cell line maintained in suspension culture offer certain advantages over cultured skin fibroblasts, and, in addition, provide a second tissue for the study of genetic abnormalities from the same patient.

Reduction in cholesterol synthesis in response to serum starvation in lymphoblasts of a patient with Barth syndromeThis paper is one of a selection of papers published in this special issue entitled “Second International Symposium on Recent Advances in Basic, Clinical, and Social Medicine” and has undergone the Journal's usual peer review process.

“…Control and DTAZ1 lymphoblasts were incubated in the absence or presence of serum for 16 h, microsomal fractions were prepared, and HMGR enzyme activity was determined. The HMGR enzyme activity observed in these cells was within the range of enzyme activity previously reported in lymphoid cell lines (Kayden et al 1976). Upon serum removal, HMGR in vitro activity was elevated 50% in control lymphoblasts (Fig.…”

Low density lipoprotein receptor mutations in South African homozygous familial hypercholesterolemic patients.

“…The presence of functional LDL receptors on long-term phytohemagglutinin-activated lymphoid cell lines was detected by Kayden et al 24 and Ho et al 15 The latter workers also showed that cells from one homozygous FH patient were devoid of such receptors. Our studies, which analyzed a number of cell lines derived from unaffected subjects and FH patients, establish the validity of using virus-transformed lymphoblastoid cell lines to detect mutant LDL receptor alleles in human subjects and to quantify receptor-mediated LDL binding and metabolism.…”

Mutations affecting the binding, internalization, and lysosomal hydrolysis of low density lipoprotein in cultured human fibroblasts, lymphocytes, and aortic smooth muscle cells

“…All of the steps of the LDL pathway in human fibroblasts also have been identified in long-term human lymphoid cells maintained in suspension culture (23,24) and in human lymphocytes freshly isolated from the bloodstream (25,26). same constellation of secondary defects (i.e., defective LDL uptake and degradation, overproduction of cholesterol, and failure t o induce cholesteryl ester formation) as d o the receptor-negative FH homozygote fibroblasts.…”