title: Improved detection of familial hypercholesterolemia by determining low density lipoprotein receptor expression in mitogen-induced proliferating lymphocytes.
abstract: In view of the presence of some 190 mutations in the low density lipoprotein receptor (LDL-R) gene and a lack of simple detection methods, we have developed an improved assay system for detecting familial hypercholesterolemia (FH) using mitogen-induced proliferating lymphocytes. Freshly isolated mononuclear cells were cultured for 3 days in RPMI 1640 supplemented with 10% human lipoprotein-deficient serum (LPDS) and 1% phytohemagglutinin (PHA). LDL-R expression was measured by flow cytometry using a monoclonal anti-LDL-R antibody or DiI-LDL. Mitogenic responses were monitored by cell size (FSC), interleukin-2 receptor (IL2-R) expression, and stimulation index (SI). The LDL-R expression in PHA-stimulated lymphocytes was significantly higher than lymphocytes or monocytes cultured without PHA (15.2- and 3.6-fold, respectively). The gradation of the LDL-R expression was highly correlated to FSC, IL2-R expression, and SI (r > 0.9 in each case). However, no difference in FSC, IL2-R expression, or SI existed between 30 clinically diagnosed FH and 42 normolipemic control subjects. The significantly lower LDL-R expression in the FH group (45.2 +/- 15.3% versus 100 +/- 14.1%; unpaired t test, P < 0.0001) indicated the presence of genetic defects. Normocholesterolemic first degree relatives and non-FH hypercholesterolemic subjects demonstrated normal LDL-R expression as did the controls. The assay carries an efficiency of 97% and both sensitivity and specificity of 98.5%. Measurement of low density lipoprotein receptor expression in phytohemagglutinin- and lipoprotein-deficient serum-stimulated lymphocytes offers a simple method for detecting familial hypercholesterolemia with improved sensitivity.
authors.affiliation: Department of Pediatrics and Medical Genetics, University of Calgary, Health Sciences Centre, Calgary, Alberta, Canada T2N 4N1.
authors.forename: P C
authors.initials: PC
authors.lastname: Chan
authors.forename: A
authors.initials: A
authors.lastname: Edwards
authors.forename: R
authors.initials: R
authors.lastname: Lafrenière
authors.forename: H G
authors.initials: HG
authors.lastname: Parsons
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D002469:Cell Separation; D002648:Child; D005260:Female; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D006938:Hyperlipoproteinemia Type II / Q000175:diagnosis*; D018950:Lipopolysaccharide Receptors / Q000032:analysis; D008213:Lymphocyte Activation; D008214:Lymphocytes / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D008934:Mitogens / Q000494:pharmacology*; D015375:Receptors, Interleukin-2 / Q000032:analysis; D011973:Receptors, LDL / Q000096:biosynthesis*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D015854:Up-Regulation
chemical_list: D018950:Lipopolysaccharide Receptors; D008934:Mitogens; D015375:Receptors, Interleukin-2; D011973:Receptors, LDL; D013755:Tetradecanoylphorbol Acetate