Flow cytometric assessment of LDL receptor activity in peripheral blood mononuclear cells compared to gene mutation detection in diagnosis of heterozygous familial hypercholesterolemia

BACKGROUND Studies indicate that human peripheral blood mononuclear cells mirror low-density lipoprotein (LDL) receptor activity of other cells in the body. To measure LDL receptor activity in patients with heterozygous familial hypercholesterolemia (FH), we prepared peripheral blood mononuclear cells from individuals with molecularly verified LDL receptor defective (Trp66-Gly mutation, n = 18) or receptor negative (Trp23-stop mutation, n = 17) heterozygous FH and from healthy individuals (n = 24). METHODS The cells were stimulated to express maximum LDL receptor by preincubation in lipoprotein-free medium. They were then incubated at 4 degrees or 37 degrees C with fluorescently conjugated LDL (DiI-LDL). T-lymphocytes and monocytes were identified by fluorescently conjugated monoclonal antibodies. DiI-LDL bound (at 4 degrees C) or internalized (at 37 degrees C) by the cells was measured using flow cytometry. Knowing the LDL receptor gene mutation of the FH patients allowed us to compare the diagnostic capability of our functional assay with the DNA diagnosis. RESULTS The diagnostic accuracy did not allow our assay to be used for diagnosis of individual cases of heterozygous FH. CONCLUSIONS We suggest that our two-color fluorescence flow cytometry assay can be used to characterize functionally gene mutations causing LDL receptor dysfunction in patients with heterozygous FH.