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Masaji Murakami, Yukitaka Ushio, Yosuke Mihara, Jun-Ichi Kuratsu, Seikoh Horiuchi, and Yoshimasa Morino

✓ Low-density lipoprotein (LDL) is a carrier of the cholesterol found in human plasma. Cells utilize cholesterol for membrane synthesis by taking up LDL via receptor-mediated endocytosis. In the present study, interactions of LDL with human malignant glioma cell lines (U-251MG and KMG-5) were investigated biochemically and morphologically. The LDL, labeled with the fluorescent dyes 1, l′-dioctadecy1–3,3,3′,3′-tetramethylindocarbocyanine (DiI) and fluorescein isothiocyanate (FITC), was internalized by both cell processes and cell bodies. Reductive methylation of DiI-labeled LDL, which abolishes the ability of the cell to bind to the LDL receptor, prevented the internalization of the cholesterol moiety of LDL. Cellular binding of 125I-LDL to U-251MG cells at 4°C revealed the presence of a specific saturable-associated receptor (dissociation constant (Kd) approximately 38 µg/ml). Endocytic uptake of 125I-LDL or 3H-cholesterol oleate-labeled LDL (3H-LDL) at 37°C demonstrated the cell-associated 125I-LDL and 3H-LDL increase. The intracellular degradation of protein moiety increased linearly with time. Reductive methylation of 3H-LDL led to a remarkable decrease in the cell-associated cholesterol moiety of LDL. The difference in uptake of the cholesterol moiety of LDL between U-251MG cells and KMG-5 cells showed that the U-251MG cells, which proliferate more actively than KMG-5 cells, take up more of the cholesterol moiety of LDL than do the KMG-5 cells.

Thus, LDL cholesterol seems to be endocytosed predominantly via the LDL receptor present on the plasma membrane of malignant glioma cells. In addition, for growth, these cells may require large amounts of the cholesterol moiety of LDL.