HIGH-DENSITY-LIPOPROTEIN AND LOW-DENSITY LIPOPROTEIN-MEDIATED SIGNAL-TRANSDUCTION IN CULTURED HUMAN SKIN FIBROBLASTS

The signalling mechanisms whereby high-density lipoproteins (HDL) and low-density lipoproteins (LDL) affect a number of cellular functions i n fibroblasts are unclear. This study has analyzed the influence of HD L(3) and LDL on the phosphatidylinositol specific phospholipase C path way in human skin fibroblasts. Exposure of myo-[2-H-3]-inositol prelab elled fibroblasts to HDL(3) or LDL elicited major increases in IP1 and minor increases in IP2 and IP3 within 30 s. In fura-2 loaded suspende d fibroblasts, HDL(3) and LDL increased intracellular Ca2+ concentrati ons ([Ca2+](i)) with comparable rapid, transient kinetics. The dose-pr ofiles for HDL(3)- and LDL-induced increases in [Ca2+](i) were also co mparable, with half-maximally and maximally effective concentrations b eing approximate to 15 mu g/mL and approximate to 50 mu g/mL, respecti vely. HDL(3)- and LDL-induced increases in [Ca2+](i) were diminished b y approximate to 60% (vs. control fibroblasts) in thapsigargin-pretrea ted fibroblasts, indicating that release of Ca2+ from intracellular po ols is the major contributor toward lipoprotein-induced increases in [ Ca2+](i). Pertussis toxin-pretreatment of cells completely abolished l ipoprotein induced Ca2+-transient, indicating the involvement of a gua nine nucleotide-binding protein in the signalling process. In [3H]-pal mitic acid-prelabelled fibroblasts, both HDL(3) and LDL were observed to stimulate production of DAG. Activation of protein kinase C (PKC) w as analysed by determining the cytosol-to-membrane translocation of bo th enzymatic activity and immunoreactivity of specific PKC isoforms (a lpha, delta, epsilon, and zeta). Stimulation with HDL(3) and LDL evoke d a rapid (within 2.5 min) translocation of PKC activity, with PKC alp ha and PKC epsilon being the isoforms translocated. It is concluded th at HDL(3) and LDL acutely stimulate a phosphoinositide-specific phosph olipase C pathway in human skin fibroblasts. However, the specific cel l membrane events mediating this signal transduction remain to be furt her elucidated.