Inhibition of the intracellular Ca2l-ATPase by thapsigargin causes the inositol 1,4,5trisphosphate (InsP3)-sensitive Ca2 pools of DDTiMF-2 smooth muscle cells to empty. Pool emptying causes cells to cease division and enter a stable, quiescent Go-like state. Although thapsigargin blocks pumps essentially irreversibly, high serum treatment of these pool-depleted quiescent cells induces synthesis of new Ca2+ pump protein, refilling of InsPs-sensitive Ca2 pools, and re-entry of cells into the cell cycle. Here we reveal th£ this activity is mediated by specific lipid metabolites, namely eicosanoids. The essential fatty acids linolenic acid, linoleic acid, and arachidonic acid (AA) each induced recovery of Ca pools and re-entry of cells into the cell cycle. Each essential fatty acid induced recovery with an EC of 5 nM; the action of each was dependent on protein synthesis. All non-essential fatty acids and growth factors tested did not promote recovery. As each essential fatty acid can serve as a precursor to eicosanoids, each synthetic pathway was investigated for a possible role in the recovery process. Inhibitors of the prostanoid and lipoxygenase metabolism pathways had no effect on essential fatty acid-induced Ca2pool or growth recovery. However, the cytochrome P-450 inhibitors SKF-525A, metyrapone, and nordihydroguaiaretic acid each prevented the action of AA. Importantly, treatment of quiescent cells with either of 8,9- or 11,12-epoxyeicosatrienoic acid (EET), two of the primary four regiospecific cytochrome P-450 metabolites of arachidonic acid, also induced recovery. However, the other regiospecific EETs and the dihydroxyeicosatrienoic acid metabolites of the functional EETs were unable to induce recovery. Evidence suggests that these EETs do not work via the action of protein kinase C or cyclic nucleotides. These results are important in understanding the role of cytochrome P-450 derived eicosanoids in the regulation of the expression of intracellular Ca2 pumps and hence in the regulation of cellular function. (Supported by NIH grant HL55426 and NSF grant MCB9307746.).
|Original language||English (US)|
|State||Published - Dec 1 1996|
All Science Journal Classification (ASJC) codes
- Molecular Biology