TY - JOUR
T1 - Arachidonic acid directly activates members of the mitogen-activated protein kinase superfamily in rabbit proximal tubule cells
AU - Alexander, Larry D.
AU - Cui, Xiao Lan
AU - Falck, J R
AU - Douglas, Janice G.
N1 - Funding Information:
This work was supported by the following National Institutes of Health grants: HL41618 (Douglas), HL 07714 (Douglas), GM31278 (Falck), and a UNCF/Merck Postdoctoral Research Fellowship (Alexander). We thank Ns. Nnennaya Nkemere and Ms. Pearl Whitley for technical assistance in isolation and tissue culture of rabbit proximal tubule cells and Dr. Michal Schwartzman for valuable discussions. We are indebted to Dr. Byron Kemper for gifts of pCMV plasmids. This work was presented in part at the Annual Meeting of the American Heart Association (Dallas, TX, USA, November, 1998) and at the Experimental Biology Scientific Meeting (New Orleans, LA, USA, April, 1997).
PY - 2001
Y1 - 2001
N2 - Background. To explore the roles of eicosanoids in arachidonic acid-induced mitogen-activated protein kinase (MAPK) signal transduction, we have shown that exposure of proximal tubular cells to arachidonic acid induces phosphorylation of c-Jun NH2-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK), two members of the MAPK superfamily. We observed that ketoconazole, an inhibitor of the cytochrome P450 pathway, blocked ERK but not JNK activation. Methods. Direct regulation of arachidonic acid on mitogen-activated protein kinase (MAPK) signaling pathways was evaluated more directly by utilizing specific enzyme inhibitors of the cytochrome P450 metabolic pathway and by comparing the relative efficacy of arachidonic acid versus its cytochrome P450 metabolites (exogenous and endogenous), eicosatetraynoic acid (ETYA), and other fatty acids on the phosphorylation of members of the MAPK superfamily (ERKs, JNK, and p38MAPK), by utilizing early passage rabbit proximal tubular epithelial cells. Results. Arachidonic acid activated p38MAPK, a third member of the MAPK superfamily, in a time- and concentration-dependent manner. Studies designed to evaluate the ability of arachidonic acid and its cytochrome P450 metabolites (endogenously and exogenously) to stimulate ERKs, JNK, and p38MAPK found four conclusions. First, the metabolites of arachidonic acid generated endogenously by cytochrome P450 2C1 significantly augmented basal ERK activity, whereas the metabolites generated by the 2C2 isozyme significantly augmented basal p38MAPK activity. However, their effects were less profound than arachidonic acid itself. In contrast, there were no significant effects with transfection of either isozyme on basal JNK activity. Second, a variety of exogenous cytochrome P450 products were less potent than arachidonic acid on a molar basis in stimulating the activity of all three MAPKs. Third, ketoconazole and 17-octadecynoic acid, inhibitors of the cytochrome P450 pathway, as well as PPOH and DDMS, inhibitors of the epoxygenase and ω-hydroxylase pathways, respectively, failed to significantly reduce the effects of arachidonic acid to activate ERK and p38MAPK (JNK was not evaluated). Finally, arachidonic acid, its inactive analog ETYA, and other fatty acids with differing chain lengths and degrees of saturation stimulated the activity of all three MAPKs. Conclusions. These observations substantiate a role for arachidonic acid and other fatty acids in signaling linked to the MAPK superfamily in rabbit proximal tubular epithelium without the necessity of conversion to cytochrome P450 metabolites.
AB - Background. To explore the roles of eicosanoids in arachidonic acid-induced mitogen-activated protein kinase (MAPK) signal transduction, we have shown that exposure of proximal tubular cells to arachidonic acid induces phosphorylation of c-Jun NH2-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK), two members of the MAPK superfamily. We observed that ketoconazole, an inhibitor of the cytochrome P450 pathway, blocked ERK but not JNK activation. Methods. Direct regulation of arachidonic acid on mitogen-activated protein kinase (MAPK) signaling pathways was evaluated more directly by utilizing specific enzyme inhibitors of the cytochrome P450 metabolic pathway and by comparing the relative efficacy of arachidonic acid versus its cytochrome P450 metabolites (exogenous and endogenous), eicosatetraynoic acid (ETYA), and other fatty acids on the phosphorylation of members of the MAPK superfamily (ERKs, JNK, and p38MAPK), by utilizing early passage rabbit proximal tubular epithelial cells. Results. Arachidonic acid activated p38MAPK, a third member of the MAPK superfamily, in a time- and concentration-dependent manner. Studies designed to evaluate the ability of arachidonic acid and its cytochrome P450 metabolites (endogenously and exogenously) to stimulate ERKs, JNK, and p38MAPK found four conclusions. First, the metabolites of arachidonic acid generated endogenously by cytochrome P450 2C1 significantly augmented basal ERK activity, whereas the metabolites generated by the 2C2 isozyme significantly augmented basal p38MAPK activity. However, their effects were less profound than arachidonic acid itself. In contrast, there were no significant effects with transfection of either isozyme on basal JNK activity. Second, a variety of exogenous cytochrome P450 products were less potent than arachidonic acid on a molar basis in stimulating the activity of all three MAPKs. Third, ketoconazole and 17-octadecynoic acid, inhibitors of the cytochrome P450 pathway, as well as PPOH and DDMS, inhibitors of the epoxygenase and ω-hydroxylase pathways, respectively, failed to significantly reduce the effects of arachidonic acid to activate ERK and p38MAPK (JNK was not evaluated). Finally, arachidonic acid, its inactive analog ETYA, and other fatty acids with differing chain lengths and degrees of saturation stimulated the activity of all three MAPKs. Conclusions. These observations substantiate a role for arachidonic acid and other fatty acids in signaling linked to the MAPK superfamily in rabbit proximal tubular epithelium without the necessity of conversion to cytochrome P450 metabolites.
KW - Cytochrome P450
KW - Eicosanoids
KW - Extracellular signal-regulated kinase
KW - Ketoconazole
KW - Signal transduction
KW - c-Jun NH-terminal kinase
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U2 - 10.1046/j.1523-1755.2001.0590062039.x
DO - 10.1046/j.1523-1755.2001.0590062039.x
M3 - Article
C2 - 11380805
AN - SCOPUS:0035008791
SN - 0085-2538
VL - 59
SP - 2039
EP - 2053
JO - Kidney international
JF - Kidney international
IS - 6
ER -