TY - JOUR
T1 - Role of arachidonyl triglycerides within lipid bodies in eicosanoid formation by human polymorphonuclear cells
AU - Johnson, Margaret M.
AU - Vaughn, Benjamin
AU - Triggiani, Massimo
AU - Swan, Dennis D.
AU - Fonteh, Alfred N.
AU - Chilton, Floyd H.
PY - 1999
Y1 - 1999
N2 - Increasing evidence suggests that the subcellular and glycerolipid localization of esterified arachidonic acid (AA) is a key factor in regulating its availability to lipases. The goal of the current study was to determine the potential of AA stored in triglycerides (TG) to serve as a substrate for lipases and 5-lipoxygenase during neutrophil (polymorphonuclear leukocytes, PMN) activation. PMN containing high concentrations of AA in TG were generated by culturing PMN in vitro with high concentrations of exogenous AA (eAA) for 12 h. Cellular AA increased 2- and 4-fold in PMNs incubated with 5 and 20 μM AA, respectively, and this increase was almost exclusively observed in neutral lipids (NL). Further analysis revealed that 88% of the AA in the NL fraction was associated with TG. Subsequent experiments were designed to determine whether this AA in TG could be mobilized and metabolized to eicosanoids during cell activation. TG pools of AA were increased as previously described and then PMN were stimulated with ionophore, A23187. In contrast to the 43-fold increase in TG AA after eAA loading (20 μM), free AA increased by only 1.9-fold after cell stimulation. Similarly, leukotriene (LT)B4 production increased only 2-fold after loading TG with large quantities of AA. The magnitude of increase in free AA released and in LTB4 formation was similar to the magnitude of increase in AA mass in phospholipase (PL), suggesting that PL, and not TG, served as the source of released AA and subsequent product generation. To confirm that AA in TG did not serve as a source tor eicosanoid production, cellular pools of AA were differentially labeled with [14C]AA and [3H]AA, and the [3H]AA-to-[14C]AA ratio of LTB4 and 20-hydroxyl LTB4 produced during cell stimulation was measured. The [3H]AA/[14C]AA ratios of LTs were markedly different from the ratios in TG, thus providing further evidence that AA pools in TG are not a major source of AA for LT generation.
AB - Increasing evidence suggests that the subcellular and glycerolipid localization of esterified arachidonic acid (AA) is a key factor in regulating its availability to lipases. The goal of the current study was to determine the potential of AA stored in triglycerides (TG) to serve as a substrate for lipases and 5-lipoxygenase during neutrophil (polymorphonuclear leukocytes, PMN) activation. PMN containing high concentrations of AA in TG were generated by culturing PMN in vitro with high concentrations of exogenous AA (eAA) for 12 h. Cellular AA increased 2- and 4-fold in PMNs incubated with 5 and 20 μM AA, respectively, and this increase was almost exclusively observed in neutral lipids (NL). Further analysis revealed that 88% of the AA in the NL fraction was associated with TG. Subsequent experiments were designed to determine whether this AA in TG could be mobilized and metabolized to eicosanoids during cell activation. TG pools of AA were increased as previously described and then PMN were stimulated with ionophore, A23187. In contrast to the 43-fold increase in TG AA after eAA loading (20 μM), free AA increased by only 1.9-fold after cell stimulation. Similarly, leukotriene (LT)B4 production increased only 2-fold after loading TG with large quantities of AA. The magnitude of increase in free AA released and in LTB4 formation was similar to the magnitude of increase in AA mass in phospholipase (PL), suggesting that PL, and not TG, served as the source of released AA and subsequent product generation. To confirm that AA in TG did not serve as a source tor eicosanoid production, cellular pools of AA were differentially labeled with [14C]AA and [3H]AA, and the [3H]AA-to-[14C]AA ratio of LTB4 and 20-hydroxyl LTB4 produced during cell stimulation was measured. The [3H]AA/[14C]AA ratios of LTs were markedly different from the ratios in TG, thus providing further evidence that AA pools in TG are not a major source of AA for LT generation.
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U2 - 10.1165/ajrcmb.21.2.3489
DO - 10.1165/ajrcmb.21.2.3489
M3 - Article
C2 - 10423409
AN - SCOPUS:0033172895
SN - 1044-1549
VL - 21
SP - 253
EP - 258
JO - American journal of respiratory cell and molecular biology
JF - American journal of respiratory cell and molecular biology
IS - 2
ER -