sn-1,2-Diacylglycerols and phorbol diesters: Uptake, metabolism, and subsequent assimilation of the diacylglycerol metabolites into complex lipids of cultured cells
✍ Scribed by Clement J. Welsh; Myles C. Cabot
- Book ID
- 102880215
- Publisher
- John Wiley and Sons
- Year
- 1987
- Tongue
- English
- Weight
- 944 KB
- Volume
- 35
- Category
- Article
- ISSN
- 0730-2312
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✦ Synopsis
The cell-permeable diacylglycerol mediators have been shown to mimic partially the effects of 12-0-tetradecanoylphorbol-13-acetate (TPA) on cultured cells. In order to evaluate the metabolic stability of the lipid mediators, several radiolabeled diacylglycerols were synthesized and their uptake and intracellular fate in cultured HL-60 (human promyelocytic leukemia) cells was compared with TPA. In addition to whole cell assessment, the stability of diacyl lipids and TPA was evaluated in a bufferjwater system and in the presence of serum and subcellular fractions. The compounds studied include 1,2-dioleoyl-sn-glycerol (DiOG), 1 -0leoy1-2-acetyl-sn-glycerol (OaG), 1-palmitoyl-2-acetyl-sn-glycerol (PaG), the ether-linked analog 1 -palmityl-2-acetyl-sn-glycerol (ePaG), and TPA. TPA was comparatively stable to lipase hydrolysis in all systems examined. First, the data show that within 5 min at pH 7.9, nearly 50% of the PaG (originally > 92% 1,2-isomer) had isomerized, and rapid formation of the 1,Zisomer also occurred with OaG and ePaG. The metabolism of OaG and PaG by serum hydrolases, using a reaction medium containing 10% serum, was chiefly by acetate hydrolysis; however, fatty acid was also liberated. After a 60-min incubation 68% of the [14C]OaG was converted, by serum enzymes, to monooleoylglycerol plus oleic acid. Heatinactivation of serum reduced the enzymatic formation of fatty acid by 60-70%. ePaG was also metabolized by serum enzymes, but the ether-linked alkylglycerol product was stable. The results of cell-free studies (postmitochondrial supernatant) showed that cellular enzymes were present that could, like serum, convert the diacylglycerols to monoacylglycerols and free fatty acids. Studies using cultured cells showed that radiolabeled OaG, PaG, and ePaG were rapidly taken up by the cells and metabolized. Labeled metabolic products from the diacylglycerols appeared, in a time-dependent manner, in cellular phospholipids and triacylglycer-01s. The results from experiments employing l-acyl-2-acetyl-sn-[3H]glycerol and [3H]acyl-2-acetyl-sn-glycerol indicate that the intracellular mode of mediator metabolism is via complete hydrolysis with subsequent incorporation of 3H-acyl