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Platelet-derived growth factor stimulates synthesis of 1,2-diacylglycerol from monoacylglycerol in Balb/c 3T3 cells.

ABSTRACT: 1,2-Diacylglycerol (1,2,-DAG) plays an important role in the protein kinase C-mediated signal-transduction system. Several reports have shown that 1,2-DAG is generated through various pathways other than classical phospholipid hydrolysis. We observed a rapid incorporation of [3H]myristate into 1,2-DAG in platelet-derived-growth-factor (PDGF)-treated Balb/c 3T3 cells. [14C]Palmitate was similarly incorporated into 1,2-DAG. The effect of PDGF, which was inhibited by cycloheximide, became maximal after 60 min treatment with PDGF, and disappeared 300 min after removal of PDGF. Treatment with triacylglycerol lipase revealed that labelled saturated fatty acid was incorporated into the sn-1 position. PDGF barely stimulated incorporation of [3H]glycerol or [14C]glucose into 1,2-DAG. Incorporation of [3H]myristate into 1,2-DAG preceded that into triacyglycerol and phospholipids. These results suggest that synthesis of 1,2-DAG from monoacylglycerol is enhanced in PDGF-treated cells. However, there is no significant difference in the activity of monoacylglycerol acyltransferase measured in vitro in quiescent and PDGF-treated cells. The reason for this discrepancy is discussed.

SUBMITTER: Hata Y

PROVIDER: S-EPMC1133364 | biostudies-other | 1989 Sep

REPOSITORIES: biostudies-other

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Project description:IntroductionThe objective of this study was to model the effects of transforming growth factor beta (TGF-beta) and platelet-derived growth factor (PDGF), both present in rheumatoid arthritis (RA) synovia, on the behavior of fibroblast-like synoviocytes (FLS) in response to pro-inflammatory cytokine (interleukin (IL)1beta, tumor necrosis factor-alpha (TNFalpha)) challenge.MethodsGene and protein expression by fibroblast-like synoviocytes in vitro was studied by quantitative Polymerase Chain Reaction (qPCR), ELISA and multiplex bead cytokine assays. Intracellular signaling pathway activation was determined by Western blot for phospho-kinases and the use of specific inhibitors.ResultsIn combination, TGF-beta and PDGF (2GF) synergistically augmented TNFalpha- or IL1beta-induced matrix metalloproteinase 3 (MMP3), IL6, IL8, and macrophage inflammatory protein 1 alpha (MIP1alpha) secretion by FLS. Other FLS-derived mediators remained unaffected. Individually, neither growth factor significantly potentiated TNFalpha or IL1beta-induced MMP3 secretion, and only slightly enhanced IL6. The effect of 2GF on TNFalpha-induced gene expression was transcriptionally mediated; blocked by imatinib mesylate; and occurred even if 2GF was added as much as four hours prior to TNFalpha. In addition, a 15-minute pulse of 2GF four hours prior to TNFalpha stimulation yielded a synergistic response. The extracellular-signal-regulated kinase (ERK) and phosphoinositide 3-kinase (PI3K) signaling pathways were induced for at least four hours by 2GF, as demonstrated by persistently upregulated levels of phospho-Akt and phospho-ERK. However, pharmacologic inhibitor studies demonstrated that the potentiating action of 2GF was dependent on PI3 kinase only, and not on ERK.ConclusionsThe combination of PDGF and TGF-beta dramatically potentiates FLS response to cytokines in a receptor-mediated and PI3 kinase-dependent fashion. These data suggest that 2GF contribute to synovitis by directing synovial fibroblasts toward a more aggressive phenotype in response to TNFalpha. Therefore, inhibition of growth factor signaling may constitute a complementary therapeutic approach to cytokine-targeted treatments for RA.

Project description:Bacterial microcompartments (BMCs) are proteinaceous organelles that optimize specific metabolic pathways referred to as metabolosomes involving transient production of toxic volatile metabolites such as aldehydes. Previous bioinformatics analysis predicted the presence of BMCs in 23 bacterial phyla including foodborne pathogens and a link with gene clusters for the utilization of host-derived substrates such as 1,2-propanediol utilization, i.e., the Pdu cluster. Although, transcriptional regulation of the Pdu cluster and its role in Listeria monocytogenes virulence in animal models have recently been reported, the experimental identification and the physiological role of BMCs in L. monocytogenes is still unexplored. Here, we ask whether BMCs could enable utilization of 1,2-propanediol (Pd) in L. monocytogenes under anaerobic conditions. Using L. monocytogenes EGDe as a model strain, we could demonstrate efficient utilization of Pd with concomitant production of 1-propanol and propionate after 24 h of anaerobic growth, while the utilization was significantly reduced in aerobic conditions. In line with this, expression of genes encoding predicted shell proteins and the signature enzyme propanediol dehydratase is upregulated more than 20-fold in cells anaerobically grown in Pdu-induced versus non-induced control conditions. Additional proteomics analysis confirmed the presence of BMC shell proteins and Pdu enzymes in cells that show active degradation of Pd. Furthermore, using transmission electron microscopy, BMC structures have been detected in these cells linking gene expression, protein composition, and BMCs to activation of the Pdu cluster in anaerobic growth of L. monocytogenes. Studies in defined minimal medium with Pd as an energy source showed a significant increase in cell numbers, indicating that Pdu and the predicted generation of ATP in the conversion of propionyl-phosphate to the end product propionate can support anaerobic growth of L. monocytogenes. Our findings may suggest a role for BMC-dependent utilization of Pd in L. monocytogenes growth, transmission, and interaction with the human host.

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Platelet-derived growth factor stimulates synthesis of 1,2-diacylglycerol from monoacylglycerol in Balb/c 3T3 cells.

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