RESUMEN
Herein, an efficient synthesis of BODIPY-α-Galactosylceramide 3, which can be used to study the cellular uptake of the potent immunostimulatory parent compound α-Galactosylceramide, is reported. Key in our synthetic strategy is the six-step synthesis of the core BODIPY scaffold (64% yield overall) and its quantitative conversion to an N-hydroxysuccinimidyl ester to facilitate conjugation and purification of the target glycolipid. For the preparation of the core of the glycolipid, the solubility of the lipid acceptor proved to be critical. The ability of BODIPY-αGalCer 3 to activate invariant natural killer cells was then demonstrated in vitro.
Asunto(s)
Compuestos de Boro/química , Compuestos de Boro/síntesis química , Galactosilceramidas/química , Galactosilceramidas/metabolismo , Sondas Moleculares/química , Sondas Moleculares/síntesis química , Transporte Biológico , Línea Celular , Técnicas de Química SintéticaRESUMEN
The C22 and C26 trehalose monoesters, each containing a single acyl chain, were synthesised in good overall yields and found to activate macrophages in a Mincle-dependent manner. The activities of the monoesters paralleled those of their diester counterparts, and both mono- and diesters could activate the immune response in the absence of priming. This is the first time that trehalose monoesters have been found to activate macrophages, and these studies thus provide an important framework for the rational design of other Mincle agonists.
Asunto(s)
Glucolípidos/química , Macrófagos/efectos de los fármacos , Trehalosa/análogos & derivados , Trehalosa/química , Animales , Células de la Médula Ósea/citología , Ésteres/farmacología , Glucolípidos/síntesis química , Glucolípidos/farmacología , Interleucina-6/metabolismo , Lectinas Tipo C/deficiencia , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Macrófagos/inmunología , Macrófagos/metabolismo , Proteínas de la Membrana/deficiencia , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Óxido Nítrico/metabolismo , Receptor Toll-Like 2/deficiencia , Receptor Toll-Like 2/genética , Receptor Toll-Like 2/metabolismo , Receptor Toll-Like 4/deficiencia , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/metabolismo , Trehalosa/síntesis química , Trehalosa/farmacologíaRESUMEN
In this study, the role of lipoteichoic acid (LTA) anchors in the activation of the innate immune response was investigated through the chemical synthesis of a series of LTA derivatives and the determination of their ability to induce NO production in bone marrow-derived macrophages (BMM). To this end, an efficient synthesis of the sn-3-O-(α-D-galactofuranosyl)-1,2-di-O-acylglycerol LTA core was developed, which was then used as a key structure to produce both phosphate and glycerylphosphate-funtionalised LTA anchors, as well as galactofuranosyldiglycerides with different fatty acid chain lengths. With a series of LTA anchors in hand, we then determined the effect of these glycolipids on the innate immune response by exploring their capacity to activate macrophages. Here, we report that several of the LTA-derivatives were able to induce NO production by BMMs. In general, the unnatural (sn-1) core glycolipid anchors showed lower levels of activity than the corresponding natural (sn-3) analogues, and the activity of the glycolipids also appears to be dependent on the length of lipid present, with an optimum lipid length of C20 for the sn-3 derivatives. Interestingly, a triacylated anchor and the 6-O-phosphorylated anchor, showed only modest activity, while the 6-O-glycerophosphorylated derivative was unable to induce NO production. Taken as a whole, our results highlight the subtle effects that glycolipid length can have on the ability to activate BMMs.
Asunto(s)
Glucolípidos/química , Lipopolisacáridos/farmacología , Streptococcus/química , Ácidos Teicoicos/farmacología , Animales , Glucolípidos/metabolismo , Lipopolisacáridos/síntesis química , Lipopolisacáridos/química , Macrófagos/química , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Óxido Nítrico/biosíntesis , Ácidos Teicoicos/síntesis química , Ácidos Teicoicos/químicaRESUMEN
Going to any length? Trehalose diesters of various chain lengths have been synthesised in order to determine the effect of lipid length on innate immune recognition, as determined by NO and cytokine production by macrophages. In this work, we show that longer lipids (C(20) -C(26)) are required for macrophage activation, with C(22) giving optimal activity.
Asunto(s)
Inmunidad Innata , Lípidos/química , Macrófagos/metabolismo , Trehalosa/química , Animales , Factores Cordón/química , Factores Cordón/inmunología , Citocinas/metabolismo , Lípidos/inmunología , Activación de Macrófagos , Macrófagos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Óxido Nítrico/metabolismo , Trehalosa/inmunologíaRESUMEN
A highly efficient synthesis of the biologically important fluorescent probe dansyl α-GalCer is presented. Key in our strategy is the incorporation of the fluorescent dansyl group at an early stage in the synthesis to facilitate in the monitoring and purification of intermediates via TLC and flash column chromatography, respectively, and the use of a high yielding α-selective glycosylation reaction between the phytosphingosine lipid and a galactosyl iodide donor. The ability of dansyl α-GalCer to activate iNKT cells and to serve as a fluorescent marker for the uptake of glycolipid by dendritic cells is also presented.