Structure-function analysis of Leishmania lipophosphoglycan. Distinct domains that mediate binding and inhibition of endothelial cell function
Ho, J.L.; Kim, H.K.; Sass, P.M.; He, S.; Geng, J.; Xu, H.; Zhu, B.; Turco, S.J.; Lo, S.K.
Journal of Immunology 157(7): 3013-3020
1996
ISSN/ISBN: 0022-1767 PMID: 8816410 Document Number: 464841
The effect of specific domains of Leishmania donovani lipophosphoglycan (LPG) on endothelial cell activation was investigated. Endotoxin (10 ng/ml, 4 h) consistently caused endothelium to increase monocyte adhesion (~20-fold) but this was completely blocked by LPG pretreatment (2 micro M, 2 h). LPG did not grossly suppress endothelial functions because tumour necrosis factor- alpha - and interleukin-1 beta -mediated adhesion toward monocytes was not affected. Four highly purified LPG fragments (repeating phosphodisaccharide (PGM), phosphoglycan, phosphosaccharide core-lyso-alkyl-phosphatidylinositol (core-PI) and lyso-alkyl-phosphatidylinositol (lyso-PI)) were used to examine whether these fragments can independently inhibit endothelial adhesion, but neither the 4 fragments (2 micro M, 2 h) independently nor the co-addition of phosphoglycan and core-PI fragments blocked the endotoxin-mediated adhesion to monocytes. To determine whether the fragments can reverse the effect of intact LPG, endothelial cells were first pretreated with the LPG fragments (10 micro M, 15 minutes), followed by the addition of LPG (2 micro M). All 4 fragments fully reversed the effect of LPG. Simultaneous addition of LPG fragments and intact LPG caused only partial suppression (~45%), while the addition of LPG fragments 15 minutes later had no reversal effect. Flow cytometry revealed that only core-PI and lyso-PI competitively inhibited (~30%) LPG binding. Conversely, LPG competed with the binding of [superscript 3H]lyso-PI (~30%). Monoclonal antibody against the PGM reversed (~70%) the effect of LPG. Thus, the lyso-PI domain on LPG mediates binding to endothelial cells, whereas the PGM domain mediates the cell inhibitory effect.