Publications by Year: 1996

1996
Finger EB, Puri KD, Alon R, Lawrence MB, von Andrian UH, Springer TA. Adhesion through L-selectin requires a threshold hydrodynamic shear. Nature. 1996;379 (6562) :266-9.Abstract
Selectins are cell adhesion molecules that bind carbohydrate ligands and promote interaction between leukocytes and the vessel wall in vascular shear flow. Selectin-ligand bonds have high mechanical strength, allowing initial tethering to the vessel wall through one or few bonds, and have fast on and off rates, permitting rolling in response to hydrodynamic drag. The L-selectin molecule on leukocytes binds to peripheral node addressin on high endothelial venules of lymph nodes to mediate leukocyte rolling and binds to a ligand on neutrophils to mediate rolling of leukocytes over one another. Here we describe a surprising mechanism for regulation of these interactions, both in vitro and in vivo. Shear above a critical threshold is required to promote and maintain rolling interactions through L-selectin, but not through E-selectin, P-selectin or VCAM-1. The shear threshold requirement for L-selectin may be physiologically important in low shear to prevent inappropriate aggregation of leukocytes and interaction with the vessel wall.
379266a0.pdf
Malý P, Thall A, Petryniak B, Rogers CE, Smith PL, Marks RM, Kelly RJ, Gersten KM, Cheng G, Saunders TL, et al. The alpha(1,3)fucosyltransferase Fuc-TVII controls leukocyte trafficking through an essential role in L-, E-, and P-selectin ligand biosynthesis. Cell. 1996;86 (4) :643-53.Abstract
alpha(1,3)Fucosylated oligosaccharides represent components of leukocyte counterreceptors for E- and P-selectins and of L-selectin ligands expressed by lymph node high endothelial venules (HEV). The identity of the alpha(1,3)fucosyltransferase(s) required for their expression has been uncertain, as has a requirement for alpha(1,3)fucosylation in HEV L-selectin ligand activity. We demonstrate here that mice deficient in alpha(1,3) fucosyltransferase Fuc-TVII exhibit a leukocyte adhesion deficiency characterized by absent leukocyte E- and P-selectin ligand activity and deficient HEV L-selectin ligand activity. Selectin ligand deficiency is distinguished by blood leukocytosis, impaired leukocyte extravasation in inflammation, and faulty lymphocyte homing. These observations demonstrate an essential role for Fuc-TVII in E-, P-, and L-selectin ligand biosynthesis and imply that this locus can control leukocyte trafficking in health and disease.
1-s2.0-s0092867400801373-main.pdf
von Andrian UH. Intravital microscopy of the peripheral lymph node microcirculation in mice. Microcirculation. 1996;3 (3) :287-300.Abstract
OBJECTIVE: The purpose of this study was to develop a model for microscopic in situ observation of the murine peripheral lymph node (LN) microcirculation and to characterize the function of the lymphocyte homing receptor L-selectin (CD62L) and the peripheral node addressin (PNAd). The latter is a high-affinity ligand for L-selectin in LN high endothelial venules (HEV). METHODS: The subiliac (superficial inguinal) LN was microsurgically dissected in anesthetized adult mice. The nodal microvascular architecture and venular hemodynamics were characterized by bright field and epifluorescence video microscopy. L1-2 pre-B cells that were either mock transfected (L1-2Vector) or stably transfected to express human L-selectin (L1-2L-selectin) were labeled fluorescently and injected into a feeding artery. Cell adhesion in LN venules was studied in both the presence and absence of neutralizing monoclonal antibodies (MAb) to PNAd and L-selectin. RESULTS: The preparation allowed a detailed analysis of hemodynamic parameters and leukocyte adhesion in LN microvessels. L1-2Vector cells did not interact with LN microvessels. In contrast, L1-2L-selectin cells rolled efficiently in venules but not in arterioles or capillaries. Rolling was most prominent in subcortical HEV (orders III to V) and was less frequent but consistently detectable in downstream medullary and hilus venules (orders I and II). Rolling interactions were abrogated by MAb DREG-56 to the lectin domain of L-selectin and were markedly reduced by the anti-PNAd MAb MECA-79. CONCLUSIONS: The present study develops a new intravital microscopy model for in vivo visualization of leukocyte interactions with microvessels in murine LN. The preparation permitted an analysis of biophysical and molecular mechanisms of leukocyte adhesion to high endothelial cells. The data support the concept that L-selectin and PNAd are the predominant receptor/ligand pair responsible for lymphocyte rolling in HEV. The model will be useful for high-resolution analysis of intra- and extravascular events in living LN.
Coxon A, Rieu P, Barkalow FJ, Askari S, Sharpe AH, von Andrian UH, Arnaout MA, Mayadas TN. A novel role for the beta 2 integrin CD11b/CD18 in neutrophil apoptosis: a homeostatic mechanism in inflammation. Immunity. 1996;5 (6) :653-66.Abstract
In mice selectively deficient in CD11b/CD18, a beta 2 integrin, chemoattractant-induced leukocyte adhesion to microvascular endothelium in vivo was reduced. Paradoxically, thioglycollate-induced neutrophil accumulation in the peritoneal cavity was increased and was associated with a significant delay in apoptosis of extravasated cells. The extravasated cells had a near absence of neutrophil phagocytosis and a reduction in oxygen free radical generation, which may contribute to the observed defect in apoptosis. This is supported by our in vitro studies, in which phagocytosis of opsonized particles by human neutrophils rapidly induced apoptosis that could be blocked with CD11b/ CD18 antibodies. Reactive oxygen species are the intracellular link in this process: phagocytosis-induced apoptosis was blocked both in neutrophils treated with the flavoprotein inhibitor diphenylene iodonium and in neutrophils from patients with chronic granulomatous disease, which lack NADPH oxidase. Thus, CD11b/CD18 plays a novel and unsuspected homeostatic role in inflammation by accelerating the programmed elimination of extravasated neutrophils.
1-s2.0-s1074761300802782-main.pdf
Diacovo TG, Puri KD, Warnock RA, Springer TA, von Andrian UH. Platelet-mediated lymphocyte delivery to high endothelial venules. Science. 1996;273 (5272) :252-5.Abstract
Circulating lymphocytes gain access to lymph nodes owing to their ability to initiate rolling along specialized high endothelial venules (HEVs). One mechanism of rolling involves L-selectin binding to peripheral node addressin (PNAd) on HEVs. Activated platelets are shown to bind to circulating lymphocytes and to mediate rolling in HEVs, in vivo, through another molecule, P-selectin, which also interacts with PNAd. In vitro, activated platelets enhanced tethering of lymphocytes to PNAd and sustained lymphocyte rolling, even in the absence of functional L-selectin. Thus, a platelet pathway operating through P-selectin provides a second mechanism for lymphocyte delivery to HEVs.
Stoddart JH, Jasuja RR, Sikorski MA, von Andrian UH, Mier JW. Protease-resistant L-selectin mutants. Down-modulation by cross-linking but not cellular activation. J Immunol. 1996;157 (12) :5653-9.Abstract
The adhesion molecule L-selectin (CD62L) is rapidly shed from the plasma membrane during leukocyte activation as a result of proteolytic cleavage between Lys321 and Ser322 within the extracellular domain. L-selectin is also down-modulated from the surface in response to cross-linking, possibly through a similar mechanism. To further characterize the mechanism of down-modulation, several L-selectin mutants were generated and transfected into COS cells. Wild-type L-selectin as well as mutants with one or two amino acid substitutions at the cleavage site were nearly quantitatively shed into the culture supernatant. However, mutants in which a nine-amino acid stretch that included the protease-sensitive site was either deleted or replaced with a polyglycine spacer or a comparable region of E-selectin were retained on the cell surface and not detected in the supernatant. These results are consistent with other reports describing protease resistant L-selectin mutants. We also demonstrate that when expressed in L1-2 pre-B cells, the L-selectin nine-amino acid deletion mutant (321del.9), but not wild-type L-selectin, is resistant to down-regulation induced by PMA. However, both wild-type and mutant 321del.9 are completely lost from the cell surface in response to cross-linking with an L-selectin Ab. PMA-induced- but not L-selectin cross-linking-induced down-modulation was inhibited by staurosporine. These data are consistent with the idea that the L-selectin protease(s) can tolerate minor structural alterations at the cleavage site, and that L-selectin down-modulation can be induced by more than one mechanism, at least one of which (cross-linking) is protein kinase C independent.