Endothelial Ligands For L-selectin Research Articles

Endothelial L-selectin ligand expression in nasal polyps

L-selectins on leukocytes and their counter-receptors on endothelial cells have been shown to be involved in leukocyte recruitment in chronic rhinosinusitis without nasal polyps (NP). The purpose of this study was to evaluate the expression level of functionally active endothelial L-selectin ligands in NP obtained from patients with NP of different etiology [simple NP, antro-choanal polyps (ACP) and cystic fibrosis (CF) NP] and inferior turbinate specimens of healthy controls and to compare these levels to the presence of various leukocyte subsets. Nasal polyp specimens and healthy nasal mucosa specimens were obtained from patients undergoing surgery and were immunohistochemically stained with monoclonal antibodies detecting CD34, sialyl Lewis x (sLe(x)) of sulfated extended core 1 lactosamines and various leukocyte subsets. All NP are characterized by a decrease in the number of CD34+ vessels. The number of eosinophils and the percentage of vessels expressing endothelial sulfated sLe(x) epitopes is upregulated in all groups of simple NP. Tissue eosinophilia is increased in those patients with increased disease severity (acetyl salicylic acid intolerance), but the percentage of endothelial sulfated sLe(x) epitopes is not. Results on CF NP are similar to those observed for simple NP. Antro-choanal polyps, on the contrary, are characterized by low numbers of tissue eosinophils and relatively few vessels expressing endothelial sulfated sLe(x) epitopes. Our results suggest that functionally active L-selectin ligands might play a role in guiding leukocyte traffic into NP in patients with simple NP and CF NP but not ACP.

Mucosal Eosinophils and L-Selectin Ligands are Associated with Invasive and Noninvasive Sinus Surgery Outcomes

Chronic rhinosinusitis (CRS) is characterized by persistent inflammation of the nasal and paranasal mucosa with numerous emigrated leukocytes. L-Selectin on leukocytes and its endothelial glycosylated ligands initiate leukocyte infiltration into inflamed tissues. Endoscopic sinus surgery (ESS) is the major approach for restoring sinus physiology after failure of conservative therapy; however, the effect of enlarging the maxillary sinus ostium is still unknown. Here, we compared two histological markers of local inflammation, the number of mucosal eosinophils, and the expression of endothelial L-selectin ligands, with clinical outcomes after enlarging or saving the maxillary sinus ostium. Twenty-three patients with CRS underwent uncinectomy on one side and additional middle meatal antrostomy on the other side. Maxillary sinus mucosa biopsy specimens from these patients and nine healthy subjects were taken for immunohistochemical evaluations of the number of mucosal eosinophils and endothelial L-Selectin ligands. Also, symptoms and mucociliary clearance were measured. The postoperative reduction of the endothelial L-Selectin ligands was independent of the operation technique. There was a correlation between postoperative number of mucosal eosinophils and symptom score, which was also independent of the surgical technique. The postoperative decrease of mucosal eosinophils, as well as the correlation of the intraoperative eosinophils with the postoperative symptom score, was found only on antrostomy side. ESS decreases the expression of endothelial L-Selectin ligands, which might lead to decreased eosinophil traffic into maxillary sinus mucosa, putatively more when enlarging the maxillary sinus ostium. Both intra- and postoperative low number of eosinophils seem to be indicators of good subjective recovery.

Inflammatory Blues Turns Velvet Skin Into Rawhide

The directed homing of leukocytes is crucial during immune surveillance or inflammation and is accomplished by a complex cooperation of signaling and adhesion molecules.1,2 Its pathophysiologic relevance is exemplified by various inflammatory diseases such as atherosclerosis.3 At present, the course of leukocyte trafficking is well established through the classical multistep cascade of initial tethering and rolling of leukocytes over vascular endothelium, followed by firm adhesion and their subsequent spreading and transendothelial migration.1,2 Leukocyte rolling is mediated by a subgroup of C-type lectins; E-, L-, and P-selectin, through shear-resistant binding to particular cell-surface carbohydrates. E- and L-selectin are expressed on activated endothelial cells and most leukocytes, respectively, while P-selectin is stored in the secretory compartments of platelets and endothelial cells to become rapidly upregulated on cell activation. The best characterized ligand for selectins is P-selectin glycoprotein ligand-1 (PSGL-1),4 a heavily posttranslationally modified homodimeric transmembrane glycoprotein. PSGL-1 contains functionally essential sialylated and fucosylated carbohydrate moieties, known as sialyl Lewis X (sLex) groups, and is expressed on blood cells such as neutrophils, monocytes, and platelets.5 The importance of PSGL-1 for cell recruitment is highlighted by studies using transgenic mice deficient in PSGL-1, which revealed a crucial contribution to P-selectin–dependent rolling on inflamed endothelium, indicating a function of PSGL-1 in early inflammatory responses.6 Presented by endothelium-bound leukocytes, PSGL-1 supports the initial L-selectin–dependent tethering of blood-borne leukocytes to the already adherent leukocytes at the inflamed vessel wall followed by E-selectin-mediated rolling on the endothelium.7 Besides supporting leukocyte–endothelium interactions, PSGL-1 promotes the P-selectin–dependent initial tethering of platelets to monocytes8,9 followed by more stable interactions through integrins expressed on both platelets and monocytes. In addition, the interaction between PSGL-1 and P-selectin also plays an essential role in the delivery …

No endothelial L‐selectin ligand mediates leukocyte rolling in inflammation

The multistep process of leukocyte recruitment guides leukocyte trafficking and immune function. The leukocyte cell adhesion molecule (CAM) L-selectin is a key molecule in this process. Its role in inflammation has largely been attributed to the presence of an endothelial L-selectin ligand expressed on inflamed endothelium. However, the identity of this ligand(s) has been elusive. Here, we used intravital microscopy to show that the reduction of leukocyte rolling in inflammation following blockage of L-selectin is entirely dependent on loss of L-selectin/PSGL-1-dependent leukocyte-mediated secondary capture. Secondary capture initiates rolling mainly on endothelial E-selectin and function-inhibition of L-selectin or E-selectin thus blocks rolling of largely the same pool of rolling leukocytes. All rolling along endothelium in venules following six distinct treatment regimens were abolished after simultaneous blockage of P-selectin, E-selectin and VCAM-1 and no L-selectin ligand activity on endothelium was detected. These data show that there is no ligand for L-selectin mediating rolling along endothelium in inflamed venules. These findings may conclude the search for L-selectin ligands capable of mediating leukocyte rolling in inflammation.

α2,3‐Sialyltransferase‐IV is essential for L‐selectin ligand function in inflammation

L-selectin belongs to the C-type lectin family of glycoproteins and is constitutively expressed on most leukocytes. L-selectin mediates leukocyte rolling in inflamed microvessels and high endothelial venules (HEV) via binding to specific carbohydrate structures on selectin ligands. Previous studies using sialidase treatment suggested a role of sialic acid residues in L-selectin-dependent rolling. To investigate the role of the alpha2,3-sialyltransferase (ST3Gal)-IV on L-selectin ligand activity in vivo, we studied leukocyte rolling in inflamed venules of the cremaster muscle and in Peyer's patch HEV of ST3Gal-IV-deficient mice and littermate control mice. In cremaster muscle venules with or without TNF-alpha treatment, L-selectin-dependent rolling was almost completely abolished in ST3Gal-IV(-/-) mice. In both models, L-selectin interacts with P-selectin glycoprotein ligand-1 (PSGL-1) presented by adherent leukocytes and leukocyte fragments, but not with endothelial L-selectin ligands. In contrast, L-selectin-dependent rolling in Peyer's patch HEV, which is mediated by unknown endothelial L-selectin ligands, was not impaired in the absence of ST3Gal-IV. Our in vivo data show that PSGL-1, the molecule responsible for L-selectin-mediated leukocyte interactions in inflammation, is dependent on ST3Gal-IV, while alpha2,3-sialylation by ST3Gal-IV is not necessary for L-selectin ligand activity on high endothelial cells of Peyer's patch HEV.

Endothelial L-Selectin Ligands in Sinus Mucosa during Chronic Maxillary Rhinosinusitis

Chronic rhinosinusitis is characterized by persistent inflammation of the nasal and paranasal mucosa with numerous emigrated leukocytes. L-selectin on leukocytes and its endothelial glycosylated ligands initiate organ-specific leukocyte infiltration into inflamed tissues. The purpose of this study was to evaluate the endothelial expression of functionally active endothelial L-selectin ligands, sulfated sialyl Lewis x, in maxillary sinus mucosa from patients with chronic rhinosinusitis and from normal control subjects. Maxillary sinus mucosa specimens (116) were obtained surgically and immunohistochemically stained with monoclonal antibodies detecting sialyl Lewis x or sulfated extended core 1 lactosamines. The severity of the inflammation was determined by intraoperative endoscopic findings, computed tomography scans, and histopathologic assessment of the specimens. The percentage of vessels expressing endothelial sulfated sialyl Lewis x epitopes increased during chronic rhinosinusitis compared with uninflamed control tissue, especially in patients with additional allergic rhinitis, and decreased in specimens from aspirin-intolerant patients with preoperative oral corticosteroid treatment. In addition, the expression level of endothelial sulfated sialyl Lewis x epitopes and the number of mucosal eosinophils correlated with the severity of the inflammation, and decreased in specimens taken 9 months postoperatively compared with intraoperative samples, especially in patients with intranasal corticosteroid treatment. Our results suggest that functionally active L-selectin ligands might guide leukocyte traffic into maxillary sinus mucosa preferentially in patients with severe findings of chronic maxillary rhinosinusitis, thus leading to aggravation of the inflammation.

Therapeutic Targeting of Endothelial Ligands for L-selectin (PNAd) in a Sheep Model of Asthma

The homing of lymphocytes to peripheral lymph nodes is initiated by an adhesive interaction between L-selectin on lymphocytes and PNAd, a set of sialomucins that are constitutively displayed on high endothelial venules of lymph nodes. PNAd is defined by monoclonal antibody MECA-79 that recognizes a sulfated oligosaccharide carried by the sialomucins. This epitope overlaps with 6-sulfo sialyl Lewis x, a recognition determinant for L-selectin. Previous work has shown that administration of a L-selectin monoclonal antibody blocks both late-phase airway responses and airway hyperresponsiveness in a sheep model of asthma. We show here that airway-associated lymphoid collections from lungs of allergic sheep exhibited PNAd(+) venules as detected by immunostaining with MECA-79. The same vessels also expressed a GlcNAc-6-O-sulfotransferase known as HEC-GlcNAc6ST, which is known to contribute to the formation of the MECA-79 epitope in high endothelial venules of mouse lymph nodes. Intravenous administration of MECA-79 to allergic sheep significantly blunted both the late-phase airway response and airway hyperresponsiveness induced by airway allergen challenge. Furthermore, MECA-79 inhibited the accumulation of all classes of leukocytes in bronchoalveolar lavage fluid. These findings represent the first demonstration that targeting of PNAd has therapeutic efficacy in an inflammatory disease.

Optical molecular imaging of lymph nodes using a targeted vascular contrast agent

We develop a highly specific antibody-dye conjugate for optical imaging of peripheral lymph nodes. The contrast agent consists of the monoclonal antibody recognizing endothelial ligands for the lymphocyte homing receptor L-selectin, MECA-79, and a near-infrared (near-IR) fluorescent indotricarbocyanine dye. The targeting and biodistribution behavior of MECA-79 is studied after radio-iodination and intravenous injection into mice demonstrating specific uptake in lymph nodes and accumulation in high endothelial venules (HEV). After conjugation of MECA-79 with indotricarbocyanine dye, the fluorescence imaging properties of the MECA-79 dye conjugate are examined by intravenous injection in nude mice and laser-induced fluorescence whole-body imaging in vivo. The MECA-79 antibody-dye conjugate accumulates in peripheral lymph nodes, whereas an isotype antibody-dye conjugate does not. Specific lymph node near-IR fluorescent signals become detectable within minutes after injection, and stable imaging persists for more than 24 h. The results demonstrate that vascular targeting of endothelial expression of glyocproteins is feasible to visualize the accumulation of near-IR fluorescent MECA-79 in lymph nodes, making this technology potentially useful to characterize processes of inflammation.

A novel endothelial L-selectin ligand activity in lymph node medulla that is regulated by alpha(1,3)-fucosyltransferase-IV.

Lymphocytes home to peripheral lymph nodes (PLNs) via high endothelial venules (HEVs) in the subcortex and incrementally larger collecting venules in the medulla. HEVs express ligands for L-selectin, which mediates lymphocyte rolling. L-selectin counterreceptors in HEVs are recognized by mAb MECA-79, a surrogate marker for molecularly heterogeneous glycans termed peripheral node addressin. By contrast, we find that medullary venules express L-selectin ligands not recognized by MECA-79. Both L-selectin ligands must be fucosylated by α(1,3)-fucosyltransferase (FucT)-IV or FucT-VII as rolling is absent in FucT-IV+VII−/− mice. Intravital microscopy experiments revealed that MECA-79–reactive ligands depend primarily on FucT-VII, whereas MECA-79–independent medullary L-selectin ligands are regulated by FucT-IV. Expression levels of both enzymes paralleled these anatomical distinctions. The relative mRNA level of FucT-IV was higher in medullary venules than in HEVs, whereas FucT-VII was most prominent in HEVs and weak in medullary venules. Thus, two distinct L-selectin ligands are segmentally confined to contiguous microvascular domains in PLNs. Although MECA-79–reactive species predominate in HEVs, medullary venules express another ligand that is spatially, antigenically, and biosynthetically unique. Physiologic relevance for this novel activity in medullary microvessels is suggested by the finding that L-selectin–dependent T cell homing to PLNs was partly insensitive to MECA-79 inhibition.

Sulphated endothelial ligands for L-selectin in lymphocyte homing and inflammation.

Lymphocytes from the blood home to secondary lymphoid tissues through a process of tethering, rolling, firm adhesion and transmigration. Tethering and rolling of lymphocytes is mediated by the interaction of L-selectin on lymphocytes with sulphated ligands expressed by the specialized endothelial cells of high endothelial venules (HEVs). The sulphate-dependent monoclonal antibody MECA79 stains HEVs in peripheral lymph nodes and recognizes the complex of HEV ligands for L-selectin termed peripheral node addressin. High endothelial cell GlcNAc-6-sulphotransferase/L-selectin ligand sulphotransferase is a HEV-expressed sulphotransferase that contributes to the formation of the MECA79 epitope and L-selectin ligands on lymph node HEVs. MECA79-reactive vessels are also common at sites of chronic inflammation, suggesting mechanistic parallels between lymphocyte homing and inflammatory trafficking.

Development of a simple homogeneous assay to screen for inhibitors of N-acetylglucosamine-6-sulfotransferases

L-selectin, a leukocyte adhesion molecule, plays a central role in lymphocyte homing to secondary lymphoid tissue and to certain sites of inflammation. Carbohydrate sulfation was implicated in this process, when it was demonstrated that carbohydrate sulfotransferase-mediated sulfation of N-acetylglucosamine (GlcNAc) within sialyl Lewis X of cognate endothelial ligands for L-selectin was an essential modification for L-selectin binding. The recently identified GlcNAc-6-sulfotransferases GlcNAc6ST-1 and -2, which facilitate GlcNAc sulfation by catalyzing the transfer of a sulfonyl group from 3 ′-phosphoadenosine 5 ′-phosphosulfate (PAPS) to the 6-hydroxy group of the acceptor GlcNAc moiety, contribute to the biosynthesis of the 6-sulfosialyl Lewis X motif. Due to their pivotal role in L-selectin ligand biosynthesis, this enzyme class has recently emerged as an important and relatively unexplored class of potential targets for anti-inflammatory therapy. However, no inhibitors have been reported to date and screening for lead inhibitors has been hampered by the lack of simple assay formats suitable for high-throughput screening. Here, we report the development of a simple homogeneous in vitro sulfotransferase assay using a newly synthesized biotinylated glycoside as a substrate. The assay is based on GlcNAc6ST-2-mediated [ 35S]sulfate transfer from [ 35S]PAPS to the biotinylated glycoside and subsequent detection using streptavidin-coated SPA beads. K m values with partially purified GlcNAc6ST-2 for PAPS and the biotinylated glycoside were estimated to be 8.4 and 34.5 μM, respectively. The sulfotransferase reaction could be inhibited by 3 ′,5 ′-ADP with an IC 50 of 2.1 μM. The assay can be operated in 384-well format; is characterized by a high signal-to-noise ratio, low variation, and excellent Z factors; and is highly suitable for high-throughput screening.

Differential requirements for core2 glucosaminyltransferase for endothelial L-selectin ligand function in vivo.

L-selectin is a calcium-dependent lectin on leukocytes mediating leukocyte rolling in high endothelial venules and inflamed microvessels. Many selectin ligands require modification of glycoproteins by leukocyte core2 beta1,6-N-acetylglucosaminyltransferase (Core2GlcNAcT-I). To test the role of Core2GlcNAcT-I for L-selectin ligand biosynthesis, we investigated leukocyte rolling in venules of untreated and TNF-alpha-treated cremaster muscles and in Peyer's patch high endothelial venules (HEV) of Core2GlcNAcT-I null (core2(-/-)) mice. In the presence of blocking mAbs against P- and E-selectin, L-selectin-mediated leukocyte rolling was almost completely abolished in cremaster muscle venules of core2(-/-) mice, but not littermate control mice. By contrast, leukocyte rolling in Peyer's patch HEV was not significantly different between core2(-/-) and control mice. To probe L-selectin ligands more directly, we injected L-selectin-coated beads. These beads showed no rolling in cremaster muscle venules of core2(-/-) mice, but significant rolling in controls. In Peyer's patch HEV, beads coated with a low concentration of L-selectin showed reduced rolling in core2(-/-) mice. Beads coated with a 10-fold higher concentration of L-selectin rolled equivalently in core2(-/-) and control mice. Our data show that endothelial L-selectin ligands relevant for rolling in inflamed microvessels of the cremaster muscle are completely Core2GlcNAcT-I dependent. In contrast, L-selectin ligands in Peyer's patch HEV are only marginally affected by the absence of Core2GlcNAcT-I, but are sufficiently functional to support L-selectin-dependent leukocyte rolling in Core2GlcNAcT-I-deficient mice.

De novo induction of endothelial L-selectin ligands during kidney allograft rejection.

Acute kidney allograft rejection is characterized by a lymphocyte infiltration. L-selectin on lymphocytes and its endothelial glycosylated ligands are instrumental in the initiation of lymphocyte extravasation to sites of inflammation. From more than 500 core biopsy specimens taken from kidneys after transplantation, 250 biopsies were graded to have signs of acute rejection. Of these, 52 biopsies with various grades of histologic signs of acute rejection were selected for the study. Controls were 15 biopsies taken within 30 min after revascularization and 10 specimens from well-functioning allografts showing no clinical or histologic evidence of rejection. Immunochemical stainings with monoclonal antibodies against functionally active decorated L-selectin ligands. i.e., sialyl-Lewis x (sLex, 2F3 and HECA-452) or sulfated lactosamine (MECA-79) were performed. Although no endothelial 2F3 and MECA-79 epitopes were detected in nonrejecting control specimens, the expression was induced at the onset and during acute allograft rejections. The level of expression (in semi-quantitative score) of 2F3 reactivity correlated with the severity of rejection (P<0.0001, grade I versus grade IIB), and the level of expression decreased as the rejection resolved. Kidney biopsies taken shortly after revascularization and thus undergoing reperfusion injury showed endothelial staining with another anti sLex antibody, HECA-452. This staining disappeared from well-functioning grafts and reappeared at the onset of an acute allograft rejection. These results suggest that expression of functionally active, properly glycosylated L-selectin ligands might have a role in reperfusion injury and in the initiation of acute rejections after human kidney allograft transplantation.

Minimal Sulfated Carbohydrates for Recognition by L-selectin and the MECA-79 Antibody

Sulfated forms of sialyl-Le(X) containing Gal-6-SO(4) or GlcNAc-6-SO(4) have been implicated as potential recognition determinants on high endothelial venule ligands for L-selectin. The optimal configuration of sulfate esters on the N-acetyllactosamine (Galbeta1-->4GlcNAc) core of sulfosialyl-Le(X), however, remains unsettled. Using a panel of sulfated lactose (Galbeta1-->4Glc) neoglycolipids as substrates in direct binding assays, we found that 6',6-disulfolactose was the preferred structure for L-selectin, although significant binding to 6'- and 6-sulfolactose was observed as well. Binding was EDTA-sensitive and blocked by L-selectin-specific monoclonal antibodies. Surprisingly, 6', 6-disulfolactose was poorly recognized by MECA-79, a carbohydrate- and sulfate-dependent monoclonal antibody that binds competitively to L-selectin ligands. Instead, MECA-79 bound preferentially to 6-sulfolactose. The difference in preferred substrates between L-selectin and MECA-79 may explain the variable activity of MECA-79 as an inhibitor of lymphocyte adhesion to high endothelial venules in lymphoid organs. Our results suggest that both Gal-6-SO(4) and GlcNAc-6-SO(4) may contribute to L-selectin recognition, either as components of sulfosialyl-Le(X) capping groups or in internal structures. By contrast, only GlcNAc-6-SO(4) appears to contribute to MECA-79 binding.

Carbohydrate sulfotransferases in lymphocyte homing.

Sulfation is a critical modification in many instances of biological recognition. Early work in lymphocyte homing indicated that the endothelial ligands for L-selectin depended upon sulfation modifications. Subsequent studies showed that the two specific modifications, Gal-6-SO4 and GlcNAc-6-SO4, were present on actual biological ligands. Recently, a family of carbohydrate sulfotransferases capable of generating these modifications has been identified at the molecular level. Reconstitution experiments implicate members of this family as critical participants in lymphocyte homing.

Endothelial Ligands for L-Selectin: From Lymphocyte Recirculation to Allograft Rejection

The article by Toppila et al 1 in this issue of The American Journal of Pathology raises the provocative possibility that the adhesion molecule L-selectin may play a significant role in the recruitment of lymphocytes to human heart allografts during rejection. The case made by these authors relies strongly on current knowledge about the high-endothelial-venule-expressed ligands (HEV ligands) for L-selectin, which participate in the constitutive homing of lymphocytes to secondary lymphoid organs. The purpose of this Commentary is to summarize and update this rapidly evolving field.

Endothelial L-Selectin Ligands Are Likely to Recruit Lymphocytes into Rejecting Human Heart Transplants

L-selectin-dependent lymphocyte extravasation is a hallmark of acute heart allograft rejection in rats. On screening over 600 endomyocardial biopsies (EMBs), taken at different time points after heart transplantation in man, we identified 91 samples with histological signs of acute rejection. Rejection and nonrejection EMBs were analyzed for the presence of properly glycosylated, ie, sulfated sialyl Lewis-x (sLex) decorated L-selectin ligands. Two anti-sLex (2F3 and HECA-452) and one anti-6- or 6'-sulfated and/or 6, 6'-bisulfation (MECA-79) monoclonal antibodies were used. Nonrejecting heart endothelium did not express, or expressed only weakly, sulfated and or sLex decorations of L-selectin ligands. On the contrary, these epitopes were readily detectable on endothelium of capillaries and venules at the onset and during acute rejection episodes. The more intense the sulfated sLex expression was, the more severe the rejection episode was in histological grading. The endothelial expression of L-selectin ligands decreased to background levels as the rejection resolved. Our data demonstrate a complete correlation between the level of expression of the sulfated sLex-decorated ligands on the one hand and the histological severity of acute heart allograft rejection on the other hand. These data suggest that functionally active endothelial L-selectin ligands are instrumental in lymphocyte extravasation into the human heart allografts at the onset and during acute rejection episodes.

Sulfotransferases of two specificities function in the reconstitution of high endothelial cell ligands for L-selectin.

L-selectin, a lectin-like receptor, mediates rolling of lymphocytes on high endothelial venules (HEVs) in secondary lymphoid organs by interacting with HEV ligands. These ligands consist of a complex of sialomucins, candidates for which are glycosylation- dependent cell adhesion molecule 1 (GlyCAM-1), CD34, and podocalyxin. The ligands must be sialylated, fucosylated, and sulfated for optimal recognition by L-selectin. Our previous structural characterization of GlyCAM-1 has demonstrated two sulfation modifications, Gal-6-sulfate and GlcNAc-6-sulfate in the context of sialyl Lewis x. We now report the cloning of a Gal-6-sulfotransferase and a GlcNAc-6-sulfotransferase, which can modify GlyCAM-1 and CD34. The Gal-6-sulfotransferase shows a wide tissue distribution. In contrast, the GlcNAc-6-sulfotransferase is highly restricted to HEVs, as revealed by Northern analysis and in situ hybridization. Expression of either enzyme in Chinese hamster ovary cells, along with CD34 and fucosyltransferase VII, results in ligand activity, as detected by binding of an L-selectin/IgM chimera. When coexpressed, the two sulfotransferases synergize to produce strongly enhanced chimera binding.

Functional distribution and further characterization of human endothelial ligand for cellular L-selectin

In the present study we examine the functional distribution of the human endothelial L-selectin ligand, which determines the sites of extravasation of L-selectin-positive cells. A murine cell line transfected with human L-selectin adhered preferentially to the high endothelial venules (HEV) of human peripheral lymph nodes compared to the HEV of mucosal lymphoid tissues (mean of 0.83 compared to a mean of 0.07 cells per HEV respectively). In addition, an antibody against L-selectin differentially inhibited the adhesion of human lymphocytes to peripheral lymphoid tissue versus mucosal lymphoid tissue HEV (mean 41 and 5% inhibition respectively). Although both sulfoglucuronyl-containing glycolipids and sialyl-Lewis X have been proposed as endothelial ligands for L-selectin, an antibody against the former did not bind to peripheral lymph node endothelium, and an antibody against the latter did not block adhesion of L-selectin-expressing cells. The enzyme O-sialoglycoprotein endopeptidase caused up to an 84% reduction in L-selectin-dependent binding, indicating that sialylated glycoproteins containing O-linked glycans are essential for a large majority of adhesion via L-selectin.

Selective induction of endothelial L-selectin ligand in human lung inflammation.

During inflammation, leukocyte emigration from the circulation can be directed by the endothelium, in part by the inducible endothelial adhesion ligand for L-selectin. In this study, endothelial L-selectin ligand expression was localized by immunohistochemistry in human lung in several different types of lung inflammation and in systemic inflammation. Endothelial L-selectin ligand was not seen in normal lung or in acute pneumonia involving neutrophil accumulation. However, the endothelial ligand was seen in most cases of chronic interstitial pneumonia with mononuclear cell accumulation (a mean of 5.9% of microvessels positive). Regarding granulomatous conditions, in sarcoidosis the endothelial ligand was not identified, but in tuberculous infection some expression was seen in a minority of cases (mean 3.3% of microvessels positive). In contrast, consistent, typically extensive ligand induction (mean 33.4% of microvessels positive) was present in bronchiectatic lung showing prominent lymphocytic accumulation and venules with thickened (high) endothelium, the latter being normally characteristic of lymphoid tissue in which L-selectin ligand is known to be constitutively expressed. Lung from subjects with systemic infection was negative for endothelial expression of the ligand. These studies show how in a defined extralymphoid tissue induction of endothelial L-selectin ligand depended not only on the presence or absence of an inflammatory state, but also on the nature of the inflammation.