Adhesion Receptors ICAM-1 Research Articles

Impact of radiation therapy on vascular endothelial adhesion receptor expression

The adjunction of PD-1/PD-L1 immunotherapy and radiation therapy (RT) has proven to be an effective approach to combat cancer. Clinical studies recently confirmed that patients show increased response to immune checkpoint inhibitors if also treated with RT. One effect of RT in this setting is the facilitation of recruitment of immune cells to tumors in which T cell infiltration is low or absent. Recruitment of immune cells is primarily dependent on endothelial activation: activated endothelial cells exhibit an increased expression of endothelial adhesion receptors that mediate recruitment from the blood. Despite evidence that RT can boost T cell recruitment to tumors an in-depth analysis of endothelial adhesion receptor expression and which fractionation doses are best suited in this context are unknown. We hypothesize that RT leads to activation of the endothelium. Using the endothelial mouse cell line bEnd5, we investigated the expression of the adhesion receptors ICAM-1, ICAM-2, VCAM-1, E-selectin, and P-selectin following RT ranging from 0.5 to 10 Gy at different times points (1.5h, 4h, and 24h) with or without prior stimulation with TNF-α by qPCR, flow cytometry, and immunofluorescence. We observed upregulation of E-selectin, P-selectin, and VCAM-1 as early as 1.5h after RT. Early upregulation of selectins was independent of dose; however, sustained upregulation of selectins and upregulation of VCAM-1 was dose-dependent. Following preceding stimulation with TNF-α led to an increased baseline expression of E-Selectin and VCAM-1. E-selectin, P-selectin, and VCAM-1 expression was further increased after RT, and to a much greater extent than without prior stimulation. Again, effects were more pronounced at early time points and showed a dose dependency for VCAM-1. In contrast, while ICAM-2 was not changed by RT or was even downregulated, the addition of RT following TNF-α treatment did increase ICAM-1 levels. This effect was not observed without prior TNF-α stimulation. In vivo experiments of KrasG12D;Tp53fl/fl NSCLC tumors revealed changes in P-selectin, E-selectin, and VCAM-1 that mimicked changes of in vitro experiments with prior TNF-α stimulation. Performing adoptive T cell transfer experiments, we found that both timing and dose of RT is important for successful recruitment of transferred cells to the tissue. Our study demonstrates that RT alters endothelial receptor display on endothelial cells in vitro and in vivo and is influenced significantly by the local cytokine milieu.

Integrated proteomic analysis of tumor necrosis factor α and interleukin 1β-induced endothelial inflammation

The vascular endothelium provides a unique interaction plane for plasma proteins and leukocytes in inflammation. The pro-inflammatory cytokines Tumor Necrosis Factor α (TNFα) and interleukin 1β (IL-1β) have a profound effect on endothelial cells, which includes increased levels of adhesion molecules and a disrupted barrier function. To assess the endothelial response to these cytokines at the protein level, we evaluated changes in the whole proteome, cell surface proteome and phosphoproteome after 24 h of cytokine treatment. The effects of TNFα and IL-1β on endothelial cells were strikingly similar and included changes in proteins not previously associated with endothelial inflammation. Temporal profiling revealed time-dependent proteomic changes, including a limited number of early responsive proteins such as adhesion receptors ICAM1 and SELE. In addition, this approach uncovered a greater number of late responsive proteins, including proteins related to self-antigen peptide presentation, and a transient increase in ferritin. Peptide-based cell surface proteomics revealed extensive changes at the cell surface, which were in agreement with the whole proteome. In addition, site-specific changes within ITGA5 and ICAM1 were detected. Combined, our integrated proteomic data provide detailed information on endothelial inflammation, emphasize the role of the extracellular matrix therein, and include potential targets for therapeutic intervention. SignificancePro-inflammatory cytokines induce the expression of cell adhesion molecules in vascular endothelial cells. These molecules mediate the adhesion and migration of immune cells across the vessel wall, which is a key process to resolve infections in the underlying tissue. Dysregulation of endothelial inflammation can contribute to vascular diseases and the vascular endothelium is therefore an attractive target to control inflammation. Current strategies targeting endothelial adhesion molecules, including PECAM, CD99, ICAM1 and VCAM1 do not completely prevent transmigration. To identify additional therapeutic targets, we mapped the endothelial proteome after pro-inflammatory cytokine treatment. In addition to the whole proteome, we assessed the surface proteome to focus on cell adhesion molecules, and the phosphoproteome to uncover protein activation states. Here, we present an integrated overview of affected processes which further improves our understanding of endothelial inflammation and may eventually aid in therapeutic intervention of imbalanced inflammation.

Induction of peripheral Tregs by homotypic T cell interaction (89.3)

Abstract The mechanisms by which regulatory T cells are generated in the periphery are incompletely understood. Here we show that homotypic T cell interaction results in the generation of T cells with a potent regulatory capacity. CD4 memory T cells that were cultured together with activated effector T cells proliferated and produced various cytokines depending on the nature of the activated effector T cell. Whereas Th1 cells promoted the development of IL-10 and/or IFNγ producing cells, T cells cultured with Th2 cells produced IL-4. The interaction between the adhesion receptors ICAM-1, 2, 3 and LFA-1, respectively was crucial for T-T cell communication with regard to proliferation and cytokine production. Blocking of ICAM-1 diminished IFNγ production, whereas ICAM-3 was important for IL-4 secretion. Interestingly, the resulting cells strongly inhibited proliferation of CD25- T cells in a dose dependent manner independent of their cytokine secretion profile. Moreover, in vitro generated T-T cell induced T cells with a TCR specific for OVA prevented the expansion of OVA specific effector T cells in Balb/c mice in vivo upon antigen challenge. The results indicate that homotypic T cell interaction induces the generation of T cells with a regulatory phenotype, providing a novel negative feedback mechanism to control sustained T cell driven immunity.

Lymphocyte transcellular migration occurs through recruitment of endothelial ICAM-1 to caveola- and F-actin-rich domains

During inflammation, leukocytes bind to the adhesion receptors ICAM-1 and VCAM-1 on the endothelial surface before undergoing transendothelial migration, also called diapedesis. ICAM-1 is also involved in transendothelial migration, independently of its role in adhesion, but the molecular basis of this function is poorly understood. Here we demonstrate that, following clustering, apical ICAM-1 translocated to caveolin-rich membrane domains close to the ends of actin stress fibres. In these F-actin-rich areas, ICAM-1 was internalized and transcytosed to the basal plasma membrane through caveolae. Human T-lymphocytes extended pseudopodia into endothelial cells in caveolin- and F-actin-enriched areas, induced local translocation of ICAM-1 and caveolin-1 to the endothelial basal membrane and transmigrated through transcellular passages formed by a ring of F-actin and caveolae. Reduction of caveolin-1 levels using RNA interference (RNAi) specifically decreased lymphocyte transcellular transmigration. We propose that the translocation of ICAM-1 to caveola- and F-actin-rich domains links the sequential steps of lymphocyte adhesion and transendothelial migration and facilitates lymphocyte migration through endothelial cells from capillaries into surrounding tissue.

Leukocyte migration in adipose tissue of mice null for ICAM-1 and Mac-1 adhesion receptors.

To determine whether the leukocyte adhesion receptors ICAM-1 and Mac-1, regulators of immune cell migration, have an intrinsic role within adipose tissue by 1) analyzing the expression of ICAM-1 in adipose tissue, 2) identifying leukocyte populations within adipose tissue, and 3) determining whether ICAM-1 and Mac-1 mutant mice exhibit abnormal numbers of adipose tissue leukocytes. Wild-type, ICAM-1(-/-), and Mac-1(-/-) mice were fed a long-term high-fat diet. ICAM-1 expression was analyzed by Northern blot and immunohistochemistry. Leukocytes within adipose tissue were identified by immunohistochemistry and flow cytometry. ICAM-1 was expressed in adipose tissue and localized to the vascular endothelium. Macrophages and lymphocytes were prevalent within the stromal-vascular cell fraction of adipose tissue, and gender-specific differences were observed, with adipose tissue from female mice containing significantly more macrophages than tissue from male mice. Numbers of leukocytes in ICAM-1(-/-) and Mac-1(-/-) mice were not different from wild-types, however, indicating that these adhesion receptors are not required for leukocyte migration into adipose tissue. Our results documented leukocyte populations within adipose tissue, which may be involved in the development of heightened inflammation that is characteristic of obesity.

In situ hybridisation and immunocytochemical localisation of osteolytic cytokines and adhesion molecules in ameloblastomas.

Ameloblastomas produce interleukin-1-like activity that could explain some part of their osteolytic capability. However, the cellular source of this osteolytic activity is unknown. In the present study, cytokines with known inflammatory and osteolytic activity, i.e., interleukin-1 (IL-1), tumour necrosis factor (TNF), and interleukin-6 (IL-6), have been localised by immunocytochemistry and in situ hybridisation. The cellular adhesion receptors ICAM-1, E-selectin and VCAM-1 have also been immunolocalised. Immunocytochemistry demonstrated that all seven specimens showed positive staining for IL-1alpha and IL-6 with these cytokines being located in the stellate reticulum-like cells and vascular endothelium. Very faint staining for IL-1beta was seen in four of seven specimens. No reaction was seen for TNF-alpha. All specimens demonstrated E-selectin staining in the vascular endothelium and ICAM-1 and VCAM-1 staining in the stellate reticulum-like cells and the endothelium. In situ hybridisation for the cytokines showed the presence of mRNA of both IL-1alpha and IL-6 in the stellate reticulum-like cells. Faint staining for IL-1beta was also seen. No staining was seen for TNF. These findings show that ameloblastomas synthesize two bone-modulating cytokines, IL-1alpha and IL-6, and that these are synthesized mainly by the stellate reticulum-like cells. These tumours also contain a proportion of activated blood vessels in which endothelial cells express the cellular adhesion receptors ICAM-1, E-selectin and VCAM-1.

Cell infiltrate in progressive pigmented purpura (Schamberg's disease): immunophenotype, adhesion receptors, and intercellular relationships.

Progressive pigmented purpura (Schamberg's disease), a form of purpura pigmentosa chronica, is a lymphocytic capillaritis of unknown etiology and obscure pathogenesis. Our purpose was to assess the expression of cell membrane antigens (CD3, CD4, CD1a, CD36), of adhesion receptors (leukocyte function adhesion 1, LFA-1, endothelial leukocyte adhesion molecule 1, ELAM-1) intercellular adhesion molecule 1, ICAM-1), and the intercellular relationships in the early phase of the disease. Quantitative immunohistochemistry and electron-microscopy were performed on specimens of five subjects, aged 45 to 63 years. These studies were repeated in two patients after treatment with topical corticosteroid (betamethasone valerate cream 0.1%) and psoralen-ultraviolet A (PUVA). The infiltrate consisted mainly of CD4+ lymphocytes and CD1a+ dendritic cells. Electron-microscopic investigation showed typical lymphocytes and two distinct types of dendritic cells. In the very early phase of the disease the adhesion receptors LFA-1 and ICAM-1 were expressed intensely by all infiltrating cells; the adhesion receptors ICAM-1 and ELAM-1 were expressed by endothelial cells. Close contact occurred between lymphocytes and dendritic cells. After PUVA (120 J per cm2) and topical steroid therapy the infiltrate disappeared completely. These data suggest that a cell-mediated immune mechanism may be important in progressive pigmented purpura and that the early endothelial expression of adhesion receptors may determine the pattern of organization of the pericapillary infiltrate.

Immunocytochemical localization of inflammatory cytokines and vascular adhesion receptors in radicular cysts

Odontogenic cysts are one of the commonest bone destroying lesions of the maxillofacial skeleton, with the inflammatory radicular cyst being the commonest jaw cyst. Explants of radicular cysts produce an interleukin-1-like activity which could explain the osteolysis seen with these tumours though the cellular source of this osteolytic activity is unknown. In the present study, cytokines with known inflammatory and osteolytic activity: interleukin-1 (IL-1), tumour necrosis factor (TNF), interleukin-6 (IL-6), and the chemotactic cytokine interleukin-8 (IL-8) have been localized immunocytochemically in radicular cysts. The cellular adhesion receptors ICAM-1 and ELAM-1 have also been immunolocalized. All specimens showed positive staining for IL-1 (alpha and beta) and IL-6, with these cytokines being located in epithelial and vascular endothelial cells. Only two specimens demonstrated TNF and IL-8 staining, which was located in macrophages. All specimens demonstrated ELAM-1 staining in endothelium and ICAM-1 staining in epithelium, endothelium and mononuclear cells. These findings show that radicular cysts contain two bone-modulating cytokines, IL-1 and IL-6, and that these appear to be synthesized mainly by the epithelial cells. Cysts also contain a proportion of activated blood vessels whose endothelial cells express the cellular adhesion receptors ICAM-1 and ELAM-1.