C-reactive protein induced expression of adhesion molecules in cultured cerebral microvascular endothelial cells

Ischemic stroke can trigger an acute phase response resulting in a rise of plasma concentration of C-reactive protein (CRP). Clinical data about the relationship between CRP and prognosis suggest that CRP might be involved in the pathogenesis of cerebral ischemia. In the present work, a significant increase of circulating level of CRP was observed in an vivo rat brain ischemia model of middle cerebral artery occlusion. To determine the possible effects of CRP on brain microvessel endothelium, we performed a dose-dependent experiment in mouse brain microvascular endothelial cells (bEnd.3 cells) with emphasis on its relation to cell adhesions molecules. Incubation with CRP (1-75 mg/L) for 24 h significantly increased Lactate dehydrogenase (LDH) leakage from bEnd.3 cells (P<0.01) in a dose-dependent manner, and induced significant up-regulations of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expressions analyzed by Western blotting (P<0.01). In contrast to earlier report, CRP also induced significant increase in ICAM-1 expression in the absence of serum (P<0.01). In conclusion, the present results suggest that CRP may be involved directly in the development of inflammation in response to cerebral ischemia.     

Co-stimulatory and adhesion molecules of dendritic cells in rheumatoid arthritis

Dendritic cells (DCs) in the rheumatoid arthritis (RA) joint mediate the immunopathological process and act as a potent antigen presenting cell. We compared the expression of co-stimulatory and adhesion molecules on DCs in RA patients versus controls with traumatic joint lesions and evalulated the correlation between the immunophenotypical presentation of DCs and the clinical status of the disease. Samples of peripheral venous blood, synovial fluid (SF) and synovial tissue (ST) were obtained from 10 patients with RA at the time of hip or knee replacement and from 9 control patients with knee arthroscopy for traumatic lesions. Clinical status was appreciated using the DAS28 score. Blood, SF and dissociated ST cell populations were separated by centrifugation and analyzed by flow cytometry. Cells phenotypes were identified using three-color flow cytometry analysis for the following receptors HLA-DR, CD80, CD83, CD86, CD11c, CD18, CD54, CD58, CD3, CD4, CD8, CD19, CD20, CD14, CD16, CD56. HLA-DR molecules, co-stimulatory receptors CD80, CD86, CD83 and adhesion molecules CD18, CD11c, CD54, CD58, were analyzed by two-color immunofluorescence microscopy on ST serial sections. In patients with active RA (DAS28>5.1) we found a highly differentiated subpopulation of DCs in the ST and SF that expressed an activated phenotype (HLA-DR, CD86+, CD80+, CD83+, CD11c+, CD54+, CD58+). No differences were found between circulating DCs from RA patients and control patients. Our data suggest an interrelationship between clinical outcome and the immunophenotypical presentation of DCs. Clinical active RA (DAS28>5.1) is associated with high incidence of activated DCs population in the ST and SF as demonstrated by expression of adhesion and co-stimulatory molecules.     

Simvastatin modulates TNFalpha-induced adhesion molecules expression in human endothelial cells

Adhesion and transendothelial migration of leukocytes into the vascular wall is a crucial step in atherogenesis. Expression of cell adhesion molecules by endothelial cells plays a leading role in this process. We investigated the effect of simvastatin, an inhibitor of HMG-CoA reductase administered to reduce plasma levels of LDL-cholesterol, on the expression of vascular cell adhesion molecule-1 (VCAM-1) and intracellular cell adhesion molecule-1 (ICAM-1) by human umbilical vein endothelial cells (HUVEC) stimulated with tumor necrosis factor alpha (TNFalpha). We found the expression to be significantly inhibited by the drug in a time and concentration-dependent manner and to a greater extent in the case of VCAM-1 as compared with ICAM-1. In TNFalpha-stimulated HUVEC, simvastatin decreased VCAM-1 and ICAM-1 mRNA levels, inhibited TNFalpha-induced activation of nuclear factor kappaB (NF-kappaB) and enhanced expression of peroxisome proliferator-activated receptor alpha (PPARalpha). These effects were associated with reduction of adherence of monocytes and lymphocytes to HUVEC. The present findings suggest that the benefits of statins in vascular disease may include the inhibition of expression of VCAM-1 and ICAM-1 through effects on NF-kappaB.     

Soluble intercellular adhesion molecules, soluble vascular cell adhesion molecules, and risk of coronary heart disease

Objective: We examined the association of circulating levels of soluble intercellular adhesion molecules (sICAM‐1) and soluble vascular cell adhesion molecules (sVCAM‐1) with coronary heart disease (CHD) risk factors and whether the adhesion molecules alone, and in combination, can serve as predictors of coronary CHD. Research Methods and Procedures: Among 18,225 men from the Health Professional Follow‐up Study who provided blood in 1994, we documented 266 incidents of non‐fatal myocardial infarction or fatal CHD during 6 years of follow‐up. The cases were matched 1:2 with non‐cases on age, smoking, and month of blood draw. We found both adhesion molecules directly associated with BMI, inflammatory biomarkers, and triglycerides and inversely associated with high‐density lipoprotein and alcohol intake (p < 0.05). After adjustment for C‐reactive protein, cholesterol‐to‐high‐density lipoprotein ratio, age, smoking, BMI, physical activity, alcohol intake, history of diabetes, parental history of CHD, aspirin use, antihypertensive drug use, and fasting status, the relative risk of CHD was 1.69 [95% confidence interval (CI), 1.14 to 2.51] for sICAM‐1 and 1.34 (95% CI, 0.91 to 1.96) for sVCAM‐1, when comparing the top quintile with the lower four quintiles. Control for other inflammatory or lipid biomarkers did not appreciably attenuate the associations. When we cross‐classified participants based on their sICAM‐1 and sVCAM‐1 levels, only the men in the top quintile of both biomarkers [relative risk = 2.39 (95% CI, 1.45 to 3.91)] had a significantly elevated risk of CHD (P interaction = 0.01, multivariate model). Discussion: sICAM‐1 and sVCAM‐1 are directly associated with obesity and other CHD risk factors. The combination of high levels of both adhesion molecules might be associated with the development of CHD, independent of other CHD risk factors.     

细胞粘附与树突状细胞迁移机制

树突状细胞（dendritic cell,DC)是迄今已知功能最强的抗原递呈细胞,DC体内迁移机制与其分化成熟。表型转换及生物学功能发挥密切相关。粘附分子及其介导的细胞粘附参与了DC的迁移机制。深入研究DC迁移的分子基础。有助于进一步阐明DC的生物学特性和功能。也可以通过DC进行免疫调控用于临床疾病防治提供重要理论基础。     

Interaction of rat sertoli cells with a collagen lattice in vitro

Summary Sertoli cells from rats aged 15, 20, and 25 d were subcultured onto collagen-coated, plastic dishes. If the collagen was released from the plastic surface by rimming, the floating rats of collagen showed uniform shrinkage. If the collagen was allowed to remain attached to the plastic, holes appeared in the collagen with cells from rats aged 25 d but not with those of 15 d. Cells from rats aged 20 d caused fewer and smaller holes to appear. The holes were associated with the formation of clumps of spherical cells from which elongated Sertoli cells extended into the surrounding collagen to end near holes. Rhodamine-phalloidin revealed diffusely distributed actin in the spherical cells in contrast to well-developed microfilaments in the peripheral elongated cells. Addition of cytochalasin B (5 μg/ml) to the medium prevented contraction of the floating rats and the development of holes in the attached collagen. In addition, cytochalasin B caused the peripheral cells to become spherical and to separate from the clumps. Moreover, rhodamine-phalloidin revealed that actin in the peripheral cells occurred as clumps without microfilaments when cytochalasin B was present. When Sertoli cells were subcultured onto silicone rubber films, the cells produced wrinkling of the rubber surface within 4 h of plating. These observations were interpreted to mean that Sertoli cells exert local tractional forces on various substrata. These forces require actin, presumably acting by a contractile mechanism. When the collagen is attached to plastic and the cells are organized into clumps with radiating elongated cells (cells from rats aged 25 d), the tractional forces in the elongated cells reorganize the collagen fibers to produce holes. When cells are uniformly distributed (cells from rats aged 15 d), holes are not formed. When the collagen is released from the plastic surface, tractional forces cause the floating rafts to shrink. These interactions of the cells with collagen are likely to be important in determining the shape of the Sertoli cell in vivo, the polarity of the cell, and its biochemical differentiation. This investigation was supported by grants HD 16525, AM 32236, and GM 32705 from the National Institutes of Health, and from the Shriners of North America.     

Hormone interactions in the sertoli cells

Summary The effects of follicle-stimulating hormone (FSH) and testosterone in rat Sertoli cells were investigated in vitro by means of isolated cell populations. The Sertoli cells selectively bind FSH, and respond to FSH stimulation with increased accumulation of endogenous cyclic AMP and secretion of androgen-binding protein (ABP). FSH binding and cyclic AMP response in the Sertoli cells change dramatically during sexual maturation. Cyclic AMP response decreases despite an increase in FSH-binding receptors per cell. Evidence has been provided for the existence of cytoplasmic and nuclear androgen receptors and chromatin acceptor-sites that specifically bind the androgen-receptor complex in the Sertoli cells. A model has been proposed for the hormonal interactions in the seminiferous tubule and the possible role of Sertoli cells in mediating the hormonal effects on spermatogenesis. Presented in the formal symposium on Sexual Differentiation in Vitro and in Vivo at the 29th Annual Meeting of the Tissue Culture Association, Denver, Colorado, June 4–8, 1978. This work was supported by Grant P50 HD08338 from the NICHHD. Dr. barbara M. Sanborn is a recipient of Research Career Development Award 1-K04-HD00126 (NIH).     

Modulation of phagocytic activity in cultured sertoli cells

Phagocytic activity of rat Sertoli cells that were cultured in vitro has been evaluated as the ability to internalize polystyrene beads. Our data demonstrate that these ceils are active phagocytes and that such phagocytic activity is under negative control by follicle-stimulating hormone (FSH). The hormonal control is mediated by increased intracellular levels of cAMP. Moreover Sertoli cdl responds to tuftsin, an oligopeptide known to act only on macrophages and granulocytes, by increasing up to five times its phagocytic activity. Phagocytic uptake of polystyrene beads is associated with dramatic changes of the cellular shape. Such morphological modifications are significantly reduced under FSH stimulation. The physiological implications of the data are discussed.     

Effect of selenium supplementation in asthmatic subjects on the expression of endothelial cell adhesion molecules in culture

Endothelial cells play a major role in immunologic reactions, in which cellular adhesion molecules P-selectin, ICAM-1, VCAM-1, and ELAM-1 are important mediators in the recruitment of leukocytes in pulmonary inflammation. Selenium (Se) is known to modulate immunological mechanisms of asthma. The aim of our investigation was to examine whether Se supplementation in cortico-dependent asthmatic patients may modulate adhesion molecule expression in cultured endothelium. Our findings indicated that P-selectin, VCAM-1, and ELAM-1 expression on human umbilical vein endothelial cells stimulated with peripheral blood mononuclear cells obtained from asthmatics before supplementation with Se was significantly higher than from healthy donors (p < 0.05). The production of ICAM-1 showed only slight augmentation. The levels of VCAM-1 and ELAM-1 expression were significantly decreased after 3 mo of Se supplementation (p < 0.05). After 6 mo of intervention period the intensity of P-selectin and ICAM-1 expression was also significantly reduced (p < 0.05 andp < 0.01, respectively). The inhibitory effect of Se on the adhesion molecule expression was studied in cultured endothelial cells after interferon-γ stimulation. Our data suggest that Se affects the expression of P-selectin, ICAM-1, VCAM-1, and ELAM-1 in a dosedependent manner and the half-maximal inhibitory concentrations were 3.4, 0.5, 4, and 3.8 μg/mL, respectively. The maximal inhibitions (greater than 80%) were observed in vitro with 10 μg/mL Se (p < 0.01). Regulation of adhesion molecule expression may be an important mechanism through which the inflammation may be controlled.     

Detection of intracellular cytokines during antioxidant supplementation in corticoid-dependent asthmatics and modulation of adhesion molecule expression on cultured endothelial cells

In this study, we report on the interferon-γ (IFN-γ) and interleukin-4 (IL-4) cytokine responses to phorbol myristate acetate (PMA)+ionomycin-stimulated CD3+ lymphocytes in asthmatic subjects when compared with normal donors. There was a significantly lower production of intracellular IFN-γ in asthmatic patients. No difference was found for IL-4 production between these two groups. After administration of a multivitamin-mineral supplement containing selenium, zinc, vitamin A, vitamin B₆, vitamin C, and vitamin E for 6 mo, a significant increase in the percentage of CD3+/IL-4 positive cells (p<0.05) was found. The induction of endothelial cell adhesion molecule (CAM) expression in cultured human umbilical vein endothelial cells (HUVEC) and whole-blood mixture was studied using flow cytometry. The ICAM-1 and VCAM-1 expressions were higher in the patients than in control donors (p<0.05). There is a correlation between the increased percentage of CD3+/IFN-γ positive cells and reduced endothelial ICAM-1 and VCAM-1 expression after 6 mo of intervention period. No apparent effect of supplementation on CAM expression was found, suggesting that these changes do not arise from an antioxidant mechanism. This newly developed whole-blood technique for the assessment of CAM expression can be of use for monitoring therapy in inflammatory diseases.     

Methyl mercury triggers endothelial leukocyte adhesion and increases expression of cell adhesion molecules and chemokines

Cardiovascular disease is the leading cause of morbidity, mortality, and health care costs in the USA, and around the world. Among the various risk factors of cardiovascular disease, environmental and dietary exposures to methyl mercury, a highly toxic metal traditionally labeled as a neurotoxin, have been epidemiologically linked to human cardiovascular disease development. However, its role in development and promotion of atherosclerosis, an initial step in more immediately life-threatening cardiovascular diseases, remains unclear. This study was conducted to examine the role that methyl mercury plays in the adhesion of monocytes to human microvascular endothelial cells (HMEC-1), and the underlying mechanisms. Methyl mercury treatment significantly induced the adhesion of monocyte to HMEC-1 endothelial cells, a critical step in atherosclerosis, while also upregulating the expression of proinflammatory cytokines interleukin-6, interleukin-8. Further, methyl mercury treatment also upregulated the chemotactic cytokine monocyte chemoattractant protein-1 and intercellular adhesion molecule-1. These molecules are imperative for the firm adhesion of leukocytes to endothelial cells. Additionally, our results further demonstrated that methyl mercury stimulated a significant increase in NF-κB activation. These findings suggest that NF-κB signaling pathway activation by methyl mercury is an important factor in the binding of monocytes to endothelial cells. Finally, by using flow cytometric analysis, methyl mercury treatment caused a significant increase in necrotic cell death only at higher concentrations without initiating apoptosis. This study provides new insights into the molecular actions of methyl mercury that can lead to endothelial dysfunction, inflammation, and subsequent atherosclerotic development.     

Alcohol exposure alters the expression pattern of neural cell adhesion molecules during brain development

Neural cell adhesion molecules (NCAMs) play critical roles during development of the nervous system. The aim of this study is to investigate the possible effect of ethanol exposure on the pattern of expression and sialylation of NCAM isoforms during postnatal rat brain development because alterations in NCAM content and distribution have been associated with defects in cell migration, synapse formation, and memory consolidation, and deficits in these processes have been observed after in utero alcohol exposure. The expression of NCAM isoforms in the developing cerebral cortex of pups from control and alcohol-fed mothers was assessed by western blotting, ribonuclease protection assay, and immunocytochemistry. The highly sialylated form of NCAM [polysialic acid (PSA)-NCAM] is mainly expressed during the neonatal period and then is down-regulated in parallel with the appearance of NCAM 180 and NCAM 140. Ethanol exposure increases PSA-NCAM levels during the neonatal period, delays the loss of PSA-NCAM, decreases the amount of NCAM 180 and NCAM 140 isoforms, and reduces sialyltransferase activity during postnatal brain development. Neuraminidase treatment of ethanol-exposed neonatal brains leads to more intense band degradation products, suggesting a higher content of NCAM polypeptides carrying PSA in these samples. However, NCAM mRNA levels are not changed by ethanol. Immunocytochemical analysis demonstrates that ethanol triggers an increase in PSA-NCAM immunolabeling in the cytoplasm of astroglial cells, accompanied by a decrease in immunogold particles over the plasma membrane. These findings indicate that ethanol exposure during brain development alters the pattern of NCAM expression and suggest that modification of NCAM could affect neuronal-glial interactions that might contribute to the brain defects observed after in utero alcohol exposure.     

Fine structure of leydig and sertoli cells in the testis of immature and mature spotted ray Torpedo marmorata

An ultrastructural investigation revealed the presence of true Leydig cells in the testis of sexually mature specimens of Torpedo marmorata. They showed the typical organization of steroid-hormone-producing cells, which, however, changed as spermatocysts approached maturity. In fact, they appeared as active cells among spermatocysts engaged in spermatogenesis, while in regions where spermiation occurred, they progressively regressed resuming the fibroblastic organization typically present in the testis of immature specimens. Such observations strongly suggest that these cells might be engaged in steroidogenesis and actively control spermatogenesis. Sertoli cells, too, appeared to play a role in spermatogenesis control, since, like Leydig cells, they showed the typical aspect of steroidogenic cells. In addition, the presence of gap junctions between Sertoli cells suggests that their activity might be coordinated. After sperm release, most Sertoli cells were modified and, finally, degenerated, but few of them changed into round cells (cytoplasts) or round cell remnants, which continued their steroidogenic activity within the spermatocyst and the genital duct lumen. From the present observations, it can be reasonably concluded that, in T. marmorata, spermatogenesis depends on both Leydig and Sertoli cells, and, as postulated by Callard (1991), in cartilaginous fish, the function of the Leydig cells as producers of steroids might be more recent and subsequent to that of Sertoli cells. In this regard, it is noteworthy that, in immature males, when Leydig cells showed a fibroblastic organization, Sertoli cells already displayed the typical organization of a steroidogenic cell.     

Characterization of a 140Kd cell surface glycoprotein involved in myoblast adhesion

Two monoclonal antibodies that cause changes in the morphology of cultured chick myogenic cells have been described previously [8]. In this paper, these antibodies are shown to interact with the same 140Kd protein. The 140Kd protein has been further characterized as a cell-surface glycoprotein by lactoperoxidase-catalyzed iodinations and lectin affinity chromatography. The protein is resistant to digestion by trypsin and collagenase and has been shown to be unrelated to fibronectin by immunoprecipitation studies and by peptide mapping. A second protein, of approximately 170Kd MW, is also immunoprecipitated by the monoclonal antibodies. This protein is probably unrelated to the 140Kd protein since the peptide maps are quite distinct.     

Expression of polysialylated neural cell adhesion molecules on adult stem cells after neuronal differentiation of inner ear spiral ganglion neurons

During brain development, polysialylated (polySia) neural cell adhesion molecules (polySia–NCAMs) modulate cell–cell adhesive interactions involved in synaptogenesis, neural plasticity, myelination, and neural stem cell (NSC) proliferation and differentiation. Our findings show that polySia–NCAM is expressed on NSC isolated from adult guinea pig spiral ganglion (GPSG), and in neurons and Schwann cells after differentiation of the NSC with epidermal, glia, fibroblast growth factors (GFs) and neurotrophins. These differentiated cells were immunoreactive with mAb’s to polySia, NCAM, β-III tubulin, nestin, S-100 and stained with BrdU. NSC could regenerate and be differentiated into neurons and Schwann cells. We conclude: (1) polySia is expressed on NSC isolated from adult GPSG and on neurons and Schwann cells differentiated from these NSC; (2) polySia is expressed on neurons primarily during the early stage of neuronal development and is expressed on Schwann cells at points of cell–cell contact; (3) polySia is a functional biomarker that modulates neuronal differentiation in inner ear stem cells. These new findings suggest that replacement of defective cells in the inner ear of hearing impaired patients using adult spiral ganglion neurons may offer potential hope to improve the quality of life for patients with auditory dysfunction and impaired hearing disorders.     

Mesenchymal entactin-1 (nidogen-1) is required for adhesion of peritubular cells of the rat testis in vitro

Epithelial-like Sertoli cells isolated from immature rat testis aggregate to form tubule-like structures when cultured on a monolayer of mesenchyme-derived peritubular cells. At the end of this morphogenetic process both cell types are separated by a basement membrane. In this study the gene expression of monocultures and direct cocultures of peritubular cells and Sertoli cells was examined using DD-RT-PCR. One of the isolated cDNA clones showed high homology to the cDNA encoding the basement membrane component entactin-1 (nidogen-1). Even though the entactin-1 (nidogen-1) gene is transcribed in peritubular cells, Sertoli cells, and in direct cocultures, the mRNA is translated only by the peritubular cells. No entactin-1 (nidogen-1) was detected in the Sertoli cells by Western blotting. Moreover, peritubular cell monocultures and cocultures showed the presence of one single band at 152 kDa in the supernatant, whereas in cell lysates two bands were detectable at 152 kDa and 150 kDa. Perturbation experiments using monoclonal antibodies directed against entactin-1 (nidogen-1) were performed with peritubular cells and Sertoli cells, respectively, and demonstrated loss of cell adhesion of the peritubular cells, while the Sertoli cells remained adherent. From these data we conclude that entactin-1 is exclusively produced and secreted by mesenchymal peritubular cells, and affects adhesion of peritubular cells in an autocrine manner.     

Changes in the sulfation extent of membrane-associated proteoglycans produced by sertoli cells in culture

Sertoli cells in culture synthesize two different membrane-associated proteoglycans (MA-PG): a proteoglycan containing heparan sulfate (HS) and chondroitin sulfate (CS) glycosaminoglycan (GAG) chains and a CS-PG containing only CS-GAG chains. The structure of these molecules is regulated by the presence of fetal calf serum (FCS) in the culture medium. Changes in the concentration of FCS resulted in changes in the total ³⁵SO₄ incorporation into MA-PG and a shift in the elution profile of each component subjected to ion-exchange chromatography. Thus, without FCS, the incorporation was low, while in 1% and 10% FCS, the uptake of the precursor was 1.7 and 4.5 times higher, respectively. MA-PG synthesized by Sertoli cells cultured in 10% FCS eluted from DEAE-Sephacel columns at higher salt concentration than the MA-PG synthesized by cells cultured in 0% or 1% FCS. Double-labeled experiments showed that the ³⁵SO_4/3H-glucosamine ratio incorporated into MA-PG produced by Sertoli cells, increased from 17.6 to 23.6 and 50.9 in cells cultured at 0, 1, and 10% FCS, respectively. However, the presence of FCS affected neither the hydrodynamic size nor the chemical nature of GAG chains of MA-PG. These results show that changes in the FCS concentration promote changes in the sulfation extent of MA-PG molecules produced by Sertoli cells.     

Murine junctional adhesion molecules JAM-B and JAM-C mediate endothelial and stellate cell interactions during hepatic fibrosis

Classical junctional adhesion molecules JAM-A, JAM-B and JAM-C influence vascular permeability, cell polarity as well as leukocyte recruitment and immigration into inflamed tissue. As the vasculature becomes remodelled in chronically injured, fibrotic livers we aimed to determine distribution and role of junctional adhesion molecules during this pathological process. Therefore, livers of naïve or carbon tetrachloride-treated mice were analyzed by immunohistochemistry to localize all 3 classical junctional adhesion molecules. Hepatic stellate cells and endothelial cells were isolated and subjected to immunocytochemistry and flow cytometry to determine localization and functionality of JAM-B and JAM-C. Cells were further used to perform contractility and migration assays and to study endothelial tubulogenesis and pericytic coverage by hepatic stellate cells. We found that in healthy tissue, JAM-A was ubiquitously expressed whereas JAM-B and JAM-C were restricted to the vasculature. During fibrosis, JAM-B and JAM-C levels increased in endothelial cells and JAM-C was de novo generated in myofibroblastic hepatic stellate cells. Soluble JAM-C blocked contractility but increased motility in hepatic stellate cells. Furthermore, soluble JAM-C reduced endothelial tubulogenesis and endothelial cell/stellate cell interaction. Thus, during liver fibrogenesis, JAM-B and JAM-C expression increase on the vascular endothelium. More importantly, JAM-C appears on myofibroblastic hepatic stellate cells linking them as pericytes to JAM-B positive endothelial cells. This JAM-B/JAM-C mediated interaction between endothelial cells and stellate cells stabilizes vessel walls and may control the sinusoidal diameter. Increased hepatic stellate cell contraction mediated by JAM-C/JAM-C interaction may cause intrahepatic vasoconstriction, which is a major complication in liver cirrhosis.     

Automated technique to evaluate thymocyte adhesion to thymic epithelial cells

Thymocyte adhesion to thymic epithelial cells is a relevant issue during intrathymic T-cell differentiation, and directly intervenes in the generation and expansion of the T-cell repertoire. In view of these data, it was apparent the usefulness of an automated strategy to evaluate the degree of thymic epithelial cell-thymocyte adhesion. This prompted us to develop an ELISA procedure (using an anti-Thy1 reagent) to determine the degree of thymocyte adhesion onto cultured thymic epithelial cells. The procedure described herein is simple, non-radioactive and reproducible. Additionally, it can potentially be applied to quantitate the degree of thymocyte adhesion to any cellular or non-cellular substrate (for example, extracellular matrix). Moreover, it detected fluctuations of thymocyte adhesion secondary to glucocorticoid treatment of epithelial cells. Thus, it can be regarded as a further tool to analyze intrathymic interactions.