Biomedical subjects

K Sugars

Publications and source records attributed to K Sugars.

6 recordsLinked to original sources

PubMed2000

Resting and activated T cells induce expression of E-selectin and VCAM-1 by vascular endothelial cells through a contact-dependent but CD40 ligand-independent mechanism.

This study explored the effect on endothelial cell (EC) activation of contact with T lymphocytes, which occurs during lymphocyte emigration into inflamed tissues. Addition of T cells to umbilical vein or dermal microvascular EC monolayers stimulated expression of EC E-selectin and VCAM-1. This response required direct cell:cell contact, but not T-cell activation. The capacity of resting CD4+ T cells to activate EC was restricted to the CD45RO+ subset and could be enhanced by 6 h prestimulation of T cells with PMA and ionomycin. The EC-stimulating capacity of resting or activated T cells was independent of CD40 ligand. Furthermore, inhibition of TNF-alpha/beta and IL-1alpha/beta, together with CD40 ligand, failed to inhibit EC activation by resting T cells and only inhibited the response to PMA- and ionomycin-activated T cells by 40 +/- 18%. Our data suggest that T-cell-EC interactions can lead to EC activation through a novel contact-dependent, but CD40 ligand-independent, mechanism.

CD40 Antigens↗

PubMed1999

Induction of decay-accelerating factor by cytokines or the membrane-attack complex protects vascular endothelial cells against complement deposition.

Vascular endothelium is continuously exposed to complement-mediated challenge, and this is enhanced during inflammation. Although the complement-regulatory proteins decay-accelerating factor (DAF), CD59, and membrane cofactor protein (MCP) protect endothelial cells (ECs) against complement-mediated injury, the control of their expression and relative contributions to vascular protection is unclear. We explored the hypothesis that mechanisms exist which induce upregulation of complement-regulatory proteins on ECs to maintain vascular function in inflammation. Tumor necrosis factor alpha (TNFalpha) and interferon gamma (IFNgamma) each increased DAF expression but not CD59 or MCP expression, and a combination of these cytokines was more potent than either alone. Cytokine-induced expression depended on increased DAF mRNA and de novo protein synthesis and was maximal by 72 hours. In addition, assembly of the membrane-attack complex (MAC) on ECs induced a 3-fold increase in DAF expression, and this was enhanced by cytokines. DAF upregulation was not inhibited by protein kinase C (PKC) antagonists. The increase in DAF was functionally relevant since it reduced complement 3 (C3) deposition by 40%, and this was inhibited by an anti-DAF monoclonal antibody. These observations indicate that upregulation of DAF expression by cytokines or MAC may represent an important feedback mechanism to maintain the integrity of the microvasculature during subacute and chronic inflammatory processes involving complement activation.

Antigens, CD↗

PubMed1998

Adhesion molecule expression in the lung: a comparison between normal and diffuse interstitial lung disease.

Cellular adhesion molecules are crucial determinants of the migration of immune effector cells to the tissues. In chronic inflammatory diseases, upregulation of the expression of these molecules may contribute to the persistent inflammatory process. The aim of this study was to determine whether there is evidence of adhesion molecule expression in chronically inflamed lung. Soluble adhesion molecules in bronchoalveolar lavage fluid (BALF) were measured by enzyme-linked immunoassay in 54 patients with chronic interstitial lung diseases and 16 normal controls. Adhesion molecule expression in fibrosing alveolitis (FA) lung and in control lung was assessed using immunohistology and reverse transcription-polymerase chain reaction (RT-PCR) amplification. Soluble intercellular adhesion molecule-1 (ICAM-1) was detected in all but two subjects. There was no difference in ICAM-1 concentration between disease groups and normal subjects. In contrast, soluble E-selectin was detected in 17 of the 70 subjects and was significantly associated with the presence of lung disease (p=0.0173). Furthermore, the presence of soluble E-selectin was associated with a raised lymphocyte percentage in BALF (p=0.0069). Soluble VCAM was only detected in five of the 70 subjects (two normals, three patients). There was no difference in adhesion molecule expression in lung parenchyma between FA and controls assessed by immunohistology and RT-PCR. The most striking finding of our study was the universal expression of intercellular adhesion molecule-1 in both normal and diseased lung, emphasizing the important role of the lung in immune function. Upregulation of E-selectin may contribute to inflammatory cell accumulation in chronic interstitial lung diseases.

Adolescent↗

PubMed1997

Human umbilical vein and dermal microvascular endothelial cells show heterogeneity in response to PKC activation.

Changes in endothelial cell (EC) phenotype are central to the function of endothelium in inflammation. Although these events mainly occur in the microvasculature, previous studies have predominantly used large-vessel EC. Using enzyme-linked immunosorbent and flow cytometric assays, we compared the responses of human umbilical vein endothelial cells (HUVEC) and dermal microvascular endothelial cells (DMEC) to the activation of protein kinase C (PKC). Stimulation with phorbol 12,13-dibutyrate and more selective PKC agonists, including 12-deoxyphorbol 13-phenylacetate 20-acetate (dPPA), induced morphological changes and proliferation in both EC types. PKC activation induced a marked increase in Thy-1 expression on DMEC and only a moderate rise on HUVEC. Furthermore, heterogeneity in the induction of the adhesion molecules intercellular adhesion molecule 1, vascular cell adhesion molecule 1 IVCAM-1), and E-selectin between the two EC types following activation of PKC was demonstrated. In particular, E-selectin and VCAM-1 were significantly upregulated on HUVEC but not DMEC. The data indicate that the PKC pathway is unlikely to be important for E-selectin and VCAM-1 expression in the microvasculature but are consistent with a role for PKC in angiogenesis. This diversity in signaling in response to PKC activation may depend on differential utilization of PKC isozymes and may facilitate specialized endothelial responses.

Cell Adhesion Molecules↗

PubMed1996

Human Thy-1 is cytokine-inducible on vascular endothelial cells and is a signaling molecule regulated by protein kinase C.

Thy-1 is a glycosylphosphatidylinositol-anchored member of the Ig superfamily whose function, particularly in the human, remains unknown. We have demonstrated that human Thy-1 is expressed on endothelial cells (EC) both in situ and on the surface of cultured human umbilical vein EC and dermal microvascular EC (DMEC). The expression of the molecule decreased with serial passage but was restored by treatment of EC with PMA and phorbol-12,13-dibutyrate (PBu), which increased Thy-1 by up to 100-fold in a dose-dependent manner. This increase was first detectable at 12 h post-stimulation, peaked at 48 h, and was maintained at 72 h. In PBu-stimulated DMEC, Western blotting revealed Thy-1 to be a 29-kDa molecule, while Northern analysis demonstrated an increase in steady-state Thy-1 mRNA. Thy-1 expression was also induced on DMEC by treatment with TNF. Inhibition studies showed that the induction of Thy-1 by PBu and TNF was protein synthesis dependent. The up-regulation of Thy-1 by PBu, but not TNF, was inhibited by the protein kinase C inhibitor RO31-8220, suggesting the presence of protein kinase C-dependent and -independent pathways for Thy-1 expression. To investigate the function of Thy-1 on human EC, we studied changes in intracellular calcium concentration ([Ca2+]i) following cross-linking of Thy-1 on human umbilical vein EC. This resulted in a rapid transient rise of [Ca2+]i in EC. Our results demonstrate for the first time the presence of Thy-1 on cultured human EC. The expression on EC, the inducibility by TNF, and the ability to transmit signals suggest that Thy-1 may have an important role in inflammatory responses.

Antibodies, Monoclonal↗

PubMed1994

Measurement of mRNA for E-selectin, VCAM-1 and ICAM-1 by reverse transcription and the polymerase chain reaction.

Stimulation of cultured human umbilical vein endothelial cells by cytokines such as interleukin-1 and tumour necrosis factor induces de novo synthesis and expression of the adhesion molecules E-selectin, vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1). In general, alterations in cell surface expression of these molecules are known to be related to increased gene transcription and altered levels of mRNA. The extension of these observations to the study of inflammatory processes in different human organs necessitates the development of techniques for the quantification of mRNA in small tissue samples. Here we present a method for the quantification of mRNA for E-selectin, VCAM-1 and ICAM-1 using reverse transcription and the polymerase chain reaction (RT-PCR). For each molecule of interest a mutant RNA was synthesised consisting of the wild-type sequence deleted of 15-20 bases. The mutant and wild-type RNA sequences are recognised by the same primers, and can therefore be amplified competitively in the same tube by RT-PCR. As the mutant and wild-type RNAs compete for the primers, the amount of wild-type RNA can be determined by the size of the dominant product that results after addition of known quantities of mutant RNA. Using this detection and quantification method we have examined the dose dependency and time course of mRNA accumulation following TNF-alpha stimulation of HUVEC. Similar time-courses of E-selectin, ICAM-1 and VCAM-1 mRNA accumulation were observed by competitive RT-PCR as by laser densitometry of Northern blots. Finally we were able to show that the technique could measure changes in levels of mRNA for these three molecules in human skin biopsies taken at different times during the development of a delayed hypersensitivity response to tuberculin purified protein derivative. This technique should be useful for the study of adhesion molecule mRNA in small tissue culture samples and in biopsies.

Base Sequence↗