Immune mechanisms in atherosclerosis.
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Animals↗
Biomedical subjects
S Stemme
Publications and source records attributed to S Stemme.
Immune mechanisms in atherogenesis.
Research in recent years has resulted in an increased understanding of the molecular mechanisms in the development of inflammatory processes. In atherosclerosis, focal expression of key adhesion molecules has been detected which may mediate the recruitment of mononuclear cells to the plaque. Local cytokine production could account for further cell migration and proliferation. The presence of substantial numbers of T lymphocytes in the plaque and local and circulating autoantibodies to modified lipoproteins suggest that T and B lymphocyte responses may play important roles in these processes.
Animals↗
Adhesion molecule expression on cerebrospinal fluid T lymphocytes: evidence for common recruitment mechanisms in multiple sclerosis, aseptic meningitis, and normal controls.
The expression of T-cell surface antigens was investigated in the cerebrospinal fluid (CSF) and peripheral blood of 11 patients with multiple sclerosis, 6 patients with aseptic meningitis, and 16 healthy subjects. A panel of monoclonal antibodies to adhesion and activation proteins was used in combination with an anti-CD3 antibody in dual-color flow cytometry. The problem of low cell numbers in the CSF from normal individuals was overcome by use of a modified staining procedure in microtiter plates, enabling analysis of as few as 5,000 cells. The majority of T cells in the CSF of the three patient groups exhibited the phenotype of memory cells (CD45RO+). CSF T cells also expressed significantly higher levels of several adhesion and activation molecules, including very late activation (VLA) antigens 3 through 6, lymphocyte function-associated (LFA) antigen 1, LFA-3, CD2, CD26, and CD44. Comparison between the different categories revealed that peripheral blood T cells from patients with multiple sclerosis expressed significantly lower amounts of the VLA integrins 4 and 5 as well as their common beta subunit CD29, compared with normal control subjects. No differences between patients with multiple sclerosis and control subjects could, however, be seen regarding the distribution of memory/naive cells or CD4+/CD8+ cells in peripheral blood. Our data support a hypothesis that memory T cells with a high expression of several adhesion molecules are selectively recruited to the central nervous system compartment, under both pathological and normal conditions.(ABSTRACT TRUNCATED AT 250 WORDS)
Adolescent↗
T lymphocytes in giant cell arteritic lesions are polyclonal cells expressing alpha beta type antigen receptors and VLA-1 integrin receptors.
Giant cell arteritis (GCA) is a common disease in the elderly. It is characterized by focal inflammatory lesions dominated by T lymphocytes and macrophages. The etiology of GCA is, however, still unknown. The aim of the present study was to determine whether lesional T cells represent clonal proliferations, and to characterize adhesion receptors that could be important for recruitment of T cells and antigen receptors involved in their activation. Temporal artery biopsies were obtained from 13 patients presenting with clinical signs of GCA. Immunohistochemistry was used to characterize cell surface receptors on CD3+ T cells in situ in the lesions of eight patients with biopsy-verified GCA. The overwhelming majority of T cells in GCA lesions expressed the TCR alpha beta receptors. In sections from three of eight patients, a small proportion of cells expressing TCR gamma delta was also seen. Almost all T cells expressed the integrin receptors, LFA-1 and VLA-1, as determined by double-staining. To characterize the clonal composition of the lesional T cell population, cells were isolated by collagenase digestion of two lesions and T cells cloned by limiting dilution in the presence of mitogenic antibodies, IL-2 and autologous feeder cells. Rearrangements of the T cell receptor (TCR) genes of the clones were analysed by Southern hybridization using probes for TCR gamma and beta genes. T cell clones established from GCA lesions exhibited heterogeneous rearrangement patterns, indicating a polyclonal origin of the cells. We conclude that GCA lesions contain T lymphocytes that are of polyclonal origin and express integrin-type adhesion receptors. This supports the hypothesis that GCA involves an inflammatory response during which polyclonal T cells adhere to arterial tissue components and accumulate in the developing lesions.
Aged↗
Lymphocyte adhesion and cellular immune reactions in chronic rejection and graft arteriosclerosis.
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Arteriosclerosis↗
Quantitation and phenotyping of T cell clones by flow cytometry.
In order to efficiently analyze large numbers of T cell clones, a method for analysis of small cell cultures by flow cytometry was developed. The aim was to assess cloning efficiency, growth rate and phenotype of T cell clones. The reliability of the flow cytometer for quantitation of cell populations was documented by repeated analysis of manually counted cell samples. In the concentration range from 3000 to 195,000 cells/ml, the correlation coefficient between counts obtained by the flow cytometer and manual cell counts was 0.999 and within-assay coefficients of variation were below 4%. Cell cultures containing less than 200 cells were reliably quantified. The technique was applied for analysis of T cell clonings with different mitogens and in the presence of varying amounts of serum. To reduce time and labour, the cultures were analyzed only 10 days after cloning, when the clones contained less than 100,000 cells. The sensitivity of the flow cytometer in the detection of immunolabeled cells made further expansion of cell cultures unnecessary, thus greatly reducing manual labour and experiment turnover time. The commonly used mitogens phytohemagglutinin (PHA) and the antibody OKT3 resulted in comparable cloning efficiencies and clone sizes. Human serum was essential for high cloning efficiency as well as for continued growth, and could not be substituted with an increased amount of fetal calf serum. When cloning with interleukin-2 at 20,000 U/ml, two growing cell types were identified. The majority of the clones contained CD3+, CD4+, or CD8+ T cells. Ten out of 60 cultures however, contained cells with the CD3-16/56+ NK cell phenotype, indicating that the culture conditions stimulated proliferation of two different cell types. The described method can be applied for rational analysis of large numbers of minute cell cultures, in for example evaluation of different cloning conditions and estimation of precursor cell frequencies in limiting dilution analysis. The simultaneous phenotyping allows precursor cell analysis under conditions that stimulate growth of more than one cell type.
Antigens, Differentiation, T-Lymphocyte↗
Adhesion of activated T lymphocytes to vascular smooth muscle cells and dermal fibroblasts is mediated by beta 1- and beta 2-integrins.
Several studies during recent years have demonstrated the potential for vascular smooth muscle cells (SMC) and dermal fibroblasts to participate in immune interactions such as antigen presentation and alloreactivity. The molecular interactions mediating lymphocyte adhesion to these mesenchymal cells have, however, not previously been characterized in detail. In the present study we demonstrate ICAM-1 (CD54) expression by cultured human SMC and its up-regulation by IL-1, IFN-gamma, and bacterial lipopolysaccharide. Monoclonal antibodies were used to define the molecular interactions in the adhesion of 51Cr-labelled T lymphoblasts to adherent SMC and fibroblasts. ICAM-1 appeared to mediate adhesion of T lymphocytes by binding to the beta 2-integrin CD11a/CD18 (LFA-1) expressed by the lymphoblasts. We present evidence for the involvement of at least three different mechanisms in the adhesion of activated T lymphocytes to cultured fibroblasts. It was found that beta 2-integrin-mediated interaction could only account for less than half of the binding activity. The remaining adhesion was partly mediated by beta 1-integrins, presumably via VLA-5 since an anti-VLA-5 antibody and an RGD-containing peptide blocked adhesion to the same degree. However, antibodies to beta 1-, beta 2-, and beta 3-integrin subunits added together only inhibited adhesion by approximately 50%. The residual adhesion could be blocked by inhibition of cell metabolism and was increased by stimulation of the lymphocytes with phorbol ester, suggesting involvement of other, as yet undefined, adhesion molecules. The molecular interactions between lymphocytes and mesenchymal cells demonstrated in this study may have implications in several inflammatory conditions such as vasculitis, atherosclerosis, and connective tissue diseases.
Antigens, CD↗
T lymphocytes in human atherosclerotic plaques are memory cells expressing CD45RO and the integrin VLA-1.
The cellular composition of human atherosclerotic plaques has been analyzed in several immunohistochemical studies in recent years. These studies have shown that the main cell types of the plaque are macrophages, smooth muscle cells, and T lymphocytes. To further characterize the T-lymphocyte population in atherosclerotic plaques, human plaque tissue was digested enzymatically and the released cells were labeled with fluorescent antibodies and analyzed by flow cytometry. Fifteen patients undergoing carotid endarterectomy were studied. Sixty-four percent of plaque T cells expressed the low-molecular-weight form (CD45RO) of the leukocyte common antigen (CD45). Many of these cells expressed the integrin very late activation antigen-1 (VLA-1), which suggests that they are in a state of late activation. In contrast, only 1% of peripheral blood T cells from the same patients expressed VLA-1. Other markers of T cell activation, such as Ta1 (CD26) and HLA-DR, were also increased on plaque T cells. The interleukin-2 receptor (CD25), which is transiently expressed after activation, was present on only a small proportion of the cells. Taken together, this analysis of plaque lymphocytes shows that the majority of plaque T cells are memory cells, many of which are in a state of late or chronic activation. This T-cell phenotype may be the result of a preferential recruitment and/or retention of activated peripheral blood T cells or local antigenic stimulation of resting T cells.
Aged↗
Flow cytometric analysis of reticulocytes using an RNA-binding fluorochrome.
The reticulocyte count is an important parameter in the diagnosis of anaemia. The commonly used microscopic counting technique is, however, laborious and hampered by poor precision and low sensitivity. An alternative method has recently been developed, which is based on flow cytometric identification and quantification of reticulocytes after staining with an RNA-binding fluorochrome, thiazol orange. The two methods were compared both in a sample of healthy individuals and in patient specimens. The correlation between the methods was good, but the flow cytometric analysis consistently gave values approximately 1.5 times higher (reticulocytes in per cent of erythrocytes) than the microscopic technique. The coefficient of variation was significantly lower for the flow cytometric technique. Reference values for the reticulocyte count were determined and the effects of storage of specimens were evaluated. It is concluded that flow cytometric analysis of reticulocytes is more sensitive and has a better precision that the standard microscopic procedure.
Anemia, Aplastic↗
Can immunocompetent cells and their cytokines play a role in atherogenesis?
The human atherosclerotic plaque represents a mixture of inflammation, lipid accumulation and fibrosis. Cholesterol deposits are predominantly found in the core of the plaque, which is surrounded by a fibrotic cap. Much of the cholesterol is intracellular in macrophage-derived foam cells, and other macrophages together with T lymphocytes are intermixed with smooth muscle cells in the fibrous cap. Recent studies have shown that cytokines released by inflammatory cells can regulate cell proliferation, cholesterol metabolism, and differentiation of vascular cells. Interferon-gamma, one of the major secretory products of activated T lymphocytes, inhibits smooth muscle proliferation and reduces expression of alpha-actin. This suggests that T cells, by releasing gamma-interferon, may inhibit the formation of arterial stenosis and spasm. Recent animal experiments support this hypothesis by demonstrating that injections of recombinant gamma-interferon reduce arterial intimal lesions after balloon catheter injury. In similar experiments, it was shown that larger intimal lesions develop in T cell-depleted rats and in athymic nude rats. This shows that T cells inhibit the vascular response to injury and supports the notion that gamma-interferon released by these T cells is an important paracrine regulator of arterial cell proliferation. The cytokines, gamma-interferon and TNF, also downregulate scavenger receptor expression in monocyte-derived macrophages, leading to an inhibition of the transformation of these cells into foam cells. Taken together, these data show that cytokines of the immune system are important regulators of cholesterol deposition and cell proliferation in atherogenesis.
Arteriosclerosis↗
Polyclonal origin of T lymphocytes in human atherosclerotic plaques.
The human atherosclerotic plaque contains large numbers of T lymphocytes and macrophages; this indicates that immune and inflammatory mechanisms may be important factors in the pathogenesis of atherosclerosis. A significant proportion of the T lymphocytes express activation markers, which suggests that they may be stimulated by local antigens, proliferate in response to such antigens, and secrete lymphokines that could serve as paracrine factors in the arterial tissue. However, it is not known, whether plaque T lymphocytes constitute a homogeneous population of clonally proliferating cells or represent a heterogeneous mixture of cells with different immunologic specificities. Clonally-derived T lymphocytes can be identified since they carry identical T cell antigen receptor (TCR) genes. These genes rearrange during T cell ontogeny resulting in TCR genes and TCR proteins that are unique for each T cell clone. We have now employed TCR gene analysis to T lymphocytes derived from atherosclerotic plaques in order to determine their clonal composition. T lymphocytes were isolated from carotid endarterectomy samples and cultured after limiting dilution cell cloning. TCR genes were analyzed by Southern blotting using probes for the TCR gamma (J gamma 2) and TCR beta (C beta 1) genes. TCR gene rearrangement patterns were totally heterogeneous, indicating that plaque T lymphocytes constitute a polyclonal population of cells. This suggests that the T cells are either recruited to the plaque in an activated state or activated locally by mechanisms that do not lead to clonal proliferation.
Aged↗
MHC class II antigen expression in human vascular smooth muscle cells is induced by interferon-gamma and modulated by tumour necrosis factor and lymphotoxin.
Arterial smooth muscle cells (SMC) express major histocompatibility complex (MHC) class II antigens in experimental vasculitis and in the human atherosclerotic plaque. We have therefore studied the regulation of expression of MHC antigens in cultured human arterial SMC, using immunofluorescence, radioimmunoprecipitation and a quantitative cell-surface immunoradiometric assay. SMC expressed class I, but not class II, antigens on their cell surfaces under basal conditions. Treatment of SMC with recombinant or natural interferon-gamma (IFN-gamma) induced expression of class II antigens in the following order of intensity, DR greater than DP greater than DQ. HLA-DR protein in SMC showed the same MW as that synthesized by B-lymphoblastoid cells. Antibodies to IFN-gamma blocked all HLA-DR-inducing activity in mixed leucocyte reaction (MLR) supernatants and PHA-stimulated peripheral blood mononuclear cell (PBMC)-conditioned media, indicating that IFN-gamma is the only lymphokine secreted under these conditions that is capable of de novo induction of HLA-DR expression in SMC. Treatment of SMC with recombinant human tumour necrosis factor-alpha (TNF) or lymphotoxin (LT) did not per se induce class II antigen expression. However, both TNF and LT substantially enhanced IFN-gamma-induced expression of HLA-DQ while decreasing that of HLA-DP. TNF, but not LT, increased HLA-DR expression. Also, in dermal fibroblasts, IFN-gamma-induced HLA-DP expression was significantly inhibited in the presence of TNF. These data demonstrate that TNF and LT differentially modulate IFN-gamma-induced MHC antigen expression in mesenchymal cells. The fact that SMC can express MHC class II antigens suggests that this cell type may serve as an accessory cell in the initiation of the immune response.
Cells, Cultured↗
Immunologic control of vascular cell growth and differentiation.
Several recent studies demonstrate that T lymphocytes and monocytes enter the vessel wall during pathologic conditions and accumulate both in atherosclerotic and arteritic lesions. This brief review summarizes our phenotypic analysis of such T lymphocytes. Effects of T lymphocytes and their secretory products (lymphokines) on vascular cells are discussed on the basis of both cell culture studies and animal experiments. Finally, the role of complement activation for recruitment of blood-borne cells is evaluated.
Animals↗
Immunologic control of vascular cell growth in arterial response to injury and atherosclerosis.
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Arteriosclerosis↗
Localization of T lymphocytes and macrophages in fibrous and complicated human atherosclerotic plaques.
The cellular composition of aortic atherosclerotic plaques was analyzed by immunocytochemistry using cell type-specific monoclonal antibodies. T lymphocytes and monocytes/macrophages were detected both in early, fibrous plaques, and in more advanced, complicated ones. Many smooth muscle cells in these plaques expressed the class II MHC antigen, HLA-DR. Since this antigen is inducible by T cell products, our findings suggest that T cell-smooth muscle interactions occur during atherogenesis.
Antibodies, Monoclonal↗
Different cellular distribution of thioredoxin and subunit M1 of ribonucleotide reductase in rat tissues.
The cellular distribution of thioredoxin and protein M1 of ribonucleotide reductase in adult rat tissues was investigated with immunohistochemical techniques using specific antisera. Tissues with high or low frequency of either mitotic or meiotic cell divisions were compared. Thioredoxin was demonstrated in many cells types that showed no detectable protein M1 of ribonucleotide reductase. A few cell types with protein M1 immunoreactivity also contained immunoreactive thioredoxin. However, in most cells no such co-localization could be demonstrated. This lack of correlation between cells containing subunit M1 of ribonucleotide reductase and the thioredoxin indicates that thioredoxin is not the physiologist hydrogen donor for ribonucleotide reductase in rat tissues and that the expression of two enzymes is differently regulated.
Animals↗
Localization of IGF-I in adult rats. Immunohistochemical studies.
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Animals↗
Evidence indicating trophic importance of IGF-I in regenerating peripheral nerves.
The mechanisms influencing regeneration of peripheral nerves are incompletely known, but growth factors are supposed to play a key role. In the present study, we demonstrate, with the aid of immunohistochemical methods, that somatomedin C (Sm-C/insulin-like growth factor I/IGF-I) rapidly increased from low to high concentrations, reaching peak values in 2 weeks, in regenerating sciatic nerves of adult rats. In addition, IGF-I was demonstrated extracellularly, never observed in the control nerves. Reactive Schwann cells appeared to be the major source for IGF-synthesis. Higher concentrations were seen in tubulated nerves as compared to sutured ones. It is proposed that IGF-I exerts important growth supporting effects on regenerating peripheral nerves.
Animals↗