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Systemic sclerosis (SSc) or scleroderma is a rare, autoimmune, multi-factorial disease characterized by early microvascular alterations, inflammation, and fibrosis. Chickens from the UCD-200 line develop a hereditary SSc-like disease, showing all the hallmarks of the human disorder, which makes this line a promising model to study genetic factors underlying the disease. A backcross was generated between UCD-200 chickens and its wild ancestor - the red jungle fowl and a genome-scan was performed to identify loci affecting early (21 days of age) and late (175 days of age) ischemic lesions of the comb. A significant difference in frequency of disease was observed between sexes in the BC population, where the homogametic males were more affected than females, and there was evidence for a protective W chromosome effect. Three suggestive disease predisposing loci were mapped to chromosomes 2, 12 and 14. Three orthologues of genes implicated in human SSc are located in the QTL region on chromosome 2, TGFRB1, EXOC2-IRF4 and COL1A2, as well as CCR8, which is more generally related to immune function. IGFBP3 is also located within the QTL on chromosome 2 and earlier studies have showed increased IGFBP3 serum levels in SSc patients. To our knowledge, this study is the first to reveal a potential genetic association between IGFBP3 and SSc. Another gene with an immunological function, SOCS1, is located in the QTL region on chromosome 14. These results illustrate the usefulness of the UCD-200 chicken as a model of human SSc and motivate further in-depth functional studies of the implicated candidate genes., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Atherosclerosis is a multifactorial, chronic-inflammatory disease for which the underlying cause remains unknown. It is also well documented that T-cells are among the first cells to migrate into the arterial intimal vessel layer, but their function there is still unexplained. Clinical and experimental data have provided evidence that atherosclerosis starts as an autoimmune reaction based on humoral and cellular immunity against a phylogenetically highly conserved stress protein, heat shock protein 60 (HSP60). In the present study, we phenotypically characterized T-cells from endarterectomized specimens of the carotid artery, and tested their reactivity to human HSP60. In addition, the T-cell receptor repertoire of the T-cell lines was defined by immunoscope analysis. We found a mixed population of CD4(+) and CD8(+) intralesional T-cells, with a slight predominance of CD8(+) cells. IFN-gamma production prevailed over IL-4 production. The T-cell reaction against human HSP60 was significantly increased in intralesional cells compared to peripheral T-cells. The lesion-derived T-cells showed an oligoclonally-restricted repertoire, in contrast to the polyclonal pattern of PBMC. These results clearly show that HSP60 is a major antigenic candidate, and that an oligoclonal T-cell expansion takes place in advanced human atherosclerotic lesions.

Cigarette smoking reduces life span by an average of 7 years, and tobacco consumption accounts for a shortening of disease free life by 14 years. The exact mechanisms by which smoking causes disease and death are generally not well understood, but evidence continues to mount that cigarette smoking exhausts cellular defense and repair functions, leading to an accumulation of damage e.g. mutations and malfunctioning proteins. In this review, we make an attempt to ascribe many of the deleterious effects of smoking on human health to a general principle, namely the acceleration of aging processes by cigarette smoke chemicals.

Humans and animals are structurally designed as a compromise to guarantee optimal survival until the time of reproduction based on natural selection that is effective until that age. Modern conditions of life including improvement of hygiene, preventive and curative medicine as well as socio-economic and political developments have led to an increase of the mean life expectancy that allows ever larger proportions of the population to reach an age that is far beyond that of the reproductive phase. The concept of a Darwinian-evolutionary basis for the development of age-related diseases in principle postulates that genetic traits that are beneficial in younger years to allow for successful reproduction may become deleterious in the elderly, i.e. when selective pressure does not seem to be effective anymore. Examples for this mechanism of pleiotropic antagonism taken from the work of the Institute for Biomedical Aging Research in Innsbruck, Austria, include atherosclerosis, benign and malignant prostate hypertrophy, Alzheimer's disease and the reciprocal relationship between cellular senescence and cancer.

'Anti-aging' medicine has become a popular topic, in the lay press, the (semi-) scientific literature and the Internet.(1) In almost all instances, claims of drugs, health supplements and other types of intervention are not based on any evidence supported by sound scientific knowledge. Specifically, the aging process itself, in contrast to age-related diseases, has so far neither been fully understood, nor significantly influenced in higher species, notably humans. In addition to the ill-defined effects, claims of anti-aging medicine also pose a considerable economic burden on the usually poorly informed older segment of the population. The present paper is based on a recent report by the United States General Accounting Office (GAO) on the questionable and even harmful effects of anti-aging health products for seniors.

In this review, we summarize data concerning the respective preservation and deterioration of antigenic determinants of various collagenous and non-collagenous extracellular matrix (ECM) proteins in palaeontologic material of different ages. ECM proteins are the major quantitative constituents of mammalian organisms and were, therefore, selected as important representative proteins for these analyses. The specimens, studied by immunofluorescence and immunohistochemical techniques, included the skin of 500-1500 year-old human mummies from Peru, skin and striated muscle from the 5300-year-old glacier mummy ("Iceman") from Tyrol, Austria, and a 50-million-year-old bat with preserved soft body parts from the fossil excavation site of Messel, Germany. In frozen sections of the former two sources, epitopes recognized by specific antibodies for triple-helical antigenic determinants of different types of collagen resistant against conventional proteases were preserved, while non-helical domains, as well as the non-collagenous ECM proteins, could no longer be demonstrated. The fossil bat, although showing evidence of fibrous, collagen-like structures in conventional histology, revealed no collagenous or non-collagenous ECM proteins by any technique. It later turned out that this was due to the replacement of the original soft parts in these fossils by lawns of bacteria. These studies introduced immunological techniques into palaeontology and opened new approaches for studying physiologically- and pathologically-altered structures in tissues of animals and humans of considerable historical age.

Austria has a long and successful tradition in geriatric medicine, nursing and sociology. In contrast, research in experimental gerontology has only come into focus more recently. Gerontological research is performed in various laboratories and clinics of the Medical Schools of the Universities of Vienna, Graz and Innsbruck, respectively, as well as in the School of Veterinary Medicine in Vienna, the Institute for Applied Microbiology of the Vienna University for Agricultural Sciences, the Faculty of Natural Sciences of the University of Salzburg, and in extra-universitary institutions such as the Ludwig-Boltzmann-Institute for Aging Research in Vienna and the Institute for Biomedical Aging Research of the Austrian Academy of Sciences in Innsbruck. Therefore, the following brief reflections will be organized in a topographic fashion.

By definition, diseases of aging become clinically manifested in elderly patients. However, their pathogenetic basis has to be sought earlier in life. The general thread of this presentation relies on the concept of an evolutionary-Darwinian view of the development of age-related diseases. In essence, this concept states that we may have to "pay" for genetic traits that play a beneficial role earlier in life by the later development of diseases since there is no post-reproductive selective pressure that may have eliminated the potential late onset detrimental effects of such genes. Examples for this kind of trade-off are taken from diseases involving the immune system (infections), the endocrine system (andropause), the nervous system (Alzheimer's disease), the locomoter system (osteoporosis), the cardio-vascular system (atherosclerosis) and cancer.

Immune-inflammatory processes are increasingly discussed as possible pathogenetic factors involved in the development of atherosclerosis. Here, we summarize data on which we have built our "immunological" hypothesis of atherogenesis. This concept is based on the observation that nearly everybody shows protective cellular and humoral immune reactions against microbial heat shock protein 60 (HSP 60). Because a high degree of antigenic homology exists between microbial (viral, bacterial, parasitic) and human HSP 60, this protective immunity may have to be "paid for" by the danger of cross-reactivity with human HSP 60 that is expressed by endothelial cells of stressed arteries. Arterial endothelial cells are more prone to produce HSP 60 and various adhesion molecules upon exposure to stress factors, including classical risk factors for atherosclerosis, due to their life-long exposure to the high arterial as compared to venous blood pressure. Also, endothelial cells are the first potential targets encountered by circulating HSP 60-specific T cells or antibodies. This concept not only opens new avenues for diagnostic approaches, but also may form the basis for new ways of therapeutic intervention.

We investigated the involvement of mitogen-activated protein kinase (MAPK) signal transduction pathways in human endothelial cells in response to shear stress and alterations of these kinases in in vitro-propagated endothelial cells (ECs). Potent activation (10-fold) of extracellular signal-regulated kinase (ERK2), a member of the MAPK family, occurred within 10 min of shear stress (5 dynes/cm2), whereupon rapid inactivation ensued. Shear stress also induced activation of stress-activated protein kinase (SAPK) or c-Jun NH2-terminal protein kinase (JNK) in ECs. Suramin pretreatment completely inhibited shear stress stimulation of ERK2, but not SAPK/JNK, highlighting a role for growth factor receptors in ERK activation. Translocation of ERK2 from the cytoplasm to the nucleus was observed in shear-stressed endothelial cells. In addition, we compared activities of MAPKs in shear-stressed cells derived from passages 4 and 10 (older). The magnitude of ERK2 activation was significantly lower in aged ECs compared to those of passage 4, while SAPK/JNK was not altered in the in vitro aged ECs. A similar level of ERK2 activation was found in both young and older cells stimulated with phorbol-12-myristate-13-acetate (PMA), indicating an age-related alteration of the plasma membrane. Taken together, these findings suggest that MAP kinase activation may be crucial for the expression of many genes in ECs stimulated by shear stress, and that an alteration in MAPK activities could contribute to the age-related decline in proliferative capacity.

Peripheral blood lymphocytes (PBL) of healthy elderly people show increased plasma membrane viscosity compared to young subjects, that inversely correlates with lymphocyte proliferation after mitogen stimulation in vitro. Maintenance of a constant membrane viscosity, which is necessary for proper cell function, is crucially dependent on the membrane lipid composition. The cellular lipid metabolism, and thus lymphocyte function, may be subject to modulation by diet or drugs. To study the susceptibility of membrane viscosity to environmental conditions, we established an in vivo model using severe combined immunodeficiency (SCID) mice: human peripheral blood lymphocytes from healthy young and old subjects were engrafted for three days intraperitoneally into SCID mice to offer identical environmental conditions. First, we demonstrate that human lymphocytes can take up and utilize murine lipoproteins: engrafted human PBL can participate in the mouse lipid metabolism, and an exchange of membrane lipids in vivo is, therefore, possible. Second, plasma membrane viscosity was determined before and after engraftment: before engraftment, PBL from the elderly showed a significantly higher membrane viscosity than that from young controls, but this difference vanished during engraftment into SCID mice, wherein cells from both age groups exhibited nearly identical values. It was, therefore, concluded that lymphocyte membrane viscosity is influenced by environmental factors, and that the age-related increase is, in principle, reversible.

The function of the immune system is known to decline in the elderly. The present communication will show that, similar to the situation in children, it is necessary to distinguish between primary and secondary alterations of the immune reactivity in the elderly. The primary immunological change in the elderly is the age-dependent intrinsic decline of immune responsiveness that is also observed in healthy persons, i.e., those selected according to the criteria of the SENIEUR Protocol of EURAGE (European Community Concerted Action on Aging). Secondary immunological changes are due to underlying diseases and various environmental factors, including diet, drug intake, physical activity, etc. While primary immunodeficiencies of the elderly are not, or only to a very minor extent, presently influenced by therapeutic measures, secondary alterations of the immune function offer further possibilities for corrective measures. Clinically, the consequences of impaired immune function in the elderly include increased susceptibility to infectious disease, the emergence of tumors, and increased autoimmune reactions, the latter albeit often without concomitant autoimmune disease. In fact, autoimmune diseases in generally begin to develop at younger ages, but their consequences are major factors affecting the quality of life of the elderly. Investigations of basic molecular and cellular aspects of the aging immune system are, therefore, of theoretical, clinical and socioeconomic interest.

Cholera toxin (CT) and Escherichia coli heat-labile enterotoxin (LT) are the strongest mucosal immunogens identified to date and are also good adjuvants when given orally together in combination with unrelated antigens. We used these potent immunogens to monitor local and systemic immune responses following oral immunization of BALB/c mice, and compared their action on the following: (a) immunoglobulin production rates (IgG, IgM and IgA) in mucosal inductive (Peyer's patches-PPs), effector (intestinal lamina propria-LP, respiratory tract) and systemic (spleen) sites; (b) analysis of systemic antigen-specific antibodies (IgG subclasses, IgA and IgE); (c) time monitoring of fecal anti-CT and anti-LT antibodies, and (d) in vivo relevance of interleukin-6 (IL-6) to mucosal responses. Both mucosal immunogens elicited specific antibody responses (IgA, IgG) not only in the gastrointestinal tract (PP's and intestinal LP), but also in the respiratory tract and spleens of orally immunized mice. These mucosal responses were accompained by elevated secretion of IL-6 in all investigated tissues, indicating involvement of this cytokine in B-cell maturation processes. Furthermore, oral immunization with CT and LT induced elevated serum titers of IgG1 followed by IgG2a, IgG2b, IgG3 and IgA, while high antigen-specific IgA and IgG1 responses were found in fecal extracts. These findings illustrate the action of orally administered CT and LT, respectively, on several humoral and cellular immune responses not only at the gastrointestinal tract, the application site, but also in distant mucosal effector sites such as the respiratory tract. These data suggest the potential use of these mucosal adjuvants in oral immunization strategies to improve the local immune response in remote mucosal tissues, in accordance with the concept of a common mucosa-associated immune system.

Thymic nurse cells are multicellular complexes located in the subcortical area of the thymus of all avian, mammalian and amphibian species investigated so far. Since their first description in 1980 many studies have been carried out to characterize their morphological and functional properties. The purpose of this review is to summarize recent morphological as well a functional analyses of chicken thymic nurse cells which suggest a role of these cell complexes in T cell selection.

Monoclonal antibody INN-CH 16 recognizes a surface determinant exclusively present on activated chicken T lymphocytes. The present experiments suggest that this chicken activated T lymphocyte antigen (CATLA) represents the chicken IL-2 receptor or an associated structure, as (i) the kinetics of CATLA expression during T cell activation are analogous to those described for the mammalian receptor for IL-2, (ii) the IL-2 dependent proliferation of mitogen prestimulated chicken lymphocytes is competitively inhibited by INN-CH 16, and (iii) pretreatment of T lymphoblasts with INN-CH 16 drastically reduces their capacity to absorb IL-2 activity from supernatants of mitogen activated chicken lymphocytes.

Thymic nurse cells (TNC) are multicellular complexes composed of morphologically intact thymocytes internalized within epithelial cells. Due to their unusual features, they have been suspected to provide an appropriate microenvironment for T-cell repertoire acquisition. In recent functional studies in the chicken system, we have shown that intra-thymic nurse cell-lymphocytes (TNC-L) are highly efficient inducers of a graft-versus-host reaction (GvHR) in an allogeneic chorionallantoic membrane (CAM) assay, manifested by the formation of visible colonies, so-called pocks. Immunohistochemical and electron microscopical assessment of pocks revealed no differences between the cellular components and ultrastructural features of GvHR induced by TNC-L or peripheral blood lymphocytes (PBL). In addition, the stimulation of this local GvHR by interleukin 2 (IL-2) was demonstrated.

In contrast to systemic autoimmunity, spontaneous autoimmune thyroiditis of Obese strain (OS) chickens is associated with a marked T cell hyperreactivity in vitro, i.e. an increased proliferation and interleukin 2 (IL 2) secretion in response to Concanavalin A (ConA). In the present study we report an enhanced capacity of OS peripheral lymphoid cells (splenocytes and peripheral blood lymphocytes, PBL) to adsorb fluorescein isothiocyante (FITC) labelled ConA, but not phytohemagglutinin (PHA). However, the elevated ConA binding cannot be a prerequisite for in vitro ConA hyperreactivity as OS thymocytes are normal with respect to ConA binding but nonetheless exhibit elevated responses to this mitogen. Moreover, ConA binding does not correlate with the frequency of cells able to express IL 2 receptors upon short term ConA stimulation. The percentage of ConA activatable cells was found to be increased in OS- PBL as compared to normal control PBL, but was unaltered in OS splenocytes. This finding points to a further mechanism of T cell hyperreactivity in OS chicks in addition to the previously reported defects in nonspecific immunosuppression. Finally, enumeration of cells in the S phase revealed that enhanced proliferation of OS T lymphocytes was not restricted to the in vitro response to ConA and phytohemagglutinin (PHA) but also occurs in vivo.

Specific antisera reacting with B-L (Ia-like) antigens were prepared by reciprocal immunization of animals from the congeneic lines CB and CC. The resulting antisera were tested either in direct or indirect immunofluorescence tests and stained 10-16% of peripheral blood cells (PBL). Of the B-L+ cells, 90% were B cells and 8% were T cells. After in vitro stimulation of PBL with ConA, 58% were B-L+ and 91% of these were T cells. B and T cells were defined by means of rabbit antisera raised against bursa and thymus cells made specific by absorption with the relevant tissues. Antigens determined by anti-B-L antisera, rabbit anti-bursa (ABS) and rabbit anti-thymus (ATS) sera showed an independent distribution on the membrane of PBL. The tissue distribution of B-L+ cells, defined by means of allo-antisera and monoclonal antibodies, was studied by direct and indirect immunofluorescence on sections of skin, liver, kidney and brain. In all organs, in addition to B cells and a small number of, presumably activated, T cells, macrophages and dendritic cells were positive. Notably, glia cells in the brain were also shown to express B-L antigen.

The present study provides first evidence for the presence of a thymic cell growth factor (TCGF) in supernatants (20 hrs) of mitogen (Con A) stimulated chicken spleen cells. 2 out of 3 batches of supernatant prepared according to a procedure originally described for the mouse system (10) induced a vigorous proliferative response of Con A prestimulated chicken spleen cells without significant mitogenic effect on unstimulated lymphocytes. No crossreactivity of this chicken-TCGF was observed with prestimulated murine lymphocytes, nor did potent mouse-TCGF preparations exhibit any proliferative effect on chicken cells. The implications of these data for both phylogenetic as well as practical experimental aspects are discussed.

Different procedures for enrichment of peripheral blood leukocytes (PBL) from whole chicken blood have been compared in terms of the percentage yield of lymphocytes and thrombocytes. The yield of thrombocytes was low in the buffy coat cells of heparinised blood, but high in PBL enriched from heparinised blood over a Ficoll-Paque density gradient and in PBL prepared from citrated blood by either technique. The fluorescence activated cell sorter (FACS) has been used to separate thrombocytes from lymphocytes in PBL by sorting of the negative cells after staining with specific anti-T and anti-B cell sera (or anti-immunoglobulin). The sorted cells were more than 99% viable, more than 98% thrombocytes by antigenic and morphological criteria and did not respond in vitro to T and B cell mitogens. Absorption studies on a turkey anti-bursa cell serum (ABST) which is cross-reactive for B cells and thrombocytes (8) indicated that the cross-reactivity is attributable to two (or more) antibody specificities in the serum rather than to one antibody directed at shared determinants on the two cell types.