Research

A. The role of macrophages in tissue remodeling
B. Emerging disease biomarkers
C. Tissue regeneration by progenitor- and stem cells

A. The role of macrophages in tissue remodeling

Macrophages comprise a major regulatory cell type in many inflammatory diseases and have been associated with the efficient repair of a tissue injury via phagocytosis and secretion of healing growth factors, cytokines and proteases, as well as with the uncontrolled deposition of extracellular matrix components, thus contributing to the pathophysiology of multiple fibrosis-dependent disease states. Their ability to modify their function (plasticity) depending on the tissue context combined with the differential expression and/or secretion pattern of certain cytokines and surface receptors appears to be of crucial importance for tissue injury and remodeling. Characteristically, during sequential phases of tissue inflammation and remodeling different macrophage subtypes are present. Classically activated macrophages (M1) generally promote phagocytosis and modify the inflammatory response, whereas alternatively activated macrophages (M2) regulate the restoration of extracellular structures during the progress to tissue remodeling and repair. Stromal cells, fibroblasts and myofibroblasts, serve as targets of the potent remodeling-promoting secretions of macrophages and of activated signaling pathways exemplifying the final effect, i.e. either a successive repair or an adverse remodeling response (fibrosis, extended angiogenesis, etc). Numerous question marks and black boxes in the above oversimplified scheme are further perplexed by the eventual presence of a modified immune background, exogenous insults (infections), etc. Despite some unique pathways and tissue-specific differences, similar molecules and mechanisms may characterize the pathophysiology of fibrosis and organ dysfunction in inflammatory diseases.

One important molecule that seems to regulate and mediate some of the macrophage’s actions is galectin-3. This lectin forms oligomers and recognizes N-acetyllactosamine sequences, thus binding to membrane receptor clusters. Both exogenous and cytoplasmic or nuclear galectin-3 exerts important functions that affect cellular proliferation and death, immune response and inflammation, as well as tissue fibrosis. However, its action may vary from beneficial to detrimental, depending on the disease background. Our recent study has shown that the levels of galectin-3 that are secreted by activated macrophages depend on the presence of osteopontin, a cytokine that is also important for tissue remodeling. In the same study, reduced expression and release of galectin-3 by macrophages infiltrating the myocardium in a genetic model of heart failure was associated with less fibrosis and improved cardiac function. Using primary macrophage cultures under different conditions of activation, the mechanism(s) underlying the dependence of galectin-3 expression and/or production on osteopontin and other cytokines is being investigated. Furthermore, using primary fibroblast cultures originated from different tissues, we investigate the eventual paracrine action of macrophages in fibroblast activation and profibrotic response, as well as the role of galectin-3, osteopontin, additional cytokines and miRNAs in these processes (Supported by the Hellenic Cardiological Society to S.P.)

In parallel, mice with galectin-3 deficiency have been crossed to mice with desmin deficiency to investigate in vivo the function of this molecule in a well characterized animal model of tissue remodeling, cardiac fibrosis and macrophage inflammation (Supported by the Hellenic Cardiological Society to S.P.). This study is being conducted in collaboration with Dr. Capetanaki, BRFAA, as well as Dr. C. Davos, BRFAA.

In addition, the role of galectin-3, and generally the role of macrophages in airway remodeling will be studied using alveolar and interstitial macrophages of the lung in mouse models of allergic and virus-induced asthma. This study is going to e conducted with Dr. E. Andreakos, BRFAA, as well as Assoc. Prof. N.G. Papadopoulos, NCUA (Supported by Thalis-MIDAS GGET programme to E.A.). In parallel, the study of paracrine action of bronchial epithelium in airway remodeling is being continued in relevant in vitro models of virus-induced asthma.

Furthermore, the contribution of macrophages in models of renal fibrosis, such as unilateral ureteral obstruction (UUO) is also important. In collaboration with Dr. A Charonis, BRFAA, and in continuation of previous studies, we investigate the presence of macrophages and other immunocytes in this model, in conjunction with the expression and action of other molecules (e.g. transgelin, calreticulin) by and on renal cells.

Finally, to study the role of macrophages in a model of cardiac fibrosis, experiments aiming to the characterization of macrophage subtypes, their selective depletion and analysis of their effects in mice deficient in desmin will be conducted, in collaboration with Dr. Y. Capetanaki, BRFAA (Supported by EPAN-GGET-Synergasia TREAT HEART to Y.C.).

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B. Emerging disease biomarkers

The emergence of novel biomarkers of particular pathologies forms an important strategic target for disease prognosis and treatment. Biomarkers related to inflammation and remodeling that will be measurable with non interventional techniques are of particular interest. Characteristically, circulating galectin-3 appears to be an effective biomarker for cardiomyopathies, including heart failure. On the other hand, in a recent study in which we participated the levels of osteopontin in plasma were correlated with the oxidative stress in patients with coronary disease. Given the association of those two molecules with macrophages and the osteopontin-dependent expression of galectin-3 in the desmin deficient mouse, we further investigate in this genetic heart failure model the eventual correlation of their circulating levels to the progress of heart remodeling and failure (Supported by Hellenic Cardiological Society to S.P.). This study is being conducted in collaboration with Dr. Y. Capetanaki, BRFAA.

In addition, recent studies show that regulatory miRNAs are secreted and can be identified in the circulation and may be useful biomarkers of several disease states. A series of inflammation- and fibrosis-related biomarkers, as well as secreted miRNAs will be studied in samples from heart failure patients, in collaboration with Prof. D. Cokkinos, BRFAA.

Moreover, using the UUO model in the rat and the mouse, a whole genome analysis has been planned in collaboration with Dr. A. Charonis, BRFAA, aiming to the identification of novel biomarkers of renal fibrosis and chronic kidney disease.

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C. Tissue regeneration by progenitor- and stem cells

The recent progress in the field of progenitor and stem cell biology recruited our interest and attention to therapeutic strategies that would take advantage of the abilities of such cells aiming to the regeneration of injured tissues. Thus, within the frames of our participation in the Synergasia-ANAGENNISI MYOKARDIOU project (supported by EPAN-GGET grant to Dr. Y. Capetanaki, BRFAA) the characteristics of cardiac progenitor cell populations will be studied in vivo, as well as after their isolation and culture. Emphasis will be given to the effect of an inflammatory milieu on the identities and characteristics of such cells, and as model we will use the environment prevailing within the degenerating myocardium of the cardiac fibrosis and failure des-/- model.

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