hLMSC Secretome Affects Macrophage Activity Differentially Depending on Lung-Mimetic Environments

Mesenchymal stromal cell (MSC)-based therapies for inflammatory diseases rely mainly on the paracrine ability to modulate the activity of macrophages. Despite recent advances, there is scarce information regarding changes of the secretome content attributed to physiomimetic cultures and, especially, how secretome content influence on macrophage activity for therapy. hLMSCs from human donors were cultured on devices developed in house that enabled lung-mimetic strain. hLMSC secretome was analyzed for typical cytokines, chemokines and growth factors. RNA was analyzed for the gene expression of CTGF and CYR61. Human monocytes were differentiated to macrophages and assessed for their phagocytic capacity and for M1/M2 subtypes by the analysis of typical cell surface markers in the presence of hLMSC secretome. CTGF and CYR61 displayed a marked reduction when cultured in lung-derived hydrogels (L-Hydrogels). The secretome showed that lung-derived scaffolds had a distinct secretion while there was a large overlap between L-Hydrogel and the conventionally (2D) cultured samples. Additionally, secretome from L-Scaffold showed an HGF increase, while IL-6 and TNF-α decreased in lung-mimetic environments. Similarly, phagocytosis decreased in a lung-mimetic environment. L-Scaffold showed a decrease of M1 population while stretch upregulated M2b subpopulations. In summary, mechanical features of the lung ECM and stretch orchestrate anti-inflammatory and immunosuppressive outcomes of hLMSCs

Versican in inflammation and tissue remodelling: the impact on lung disorders.

Versican is a proteoglycan that has many different roles in tissue homeostasis and inflammation. The biochemical structure is comprised of four different types of the core protein with attached glycosaminoglycans that can be sulphated to various extents and has the capacity to regulate differentiation of different cell types, migration, cell adhesion, proliferation, tissue stabilization and inflammation. Versican's regulatory properties are of importance during both homeostasis and changes that lead to disease progression. The glycosaminoglycans that are attached to the core protein are of the chondroitin sulfate/dermatan sulfate type and are known to be important in inflammation through interactions with cytokines and growth factors. For a more complex understanding of versican it is of importance to study the tissue niche, where the wound healing process in both healthy and diseased conditions take place. In previous studies our group has identified changes in the amount of the multifaceted versican in chronic lung disorders such as asthma, chronic obstructive pulmonary disease and bronchiolitis obliterans syndrome, which could be a result of pathologic, transforming growth factor β driven, on-going remodelling processes. Reversely, the context of versican in its niche is of great importance since versican has been reported to have a beneficial role in other contexts e.g. emphysema. Here we explore the vast mechanisms of versican in healthy lung and in lung disorders

Enhanced ROCK1 dependent contractility in fibroblast from chronic obstructive pulmonary disease patients

Background: During wound healing processes fibroblasts account for wound closure by adopting a contractile phenotype. One disease manifestation of COPD is emphysema which is characterized by destruction of alveolar walls and our hypothesis is that fibroblasts in the COPD lungs differentiate into a more contractile phenotype as a response to the deteriorating environment. Methods: Bronchial (central) and parenchymal (distal) fibroblasts were isolated from lung explants from COPD patients (n = 9) (GOLD stage IV) and from biopsies from control subjects and from donor lungs (n = 12). Tissue-derived fibroblasts were assessed for expression of proteins involved in fibroblast contraction by western blotting whereas contraction capacity was measured in three-dimensional collagen gels. Results: The basal expression of rho-associated coiled-coil protein kinase 1 (ROCK1) was increased in both centrally and distally derived fibroblasts from COPD patients compared to fibroblasts from control subjects (p < 0.001) and (p < 0.01), respectively. Distally derived fibroblasts from COPD patients had increased contractile capacity compared to control fibroblasts (p < 0.01). The contraction was dependent on ROCK1 activity as the ROCK inhibitor Y27632 dose-dependently blocked contraction in fibroblasts from COPD patients. ROCK1-positive fibroblasts were also identified by immunohistochemistry in the alveolar parenchyma in lung tissue sections from COPD patients. Conclusions: Distally derived fibroblasts from COPD patients have an enhanced contractile phenotype that is dependent on ROCK1 activity. This feature may be of importance for the elastic dynamics of small airways and the parenchyma in late stages of COPD

Mesenchymal stromal cells in lung tissue

Mesenchymal stromal cells (MSC) are multipotent cells with immunomodulatory and regenerative properties. During recent years, the interest in using MSC for clinical approaches for various diseases have increased. The lung field is no exception, and several clinical trials using MSC as a cell-based therapy for severe lung diseases such as chronic obstructive pulmonary disease, idiopathic pulmonary fibrosis, and severe emphysema have been performed. Another severe lung disease where MSC therapy might be an alternative is bronchiolitis obliterans syndrome (BOS), a chronic type of rejection affecting approxamately 50% of lung transplanted patients within five years after transplantation. The exact pathology of BOS is at present not known, but inflammation is thought to be an important driving factor. Despite an increased interest in using MSC as a therapy, the in vivo biological function of endogenous MSC is not completely known. Furthermore, the cellular identity of primary MSC in human lungs has not been reported. The aim of this thesis work has been to provide new insights of MSC in lung tissue, especially after lung transplantation, with respect to origin, tissue-specificity, and extracellular matrix production. Primary and culture-derived MSC were isolated from lung biopsies obtained from lung transplanted patients, fetal lung tissue, and bone marrow aspirates and evaluated using a comprehensive panel of in vitro and in vivo assays. The studies show that lung-derived MSC are tissue-resident cells with tissue-specific properties. Lung-derived MSC have different gene expression- and cytokine patterns, higher proliferation rate, higher colony-forming capacity, and smaller size compared with bone marrow-derived MSC. Interestigly, lung-derived MSC do not have in vivo bone formation capacity. Furthermore, we demonstrate for the first time that primary lung-derived MSC are enriched in the CD90+/CD105+ mononuclear cell fraction, which have a perivascular location in situ. Additionally, we demonstrate that MSC are prominent extracellular matrix producers, and that bone marrow- and lung-derived MSC produce distinct extracellular matrix profiles. There were no significant differenses between patients with chronic and acute rejections compared with good outcome recipients regarding EDA-fibronectin expression. However, increased EDA-fibronectin expression was found in acute rejections grade A2-A3 and in biopsies from patients with infection. Finally, these studies show that tissue-resident MSC do not have an altered phenotype after BOS development and that the number of colony-forming cells i.e. MSC do not correlate with the onset of BOS. In summary, this thesis work provide novel insights of tissue-resident MSC within the lung, an important step in identifying the functional role of MSC in normal lung physiology and during disease. Hopefully in the future, this knowledge will improve clinical strategies to treat severe lung diseases such as BOS

MSCs interaction with the host lung microenvironment : An overlooked aspect?

Mesenchymal stromal cells (MSCs) were identified more than 50 years ago, and research advances have promoted the translation of pre-clinical studies into clinical settings in several diseases. However, we are only starting to uncover the local factors that regulate cell phenotype, cell function, and cell viability across tissues following administration in different diseases. Advances in pre-clinical and translational studies suggest that the host environment, especially inflammatory active environments, plays a significant role in directing the infused MSCs towards different phenotypes with different functions. This can significantly effect their therapeutic efficacy. One way to study this interaction between the host environment and the infused cells is to expose MSCs ex vivo to patient samples such as serum or bronchoalveolar lavage fluid. Using this approach, it has been demonstrated that MSCs are very sensitive to different host factors such as pathogens, inflammatory cytokines, and extra cellular matrix properties. By understanding how different local host factors effect MSC function it will open possibilities to select specific patient sub-groups that are more likely to respond to this type of treatment and will also open possibilities to prime the local host environment to increase viability and to enrich for a specific MSC phenotype. Here, we aim to review the current understanding of the interaction of MSCs with the host microenvironment. To narrow the scope of this mini review, the focus will be on the pulmonary microenvironment, with a specific focus on the diseases acute respiratory distress syndrome (ARDS) and cystic fibrosis (CF)

Challenges and opportunities for regenerating lung tissue using three-dimensional lung models

Lung transplantation is currently the only option for patients with end-stage respiratory diseases, but owing to various complications and adverse effects of the treatment in combination with the increasing demand for and limited access to donor organs, new strategies are needed. Recent advances in materials science, culture techniques, cell-phenotyping and isolation techniques, bioreactor engineering, and imaging techniques have opened up new possibilities for generating advanced three-dimensional lung models. The lung is a complex organ to rebuild, consisting of more than 40 different cell types working together to create a functional unit that allows for proper gas exchange and protection against the external environment. However, it is well known that the cells do not work alone. It was long thought that the only function of the extracellular matrix (ECM) was to provide the organ and cells with structural support, but we now know that the ECM plays a much more vital role. Despite this knowledge, the majority of studies are still performed by using traditional two-dimensional (2D) cultures on plastic. There are, of course, occasions when 2D cultures are necessary; however, it is important to remember that essential features, including orientation, mechanical properties, and cell-cell and cell-matrix interactions, are missing. These are important properties that will most likely affect the final results and the possibility of translating in vitro findings into in vivo models

3D Lung Models for Regenerating Lung Tissue

3D Lung Models for Regenerating Lung Tissue is a comprehensive summary on the current state of art 3D lung models and novel techniques that can be used to regenerate lung tissue. Written by experts in the field, readers can expect to learn more about 3D lung models, novel techniques including bioprinting and advanced imaging techniques, as well as important knowledge about the complexity of the lung and its extracellular matrix composition.Structured into 15 different chapters, the book spans from the original 2D cell culture model on plastic, to advanced 3D lung models such as using human extracellular matrix protein. In addition, the last chapters cover new techniques including 3D printing, bioprinting, and artificial intelligence that can be used to drive the field forward and some future perspectives. This highly topical book with chapters on everything from the complexity of the lung and its microenvironment to cutting-edge 3D lung models, represents an exciting body of work that can be used by researchers during study design, grant writing, as teaching material, or to stay updated with the progression of the field.Key Features A comprehensive summary of advanced 3D lung models written by the experts in the respiratory field Explore novel techniques that can be used to evaluate and improve 3D lung models, including techniques such as 3D printing, bioprinting, and artificial intelligence Explains what extracellular matrix is, the complexity of the lung microenvironment, and why this knowledge is important for creating a functional bioartificial lun

Bioartificial lungs based on de-and recellularisation approaches : A historical perspective

For patients with end-stage respiratory diseases, such as COPD, interstitial lung diseases and cystic fibrosis, lung transplantation remains the only treatable option. However, due to increasing demand and limited availability of donor lungs, risk of complications such as acute and chronic rejection, and adverse effects of immunosuppressive treatments, this is not an alternative for the majority of this patient group [1, 2]. To meet the rising clinical demand new strategies to increase the number of available lungs for transplantation are needed [2]. One such strategy involves creating a functional lung ex vivo using different de- and recellularisation approaches. In this article, we will provide an overview of three landmark studies on bioartificial lungs published during 2010 that set the base for the direction of this relatively young field