Lung inflammatory environments differentially alter mesenchymal stromal cell behavior.

Mesenchymal stromal (stem) cells (MSCs) are increasingly demonstrated to ameliorate experimentally induced lung injuries through disease-specific anti-inflammatory actions, thus suggesting that different in vivo inflammatory environments can influence MSC actions. To determine the effects of different representative inflammatory lung conditions, human bone marrow-derived MSCs (hMSCs) were exposed to in vitro culture conditions from bronchoalveolar lavage fluid (BALF) samples obtained from patients with either the acute respiratory distress syndrome (ARDS) or with other lung diseases including acute respiratory exacerbations of cystic fibrosis (CF) (non-ARDS). hMSCs were subsequently assessed for time- and BALF concentration-dependent effects on mRNA expression of selected pro- and anti-inflammatory mediators, and for overall patterns of gene and mRNA expression.

Comparative Decellularization and Recellularization of Wild-Type and Alpha 1,3 Galactosyltransferase Knockout Pig Lungs: A Model for Ex Vivo Xenogeneic Lung Bioengineering and Transplantation.

Background: A novel potential approach for lung transplantation could be to utilize xenogeneic decellularized pig lung scaffolds that are recellularized with human lung cells. However, pig tissues express several immunogenic proteins, notably galactosylated cell surface glycoproteins resulting from alpha 1,3 galactosyltransferase (α-gal) activity, that could conceivably prevent effective use. Use of lungs from α-gal knock out (α-gal KO) pigs presents a potential alternative and thus comparative de- and recellularization of wild-type and α-gal KO pig lungs was assessed.

CD11b+ and Sca-1+ Cells Exert the Main Beneficial Effects of Systemically Administered Bone Marrow-Derived Mononuclear Cells in a Murine Model of Mixed Th2/Th17 Allergic Airway Inflammation.

Unlabelled: Systemic administration of bone marrow-derived mononuclear cells (BMDMCs) or bone marrow-derived mesenchymal stromal cells (MSCs) reduces inflammation and airway hyperresponsiveness (AHR) in a murine model of Th2-mediated eosinophilic allergic airway inflammation. However, since BMDMCs are a heterogeneous population that includes MSCs, it is unclear whether the MSCs alone are responsible for the BMDMC effects. To determine which BMDMC population(s) is responsible for ameliorating AHR and lung inflammation in a model of mixed Th2-eosinophilic and Th17-neutrophilic allergic airway inflammation, reminiscent of severe clinical asthma, BMDMCs obtained from normal C57Bl/6 mice were serially depleted of CD45, CD34, CD11b, CD3, CD19, CD31, or Sca-1 positive cells.

Residual Detergent Detection Method for Nondestructive Cytocompatibility Evaluation of Decellularized Whole Lung Scaffolds.

The development of reliable tissue engineering methods using decellularized cadaveric or donor lungs could potentially provide a new source of lung tissue. The vast majority of current lung decellularization protocols are detergent based and incompletely removed residual detergents may have a deleterious impact on subsequent scaffold recellularization. Detergent removal and quality control measures that rigorously and reliably confirm removal, ideally utilizing nondestructive methods, are thus critical for generating optimal acellular scaffolds suitable for potential clinical translation.

Systemic Administration of Human Bone Marrow-Derived Mesenchymal Stromal Cell Extracellular Vesicles Ameliorates Aspergillus Hyphal Extract-Induced Allergic Airway Inflammation in Immunocompetent Mice.

Unlabelled: An increasing number of studies demonstrate that administration of either conditioned media (CM) or extracellular vesicles (EVs) released by mesenchymal stromal cells (MSCs) derived from bone marrow and other sources are as effective as the MSCs themselves in mitigating inflammation and injury. The goal of the current study was to determine whether xenogeneic administration of CM or EVs from human bone marrow-derived MSCs would be effective in a model of mixed Th2/Th17, neutrophilic-mediated allergic airway inflammation, reflective of severe refractory asthma, induced by repeated mucosal exposure to Aspergillus hyphal extract (AHE) in immunocompetent C57Bl/6 mice. Systemic administration of both CM and EVs isolated from human and murine MSCs, but not human lung fibroblasts, at the onset of antigen challenge in previously sensitized mice significantly ameliorated the AHE-provoked increases in airway hyperreactivity (AHR), lung inflammation, and the antigen-specific CD4 T-cell Th2 and Th17 phenotype.

Recellularization of decellularized lung scaffolds is enhanced by dynamic suspension culture.

Strategies are needed to improve repopulation of decellularized lung scaffolds with stromal and functional epithelial cells. We demonstrate that decellularized mouse lungs recellularized in a dynamic low fluid shear suspension bioreactor, termed the rotating wall vessel (RWV), contained more cells with decreased apoptosis, increased proliferation and enhanced levels of total RNA compared to static recellularization conditions. These results were observed with two relevant mouse cell types: bone marrow-derived mesenchymal stromal (stem) cells (MSCs) and alveolar type II cells (C10).

Freshly thawed and continuously cultured human bone marrow-derived mesenchymal stromal cells comparably ameliorate allergic airways inflammation in immunocompetent mice.

Unlabelled: Recent data suggest that freshly thawed previously frozen mesenchymal stromal cells (MSCs) may not have the same effectiveness or breadth of anti-inflammatory activities as do continuously cultured MSCs. This has significant implications for clinical use, in which many infusion schemes use frozen cells thawed at the bedside for administration. The available data, however, predominantly evaluate in vitro MSC properties, and so far there has been limited in vivo analysis.

Design and Synthesis of an Artificial Pulmonary Pleura for High Throughput Studies in Acellular Human Lungs.

Whole organ decellularization of complex organs, such as lungs, presents a unique opportunity for use of acellular scaffolds for tissue engineering or for studying cell-extracellular matrix interactions . A growing body of literature investigating decellularizing and recellularizing rodent lungs has provided important proof of concept models and rodent lungs are readily available for high throughput studies. In contrast, comparable progress in large animal and human lungs has been impeded owing to more limited availability and difficulties in handling larger tissue.

Multipotent adult progenitor cells decrease cold ischemic injury in ex vivo perfused human lungs: an initial pilot and feasibility study.

Background: Primary graft dysfunction (PGD) is a significant cause of early morbidity and mortality following lung transplantation. Improved organ preservation techniques will decrease ischemia-reperfusion injury (IRI) contributing to PGD. Adult bone marrow-derived adherent stem cells, including mesenchymal stromal (stem) cells (MSCs) and multipotent adult progenitor cells (MAPCs), have potent anti-inflammatory actions, and we thus postulated that intratracheal MAPC administration during donor lung processing would decrease IRI.

Comparative decellularization and recellularization of normal versus emphysematous human lungs.

Acellular whole human lung scaffolds represent a unique opportunity for ex vivo tissue engineering. However, it remains unclear whether lungs from individuals with chronic lung diseases such as chronic obstructive pulmonary disease (COPD) can be appropriately decellularized and recellularized. To assess this, cadaveric human lungs from normal (non-smoking) patients and from patients with COPD (smoking history) were decellularized and found by histochemical and immunohistochemical staining, electron microscopy, and mass spectrometry to retain characteristic histological architecture and extracellular matrix components (ECM) reflecting either normal or COPD, particularly emphysematous, origin.

Mesenchymal stromal cells mediate Aspergillus hyphal extract-induced allergic airway inflammation by inhibition of the Th17 signaling pathway.

Systemic administration of mesenchymal stromal cells (MSCs) suppresses airway inflammation and methacholine-induced airway hyper-responsiveness (AHR) in mouse models of T helper cell (Th) type 2-mediated eosinophilic allergic airway inflammation (AAI); however, the efficacy of MSCs in mouse models of severe Th17-mediated neutrophilic AAI has not yet been demonstrated. We assessed MSC effects in a mouse model of mixed Th2/Th17 AAI produced by mucosal exposure to Aspergillus fumigatus hyphal extract (AHE). Following sensitization produced by oropharyngeal AHE administration, systemic (tail vein) administration of syngeneic MSCs on the first day of challenge significantly reduced acute AHR predominantly through reduction of Th17-mediated airway inflammation.

Three-dimensional scaffolds of acellular human and porcine lungs for high throughput studies of lung disease and regeneration.

Acellular scaffolds from complex whole organs such as lung are being increasingly studied for ex vivo organ generation and for in vitro studies of cell-extracellular matrix interactions. We have established effective methods for efficient de and recellularization of large animal and human lungs including techniques which allow multiple small segments (∼ 1-3 cm(3)) to be excised that retain 3-dimensional lung structure. Coupled with the use of a synthetic pleural coating, cells can be selectively physiologically inoculated via preserved vascular and airway conduits.

The effect of age and emphysematous and fibrotic injury on the re-cellularization of de-cellularized lungs.

Use of de-cellularized cadaveric lungs as 3-dimensional scaffolds for ex vivo lung tissue generation offers a new potential therapeutic approach for clinical lung transplantation. However, it is likely that some of the available cadaveric human lungs may be from older donors or from donors with previously existing structural lung diseases such as emphysema or pulmonary fibrosis. It is not known whether these lungs will be suitable for either de-cellularization or re-cellularization.

The effects of storage and sterilization on de-cellularized and re-cellularized whole lung.

Despite growing interest on the potential use of de-cellularized whole lungs as 3-dimensional scaffolds for ex vivo lung tissue generation, optimal processing including sterilization and storage conditions, are not well defined. Further, it is unclear whether lungs need to be obtained immediately or may be usable even if harvested several days post-mortem, a situation mimicking potential procurement of human lungs from autopsy. We therefore assessed effects of delayed necropsy, prolonged storage (3 and 6 months), and of two commonly utilized sterilization approaches: irradiation or final rinse with peracetic acid, on architecture and extracellular matrix (ECM) protein characteristics of de-cellularized mouse lungs.

A nonhuman primate model of lung regeneration: detergent-mediated decellularization and initial in vitro recellularization with mesenchymal stem cells.

Currently, patients with end-stage lung disease are limited to lung transplantation as their only treatment option. Unfortunately, the lungs available for transplantation are few. Moreover, transplant recipients require life-long immune suppression to tolerate the transplanted lung.

Comparative assessment of detergent-based protocols for mouse lung de-cellularization and re-cellularization.

Several different detergent-based methods are currently being explored for de-cellularizing whole lungs for subsequent use as three-dimensional scaffolds for ex vivo lung tissue generation. However, it is not yet clear which of these methods may provide a scaffold that best supports re-cellularization and generation of functional lung tissue. Notably, the detergents used for de-cellularization activate matrix metalloproteinases that can potentially degrade extracellular matrix (ECM) proteins important for subsequent binding and growth of cells inoculated into the de-cellularized scaffolds.

Initial binding and recellularization of decellularized mouse lung scaffolds with bone marrow-derived mesenchymal stromal cells.

Recellularization of whole decellularized lung scaffolds provides a novel approach for generating functional lung tissue ex vivo for subsequent clinical transplantation. To explore the potential utility of stem and progenitor cells in this model, we investigated recellularization of decellularized whole mouse lungs after intratracheal inoculation of bone marrow-derived mesenchymal stromal cells (MSCs). The decellularized lungs maintained structural features of native lungs, including intact vasculature, ability to undergo ventilation, and an extracellular matrix (ECM) scaffold consisting primarily of collagens I and IV, laminin, and fibronectin.

Slam haplotypes modulate the response to lipopolysaccharide in vivo through control of NKT cell number and function.

CD1d-restricted NKT cells make up an innate-like T cell subset that plays a role in amplifying the response of innate immune leukocytes to TLR ligands. The Slam locus contains genes that have been implicated in innate and adaptive immune responses. In this study, we demonstrate that divergent Slam locus haplotypes modulate the response of macrophages to the TLR4 ligand LPS through their control of NKT cell number and function.