Mesenchymal stem cells (MSCs) are among the best investigated stem cell populations (3). BM-derived MSCs are the most frequently studied MSCs. The international society for cell therapies has set minimum requirements for the definition of MSC: positive for surface markers CD73, CD90, and CD105 and negative for CD14, CD34, and CD45 (4). In addition, MSCs also need to be able to generate adipocytes, osteoblasts, and chondroblasts in lineage differentiation assays, in vitro (4). This criterion shows the multipotency of these cells, meaning that MSCs are able to give rise to several cell types within the mesenchymal lineage. MSCs have a typical fibroblast-like morphology when cultured in vitro and grow in colony-forming units-fibroblast-like (CFU-F) when seeded in limiting dilution (5). Because of their lack of human leukocyte antigen (HLA) expression, the immune system is largely blind toward MSC, leading to what has been called the “immune-privileged status” of MSC (6). This immune-privileged status has made the MSC an ideal candidate cell for transplantation approaches, as HLA incompatibility is not an issue with MSC. BM-MSCs have shown great promise for cell therapy in various animal models of lung disease and undergone testing for safety and efficacy in several patient groups of lung diseases as will be outlined in the following paragraphs.

In addition to BM-MSCs, lung resident MSCs or mesenchymal stromal progenitors have raised attention due to their potential contribution to several disease processes (7–12). These cells are less clearly defined in their phenotype, although authors use many of the criteria that define BM-MSC.

Fibrocytes have been discovered as a population of hematopoietic cells with ability to produce collagen and to differentiate to myofibroblasts (64). Although shown to accumulate in multiple organ systems during injury repair and fibrosis, fibrocytes have received particular attention in the field of chronic lung diseases, because of a possible contribution of fibrocytes to disease development and progression: such diseases include ILD of the adult, such as IPF, where high levels of circulating fibrocytes have been demonstrated to indicate a poor prognosis (48). In addition, histological evidence for fibrocytes is present in lung tissue from IPF patients (46). Homing of fibrocytes to fibrotic lung tissue depends upon CXC chemokine ligand 12 (CXCL12), a finding that provides an interesting option to block tissue accumulation of fibrocytes as a potential therapeutic avenue (65). Fibrocytes may also represent a target of therapy in PAH, as evidenced by reduced fibrocyte accumulation as a response to treprostinil therapy in an animal model (66).

Different cell entities have been summarized under the term “EPC”: EPC have been recently shown as resident cells in the lung circulation and initially as circulating cells (67–71). There are two main ways to identify circulating EPC: first, EPC can be detected within blood mononuclear cells by flow cytometry with specific surface markers (72). The most frequently used marker combination is CD34⁺ CD133⁺ VEGFR2⁺. The identity of these cells as “true” EPC has been controversial for quite a while (68, 73). Second, EPC can be detected by growth: depending on culture principles, early outgrowth CFU-Hills have been distinguished from late outgrowth endothelial colony-forming cells (ECFC) (71, 74, 75). Today, the consensus is that CFU-Hills are rather mononuclear cells with endothelial markers that exert protective effects on endothelial cells largely in a paracrine fashion (76), whereas ECFC are “true” EPC with the ability to replace damaged EC by engraftment (74). Different results have been obtained in preclinical studies for PAH, largely depending on the type of cell that the investigators used: Whereas endogenous EPC seem to be functionally impaired with higher than control proliferation rate but lacking ability for angiogenesis (72), therapy with genetically modified EPC has been successful for treatment in animal models of PAH (77). A clinical trial to determine whether genetically modified EPC have the same therapeutic benefit on PAH patients failed to provide the expected results (78).

In addition to circulating EPC, there is evidence for several EPC populations in the adult lung: Alphonse et al. have shown that ECFC can be isolated from the lung periphery and that these ECFC are functionally impaired in hyperoxia lung injury (79, 80). There is also evidence for EPC with high ability for self-renewal in the adult murine lung (81). These cells have been named “vascular endothelial stem cells” and represent EPC that are on the very top of the lung EC hierarchy. The finding of these endothelial stem cells in the blood vessels of the lung is consistent with previous reports of systemic blood vessels harboring a complete hierarchy of EPC (82). EPC at various levels of self-renewal and differentiation could indeed provide a basis for extensive vascular regeneration. On the other hand, because of the high ability to grow, some of these EPC or endothelial stem cells may also contribute to generate the apoptosis-resistant EC that are found in the complex vascular lesions of PAH patients (83). Evidence for mono- and polyclonal EC proliferation in complex lesions of pulmonary arteries in PAH indeed suggests a contribution of EPC or endothelial stem cells to the development of plexiform lesions in PAH (84). Much work needs to be done to characterize the EC/EPC/endothelial stem cell hierarchy in the lung and to identify the potential of EPC/EC stem cells for regenerative medicine or disease pathobiology.

Adult stem cells are mostly understood as multipotent cells, meaning that adult stem cells can give rise to multiple cell types within a given lineage (85). The classical example for a multipotent stem cell is the MSC with the ability to produce cells of multiple skeletal cell types, such as osteoblasts, chrondroblasts, adipocytes, and muscle cells (3, 86). In contrast, during embryonic development, there is an early population of stem cells that have the ability to make cell lineages from all three germinal layers (87). This ability is called pluripotency, and the cells are described as embryonic stem cells. No single marker or transcription factor can define pluripotency, but instead a panel of functional assays in combination with activity of several transcription factors, e.g., Oct-4, Nanog, Klf4, and others, can help to identify pluripotent cells (87–90). In recent years, a variety of strategies have been shown to use a mature cell, such as fibroblasts, and induce pluripotency in such a cell. The strategies range from simultaneous overexpression of several transcription factors to chemical stimulation methods (88–94). So why is it an advantage to generate such iPSC? First, pluripotent cells are very powerful cells that can give rise to almost any cell lineage in the body and, therefore, their value for regenerative medicine may be skyrocketing. But human embryonic stem cells, which are the “natural” pluripotent stem cells, are highly restricted and difficult to obtain. Hence, iPSC would provide a much easier accessible and less regulated source of pluripotent stem cells. Second, iPSC allow generating pluripotent stem cells from tissues such as skin of patients and, therefore, providing a patient-specific and genetically identical source of cells. Such approaches will likely be very useful in the dawning age of “personalized medicine.” Recent work in the vascular field has demonstrated that iPSC can give rise to different vascular lineages, including EC (95, 96). The ability to generate genetically identical EC from, e.g., skin fibroblasts will provide a unique tool for regenerative medicine, including seeding of decellularized lung matrices with genetically engineered but patient-specific lung cells as prerequisite for using these recellularized lungs as transplant organs (97). Despite all the enthusiasm and hopes that flare up with such exciting and groundbreaking progress, these so powerful iPSC also need to be careful evaluated for adverse effects when transplanted to patients and seeded in organ matrices to generate future transplant organs: because iPSC can give rise to so many different lineages and have a high self-renewal potential, iPSC harbor the danger of giving rise to produce genetic and epigenetic changes, immunogenicity, and tumor cells (98). Even the possibility of such adverse effect of iPSC therapy needs to be carefully considered, and the risks have to be carefully weighted against the clear and exciting potential to benefit the patient by using iPSC.

As science progresses with a better understanding of the role of the various stem or progenitor cell types in contribution to lung disease, repair, or regeneration, the mechanical microenvironment has emerged as a major player. Mechanical signal transduction through focal adhesions, the glycocalyx, mechanically activated ion channels, primary cilia, and force across cell–cell junctions may all play a role in the contribution of stem cells to pulmonary vascular remodeling (99). The dynamic mechanical environment of the lung lends itself to study the effects of mechanobiology in normal physiology and pathology of the lung microvasculature. The mechanical environment is essential to stem cell behavior during lung development and during pathological vascular remodeling events, which occur in BPD, IPF, COPD, and PAH.

During development of the pulmonary microvasculature, the fetal lung undergoes hydrostatic pressure between 1–2 mmHg, which maintains the lung in a distended state (100). Fetal breathing movements cause cyclic mechanical loading to the developing alveoli and microvasculature causing shear stress (101). Following the development of the primitive circulatory system, blood velocity and shear stress is critical for embryonic vascular patterning, remodeling, and proper development (102). Lack of proper mechanotransduction at any stage in the developmental process can lead to structural abnormalities and an underdeveloped lung.

Bronchopulmonary dysplasia is characterized by impaired alveolarization and vascularization and often occurs following mechanical ventilation. The impaired structure of the lung microvasculature leads to abnormal fluid flow and reduced gas exchange. High air flow from mechanical ventilation can increase lung injury, leading to the development of BPD (103).

Idiopathic pulmonary fibrosis is associated with progressive fibrotic scarring and stiffening of the lung ECM (104). This scarring is also associated with loss of compliance. The overall stiffness of the alveolar septa is increased by two- to threefold as measured by atomic force microscopy (105). IPF is also associated with increased vascular remodeling and may be associated with increased PH.

In PAH, pulmonary vascular remodeling is influenced by both hemodynamic and ECM changes. By its very nature, the remodeling in PAH causes increases in the pulmonary vasculature pressure as shown by animal models of severe PAH (106). But increased pulmonary artery pressure, e.g., by vasoconstriction, may also contribute to remodeling of pulmonary arteries and even foster the development of complex lesions in pulmonary arteries (107, 108). Furthermore, the ECM changes that occur in remodeling lend themselves to increased substrate stiffness (109, 110).

Chronic obstructive pulmonary disease also causes changes in lung mechanics and vascular remodeling. A reduction of vascularity in the alveolar septa occurs in emphysema (111). Loss of alveoli causes lung hyperinflation and overdistension of the remaining alveoli and capillaries. Furthermore, pulmonary arterial stiffness and blood pressure are increased in COPD patients and are not reduced with pulmonary rehabilitation (112).

These altered mechanical environments and their effects on stem cell contribution to pulmonary vascular remodeling warrants further study. The general physiological response of stem cells to mechanical signaling has been studied greatly in vitro, with in vivo studies largely lacking. Furthermore, specific studies of the mechanical environment on stem cells residing in the lung vasculature are especially absent. Information may be gleaned by understanding stem and progenitor cell response to general mechanical conditions. We will briefly review the state of the science of stem cell response to each of three mechanical conditions: shear stress, cyclic strain, and substrate stiffness as they may pertain to pulmonary vascular remodeling and stem cell-based therapies. These mechanical loading conditions and their effect on cell signaling and phenotype are summarized in Table 1.