Cancer Conditioned Medium Modulates Functional and Phenotypic Properties of Human Decidua Parietalis Mesenchymal Stem/Stromal Cells.

Background: Mesenchymal Stem/Stromal Cells (MSCs) from the decidua parietalis (DPMSCs) of human term placenta express several molecules with important biological and immunological properties. DPMSCs induce natural killer cell expression of inflammatory receptors and their cytotoxic activity against cancer cells. These properties make DPMSCs promising therapeutical agent for cancer. The successful development of MSCs as an anti-cancer therapeutic cells rely on their ability to function in a hostile inflammatory and oxidative stress cancer environment. Here, we studied the effects of conditioned medium obtained from the culture of breast cancer cells (CMMDA-231) on the functional and phenotypic properties of DPMSCs. Methods: DPMSCs were cultured with CMMDA-231 and important functions of DPMSCs were measured. The effect of CMMDA-231 on DPMSC expression of several genes with different functions was also evaluated. Results: DPMSCs were able to function in response to CMMDA-231, but with reduced proliferative and adhesive potentials. Preconditioning of DPMSCs with CMMDA-231 enhanced their adhesion while reducing their invasion. In addition, CMMDA-231 modulated DPMSC expression of many genes with various functional (i.e., proliferation, adhesion, and invasion) properties. DPMSCs also showed increased expression of genes with anti-cancer property. Conclusion: These data show the ability of DPMSCs to survive and function in cancer environment. In addition, preconditioning of DPMSCs with CMMDA-231 enhanced their anti-cancer properties and thus demonstrating their potential as an anti-cancer therapeutic agent. However, future studies are essential to reveal the mechanism underlying the effects of MDA-231 on DPMSC functional activities and also to confirm the anti-cancer therapeutic potential of DPMSCs.

Preconditioning human natural killer cells with chorionic villous mesenchymal stem cells stimulates their expression of inflammatory and anti-tumor molecules.

BACKGROUND: Mesenchymal stem cells derived from the chorionic villi of human placentae (pMSCs) produce a unique array of mediators that regulate the essential cellular functions of their target cells. These properties make pMSCs attractive candidates for cell-based therapy. Here, we examined the effects of culturing human natural killer (NK) cells with pMSCs on NK cell functions. METHODS: pMSCs were cultured with IL-2-activated and non-activated NK cells. NK cell proliferation and cytolytic activities were monitored. NK cell expression of receptors mediating their cytolytic activity against pMSCs, and the mechanisms underlying this effect on pMSCs, were also investigated. RESULTS: Our findings show that IL-2-activated NK cells, but not freshly isolated NK cells, efficiently lyse pMSCs and that this response might involve the activating NK cell receptor CD69. Interestingly, although pMSCs expressed HLA class I molecules, they were nevertheless lysed by NK cells, suggesting that HLA class I antigens do not play a significant role in protecting pMSCs from NK cell cytolytic activity. Co-culturing NK cells with pMSCs also inhibited NK cell expression of receptors, including CD69, NKpG2D, CD94, and NKp30, although these co-cultured NK cells were not inhibited in lysing cancer cells in vitro. Importantly, co-cultured NK cells significantly increased their production of molecules with anti-tumor effects. CONCLUSIONS: These findings suggest that pMSCs might have potential applications in cancer therapy.

Human decidua basalis mesenchymal stem/stromal cells protect endothelial cell functions from oxidative stress induced by hydrogen peroxide and monocytes.

BACKGROUND: Human decidua basalis mesenchymal stem/multipotent stromal cells (DBMSCs) inhibit endothelial cell activation by inflammation induced by monocytes. This property makes them a promising candidate for cell-based therapy to treat inflammatory diseases, such as atherosclerosis. This study was performed to examine the ability of DBMSCs to protect endothelial cell functions from the damaging effects resulting from exposure to oxidatively stress environment induced by H2O2 and monocytes. METHODS: DBMSCs were co-cultured with endothelial cells isolated from human umbilical cord veins in the presence of H2O2 and monocytes, and various functions of endothelial cell were then determined. The effect of DBMSCs on monocyte adhesion to endothelial cells in the presence of H2O2 was also examined. In addition, the effect of DBMSCs on HUVEC gene expression under the influence of H2O2 was also determined. RESULTS: DBMSCs reversed the effect of H2O2 on endothelial cell functions. In addition, DBMSCs reduced monocyte adhesion to endothelial cells and also reduced the stimulatory effect of monocytes on endothelial cell proliferation in the presence of H2O2. Moreover, DBMSCs modified the expression of many genes mediating important endothelial cell functions. Finally, DBMSCs increased the activities of glutathione and thioredoxin reductases in H2O2-treated endothelial cells. CONCLUSIONS: We conclude that DBMSCs have potential for therapeutic application in inflammatory diseases, such as atherosclerosis by protecting endothelial cells from oxidative stress damage. However, more studies are needed to elucidate this further.

Human chorionic villous mesenchymal stem/stromal cells protect endothelial cells from injury induced by high level of glucose.

BACKGROUND: Mesenchymal stem/stromal cells derived from chorionic villi of human term placentae (pMSCs) protect human endothelial cells from injury induced by hydrogen peroxide (H2O2). In diabetes, elevated levels of glucose (hyperglycaemia) induce H2O2 production, which causes the endothelial dysfunction that underlies the enhanced immune responses and adverse complications associated with diabetes, which leads to thrombosis and atherosclerosis. In this study, we examined the ability of pMSCs to protect endothelial cell functions from the negative impact of high level of glucose. METHODS: pMSCs isolated from the chorionic villi of human term placentae were cultured with endothelial cells isolated from human umbilical cord veins in the presence of glucose. Endothelial cell functions were then determined. The effect of pMSCs on gene expression in glucose-treated endothelial cells was also determined. RESULTS: pMSCs reversed the effect of glucose on key endothelial cell functions including proliferation, migration, angiogenesis, and permeability. In addition, pMSCs altered the expression of many genes that mediate important endothelial cell functions including survival, apoptosis, adhesion, permeability, and angiogenesis. CONCLUSIONS: This is the first comprehensive study to provide evidence that pMSCs protect endothelial cells from glucose-induced damage. Therefore, pMSCs have potential therapeutic value as a stem cell-based therapy to repair glucose-induced vascular injury and prevent the adverse complications associated with diabetes and cardiovascular disease. However, further studies are necessary to reveal more detailed aspects of the mechanism of action of pMSCs on glucose-induced endothelial damage in vitro and in vivo.

Preconditioning by Hydrogen Peroxide Enhances Multiple Properties of Human Decidua Basalis Mesenchymal Stem/Multipotent Stromal Cells.

Stem cell-based therapies rely on stem cell ability to repair in an oxidative stress environment. Preconditioning of mesenchymal stem cells (MSCs) to a stress environment has beneficial effects on their ability to repair injured tissues. We previously reported that MSCs from the decidua basalis (DBMSCs) of human placenta have many important cellular functions that make them potentially useful for cell-based therapies. Here, we studied the effect of DBMSC preconditioning to a stress environment. DBMSCs were exposed to various concentrations of hydrogen peroxide (H2O2), and their functions were then assessed. DBMSC expression of immune molecules after preconditioning was also determined. DBMSC preconditioning with H2O2 enhanced their proliferation, colonogenicity, adhesion, and migration. In addition, DBMSCs regardless of H2O2 treatment displayed antiangiogenic activity. H2O2 preconditioning also increased DBMSC expression of genes that promote cellular functions and decreased the expression of genes, which have opposite effect on their functions. Preconditioning also reduced DBMSC expression of IL-1ß, but had no effects on the expression of other immune molecules that promote proliferation, adhesion, and migration. These data show that DBMSCs resist a toxic environment, which adds to their potential as a candidate stem cell type for treating various diseases in hostile environments.

Characterization of the interaction between human decidua parietalis mesenchymal stem/stromal cells and natural killer cells.

BACKGROUND: Human decidua parietalis mesenchymal stem/multipotent stromal cells (DPMSCs) have unique phenotypic and functional properties that make them promising candidates for cell-based therapy. Here, we investigated DPMSC interaction with natural killer (NK) cells, and the effects of this interaction on NK cell phenotypic characteristics and functional activities. METHODS: DPMSCs isolated from the decidua parietalis of human fetal membranes were cultured with interleukin (IL)-2-activated and IL-2-unactivated NK cells isolated from healthy human peripheral blood. NK cell proliferation and cytolytic activities were then examined using functional assays. NK cell expression of receptors mediating the cytolytic activity against DPMSCs, and the mechanism underlying this effect on DPMSCs, were also examined using flow cytometry and light microscopy, respectively. RESULTS: DPMSCs stimulated IL-2-induced proliferation of resting NK cells and the proliferation of activated NK cells. Moreover, IL-2-activated NK cells, but not freshly isolated NK cells, efficiently lysed DPMSCs. The induction of this NK cell cytolytic activity against DPMSCs was mediated by the activating NK cell receptors NKG2D, CD69, NKp30, and NKp44. However, DPMSCs showed a direct induction of NK cell cytolytic activity through CD69. We also found that DPMSCs expressed the ligands for these activating NK cell receptors including Nectin-2, ULBP-2, MICA, and MICB. Although DPMSCs expressed HLA class I molecules, they were susceptible to lysis by NK cells, suggesting that HLA class I antigens do not play a significant role in NK cell cytolytic action. In addition, DPMSCs did not inhibit NK cell cytolytic activity against cancer cells. Importantly, DPMSCs significantly increased NK expression of inflammatory molecules with anticancer activities. CONCLUSIONS: We conclude that DPMSCs have potential for therapeutic application in cancer therapy, but not in transplantation or immunological diseases.

Human chorionic villous mesenchymal stem/stromal cells modify the effects of oxidative stress on endothelial cell functions.

Mesenchymal stem/stromal cells derived from chorionic villi of human term placentae (pMSCs) produce a unique combination of molecules, which modulate important cellular functions of their target cells while concurrently suppressing their immune responses. These properties make MSCs advantageous candidates for cell-based therapy. Our first aim was to examine the effect of high levels of oxidative stress on pMSC functions. pMSCs were exposed to hydrogen peroxide (H2O2) and their ability to proliferate and adhere to an endothelial cell monolayer was determined. Oxidatively stressed pMSCs maintained their proliferation and adhesion potentials. The second aim was to measure the ability of pMSCs to prevent oxidative stress-related damage to endothelial cells. Endothelial cells were exposed to H2O2, then co-cultured with pMSCs, and the effect on endothelial cell adhesion, proliferation and migration was determined. pMSCs were able to reverse the damaging effects of oxidative stress on the proliferation and migration but not on the adhesion of endothelial cells. These data indicate that pMSCs are not only inherently resistant to oxidative stress, but also protect endothelial cell functions from oxidative stress-associated damage. Therefore, pMSCs could be used as a therapeutic tool in inflammatory diseases by reducing the effects of oxidative stress on endothelial cells.

Immunomodulatory properties of human placental mesenchymal stem/stromal cells.

Mesenchymal stem/stromal cells (MSCs) are isolated from various fetal and adult tissues such as bone marrow, adipose tissue, cord blood and placenta. Placental MSCs (pMSCs), the main focus of this review, are relatively new MSC types that are not as intensively studied compared with bone marrow-derived MSCs (BMMSCs). MSCs modulate the immune functions of important immune cells involved in alloantigen recognition and elimination, including antigen presenting cells (APCs), T cells, B cells and natural killer (NK) cells. Clinical trials, both completed and underway, employ MSCs to treat various human immunological diseases, such as multiple sclerosis (MS) and type 1 diabetes. However, the mechanisms that mediate the immunosuppressive effects of pMSCs are still largely unknown, and the safety of pMSC use in clinical settings needs further confirmation. Here, we review the current knowledge of the immunosuppressive properties of placental MSCs.

Phenotypic and Functional Characterization of Mesenchymal Stem/Multipotent Stromal Cells from Decidua Basalis of Human Term Placenta.

Mesenchymal stem cell (MSC) therapies for the treatment of diseases associated with inflammation and oxidative stress employ primarily bone marrow MSCs (BMMSCs) and other MSC types such as MSC from the chorionic villi of human term placentae (pMSCs). These MSCs are not derived from microenvironments associated with inflammation and oxidative stress, unlike MSCs from the decidua basalis of the human term placenta (DBMSCs). DBMSCs were isolated and then extensively characterized. Differentiation of DBMSCs into three mesenchymal lineages (adipocytes, osteocytes, and chondrocytes) was performed. Real-time polymerase chain reaction (PCR) and flow cytometry techniques were also used to characterize the gene and protein expression profiles of DBMSCs, respectively. In addition, sandwich enzyme-linked immunosorbent assay (ELISA) was performed to detect proteins secreted by DBMSCs. Finally, the migration and proliferation abilities of DBMSCs were also determined. DBMSCs were positive for MSC markers and HLA-ABC. DBMSCs were negative for hematopoietic and endothelial markers, costimulatory molecules, and HLA-DR. Functionally, DBMSCs differentiated into three mesenchymal lineages, proliferated, and migrated in response to a number of stimuli. Most importantly, these cells express and secrete a distinct combination of cytokines, growth factors, and immune molecules that reflect their unique microenvironment. Therefore, DBMSCs could be attractive, alternative candidates for MSC-based therapies that treat diseases associated with inflammation and oxidative stress.

Phenotypic and Functional Characterization of Mesenchymal Stem/Multipotent Stromal Cells From Decidua Parietalis of Human Term Placenta.

Mesenchymal stem/multipotent stromal cells (MSCs) from the human placenta show stem cell-like properties useful for regenerative medicine. Previously, we reported that MSCs isolated from the fetal part of human term placentae have characteristics, which make them a potential candidate for regenerative medicine. In this study, we characterized MSC isolated from the maternal part of human term placenta. The MSCs were isolated from the decidua parietalis (DPMSCs) of human placenta using a digestion method and characterized by colony-forming unit assay and the expression of MSC markers by flow cytometry technique. In addition, DPMSC differentiation into the 3 mesenchymal lineages was also performed. Moreover, the gene and protein expression profiles of DPMSCs were identified by real-time polymerase chain reaction and flow cytometry techniques, respectively. Furthermore, proteins secreted by DPMSCs were detected by sandwich enzyme-linked immunosorbent assays. Finally, the proliferation and migration potentials of DPMSCs were also determined. The DPMSCs were positive for MSC markers and negative for hematopoietic and endothelial markers, as well as costimulatory molecules and HLA-DR. Functionally, DPMSCs formed colonies and differentiated into chondrocytes, osteocytes, and adipocytes. In addition, they proliferated and migrated in response to different stimuli. Finally, they expressed and secreted many biological and immunological factors with multiple functions. Here, we carry out an extensive characterization of DPMSCs of human placenta. We report that these cells express and secrete a wide range of molecules with multiple functions, and therefore, we suggest that these cells could be an attractive candidate for cell-based therapy.

Human Chorionic Villous Mesenchymal Stem Cells Modify the Functions of Human Dendritic Cells, and Induce an Anti-Inflammatory Phenotype in CD1+ Dendritic Cells.

BACKGROUND: Mesenchymal stem cells derived from the chorionic villi of human term placenta (pMSCs) have drawn considerable interest because of their multipotent differentiation potential and their immunomodulatory capacity. These properties are the foundation for their clinical application in the fields of stem cell transplantation and regenerative medicine. Previously, we showed that pMSCs induce an anti-inflammatory phenotype in human macrophages. In this study, we determined whether pMSCs modify the differentiation and maturation of human monocytes into dendritic cells (DCs). The consequences on dendritic function and on T cell proliferation were also investigated. METHODS: Interleukin-4 (IL-4) and granulocyte-macrophage colony stimulating factor (GM-CSF) were used to stimulate the differentiation of monocytes into immature dendritic cells (iDCs), which were subsequently co-cultured with pMSCs. Lipopolysaccharide (LPS) was used to induce maturation of iDCs into mature dendritic cells (mDCs). Flow cytometry and enzyme-linked immunosorbent assays (ELISA) were used to quantify the effect pMSC co-culturing on DC differentiation using CD1a, a distinctive marker of DCs, as well as other molecules important in the immune functions of DCs. The phagocytic activity of iDCs co-cultured with pMSCs, and the effects of iDCs and mDC stimulation on T cell proliferation, were also investigated. RESULTS: Monocyte differentiation into iDCs was inhibited when co-cultured with pMSCs and maturation of iDCs by LPS treatment was also prevented in the presence of pMSCs as demonstrated by reduced expression of CD1a and CD83, respectively. The inhibitory effect of pMSCs on iDC differentiation was dose dependent. In addition, pMSC co-culture with iDCs and mDCs resulted in both phenotypic and functional changes as shown by reduced expression of costimulatory molecules (CD40, CD80, CD83 and CD86) and reduced capacity to stimulate CD4(+) T cell proliferation. In addition, pMSC co-culture increased the surface expression of major histocompatibility complex (MHC-II) molecules on iDCs but decreased MHC-II expression on mDCs. Moreover, pMSC co-culture with iDCs or mDCs increased the expression of immunosuppressive molecules [B7H3, B7H4, CD273, CD274 and indoleamine-pyrrole 2,3-dioxygenase (IDO). Additionally, the secretion of IL-12 and IL-23 by iDCs and mDCs co-cultured with pMSCs was decreased. Furthermore, pMSC co-culture with mDCs decreased the secretion of IL-12 and INF-γ whilst increasing the secretion of IL-10 in a T cell proliferation experiment. Finally, pMSC co-culture with iDCs induced the phagocytic activity of iDCs. CONCLUSIONS: We have shown that pMSCs have an inhibitory effect on the differentiation, maturation and function of DCs, as well as on the proliferation of T cells, suggesting that pMSCs can control the immune responses at multiple levels.

Human placental mesenchymal stem cells (pMSCs) play a role as immune suppressive cells by shifting macrophage differentiation from inflammatory M1 to anti-inflammatory M2 macrophages.

BACKGROUND: Mesenchymal stem cells (MSCs) have a therapeutic potential in tissue repair because of capacity for multipotent differentiation and their ability to modulate the immune response. In this study, we examined the ability of human placental MSCs (pMSCs) to modify the differentiation of human monocytes into macrophages and assessed the influence of pMSCs on important macrophage functions. METHODS: We used GM-CSF to stimulate the differentiation of monocytes into the M1 macrophage pathway and then co-cultured these cells with pMSCs in the early stages of macrophage differentiation. We then evaluated the effect on differentiation by microscopic examination and by quantification of molecules important in the differentiation and immune functions of macrophages using flow cytometry and ELISA. The mechanism by which pMSCs could mediate their effects on macrophage differentiation was also studied. RESULTS: The co-culture of pMSCs with monocytes stimulated to follow the inflammatory M1 macrophage differentiation pathway resulted in a shift to anti-inflammatory M2-like macrophage differentiation. This transition was characterized by morphological of changes typical of M2 macrophages, and by changes in cell surface marker expression including CD14, CD36, CD163, CD204, CD206, B7-H4 and CD11b, which are distinctive of M2 macrophages. Co-culture with pMSCs reduced the expression of the costimulatory molecules (CD40, CD80 and CD86) and increased the expression of co-inhibitory molecules (CD273, CD274 and B7-H4) as well as the surface expression of major histocompatibility complex (MHC-II) molecules. Furthermore, the secretion of IL-10 was increased while the secretion of IL-1ß, IL-12 (p70) and MIP-1α was decreased; a profile typical of M2 macrophages. Finally, pMSCs induced the phagocytic activity and the phagocytosis of apoptotic cells associated with M2- like macrophages; again a profile typical of M2 macrophages. We found that the immunoregulatory effect of pMSCs on macrophage differentiation was mediated by soluble molecules acting partially via glucocorticoid and progesterone receptors. CONCLUSIONS: We have shown that pMSCs can transition macrophages from an inflammatory M1 into an anti-inflammatory M2 phenotype. Our findings suggest a new immunosuppressive property of pMSCs that may be employed in the resolution of inflammation associated with inflammatory diseases and in tissue repair.