*2.2.2 Exosomal pro- and anti-angiogenic factors*

MSCs can secret signal transducer and activator of transcription-3 (STAT3) mRNAs via exosomes that augment the transcription of hepatocyte growth factor (HGF), IL-6, and VEGF, promoting proliferation and migration of ECs [99]. In this context, MSC exosomes abundantly are enriched with VEGF factor that increases neovascularization through the Wnt4/β-catenin pathway in epithelial cells [100, 101]. The pro-angiogenic propriety of MSC exosomes has been previously shown in myocardial ischemia/reperfusion injury experiments following acute myocardial infarction [102–104]. In contrast, MSC exosomes may contain abundant anti-angiogenic factors that could regulate tumor angiogenesis rate. Lee et al. showed that exosomes from MSCs significantly downregulated the expression of VEGF in breast cancer cells, leading to the abortion of angiogenesis [91]. However, there are contradicting results. For example, human bone marrow MSC

exosomes promoted VEGF synthesis in colonic and gastric carcinomas through the activation of extracellular signal-regulated kinase1/kinase2 (ERK1/ERK2) and p38 MAPK pathways [105]. Taken together, these issues show a fact that exosomes from various MSC types can mediate physiological and pathological angiogenesis and could be considered as a suitable bio-shuttle for establishing promising therapeutic approaches in an individual with cancers and ischemic pathologies. The feasibility of exosome uptake by recipient cells, make these cell products for introducing in clinical approaches. Xue and colleagues investigated the effects of cord blood and adipose-derived MSC exosomes on human EC angiogenesis capacity under hypoxic and normal conditions [106, 107]. They noted the potency of isolated exosomes in triggering angiogenesis rate especially under the hypoxic condition compared to exosome counterpart originated from normal milieu. Based on their data, the transcription level of genes related to angiogenesis such as angiopoietin-1 (Ang-1) and VEGF receptor-2 (also termed FLK-1) was induced significantly after exposure to exosomes collected from hypoxic MSCs rather than that of normal cells. Following the induction of Ang-1 and FLK-1, the status of some downstream effectors would be turned to an activated form. For instance, it was found that protein kinase A (PKA) is indirectly triggered after the activation of genes *Ang-1* and *VEGFR-2*. Along with changes, the transcription level of angiogenesis inhibitory gene like Vash1 is completely suppressed. The inhibitory angiogenesis potential of MSCs was investigated on cancer cells or progenitors residing inside tumor mass. Both anti-inflammatory and pro-angiogenesis property of MSC-derived exosomes were shown in cardiovascular disease [92, 97]. In addition to the promotion of cell surface receptors, exosomes could augment the synthesis of VEGF factor in targeted cells. Doeppner et al*.* also previously demonstrated that MSC-derived exosomes initiated healing processes after the onset of neurological diseases by increasing angiogenesis and blood supply which led to the neurological recovery and neurogenesis [108]. Other experiments added notion on the potency of exosomes to reduce neuroinflammation in traumatic brain injury [109]. However, some contradictory facts exist regarding the sole application of exosomes in the context of tumor cells. The superior stimulatory effect of MSC-derived exosomes on tumor angiogenesis was also addressed by different authors [110]. For example, Zhu et al. demonstrated the vasculogenic role of MSC exosomes after addition to human gastric carcinoma (SGC-7901) and colon cancer (SW480) cell lines [105, 111]. They found that the normal status of signaling effectors such as phosphorylated ERK1/ ERK2, Bcl-2, and VEGF proteins; alpha-smooth muscle actin (α-SMA); CXCR-4; and mouse double minute 2 homolog (MDM2) mRNA was modulated in the favor of angiogenesis in a mouse cancer model. In addition to the direct fast action on recipient cells, it is reasonable to hypothesize that exosomes are able to dictate pro-/ anti-angiogenesis pattern in distinct cells by provoking specific signaling pathways and effectors such as ERK1/ERK2 and p38 MAPK kinase routes.

The engagement of factors such as AKT, STAT3, Wnt/β-catenin, and ERK happens following cutaneous wound regeneration treated with MSC exosomes. Proteomic analysis revealed that the protein content of growth factors IL-6, stromal cell-derived factor-1α (SDF-1α), IGF-1α, STAT3, and HGF contributed to cell proliferation, migration, and angiogenesis, improving reepithelialization in wound sites [112]. The modulation of Wnt/β-catenin pathway targeting Wnt4 diminishes the number of cells with apoptotic changes with the levels of pro-angiogenic factors such as IL-6 and IL-8, granulocyte-colony-stimulating factor (G-CSF), PDGF-BB, MCP-1, and VEGF are increased. In response to treatment with exosomes, phosphorylation of glycogen synthase kinase 3β (GSK3β) as a main negative regulator of Wnt signaling pathway is initiated, resulting in the progression of a cell from phase G1 to S and cutaneous cell proliferation [113]. An enhanced angiogenesis rate

**109**

*The Angiogenic Paracrine Potential of Mesenchymal Stem Cells*

with reduced cardiomyocyte apoptosis was reported following the administration of MSCs to infarct area. After being exposed to the ischemic/hypoxic condition, MSCs were programmed to secrete exosomes. Under these conditions, GATA-4 is induced which prevents cell apoptosis, reducing the infarct size. Meanwhile, the level of an anti-apoptotic agent such as miR-19a and miR-22 was increased in the target sites [114]. In another experiment conducted by Teng et al., it was shown that MSC-derived exosomes harboring miRNA-132 efficiently are delivered to human umbilical vein ECs (HUVECs). Therefore, it could be pointed out that MSCs could dictate prominent changes in the target cells. They also declared that endothelial Ras signaling pathway effectors are modulated by recipient cells after direct interaction of this miRNA with *RASA1* gene. Ras group genes have a basic role in controlling cell proliferation and differentiation [107]. Along with these statements, the bona fide effects of MSC exosomes need to be precisely addressed by a plethora of

In the context of tumor niche, both anti- and pro-tumorigenic features was found after the treatment of cancer cells with MSCs exosomes. The migration and proliferation of tumor cells were tightly regulated by exosome factors by the modulation of PDGFR, C-Met, and EGFR signaling. Ex vivo modulation showed this fact that MSC exosomes could activate or phosphorylate intracellular kinase domain of relevant receptors, thereby triggering Akt, PKC/PKB, and MAP signaling pathways, leading to proliferation and migration of gastric tumor cells [115]. Exosomes released by human bone marrow MSCs augmented VEGF in colonic carcinoma and gastric carcinoma tumor cells through the activation of ERK1/ERK2 and p38 MAPK pathways [105]. This hypocrisy generates doubts on the definite therapeutic effect of exosomes from MSC source in various niches. In an experiment, the lack of cell response was approved in dormant-like tumor-initiating cells [116]. The differences in tumor cells to MSC secretome may relate to the divergence of factors and dynamic growth of target cells inside tumor niche [116]. In light of various genetic and proteomic reservoir, the target signaling and possible side effects of exosome treatment are required to be investigated in relation to specific distinct signaling pathway. It seems that exosome therapy is at the beginning step, and the type and source of cells have a superior role in the orientation of target cell behavior. A more deep understanding of the regulatory signaling pathways and precise inquiry in profiling of components transferred by exosomes is required to enroll and engineer the exosomes for therapeutic angiogenesis or targeted therapy

**3. The application of MSCs and secretome in ischemic cardiac disease**

Cardiovascular diseases remain the leading cause of mortality and morbidity in worldwide. Various investigators have continued to assess a large number of cell types injected through several routes to promote cardiac repair in patients with cardiovascular diseases in both the preclinical and clinical stages. Clinical studies have largely been focused on the administration of MSCs [117, 118]. For instance, intracoronary injection of bone marrow MSCs led to an improved function of the left ventricle in subjects with acute myocardial infarction [119]. Mechanisms of action of MSCs administrated to the injured myocardium include accelerating angiogenesis process, diminished fibrosis, and regulation of immune response [102, 120]. Both in vitro and in vivo investigations have confirmed the trans-differentiating capacity of MSCs to effective cardiomyocytes in injured cardiac tissue [50]. In addition, documents revealed that MSCs from different sources release greater amounts of angiogenic factors (HGF, VEGF, and other growth factors), cell migration chemokine (SDF-1α), immune-signaling elements (IL-6, IL-8, and

*DOI: http://dx.doi.org/10.5772/intechopen.84433*

various experiments.

#### *The Angiogenic Paracrine Potential of Mesenchymal Stem Cells DOI: http://dx.doi.org/10.5772/intechopen.84433*

*Update on Mesenchymal and Induced Pluripotent Stem Cells*

and effectors such as ERK1/ERK2 and p38 MAPK kinase routes.

The engagement of factors such as AKT, STAT3, Wnt/β-catenin, and ERK happens following cutaneous wound regeneration treated with MSC exosomes. Proteomic analysis revealed that the protein content of growth factors IL-6, stromal cell-derived factor-1α (SDF-1α), IGF-1α, STAT3, and HGF contributed to cell proliferation, migration, and angiogenesis, improving reepithelialization in wound sites [112]. The modulation of Wnt/β-catenin pathway targeting Wnt4 diminishes the number of cells with apoptotic changes with the levels of pro-angiogenic factors such as IL-6 and IL-8, granulocyte-colony-stimulating factor (G-CSF), PDGF-BB, MCP-1, and VEGF are increased. In response to treatment with exosomes, phosphorylation of glycogen synthase kinase 3β (GSK3β) as a main negative regulator of Wnt signaling pathway is initiated, resulting in the progression of a cell from phase G1 to S and cutaneous cell proliferation [113]. An enhanced angiogenesis rate

exosomes promoted VEGF synthesis in colonic and gastric carcinomas through the activation of extracellular signal-regulated kinase1/kinase2 (ERK1/ERK2) and p38 MAPK pathways [105]. Taken together, these issues show a fact that exosomes from various MSC types can mediate physiological and pathological angiogenesis and could be considered as a suitable bio-shuttle for establishing promising therapeutic approaches in an individual with cancers and ischemic pathologies. The feasibility of exosome uptake by recipient cells, make these cell products for introducing in clinical approaches. Xue and colleagues investigated the effects of cord blood and adipose-derived MSC exosomes on human EC angiogenesis capacity under hypoxic and normal conditions [106, 107]. They noted the potency of isolated exosomes in triggering angiogenesis rate especially under the hypoxic condition compared to exosome counterpart originated from normal milieu. Based on their data, the transcription level of genes related to angiogenesis such as angiopoietin-1 (Ang-1) and VEGF receptor-2 (also termed FLK-1) was induced significantly after exposure to exosomes collected from hypoxic MSCs rather than that of normal cells. Following the induction of Ang-1 and FLK-1, the status of some downstream effectors would be turned to an activated form. For instance, it was found that protein kinase A (PKA) is indirectly triggered after the activation of genes *Ang-1* and *VEGFR-2*. Along with changes, the transcription level of angiogenesis inhibitory gene like Vash1 is completely suppressed. The inhibitory angiogenesis potential of MSCs was investigated on cancer cells or progenitors residing inside tumor mass. Both anti-inflammatory and pro-angiogenesis property of MSC-derived exosomes were shown in cardiovascular disease [92, 97]. In addition to the promotion of cell surface receptors, exosomes could augment the synthesis of VEGF factor in targeted cells. Doeppner et al*.* also previously demonstrated that MSC-derived exosomes initiated healing processes after the onset of neurological diseases by increasing angiogenesis and blood supply which led to the neurological recovery and neurogenesis [108]. Other experiments added notion on the potency of exosomes to reduce neuroinflammation in traumatic brain injury [109]. However, some contradictory facts exist regarding the sole application of exosomes in the context of tumor cells. The superior stimulatory effect of MSC-derived exosomes on tumor angiogenesis was also addressed by different authors [110]. For example, Zhu et al. demonstrated the vasculogenic role of MSC exosomes after addition to human gastric carcinoma (SGC-7901) and colon cancer (SW480) cell lines [105, 111]. They found that the normal status of signaling effectors such as phosphorylated ERK1/ ERK2, Bcl-2, and VEGF proteins; alpha-smooth muscle actin (α-SMA); CXCR-4; and mouse double minute 2 homolog (MDM2) mRNA was modulated in the favor of angiogenesis in a mouse cancer model. In addition to the direct fast action on recipient cells, it is reasonable to hypothesize that exosomes are able to dictate pro-/ anti-angiogenesis pattern in distinct cells by provoking specific signaling pathways

**108**

with reduced cardiomyocyte apoptosis was reported following the administration of MSCs to infarct area. After being exposed to the ischemic/hypoxic condition, MSCs were programmed to secrete exosomes. Under these conditions, GATA-4 is induced which prevents cell apoptosis, reducing the infarct size. Meanwhile, the level of an anti-apoptotic agent such as miR-19a and miR-22 was increased in the target sites [114]. In another experiment conducted by Teng et al., it was shown that MSC-derived exosomes harboring miRNA-132 efficiently are delivered to human umbilical vein ECs (HUVECs). Therefore, it could be pointed out that MSCs could dictate prominent changes in the target cells. They also declared that endothelial Ras signaling pathway effectors are modulated by recipient cells after direct interaction of this miRNA with *RASA1* gene. Ras group genes have a basic role in controlling cell proliferation and differentiation [107]. Along with these statements, the bona fide effects of MSC exosomes need to be precisely addressed by a plethora of various experiments.

In the context of tumor niche, both anti- and pro-tumorigenic features was found after the treatment of cancer cells with MSCs exosomes. The migration and proliferation of tumor cells were tightly regulated by exosome factors by the modulation of PDGFR, C-Met, and EGFR signaling. Ex vivo modulation showed this fact that MSC exosomes could activate or phosphorylate intracellular kinase domain of relevant receptors, thereby triggering Akt, PKC/PKB, and MAP signaling pathways, leading to proliferation and migration of gastric tumor cells [115]. Exosomes released by human bone marrow MSCs augmented VEGF in colonic carcinoma and gastric carcinoma tumor cells through the activation of ERK1/ERK2 and p38 MAPK pathways [105]. This hypocrisy generates doubts on the definite therapeutic effect of exosomes from MSC source in various niches. In an experiment, the lack of cell response was approved in dormant-like tumor-initiating cells [116]. The differences in tumor cells to MSC secretome may relate to the divergence of factors and dynamic growth of target cells inside tumor niche [116]. In light of various genetic and proteomic reservoir, the target signaling and possible side effects of exosome treatment are required to be investigated in relation to specific distinct signaling pathway. It seems that exosome therapy is at the beginning step, and the type and source of cells have a superior role in the orientation of target cell behavior. A more deep understanding of the regulatory signaling pathways and precise inquiry in profiling of components transferred by exosomes is required to enroll and engineer the exosomes for therapeutic angiogenesis or targeted therapy
