**1. Introduction**

182 Myocarditis

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> In response to ischemic damage, the heart undergoes vicious process of remodeling wherein the damaged myocardium is replaced by scar tissue and as compensatory mechanism, its existing collateral vessels and neovascularization with concomitant changes in cell recruitment, multiplication and cytokine/growth factor action. Angiogenesis is a complex process which involves an interplay between multiple pro- and anti-angiogenic factors and a harmonized interaction between endothelial progenitor cells, smooth muscle cells, pericytes and supportive environment. Besides Vegf/ Vegf receptor system, angiopoietin family of pro-angiogenic growth factors in conjunction with their receptor system are critical for vascular protection, remodeling, proliferation and maturation beside preservation of the integrity of newly formed vascular structures for functional activity (Thurston et al. 2000; Saharinen et al. 2005; Brindle, Saharinen et al. 2006).

> An outside intervention to support the inefficient intrinsic myocardial repair processes by administration of stem/ progenitor cells has emerged as a promising strategy for the treatment of ischemic heart diseases. The transplanted stem cells have shown both myogenic as well as vasculogenic differentiation potential and participate in the myocardial regeneration *via* angiomyogenesis (Chen et al. 2010; Uemura et al. 2006; Eguchi et al. 2007). In addition to differentiation, stem cells can also ameliorate inflammation, migrate to ischemic regions and secrete bioactive molecules as a part of their paracrine activity and significantly contribute myocardial protection and angiogenesis. Alternatively, multimodal therapeutic strategies have also been adopted to accentuate the angiomyogenic potential of stem cells. This includes preconditioning of stem cells with growth factor treatment, their genetic modification with plasmids encoding for various angiogenic growth factors and concomitant administration of recombinant angiogenic growth factor proteins (Jiang et al. 2006; Haider et al. 2008; Kim et al. 2009; Lu et al. 2009). Such multimodal treatment strategies have elicited beneficial effects in terms of improving stem cell survival and enhancing their paracrine behavior besides stimulation of angiogenesis through direct recruitment, proliferation and maturation of precursor cells such as endothelial progenitor cells, mesenchymal stem cells and monocytes to the ischemic heart (Banai et al. 1994; Hiasa et al. 2004; Elmadbouh et al. 2007; Haider et al. 2008). We discuss here the biological regulation of angiopoietin-1 expression, its interaction with specific receptor system and the advantages of transgenic over expression of angiopoietin-1 either alone or in combination

al. 2011).

and conditional environments.

Angiopoietin-1 for Myocardial Angiogenesis 185

and resulted in early embryonic death due to failure of vascular branching and differentiation. Homozygous mutated embryos also displayed abnormalities in the development of the heart (Dumont et al. 1994; Sato et al. 1995). Lack of Tie2 also resulted in angiogenic defects in term of vessel branching and remodeling, and displayed defects in the developing vessels to have scarce peri-endothelial cells and thinner collagen-like fibers (Suri et al. 1996). Dysregulated expression of Tie2 have also been observed in several clinical diseases including venous malformations, intramuscular hemangiomas, pulmonary hypertension and infantile hemangiomas (Yu et al. 2001; Wang et al. 2004; Morris et al. 2005). On the contrary, overexpression of angiopoietin-1 in the skin of experimental animal models led to the formation of highly branched and larger vessels, and resulted in reduction of microvascular leakage (Suri et al. 1998; Thurston et al. 1999). Tie2 over expression in the skin caused psoriasis-like phenotype after birth and persisted throughout adulthood, and was featured by epidermal hyperplasia, accumulation of inflammatory cells and altered dermal angiogenesis (Voskas et al. 2005). These findings clearly suggested that a delicate level of Tie2 receptor was required for physiological functioning and any unregulated induction or loss of Tie2 resulted in potentially worsened effects. Despite considerable similarity with Tie1, experiments with Tie1- or Tie2-deficient mice have provided evidence of their distinct functions in response to different members of angiopoietin family (Seegar et

It is interesting to note that angiopoietin-1 and angiopoietin-2 have different effects on vascular formation and development, however, they bind to Tie2 receptor with distinct kinetics of release following binding thus indicating that activation of Tie2 receptor is regulated independently by these two molecules. In fact, angiopoietin-2, a natural antagonist of angiopoietin-1 (Maisonpierre et al. 1997), binds to Tie2 receptor without its activation (Davis et al. 2003). Similarly, structural characteristics and distinguishable interaction with other molecules in the extracellular environment of the ligands may also essentially contribute to their counteractive properties (Kim et al. 2005). More recent studies have shown that the effects of angiopoietin-1 and angiopoietin-2 on the receptor tyrosine kinase Tie2 are differentially regulated at the endothelial cell surface (Hansen et al. 2010) and a critical balance is maintained between angiopoietin-1 and angiopoietin-2 expression by sonic hedgehog and fibroblast growth factor-2 during angiogenesis (Fujii & Kuwano, 2010). Phosphorylation of tyrosine residues of Tie2 occurs subsequent to binding with angiopoietin-1 and activates kinase domain of the receptor to initiate various downstream intracellular signaling cascades (Murray et al. 2001). The phosphorylation of tyrosine residues on the intracellular domain of Tie2 receptor interacts with the p85 subunit of PI3K *via* Src homology 2 or phosphotyrosine binding domain. These molecular changes result in activation of PI3K and its downstream Akt in the endothelial cells and ultimately lead to multiple responses such as cell survival, differentiation and chemotaxis (Witzenbichler et al. 1998; Fujikawa et al. 1999; Abdel-Malak et al. 2009; Bai et al. 2009). Although some studies have already demonstrated that angiopoietin-1 mediated activation of Tie2 does not cause mitogenesis of endothelial cells, the others have reported a pro-proliferative effect of angiopoietin-1 on vascular cells (Kanda et al. 2005; Abdel-Malak et al. 2009). These contradictions in the data may be explained on the basis of the observation that angiopoietin-1 may induce various effects on endothelial cells depending on the tissue type

**2.2 Angiopoietin-1, Tie2 receptor and intracellular signaling** 

with Vegf to support angiogenesis as a therapeutic option for the treatment of ischemic heart disease.
