**5. Role of Stem Cells in Coronary Collateral Growth**

In addition to the aforementioned chemical mediators that mitigate the consequences of vascular occlusive diseases by stimulating collateral growth, in recent years, stem cell-based therapy has been implicated as a possible avenue for vascular regeneration. Stem cells have the unique potential of developing into many different cell types in the body. Under certain physiologic and experimental conditions, they can be manipulated to grow into specific tissues and organ cells with exclusive functionality. This revolutionary discovery for stem cells has demonstrated a clinical potential to create new networks of blood-perfused vessels and treat human patients with cardiovascular and vascular diseases [55]. The current theory is that stem cells may release a series of angiogenic factors, such as VEGF and bFGF, which mobilize vascular endothelial cells through a paracrine effect [56]. In this section, we will summarize the current state of regenerative approaches using stem cells to stimulate coronary collateral growth.

A 2012 study programmed endothelial cells to develop into induced vascular progenitor cells (iVPCs) and assessed their ability to induce coronary collateral growth in a rat model in efforts to increase blood flow to the collateral-dependent region of risk [57]. iVPCs are also known to be less tumorigenic compared with induced pluripotent cells (iPSCs) and are more likely to commit to a line of vascular differentiation (they will not turn into cardiomyocytes) [57]. When the iVPCs were transplanted into myocardium, they formed blood vessels and improved blood flow markedly better than did natural endothelial cells, mesenchymal stem cells, or iPSCs [57]. However, while results showed that partial programming of the endothelial cells was promising enough to sprout new blood vessels in the myocardium, one big challenge persists: how to maintain the partial programmed state of the cells until they get to their intended destination [57].

In addition, current literature posits that bone marrow-derived stem cells and endothelial progenitor cells in arteriogenesis do not physically deposit onto the walls of newly generated arteries but rather play the role of supporting cells [58]. The therapeutic induction of collateral growth from already established arteries improves any blood flow deficiencies caused by blockage in major arteries. Transplanted bone marrow-derived cells act as "cytokine factors" and secrete specific growth factors that mediate their effects through paracrine activity [59]. As Dr. Matthias Heil of the Netherlands puts it, "bone marrow stem cells provide the software and not the hardware in vascular growth" [59]. His group's study on the hindlimb ischemic model with mice revealed that GFP-tagged bone marrow was not localized to endothelial and smooth cell markers, but around burgeoning collaterals that were secreting chemokines and growth factors [59]. Hence, therapeutic arteriogenesis functions to boost the body's natural angiogenic ability by stimulating the release of pro-angiogenic factors rather than actually providing the buildings block for a new artery. A caveat to this is if the processes in the heart are different from those in the peripheral circulation.

While there is a continued debate on whether bone marrow-derived multipotent stromal cells (MSCs) exert their effect via transdifferentiation or through paracrine activity, there is unequivocal evidence showing that MSCs must first travel to ischemic tissue to achieve a therapeutic benefit [60]. MSCs localize to injured tissues by adhering to endothelial cells and migrating across the cell wall. Homing of MSCs to injured tissues is optimized by an expression of ligands on endothelial cells [60]. A 2009 study showed the importance of epidermal growth factor (EGF) and heparinbinding epidermal growth factor-like growth factor (HB-EGF) in inducing increased expression of these ligands [60]. Specifically, phosphorylation of the EGF-R leads to higher expression of ligands, VCAM-1, and ICAM-1 that enhanced MSC adherence and ultimately stimulated coronary collateral growth in rats that had undergone repetitive instances of myocardial ischemia [60]. Coronary collateral growth was assessed with the ratio of collateral dependent flow (CZ) to normal zone flow (NZ). Exposure of both MSCs and coronary endothelial cells (CECs) to a 100 ng/mL dose of EGF for 16 hours maximally increased expression of adhesion molecules compared with samples untreated with EGF [60]. The CZ/NZ ratio increased in rats whose MSCs were treated with EGF and showed improved cardiac function and decreased left ventricular remodeling compared to rats without EGF treatment of MSCs [60].

Another 2009 study involving a rat model of repetitive myocardial ischemia showed that granulocyte-colony stimulating factor (G-CSF), a glycoprotein responsible for hematopoietic cell proliferation and differentiation of neutrophil granulocytes, also stimulates coronary collateral growth [61]. G-CSF mounts a series of defenses against infectious agents, one of which is promotion of neutrophils to release reactive oxygen species (ROS) [61]. This generation of ROS was studied both *in vivo* and *in vitro* and was shown to directly act on injured cardiomyocytes. Cardiomyocytes under the influence of G-CSF-induced ROS generate angiogenic factors that lead to vascular growth and tube formation in levels comparative to cardiomyocytes induced by VEGF [61]. To the surprise of researchers, this study also demonstrated that G-CSF can promote coronary collateral growth without the impetus of repetitive ischemia and hence this cytokine can act as a surrogate for ischemia [61].

Majority of the recent clinical trials in humans purport that stem cell-based therapy adequately facilitates angiogenesis in patients suffering from peripheral arterial disease and promotes wound healing [55]. Specifically, bone marrow-derived stem cell transplantation has shown to improve ischemic symptoms, such as claudication, ischemic rest pain, and has augmented wound healing in ulcer-related conditions [55]. Nonetheless, these studies have been limited by a lack of care standardization, absence of a control group, small sample sizes, dissimilar inclusion criteria, and inconsistencies in methods of outcome assessment [55]. In other cases, the absence of follow-up procedures has prevented elucidation of long-term effects of treating peripheral artery disease with stem cells [55]. While the central issues of public safety and treatment efficacy linger over the field, progress, albeit limited, has been made in the arena of coronary collateral growth.

## **6. Summary**

will be extremely valuable in understanding the overall chemical mechanism behind collat-

In addition to the aforementioned chemical mediators that mitigate the consequences of vascular occlusive diseases by stimulating collateral growth, in recent years, stem cell-based therapy has been implicated as a possible avenue for vascular regeneration. Stem cells have the unique potential of developing into many different cell types in the body. Under certain physiologic and experimental conditions, they can be manipulated to grow into specific tissues and organ cells with exclusive functionality. This revolutionary discovery for stem cells has demonstrated a clinical potential to create new networks of blood-perfused vessels and treat human patients with cardiovascular and vascular diseases [55]. The current theory is that stem cells may release a series of angiogenic factors, such as VEGF and bFGF, which mobilize vascular endothelial cells through a paracrine effect [56]. In this section, we will summarize the current state of regenerative approaches using stem cells to stimulate coronary collateral growth.

A 2012 study programmed endothelial cells to develop into induced vascular progenitor cells (iVPCs) and assessed their ability to induce coronary collateral growth in a rat model in efforts to increase blood flow to the collateral-dependent region of risk [57]. iVPCs are also known to be less tumorigenic compared with induced pluripotent cells (iPSCs) and are more likely to commit to a line of vascular differentiation (they will not turn into cardiomyocytes) [57]. When the iVPCs were transplanted into myocardium, they formed blood vessels and improved blood flow markedly better than did natural endothelial cells, mesenchymal stem cells, or iPSCs [57]. However, while results showed that partial programming of the endothelial cells was promising enough to sprout new blood vessels in the myocardium, one big challenge persists: how to maintain the

partial programmed state of the cells until they get to their intended destination [57].

are different from those in the peripheral circulation.

In addition, current literature posits that bone marrow-derived stem cells and endothelial progenitor cells in arteriogenesis do not physically deposit onto the walls of newly generated arteries but rather play the role of supporting cells [58]. The therapeutic induction of collateral growth from already established arteries improves any blood flow deficiencies caused by blockage in major arteries. Transplanted bone marrow-derived cells act as "cytokine factors" and secrete specific growth factors that mediate their effects through paracrine activity [59]. As Dr. Matthias Heil of the Netherlands puts it, "bone marrow stem cells provide the software and not the hardware in vascular growth" [59]. His group's study on the hindlimb ischemic model with mice revealed that GFP-tagged bone marrow was not localized to endothelial and smooth cell markers, but around burgeoning collaterals that were secreting chemokines and growth factors [59]. Hence, therapeutic arteriogenesis functions to boost the body's natural angiogenic ability by stimulating the release of pro-angiogenic factors rather than actually providing the buildings block for a new artery. A caveat to this is if the processes in the heart

While there is a continued debate on whether bone marrow-derived multipotent stromal cells (MSCs) exert their effect via transdifferentiation or through paracrine activity, there is unequivocal

eral vessel growth and how to apply this knowledge to a clinical setting.

**5. Role of Stem Cells in Coronary Collateral Growth**

142 Physiologic and Pathologic Angiogenesis - Signaling Mechanisms and Targeted Therapy

The process of coronary collateral growth is being better understood year by year. The role that the many chemical factors, mechanical factors, and stem cells play in the process is still incompletely understood. The study of these factors in "normal" preclinical models may be an oversimplification, because under conditions with risk factors for coronary disease, there may be shifts in the normal control mechanisms. We advocate that future studies incorporate models of cardiovascular disease and aging to better understand the mechanisms by which this adaptive process is abrogated in the majority of patients with ischemic heart disease.

## **Author details**

Bhamini Patel, Peter Hopmann, Mansee Desai, Kanithra Sekaran, Kathleen Graham, Liya Yin and William Chilian\*

\*Address all correspondence to: wchilian@neomed.edu

Department of Integrative Medical Science, Northeast Ohio Medical University, Rootstown, Ohio, USA

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an oversimplification, because under conditions with risk factors for coronary disease, there may be shifts in the normal control mechanisms. We advocate that future studies incorporate models of cardiovascular disease and aging to better understand the mechanisms by which this adaptive process is abrogated in the majority of patients with ischemic heart

Bhamini Patel, Peter Hopmann, Mansee Desai, Kanithra Sekaran, Kathleen Graham, Liya Yin

Department of Integrative Medical Science, Northeast Ohio Medical University, Rootstown,

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**Author details**

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\*Address all correspondence to: wchilian@neomed.edu

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**Provisional chapter**
