Regulation of blood vessel growth so that normal tissues receive sufficient oxygen and nutrients during development, homeostasis, and wound repair while is one of the central functions of the endothelium. During tumor growth, normal regulatory mechanisms of vessel growth are altered, and new vessels formation, via sprouting angiogenesis, occur allowing aggressive growth of the malignant tissue. Ischemic vascular disease and cancer are the first and second, respectively, leading cause of mortality in the United States. As our treatments for these disease states are insufficient, a better understanding of the cellular signaling pathways that control them is of considerable scientific and therapeutic interest. Kruppel-like Factors (KLFs) are a family of zinc finger proteins that regulate transcription and are implicated in a wide spectrum of biologic processes. However, a role for the KLF family in blood vessel growth is unknown. Based on our preliminary studies, we find that the 4th member of this family (KLF4) regulates tumor angiogenesis and arteriogenesis. Gain- and loss-of-function studies reveal that KLF4 regulates several critical angiogenic factors including members of the Notch signaling pathway. Mice with endothelial-specific overexpression of KLF4 have increased tumor blood vessel density, but smaller tumor size. Assessment of the tumor vasculature suggests that KLF4 overexpression leads to ineffective hypervascularity- angiogenesis is enhanced , but the new vessels are hypoperfused, limiting tumor growth. In hindlimb ischemia studies endothelial KLF4 overexpression leads to poor blood flow immediately post femoral artery ligation (decreased function of native collaterals), but enhanced collateral remodeling during the recovery period, with blood flow quickly becoming equal to WT animals. Assessment of cerebral pial collateral vessels shows decreased collateral density, consistent with the presumed decrease in skeletal muscle collaterals that cause the limited distal flow immediately post ligation. Cerebral microvessels and tracheal mucosal capillaries are increased in density; the cause of this is yet unknown and will be a topic for futue studies. The neovascularization phenotypes seen with KLF4 overexpression are reminiscent of those described in animals with altered Notch signaling. Notch activation has a well-established central role in several modes of neovascularization, and Notch-modulating therapies are under consideration for clinical use. These observations provide the foundation for the central hypothesis the KLF4 is a novel regulator of neovascularization, and functions as an upstream regulator of Notch. To better understand the precise role of KLF4 in this process, three aims are proposed.
In Aim 1, we will define the role of KLF4 in sprouting angiogenesis and vascular patterning.
In Aim 2 we will determine the role of EC KLF4 overexpression and deficiency on collateralization.
In Aim 3, we will determine the precise molecular relationship between KLF4 and Notch. Collectively, these studies will define a novel pathway, from nucleus to vascular network, that regulates blood vessel growth. The results of these studies may provide new therapies beneficial in the treatment of tumor growth or atherosclerotic vascular disease.
Abnormal or deficient blood vessel growth creates disease states such tumor growth or tissue injury and loss from cerebral, coronary and peripheral arterial disease. Our studies identify a new genetic factor that regulates blood vessel formation in tumors and in normal tissues. This proposal seeks to understand the molecular mechanisms underlying the function of this factor with the goal of developing novel therapies for the treatment of diseases such as cancer and atherosclerotic disease.
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