Glutaminase in Arterial Injury and Disease

点击翻译按钮获取中文摘要

PROJECT SUMMARY The broad long-term goal of this proposal is to establish the enzyme glutaminase-1 (GLS1) as a critical mediator of lesion formation in arterial injury and disease. GLS1 is a mitochondrial enzyme that metabolizes glutamine (Gln) to glutamate (Glu) and ammonia (NH3). GLS1 plays a critical role in neurotransmission, acid- base balance, angiogenesis, hepatic fibrosis, and tumor growth, but little is known regarding the function of GLS1 in vascular smooth muscle cells (SMCs). However, preliminary studies demonstrated that the proliferation, migration, and survival of SMCs is highly dependent on the presence of Gln. It was also discovered that SMCs exclusively express the GLS1 isoform, that overexpression of GLS1 stimulates SMC migration, and that inhibition of GLS1 activity or expression blocks SMC DNA synthesis or migration, respectively. Moreover, Glu, but not NH3, substitutes for Gln in promoting SMC proliferation and collagen synthesis. Additional experiments revealed that growth factors and glucose stimulate GLS1 activity in vascular SMCs. Final pilot experiments also demonstrated that arterial injury and diabetes induces the expression of GLS1 in the vessel wall, and that inhibition of GLS1 activity or genetic depletion of GLS1 attenuates neointima formation following arterial injury as well as arterial fibrosis and stiffening in diabetic animals. Based on these findings, it is proposed that GLS1 plays an integral role in aberrant arterial remodeling by stimulating SMC proliferation, migration, collagen synthesis, and survival by metabolizing Gln to Glu and NH3. This hypothesis will be tested in three interrelated specific aims. In aim 1, the role of GLS1 in regulating SMC function will be determined. These studies will investigate the effect of GLS1 gene delivery on SMC proliferation, migration, collagen synthesis and survival, and determine the role of the various GLS1 products on these processes. They will also explore if the induction of GLS1 by growth factors contributes to their ability to promote SMC proliferation, migration, and collagen synthesis. In aim 2, the role of GLS1 in regulating the arterial response to injury will be established. These studies will examine the time-course of GLS1 expression in injured rodent carotid arteries, and determine if pharmacological inhibition of GLS1 activity, silencing GLS1 expression, or genetic deletion of GLS1 in SMCs attenuates the remodeling response following arterial injury. In aim 3, the role of GLS1 in aberrant arterial remodeling and hypertension in diabetes will be investigated. These studies will examine the effect of glucose on GLS1 expression both in cultured vascular SMCs and in arteries from diabetic mice. In addition, they will examine if GLS1 contributes to glucose-mediated alterations in SMC function, arterial remodeling, and hypertension in diabetic mice. It is anticipated that these studies will establish GLS1 as a key regulator of SMC proliferation, migration, collagen synthesis, and survival. They may also also identify GLS1 and its products as novel contributors to lesion formation following arterial injury, and establish GLS1 as a new translational target in treating abnormal remodeling and hypertension in diabetes.