Our laboratory is interested in the molecular genetics of vascular diseases. We utilize several approaches, including molecular and cellular biology studies, genetic studies in mice, and clinical investigations in patients with vascular diseases. Our focus is on signaling pathways and cell cycle proteins which regulate vascular proliferation and inflammation. The cyclin-dependent kinase inhibitor p27 arrests cell cycle progression through G1 and S phases and is regulated by phosphorylation of serine/threonine residues. Recently, we identified a new serine/threonine kinase, KIS, which phosphorylates p27 on serine 10 leading to nuclear export of p27 and protein degradation. However, the basis for the regulation of KIS gene expression is not known. To investigate the mechanism of transcriptional activation of human KIS gene expression, we isolated the genomic DNA fragment of the 5' flanking region of the KIS gene and have characterized transcriptional activation of the KIS promoter. We have identified GABP binding to Ets-binding sites as an essential factor for promoter activation, leading to KIS gene expression. Ongoing studies are investigating the phenotype of KIS knock-out mice, and knock-in mutations of the different phosphorylation sites of p27 in mice in order to better understand the molecular regulation of p27 by KIS. Our studies of signaling pathways that regulate vascular function have lead us to discover a novel function for heme oxygenase-1 (HO-1) in vascular biology, namely, protection against arterial thrombosis during vascular damage. We have dissected the mechanisms for this effect in HO-1 null mice, which include: endothelial cells in HO-1 null arteries are more susceptible to apoptosis and denudation, leading to platelet rich micro-thrombi in the subendothelium; arterial tissue factor and plasma von Willebrand?s Factor are significantly elevated in HO-1 null mice, consistent with EC loss; and inhaled carbon monoxide, a by-product of heme metabolism by HO-1, rescues the prothrombotic phenotype in HO-1 null mice. Our findings suggest an interesting role for HO-1 in the regulation of arterial thrombosis and suggest that induction of HO-1 or inhaled CO may be beneficial in the prevention of arterial thrombosis associated with vascular oxidant stress and inflammation. We are continuing data analysis of a clinical study of in-stent restenosis in order to understand the genetic susceptibility of this complex, common cardiovascular disease. Genotyping and gene expression profiling has been completed, and a clinical predictor model is being developed. The goal is to identify gene, RNA and protein profiles of patients with recurrent in-stent restenosis in order to better diagnose and triage patients undergoing these procedures and to potentially refine therapeutics.
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