Showing papers by "Brian R. Wamhoff published in 2009"

Molecular Mechanisms of Collagen Isotype-Specific Modulation of Smooth Muscle Cell Phenotype

Abstract: Objective— Smooth muscle cell (SMC) phenotypic modulation, an important component of atherosclerosis progression, is critically regulated by the matrix, with normal components of the healthy SMC matrix limiting modulation and atherosclerosis-associated transitional matrix proteins promoting phenotypic modulation. We sought to determine how collagen IV (which comprises the healthy artery wall) and monomeric collagen I (which comprises atherosclerotic lesions) differentially affect SMC phenotype. Methods and Results— Plating SMCs on collagen IV resulted in elevated expression of SMC contractility proteins compared to collagen I. Concurrent with enhanced contractile gene expression, collagen IV stimulates binding of SRF to CArG boxes in the promoters of smooth muscle actin and smooth muscle myosin heavy chain. Coll IV also stimulated the expression of myocardin, a critical SRF coactivator required to drive expression of SMC specific genes. In contrast to collagen IV, collagen I stimulated enhanced expression of the inflammatory protein vascular cell adhesion molecule (VCAM)-1. NF-κB and NFAT-binding sites in the VCAM-1 promoter are critical for collagen I-mediated expression of VCAM-1 promoter activity. However, only inhibitors of NFAT, not NF-κB, were able to reduce collagen I-associated VCAM expression, and collagen I but not collagen IV stimulated NFAT transcriptional activity. Conclusion— These results show for the first time that collagen IV and collagen I differentially affect smooth muscle phenotypic modulation through multiple pathways.

PDGF-DD, a novel mediator of smooth muscle cell phenotypic modulation, is upregulated in endothelial cells exposed to atherosclerosis-prone flow patterns

Abstract: Platelet-derived growth factor (PDGF)-BB is a well-known smooth muscle (SM) cell (SMC) phenotypic modulator that signals by binding to PDGF αα-, αβ-, and ββ-membrane receptors. PDGF-DD is a recentl...

Human IL-8 Regulates Smooth Muscle Cell VCAM-1 Expression in Response to Endothelial Cells Exposed to Atheroprone Flow

Abstract: Objective— Interleukin-8 (IL-8) is a soluble human-specific chemokine implicated in the development of the chronic inflammatory disease atherosclerosis. Recently, we showed that atheroprone hemodynamics induced IL-8 secretion from endothelial cells (ECs) concurrent with increased EC/smooth muscle cell (SMC) VCAM-1 expression in a human hemodynamic coculture model. Despite an IL-8 association with inflammation, we show here that blocking IL-8 activity during atheroprone flow resulted in increased levels of EC/SMC VCAM-1 expression. We tested the hypothesis that IL-8 limits SMC VCAM-1 expression in response to inflammatory stimuli, either atheroprone flow or cytokine interleukin-1β (IL-1β) addition. Methods and Results— Atheroprone flow increased monocyte adhesion in both EC/SMCs, concurrent with the induction of VCAM-1 protein. VCAM-1 antisera attenuated this response. IL-1β upregulated VCAM-1 in SMCs by 3-fold, a response inhibited by the addition of IL-8 at 24 hours. Neither IL-1β nor IL-8 induced proliferation or migration. Neutralization of the IL-8 receptor, CXCR2, further induced VCAM-1 in the presence of IL-1β, and phospho-p38 was required for NF-κB activation and VCAM-1 expression. Additionally, IL-8 reduced p38 activation and NF-κB activity induced by IL-1β alone. Conclusions— Together, these findings provide evidence for a novel role whereby IL-8 limits the inflammatory response in ECs/SMCs via VCAM-1 modulation.

Intravascular ultrasound detection and delivery of molecularly targeted microbubbles for gene delivery

Abstract: We have investigated microbubble based targeted delivery and combined intravascular ultrasound (IVUS) imaging as potential therapy to reduce incidence of restenosis following stent placement in atherosclerotic coronary arteries. The goal of these studies was to determine whether IVUS could be used to detect targeted microbubbles and enhance drug/gene delivery through targeting. Fluorescently labeled microbubbles targeted to the inflammatory cell surface marker VCAM-1 were combined with cells under flow to measure adhesion compared to control bubbles. Gene delivery was performed using targeted bubble constructs and 1MHz ultrasound at 200 and 300 kPa - acoustic pressures which can be generated by a modified commercial IVUS catheter. Detection of adherent microbubbles to inflamed cells in culture and flow chambers was measured using a clinical IVUS catheter. VCAM-1 targeted microbubbles enhanced adhesion to inflamed cells up to 100 fold over non-targeted microbubbles. Compared to non-inflamed cells VCAM-1 targeted bubbles exhibited a 7.9 fold increase in adhesion to IL- 1beta treated cells. Targeted microbubbles resulted in a 5.5 fold increase in plasmid DNA transfection over non targeted bubbles in conjunction with a focused 1-inch diameter 1MHz transducer and 1.5 fold increase following insonation from a fabricated IVUS transducer at 1.5 MHz. At an equivalent density of 3×104 bubbles/mm2, IVUS image intensity increased 4.3 fold over that of non-bubble-coated surfaces. Rupture of microbubbles from the modified IVUS transducer resulted in a 53% reduction in image intensity. Taken together, these results indicate IVUS may be used to detect targeted microbubbles to inflamed vasculature and subsequently deliver a gene/drug locally.

Poisson inverse gradient approach to vascular myocyte detection and segmentation

Abstract: This paper addresses the detection and segmentation of vascular myocytes. The detection and segmentation of these cells are critical to the investigation of atherosclerosis among other cardiovascular diseases. Our approach to detection is unique in that it attempts to compute the underlying external energy in an active contour model. Isolines in this computed external energy can be employed to localize boundaries of the cell nuclei. The process used to solve the inverse problem of obtaining the energy from the force vectors is called Poisson inverse gradient due to the Poisson-based solution. From the initial contours given by the isolines in the computed energy, parametric active contours are used to find the subtle cell boundaries. The results indicate that the Poisson inverse gradient improves detection accuracy and reduces false positives compared to existing morphological methods. Furthermore, the contour-based detection allows segmentation of the cell boundaries.