Molecular Mechanisms of Interleukin-1beta-Stimulated Regulation of Angiogenesis in Cardiac Microvascular Endothelial Cells.

<p>Angiogenesis, the formation of new vessels from a preexisting vasculature, is critical for supplying a healing myocardium with oxygen and nutrients to sustain metabolism post myocardial infarction (MI). Interleukin-1&#946; (IL-1&#946;), a proinflammatory cytokine increased in the heart post-MI, is considered essential for angiogenesis in tumor growth and metastasis, arthritis, endometriosis, and wound healing. Matrix metalloproteinases (MMPs) are implicated in angiogenesis because of their ability to selectively degrade components of the extracellular matrix. Vascular endothelial growth factors (VEGFs) play a vital role in angiogenesis because of their involvement in the recruitment and proliferation of endothelial cells. The current study explores IL-1&#946;-stimulated regulation of angiogenic genes in cardiac microvascular endothelial cells (CMECs), the signaling mechanisms involved, and the implications in the processes of angiogenesis. DNA microarray analysis indicated IL-1&#946; modulates the expression of numerous angiogenesis-related genes, notably upregulating MMP-2 and downregulating VEGF-D expression. RT-PCR and Western blot analyses confirmed the differential expression in response to IL-1&#946;. In-gel zymographic analysis demonstrated IL-1&#946;-stimulated increase in MMP-2 activity. IL-1&#946; activated ERK1/2 and JNKs, not p38 kinase, and activated PKC&#945;/&#946;₁ independent of MAPKs. IL-1&#946; inactivated GSK3&#946; via ERK1/2. Pharmacological inhibition of these signaling cascades indicated IL-1&#946;-stimulated regulation of MMP-2 and VEGF-D occurs via ERK1/2, JNKs, and PKC&#945;/&#946;₁-dependent mechanisms. In addition, inactivation of GSK3&#946; inhibited basal VEGF-D expression. H₂O₂ significantly increased MMP-2 protein levels while IL-1&#946;-induced VEGF-D downregulation was further potentiated by ROS scavenging compounds and inhibition of NF-&#954;B. Phalloidin-FITC stain indicated a sharp reduction in fibrillar actin in the cytoskeleton of IL-1&#946;-stimulated cells. Wounding assays revealed that IL-1&#946; induced CMEC migration but prevented cell-to-cell contact and restoration of the monolayer. Flow cytometric analysis revealed a G₀/G₁ phase cell cycle arrest in IL-1&#946;-stimulated cells, indicative of decreased proliferation. IL-1&#946; inhibited three-dimensional <em>in vitro</em> tube formation by CMECs. Lastly, IL-1&#946; inhibited microvessel sprouting from aortic rings, an assay examining the collective response of multiple cell types. Collectively, the data presented in this study provide evidence that IL-1&#946; differentially regulates important angiogenesis-related genes in CMECs. This differential regulation may lead to interruptions in the processes of angiogenesis, ultimately creating a dysfunctional phenotype for myocardial vessel formation.</p>

Name thesis:degree_name

PhD (Doctor of Philosophy)

Level thesis:degree_level

Dissertation - restricted

Discipline thesis:degree_discipline

Biomedical Sciences

Year dc:date.issued

2007

Author dc:creator

• Mountain, Deidra Jill Hopkins

• angiogenesis
• interleukin-1beta
• VEGF
• MMP
• endothelial cells
• Amino Acids, Peptides, and Proteins
• Chemicals and Drugs
• Medicine and Health Sciences

Statement dc:rights

• Copyright by the authors.

Repository record dc:identifier

https://dc.etsu.edu/etd/2136

OAI identifier oai:identifier

oai:dc.etsu.edu:etd-3497