Vascular Molecular Bioengineering Laboratory

The Vascular Bioengineering Laboratory at UCSD Bioengineering was established in 1988 when Dr. Shu Chien and several of his colleagues moved from Columbia University to San Diego. The Laboratory has grown from less than ten staff members to over twenty over the past decade. Our research goal is to elucidate the molecular mechanisms of the regulation of vascular functions, with a special emphasis on the transduction of mechanical stimuli, such as pressure and flow, into intracellular signaling and the ensuing gene expression and functional responses in health and disease.

Vascular endothelial cells (ECs) are continuously exposed to blood flow and distending pressure, which exert significant influences on physiological and pathophysiological processes. For instance, the regional variations in blood flow pattern play a significant role in the preferential localization of atherosclerosis at arterial branch points. Studies are conducted at different levels of the biological hierarchy, from molecules and cells to tissues and animal models, by using a combination of biological and engineering approaches. Our current emphases are on the following subjects:

Use of DNA Microarray Technology to Study Gene Expression: Recent advances in DNA microarray technology provide a powerful and efficient tool to decipher the complex patterns of gene expression in response to physical and chemical stimuli by comparing expression levels of thousands of genes under controlled experimental conditions. We have investigated the modulation of gene expression in ECs by shear stress and in smooth muscle cells (SMCs) by matrix geometry. We have demonstrated that long-term laminar shear stress (24 hr) down-regulates a number of genes to keep ECs in a relatively quiescent stage. Only a few genes are up-regulated, mainly those related to EC survival, angiogenesis and vascular remodeling. These findings provide a better understanding of the molecular basis of the protection of ECs from atherogenesis by long-term laminar shear stress, which occurs in the straight part of the arterial tree.  We currently have a dedicated facility that supports all microarray-related experimentations, with human, rat and mouse cDNA and oligonucleotide libraries and a platform to perform bioinformatics analysis.

Mechanotransduction in Endothelial Cells: We are interested in the mechanisms by which cells sense mechanical stimuli and transduce them into biochemical signals,  especially the mechano-sensor system that initiates the transduction process. Membrane receptors on both the luminal side (e.g., Flk-1), abluminal side (e.g., integrins) of the EC have been found to be activated by shear stress and play a role in mechanotransduction. Our results indicate that these receptors interact with each other as a network to regulate the downstream signaling pathways. Integrins trans-activate Flk-1 in response to shear stress, but not vice versa. Integrins also play a role in the control of integrity of intercellular junction proteins.  We studied membrane fluidity on the EC luminal surface as another potential source of mechano-sensing.  Besides shear stress, we are investigating the mechanism by which mechanical strain affects molecular and cellular events. We found that equibiaxial strain with high magnitude (>25%) induced SMC apoptosis (programmed cell death), whereas lesser strain within physiological range (<10%) had a protective effect. SMCs subjected to uniaxial strain aligned in the direction perpendicular to the strain vector, and the induced JNK expression was transient. In contrast, SMCs subjected to equibiaxial strain did not undergo any alignment, and the induced JNK expression was sustained. Thus, uniaxial and equibiaxial strains have different effects on cell morphologies and cell signaling. We are currently investigating the molecular mechanisms causing such differential cellular responses to different modes of mechanical stimulation.

Endothelial Cell Migration: The migration of vascular cells plays an important role in many physiological processes. We used a wound-healing model to investigate the mechanical and molecular mechanisms regulating EC migration by scraping a strip in an EC monolayer and subsequently exposing this monolayer to flow. EC strip wounding induced the dissociation of cell junctions at wound edges. Under laminar flow, more of the cells upstream to the wound lost their cell junctions, became dissociated from EC monolayer, and migrated with a great speed into the wound area along the flow direction, as compared with static cells. The results suggest that laminar flow promotes EC migration through the activation of signaling pathways, in addition to a mechanical pushing effect.  In order to dissect out the various factors, we studied the migration of subconfluent ECs, which do not have cell-cell adhesions. The migration speed of cells seeded on different concentrations of extracellular matrix (ECM) proteins showed a bell-shaped curve, with a peak at an optimal ECM concentration, indicating that a balance between the attachment at the cell front and detachment at the cell rear is important for cell migration. By studying EC migration on ECM proteins seeded on top of an elastic gel embedded with fluorescently labeled beads, we are mapping the mechanical strain induced in the membrane and the force exerted by the migrating cell. The role of Rho small GTPases, which play significant roles in regulating the cytoskeleton (especially actin filaments), in cell migration is also being investigated. The dominant negative mutants of both RhoA and Rac significantly retarded EC migration and reduced the wound-healing rate. Microtubules, as one of the components of cytoskeleton, also play an important role in EC migration, as evidenced by the inhibition of migration with Taxol, a microtubule-stabilizing agent.

In-vivo Investigations: We applied our knowledge from in vitro studies to in vivo animal models. We found that a dominant negative mutant of Ras (Ras N17) blocked the re-stenosis induced by balloon injury of rat arteries, suggesting a potential role of Ras in gene therapy for the prevention of re-stenosis after balloon injury. Investigations on signaling pathways in vivo are being conducted in animal models of balloon injury and hypertension, which is created by banding of the abdominal aorta. The effects on cellular signaling and gene expression are correlated with physiological changes.

In summary, by using a systematic approach to study the effects of mechanical stimuli on mechanosensing, signal transduction, gene expression, cellular dynamics, and physiological functions, we aim at elucidating the molecular basis of mechanotransduction in health and disease.

 

See our publications online!