Biomechanical Regulation of Endothelium-dependent Events Critical for Adaptive Remodeling

Abstract

Alterations in hemodynamic shear stress acting on the vascular endothelium are critical for adaptive arterial remodeling. The molecular mechanisms regulating this process, however, remain largely uncharacterized. Here, we sought to define the responses evoked in endothelial cells exposed to shear stress waveforms characteristic of coronary collateral vessels and the subsequent paracrine effects on smooth muscle cells. A lumped parameter model of the human coronary collateral circulation was used to simulate normal and adaptive remodeling coronary collateral shear stress waveforms. These waveforms were then applied to cultured human endothelial cells (EC), and the resulting differences in EC gene expression were assessed by genome-wide transcriptional profiling to identify genes distinctly regulated by collateral flow. Analysis of these transcriptional programs identified several genes to be differentially regulated by collateral flow, including genes important for endothelium-smooth muscle interactions. In particular, the transcription factor KLF2 was up-regulated by the adaptive remodeling coronary collateral waveform, and several of its downstream targets displayed the expected modulation, including the down-regulation of connective tissue growth factor. To assess the effect of endothelial KLF2 expression on smooth muscle cell migration, a three-dimensional microfluidic assay was developed. Using this three-dimensional system, we showed that KLF2-expressing EC co-cultured with SMC significantly reduce SMC migration compared with control EC and that this reduction can be rescued by the addition of exogenous connective tissue growth factor. Collectively, these results demonstrate that collateral flow evokes distinct EC gene expression profiles and functional phenotypes that subsequently influence vascular events important for adaptive remodeling.Coronary collateralization or arteriogenesis is a vascular adaptive remodeling process that occurs in the context of arterial occlusion when preexisting collateral arterioles remodel to form larger diameter bypass arteries. The current arteriogenesis paradigm implicates local hemodynamics, recruitment/activation of monocytes, and structural remodeling of the vascular wall as key steps in collateralization (1). Two primary stages of collateralization have been described based predominantly on observations from femoral artery ligation models that induce collateral remodeling via an instantaneous increase in collateral flow. The initial stage of collateralization is characterized by a wave of monocyte-driven inflammation and rapid vasodilation, an immediate compensatory response to increased collateral flow. In the second phase of collateralization, the initial inflammatory response is resolved, and the newly formed conductance artery wall is stabilized (2). To date, considerable attention has been dedicated to investigating the inflammatory regulation of this process (i.e. MCP-1 and GM-CSF), but despite evidence on the importance of the endothelium and endothelium-derived ...

Molecular Biology of Atherosclerosis

“…Numerous studies have pointed to the induction of the transcription factor KLF2 (with KLF4 more recently added) as a critical step in the conversion to and maintainance of an athero-protective state (1106,1332,1881), even in microvessels (330). Indeed, KLF2 (and possibly KLF4) and Nrf2 are thought to act together to control as many as 70% of the genes upregulated by laminar flow (1291).…”

Biomechanical regulation of vascular smooth muscle cell functions: from in vitro to in vivo understanding

“…A three-dimensional in vitro perfusion system allows direct investigation of shear stress effects on the development of microvasculature in vitro [102]. A new type of 'on chip and in situ' micropatterning technique has been developed [103][104][105] and this technique is even shown to control single cell co-cultures. Single cell co-cultures are in close proximity to the formation of connexion structures and the study of contact modes of communication [106].…”

“…Shear stress also delays the migration of either cell type in a dose-dependent manner in an EC-VSMC co-cultured microsystem [104]. In a three-dimensional EC-VSMC co-cultured system, ECs with a high expression of Krü ppel-like factor 2 significantly reduce VSMC migration [103]. Sustained pulsatile flow regulates endothelial NO synthase and cyclooxygenase expression in co-cultured vascular ECs and VSMCs [126], and NO decreases VSMC proliferation and increases apoptosis [125,127].…”

Development and Applications of Microfluidic Devices for Cell Culture in Cell Biology

“…The two-independent-channel types, which separated by a gel/scaffold compartment, has been used often to control [31] or study the influence of shear stress on cultured cells. The microfluidics shows advantages for allowing the flow to be controlled precisely in the channels as well as across the gel compartment [32,33]. Both micro environmental components and intricate architecture of the micro-vasculature networks strongly influence the fluid shear stress.…”