Microfluidic endothelial cell culture model to replicate disturbed flow conditions seen in atherosclerosis susceptible regions

Abstract

Atherosclerotic lesions occur non-randomly at vascular niches in bends and bifurcations where fluid flow can be characterized as "disturbed" (low shear stress with both forward and retrograde flow). Endothelial cells (ECs) at these locations experience significantly lower average shear stress without change in the levels of pressure or strain, which affects the local balance in mechanical stresses. Common in vitro models of atherosclerosis focus primarily on shear stress without accounting for pressure and strain loading. To overcome this limitation, we used our microfluidic endothelial cell culture model (ECCM) to achieve accurate replication of pressure, strain, and shear stress waveforms associated with both normal flow seen in straight sections of arteries and disturbed flow seen in the abdominal aorta in the infrarenal segment at the wall distal to the inferior mesenteric artery (IMA), which is associated with high incidence of atherosclerotic lesion formation. Human aortic endothelial cells (HAECs) were cultured within the ECCM under both normal and disturbed flow and evaluated for cell shape, cytoskeletal alignment, endothelial barrier function, and inflammation using immunofluorescence microscopy and flow cytometry. Results clearly demonstrate quantifiable differences between cells cultured under disturbed flow conditions, which are cuboidal with short and randomly oriented actin microfilaments and show intermittent expression of b-Catenin and cells cultured under normal flow. However, in the absence of pro-inflammatory stimulation, the levels of expression of activation markers: intra cellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), platelet endothelial cell adhesion molecule-1 (PECAM-1), and vascular endothelial cell growth factor -receptor 2 (VEGF-R2) known to be involved in the initiation of plaque formation were only slightly higher in HAECs cultured under disturbed flow in comparison to cells cultured under normal flow.

Targeting cell adhesion molecules with nanoparticles usingin vivoand flow-basedin vitromodels of atherosclerosis

“…72 By exposing HAECs to these types of flow, the authors concluded that in the absence of pro-inflammatory stimulation, PECAM-1, VCAM-1, and ICAM-1 were not significantly upregulated by flow condition. Cells cultured under disturbed flow exhibited round shape and random orientation.…”

“…Disturbed and non-laminar flow below healthy physiological levels of shear stress (10-70 dyn/cm 2 ) can cause EC phenotype changes resulting in pro-inflammatory endothelium. [70][71][72] Here, we review in vitro models developed for determining the role of disturbed flow patterns on CAM expression and leukocyte recruitment, and conclude with the testing of targeted particles against CAMs in such models.…”

Preface to Special Topic: Microsystems for manipulation and analysis of living cells

“…8 Endothelial cells form an inside lining of blood vessels and are important participants in plaque formation. Function of these cells may be affected by the disturbed flow patterns associated with partially occluded blood vessels.…”

“…1,2 This special topics section provides a sample of the types of microsystems being developed for cell analysis and manipulation. The research directions covered in this thematic section include sorting of mammalian cells, 3,4 pairing of mammalian cells, 5 sorting of viral particles, 6 concentration of bacteria, 7 cultivation of cells in microfluidic devices, 8,9 on-chip monitoring of cell function, 10 and on-chip flow cytometry.…”

Mimicking arterial thrombosis in a 3D-printed microfluidic in vitro vascular model based on computed tomography angiography data

“…This type of 3D complexity is essential to accurately mimic the in vivo flow patterns which are fundamental in the mechanism of thrombosis. 14,25,26 Three-dimensional printing has the potential to revolutionize the field of microfluidics by allowing to overcome the limitations posed by conventional 2D wafer-based microfabrication techniques. 22 Furthermore, it allows the fabrication of fully 3D structures, which closely mimic the natural shape of complex human tissue structures and could, therefore, also be useful in replicating the architecture of human blood vessels.…”