Devlopment of a Cell Coculture Microfluidic Shear Device for Mechano-Transmission Study

We have developed a microfluidic shear device that allows for the study of cell communication in a dynamically controlled biochemical and biomechanical environments simulating cells’ living environments in vivo. Such study may help to improve our understanding in the effects of hypertension-relevant and vascular development-relevant flow shear stress on cell behaviors. Endothelial cells may be a key factor for transmitting the blood flow conditions from the endothelial lining to interstitial layers and smooth muscle cells. The interstitial flow stress and the shear stress induced signaling factors may greatly alter vascular biology of these deep layers. Endothelial cells act as a mechano-transducer by converting shear stress into biochemical signaling factors. The biochemical factors diffuse to smooth muscle cells and further alter the biological structure of vascular tissues. Also, the flow shear stress will be transmitted to the interstitial tissue layer through the pores resulted from the pores in the fenestrated endothelial lining. Studies in both the mechano-transduction process and the mechano-transmission process will benefit from a biomimetic flow shear device with co-cultured cells. Our device will allow the co-culture of endothelial cells and smooth muscle cells to study these biomechanical processes. The pulmonary arterial cells are used as a model in the study. The microfluidic device developed here will be used to enhance the understanding of pulmonary vascular disease pathogenesis due to the variations in the flow shear stress.

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Flow shear stress enhances intracellular Ca2+ signaling in pulmonary artery smooth muscle cells from patients with pulmonary arterial hypertension

AJP Cell Physiology ◽

10.1152/ajpcell.00115.2014 ◽

2014 ◽

Vol 307 (4) ◽

pp. C373-C383 ◽

Cited By ~ 33

Author(s):

Shanshan Song ◽

Aya Yamamura ◽

Hisao Yamamura ◽

Ramon J. Ayon ◽

Kimberly A. Smith ◽

...

Keyword(s):

Blood Flow ◽

Shear Stress ◽

Smooth Muscle ◽

Pulmonary Arterial Hypertension ◽

Smooth Muscle Cells ◽

Muscle Cells ◽

Flow Shear ◽

Pulmonary Vasoconstriction ◽

Pulmonary Arterial ◽

Flow Shear Stress

An increase in cytosolic Ca2+ concentration ([Ca2+]cyt) in pulmonary arterial smooth muscle cells (PASMC) is a major trigger for pulmonary vasoconstriction and an important stimulus for pulmonary arterial medial hypertrophy in patients with idiopathic pulmonary arterial hypertension (IPAH). Vascular smooth muscle cells (SMC) sense the blood flow shear stress through interstitial fluid driven by pressure or direct exposure to blood flow in case of endothelial injury. Mechanical stimulus can increase [Ca2+]cyt. Here we report that flow shear stress raised [Ca2+]cyt in PASMC, while the shear stress-mediated rise in [Ca2+]cyt and the protein expression level of TRPM7 and TRPV4 channels were significantly greater in IPAH-PASMC than in normal PASMC. Blockade of TRPM7 by 2-APB or TRPV4 by Ruthenium red inhibited shear stress-induced rise in [Ca2+]cyt in normal and IPAH-PASMC, while activation of TRPM7 by bradykinin or TRPV4 by 4αPDD induced greater increase in [Ca2+]cyt in IPAH-PASMC than in normal PASMC. The bradykinin-mediated activation of TRPM7 also led to a greater increase in [Mg2+]cyt in IPAH-PASMC than in normal PASMC. Knockdown of TRPM7 and TRPV4 by siRNA significantly attenuated the shear stress-mediated [Ca2+]cyt increases in normal and IPAH-PASMC. In conclusion, upregulated mechanosensitive channels (e.g., TRPM7, TRPV4, TRPC6) contribute to the enhanced [Ca2+]cyt increase induced by shear stress in PASMC from IPAH patients. Blockade of the mechanosensitive cation channels may represent a novel therapeutic approach for relieving elevated [Ca2+]cyt in PASMC and thereby inhibiting sustained pulmonary vasoconstriction and pulmonary vascular remodeling in patients with IPAH.

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Surface Engineering in Microfluidic Devices for the Isolation of Smooth Muscle Cells and Endothelial Cells

MRS Proceedings ◽

10.1557/proc-1004-p01-03 ◽

2007 ◽

Vol 1004 ◽

Author(s):

Shashi Murthy ◽

Brian Plouffe ◽

Milica Radisic

Keyword(s):

Tissue Engineering ◽

Endothelial Cells ◽

Shear Stress ◽

Smooth Muscle ◽

Cell Adhesion ◽

Smooth Muscle Cells ◽

Cell Sorting ◽

Microfluidic Devices ◽

Muscle Cells ◽

Small Sample

AbstractMicrofluidic cell separation systems have emerged as attractive alternatives to traditional techniques in recent years. These systems offer the advantages of being able to handle small sample volumes and at the same time achieve highly selective separation. Conventional separation techniques, including both fluorescence-activated cell sorting (FACS) and magnetic-activated cell sorting (MACS), typically require a pre-processing incubation step to attach ligated tags (such as fluorescent dyes or magnetic beads) to cell surfaces prior to separation. These techniques are also constrained by infrastructure and high cost. Microfluidic devices with surface-immobilized adhesion molecules eliminate the need for pre-processing incubation and are a low cost alternative.We describe the selective adhesion of smooth muscle cells and endothelial cells in microfluidic devices coated with adhesion peptides. The device geometry is such that the shear stress varies linearly as a function of flow channel length, allowing simultaneous evaluation of the effects of surface chemistry and fluid shear on cell adhesion. The adhesion peptides, val-ala-pro-gly (VAPG) and arg-glu-asp-val (REDV), are known to bind selectively to smooth muscle cells and endothelial cells, respectively. These peptides were tethered to the device surface using silane chemistry and NHS-ester coupling. Cell adhesion was examined in a shear stress range of 1.3-4.0 dyn/cm2. Under these conditions, endothelial cells show significantly higher adhesion to REDV-coated devices compared to smooth muscle cells and fibroblasts. Correspondingly, smooth muscle cell adhesion in VAPG-coated devices is much greater than that of endothelial cells and fibroblasts. This selective binding behavior is also observed when mixed suspensions of the three cell types are flowed into both types of peptide-coated microfluidic devices. These results suggest that microfluidic devices coated with REDV and VAPG can be used as effective separation tools in various applications, such as tissue engineering. Specific examples of applications in cardiac and skin tissue engineering will be discussed.

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