Phenotype Transformation of Aortic Valve Interstitial Cells Due to Applied Shear Stresses Within a Microfluidic Chip XINMEI WANG, 1 JOOHYUNG LEE, 2 MIR ALI, 1 JUNGKYU KIM, 2 and CARLA M. R. LACERDA 1,3 1 Department of Chemical Engineering, Texas Tech University, Lubbock, TX 79409, USA; 2 Department of Mechanical Engineering, Texas Tech University, Lubbock, TX 79409, USA; and 3 Department of Chemical Engineering, Texas Tech University, 6th and Canton Avenue, Box 43121, Lubbock, TX 79409-3121, USA (Received 2 May 2017; accepted 8 June 2017) Associate Editor Umberto Morbiducci oversaw the review of this article. Abstract—Despite valvular heart diseases constituting a sig- nificant medical problem, the acquisition of information describing their pathophysiology remains difficult. Due to valvular size, role and location within the body, there is a need for in vitro systems that can recapitulate disease onset and progression. This study combines the development of an in vitro model and its application in the mechanical stimulation of valvular cell transformation. Specifically, porcine aortic valvular interstitial cells (PAVIC) were cultured on poly- dimethylsiloxane microfluidic devices with or without expo- sure to shear stresses. Mechanobiological responses of valvular interstitial cells were evaluated at shear stresses ranging from 0 to 4.26 dyn/cm 2 . When flow rates were higher than 0.78 dyn/ cm 2 , cells elongated and aligned with the flow direction. In addition, we found that shear stress enhanced the formation of focal adhesions and up-regulated PAVIC transformation, assessed by increased expression of a-smooth muscle actin and transforming growth factor b. This study reveals a link between the action of shear forces, cell phenotype transfor- mation and focal adhesion formation. This constitutes the first step towards the development of co-cultures (interstitial- endothelial cells) on organ-on-a-chip devices, which will enable studies of the signaling pathways regulating force- induced valvular degeneration in microtissues and potential discovery of valvular degeneration therapies. Keywords—Heart valve degeneration, Microfluidic in vitro model, Organ-on-a-chip, Mechanotransduction. INTRODUCTION According to the American Heart Association, the prevalence of valvular heart diseases in the US is 2.5% and this rate increases significantly with age. 36 Recent studies of in vitro mechanical stimulation of valvular cells and tissues showed that abnormal mechanical stresses may contribute to the initiation and progres- sion of valvular heart disease. 4,13,28 It is increasingly accepted that cyclic strains alter aortic valvular inter- stitial cell (VIC) orientation and extracellular matrix (ECM) fiber alignment, enhance cell proliferation and apoptosis, and regulate cell phenotype transforma- tion. 19,41 In addition, mechanical stresses have been shown to induce ECM remodeling and cell phenotype transformation in other cardiac valves. 20,29,54 At the tissue level, shear stresses are known to in- duce ECM remodeling in aortic valves by modifying activity and expression levels of collagenases and gelatinases. 41 Even though both tensile and shear stresses result in pathological changes within heart valve tissues, in vitro shear stresses seem to promote an overall increase in ECM synthesis. 41 Other than impacting the ECM, increased shear stresses generated by altered hemodynamic conditions can be linked to calcific degeneration of aortic valves 57 and present variable effects on each valve side. 6,56 In shear stress- induced valvular degeneration, interstitial cells are responsible for propagating signals of ECM remodel- ing, largely without disruption of the endothelial cell lining. 18,46,47 These changes induced by abnormal mechanical stresses in tissues are reproducible in vitro and consistent with long-term disease development. Thus far, at the cell level, an extensive body of shear stress work focuses on valvular endothelial cells, 10,11 due to their location and function in vivo. It has been previously demonstrated that valvular endothelial cells align perpendicular to the direction of shear force application, while vascular endothelial cells align in parallel. 12 Mechanical forces have been shown to alter valvular endothelial cell morphology and expression Address correspondence to Carla M. R. Lacerda, Department of Chemical Engineering, Texas Tech University, 6th and Canton Avenue, Box 43121, Lubbock, TX 79409-3121, USA. Electronic mail: carla.lacerda@ttu.edu Annals of Biomedical Engineering (Ó 2017) DOI: 10.1007/s10439-017-1871-z Ó 2017 Biomedical Engineering Society