Cilia Have a Significant Role in Regulating Cell Size in Response to Fluid Flow Induced Shear Stress in a Flow Chamber

Cilia are hair-like protrusions on the apical surface of cells. Their function is to relay mechanical signals like shear stress from extracellular into intracellular environment and thereby maintain cellular homeostasis. Ciliary dysfunctions include polycystic kidney disease and new therapeutic interventions based on ciliary function are under investigation. The current study evaluates the use of a custom designed fluid flow chamber’s ability to study the role of cilia in regulating cell size in response to shear stress. A fluid flow chamber that continually maintains laminar flow at different flow rates and temperature was designed. Endothelial wild type cells (ETWT) that have cilia and polycystic kidney disease cells (PKD) that lost their ciliary function are grown on different glass slides. Cells on each glass slide are then exposed to continuous flow of phosphate-buffered saline at 37oC in the flow chamber. The optimal flow rate and duration of flow were first determined by measuring the total protein concentration before and after exposing the cells. Cell radius and area before and after exposing them to flow are measured using the NIS Software available on the microscope. The results from protein concentrations (n=12) indicate that cells are still attached at normal physiological flow rate 467 mL/min (2.8 µg/µL) and did not significantly differ from 60 mL/min (4.08 µg/µL) or 600 mL/min (2.73 µg/µL). The results for duration of fluid flow (n=22) show that 60 minutes (0.09 + 0.01 µg/µL) is optimal compared to 120 minutes (0.06 + 0.01 µg/µL) or 180 minutes (0.10 + 0.02 µg/µL). Under these optimal conditions, the average area of ETWT cells (n=300) measured from different slides before and after the flow is 4420.81+ 67.40 µm2 and 4678.17 + 87.15 µm2 (n=200) respectively. For PKD cells, the average area before and after the flow (n=300) is 5682.46 + 105.48 µm2 and 4173.74 + 263.97 µm2 (n=250). These results are in agreement with the published literature on the ability of cilia to maintain cell size in ETWT cells in response to shear stress that is similar to normal blood flow. However, under similar conditions, PKD cells could not maintain their cell size as the mechano-chemical signaling pathway that communicates external signals to prepare appropriate intracellular response is disrupted. These results provide confirmation that the custom designed parallel plate fluid flow chamber is a reliable tool to investigate the specific targets in the mechano-chemical cell signaling pathways