We have demonstrated that 3D spatial patterning of a relatively bland material such as gelatin can influence the function of hepatocytes seeded within scaffold pores by varying a simple feature such as scaffold geometry. The increased expression of importer proteins (SLCO1B3) and canalicular exporter proteins (ABCC2) in 2D, and a lack of additional canalicular spaces, is additional evidence for the discrepancy between expression and function, and is likely a manifestation of the sensitivity of HUH7 cells and hepatocytes in general to 3D culture. Using confocal microscopy and image analysis, we observed a significantly higher amount of canaliculus formation in the pores of more tortuous 60⁰ geometries. To test this, we cultured hepatocytes in the presence of a fluorescent bile salt, cholyl-lysyl-fluorescein (CLF), which localizes to bile canaliculi (Figure 2B). We hypothesized that this was due to the structure of in vivo hepatocytes, and that a 3D environment is necessary for normal hepatocyte polarization and membrane trafficking. However, 2D controls began to demonstrate a disparity between gene expression and assayed function of cytochrome P450 enzymes (Figure 2A). Evaluation of albumin secretion and gene expression indicate superior hepatocyte-function in 60⁰ geometries (Figure 1C). 2D coatings of gelatin were used as a control. When seeded on 3D-printed scaffolds of two different geometries (60⁰ and with 90⁰ strut orientations) an undifferentiated hepatocyte cell line (HUH7) demonstrated high viability (Figure 1B). In this study, we show the ability to precisely control pore geometry of 3D-printed gelatin scaffolds (Figure 1A). However, the effect of differing geometries, while controlling for pore size, has yet to be investigated in the context of hepatocyte function. The creation of uniform and geometrically repetitive tissue scaffolds can allow for the control over cellular aggregation and nutrient diffusion. Three dimensional (3D) printing is a new method amenable to the fabrication of TE organs of a repetitive microstructure such as the liver. Tissue engineering (TE) holds promise to mitigate the transplant shortage by providing transplantable tissues and organs, however traditional TE methods have yet to produce significant clinical results. Transplantation is unfortunately severely limited by the deficit of available donor organs. Third Floor, Feinberg Pavilion, Northwestern Memorial Hospital B174 - Basic Science 3D-Printed gelatin scaffold geometry modulates hepatocyte function and gene expressionĬurrently the only viable treatment for end stage liver disease is orthotropic liver transplantation. Liver, Tissue Engineering, 3D-Printing, Regenerative Medicine, Scaffold Location:
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