Date of Award
Tissue engineering, Biomaterials, Regenerative
There is deficit in the number of human livers that are available for transplantation. Additionally, there is no reliable model for the liver environment available for laboratory research. Tissue engineering provides alternatives for organ transplantation. We have created a supramolecular biomaterial out of simple peptide and saccharide building blocks to act as a hydrogel scaffold to support culture of liver cells. Using a self-‐assembling peptide fiber decorated with functional molecules, we were able to culture primary human hepatocytes demonstrate their viability and maintenance of phenotype. The hydrogel material was formed by co-assembly in aqueous buffer and characterized using transmission electron microscopy (TEM), fluorescence analysis, and rheology. TEM was employed to determine the nanoscale structure of the material. In order to verify that the peptide-‐based structural fiber, cell adhesion peptide, and amino sugar components of the gel were integrating together to form the fibers, we used fluorescence analysis; this revealed that the components were assembled. Rheology revealed that the stiffness of the most successful of the three gels tested is softer than that of healthy liver tissue, indicating that the cells are not receiving all of the native environmental signals. Light microscopy was used in order to determine cell binding and live/dead cell staining was performed to indicate cell viability. An XTT assay provided a quantification of cell health based on metabolic activity and showed survivorship of the hepatocytes cultured in the customized hydrogel. We used an enzyme-‐linked immunosorbent assay of cell culture supernatant (ELISA) to verify that the hepatocytes were producing the characteristic proteins that provide anindication of hepatocyte viability. Analysis of RNA levels combined with 5 quantitative polymerase chain reaction (qPCR) demonstrated an increase in gene expression for hepatocyte-‐specific markers.
MacPherson, Douglas, "Synthetic Functionalized Scaffolds for Culture and Maintenance of Primary Human Hepatocytes" (2016). CUNY Academic Works.