Discipline: Technology and Engineering
Subcategory: Biomedical Engineering
Farrah-Amoy Fullerton - Mary Baldwin University
Co-Author(s): Jasqueline Staton, Louisiana State University, Shreveport, Louisiana
Recently, decellularized scaffolds derived from native tissues have gained significant attentions in the field of tissue engineering. The advantages of the decellularization approach include the efficiency of removing all cellular and nuclear content and the preservation of extracellular matrix (ECM), vascular channels, and many biologically active matrix components. In this project, we focused on qualitatively assessing the capability of the detergent based decellularization method to effectively remove cells from the native porcine livers as well as preserve the ECM structures as compared to their native counterparts. Retention of protein structure is of substantial importance in tissue scaffolds because of its close relation to the microenvironment of the new tissue and, subsequently, its function. The primary solution used to decellularize the 3.75 cm3 porcine liver samples was 2.5% Triton™ X-100 with 0.25% NH4OH. These decellularized samples, in addition to native porcine liver tissue samples, were manually fixed, dehydrated, infiltrated with & embedded into paraffin, and mounted to slides for microscopic analysis. Immunofluorescence microscopy allowed the slides to be stained for the main components of the extracellular matrix: collagen, fibronectin, and laminin. Cell nuclei were made visible with DAPI. Hematoxylin and eosin (H&E) staining of decellularized livers showed that large spaces once occupied by hepatocytes were turned out, leaving behind an intact ECM. Immunohistochemistry staining of the liver ECM showed the preservation of collagen, fibronectin, and laminin when compared with the native liver. These protein-rich scaffolds have promising applications in the field of tissue engineering, as they function as suitable microenvironments for developing cells.
Funder Acknowledgement(s): This work was supported by NSF-REU (DBI-1358923) to Komal Vig (PI) and by NSF- CREST (HRD-1241701) to Shree S. Singh (PI).
Faculty Advisor: Bo Wang, firstname.lastname@example.org
Role: My role was to find viable porcine liver tissue samples, decellularize these samples, histologically process them, and use various staining procedures to visualize the ECM proteins, vascular channels, and cell nuclei. Immunofluorescence microscopy revealed retention of all ECM proteins and DAPI staining of cell nuclei confirmed removal of cells in decellularized samples. Hematoxylin and eosin staining of these samples showed the preservation of vascular channels and other functional structures. With these results, it was shown that the decellularization process used was efficient in removing native cells while retaining the structure and proteins of the tissue.