Heparin-based hydrogel as a matrix for encapsulation and cultivation of primary hepatocytes

Abstract

Primary hepatocytes are commonly used as liver surrogates in toxicology and tissue engineering fields, therefore, maintenance of functional hepatocytes in vitro is an important topic of investigation. This paper sought to characterize heparin-based hydrogel as a three-dimensional scaffold for hepatocyte culture. The primary rat hepatocytes were mixed with a prepolymer solution comprised of thiolated heparin and acrylated poly(ethylene glycol) (PEG). Raising the temperature from 25° to 37°C initiated Michael addition reaction between the thiol and acrylated moieties and resulted in formation of hydrogel with entrapped cells. Analysis of liver-specific products, albumin and urea, revealed that the heparin hydrogel was non-cytotoxic to cells and, in fact, promoted hepatic function. Hepatocytes entrapped in the heparin-based hydrogel maintained high levels of albumin and urea synthesis after three weeks in culture. Because heparin is known to bind growth factors, we incorporated hepatocyte growth factor (HGF) -an important liver signaling molecule -into the hydrogel. HGF release from heparin hydrogel matrix was analyzed using enzyme linked immunoassay (ELISA) and was shown to occur in a controlled manner with only 40% of GF molecules released after 30 days in culture. Importantly, hepatocytes cultured within HGF-containing hydrogels exhibited significantly higher levels of albumin and urea synthesis compared to cells cultured in the hydrogel alone. Overall, heparin-based hydrogel showed to be a promising matrix for encapsulation and maintenance of difficult-to-culture primary hepatocytes. In the future, we envision employing heparin-based hyrogels as matrices for in vitro differentiation of hepatocytes or stem cells and as vehicles for transplantation of these cells.

Xenobiotic Metabolism and Disposition in Human Lung Cell Models: Comparison with In Vivo Expression Profiles

“…However, our results indicate that primary airway epithelial cell cultures seem to be the most relevant experimental model because they displayed the highest homology in expression pattern and the lowest number of dysregulated genes compared with BM. Improvements of culture conditions, as previously discussed for primary hepatocytes with the change of extracellular matrix or the use of new three-dimensional in vitro models (Kim et al, 2010;Lang et al, 2011), could allow primary airway epithelial cells to present gene expression profiles that accurately reflect those of bronchial tissue, and to be used with full confidence for toxicological and pharmacological studies. It should be noted that the interindividual variability of expression observed for some genes in primary cells may affect the reproducibility of experiments.…”

Introduction to cell–hydrogel mechanosensing

“…These types of hydrogel are particularly useful in examining cellular behaviour owing to their biocompatibility, biodegradability and biofunctionality [68]. cornea agarose-fibrin [29] alginate-gelatin nanofibre [163] collagen [146,149,150] collagen-PLGLA nanofibre [160] alginate [34] cellulose [35] heparin [36] alginate-hyaluronic acid [33] collagen [190] collagen-chitosan [32] dextran [30,31] hyaluronic acid [44] alginate [27] collagen [23,24] dextran [25] gelatin [26] collagen [17,97] fibrin [37] agarose [51,175,179,196] alginate [126] chitosan [20,50] fibrin [42,51] gellan gum [22,51] hyaluronic acid [110] PEG [60] A key factor when examining cell-material mechano-interactions is the mechanisms of cell adhesion to the hydrogel. Cell surface receptors such as integrins bind to ligands within the hydrogel if such adhesion sites exist.…”

“…This remodelling behaviour is influenced by several factors, in particular, the type of hydrogel, the type of cell and the absence or presence of biochemical and mechanical stimuli. To date, cell-seeded hydrogels have been under investigation to repair or replace several tissue types, including cartilage [20][21][22], skin [23][24][25][26][27], cornea [28,29] and vascular tissues [30][31][32][33] among other tissues [34][35][36][37] (figure 1). As this field has grown, it has become evident that a greater understanding of the cell-material interactions in hydrogel systems is required to properly explain the mechanical mechanism that cells use to remodel their surroundings and to exploit this behaviour to develop functional tissues and tissue repair strategies.…”

Characterization of a Liver Organoid Tissue Composed of Hepatocytes and Fibroblasts in Dense Collagen Fibrils

“…Primary hepatocytes have been cultured on biomaterials in vitro, for example, collagen gels, [3][4][5][6] Engelbreth-Holm-Swarm gels, [7][8][9] and other materials. [10][11][12][13] There are also some recent studies using collagen sandwich hepatocyte cultures that have achieved long culture periods and the maintenance of hepatic functions. [14][15][16] However, no studies have described the reconstruction of highly concentrated connective tissue in vitro with properties similar to endogenous liver tissue.…”