Self‐supporting nanoporous alumina membranes as substrates for hepatic cell cultures

Abstract

Membranes made from nanoporous alumina exhibit interesting properties for their use in biomedical research. They show high porosity and the pore diameters can be easily adjusted in a reproducible manner. Nanoporous alumina membranes are thus ideal substrates for the cultivation of polar cells (e.g., hepatocytes) or the establishment of indirect co-cultures. The porous nature of the material allows supply of nutrients to both sides of adherent cells and the exchange of molecules across the membrane. However, it is well-known that surface features in the nanometer range affect cellular behavior. In this study, the response of HepG2 cells to nanoporous alumina membranes with three different pore diameters, ranging from 50 to 250 nm, has been evaluated. The cellular interactions with the nanoporous materials were assessed by investigating cell adhesion, morphology, and proliferation. Cell functionality was measured by means of albumin production. The membranes supported good cell adhesion and spreading. Compared to tissue culture plastic, the cells on the porous substrates developed distinct focal adhesion sites and actin stress fibers. Additionally, electron microscopical investigations revealed the penetration of cellular extensions into pores with diameters bigger than 200 nm. Furthermore, cell proliferation significantly increased with an increase in pore diameter, whereas the albumin production followed a reverse trend. Thus, it seems to be possible to direct cellular behavior of HepG2 cells growing on nanoporous alumina by changing the pore diameter of the material. Hence, nanoporous alumina membranes can be useful culture substrates to develop new approaches in the field of liver tissue engineering.

Mesoporous alumina as a biomaterial for biomedical applications

“…Cell proliferation increased with an increase in pore diameter and was highest on substrates with 200 nm pores. In contrast, cell functionality was fostered on membranes with small pore diameters of around 50 nm [21]. These results agree with the results obtained using 50-150 nm PAA and NIH-3T3 fibroblast cells [22], 20-200 nm pore size alumina and human osteoblast-like MG63 cells [23], and 75-300 nm pore size alumina and NIH 3T3 cells [24].…”

Cytotoxicity of cultured macrophages exposed to antimicrobial zinc oxide (ZnO) coatings on nanoporous aluminum oxide membranes

“…The ZnO-coated membranes examined in this study were previously shown to possess antibacterial activity; 16 however, the cytotoxicity of these materials was not previously examined. It is evident from the MTT cell viability assay results that the ZnO-coated membranes are cytotoxic to macrophages after 48 h of incubation.…”

“…Previous studies have indicated that the pore size of nanoporous anodized aluminum oxide membranes and coated membranes does not have a significant effect on cell viability 24 . As noted in the introduction, our group quantified Zn ion release from 20 nm and 100 nm anodized aluminum oxide membranes with atomic layer-deposition grown 8 nm-thick ZnO coatings into DMEM culture media containing 10% fetal bovine serum by means of inductively coupled plasma mass spectrometry 16 . This study confirms that varying the pore size between 20 and 200 nm does not alter cell viability as measured by the MTT assay.…”

“…They showed that membranes with ALD-grown ZnO coatings leach Zn ions in vitro. The extract from the ZnO-coated 20 nm and 100 nm anodized aluminum oxide membranes contained Zn at a concentration of approximately 90 µg/ml 16 . This study raised the question about the cytotoxic potential of the released Zn ions to eukaryotic cells.…”

“…The underlying inert substrate used in the study was anodized aluminum oxide, which is known to have good cell adhesion and proliferation in vitro compared with a standard tissue culture well. Hoess et al showed that cells interact with 200 nm pores and thus may exhibit different responses to membranes with dissimilar pore diameters 16 . Ferraz et al indicated that differences in nanoporosity elicited dissimilar immune responses for 20 and 200 nm diameter anodized aluminum oxide in vivo 17 .…”