Nanomedicine, Volume IIA: Biocompatibility
© 2003 Robert A. Freitas Jr. All Rights Reserved.
Robert A. Freitas Jr., Nanomedicine, Volume IIA: Biocompatibility, Landes Bioscience, Georgetown, TX, 2003
220.127.116.11 Cell Response to Bulk Alumina and Sapphire
A number of experiments have been performed to determine the response of fibroblasts to alumina and sapphire surfaces. For example, alumina ceramic surface has shown excellent in vitro biocompatibility in a tissue culture of rabbit fibroblasts  and cultured embryonic mouse fibroblasts . Colony-forming Chinese hamster V79 fibroblast cells proliferated equally well on alumina ceramic and control surfaces . Fibroblast-like mesenchymal cell populations cultured on solid alumina ceramic surfaces induced no cytotoxic or antiproliferative effects on monolayer populations in vitro, leading the researchers  to conclude that “the aluminum oxide ceramic presents itself as an absolutely bioinert material.” A scanning electron microscopic study  was conducted on the adhesion, spreading and formation of confluent cell monolayers from fibroblasts and epithelioid cells on Al2O3 ceramics. The study found that the cells adhered, spread, migrated and proliferated on the surfaces tested, leading to the conclusion that this implant material is compatible with cells . In general, such cells adhere well to single-crystal or polycrystalline alumina . Experiments by Mawn et al  found that human orbital fibroblasts grown on alumina bioceramic implant were free of debris and had the largest cell count, whereas cells grown on hydroxyapatite or porous polyethylene implants had cellular debris associated with them.
The response of bone cells to alumina ceramic has also been examined. For instance, the nature of the contact sites (including focal contact formation and cytoskeletal organization) formed during the adhesion of neonatal rat calvarial (cranial) osteoblasts attaching to and spreading on alumina orthopedic implant materials was investigated by fluorescence microscopy . Focal contacts are regions where the plasma membrane approaches the substrate to within 10-15 nm and where bundles of cytoskeletal microfilaments terminate. Fluorescent-labeling of F-actin-containing microfilaments demonstrated a typical sequence of events as rounded, suspended osteoblasts spread onto the alumina substrates, initially showing the formation of streak-like vinculin-mediated focal patches . In another study, the morphological responses of individual osteoblasts as they attached and spread on alumina surfaces in vitro were examined with scanning electron microscopy . The cells were round after 30 minutes, then spread radially during the next 1.5 hours until they were almost flat with a nuclear bulge on both rough and polished alumina . More recently, Josset et al  confirmed that normal biochemical and biological functions of cultured human osteoblasts are preserved in the presence of 6-mm-diameter 1.2-mm-thick alumina disks. Their results also suggested the absence of a mutagenic or carcinogenic effect on cells during the 30-day testing period, given that DNA image cytometry and interphase silver-NOR quantification showed no changes in cell ploidy, growth rates, or DNA replication compared to controls . Another recent experiment  found no differences in cell viability between human osteoblasts cultured on polished surfaces of alumina or hydroxyapatite after 48 hours. However, osteoblast adhesion  and osteoclast-like cell function  are increased on nanophase alumina (grain sizes <100 nm) compared to conventional alumina. The responses of human osteoblasts cultured on an alumina surface and subjected to cyclic stretching has also been examined , and included unchanged alkaline phosphatase activity and increased synthesis of collagen and total protein.
Others have investigated the response of various oral cells to sapphire dental implant surfaces. In one study , the influence of single-crystal sapphire on the behavior of human epithelial cells and fibroblasts derived from biopsies of the oral mucosa was studied. Compared to control cultures, no effects on cell morphology and growth characteristics were observed. Another study  sought to elucidate the ultrastructure of peri-implant junctional epithelium (IJE) on single-crystal sapphire dental implants connected to adjacent teeth by a metal superstructure, by examining the peri-implant gingivae of ten monkeys using a transmission electron microscope at 3, 6 and 12 months after implant insertion. At the time of examination, the ultrastructural features of the IJE were almost identical to those of the natural junctional epithelium attached to natural teeth. These features included developed Golgi complexes, rough-surfaced endoplasmic reticulum, numerous free ribosomes and mitochondria. The innermost cells of IJE were attached to the implant surface by means of 50-100-nm thick basal lamina-like structures and hemidesmosomes, but lacked a dental cuticle as seen on teeth. This epithelial attachment of the IJE was often indistinct or absent at the apical portion of the IJE which terminated at the level of alveolar crest. In yet another study , amorphous alumina was found to be slightly bioactive but more cytocompatible than titanium for human alveolar (tooth socket) bone osteoblasts and gingival fibroblasts. Cytocompatibility was assessed at the level of both the basic (attachment, proliferation, cell protein content) and the specific features (intracellular alkaline phosphatase activity, cytoskeleton) of the cells that were in direct contact with the coating .
Surface chemistry modifies cell response. For instance, a comparison of the response of costochondral (rib cartilage) chondrocytes at two stages of endochondral development demonstrated that the effects of various materials were surface- and cell-maturation-dependent. Cells cultured on titanium exhibited increased alkaline-phosphatase-specific activity, whereas those cultured on Al2O3 showed decreased enzyme activity . Another in vitro study  investigated the effect of surface chemistry modification of bioceramics on human bone-derived cells grown on biomaterial surfaces for 2 weeks. Cells were cultured on either pure alumina (Al2O3), alumina doped with magnesium ions ([Mg]-Al2O3), hydroxyapatite (HAP) or tissue culture polystyrene (TCPS). The researchers measured expression of alkaline phosphatase (ALP), thrombospondin (Tsp), osteopontin (OP), osteocalcin (OC), osteonectin (ON/SPARC), type I collagen (Col I), and bone sialoprotein (BSP). Protein levels for ALP, OP, OC, and BSP were significantly greater at day 5 in cells cultured on [Mg]-Al2O3 than in cells grown on pure Al2O3. By day 14, the levels of ALP, Tsp, Col I, OP, ON/SPARC, and BSP rose significantly above those occurring in cells grown on pure Al2O3, HAP, and TCPS. This suggests both that cells from the same patient respond to differences in surface chemical groups, and that substratum chemistry which facilitates cellular adhesion will enhance cellular differentiation  – though there is evidence that Al2O3 cannot act as a co-carcinogenic carrier for polycyclic aromatic hydrocarbons (PAHs) .
As with other materials, the interaction of cells with alumina implant materials is usually protein-mediated. For example, the adherence of Streptococcus mutans OMZ-176 bacteria was the lowest on uncoated polycrystalline alumina and on single-crystal alumina (sapphire) precoated with human serum or saliva, of six common implant materials tested . Surface free energy of uncoated material was strongly (negatively) correlated with S. mutans bacterial adherence . However, the correlation disappeared when coated materials were tested . This suggests that other binding mechanisms (e.g., protein-surface interactions) are commonly of greater importance to microbial adhesion to implant surfaces in vivo, although hydrophobic interactions may sometimes play an important role [1108-1110]. Another study  found that differences in surface energy achieved by changing implant material composition of a ternary mixture of Al2O3, SiO2, and TiO2 could not be correlated to varying cell responses, although overall biocompatibility (in terms of cell proliferation and metabolic activity) was good.
What about blood cells? Alumina ceramic male pivots used in a totally implantable centrifugal artificial heart were evaluated for vitro platelet adhesion and activation, events which may play key roles in thrombogenesis on foreign surfaces . Platelet adhesion on alumina, assessed using monoclonal antibody (CD61) directed against glycoprotein IIIa, was found to be about the same as for pure titanium, silicon carbide, and ultrahigh molecular weight polyethylene, somewhat higher than for Ti-6A1-4V alloy, but much lower than for polycarbonate. Platelet activation on alumina was evaluated  by measuring P-selectin (GMP-140) released from irreversibly activated platelets. GMP-140 levels for all tested materials were not significantly different from the control value of 45.9 nanogram/cm3, and platelet activation by alumina was not observed under the static conditions in this work . Another study found only 0.5 platelets/mm2 adhered to alumina surfaces that had been exposed to human whole blood, although significant fibrin was also adhered .
Alumina-coated surfaces have also been found to significantly reduce adhesion of Porphyromonas gingivalis ATCC33277 , an oral anaerobic bacterium important in periodontal disease and oral malodor.
Last updated on 30 April 2004