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Transfection System of Amino-Functionalized Calcium Phosphate Nanoparticles: In Vitro Efficacy, Biodegradability, and Immunogenicity Study
- Mostaghaci, Babak, Susewind, Julia, Kickelbick, Guido, Lehr, Claus-Michael, Loretz, Brigitta
- ACS Applied Materials & Interfaces 2015 v.7 no.9 pp. 5124-5133
- biocompatibility, biodegradability, cell culture, complement, confocal laser scanning microscopy, crystal structure, cytokines, cytotoxicity, flow cytometry, hydroxyapatite, immune response, innate immunity, macrophages, nanoparticles, nucleotides, pH, transfection
- Many methods have been developed in order to use calcium phosphate (CaP) for delivering nucleotides into living cells. Surface functionalization of CaP nanoparticles (CaP NPs) with N-(2-aminoethyl)-3-aminopropyltrimethoxysilane was shown recently to achieve dispersed NPs with a positive surface charge, capable of transfection (Chem. Mater. 2013, 25 (18), 3667). In this study, different crystal structures of amino-modified CaP NPs (brushite and hydroxyapatite) were investigated for their interaction in cell culture systems in more detail. Qualitative (confocal laser scanning microscopy) and quantitative (flow cytometry) transfection experiments with two cell lines showed the higher transfection efficacy of brushite versus hydroxyapatite. The transfection also revealed a cell type dependency. HEK293 cells were easier to transfect compared to A549 cells. This result was supported by the cytotoxicity results. A549 cells showed a higher degree of tolerance toward the CaP NPs. Further, the impact of the surface modification on the interaction with macrophages and complement as two important components of the innate immune system were considered. The amine surface functionalization had an effect of decreasing the release of proinflammatory cytokines. The complement interaction investigated by a C3a complement activation assay did show no significant differences between CaP NPs without or with amine modification and overall weak interaction. Finally, the degradation of CaP NPs in biological media was studied with respect to the two crystal structures and at acidic and neutral pH. Both amino-modified CaP NPs disintegrate within days at neutral pH, with a notable faster disintegration of brushite NPs at acidic pH. In summary, the fair transfection capability of this amino functionalized CaP NPs together with the excellent biocompatibility, biodegradability, and low immunogenicity make them interesting candidates for further evaluation.