RGD-FUNCTIONALIZATION OF ANODIZED ZIRCONIUM AS A POTENTIAL STRATEGY FOR BIOMEDICAL APPLICATION: AN IN VITRO STUDY

MARIA FLORENCIA TANO DE LA HOZ; ANDREA GOMEZ SANCHEZ; VALENTINA MARISCOTTI; RAUL PROCACCINI; MELISA MICHELETTI; LUCIANA BARBINI; MARIA R KATUNAR; SILVIA CERE

San Pablo

Congreso; XI COLAOB and XVII LASAO XI Latin American Congress of Artificial Organs and Biomaterials; 2021

Surface functionalization is an increasingly promising approach for property enhancement of biomaterials [1, 2]. This type of superficial modification offers the possibility of allowing greater control over the biological interactions between the implant and the surrounding tissue [3]. In the present in vitro study, the effect of functionalization of anodized zirconium (Zr) with RGD peptides (arginine-glycine-aspartic acid) on cell behavior as a feasible approach to stimulate its bioactivity is presented. The surface of Zr anodized at 60 V in H3PO4 was functionalized using silanes (APTES) as coupling agents to form a chemical bond between the anodized metal and the organic molecules. The surface modifications were evaluated by Raman spectroscopy and X-ray photoelectron spectroscopy (XPS). Electrochemical tests were performed in simulated body fluid solution (SBF) in order to determine the effect of functionalization with the organic molecules on the stability of the anodized zirconium oxide film. Finally, the biocompatibility of the material was evaluated in an in vitro cell model and cell spreading area, elongation factor and circularity parameters were calculated by image analysis. For this, the human osteosarcoma cell line MG-63 (ATCC® CRL-1427TM) was used. Surface analysis by Raman spectroscopy showed that the crystal structure of the oxide formed by anodization is predominantly monoclinic and that the functionalization treatment does not alter the chemical composition of the anodic oxide film. XPS showed the presence of peptides on the surface after functionalization, although it is likely that the deposition of the organic film was not uniform, possibly exposing the underlying anodic zirconium oxides that have incorporated phosphates from the anodizing solution. Anodic polarization curves and electrochemical impedance spectroscopy results indicated that both anodized and functionalized with the RGD peptide surfaces present excellent corrosion resistance in SBF. Electrochemical impedance spectroscopy results evidences that both surfaces behave as non-ideal capacitors, characteristic of passive films on valve metals. Scanning electron microscopy analysis showed greater cell adhesion on the functionalized Zr surfaces compared to the material without the peptide coating. The morphometric analysis indicated that functionalization treatment induces a significant increase in the mean cell area of MG-63 cells attached to RGD-coated samples compared to anodized surfaces. This increase in cell area was accompanied by a drop in roundness, indicating that cells spread more on RGD-coated surfaces compared to non-funcionalized samples. The results presented demonstrate that the functionalization treatment with RGD peptide does not cause any detrimental effect on the anodic oxide formed on zirconium during anodization and, in turn, increases the in vitro biocompatibility of the material at short times. Thus, the immobilization of RGD peptides on pieces of anodized Zr would represent an optimal strategy to promote and increase the bioactivity of the material and the osseointegration process.