Enhanced osteoblastic cell response on zirconia by bio-inspired surface modification

https://doi.org/10.1016/j.colsurfb.2013.01.023Get rights and content

Abstract

Excellent esthetic properties and limited plaque adhesion make zirconia ceramics an ideal material for implants in the fields of dentistry and orthopedics. Unfortunately, the physicochemical stability of zirconia makes it difficult to improve biocompatibility through surface modification. The dopamine-derived residue, 3,4-dihydroxy-l-phenylalanine (l-DOPA), has been identified as an important molecule secreted by marine mussels for the formation of adhesive pads. This study coated zirconia with l-DOPA to improve the biocompatibility of ZrO2. As confirmed by contact angle and X-ray photoelectron spectroscopy (XPS), the formation of l-DOPA film can be controlled by varying the process temperature. Results from scanning electron microscopy (SEM) and atomic force microscopy (AFM) show that the topography of the zirconia substrate was preserved after being coated with a film of l-DOPA. Specifically, the thickness of the coating and initial cell spreading ability were both enhanced by preparing samples at higher temperatures. l-DOPA coated zirconia demonstrated better cyto-compatibility than uncoated specimens, as indicated by cell responses such as cell spreading and proliferation. These preliminary results suggest that l-DOPA film could be used to improve the cyto-compatibility of zirconia and further has the potential to immobilize other biofunctional molecules in biomedical applications.

Highlights

► We present a bio-inspired technique of coating bio-inert zirconia with a film of l-DOPA in order to enhance biocompatibitliy. ► l-DOPA molecules change the chemical properties of the substrate without altering the physical properties, resulting in excellent cell performance. ► Temperature could be used to control the properties of l-DOPA films.

Introduction

Previous success in the application of zirconia ceramics as a framework material is a clear demonstration of its applicability in all-ceramic esthetic restoration [1], [2]. Thanks to its high mechanical strength and excellent fracture toughness, zirconia is widely used in orthopedic and dental applications such as artificial knees, dental implants, and dental crowns [3]. However, zirconia is categorized as a bio-inert material that resists the formation of chemical bonds with bone tissue following implantation, which restricts its application in the field of biomedicine [4]. To overcome this problem, surface modification is used to alter the chemical and morphological properties of implants to improve tissue reactions and shorten the duration required for bones to heal [5]. Previous studies have indicated that surface modification enhances the bioactivity of zirconia. Nanostructured ZrO2 coatings on a zirconium substrate have been fabricated via micro-arc oxidation (MAO) [6]. MAO-formed zirconia films facilitate the formation of apatite and promote good cell responses. However, the MAO process requires high-voltage and is thus only applicable to conductive materials, such as zirconium rather than zirconia. Other proposed methods, such as optimizing the micro-roughness (sandblasting and acid-etching; SLA), have been applied to modify the surface of zirconia in order to increase the interface area between the implant and bone tissue [7]. SLA is an acid-etching procedure which requires strong acids that can be harmful to the environment; in addition, the residual chemicals may damage cultured cells [5]. Bioactive coatings, such as calcium phosphate [8] and fluor-hydroxyapatite [9], have also been deposited on zirconia via the sol-gel method. Unfortunately, these bioactive coatings often exhibit poor adhesion to the substrate, resulting in rapid dissolution and degradation of the coatings [10]. Ultraviolet (UV) light treatment is another emerging method used to create hydrophilic surfaces with increased Zr–OH groups [11], [12]. However, photoactive treatment requires specialized laboratory conditions and equipment. Therefore, developing a more effective method to enhance the biocompatibility of zirconia requires further study.

The neurotransmitter, 3,4-dihydroxy-l-phenylalanine (l-DOPA), is an important component in the adhesive structure of mussels [13]. Previously, the properties of adhesives found in mussels have inspired a simple, versatile approach to surface coating [14]. This technique involves the application of a thin film coating of l-DOPA on a substrate via oxidative polymerization [15]. The properties of l-DOPA film can be precisely controlled by adjusting a number of parameters. For example, the thickness of the film can be controlled in a time-dependent manner to increase overall roughness [16]. In fact, cell adhesion can be improved through the application of dopamine for only 5 min. However, no study has yet addressed whether modulating the coating temperature during the deposition of l-DOPA has a direct or indirect influence on cell behavior.

Mussel-inspired methods enable the creation of an interface on biomedical devices with complex geometry, such as dental implants with screws. Moreover, the l-DOPA-based film could serve biomedical applications by acting as an intermediate layer which immobilizes other biofunctional molecules [17], [18]. However, the effectiveness of applying l-DOPA film to dental and orthopedic implants has yet to be investigated. This study applied an l-DOPA film on a zirconia surface using a one-step process and evaluated the thickness, composition, and surface morphology of the resulting film. The influence of processing temperature on various characteristics of the film was also investigated. To evaluate the biological response of the specimens, we analyzed the in vitro protein adsorption, cell morphology and initial spreading, cytoskeletal development, cell numbers, and alkaline phosphatase (ALP) activity.

Section snippets

Materials

3,4-Dihydroxy-l-phenylalanine (l-DOPA) and tris(hydroxymethyl)aminomethane (TRIS) were purchased from Sigma–Aldrich. All other reagents were of analytical grade and used without additional purification. The specific surface area and mean particle diameter of the purchased zirconia (Aplustek Tech, Taiwan) were 9–11 m2/g and 0.15 μm, respectively. The zirconia discs were sintered at 500 °C for 10 h and maintained at 1500 °C for 5 h in a box furnace (LindBerg/Blue M, Thermo, USA) before being cooled to

Morphologies and characteristics

Initially, the zirconia powders presented a monoclinic phase (Fig. 1a: i, XRD pattern); however, this transformed into a tetragonal phase after sintering temperature reached 1500 °C (Fig. 1a: ii). Fig. 1(b) presents the results of EDS analysis, in which the EDS spectrum revealed the existence of zirconium and oxygen. Fig. 1(c) presents the composition of the specimens. The atomic concentrations of oxygen and zirconium are 70.59% and 29.41%, respectively, and the O/Zr ratio is close to the

Discussion

Zirconia bioceramic implants require biocompatibility and appropriate mechanical properties for use in the fields of dentistry and orthopedics. Zirconia has excellent esthetics and mechanical properties, including strength and fracture toughness exceeding those of Al2O3 [20]. In addition, zirconia does not promote inflammation or ion-release in the body and therefore has considerable potential as a next-generation material for dental applications [21], [22]. However, the bio-inert nature of

Conclusion

This study proposes an easy, efficient, solvent-free process for the coating of l-DOPA film on a zirconia surface. The bio-inspired l-DOPA film altered the chemical properties of the zirconia surface considerably without affecting its topography. The l-DOPA coating also outperformed uncoated zirconia in osteoblast responses, including cell adhesion, cytoskeleton development, and cell number increase. During the coating process, temperature could be used to control the properties of l-DOPA

Acknowledgment

This work was financially supported by the National Science Council, Taiwan, under grant NSC 100-2221-E-006-263.

References (35)

  • F. Zarone et al.

    Dent. Mater.

    (2011)
  • L. Le Guéhennec et al.

    Dent. Mater.

    (2007)
  • H.W. Kim et al.

    Biomaterials

    (2004)
  • H.C. Gledhill et al.

    Biomaterials

    (2001)
  • W. Att et al.

    Biomaterials

    (2009)
  • Z. Zhang et al.

    Colloids Surf. B: Biointerfaces

    (2012)
  • Y.M. Shin et al.

    Colloids Surf. B: Biointerfaces

    (2011)
  • K.E. Smith et al.

    Biomaterials

    (2010)
  • C. Piconi et al.

    Biomaterials

    (1999)
  • P.F. Manicone et al.

    J. Dent.

    (2007)
  • K. Yang et al.

    Biomaterials

    (2012)
  • J. Chen et al.

    J. Membr. Sci.

    (2009)
  • H. Zeng et al.

    Biomaterials

    (1999)
  • C.J. Wilson et al.

    Tissue Eng.

    (2005)
  • S.H. Ku et al.

    Biomaterials

    (2010)
  • S.K. Hsu et al.

    Ultrasound Med. Biol.

    (2011)
  • N.G. Rim et al.

    Colloids Surf. B: Biointerfaces

    (2012)
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