Elsevier

Biomaterials

Volume 28, Issue 36, December 2007, Pages 5418-5425
Biomaterials

The effect of hydrofluoric acid treatment of TiO2 grit blasted titanium implants on adherent osteoblast gene expression in vitro and in vivo

https://doi.org/10.1016/j.biomaterials.2007.08.032Get rights and content

Abstract

It is widely accepted that implant surface factors affect the quality of the bone-to-implant interface. Recent additional treatments superimposed on moderately rough cpTitanium surface provide further enhancement of bone-to-implant contact. The aim of this study was to compare osteoinductive and bone-specific gene expression in cells adherent to titanium dioxide-grit blasted (TiO2) versus TiO2 grit blasted and HF treated (TiO2/HF) cpTitanium implant surfaces. MC3T3-E1 cells were grown in osteogenic supplements on the titanium disk surfaces for 1–14 days. Real-time PCR was used to measure RUNX-2, Osterix, and bone sialoprotein (BSP) mRNA levels. Implants were placed in rat tibia and, following harvesting at 1–7 days after placement, real-time PCR was used to measure RUNX-2, alkaline phosphatase (ALP), and BSP mRNA levels in implant adherent cells. In cell culture, RUNX-2 and Osterix levels were significantly increased (p<0.05) on the TiO2/HF surfaces as compared to the TiO2 and smooth surfaces through the cultural period, while BSP expression was elevated on both TiO2 and TiO2/HF surfaces when compared to a machined surface control. In cells adherent to implants retrieved from rat tibia, RUNX-2 mRNA levels were 2-fold and 8-fold greater on the TiO2/HF surfaces at 1–3 and 7 days following implantation. This was paralleled by significantly greater levels of ALP at 3 and 7 days and BSP mRNA at 7 days following implantation. As a marker of osteoinduction, the increased levels of RUNX-2 in cells adherent to the TiO2/HF surfaces suggest that the additional HF treatment of the TiO2 grit blasted surface results in surface properties that support adherent cell osteoinduction. In vivo assessments of implant adherent cell phenotypes provide further insight into the mechanisms affecting alloplast–tissue interactions.

Introduction

Endosseous implants are of growing importance in medicine and dentistry. The expanding use of titanium implants as anchorage for orthopedic and dental prostheses is supported by favorable clinical responses. In the field of dental implants, long-term endosseous implant success is dependent on the phenomenon of osseointegration, defined as the formation of a direct bone-to-implant interface observed at the light microscopic level [1]. The required formation of bone on the endosseous surface of the cpTitanium implant is due to the activity of adherent and adjacent mesenchymal cells [2], [3]. Recent histological evaluation of this process in the canine model demonstrated that the formation of osteoid and its mineralization occurs within weeks of implant placement [4]. Implied is a functional relationship between the adherence of cells to the cpTitanium surface and their bone forming activity. In fact, the endosseous implant surface is one of the principal factors affecting the process of osseointegration [5]. The effect of surface topography and surface chemistry on osteogenesis is demonstrable at the cellular level in cell culture studies [6].

Osseointegration at endosseous implants requires activation of the key regulatory pathways affecting osteoblastogenesis, promotion of osteoblastic differentiation from uncommitted mesenchymal stem cells. The bone morphogenetic proteins (BMPs) are prominent signaling molecules of osteogenesis [7]. The main intracellular switch for osteogenic pathway in the mesenchymal stem cell is RUNX-2, a transcription factor necessary for osseous skeletal development and bone repair [8]. A second intracellular switch controlling osteoinduction is Osterix, another transcription factor required for osseous skeletal development [9]. In mouse studies, knockout of RUNX-2 [10] or Osterix [11] expression precluded osseous skeletal development and osteoblastogenesis. While many other signaling molecules and intracellular signaling pathways are important for regulation of bone formation, RUNX-2 and Osterix are clearly essential to the process of osteogenesis.

Recently, the regulation of RUNX-2 and Osterix expression in titanium surface-adherent osteoprogenitor cells was reported to be influenced by adherent surface parameters [12]. The aim of the present study was to determine the effect of a specific endosseous titanium surface modification on RUNX-2 and Osterix expression in adherent osteoprogenitor cells in vitro and in vivo.

Section snippets

Implant preparation and scanning electron microscopy (SEM) analysis

About 8 mm cpTitanium (grade IV) disks were prepared by machining. Other disks were subsequently treated by TiO2 grit blasting using 75 μm titanium dioxide (TiO2) particles or by identical TiO2 grit blasting with treatment by hydrofluoric acid immersion (AstraTech AB). The threaded implants (1.5 mm×1.0 mm; 0.2 mm pitch) were machined from cpTitanium and TiO2 grit blasted using 75 μm TiO2 particles. All implants and disks were manufactured for AstraTech AB and subsequently cleaned and sterilized

Results

The machined, TiO2 grit blasted (TiO2), and TiO2 grit blasted and HF treated (TiO2/HF) implants were examined by SEM (Fig. 1b–d). At 5000× magnification, the moderately rough surface of the TiO2 grit blasted surface was readily apparent (Fig. 1c). The HF treatment imparted nanofeatures to the surface with dimensions of approximately 100 nm in diameter (Fig. 1d). Comparison of the chemical composition of the TiO2 versus TiO2/HF surfaces by X-ray photoelectron spectroscopy (XPS) analysis revealed

Discussion

The interaction of cells and tissues with the endosseous implant surface has been the focus of experimental scrutiny for several decades. In vitro investigations at the cellular and molecular level [3], [6] and histological interpretation of these events [19] suggest that implant surface properties influence interfacial bone formation by modulating the adherent cell phenotype. This study provides cell culture-derived data and novel in vivo evidence corroborating the observations that

Conclusions

Scanning electron micrographs revealed that the HF treatment TiO2 grit blasted cpTitanium surfaces provided the treated surface with nanometer scale surface features. Both cell culture and in vivo comparison of the implant surface adherent cell phenotype at the level of bone-specific mRNA expression indicated that the treatment of TiO2 grit blasted cpTitanium surfaces using HF results in a surface that enhanced the adherent cell osteoinductive and bone-specific mRNA expression. Ongoing analysis

Acknowledgment

The authors wish to thank AstraTech AB for providing the disks and the associated analyses of the prepared surfaces.

References (35)

  • R. Branemark et al.

    Biomechanical characterization of osseointegration during healing: an experimental in vivo study in the rat

    Biomaterials

    (1997)
  • P.I. Branemark et al.

    Osseointegrated implants in the treatment of the edentulous jaw. Experience from a 10-year period

    Scand J Plast Reconstr Surg Suppl

    (1977)
  • J.E. Davies

    Understanding peri-implant endosseous healing

    J Dent Educ

    (2003)
  • T. Berglundh et al.

    De novo alveolar bone formation adjacent to endosseous implants

    Clin Oral Implants Res

    (2003)
  • T. Albrektsson et al.

    Osteoinduction, osteoconduction and osseointegration

    Eur Spine J

    (2001)
  • Z. Schwartz et al.

    Implant surface characteristics modulate differentiation behavior of cells in the osteoblastic lineage

    Adv Dent Res

    (1999)
  • D. Chen et al.

    Bone morphogenetic proteins

    Growth Factors

    (2004)
  • Cited by (0)

    View full text