In Vitro Analysis of Fibronectin-Modified Titanium Surfaces

Abstract

Background: Glow discharge plasma (GDP) procedure is an effective method for grafting various proteins, including albumin, type I collagen, and fibronectin, onto a titanium surface. However, the behavior and impact of titanium (Ti) surface modification is yet to be unraveled.

Purpose: The purpose of this study is to evaluate and analyze the biological properties of fibronectin-grafted Ti surfaces treated by GDP.

Materials and methods: Grade II Ti discs were initially cleaned and autoclaved to obtain original specimens. Subsequently, the specimens were GDP treated and grafted with fibronectin to form Ar-GDP (Argon GDP treatment only) and GDP-fib (fibronectin coating following GDP treatment) groups. Blood coagulation test and MG-63 cell culture were performed to evaluate the biological effects on the specimen.

Results: There was no significant difference between Ar-GDP and GDP-fib groups in blood compatibility analysis. While in the MTT test, cellular proliferation was benefited from the presence of fibronectin coating. The numbers of cells on Ar-GDP and GDP-fib specimens were greater than those in the original specimens after 24 h of culturing.

Conclusions: GDP treatment combined with fibronectin grafting favored MG-63 cell adhesion, migration, and proliferation on titanium surfaces, which could be attributed to the improved surface properties.

Fig 1. Schematic of the flow discharge plasma system.

The blue circles represented the gas ions (argon or allylamine) which were used to modify the titanium surface.

Fig 2. Schematic of sample preparation.

(a)The surfaces of Ti discs (original specimens) were cleaned with argon-based GDP. These titanium discs were then labeled as “Ar-GDP.” (b) Amine groups were grafted onto the Ar-GDP surfaces in the GDP reactor fed with allylamine (AA) gas. These specimens were subsequently treated in glutaraldehyde (GA) and fibronectin solutions and were labeled as “GDP-fib”.

Fig 3. Scanning electron microscopy images of the Ar-GDP (a) and GDP-fib (b) titanium discs.

Irregular folding of the fibronectin was revealed on the surface of fibronectin-grafted titanium disks (white arrows).

Fig 4. Blood coagulation of Ar-GDP and GDP-fib samples was expressed as percentage and compared to that of the blank plate (100%).

There was no significant difference between two groups.

Fig 5. The MTT assay of MG-63 cells.

Compared to the original specimen, both on the surface of GDP-treated and fibronectin-modified titanium, the MG-63 cells viability was significantly enhanced after 24 hours of culture (**P < 0.01).

Fig 6. Scanning electron microscopy images of MG-63 cells cultured on original specimen (a), Ar-GDP (b), and GDP-fib (c) titanium discs after 24 h.

Cells showed morphological alternation from spindle to more-stellar shapes, and extensive process (white arrows).