Research Article: Time resolved 3D live-cell imaging on implants

Date Published: October 10, 2018

Publisher: Public Library of Science

Author(s): Alexandra Ingendoh-Tsakmakidis, Lena Nolte, Andreas Winkel, Heiko Meyer, Anastasia Koroleva, Anastasia Shpichka, Tammo Ripken, Alexander Heisterkamp, Meike Stiesch, Antonio Riveiro Rodríguez.


It is estimated that two million new dental implants are inserted worldwide each year. Innovative implant materials are developed in order to minimize the risk of peri-implant inflammations. The broad range of material testing is conducted using standard 2D, terminal, and invasive methods. The methods that have been applied are not sufficient to monitor the whole implant surface and temporal progress. Therefore, we built a 3D peri-implant model using a cylindrical implant colonized by human gingival fibroblasts. In order to monitor the cell response over time, a non-toxic LIVE/DEAD staining was established and applied to the new 3D model. Our LIVE/DEAD staining method in combination with the time resolved 3D visualization using Scanning Laser Optical Tomography (SLOT), allowed us to monitor the cell death path along the implant in the 3D peri-implant model. The differentiation of living and dead gingival fibroblasts in response to toxicity was effectively supported by the LIVE/DEAD staining. Furthermore, it was possible to visualize the whole cell-colonized implant in 3D and up to 63 hours. This new methodology offers the opportunity to record the long-term cell response on external stress factors, along the dental implant and thus to evaluate the performance of novel materials/surfaces.

Partial Text

The use of dental implants constitutes a revolution in dentistry by restoring the tooth function in partially or fully edentulous patients. Approximately two million dental implants are placed worldwide each year [1,2]. Peri-implant inflammation might be induced by oral bacterial biofilms and leads to gradual tissue destruction and eventual implant loss [3]. According to a recent meta-analysis, the median prevalence of peri-implant infections is 26% for patients with at least 5 years implant function time and 21.2% with at least 10 years [4]. Therefore, novel antibacterial implant materials and surfaces are proposed in order to minimize the biofilm-related dental implant failure. For instance, surface coatings or laser-structured and liquid-infused surfaces have been shown to be antibacterial [5–9].

An intact implant-mucosa interface is necessary to avoid oral biofilm growth along the implant into the tissue [10–12]. The reaction of soft-tissue cells from up to down along the implant is of great interest, since external stress progression, like biofilm growth, is apical [33]. In order to complement the high-throughput 2D testing with spatiotemporal information, a non-invasive and time resolved imaging in a three-dimensional setting of the cell reaction would be of advantage. We established a three-dimensional peri-implant model and a LIVE/DEAD staining, which allowed the non-invasive monitoring of cell death progression along the implant over time.

We developed a method for non-invasive optical monitoring of live cell response in an in vitro 3D peri-implant model. To the best of our knowledge, this is the first demonstration of non-invasive imaging of dynamic change of the cell condition on an implant in response to external stress. The proof of principle, demonstrated in this work, allows the development of optically based imaging tools for the analysis of more sophisticated in vitro 3D cell-implant models (e.g. co-cultured with oral bacterial biofilms). Thus, this represents a unique experimental platform for tissue engineering and implant material testing in combination with oral microbiology.




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