2.1

Study design

The protocol of this prospective clinical study was approved by the Ethics Committee of the University Hospital Center (CHU) of Liege and was registered in the ClinicalTrials.gov database (Identifier NCT02150226). The study design is illustrated in Fig. 1 . Written patient consent was obtained before inclusion. The patients were treated by four experienced operators in the Department of Fixed Prosthodontics, Institute of Dentistry, CHU of Liege, Belgium. The eligibility criteria included the need for a molar or premolar crown (maximum of 6 elements per patient). The restorations were realized either on the teeth or the implants, and FDPs were also included, provided they were on the implants and limited to three elements (maximum 2 FDPs per patient). The exclusion criteria were severe or acute periodontal or carious diseases and poor oral hygiene. Patients without antagonistic teeth or those with removable prostheses as antagonists were excluded. A total of 47 patients, with a mean age of 54.3 ± 15.3 years, were recruited (14 males, 33 females). A total of 75 restorations (101 tooth elements) were included. Sample descriptions of patients, restorations, and tooth elements are presented in Table 2 . The participants received no financial compensation, although the treatment and prostheses were free of charge. All of the patient-related data were recorded in a specifically designed online database, to facilitate easy access to clinical data, pictures, and data obtained from ex vivo analyses for consultation. The database is hosted on a university hospital-secured server.

Study design.

2.2

Clinical procedure and fabrication of the restorations

2.2.1

Tooth preparation and impression for tooth or implant-supported prostheses

All clinical and technical procedures were performed strictly according to the manufacturer’s recommendations. The teeth were prepared following standardized criteria (1.0–1.5 mm occlusal depth cut to achieve the appropriate occlusal anatomy, 1.0–1.5 mm functional cusp tip reduction, 0.5 mm gingival chamfer reduction, and a 6–8 degree taper to the axial walls). A double-mix impression was performed with high- and low-viscosity A-silicone impression materials (Aquasil Heavy/XLV, Dentsply De Trey, Konstanz, Germany). An open tray impression technique was performed using the same impression material used for implant restorations. Two implant systems (NobelReplace, Nobel Biocare, Gothenburg, Sweden; and Standard Implants, Straumann, Basel, Switzerland) were used in this clinical study. The shade was registered using the Vita Classic System (Vita Zahnfabrik, Bad Säckingen, Germany). Restorations of antagonistic teeth were replaced in the presence of a deficient marginal joint, decay, or inadequate morphology.

2.2.2

Provisional restorations

Before manufacturing the zirconia restorations, CAD-CAM composite provisional crowns (Lava Ultimate, 3M ESPE, Seefeld, Germany) or PMMA-provisional FDPs were made. After die scanning the restoration was designed using CAD/CAM software: Exocad (Darmstadt, Germany) or Dental Wings (Montreal, Canada) (DPI Lava milling center, Anderlecht, Belgium). Specific buccal and palatal undercuts were added to the crown design to facilitate cemented crown removal ( Fig. 2 a). The file was transferred to a milling machine for manufacturing (Lava CNC 500, Serial Number: 07019 (2009), 3M ESPE). The provisional restorations were adjusted in the mouth and used as a mock-up for the design of zirconia restorations. Particular attention was paid to the occlusal contact point adjustment to obtain at least one flat contact surface of a minimum of 1−2 mm ² per cusp by grinding or adding a composite ( Fig. 2 a and b).

Realization of a cemented zirconia crown (tooth #16). (a) Occlusal contact points before adjustment on the Lava Ultimate provisional crown. (b) Occlusal contact points after adjustment to obtain 1–2 mm ² flat surfaces on each cusp. The provisional crown was scanned and used as a mock-up for the zirconia crown. (c) Lava Plus zirconia crown after intraoral try-in and registering of occlusal contact points. Areas that will not be glazed are landmarked with permanent ink. The palatal surface is left unglazed on a part above the undercut. d) For molars, two cusps have been randomly selected to remain unglazed: one centric cusp (unglazed centric cusp; UCC) and one non-centric (unglazed non-centric cusp; UNCC). The two other cusps were called “glazed centric cusp” (GCC) and “glazed non-centric cusp” (GNCC). (e) Glazed Lava Plus zirconia final crown. (f) Landmarking of areas to be analyzed after intraoral try-in: occlusal contact points and the axial areas on the buccal and palatal surfaces located up to the undercut created to remove the crown. (g) Intraoral occlusal view after baseline evaluation and restoration placement. (h) Buccal view.

2.2.3

Zirconia prostheses

Provisional restorations were scanned for zirconia restoration fabrication (Lava Plus, 3M ESPE, Seefeld, Germany) with the same milling system. Lava Plus is a second-generation 3Y-TZP, with a higher translucency than Lava Frame Zirconia (first-generation) due to a lower alumina content of 0.1%, optimally distributed within the material (Lava Plus, technical product profile, 3M). Sintering was performed according to the manufacturer’s instructions at 1450 °C for 2 h. The zirconia restorations were polished with a sequence of three diamond-impregnated silicone discs (Edenta AG, Hauptstrasse, Switzerland) and a diamond polishing paste (Bredent, Senden, Germany). Implant-supported restorations were bonded onto a specific titanium abutment (1000er-Serie, Medentika, Hugelsheim, Germany) with a resin composite cement (RelyX Ultimate [3M, Seefeld, Germany] in the first part of the study or Multilink Hybrid Abutment [Ivoclar Vivadent, Schaan, Liechtenstein] in the second part) after sandblasting the abutment and the zirconia restoration with 50 μm alumina particles at 2 bar according to the manufacturers’ recommendations.

Zirconia restorations were tried-in, and, if needed, occlusal contact points were adjusted with a diamond bur (ISO 524 grit size) and polished using a sequence of three zirconia-dedicated silicon gums (Diasynt Plus/Diacera Zirconium, Eve Ernst Vetter, Pforzheim, Germany), following the manufacturer’s recommendations. The need for occlusal adjustments was encoded in an online database for each occlusal area.

For restorations on implants that had no undercuts on buccal and lingual/palatal surfaces, a small groove was designed with a diamond bur ( Fig. 3 a) as a landmark for ex vivo analyses of axial surfaces, which were the control areas (free of mechanical stress). For molars, two cusps were randomly selected to remain unglazed: one centric cusp (UCC) and one non-centric cusp (UNCC). The two other cusps were called “glazed centric cusp” (GCC) and “glazed non-centric cusp” (GNCC) ( Fig. 2 c and d). For premolars, one cusp was randomly selected to remain unglazed ( Fig. 3 a). The buccal surface was systematically glazed, and the lingual/palatal surface was left unglazed on a part above the undercut or groove ( Figs. 2 c and 3 a). The glaze (IPS empress stains and eMax Ceram glaze, Ivoclar Vivadent, Schaan, Liechtenstein) was sintered at 780 °C for 1 min. Definitive bonding (bond was eliminated during glaze firing) on the specific titanium abutment was performed following a previously described procedure. The glazed restorations were tried-in, and occlusal contact point areas (one per cusp, i.e. , four on molars and two on premolars), as well as lingual/palatal and buccal areas, were marked for ex vivo analyses and registered with a picture ( Figs. 2 f and 3 b).

Screw-retained zirconia crown on an implant (tooth #34), which was realized in the same way as the cemented crown in Fig. 2 . (a) Lava Plus zirconia crown after intraoral try-in and registering of occlusal contact points. Areas that will not be glazed are landmarked with permanent ink. The lingual surface is left unglazed on a part above the groove. For premolars, one cusp is randomly selected to remain unglazed. (b) Landmarking of the areas to be analyzed after intraoral try-in of the final zirconia crown: occlusal contact points and the axial areas on the buccal and lingual surfaces are located up to the groove performed on the restoration surface. (c) Intraoral occlusal view after baseline evaluation and restoration placement. d) Buccal view.

2.2.4

Zirconia prostheses placement and removal for ex vivo analyses

Baseline ex vivo analyses of the zirconia restorations were performed before placement. Screw-retained restorations were torqued at 35 Ncm ⁻¹ ( Fig. 3 c and d). To remove the gold sputter coating, the restorations were cleaned first with an alcohol-soaked cotton ball, brushed with toothbrush and toothpaste, and finally immersed in an ultrasonic bath containing alcohol for 3 min. Cemented restorations were sealed with eugenol-free cement (RelyX Temp NE, 3 M ESPE) ( Fig. 2 g and h), and the teeth were disinfected with 2% chlorhexidine. After 6 months, the restorations were removed for ex vivo analyses. Provisional restorations replaced the zirconia restorations during the analyses. The zirconia restorations were placed again in the mouth of the patient, following the same procedure as for the first time. Evaluations were repeated after a one-year intraoral stay and after two years. The occlusal contact points were landmarked with bite registration paper before removing the prostheses and encircled with permanent ink ( Figs. 2 f and 3 b).

2.3

Ex vivo analyses

Ex vivo analyses included (1) phase transformation with Raman spectroscopy and (2) glaze wear using scanning electron microscopy (SEM). Sample descriptions of the ex vivo analyzed areas (occlusal or axial, glazed, or not glazed) (n tot = 528) are presented in Table 2 .

2.3.1

Raman spectroscopy

Five measurement points (MPs) were randomly chosen for each occlusal contact area and each axial area ( i.e. , 30 measurements for a molar and 20 measurements for a premolar). Consequently, the analyses were performed for 2640 MPs at each evaluation time (baseline, 6 months, 1 year, and 2 years). Sample descriptions of the MPs are presented in Table 2 . Raman spectra were recorded using a Raman spectrometer (Horiba Jobin Yvon, Kyoto, Japan). The excitation laser was provided by a HeNe laser (632 nm), with 1 mW power focused at the surface of the specimen, and the Raman spectra were acquired using a charge-coupled device detector (Horiba-Jobin Yvon, Kyoto, Japan) with a cm ⁻¹ spectral resolution of 1 (1800 grooves/mm grating). The Raman spectrometer was combined with an optical microscope (Olympus LX71; Olympus Corporation, Tokyo, Japan). A confocal pinhole with adjustable diameter was used for confocal detection, and an 80× objective (numerical aperture 0.75) was used to achieve a 1 μm ³ resolution (lateral × axial). A pinhole aperture of 1.99 μm was used to reach a collection depth of 1.44 μm. The depth was determined using the calibration-wedge method [ ].

The collected spectra were analyzed, and the transformed monolithic volume fraction (V _fm ), which is a measurement of the transformation volume ratio in the confocally probed volume, was estimated using Eq. (1) , which was initially proposed by Clarke and Adar [ ]

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