Histomorphometrical and CBCT Evaluation of Tissue Loss Progression Induced by Consecutive, Alternate Ligatures in Experimental Peri-Implantitis in a Dog Model: A Pilot Study

Abstract

Objectives: Soft and hard tissue breakdown was histologically and radiologically assessed around implants with alternate, consecutively placed ligatures on the same edentulous dog hemimandible. The influence of ligatured implants (LI) on adjacent non-ligatured implants (NLI, as a possible naturally induced peri-implantitis) was also evaluated.

Material and methods: Three months after tooth extraction, five dental implants were placed in the dog hemimandible. Two months after abutment placement, ligatures were placed subsequently two months apart on alternate implants, while both intermediate implants were left without ligatures. Ligatures were kept in place during the entire experiment, and no plaque control measures were taken. Eleven months post-implantation, the animal was sacrificed. Undecalcified ground sections were cut, stained with Masson Goldner and MOVAT Pentachrome and evaluated by light microscopy. Soft and hard tissue loss was assessed using histomorphometric and CBCT parameters.

Results: All NLI presented deep false peri-implant pockets on the oral aspect and pronounced vertical bone resorption on the buccal aspect. After 2, 4 and 6 months, during the breakdown period, more than 30% of the bone was lost in LI in all directions, while, despite immediate vicinity, NLI displayed less destruction. Intense inflammation, typical for induced peri-implantitis, was present, with similar intensity in LI as NLI, but in different parts of the lesions. Morphometry confirmed intense soft tissue inflammation, more bone resorption and higher amounts of infiltrated connective tissue in LI when compared with NLI.

Conclusion: Within the limits of the present pilot study, the adequacy of the experimental dog model based on ligature-induced peri-implantitis was able to be successfully challenged by non-ligature models of spontaneously occurring peri-implant inflammation, while meeting the requirements for experimental designs with a very small numbers of animals. The influence of implants with severe peri-implantitis on adjacent implants resulted in less than expected tissue loss in the latter accession numbers.

Keywords: animal study; cone-beam CT; dental implant; histomorphometry; ligature induced; naturally occurring; peri-implantitis.

Figure 1

Timeline of the experiment.

Figure 2

Intra-surgical image after the insertion of the implants. Note the integrity of the buccal bone, as a consequence of the rigorous prosthetic-driven implant placement.

Figure 3

Intra-operative view of the implants at the exposure time, 3 months post-implantation, with the healing abutments in place.

Figure 4

Post-operative image at 9-month post-implantation. Note the ligatures placed in alternating implants 1, 3 and 5, and the subsequent local inflammation.

Figure 5

Radiological follow-up immediately after the abutment connection, revealing no post-implantation pre-exposure bone loss around the screws.

Figure 6

Landmarks identified in each sagittal image at all buccal, oral, mesial and distal aspects: IS (implant shoulder), the bottom of the bone defect (BD), the alveolar bone crest (BC) and the angle between segments IS-BD and BD-BC. CBCT implant #5 (original magnification ×8).

Figure 7

Histomorphometric landmarks identified on both buccal and lingual aspects in each implant: 1—implant shoulder (IS); 2—implant surface at the level of the bone crest (imp); 3—bone crest, defined as the most coronal point of the bone (BC); 4—bottom of the bone defect (BD); 5—margin of the peri-implant mucosa (PM); 6—the apical termination of the junctional epithelium (aJE); 7—the coronal level of the infiltrated connective tissue (cICT); 8—the apical extension of the infiltrated connective tissue (aICT); 9—the most apical extension of the submarginal biofilm that was interposed between the implant and the pocket epithelium of the peri-implant mucosa (aPlaque). (Implant #5, mirroring sections, (A)—Masson Goldner Anilin blue stain, (B)—Movat Pentachrome stain, original magnification ×2).

Figure 8

“Open wound” aspect of the ICT in contact with the implants surface and a total lack of the epithelial lining. ICT is diffusely present in the soft tissue and is seen to progress into the bone marrow, without any sign of fibrous encasement. The peri-implant sulcus is deep and the intimate contact between the soft tissues and the implant is below the BC level (implant #3, MGA, bar measure is 900 µm).

Figure 9

Despite the presence of intense inflammation induced by the ligature, the deep peri-implant sulcus presents a continuous epithelial lining. The intimate contact between the soft tissues and the implant is below the BC level (implant #5, MGA, bar measure is 2 mm).

Figure 10

In the absence of the ligature, full epithelial lining is observed along the interface with the implant, with no ulceration. Note the intimate contact between the soft tissues and implant surface that coronally ends above the level of the BC, while the ICT remains above the BC-BD line. (implant #2, MGA, bar measure is 700 µm).

Figure 11

Bone in active phase of remodeling. ICT in simultaneous contact with the bone and with the implant surface with no squamous epithelium covering. Additionally, active bone remodeling is identified by the scalloped endosteal surface (Howship’s lacunae) (implant #1, MGA, bar measure is 200 µm).

Figure 12

Position and effect of the ligatures. Note the intense inflammation on the left (buccal) aspect and the accentuate resorption of the buccal plate, up to its total loss. In the oral aspect, the peri-implant sulcus is deep and wide and the implant is in intimate contact with the ICT in just a limited area above the BD (implant #1, MGA, bar measure is 2 mm).

Figure 13

ICT in direct contact with the implant surface, without an interposed epithelial lining. Note the abundant lympho-plasmocytic infiltrate, which progresses inside the bone marrow, the thin bone trabeculae left after remodeling along with prominent osteoblast rim (implant #3, MGA, bar measure is 100 µm).