Six patients, requiring bilateral sinus augmentation procedures prior to placement of implants and no contraindications for this procedure (e.g., uncontrolled diabetes, long-term steroid usage, and blood disorders) were selected from those presenting at the Aristotle University of Thessaloniki School of Dentistry, Department of Periodontology. All patients were advised of alternative treatment plans and selected the plan requiring maxillary sinus elevation. Further, the patients were informed of the requirements for participation in the study, and all had the option of withdrawing from the study at any time. The nature of the study was explained to each patient, and each signed an informed consent form that was approved by the University ethical Committee on Activities Involving Human Subjects of the University of Thessaloniki, Dental School (14/02-02-2017). Due to the pilot character of this study, no sample size or power calculation was performed and only a few patients in need for a bilateral sinus augmentation were included. The intention was to assess any trends for differences between groups, and thus to decide on the possible relevance of a larger-scale study.
Surgical procedure
On the day of surgery, a flip of a coin determined the side receiving PRGF (PRGF System 1, Vitoria, Spain) in combination with DBB (Bio-Oss/Geistlich Biomaterials, Switzerland); the contralateral side received only DBB. Briefly, MSFA was performed with a lateral window approach, where after osteotomy, the lateral bone window was lifted upwards together with the Schneiderian membrane very carefully separated from the bone, and the bone graft material was placed into the newly created space (Fig. 1). All 12 sinuses in this pilot study were grafted with cancellous DBB particles 0.25 to 1 mm in size.
PRGF preparation
Twenty milliliters of peripheral blood was collected by venipuncture directly into tubes (BTI blood collecting tubes, BTI Vitoria, Spain) containing 3.8% (wt/vol) sodium citrate as anticoagulant. The blood was then centrifuged at 1400 rpm for 8 min at room temperature (PRGF System1, Vitoria, Spain), and thus separated into its three basic components: red blood cells (at the bottom of the tube); PRGF (in the middle of the tube); and plasma poor in growth factors (at the upper part of the tube). The 0.5 ml PRGF fraction located just above the red cell fraction was collected, with care being taken to avoid the overlying buffy coat, and deposited in an Eppendorf tube. Clotting and activation were initiated by adding 50 μl calcium chloride solution (10% w/v) to the Eppendorf tube. The activated PRGF was then mixed with 1 cc of DBB in a glass dish, and after 5–8 min the material became viscous and was ready for use (Fig. 2).
Infection and pain control
All patients were prescribed systemic antibiotics (2 g amoxicillin with clavulanic acid) starting 1 day before surgery and for 6 days post-surgically. Dexamethasone (8 mg) was prescribed and administered orally right before the surgery and for the following 3 days with a decreasing dose (8, 4, and 2 mg respectively). Analgesics were also prescribed (600mgr ibuprofen) for pain control. The patients were then asked to describe the post-operative discomfort as no pain discomfort, pain controlled with painkillers, or great pain.
Histologic and histomorphometric analyses
Cylindrical biopsies were retrieved at the time of implant placement by means of trephine burs with an external/internal diameter of 3 mm/2 mm, about 6 months after MSFA (Fig. 3). Immediately after harvesting, the trephines containing the tissue cores were placed in 70% alcohol for fixation. The cores were code-masked to facilitate blind histological assessment. After 2 weeks, the biopsy core was removed from the trephine, whenever possible; otherwise, the entire trephine-biopsy complex was further processed for non-decalcified sectioning, including dehydration in a series of increasing concentrations of ethanol and embedding in methylmethacrylate. Longitudinal sections were generated with a cutting-grinding technique and then stained with van Gieson’s picro-fuchsin. Histological and histomorphometrical analysis was performed while viewing the most central section of each biopsy under a microscope with incandescent light (BH-50, Olympus, Ballerup, Denmark) fitted to a video camera (Olympus DP70, Olympus). First, the margin between pristine bone and newly formed tissue inside the sinus was histologically identified, and then the area fractions (%) of newly formed bone (mineralized tissue and bone marrow), soft connective tissue, residual biomaterial, empty spaces, and debris were estimated by a semi-automated technique based on color segmentation through the software (Adobe Photoshop; Adobe) (Fig. 4).
Summary statistics were used to describe the data, and the two-sided Wilcoxon test for paired observations was used to evaluate differences in the various tissue components between the two groups. The significance level was set at P < 0.05. The SPSS 13.0 software (SPPS Inc., Chicago, IL, USA) was used for the statistical analysis.