A step by-step procedure for bone regeneration using calcium

1. Antonello Maria Messina, DDS, Luca Marini, DDS,y Daniel S. Oh, MS,
PhD,z§
and Ettore Marini, MD, DDS
The Journal of Craniofacial Surgery , January 2019

2.  Osseointegrated dental implants are an advancement in dentistry and have
become a reliable option for the rehabilitation of partially or totally edentulous
patients, with very high success rate
 However, for a predictable long-term prognosis after such a procedure, a
potential implant recipient site must possess a sufficiently healthy volume of
bone
 Transplantation of bone tissue in a recipient site that is deficient in bone can
replace the missing skeleton or guide its reconstruction, or both.
 Different types of bone can be transplanted, for example, autologous,
homologous, or heterologous, compact or spongy, whole or without organic
substance, vascularized or not, live or dead, and undecalcified or decalcified

3.  The traditional ‘‘criterion -standard’’ transplantation is an autologous graft.
 Unlike both homologous and heterologous bone transplantation, an autologous
graft does not induce immunological reactions or cause post-implantation
resorption.
 However, the amount of autologous bone necessary for transplantation may not
be available.
 Autologous bone is also difficult to shape, which is required for its adaptation .
 Autografts must be harvested from a separate patient body donor site and,
therefore, 2 surgical procedures are required.
 Thus, autograft transplantation increases the risk of donor site pain, bleeding,
and infection

4.  HA is the main mineral component of bone and teeth and is well known for its
ability to promote bone formation.
 HA granules are used for filling bone defects, but they have poor handling
qualities and retention at the surgical site, leading to graft voids between the
granules and bone tissue and resulting in mechanical instability.
 Furthermore HA implantation is not constantly followed by favorable results,
especially when it is carried out for augmentation of the alveolar ridge
 The Marini technique (MT) is a bone regeneration procedure developed 25 years
ago by one of the authors (E.M.). Here, they have described how to create the
ideal bone recipient site and the composite graft preparation using the Marini
technique.
 Although similar techniques using HA in vivo have been reported in other
experimental or clinical investigations, the MT emphasizes surgical site
preparation using a piezoelectric device to remove necrotic debris and the use of
a mixture of coralline HA granules (CHAG) and fibrin glue
 This article provides a 2-year follow-up with histological analysis of newly formed
bone.

5.  Patients –this study was a retrospective analysis of patients who were
consecutively treated for bone regeneration using the MT.
 All patients were treated at a private clinic in Rome, Italy.All surgical procedures
were performed by both authors (E.M. andA.M.M)
 They enrolled a group of 20 randomly selected patients (12 women and 8 men).
 All patients were treated for 2 years
 The patients enrolled in this study required bone regeneration to
achieve the necessary bone volume for future implant placements.
 The exclusion criteria for our study were smoking, alcohol consumption,
uncontrolled systemic conditions, or uncontrolled periodontal disease

6.  Before any grafting, recipient-site preparation is required to allow access to the
underlying healthy bone structure, which is critical for the optimal ossification of
the CHAG and HFG complex.
 Basic principles for flap design were followed
 All necrotic and inflamed portions of both hard and soft tissues were carefully
removed.
 Removing these unhealthy tissues permits in situ plasmatic imbibition for the first
24 to 48 hours, resulting in vascular ingrowth and cellular nutrition.
 The MT to prepare the recipient site involved an initial debridement based on
with osteoplasty inserts to remove the necrotic and inflamed tissues
 The surgical field was then washed and cooled with a sterile saline solution.
 Additional meticulous debridement of the operative site was performed using
curettes, excavators, and a Beaver blade.

7.  During the debridement, several lavages with 10% hydrogen peroxide (H2O2)
were performed.
 After each lavage with H2O2, the site was rewashed with saline solution so that
the bone and soft tissue were not overly exposed to the caustic action of the
H2O2.
 To enhance bone regeneration, bleeding at the recipient site is essential.
 With this manner, the graft materials will be fertilized with cells and stromal
elements that were derived from host tissue.
 Hence, the recipient site was decorticated using piezoelectric device with an
insert for osteotomy, if needed

8. (A)A preoperation view shows the mandibular first and second
molars. (B) Pantomography shows an osteolytic lesion in the right mandible. (C)
An intraoral view shows necrotic tissue. (D) Piezosurgery to clean the surgical
site is shown.

9.  Bone regeneration was carried out using porous CHAGs with a mean pore size of
approximately 200Um
 Before surgical placement, the CHAGs were rapidly molded with the HFG Tisseel
 Tisseel is a 2-component tissue sealant that consists of sealer protein and
thrombin solutions.
 Tisseel contains fibrinogen and thrombin, 2 blood proteins that are important for
blood coagulation.
 A mixture of these proteins is used to form a clot at the application site.
 The clot that is formed usingTisseel is very similar to the clot formed during
normal blood coagulation.
 TheTisseel clot degrades similarly to an endogenous clot and does not leave any
residue.
 A synthetic protein (aprotinin) is added to the mixture to extend the life of the
clot and prevent premature degradation.
 The CHAG amount required for mixing with the fibrin glue depends on the width
of the area to be reconstructed.

10.  The granules were mixed using a mortar and a periosteal elevator.
 After careful, rapid mixing, the CHAG-HFG complex was placed Into the area that
was to be augmented, reconstructed, or both
 The working time of the mixture before the glue was completely set was 4 to 5
minutes. However, the grafting preparation could be repeated to properly fill each
bone defect; the mixture can be stratified without any inconvenience.
 Tweezers were used to transport the mixture from the mortar, and a periosteal
elevator was used to shape the graft.
 Practically, the unique biological and adhesive properties of the CHAG-HFG
complex are particularly advantageous because they allow the creation of a
compound that incorporates the granules to ultimately form a texture similar to
silicone, which is easy to shape and position.

11.  This feature is extremely important, considering that surgeons frequently
work in an antigravity position (eg, on the maxilla), need to rebuild thin
bony ridges, or fill small areas that are not easily accessible. Regardless of
these conditions, such surgery is always performed in the presence of
bleeding that is often difficult to suspend and requires the use of a surgical
aspirator.
 The aspiration commonly increases the difficulty in managing the surgical
procedures, such as the frequent aspiration can lead to dispersion of
granular biomaterials from the recipient sites.
 In contrast, the CHAG-HFG mixture, aided by the adhesive properties of
the fibrin glue, enables surgical aspiration without the removal of the
granule from the recipient site.

12. 2. (A)The site is grafted with a mixture of homologous fibrin glue and coralline hydroxyapatite
granules. (B) Primary closure of the wound is shown.
(C) A 9-month post-operation pantomography shows newly formed bone in the reconstructed
area. (D) A 9-month post-operation intraoral view of the reconstructed site.

13.  Primary closure of the flap with sutures was necessary; however, the use of fibrin
glue was extremely effective as a means of fixing the tissues after oral surgery, as
HFG is an unusually strong sealer.
 Moreover, although inflammation occurs around sutures, the fibringlue promotes
an early healing of the surgical sites.
 Notably, in esthetic zones, the results from using both sutures and fibrin glue are
better than those from using sutures alone.
 The flap was sutured and then covere the CHAG-HFG graft with an additional
protective layer of HFG to protect the graft from bacterial contamination and
promote rapid healing.
 Postoperatively, they monitored the patient wound to prevent an exposure of the
grafted area.
 In cases of graft exposure, the area was washed with saline solution, and
dispersed granules were removed.
 After lavage, the wound was covered using fibrin glue to promote a faster closure
of the recipient site

14.  Radiographic analysis was performed before graft placement, and afterward,
both a physical examination and radiographic analysis were performed.
 After the ossification process was radiographically considered to be at an
advanced stage (ie, usually from 7 to 9 months after the grafting operation), each
patient underwent to a second surgery, and 1 or more titanium implants were
placed into the newly formed bone.
 During this procedure, they obtained a small cylinder of the regenerated bone
tissue using a trephine bur, and the sample was prepared for histological
examination.

15. (A)The final prosthesis of a case with a fixed bridge. (B) A 1-year post-operative x-ray image is
shown. (C) A low magnification cross-section of the specimen shows the relative proportion
of bone trabeculae and fibrillar connective tissue; D) Reconstituted bone tissue at 100
magnification showing primary bone trabeculae

16.  The surgical sites for all 20 patients healed within 1 week to 10 days, with minimal
inflammation; no infections were observed.
 The dispersion of a few CHAGs around the proximity of the wound was observed
during the first 5 days after graft placement in five cases.
 No sequela was associated with this condition, and the remaining CHAGs were
retained by reapplying the HFG to the wounds.
 Twenty to 30 days following graft placement, a pink mucosa covered the surgical
area, and no signs of inflammation were apparent.
 x-ray examination revealed a progressive increase in bone density.
 This increase was evident at 5 months after graft placement, and bone density
became similar to that of the adjacent bone at 7 to 9 months
 Microscopic analysis of the grafted area revealed a hard consistency with a rather
compact appearance that was resistant to trephine penetration.
 Trepanation of the graft removed a small osseous cylindrical core, leaving an
extremely small cavity with a sharp outline .

17. (A)After reflection of the flap, newly formed bone can be observed.
(B) A trephine bur was used to collect bony biopsy samples of the regenerated bone. (C)Two
implants being inserted into the regenerated bone are shown. (D) An immediate post-
operation intraoral x-ray shows 2 titanium implants in the regenerated bone.

18. A 5-year postoperative pantomography shows the long-term success of the procedure

19.  For a successful graft, an exact apposition of the transplant on the recipient bed
must be achieved, and this state must be maintained until the graft has become
firmly established
 HA implants can directly bond with the surrounding bone without the presence of
an intervening fibrous shell.
 This process accounts for a significant portion of the strength of the integrated
graft.
 This chemical bonding contributes to the excellent biocompatibility and, in large
part, to the clinical utility and long-term success of these implants.
 The MT focuses on preparing the recipient site, stressing the need to remove all
traces of pathological and necrotic tissue to facilitate the healing phase and
promote bone regeneration.
 A meticulous preparation of the surgical area creates ideal conditions for the
long-term success of the regenerative bone graft with predictable results.

20.  For the preparation of the surgical area, piezoelectric surgery is less traumatic to
hard and soft tissues than traditional pneumatic, or electric hand-drills are.
 Mechanical removal of pathological tissue improves the intraoperative visibility
of the surgical field, promotes rapid hemostasis, and is a safer procedure in
critical anatomical areas
 A piezoelectric device with oscillating low-frequency precision tips can be used
for optimal anatomical dissection of the interface between bone and soft tissue
 The physical, chemical, and biological properties of the CHAG-HFG complex
result in a bone graft that has a greater potential for success than using either
individual component alone
 The newly formed bone undergoes a remodeling process similar to that of
normal bone maturation.
 With the MT approach, neither membranes nor fixation screws are necessary to
stabilize the graft, as the mixture of the CHAG-HFG complex hardens in situ and
stabilizes in 2 to 3 minutes and, thus, perfectly adheres to the surrounding
tissues.

21.  Dental implants can be vulnerable to infection or extrusion because of a lack of
chemical or biomechanical compatibility or an incomplete cellular ingrowth of the
augmented bone.
 The ideal solution appears to be the use of an interconnected, coralline porous
implant (ProOsteon 200, Zimmer Biomet, IN ) that becomes ingrown with bone
and, subsequently, undergoes slow resorption with replacement by additional
regenerated bone.
 CHAG partially biodegrades with time in response to the chemical and
biomechanical mechanisms that influence normal bone remodeling.
 Nonetheless, the biological response to augmentation of the facial skeleton with
CHAG ultimately induces the formation of an implant consisting of host bone.

22.  The bone tissue was remodeled and, thus, the CHAG-HFG graft not only
efficiently replaced and reconstituted the missing parts of the skeleton but
also fully performed the mechanical functions of the reconstituted
segments
 The addition of HFG to CHAGs improved the handling of the granules
during surgery and prevented their dislocation and dispersion during the
postoperative period.
 Furthermore, the CHAG-HFG graft-induced bone regeneration results in
only a small amount of bone resorption following alveolar ridge
augmentation and, currently, no failure has been reported for any
recipient surgical site.

23.  Within the limitation of this study, it maybe stated that the presented bone
regenerative technique is an effective procedure, and is suitable for the
reconstruction of bone defects using a calcium phosphate scaffold.
 They demonstrated that recipient site preparation is a fundamental step toward a
successful grafting procedure.
 The accurate removal of necrotic and inflamed tissues from the recipient site
enhances the healing phase and promotes new bone formation.
 The addition of fibrin glue to HA granules transforms a granular, incoherent,
easily dislodged biomaterial into a moldable paste that can be easily handled,
adapted to, and maintained within, bone defects of every shape

24.  Thank you