1 Introduction

Periodontal disease is a chronic inflammatory disease process that eventually leads to loss of periodontal attachment and an eventual, bone destruction. The objective of periodontal therapy is to regenerate the lost periodontal tissues. However, periodontal regeneration requires a sequence of biological events including cell adhesion, migration, proliferation and differentiation (Giannobile, 1996). A combination of growth factors may more effectively stimulate formation of mineralized as well as non-mineralized tissues (Lynch, 1994). Platelets are rich in growth factors that may contribute to an accelerated process of tissue regeneration. During the early stages of wound healing, platelets released growth factors, including platelet derived growth factor, insulin like growth factor-1 and transforming growth factor-β, initiate a cascade of cellular and molecular events that result in wound healing in a highly regulated and coordinated manner (Lynch, 1994). The application of these growth factors to bone and periodontal regeneration has been investigated using numerous in-vitro and in-vivo models with promising results (Pierce et al., 1991; Hollinger et al., 2008; Kaigler et al., 2011; Kao and Fiorellini, 2012). Xenografts used in the treatment of infra-bony defects can be both bovine bone and natural coral; these, also, being referred to as anorganic bone graft materials, since they are suggested to remove all cells and proteinaceous material leaving behind an inert, bioresorbable, bone scaffolding, upon which re-vascularization, osteoblast migration and woven bone formation supposedly occur (Sowmya et al., 2009). Demineralised bone matrix (DMBM) xenograft is a bone inductive, sterile, bioresorbable, xenograft composed of Type I collagen, prepared from bovine cortical samples, resulting in non-immunogenic, flowable particles, of approximately 250 μm dimensions, that are completely replaced by host bone in around 4-24 weeks (Blumenthal et al., 1986). Platelet-rich fibrin (PRF) described by Choukroun et al is a second-generation platelet concentrate which allows one to obtain fibrin membranes enriched with platelets and growth factors after starting from an anti-coagulant-free blood harvest without any artificial biochemical modification (Choukroun et al., 2000). The PRF clot forms a strong natural fibrin matrix that concentrates almost all the platelets and growth factors of the blood harvest and shows a complex architecture as a healing matrix including mechanical properties that no other platelet concentrate offers (Choukroun et al., 2000; Dohan et al., 2006). Also, as an autologous biomaterial, it has found numerous clinical applications that have been described in length in the literature (Choukroun et al., 2006; Choukroun et al., 2006; Diss et al., 2008; Mazor et al., 2009; Zhao et al., 2011). Herewith, we are presenting a case report of a combined endo-perio lesion that was treated successfully by a combination of autologous PRF with bovine derived xenograft, and that was assessed clinically and radiographically with a 12-month follow-up.

2 Case Report

A 35 year old patient reported to the department with a chief complaint of swelling in the upper front tooth region since 2 months. The patient had reported trauma in the same region 7 years back. The nature of the pain was intermittent in nature. The patient used to take analgesics to relieve the pain. The past medical history of the patient did not reveal any significant finding and the patient was systemically healthy and was not on any medication. Patient revealed no history of smoking and alcohol and other deleterious habits. On examination, there was a swelling of about 3cm x 3cm with Ellis class IV fracture in relation to 11. The tooth appeared to be discolored. (Figure 1) The pulp vitality tests revealed that the tooth was non-vital while the radiographic examination of the concerned area in the form of an intra-oral peri-apical radiograph (IOPAR) revealed that it had a large peri-apical radiolucency. (Figure 2) Periodontal probing revealed a deep periodontal pocket measuring about 15 mm in depth. (Figure 3) The provisional diagnosis arrived-at was a combined endo-perio lesion in relation to 11 region. Endodontic Therapy: At the initial visit, immediate emergency access opening was done and the pus was drained-out. The access was kept open and only a thick cotton pledget was used to close the access for a day. Next day, the patient was recalled and the cotton pledget was removed and the canal was re-irrigated with saline. The swelling had reduced. Patient was recalled for irrigation and dressing after 4 days till which time, the swelling had completely resolved. Working length and biomechanical preparation were completed with the tooth. (Figure 4) Calcium hydroxide dressing was given for 7 days and the access was closed with a temporary restoration. (Figure 5) A dressing was given again and the patient was recalled after 7 days for the follow-up and then, the patient was transferred to Department of Periodontics for opinion and needful. A thorough oral prophylaxis was done in the concerned region (Figure 6) and the patient was recalled for completion of endodontic treatment. (Figure 7)

Figure 1 Showing discolored, non-vital 11 with Ellis class IV fracture

Figure 2 Showing IOPAR revealing a large peri-apical radiolucency

Figure 3 Showing probing depth of around 15 mm as measured with the help of a periodontal probe

Figure 4 Showing working length determination as seen on IOPAR

Figure 5 Showing calcium hydroxide dressing with access opening closed with a temporary restoration as seen on IOPAR

Figure 6 Showing oral prophylaxis done in relation to maxillary arch

Figure 7 Showing completed endodontic treatment in relation to 11

Periodontal therapy: A periodontal surgery was planned after 1 month of the completion of the endodontic treatment. Routine blood investigations were advised for which the reports were found to be normal. Pre-surgical periodontal pocket depth was reduced to 7 mm. (Figure 8) After proper isolation of the surgical field, the operative site was anaesthetised using 2% xylocaine hydrochloride with adrenaline (1: 200 000). A crevicular incision was given covering 12, 11, 21 region and a full thickness muco-periosteal flap was raised. (Figure 9) The infected, necrosed bone was removed and a continuous apico-marginal defect was observed along with buccal wall with dehiscence. 3 mm of root was resected for apicoectomy and 3 mm gutta percha was removed from the apex with the help of a heated probe. (Figure 10)

Figure 8 Administrative map of Nigeria (above) showing the Hydrological map of the Lagos lagoon complex. The sample stations A & B are indicated on the Badagry axis of the lagoon

Figure 9 Showing crevicular incision given in relation to 12, 11, 21 region with raised full thickness mucoperiosteal flap

Figure 10 Showing removed infected, necrosed bone with a continuous apico-marginal defect being seen along with buccal wall with dehiscence and 3 mm of root resected for apicoectomy with 3 mm gutta percha removed from the apex with the help of a heated probe

PRF preparation: Blood sample of the patient was drawn in 10 mL test tubes without an anti-coagulant and centrifuged immediately. Blood was centrifuged using a tabletop centrifuge (REMY Laboratories, Chennai, Tamilnadu, India) for 12 min at 3000 rpm. The resultant product consisted of the following three layers (Figure11):

• the upper layer of acellular PPP (platelet-poor plasma);

• PRF clot in the middle;

• Red blood cells at the bottom.

​  Figure 11 Showing blood sample of the patient being drawn in 10 mL test tubes without an anti-coagulant for centrifugation

Platelet rich fibrin was then separated by cutting with a sterile surgical scissor and collected using a dappen dish. (Figure 12) PRF clot was placed inside the periapical defect. (Figure 13) A demineralised bone matrix (DBM) xenograft was then placed over the peri-apical defect and also, the entire root length devoid of bone. (Figure 14) A Collagen membrane was finally placed over the defect. The Collagen membrane was cut according to requirement and then placed over the defect using 4-0 vicryl resorbable suture. (Figure 15) The flap was sutured with 3-0 black braided silk. (Figure 16) A perio pack was then mixed and placed over the sutured area so that the repositioned flap be immobile till healing ensued. (Figure 17) Immediate post-operative radiograph (IOPAR) was advised. (Figure 18) Antibiotics and analgesics were prescribed for one week. After 15 days, the pack was removed. The sutures were removed and the healing was found to be uneventful. The crown preparation with 11 was done (Figure 19) and elastomeric impression was recorded. (Figure 20) All Ceramic Emax crown was fabricated and cemented. (Figure 21) The patient was asked for regular follow-up visits and the 12 months post-operative radiograph was taken which revealed apparent bone fill with resolution of the osseous defect. (Figure 22) Clinical evaluation of the periodontal pocket depth revealed a successful outcome of the procedure with a considerable reduction in pocket depth that got reduced from 15mm to 3mm depth (Figure 23).

Figure 12 Showing PRF clot in the dappen dish

Figure 13 Showing PRF clot placed inside the peri-apical defect

Figure 14 Showing demineralised bone matrix (DBM) xenograft placed over the peri-apical defect and the entire root length devoid of bone

Figure 15 Showing collagen membrane placed over the defect

Figure 16 Showing sutured flap with 3-0 black braided silk

Figure 17 Showing perio pack over the sutured area to make the repositioned flap immobile till it heals

Figure 18 Showing immediate post-operative radiograph

Figure 19 Showing crown preparation in relation to 11

Figure 20 Showing elastomeric impression registered in relation to 11

Figure 21 Showing all Ceramic Emax crown cemented in relation to 11

Figure 23 Showing pocket depth reduced from 15mm to 3mm

3 Discussion

The treatment of combined endo-perio lesions requires both endodontic therapy and periodontal regenerative procedures, as discussed in the above case report. The goal of peri-apical surgical procedures is to remove all the necrotic tissue from the peri-apical area to completely seal the pulp canal system to facilitate the regeneration of hard and soft tissues including the formation of a new attachment apparatus (Karabucak and Setzer, 2009). Many a times, there is no clear insult to the pulp noted in these types of lesions. The most common clinical/radiographic features of these lesions include peri-apical radiolucencies and deep pocket depths associated with a non-vital pulp. Traditional approaches to treat endo-perio lesions include non-surgical debridement of the root surfaces and pulp canals, as well as surgical approaches that provide better access to clean the root surfaces and apical lesions and to re-shape the surrounding bone/root apex. Bone loss caused by pulpal disease is reversible, whereas advanced bone loss caused by periodontal disease is usually irreversible (Law and Beaumont, 2004). The necessity of periodontal surgical therapy most likely remains because the periodontal bone loss is usually more advanced and is less likely to resolve after non-surgical pulp canal therapy alone (Meng, 1999). Generally, partial apical root resection has been suggested for all endodontic surgeries advised for extensive involvements (Bashutski and Wang, 2009). In this case, we had planned a regeneration therapy for bone as well as the periodontal ligament as it was supposed to lead to a better prognosis. Hence, we performed pulp canal debridement with subsequent retrograde filling and removal of granulation tissue around the root apex. Apicoectomy was done to prevent recurrence of infection as the maximum number of canal variations and complexities are seen in the apical third (Siqueira et al., 1997). PRF i.e. Platelet rich fibrin, is in the form of a platelet gel that is used in conjunction with bone grafts offering several advantages including promoting wound healing, bone growth and maturation, graft stabilization, wound sealing and haemostasis, and improving the handling properties of graft materials. It is saturated with growth factors which expediate the regenerative process during healing (Choukroun et al., 2006). Also here, using the guided tissue regeneration (GTR) technique, combined with bone graft, periodontal regeneration was clinically and radiographically evident after a 1-year follow-up. The role of bone graft in the above case was to provide space for periodontal regeneration and for inducing bone formation and attachment gain. The rationale for using GTR membrane in the above case with bone graft material was to encourage the growth of key surrounding tissues, while excluding unwanted cell types such as epithelial cells (Oh et al., 2009). GTR therapy has been implemented in endodontic surgeries as a concomitant treatment procedure during the management of combined endo-perio lesions. However, from clinical and radiographic findings, there was a significant reduction in probing depth and bone fill. Although traditional non-surgical periodontal therapy and regular endodontic therapy can be predictably used to arrest mild to moderate periodontal bone defects in combined endo-perio lesions, it might be inadequate for the treatment of diseases characterized by deep pockets and/or apico-marginal defects. Currently, regenerative techniques are widely available in terms of their predictability to regenerate the lost tissue/bone in all types of defects and for all situations.

4 Conclusion

A careful pre-operative diagnosis, appropriate case selection and knowledge of the factors that can negatively affect regeneration outcomes can help to optimize successful regenerative attempts. Treatment strategies used in this case report suggest that combined endo-perio lesions can be successfully managed with combined regenerative procedures.

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