Comparative evaluation of clinical efficacy of β-tri calcium phosphate (Septodont-RTR)™ alone and in combination with platelet rich plasma for treatment of intrabony defects in chronic periodontitis.

AIM: To assess the clinical outcome by comparing β-tri calcium phosphate (Septodont RTR)™ along with platelet rich plasma (PRP) and β-tri calcium phosphate (β-TCP) alone in intrabony defects, by clinical evaluation in a 6-month analysis.
METHODOLOGY: Ten patients participated in the study. Using a split-mouth design, interproximal bony defects were surgically treated with either platelet rich plasma (PRP) combined with β-tri calcium phosphate (β-TCP) or β-TCP alone. Plaque Index (PI), Gingival Index, Probing Pocket Depth (PPD), Clinical Attachment Level (CAL) were recorded 6 months after surgery.
RESULTS: At 6 months after therapy, the PRP combined with β-TCP group showed mean PPD reduction of 2.50, CAL mean gain of 2.60 + 1.43. The β-TCP group showed mean PPD reduction of 2.80 mm, mean CAL gain of 2.60 mm. On intra-group comparison, there was greater PPD reduction and CAL gain at 6 months in both the groups. In intergroup comparison of PRP/β-TCP and β-TCP alone, there was no statistical significant difference observed. (P = 0.55, and 0.87 for PPD and CAL gain).
CONCLUSION: Both therapies resulted in significant PPD reduction, CAL gain. The present study shows that treatment of intrabony periodontal defects with combination of PRP and β-TCP does not have additional improvements when compared with β-TCP alone within 6 months follow-up.

INTRODUCTION

Periodontitis is the inflammation of the supporting tissues of the teeth. It is usually a progressively destructive change leading to loss of bone and periodontal ligament.[1] When periodontal disease has caused a loss of the attachment apparatus, optimal care seeks to regenerate the periodontium to its pre-disease state. Intraosseous periodontal defects of varying morphology may have a varying regenerative potential depending on the extent of the source of the cells from the periodontium.[2]

The therapeutic modalities that attempt to enhance these biologic events such as cell migration, adherence, growth and differentiation have the potential to increase the success and predictability of periodontal regenerative procedures.[34]

The combination of various regenerative biologic agents and techniques has attracted the interest of researchers in the field of reconstructive periodontal surgery. The periodontal regeneration requires an orchestrated sequence of biological agents and techniques for its successful outcome.[56] Autogenous bone grafting is considered the gold standard for grafting, although it has limitations.[7]

Traditional synthetic bone grafts are ceramics of hydroxyapatite, tricalcium phosphate (TCP) or combination of two which are resorbable materials. Among the tested bone substitutes, TCP showed significantly higher percentage of bone fill at 24 weeks of healing.[8] It is well tolerated and has no adverse affects such as allergic reactions.[9]

Polypeptide growth factors (PGFs) are biological mediators that have the ability to regulate cell proliferation, chemotaxis and differentiation. Of all known PGFs, platelet-derived growth factor (PDGF) and transforming growth factor (TGF-β) has shown to exert a favorable effect on periodontal regeneration.

Platelets are very important in the wound healing process, and the alpha granules in them are the major source of PDGF and TGF-β. A convenient approach to obtain autologous PDGF and TGF-β is the use of platelet rich plasma (PRP). Researchers reported successful bone regeneration, and bone fill was achieved using PRP in periodontal and non-periodontal sites.[10] However, human clinical studies using PRP in periodontal therapy are limited. Since β-tri calcium phosphate (β-TCP) is tolerated well and has no adverse effects such as allergic reactions, it is of biologic and clinical interest to use β-TCP as a bone substitute in conjunction with PRP. It might be hypothesized that β-TCP may resorb, thus allowing the newly forming periodontal tissues to fill the available space.[11]

However, the studies on the stability and potential changes in the regenerated tissue are missing, and there is still a question about how PRP affects the quality of the new tissue. Therefore, we hypothesized that using a normally slow bioabsorbing material, such as β-TCP as a graft material to deliver PRP could reveal how much of the regeneration was attributable to PRP. The aim of the present study was to assess the clinical outcome by comparing a newly introduced bone graft material β-TCP (Septodont RTR)™ along with PRP and β-TCP alone in intrabony defects, by clinical evaluation in a 6-month analysis.

MATERIALS AND METHODS

Patient selection: After ethical committee approval, patients were selected from the patient pool attending Vishnu Dental College and Hospital at Bhimavaram. Patients with greater than 30% of the sites having attachment loss were considered as chronic generalized periodontitis. 10 patients with chronic generalized periodontitis, with the presence of contralateral intrabony defects were participated in the study after obtaining informed consent.

Inclusion criteria

Patients with chronic periodontitis within 25-50 years age and requiring periodontal therapy.

Presence of at least two interproximal periodontal pockets on contralateral sides with probing pocket depth measuring ≥6 mm, with radiographic evidence of angular bone loss in the mandibular posterior region.

Contralateral intrabony defects, with defect depth ≥3 mm.

Exclusion criteria

Patients with shallow and narrow intrabony defects, history of systemic disease, smokers, antibiotics during last six months, poor oral hygiene, drug allergy, advanced gingival recession, intrabony defects extending into furcation area, pregnant and lactating women.

Study design: The study used a split mouth design, wherein two sites in the contralateral mandibular quadrants, which required periodontal treatment with probing pocket depth ≥6mm and intrabony defect depth ≥3 mm were chosen and randomly assigned for a single blind study. A total of 20 sites from 10 patients were selected for the study. Control sites were treated with open debridement followed by placement of β-tri calcium phosphate (β-TCP) alone, and experimental sites were treated with open debridement followed by placement of β-tri calcium phosphate (β-TCP) combined with autologous platelet rich plasma (PRP).

Septodont RTR™

RTR – resorbable tissue replacement is a gradually resorbable material made of β-TCP granules of synthetic origin. It is available as granules, with size ranging between 500 μm and 1 mm. The size of macropores varies from 100 μm to 400 μm and micropores are less than 10 μm in diameter. This specific structural property allows the colonization of macropores by newly formed bone. It is designed for filling and reconstruction of bone defects in maxillofacial and dental surgeries.

Pre-surgical procedure

All the selected patients, following an initial examination and treatment planning discussion were given detailed instructions about self-performed plaque control measures and were subjected to Phase I therapy consisting of meticulous scaling and root planing. Four weeks after phase I therapy, the oral hygiene status and tissue response was evaluated using Plaque Index[12] and Gingival Index.[13] A complete periodontal examination was carried to confirm the need for periodontal surgery.

Clinical examination

Clinical examination was carried out at baseline and 6 months. Periodontal status of the patient was evaluated using Plaque Index (PI), Gingival Index (GI), Probing pocket depth (PPD) and Clinical Attachment Level (CAL). The PPD and CAL were recorded using customized acrylic stents with grooves to ensure a reproducible placement of the calibrated University of North Carolina No. 15 periodontal probe§ to the nearest mm in the defect specifically. PPD measurements were obtained at six sites (mesiobuccal, mid-buccal, distobuccal, mesio lingual, mid lingual/palatal, disto lingual) by using UNC-15 periodontal probe.

The deepest sites with PPD ≥ 6 mm, with radiographic evidence of angular bone loss have been selected. In the defect area, the measurements were taken using customized acrylic stents [Figure 1].

Figure 1

Measuring the PPD and CAL with UNC 15 probe at baseline

CAL was determined by measuring the distance from the Cemento Enamel Junction (CEJ) to the base of the gingival sulcus with an UNC-15 periodontal probe. The CEJ determination was made by using the anatomical CEJ or the apical extent of the existing restoration margin on each tooth.

PRP preparation

On the day of surgery on the experimental sites, 9 mL of blood was drawn from each patient by venipuncture of the antecubital vein. Five milliliter of patient's blood was collected in plastic test tube and was shaken gently to enhance complete mixing of blood with anticoagulant. The tube was then kept for centrifugation at 2400 RPM for 10 min. This spinning produced a top layer of Platelet Poor Plasma (PPP), which was separated from erythrocytes, platelets and leukocytes. The PPP was discarded and the remaining sample was further spun for second centrifugation at 5000 RPM for 15 min. This second cycle helps to concentrate the platelets. After second centrifugation, the supernatant of about 0.5 mL nearest to the RBC layer was collected using Pasteur pipette without being contaminated by RBC. Before using the PRP for clinical application, it has to be activated to form gel. The collected PRP was taken into a sterile dappen dish [Figure 2] and 0.2 mL of serum of the patient and an equal volume of 10% calcium chloride is added.[14]

Figure 2

Platelet rich plasma to combine with β-tri calcium phosphate

Surgical procedure: On the completion of the baseline examination, the infrabony defects were randomly assigned as experimental and control sites. Under local anesthesia, full thickness mucoperiosteal flaps were reflected on both buccal and lingual sides extending at least one tooth mesial and distal to the tooth with the infrabony defect. After debridement, cotton pledgets soaked in saline mixed with tetracycline powder were applied on the root surface and left for 2-5 minutes of time. Normal saline was used to irrigate the root surface after removing the cotton pledgets. In the control group β-TCP (Septodont RTR™) was combined with physiological saline solution and was condensed to fill the defect [Figure 3]. In the experimental site, the β-TCP granules were mixed with PRP in a sterile dappen dish and the mixture was properly condensed in the test site to the level of the surrounding bony walls [Figure 4]. Care was taken not to overfill the defects. In both the groups, following surgery the mucoperiosteal flaps were repositioned and secured in place by interrupted suturing using 3-0 black silk suture material. The surgical area was protected and covered with periodontal dressing.

Figure 3

Placement of β-tri calcium phosphate in the defect

Figure 4

Placement of β-tri calcium phosphate in combination with PRP in the defect

Postoperative care

All subjects received postoperative analgesia (Combiflam: Ibuprofen 400 mg, Paracetamol 325 mg tid for 5 days) and antibiotics (Amoxicillin 500 mg tid for 5 days). Periodontal dressing and sutures were removed 14 days after the surgery. Recall appointments were scheduled weekly for the first 6 weeks after surgery and once per month during the remainder of the observation period of 6 months. The recall appointments consisted mainly of reinforcement of oral hygiene measures and professional supragingival tooth cleaning.

Postsurgical measurements: PI, GI, were recorded at 3, 6 months after initial surgery. Soft tissue measurements were repeated with the previously used acrylic stents [Figure 5].

Figure 5

Measuring the PPD and CAL with UNC 15 probe after treatment

The obtained data were statistically analyzed using Microsoft Excel SPSS software version 16. Variables within the group were compared by paired t test and variables between different groups at baseline and 6 months were compared by unpaired t test.

RESULTS

The study evaluated and compared the clinical efficacy of β-TCP bone replacement graft alone, and in combination with PRP. Over the course of the study, there were no infectious episodes and no other adverse complications in either test or control sites. The soft tissue response in both control and study groups was excellent clinically.

The mean PI scores of the control group at baseline was 1.23 ± 0.22, reduced to 0.70 ± 0.44 after 6 months. Similarly the mean PI scores of the study group at baseline was 1.25 ± 0.31, reduced to 0.40 ± 0.67 after 6 months. On comparison, this mean reduction of the plaque scores was statistically significant (P < 0.001) as shown in Table 1.

Table 1

Comparison of mean plaque index scores from baseline to 6 months in control and study groups

The mean GI scores of the control group at baseline were 1.10 ± 0.44, reduced to 0.18 ± 0.24 after 6 months. Similarly the mean GI scores of the study group at baseline were 1.15 ± 0.41, which reduced to 0.13 ± 0.18 after 6 months. On comparison this mean reduction of the GI scores was statistically significant (P < 0.001) as shown in Table 2.

Table 2

Comparison of mean gingival index scores from baseline to 6 months in control and study groups

When the mean reduction of the PI (P = 0.20) and GI (P = 0.23) scores after 6 months were compared between control and study groups as intergroup comparison, there was a statistically insignificant value as shown in Table 3.

Table 3

Comparison of plaque index and gingival index scores between study and control groups

The mean PPD of the control group at baseline was 7.40 ± 1.65, which reduced to 4.60 ± 1.17 after 6 months. Similarly the mean PPD of the study group at baseline was 6.60 ± 0.70, which reduced to 4.10 ± 0.99 after 6 months. On comparison, this mean reduction of PPD was statistically significant in both the groups (P < 0.001) as shown in Table 4.

Table 4

Comparison of mean probing pocket depth from baseline to 6 months in control and study groups

The mean CAL in control group at baseline was 7.60 ± 1.65, which reduced to 5.00 ± 1.25 after 6 months. Similarly the mean CAL in the study group at baseline was 6.70 ± 0.82, which reduced to 4.20 ± 1.14 after 6 months. On comparison this mean difference of CAL was statistically significant (P < 0.001) as shown in Table 5.

Table 5

Comparison of mean clinical attachment level at baseline and 6 months in control and study groups

When the mean difference of PPD (P = 0.55) and CAL (P = 0.87) after 6 months were compared between control and study groups as intergroup comparison, there was statistically insignificant difference as shown in Table 6.

Table 6

Comparisons of mean difference of probing pocket depth and clinical attachment level between control and study groups

DISCUSSION

The ultimate goal of the regenerative periodontal therapy is complex and involves regeneration of lost attachment apparatus. The β form is the most stable resembles human cancellous bone in structure and composition.[15] Therefore in the present study, β-TCP has been selected as bone replacement graft to treat intrabony defects.

Periodontal regeneration therapies needs bone augmentation for a good attachment and PRP has been suggested to increase this rate of bone deposition and quality of bone regeneration when used in conjunction with bone grafts. Hence in the present study, PRP was used in combination with β-TCP for osseous grafting at the defect sites. According to the protocol suggested by Tozum and Demirlap,[14] PRP was prepared by using patient's blood after obtaining through venipuncture and processed to obtain PRP. Later it was activated by using patients own serum to obtain a gelatinous form.[16] Autologous PRP activated by the patients own serum has the advantage of eliminating the risk for disease transmission as well as preventing the hypersensitivity reactions.[17] Hence autologous serum was used in our present study for the process of activation. Treatment of intrabony defects using bone replacement grafting alone proved to be successful to achieve bone fill but a residual defect usually exists.[18] So in our present study, PRP was used in combination with β-TCP to evaluate its efficacy in the treatment of intrabony defects.

The present study showed a significant reduction in the mean PI scores and GI scores in both the control and study groups after 6 months postoperatively, which shows that soft tissue healing was clinically good in both the groups. The mean reduction PI and GI scores compared between control and study groups showed statistically insignificant difference after 6 months. These results suggest that the application of PRP in adjunct to β-TCP did not reveal any additional benefit in the reduction of clinical parameters like the PI and GI. These results are concomitant with earlier studies.[1920] This could be because of the regular and frequent recall visits in which the patients underwent regular supragingival scaling, which further minimized plaque accumulation.

The results of the study demonstrated that treatment of intrabony periodontal defects with the combination of platelet rich plasma and β-TCP, and β-TCP alone, can lead to significant clinical improvements in terms of PPD reduction and CAL gain, although statistically insignificant difference were found when the two treatment modalities were compared. The lack of adverse reactions such as allergies, abscesses, or rejection of the implanted materials suggested that all used materials were well tolerated. These observations corroborated those from previous studies, which have failed to show that any of the materials used may elicit any allergic or foreign body reaction.[21]

Limited number of studies have been documented so far to evaluate the effects of PRP in combination with β-TCP in the treatment of intrabony defects.[22232425] Studies evaluating the effect of PRP with other materials or in combination with GTR alone showed a significant positive effect on the treatment of periodontal intrabony defects.[21242526] However the results of these studies cannot be compared due to different study designs.

In our study only clinical parameters were analyzed since it was hypothesized that the use of PRP in conjunction with β-TCP may accelerate the rate of bone formation rate, and due to the limitations of ethical considerations, histological samples were not be obtained in this study in order to demonstrate if the β-TCP particles were osseointegrated or encapsulated by connective tissue.

Reduction in PPD, gain in mean CAL after 6 months postoperatively were highly significant in both the study and the control groups. The results shows that PRP in combination with β-TCP showed comparable clinical outcomes in the treatment of intrabony defects, which is concomitant with other studies reported in the literature.[1922] However, the results did not show any significant improvement over β-TCP when used alone in the defect. No statistically significant difference was found after 6 months when both study and control groups were compared. The explanations of the reason for the lack of additive effect of PRP will be speculative due to the limitations of the present study in which we did not test/detect the potential mechanisms of PRP for bone formation. The study sample is within an acceptable range for a certain amount of periodontal regenerative studies in humans.[27]

The results of our study suggested that PRP combined with β-TCP and β-TCP alone are effective treatment modalities, but combination of β-TCP and PRP does not have any additional effect on the clinical parameters for treatment of intrabony defects in humans. These results were compared well with those of two recent randomized controlled studies evaluating the additional effect of PRP on the healing of bone graft, and suggested that PRP did not have additional influence on periodontal and bone regeneration.[1922]

Additional studies with a large sample size, longer duration of follow up may be needed to detect an eventual difference between the treatments.

CONCLUSION

The present study indicated that treatment with β-TCP alone and in combination with PRP resulted in PPD reduction, CAL gain in the due course of treatment. Within the limits of the study and based on the results obtained the following conclusions can be drawn:

On intragroup comparison there was significant reduction in PPD and gain in CAL after 6 months postoperatively in both study and control groups

On intergroup comparison there was no significant difference between study and control groups in terms of reduction in PPD and gain in CAL after 6 months

The present study shows that treatment of intrabony periodontal defects with combination of PRP and β-TCP doesn’t have additional improvements in the clinical parameters measured within 6 months follow up.

Overall the internal validity of the evidence might be judged as moderate. However long-term studies and larger sample may be useful to determine the efficacy of PRP in combination with β-TCP.