Received: 15 October 2018 | Revised: 14 January 2019 | Accepted: 10 February 2019

DOI: 10.1111/jcpe.13086

RANDOMIZED CLINICAL TRIAL

Treatment of intrabony periodontal defects using rhFGF-­2

in combination with deproteinized bovine bone mineral or
rhFGF-­2 alone: A 6-­month randomized controlled trial

Atsushi Saito | Takahiro Bizenjima | Takahiro Takeuchi | Eiichi Suzuki |

Masahiro Sato | Kouki Yoshikawa | Yurie Kitamura | Daisuke Matsugami |
Hideto Aoki | Daichi Kita | Kentaro Imamura | Daisuke Irokawa | Fumi Seshima |
Sachiyo Tomita

Department of Periodontology, Tokyo

Dental College, Tokyo, Japan Abstract
Aim: To evaluate the use of recombinant human fibroblast growth factor (rhFGF)-­2 in
Correspondence
Atsushi Saito, Department of combination with deproteinized bovine bone mineral (DBBM) compared with rhFGF-
Periodontology, Tokyo Dental College, ­2 alone, in the treatment of intrabony periodontal defects.
Tokyo, Japan.
Email: [email protected] Materials and Methods: Patients with periodontitis who had received initial periodon-
tal therapy and had intrabony defects of ≥ 3 mm in depth were enrolled. Sites were
Funding information
This study was supported in part by a grant randomly assigned to receive a commercial formulation of 0.3% rhFGF-­2 + DBBM (test)
from Osteology Foundation (Advanced
or rhFGF-­2 alone (control). Clinical parameters and a patient-­reported outcome meas-
Researcher Grant No.17-­136) and a grant
from Multidisciplinary Research Center ure (PROM) were evaluated at baseline and at 3 and 6 months postoperatively.
for Jawbone Disease (MRCJD), Tokyo
Results: Twenty-­t wo sites in each group were evaluated. A significant improvement
Dental College (a MEXT Private University
Research Branding Project). in clinical attachment level (CAL) from baseline was observed in both groups at
6 months postoperatively. CAL gain was 3.16 ± 1.45 mm in the test group and
2.77 ± 1.15 mm in the control group, showing no significant difference between
groups. Radiographic bone fill was significantly greater in the test group (47.2%) than
in the control group (29.3%). No significant difference in PROM between groups was
observed.
Conclusions: At 6 months, no significant difference in CAL gain or PROM between
the two treatments was observed, although combination therapy yielded an en-
hanced radiographic outcome.

KEYWORDS
bone graft, deproteinized bovine bone mineral, FGF-2, patient-reported outcome, periodontal
regeneration, periodontitis

Trial registration: The University Hospital Medical Information Network-Clinical Trials Registry (UMIN-CTR) number UMIN 000025257

This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License, which permits use, distribution and reproduction
in any medium, provided the original work is properly cited and is not used for commercial purposes.
© 2019 The Authors. Journal of Clinical Periodontology Published by John Wiley & Sons Ltd

332 | wileyonlinelibrary.com/journal/jcpe J Clin Periodontol. 2019;46:332–341.

SAITO et al. | 333

1 | I NTRO D U C TI O N
Clinical Relevance
Thus far, many regenerative approaches have been introduced as Scientific rationale for the study: Currently, information is
therapy for periodontal defects. Of these, guided tissue regenera- limited regarding the clinical effect of combined use of rh-
tion (GTR) and treatment with enamel matrix derivative (EMD) are FGF-­2 and bone graft on periodontal healing.
widely performed and have shown considerable success (Heden Principal findings: Treatment of intrabony defects using
& Wennstöm, 2006; Kao, Nares, & Reynolds, 2015; Rösing, Aass, 0.3% rhFGF-­2 with DBBM or rhFGF-­2 alone demonstrated
Mavropoulos, & Gjermo, 2005; Tonetti et al., 2004). comparable values of clinical attachment level gain at 6
Guided tissue regeneration using a collagen barrier membrane months postoperatively. The combination therapy yielded
and deproteinized bovine bone mineral (DBBM) as a scaffold has a greater radiographic bone fill than rhFGF-­2 alone. No sig-
been reported to yield significantly greater regeneration of peri- nificant difference in PROM between groups was
odontal tissue, compared with that achieved by using each mate- observed.
rial alone (Camelo et al., 2001; Nevins, Camelo, Nevins, Schenk, & Practical implications: A significant improvement in perio-
Lynch, 2003). We previously reported favourable clinical outcomes dontal parameters can be expected by treatment using rh-
at up to 2.5 years after combination therapy in the treatment of in- FGF-­2, with or without DBBM. Further investigations are
trabony defects (Irokawa, Okubo, et al., 2017; Irokawa, Takeuchi, necessary to clarify the true benefit of adding bone graft to
et al., 2017). However, such procedure involving the use of a barrier rhFGF-­2 therapy.
membrane is technique-­sensitive and difficult to implement in some
clinical situations. Notably, limitations have also been reported in
the predictability of regenerative therapy, including bone grafts and
GTR (Kao et al., 2015; Lin, Rios, & Cochran, 2015). Montebugnoli, & De Sanctis, 2003). Additionally, treatment using
Attempts have been made to use bone substitutes with biolog- either EMD with DBBM or collagen membrane with DBBM showed
ical agents, without barrier membranes. Clinical and histological similar results with respect to non-­contained intrabony defects at
analyses in humans have revealed that treatment using recombinant 1 year postoperatively (Iorio-­Siciliano et al., 2014). However, no in-
human platelet-­derived growth factor (rhPDGF)-­BB with bone al- formation is available regarding the effect of combined use of rhF-
lograft yielded regeneration of periodontal tissues in intrabony de- GF-­2 with DBBM on periodontal healing.
fects and Class II furcations at 9 months (Nevins et al., 2003). In a This randomized, prospective study aimed to assess the use of
large multicenter randomized controlled trial (RCT), the use of beta-­ rhFGF-­2 in combination with DBBM, compared with rhFGF-­2 alone,
tricalcium phosphate (β-­TCP) with rhPDGF-­BB showed superior clin- in the treatment of intrabony periodontal defects after an observa-
ical outcomes, compared with those achieved using β-­TCP alone, at tion period of 6 months.
6 months (Nevins et al., 2005). Three-­year extension results from a
multicenter RCT showed that use of rhPDGF-­BB with β-­TCP yielded
2 | M ATE R I A L S A N D M E TH O DS
long-­term clinical and radiographic improvements (Nevins et al.,
2013). Furthermore, a systematic review indicated that EMD used
2.1 | Study design
with bone graft materials may yield additional clinical improvements,
as measured by gains in clinical attachment (CAL) and reductions This prospective, parallel-­arm, single-­blind RCT, which involved
in probing depth, compared with those achieved using EMD alone periodontal regenerative therapy in patients with periodontitis,
(Matarasso et al., 2015). was conducted at two centres: Tokyo Dental College Suidobashi
Fibroblast growth factor (FGF)-­2 exerts mitogenic and angio- Hospital (Tokyo, Japan) and Tokyo Dental College Chiba Hospital
genic effects on mesenchymal cells in the periodontal ligament (Chiba, Japan). This study was performed in accordance with the
and has been shown to regenerate periodontal tissue in pre-­clinical Declaration of Helsinki and was approved by the ethics committee
models (Ishii et al., 2013; Murakami et al., 2003; Oi, Ota, Yamamoto, of Tokyo Dental College (No.747). This RCT was registered (UMIN
Shibukawa, & Yamada, 2009; Takayama, Murakami, Shimabukuro, 000025257) and followed Consolidated Standards of Reporting
Kitamura, & Okada, 2001). Clinical trials have shown that trafermin, Trials (CONSORT) guidelines.
a form of recombinant human FGF-­2 (rhFGF-­2) is a safe and effective
regenerative therapy in patients with periodontitis (Kitamura et al.,
2.2 | Participants
2008, 2011, 2016). Notably, a novel regenerative therapy using 0.3%
rhFGF-­2 received a formal pharmaceutical approval for clinical use in Study participants, aged 20–79 years, were recruited consecu-
Japan in December 2016. tively between January 2017 and February 2018 from among pa-
A comparative controlled clinical trial showed that, at 1 year tients with moderate to severe chronic periodontitis (Armitage,
postoperatively, the use of combination therapy, comprising EMD 1999; Page & Eke, 2007), all of whom had completed initial peri-
and DBBM, yielded greater improvements in clinical and radiograph- odontal therapy (IP). All participants consented to enrolment in
ical outcomes than achieved using EMD alone (Zucchelli, Amore, this study.
334 | SAITO et al.

The inclusion criteria were as follows: presence of an intra-

2.4 | Sample size estimation
bony defect depth of ≥ 3 mm in inter-­proximal areas of teeth, inter-­
proximal sites with probing pocket depth (PPD) ≥ 4 mm, and a good Two treatment modalities were compared; a combined application
level of plaque control [mean Plaque Control Record (O'Leary, Drake, of rhFGF-­2 + DBBM (test) and rhFGF-­2 alone (control). A sample size
& Naylor, 1972) ≤ 20%]. A sufficient level of keratinized gingiva had of 20 defects in each group was needed to achieve 80% power for
to be present for complete tissue coverage. detection of a clinically relevant difference in CAL gain of 1.1 mm
Exclusion criteria were the presence of diabetes mellitus; respi- between treatment groups, assuming a SD of 1.2 mm (Losada,
ratory and cardiovascular diseases; immunodeficiency; oral cancers González, Garcia, Santos, & Nart, 2017) with an alpha level 0.05
and other cancers requiring treatment within the past 5 years; an (two-­t ailed). An overquotation of 10% was calculated to compensate
ongoing smoking habit; allergy to medication; and/or previous or for possible sample dropout. Thus, total sample size was 22 sites per
concurrent bisphosphonate or corticosteroid therapy. We excluded group.
patients who were pregnant/lactating, as well as those with general The unstratified design was used to calculate a two-­sided 95%
contraindications for surgical therapy. CI for the difference in CAL gain 6 months following surgery. The
non-­inferiority of test to control was accepted if the lower limit of
the 95% CI for between-­group difference was -­1.1 mm or greater.
2.3 | Clinical examination
Superiority of the test treatment was tested only when non-­
After obtaining systemic and dental histories, the following ex- inferiority was established.
aminations were performed by calibrated examiners who were
blinded to treatment assignment: PPD was measured using a
2.5 | Randomization, allocation
periodontal probe with a constant force applier (Gram Probe #2,
concealment, and blinding
YDM, Tokyo, Japan). PPD and gingival recession (GR) were meas-
ured in 0.5 mm increments. CAL was determined by adding PPD Periodontal defects were randomly assigned (1:1 ratio) to one of the
values to GR values. Bleeding on probing (BOP) was assessed di- two treatments, using a permuted block method by an individual
chotomously, and tooth mobility (TM) was also evaluated (Miller, who was not involved in other aspects of the study. Allocation to
1950). These parameters were re-­evaluated at 3 and 6 months test or control groups was concealed from periodontists until the
postoperatively. time of surgery. The assignment was performed using a sealed-­
In an investigator meeting that was held before initiation of the envelope method, and patients were blinded to their assigned treat-
study, six non-­study volunteers participated in a calibration exercise. ment. If a patient had more than one defect, all defects received the
Intra-­examiner reproducibility was assessed as the standard devia- same treatment assignment.
tion (SD) of the difference in triplicate measurements. All examiners
achieved an SD of ≤ 0.4 mm for PPD. Inter-­examiner variability was
2.6 | Surgical procedures
assessed as the difference from the gold standard examiner. The
kappa value ranged from 0.75 to 0.80 for PPD. Following local infiltration anaesthesia, defects were accessed using
the papilla preservation techniques (Cortellini, Pini Prato, & Tonetti,
1995, 1999). After debridement, defect morphology was recorded, and
2.3.1 | Radiographic examination
the depth and width of the intrabony defect were measured. Then,
Periapical radiographs were taken using customized film holders as root surfaces underwent scaling and root planing, and the sites were
described previously (Irokawa, Okubo, et al., 2017). Prior to surgery, rinsed with sterile saline. In the test group, 0.3% rhFGF-­2 [REGROTH®
the depth of intrabony defects was estimated on radiographs; this Dental Kit, 600 μg or 1200 μg in hydroxypropyl cellulose (HPC), Kaken
was confirmed during surgery. Pharmaceutical, Tokyo, Japan] with DBBM (Geistlich Bio-­Oss®, 0.25–
Assessment of radiographic changes (Schei et al., 1959) at the 1.0 mm granules, Geistlich Pharma AG, Wolhusen, Switzerland) was
surgical sites was performed using a method previously described applied to the defect. Prior to the application, the rhFGF-­2 solution
(Seshima et al., 2017). The difference in tooth axis height between was thoroughly mixed with DBBM in a sterile disposable dish. In the
the cemento-­enamel junction and the defect bottom was defined as control group, rhFGF-­2 alone was applied to the defect. Immediately
linear bone growth (LBG). The percentage of radiographic bone fill after application, the flaps were repositioned for complete closure
was determined by dividing the LBG by the baseline defect depth. and sutured with modified vertical mattress or interrupted sutures.
Representative treatment cases are shown in Figure 1.

2.3.2 | Patient-­reported outcome measure

2.7 | Postsurgical care
An oral health-­related quality of life (QoL) instrument, the OHRQL
(Japanese version) (Saito et al., 2010, 2011), was used. Scoring was Patients received antimicrobials (amoxicillin 750 mg/day or cefdinir
performed as previously described (Saito et al., 2010, 2011). 300 mg/day) for 4 days. Standard pain medications were prescribed
SAITO et al. | 335

(a) (b) (c) (d) (e)

(f) (g) (h) (i) (j)

(k) (l) (m) (n)

F I G U R E 1 Surgical procedure and outcomes. (a–g) 60-­year-­old female patient who received recombinant human fibroblast growth
factor-­2 (rhFGF-­2) with deproteinized bovine bone mineral (DBBM; test group). (a) Preoperative radiograph. (b) Baseline clinical view
(palatal). Probing pocket depth (PPD) at the mesial aspect of tooth #24 was 7 mm. (c) Intra-­operative view. Defect depth 3 mm, width 5 mm.
(d) After debridement and rinsing, the defect of #24 was filled with rhFGF-­2 formulation as well as DBBM that had been pre-­saturated with
rhFGF-­2. (e) Suturing. (f) Six-­month follow-­up view; PPD = 2 mm. (g) Six-­month follow-­up radiograph. (h-­n) 53-­year-­old female patient who
received rhFGF-­2 alone (control group); (h) Preoperative radiograph. (i) Baseline clinical view. PPD at the distal aspect of tooth #33 was
7 mm. (j) Intra-­operative view. Defect depth 5 mm, width 3 mm. (k) After debridement and rinsing, the defect of #33 was filled with rhFGF-­2.
(l) Suturing. (m) Six-­month follow-­up view. PPD = 3 mm. (n) Six-­month follow-­up radiograph

as needed. Patients used a mouthwash twice per day. They gently was used. A p value of 0.05 was considered to indicate statistical
cleaned the treated area with an ultrasoft toothbrush, beginning significance.
1 day postoperatively and continued for 4 weeks.
Sutures were removed after 10–14 days. The patients were then
3 | R E S U LT S
placed in supportive care programs.

3.1 | Participant demographics and clinical

2.8 | Statistical analysis parameters
Fisher's exact test was used to assess categorical variables. The A total of 34 patients were assessed for eligibility at two study cen-
Mann–Whitney U test was used to compare other non-­categorical tres; 44 sites in 32 patients were randomized between the two treat-
demographic and baseline parameters between the two groups. ment groups (Figure S1 flowchart).
The primary outcome was the change in CAL at 6 months post- Table 1 shows the participant demographics and baseline clini-
operatively. Sites were compared with baseline, as well as between cal parameters. Five patients contributed more than one defect site.
treatments. The data from the two centres were pooled for the pres- There were no significant differences in participant demographics
ent analysis. The Friedman test with post hoc analysis was used to and baseline parameters between groups.
compare intra-­group data over time. For intra-­group comparisons of
OHRQL scores over time, repeated measures analysis of variance
3.2 | Clinical outcome
(ANOVA) with Tukey–Kramer post hoc test was used to compare
intra-­group data over time, after testing on normal distribution of In each group, 22 sites of 16 patients completed the study. Healing was
data. uneventful for all participants. There were no notable adverse events.
Correlations between postoperative CAL gains and baseline Characteristics of intrabony defects in the participants are sum-
variables were determined by Spearman correlation coefficient. marized in Table 2. Overall, there were no significant differences in
Statistical software (InStat 3.10, GraphPad, La Jolla, CA, USA) maxillary versus mandibular sites, tooth type, defect morphology,
336 | SAITO et al.

TA B L E 1 Participant demographics
rhFGF-­2 rhFGF-­2 + DBBM Difference
and baseline parameters
Age (years; mean ± SD) 50.0 ± 10.9 52.3 ± 10.1 N.S.
(range, 28–69) (range, 37–76)
Gender
Men 6 7 N.S. a
Women 10 9
No. of teeth (mean ± SD) 25.7 ± 3.9 26 ± 2.2 N.S.
Clinical attachment level (CAL) (mm; mean ± SD)
Full-­mouth 3.30 ± 0.59 3.30 ± 0.63 N.S.
b
Reference site 7.07 ± 1.56 7.57 ± 1.64 N.S.
Probing pocket depth (PPD) (mm; mean ± SD)
Full-­mouth 2.86 ± 0.50 3.00 ± 0.53 N.S.
Reference siteb 6.02 ± 1.33 6.32 ± 1.25 N.S.
Gingival recession (GR) (mm; mean ± SD)
Reference siteb 1.23 ± 1.51 1.25 ± 1.38 N.S.
Bleeding on probing (BOP) positive (%)
Reference siteb 72.7 77.3 N.S. a
Tooth mobility (TM) (mean ± SD)
Reference toothb 0.18 ± 0.50 0.22 ± 0.43 N.S.

Note. Differences were assessed by the Mann–Whitney U test ( Fisher's exact test). b n = 22 per
a

group.

TA B L E 2 Distribution and configuration of intrabony defects Non-­inferiority of the test treatment to the control can be claimed,
because the lower limit of the 95% CI for the difference in CAL gain
Intrabony defect rhFGF-­2 rhFGF-­2 + DBBM
was 0.39 ± 1.73 [95% CI (−0.38 to 1.15) was greater than −1.1 mm. No
Position [n (%)]
statistically significant inter-­group difference was observed. In the test
Maxilla 9 (41) 10 (45.5) group, 22.7% of sites (n = 5) showed CAL gains of >4 mm; only 4.5% of
Mandible 13 (59) 12 (54.5) sites (n = 1) in the control group showed this level of CAL gain (Table S1).
Anterior teeth 7 (31.8) 2 (9.1) Reductions in PPD were 3.55 ± 1.35 mm in the test group and
Premolars 6 (27.3) 6 (27.3) 3.30 ± 1.20 mm in the control group; these did not significantly dif-
Molars 9 (40.9) 14 (63.6) fer between groups.
Morphology [n (%)]
1-­wall 3 (13.6) 2 (9.1) 3.3 | Correlation between CAL gain and
2-­wall 5 (22.7) 5 (22.7) baseline parameters
3-­wall 8 (36.4) 7 (31.8)
In a secondary analysis, relationships were assessed between post-
Combination 6 (27.2) 8 (36.4)
operative CAL gains at 6 months postoperatively and baseline (post-
Depth (mm; 4.66 ± 1.76 4.70 ± 1.08
­IP) variables. CAL gain at 6 months and baseline CAL or PPD values
mean ± SD) (range, 3.0–11.0) (range, 3.0–6.5)
showed significantly positive correlations in both groups (Table S2).
Width (mm; 2.80 ± 0.75 3.89 ± 1.81***
mean ± SD) (range, 2.0–5.0) (range, 2.0–10.0) In the control group, a significant positive correlation was noted be-
tween CAL gain and the number of teeth at baseline. There was no
Note. ***p = 0.007, Mann–Whitney U test.
significant correlation with other baseline variables in either group.
or defect depth between groups. However, there was a statistically
significant difference in the defect width (p = 0.007). The test group
3.4 | Radiographic outcome
had significantly wider defects.
At 3 and 6 months postoperatively, marked improvements in In both groups, a significant increase was observed in radiographic
CAL and PPD from baseline (post-­IP) were noted in both groups bone fill from 3 months to 6 months postoperatively (Table 3). At
(p < 0.001) (Table 3). 6 months, the mean value for the radiographic bone fill was signifi-
At 6 months postoperatively, CAL gains were 3.16 ± 1.45 mm in cantly greater in the test group (47.2%) than in the control group
the test group and 2.77 ± 1.15 mm in the control group (Figure 2a). (29.3%) (p = 0.013) (Figure 2b).
SAITO et al. | 337

TA B L E 3 Clinical and radiographic outcomes of treated sites (Total n = 44; n = 22 per group)

Change from baseline Change from baseline Change from 3

Variable/Group Baseline 3 months to 3 months 6 months to 6 months to 6 months

CAL (mm)
rhFGF-­2 7.07 ± 1.56 4.75 ± 1.24 p < 0.001 4.29 ± 1.33 p < 0.001 N.S.
(6.38–7.76) (4.20–5.30) (3.70–4.89)
rhFGF-­2 + DBBM 7.57 ± 1.64 4.50 ± 1.59 p < 0.001 4.41 ± 1.40 p < 0.001 N.S.
(6.80–8.39) (3.80–5.20) (3.80–5.03)
Difference N.S. N.S. N.S.
PPD (mm)
rhFGF-­2 6.02 ± 1.33 3.05 ± 0.80 p < 0.001 2.73 ± 0.84 p < 0.001 N.S.
(5.43–6.61) (2.69–3.40) (2.35–3.10)
rhFGF-­2 + DBBM 6.32 ± 1.25 2.84 ± 0.82 p < 0.001 2.77 ± 0.72 p < 0.001 N.S.
(5.76–6.87) (2.48–3.21) (2.45–3.09)
Difference N.S. N.S. N.S.
GR (mm)
rhFGF-­2 1.23 ± 1.51 1.57 ± 1.38 N.S. 1.39 ± 1.34 N.S. N.S.
(0.56–1.90) (0.96–2.18) (0.80–1.98)
rhFGF-­2 + DBBM 1.25 ± 1.38 1.68 ± 1.41 N.S. 1.64 ± 1.33 N.S. N.S.
(0.64–1.86) (1.06–2.31) (1.05–2.23)
Difference N.S. N.S. N.S.
BOP positive (%)
rhFGF-­2 72.7 22.7 p = 0.002 9.1 p < 0.001 N.S.
rhFGF-­2 + DBBM 77.3 4.5 p = 0.001 4.5 p < 0.001 N.S.
Differencea N.S. N.S. N.S.
TM
rhFGF-­2 0.18 ± 0.50 0.09 ± 0.29 N.S. 0.09 ± 0.29 N.S. N.S.
(0–0.40) (0–0.22) (0–0.22)
rhFGF-­2 + DBBM 0.22 ± 0.43 0.05 ± 0.21 N.S. 0.05 ± 0.21 N.S. N.S.
(0.09–0.04) (0–0.14) (0–0.14)
Difference N.S. N.S. N.S.
RBF (%)
rhFGF-­2 – 21.8 ± 11.9 – 29.3 ± 13.3 – p = 0.001
rhFGF-2 + DBBM – 39.3 ± 17.6 – 47.2 ± 16.0 – p < 0.001
Difference p = 0.001 p = 0.001

Notes. Data shown as mean ± SD (95% Confidence Interval) (except for BOP and RBF). Inter-­group difference at each time point was assessed by the
Mann–Whitney U test. Intra-­group difference over time was assessed by the Friedman test with Dunn's post-­test (aCategorical data were assessed by
Fisher's exact test).
CAL: clinical attachment level; PPD: probing pocket depth; GR: gingival recession; BOP: bleeding on probing; TM: tooth mobility; RBF: radiographic
bone fill.

score, no significant changes were observed at 3 or 6 months post-

3.5 | Effect of defect configurations
operatively. At each timepoint, no significant difference in total
When 6-­month postoperative CAL gains were compared between OHRQL score was observed between groups.
1–2-­wall and 3-­wall defects, no significant differences were found in
either treatment group (Table 4). Similarly, no significant differences
in radiographic bone fill were found between 1–2-­wall defects and 4 | D I S CU S S I O N
3-­wall defects in either group.
To the best of our knowledge, this is the first RCT to evaluate the use
of rhFGF-­2 with DBBM, compared with rhFGF-­2 alone in the treat-
3.6 | Patient-­reported outcome measure (PROM)
ment of intrabony defects, using both clinical and patient-­centred
In both groups, IP yielded a significant improvement in total OHRQL outcomes. We have shown no significant difference in CAL gain be-
score (Figure 3). Compared with baseline (post-­IP) total OHRQL tween the two groups at 6 months postoperatively. The combination
338 | SAITO et al.

F I G U R E 3 Change in total OHRQL scores over time. **p < 0.01,

***p < 0.001, significantly different from those at initial visit, by the
repeated measures analysis of variance with Tukey–Kramer post
hoc test. No significant difference was noted between the two
different treatment groups at any time point. IP: initial periodontal
therapy
F I G U R E 2 Clinical attachment level (CAL) gain (a) and
radiographic bone fill (RBF) (b) at 6 months. Scatter plots showing
individual data with mean (middle line) and standard deviation defects in a dog model, compared with the use of β-­TCP alone. In a
(error bars).***p = 0.001 by Mann–Whitney U test multicenter RCT evaluating the treatment of intrabony periodon-
tal defects, the use of 0.3% and 0.4% rhFGF-­2 with β-­TCP showed
therapy yielded a greater radiographic bone fill than rhFGF-­2 alone. an improved success rate (based on changes in CAL and LBG) at
Furthermore, no significant difference in PROM between groups 6 months postoperative, compared with β-­TCP alone (Cochran
was noted, as assessed by total OHRQL scores. et al., 2016). The reported CAL gain achieved using 0.3% rhFGF-­2
In clinical trials regarding treatment of periodontal defects, the with β-­TCP was 3.0 ± 1.4 mm at 6 months; this was comparable
use of rhFGF-­2 alone has yielded clinically favourable outcomes to our present findings using rhFGF-­2 with DBBM (3.2 ± 1.5 mm).
(Kitamura et al., 2008, 2011, 2016). This is remarkable considering However, the participants in the prior study exhibited consider-
that, as in the present study, prior investigators used 0.3% rhF- ably greater baseline mean CAL (8.3 mm) and PPD (7.9 mm) values,
GF-­2 with HPC, a gel-­like base material, without the support of compared with those (CAL 7.6 mm, PPD 6.3 mm) in the present
other scaffolds such as bone graft materials (Li et al., 2017). To study. In this study, the CAL gain was significantly positively
further explore the regenerative potential of rhFGF-­2 , the ef- correlated with baseline CAL or PPD values, which is consistent
fects of its combined use with β-­TCP have been studied. Oi et al. with the previous reports (Cortellini, Pini Prato, & Tonetti, 1993;
(2009) reported that, in dogs, rhFGF-­2 + β-­TCP treatment yielded Irokawa, Okubo, et al., 2017; Saito, Nanbu, Nagahata, & Yamada,
greater levels of new bone and cementum formation compared 2008). This indicates that CAL gain after regenerative therapy de-
with treatment with rhFGF-­2-­alone. In a previous study, we used pends on the initial level of destruction, which has to be taken into
rhFGF-­2 + β-­TCP for root coverage in a dog model (Ishii et al., account when comparing data from different studies.
2013); we found that the combination therapy enhanced forma- Regarding the effect of the combined use of rhFGF-­2 with de-
tion of new bone and cementum. Thus, we speculated that β-­TCP mineralized bone matrix (DBM), Hagino, Hamada, and Amemiya
provided a local environment that was suitable for periodontal re- (2001) showed that the use of 200 μg of rhFGF-­2 with DBM yielded
generation. Moreover, Anzai et al. (2010) reported that the use of early new bone formation in segmental bone defects in a rabbit
FGF-­2 with β-­TCP increased the bone mineral contents of 1-­wall model. Notably, these authors suggested that when DBM was used

TA B L E 4 Clinical attachment gain and

Defect rhFGF-­2 Difference rhFGF-­2 + DBBM Difference
radiographic bone fill at 6 months
CAL gain (mm) 3-­wall 2.78 ± 0.79 N.S. 3.43 ± 2.09 N.S. postoperatively, based on defect
1-­2-­wall 2.77 ± 0.38 3.11 ± 1.11 configuration

RBF (%) 3-­wall 29.4 ± 11.2 N.S. 41.5 ± 13.0 N.S.

1-­2-­wall 29.2 ± 15.1 49.9 ± 17.0

Notes. Data are shown as mean ± SD.

CAL: clinical attachment level; RBF: radiographic bone fill.
SAITO et al. | 339

as a carrier, a single local administration of rhFGF-­2 at an appropri- we compared the results in 3-­wall defects and 1–2-­wall defects,
ate concentration may be effective for bone formation; moreover, no significant difference was noted in CAL gain or radiographic
the release of FGF-­2 over an extended period may not be neces- bone fill. There are two possible reasons for this phenomenon: (1)
sary. When DBBM was used with EMD in vitro, the investigators rhFGF-­2 alone is clinically effective for non-­contained defects; (2)
observed enhanced attachment, proliferation and differentiation the present study was underpowered to detect this difference.
of osteoblasts and periodontal ligament cells on DBBM granules Considering that 1-­wall defects comprised only 14% and 9% of
(Miron et al., 2012). In a study using concentrated growth factors sites in the test and control groups, respectively, both possibilities
with DBBM, persistent releases of cytokines, including FGF-­2, was should be examined.
observed over a period of 28 days (Yu, Wang, Liu, & Qiao, 2018). Subjective oral health-­related QoL assessments are considered
These data indicate the potential for use of DBBM as a scaffold or true endpoints when evaluating the effect of periodontal treatment
carrier for growth factors. The adsorption of rhFGF-­2 to DBBM, as (Hujoel, 2004). In the present study, oral health-­related QoL was
well as specific cell behaviours on DBBM with rhFGF-­2 should be assessed as a PROM. Consistent with the findings of our previous
further explored. study (Makino-Oi et al., 2016), patients’ oral health-­related QoL
There are multiple opinions within the literature regarding the scores were significantly improved after IP. Regenerative therapy
effect of bone substitutes in combination regenerative therapy. In using rhFGF-­2 alone or rhFGF-­2 combined with DBBM yielded no
a multicenter RCT, Jepsen et al. (2008) reported the treatment of significant changes in oral health-­related QoL, indicating that neither
intrabony defects using EMD with alloplast or EMD alone; both surgical intervention positively or negatively affected patients’ oral
approaches showed similar results after 6 months. The similar re- health-­related QoL. Furthermore, patients’ perceptions of the rhF-
sults were confirmed at 36 months after treatment (Hoffmann, GF-­2 treatment and healing process may not have been impaired by
Al-­Machot, Meyle, Jervøe-­Storm, & Jepsen, 2016). Pietruska et al. the addition of DBBM.
(2012) also evaluated clinical outcomes following treatment of in- There were limitations in this study. For example, the test sites
trabony defects using EMD with synthetic bone graft or EMD alone; had wider defects at baseline, but our statistical analysis did not in-
they concluded that, in 2-­ and 3-­wall defects, combination therapy clude an adjustment for this. Because the number of non-­contained
did not show any advantage after 4 years. In contrast, a systematic defects was limited, the impact of the number of residual walls on
review revealed that the use of EMD with bone grafts may enhance treatment outcomes may not have been appropriately evaluated.
clinical outcomes (Matarasso et al., 2015). According to a clinical Moreover, because the observation period of 6 months is relatively
practice guideline of the Japanese Society of Periodontology (JSP, short, a longer follow-­up is needed.
2016), there is no clear evidence of an additional effect when bone In conclusion, treatment of intrabony defects with rhFGF-­2, with
graft is applied in combination with GTR or EMD. The guideline or without DBBM, yielded significant improvements in periodon-
states that careful consideration should be given to this type of ap- tal parameters at 6 months, relative to baseline measurements. No
plication. In the present study, the addition of DBBM to rhFGF-­2 significant difference in CAL gain was observed between groups,
therapy did not yield a significantly greater CAL gain at 6 months although combination therapy yielded an enhanced radiographic
(the primary endpoint). Of concern was the statistically significant outcome. There was no significant difference in PROM between
difference (p = 0.007) in baseline defect width for the test (3.89 mm) groups. Further longitudinal investigation with a larger number of
and the control (2.80 mm). This may be one reason for no signifi- participants should identify the characteristics of cases that would
cant difference in CAL gains found between groups. However, the benefit most from combined treatment using rhFGF-­2 and DBBM.
radiographic bone fill (a secondary endpoint) in the test group was
significantly greater than in the control group. The influence of dif-
C O N FL I C T O F I N T E R E S T
ference in the initial defect width on the clinical outcome remains to
be clarified. The authors declare that there are no conflicts of interest in connec-
Tonetti et al. (2002) reported that defect morphology (i.e. tion with this article.
number of bone walls) influenced the clinical results of regenera-
tive therapy with EMD. Trombelli and Farina (2008) reported that
ORCID
EMD does not maintain a space itself when used in non-­contained
defects. According to an evidence-­b ased decision tree by Atsushi Saito https://orcid.org/0000-0002-0065-2207
Cortellini and Tonetti (2000), the combined use of supportive or
filling materials is recommended to treat wide defects and/or non-­
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