Mahidol Study 1: Comparison of Radiographic and

Survival Outcomes of Immature Teeth Treated with Either

Regenerative Endodontic or Apexication Methods:
A Retrospective Study
Thanawan Jeeruphan, DDS, Grad Dip in Clin Sc (Endodontics), High Grad Dip in Dentistry
(Operative),* Jeeraphat Jantarat, DDS, MDSc, PhD,* Kallaya Yanpiset, DDS, MDSc,*
Lalida Suwannapan, DDS, Grad Dip in Clin Sc (Endodontics),*
Phannarai Khewsawai, DDS, High Grad Dip in Dentistry (Endodontics),

and Kenneth M. Hargreaves, DDS, PhD

Abstract
Introduction: There are numerous challenges in treating
immature permanent teeth with a diagnosis of pulp
necrosis. Three general treatment options are calcium
hydroxide apexication, mineral trioxide aggregate
(MTA) apexication, and revascularization. The objective
of this retrospective study was to evaluate radiographic
and clinical outcomes of immature teeth treated with 1
of these 3 methods. Methods: Clinical outcome data
and radiographs were collected from 61 cases (ie, 22
calcium hydroxide apexication cases, 19 MTA apexica-
tion cases, and 20 revascularization cases). Both tooth
survival and clinical success rates were analyzed. In addi-
tion, the preoperative and recall radiographs were
analyzed to calculate the percentage increase in root
width and length. Results: The percentage change of
root width was signicantly greater in the revasculariza-
tion group (28.2%) compared with the MTA apexication
(0.0%) and calcium hydroxide apexication groups
(1.5%). In addition, the percentage increase of root
length was signicantly greater in the revascularization
group (14.9%) compared with the MTA (6.1%) and
calcium hydroxide apexication groups (0.4%). More-
over, the survival rate of the revascularization-treated
teeth (100%) and MTA apexicationtreated teeth
(95%) were greater than the survival rates observed in
teeth treated with calcium hydroxide (77.2%). Conclu-
sions: In this study, revascularization was associated
with signicantly greater increases in root length and
thickness in comparison with calcium hydroxide apexi-
cation and MTA apexication as well as excellent overall
survival rates. (J Endod 2012;38:13301336)
Key Words
Apexication, calcium hydroxide, immature teeth, mineral trioxide aggregate, regener-
ative endodontics, retrospective studies, revascularization
T
here are numerous challenges that the clinician faces when treating infected
pulp in immature permanent teeth. The cleaning and shaping of the root canal
system is challenging because of the thin dentinal walls. Obturation is also compli-
cated because the apex is not fully developed and has a blunderbuss shape. More-
over, these teeth may be susceptible to fracture during or after treatment (1).
Traditionally, a calcium hydroxidebased apexication procedure has been advo-
cated for treating an immature permanent tooth with an open apex (2). Teeth
treated with this apexication procedure require a long-term application of calcium
hydroxide in order to create an apical barrier to prevent the extrusion of obtura-
tion materials. However, there are several drawbacks to this traditional apexication
procedure, including a potential calcium hydroxidemediated reduction in root
strength and the requirement for excellent patient compliance because of the
need for multiple visits scheduled over many months (3, 4).Thus, the traditional
calcium hydroxide treatment approach for these cases may be less than ideal
for many patients.
Alternative apexication methods have recently been proposed. Mineral trioxide
aggregate (MTA), used in a 1- or 2-step apexication procedure, has been shown to
create an articial apical barrier that permits the compaction of obturating material
and the placement of coronal restoration (5). Several case series have indicated that
MTA apexication treatment has a high success rate with a greatly decreased number
of appointments and time to completion (610). Despite these advantages, these
treatments neither strengthen the root nor foster further root development. Thus,
the roots remain thin and fragile, suggesting the need for another treatment
approach.
A recently proposed alternative treatment is to promote continued hard-tissue
formation and root growth. Revascularization is a regenerative treatment that is
biologically based and designed to allow the continuation of root development
*Department of Operative Dentistry and Endodontics, Mahidol University, Bangkok, Thailand;

Department of Dentistry, Pasang Hospital, Lamphun, Thailand; and

Department of Endodontics, University of Texas Health Science Center at San Antonio, San Antonio, Texas.
Address requests for reprints to Dr Thanawan Jeeruphan, Department of Operative Dentistry and Endodontics, Mahidol University, 6 Yothi Street, Rajthevi, Payathai,
Bangkok 10400, Thailand. E-mail address: thandent@gmail.com
0099-2399/$ - see front matter
Copyright 2012 American Association of Endodontists.
http://dx.doi.org/10.1016/j.joen.2012.06.028
Clinical Research
1330 Jeeruphan et al. JOE Volume 38, Number 10, October 2012
and strengthening of the root structure (11, 12). Many recently
published case reports (1315) indicate that this treatment has
the potential to promote the continued development of both the
root width and the overall root length in these immature cases.
However, there are no clinical data comparing the outcomes of
these 3 treatments for the immature tooth with a necrotic pulp. The
objective of this retrospective study was to compare the outcomes of
treatment in immature teeth with nonvital pulp among calcium
hydroxide apexication, MTA apexication, and revascularization and
to establish a standard protocol permitting future comparative studies.
Materials and Methods
The study protocol was approved by the Ethics Committee
of Mahidol University (ref. 2012/009.2501). The dental charts of all
patients who had treatment for an immature permanent tooth in the
Department of Operative Dentistry and Endodontics and the Department
of Pediatric Dentistry, Mahidol University, Bangkok, Thailand, between
1997 and 2009 that fullled the following criteria were collected:
1. The immature permanent tooth was treated by either calcium
hydroxide apexication, MTA apexication, or revascularization.
2. There was a preoperative, postoperative, and follow-up radiograph
of at least 6 months after the completion of treatment.
3. Clinical data of signs and symptoms of preoperative and follow-up
visits were recorded.
The data collected included patient age, sex, tooth number, clinical
symptoms and signs, pulpal and periapical diagnoses, clinical proce-
dures, size of apical radiolucency, root length, dentin thickness, the
Figure 1. A clinical checklist used for all patients at each recall visit. Clinicians were instructed to check for pain, percussion sensitivity, probing depths, swelling,
and fractures, and this information was placed under Remarks.
Figure 2. (A) The root length is measured from the CEJ to the radiographic apex. (B) The root canal width was measured at the level of two thirds of the preop-
erative root canal length. (C) The pulp space was measured at the same level, and the remaining dentin thickness was calculated by subtracting the pulp space from
the root canal width.
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JOE Volume 38, Number 10, October 2012 Radiographic and Survival Outcomes of Immature Teeth 1331
follow-up period, and, if required, the reason for extraction. The collec-
tion of these data was standardized by the use of a clinical checklist
(Fig. 1) that is completed for all patients at all postoperative visits. This
standardized method of assessment has been used at Mahidol University
since 1995 and permits the analysis of radiographic, subjective (ie, pain
and percussion sensitivity), and objective (ie, sinus tract, swelling, and
crown fracture) outcomes after the completion of treatment.
Revascularization was accomplished in a 3-appointment process
in which the rst appointment consisted of rubber dam isolation, irri-
gation with 2.5% NaOCl, and no instrumentation. The triple antibiotic
mix as described by Hoshino et al (16) was placed for a mean of
28.85 13.08 days between appointments; teeth were closed between
appointments with a temporary restoration. The second appointment
consisted of rubber dam isolation, irrigation with 2.5% NaOCl, and
laceration of the apical tissue using either a le or an endo spreader.
A blood clot was formed in the canal and a matrix (Collaplug; Zimmer
Dental, Carlsbad, CA) was placed over the blood clot if the blood clot
was formed below the cementoenamel junction (CEJ). MTA apexica-
tion was accomplished in a 1-appointment procedure and consisted of
the delivery of an apical plug of at least 3 mm MTA followed by obtura-
tion with glass ionomer cement and resin composite. Calcium
hydroxide apexication consisted of a rubber dam isolation and intra-
canal delivery of Ca(OH)
2
spread over a mean of 17 12.6 months
followed by obturation with gutta-percha.
The preoperative and postoperative images were scanned with
a Microtek scanner (Scanmaker 9800XL; Microtek International Inc,
Hsinchu, Taiwan); saved in a JPEG format; and transferred to the
imaging program software (Sopro; Acteon, La Cirotat Cedex, France)
for measurement and recording the root length, dentin thickness,
and the size of the periapical lesions. The size #2 of an intraoral radio-
graphic lm image in the Sopro software was calibrated by setting the
horizontal dimension to 31 mm and the vertical dimension to 41
mm. The calibration process permitted the measurement of changes
in root size on a millimeter scale. The root length from both the preop-
erative and recall images were measured, which were modied fromthe
procedure described by Bose et al (17). A straight line from the CEJ to
the radiographic apex of the tooth and the increasing root length was
calculated. The dentinal wall thickness for both the preoperative and
recall images were measured at the level of the apical one third of
the preoperative root canal length measured from the CEJ (Fig. 2).
The root canal width and the pulp space were measured at this level,
and the remaining dentin thickness was calculated by subtracting the
pulp space from the root canal width. The root length and the dentin
wall thickness were reported in terms of percentage change in root
length and percentage change in dentin thickness, respectively. All
radiographic measures were collected by the same investigator, and
images subjected to repeated analysis showed a kappa reliability. A
repeated measure of the same set of radiographs 1 week apart (n =
15 cases) produced an intraexaminer agreement of k = 0.9845, indi-
cating the reliability of these measures.
Treatment outcomes were assessed using both survival and
success as separate analyses. Survival was dened as retention of the
tooth in the arch at the time of the postoperative recall. Although
many factors can contribute to a lack of survival, in this case series it
was exclusively because of a catastrophic fracture that was deemed non-
restorable by the attending clinician. The second outcome analysis was
success. This was dened using the same criteria described by Fried-
man and Mor (18), which consisted of the following categories:
1. Healed: Both the clinical (subjective and objective) and radio-
graphic presentations were normal.
2. Healing: The periapical radiolucency was reduced with a normal
clinical presentation.
3. Disease: The radiolucency was either increased or persisted without
change even when the clinical presentation was normal or clinical
signs or symptoms were present regardless of the radiographic
presentation.
Statistical Analysis
All radiographic measurements were repeated after 1 week, and
the mean of the 2 sets was considered as the nal value. The data of
TABLE 1. A Summary of Patient Demographics and the Clinical Characteristics of the Study Population
Variable
Revascularization
(n = 20) MTA (n = 19)
Calcium hydroxide
(n = 22)
Teeth % Teeth % Teeth %
Sex
Female 10 50.00 6 31.58 9 40.91
Male 10 50.00 13 68.42 13 59.09
Tooth type
Incisor 7 35.00 11 57.89 18 81.82
Premolar 13 65.00 8 42.11 4 18.18
Tooth location
Maxilla 7 35.00 12 63.16 18 81.82
Mandible 13 65.00 7 36.84 4 18.18
Cause
Caries 1 5.00 4 21.05 0 0.00
Dens evaginatus 12 60.00 4 21.05 4 18.18
Trauma 7 35.00 11 57.89 18 81.82
Signs and symptoms*
Absent 3 15.00 5 26.32 1 4.55
Present 17 85.00 14 73.68 21 95.45
Apical periodontitis
Absent 1 5.00 6 31.58 8 36.36
Present 19 95.00 13 68.42 14 63.64
Age (y) (mean SD) 12.9 5.07 14.6 6.17 10.5 3.85
Follow-up time (mo) (mean SD) 21.15 11.70 14.21 7.84 27.32 30.47
SD, standard deviation.
*Signs and symptoms include pain, swelling, sinus opening, percussion, palpation, mobility, or probing depths >3 mm.
Clinical Research
1332 Jeeruphan et al. JOE Volume 38, Number 10, October 2012
dentin thickness and root length were calculated as the percentage
change from the preoperative values. The radiographic data were
analyzed initially for normality and were found to be nonnormally
distributed. The percentage change in root length and the percentage
change in the dentin of thickness were analyzed by the nonparametric
Kruskal-Wallis analysis of variance test, and the Mann-Whitney U test
was used to identify the difference among groups. A P value <.05 was
considered to be signicant, and the mean values were reported.
Results
The retrospective chart review found 79 cases meeting the
eligibility criteria, of which 61 had documented recall
Figure 3. (A) A preoperative radiograph of calcium hydroxide apexication treatment of tooth #11 and (B) a recall radiograph at 12 months. (C) A preoperative
radiograph of MTA apexication treatment of tooth #34 and (D) a recall radiograph at 24 months. (E) A preoperative radiograph of revascularization treatment of
tooth #45 and (F) a recall radiograph at 29 months.
Figure 4. (A) The percentage increase in root width after treatment with revascularization (Revasc) or with apexication by either MTA or calcium hydroxide
(Ca(OH)
2
). (B) The percentage increase in root length after treatment. ***P < .001 vs all other groups. **P < .01 vs all other groups.
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JOE Volume 38, Number 10, October 2012 Radiographic and Survival Outcomes of Immature Teeth 1333
information for a recall rate of 77%. These 61 cases included
20 revascularization cases, 19 MTA apexication cases, and 22
calcium hydroxide apexication cases. Table 1 summarizes the
patient demographics and the clinical characteristics of this
study population.
An analysis of the radiographic outcomes indicated a signicant
effect (P < .0001) for revascularization treatment on the overall
root width at the apical third. Representative cases are presented
in Figure 3, and group analyses are presented in Figure 4. As de-
picted in Figure 4A, the treatment of immature teeth with the revas-
cularization protocol produced signicantly greater percentage
increases in root width (28.2%) compared with teeth treated by
either MTA apexication (0.00%) or calcium hydroxide apexica-
tion (1.52%, P < .0001 for both comparisons). There were no
statistically signicant differences between the calcium hydroxide
apexication and the MTA apexication groups.
The treatment groups also differed signicantly (P < .001) in
the development of the root length (Fig. 4B). Teeth treated with
revascularization showed a signicantly greater percentage increase
in root length (14.9%) compared with teeth treated by either MTA
apexication (6.1%) or calcium hydroxide apexication (0.4%, P
< .01 for both comparisons).
We next conducted a subset analysis to determine whether there
were any effects of patient age or sex on radiographic changes in
root development after the revascularization procedure (Fig. 5).
Although the sample size was small, there was no trend for differences
in outcome for either increased root width (Fig. 5A) or increased root
length (Fig. 5B) in this study population.
Figure 5. An evaluation of the contribution of patient age and sex to radiographic changes in (A) root width and (B) root length after revascularization treatment.
Numbers at each bar indicate the number of patients per age group.
Clinical Research
1334 Jeeruphan et al. JOE Volume 38, Number 10, October 2012
We next evaluated clinical outcomes by both tooth survival
analysis and a clinical success analysis. In terms of tooth survival
(Table 2), revascularization had similar survival rates (20/20 teeth
[100%]) compared with teeth treated with MTA apexication
(18/19 teeth [95%]) although the follow-up times were nearly
50% longer in the revascularization cases. Both of these treatments
produced greater survival rates (P < .05) compared with teeth
treated with calcium hydroxide apexication (17/22 teeth [77%]).
The second clinical outcome analysis was based on success
dened by radiographic, objective, and subjective ndings (Table 3).
Teeth treated with the revascularization protocol had rates of complete
healing (16/20 cases [80%]) similar to teeth treated with either the
MTA (13/19 [68%]) or calcium hydroxide (17/22 [77%]) apexica-
tion methods. Alternatively, when using a looser outcome of healed +
healing, teeth treated with the revascularization protocol had similar
overall rates (20/20 teeth [100%]) compared with teeth treated with
MTA apexication (18/19 teeth [95%]), and both of these treatments
produced greater rates (P < .05) compared with teeth treated with
calcium hydroxide apexication (17/22 teeth [77%]).
Discussion
The recent demonstration of good radiographic outcomes after
revascularization treatments applied to immature permanent teeth
has greatly stimulated interest in this method. Although most of the
data are derived from case reports or relatively small case series, 1
retrospective study has shown a signicantly greater increase in
root width and length after revascularization compared with MTA
apexication or traditional nonsurgical root canal treatment (17).
The present study provides an independent replication of this prior
retrospective study and extends this nding by analyzing calcium
hydroxide apexication and performing an outcome analysis of
both tooth survival and clinical success.
There are several pertinent issues related to the present study.
First, the routine use of a standardized clinical checklist for all recall
patients (Fig. 1) permits the evaluation of subjective and objective
clinical outcomes that greatly expands an analysis of only radio-
graphic outcomes. Second, the overall recall rate (77%) is higher
than that reported in many clinical studies, suggesting that the
analyzed data may reect the actual outcomes observed in this patient
population. Third, the radiographic outcomes of the percentage
increase in root width and length for revascularization (28.2% and
14.9%, respectively) and MTA apexication (0% and 6.0%, respec-
tively) show the dramatic effect of this new treatment approach.
Moreover, these data are similar to the results of the Bose et al study
(17) in terms of relative effects (increases in radiographic root width
>increases in length) and the magnitude of the effects. Although the
Bose et al data reported median effects because of the heterogeneity
of variance in their clinical material, a reanalysis of their raw data for
mean effects reveals similar relative percentage increases in root
width and length for both revascularization (48% and 11%, respec-
tively) and MTA apexication (0.2% and 0.2%, respectively) as
seen in the present study. Collectively, these studies provide indepen-
dent assessments of the efcacy of revascularization methods. Fourth,
to our knowledge, this is the rst study to quantitatively analyze both
tooth survival and clinical success after revascularization treatment.
Taken together, these factors show the clinical usefulness of revascu-
larization protocols and establish a framework for future studies in
this eld.
This study does have issues that limit the interpretation of the
data. First, it is a retrospective study, and patients were not randomly
allocated to treatment. Second, the radiographs were not collected
using standardized methods. However, we attempted to minimize
this particular issue by controlling central beam angulation by using
parallel technique lms and a computer program to calibrate the
images.
Another clinically signicant nding in this study was the clinical
outcomes observed in teeth treated with calcium hydroxide apexica-
tion. This study found that the clinical and radiographic success rate
for calcium hydroxide apexication was 77.3% (healed), and for
MTA apexication it was 68.4% (healed). This result was somewhat
lower than that reported by El-Meligy and Avery (19), who reported
a success rate for calcium hydroxide apexication of 87% and for
MTA apexication 100%. Another nding in the present study was
differences in tooth survival. The tooth survival after calcium hydroxide
apexication was less than the other 2 treatments in this study. The
reason for extraction was nonrestorable root fracture. The duration
of calcium hydroxide apexication can range from 3 to 21 months
(19). During that time of treatment, the tooth is susceptible to reinfec-
tion from coronal leakage and may be prone to fracture because the
immature tooth with an open apex has a thin dentin wall and a short
root. In addition, Andreasen et al (3) found that long-term calcium
hydroxide dressing in the root canal weakens the root structure. In
this study, the time of a calcium hydroxide dressing in the root canal
of the calcium hydroxide apexication group was 14.5 months.
However, there were other factors that may have had an impact on
this outcome, such as restoration.
In conclusion, the present study provides an independent conr-
mation of the ndings of Bose et al (17) and indicates that revascular-
ization protocols offer the potential for treating the immature
permanent tooth with resulting resolution of signs and symptoms and
the initiation of continued root development. Moreover, the methods
described in this study, including standardized clinical follow-up check-
lists, provide the framework for continuing this study with both longer
follow-up periods and increased numbers of treated patients.
TABLE 3. The Success Rate of Immature Permanent Teeth after Treatment
Outcomes
Treatment
Revascularization
(n = 20)
MTA
(n = 19)
Calcium
hydroxide
(n = 22)
Teeth (%) Teeth (%) Teeth (%)
Healed* 16 (80.00) 13 (68.42) 17 (77.27)
Healing

4 (20.00) 5 (26.32) 0 (0.00)

Disease

0 (0.00) 1 (5.26) 5 (22.73)

*Both the clinical and radiographic presentations are normal.

The periapical radiolucency reduced combined with normal clinical presentation.

The radiolucency was either increased or persisted without change even when the clinical presen-
tation was normal or the clinical signs or symptoms were present regardless of the radiographic
presentation.
TABLE 2. The Survival Rate of Immature Permanent Teeth after Treatment
Outcome
Treatment
Revascularization
(n = 20) MTA (n = 19)
Calcium
hydroxide
(n = 22)
Teeth % Teeth % Teeth %
Survival
rate
20 100.00 18 94.70 17 77.30
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JOE Volume 38, Number 10, October 2012 Radiographic and Survival Outcomes of Immature Teeth 1335
Acknowledgments
The authors wish to thank Assistant Professor Panit Bandits-
ing, Department of Pediatric Dentistry, Mahidol University, for
providing cases for this study and Assistant Professor Chulaluk
Komoltri, Faculty of Medicine, Siriraj Hospital, for her help in statis-
tical analysis.
The authors deny any conicts of interest related to this study.
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