Salma El Ashry1, Ashraf Abu-Seida2, Amr

Bayoumi1 and Ahmed Hashem1

1Endodontic Department. Faculty of Dentistry. Ain Shams

University. 2Department of Surgery. Anesthesiology & Radiology.

Faculty of Veterinary Medicine. Cairo University

Pulp necrosis of an immature tooth as a result of caries or trauma could arrest further development of the root, leaving the tooth with thin and weak walls that are prone to fracture.

Endodontic treatment of such a tooth is

difficult because thin walls do not allow much instrumentation, and the obturation might not provide predictable apical seal due to the large opening of the apex.

Conventional treatment of such cases was

multiple-visit apexification using calcium

hydroxide. Although it was successful, it had

several disadvantages including;

unpredictability of apical closure, multiple

visits, probability of canal contamination

between visits, difficulty in follow-up and

subsequent treatment. In addition, long-term

root canal dressing by using calcium

hydroxide weakens the root structure and may

lead to future fracture of the root

Apical barrier technique was introduced as a replacement for apexification with calcium hydroxide. In the apical barrier technique a barrier material is placed at the apex to facilitate obturation procedure. Mineral trioxide aggregate (MTA) is the material of choice for this technique considering its sealing ability, biocompatibility, hard-tissue deposition potential, and the ability to set in the presence of moisture. However, the risk of future fracture may still exist

Recently, regenerative endodontic procedures have gained much attention as it allows root maturation to continue by the generated vital tissue. Presence of a smear layer inhibits the adherence of implanted pulp stem cells, potentially causing failure of the regenerative endodontic treatment. Its removal provides better sealing ability of the endodontic filling material to dentin, and prevents the leakage of microorganisms into oral tissues.

The most commonly used chemical chelating agent to remove the smear layer from root canal walls is 17% solution of EDTA that is used as a final flush. Therefore, the aim of the present study was to investigate the impact of surface modification of dentin using 17% EDTA irrigating solution on the regenerative potential following revascularization of immature permanent non-vital teeth.

This study was conducted according to

the ethical committee protocol at the

Faculty of Dentistry, Ain Shams University,

Egypt.

Sex mongrel dogs (4-6 months) were selected for

this study at Department of Surgery,

Anesthesiology, and Radiology, Faculty of

Veterinary Medicine, Cairo University-Egypt.

Sex dogs

Group I (2 dogs)

(2 weeks)

Blood Clot subgroup

(n=6 teeth)

Blood Clot+EDTA subgroup (n=6 teeth)

Group II (2 dogs)

(6 weeks)

Positive control

Subgroup (n=6 teeth).

Negative control subgroup (n=6

teeth)

Group III (2 dogs)

(12 weeks)

Induction of periapical pathosis: Under general anesthesia, an endodontic

access cavity was done in all experimental and positive control teeth.

The access cavity was left open for four

weeks. Dogs were monitored radiographically for the evidence of development of periapical pathosis. The operated dogs were given oral carprofen tablets daily for 15 days as a pain killer.

Root canal disinfection:

After the infection period, dogs were

anesthetized. The previously infected

experimental teeth were re-entered. The

canals were irrigated using 20 ml of 2.6%

sodium hypochlorite solution and filled

with 1-2 ml of the triple antibiotic paste.

The access cavity was then sealed using

temporary restoration for three weeks.

Treatment protocols:

The teeth were re-entered; the antibiotic

paste was removed by copious irrigation

using hypochlorite 2.6% solution. The

root canals were dried and treated

according to different treatment

protocols as follows:

Subgroup a (Blood Clot or

revascularization):

Hand file size no. 35 was inserted past to

the canal terminus until bleeding was

induced to fill the canal space just below

the cement-enamel junction. MTA orifice

plug was applied to seal the canal orifice.

The access cavity was then sealed using

glass ionomer filling.

Subgroup b (Blood clot and EDTA): Each canal was irrigated with 1ml of 17%

EDTA solution which was kept for 2 minutes inside the canal. The canal was rinsed with another 1ml of 17% EDTA, flushed with normal saline solution and dried by paper points. Similar blood clot induction, sealing of canal orifice and access cavity were carried out as in subgroup (a).

Subgroup c (Positive control):

This subgroup represented the pulp

exposed teeth. Cotton pellets were

inserted into the canal space and no

temporary filling was applied.

Subgroup d (Negative control):

This subgroup represented the

untouched normal teeth.

Radiographic evaluation:

Periapical radiographs were taken before and after induction of the periapical lesion and compared with follow up radiographs taken according to the group. These radiographs were digitized. TurboReg plug-in software was used to standardize these radiographs. Increase in the root length, root thickness and decrease in apical diameter was measured.

Histopathological evaluation:

Assessment of the periapical inflammatory cell scores, periapical inflammatory cell, tissues in-growth in the pulp space, new hard tissue formation, bone / root resorption and apical closure were assessed.

Statistical analysis was performed with

IBM®SPSS®.

Incre

ase

in

th

e

root

len

gth

Subgroups Group I Group II Group III

Subgroup (a) 4.9±1.2 a 13.9±1.8 a 16.3±1.3 a

Subgroup (b) 5.1±1 a 14±1.6 a 16.5±1.4 a

Subgroup (c) 0±0 b 0±0 b 0±0 b

Subgroup (d) 6.2±0.5 a 15.5±0.6 a 19.7±0.4 a

P-value <0.001* <0.001* <0.001*

Incre

ase

in

th

e

root

thic

kn

ess

Subgroup (a) 4±0.6 a 11.4±0.9 a 13.4±0.8 a

Subgroup (b) 4.7±1 a 11.6±0.9 a 13.6±1.3 a

Subgroup (c) 0±0 b 0±0 b 0±0 b

Subgroup (d) 6.1±0.7 a 12.4±1.3 a 15.8±.1 a

P-value <0.001* <0.001* <0.001*

Ap

ical

Clo

sure

Subgroup (a) 2.9±0.9 b 16.4±1.4 b 30.2±1.9 b

Subgroup (b) 3.1±0.9 b 16.5±1.1 b 30.8±1.7 b

Subgroup (c) 0±0 c 0± 0c 0±0 c

Subgroup (d) 6.2±0.6 a 20.5±1.2 a 47±1 a

P-value <0.001* <0.001* <0.001*

*: Significant at P ≤ 0.05, Different letters in the same column are statistically significantly different according to Tukey’s test

Figure 1: Upper radiographs of subgroup (a) (blood clot): preoperative (a) and 2 weeks (b), 6

weeks (c) and 3 weeks (d) following treatment. Lower radiographs of subgroup (c), pulp

exposed teeth (positive control): o days (a), 2 weeks (b), 6 weeks (c) and 3 weeks (d).

Infl

am

ma

tory

sco

res

Subgroups Group I Group II Group III

Subgroup (a) 2.1±0.9 a 1.3±0.5 b 0.4±0.5 b

Subgroup (b) 2.3±0.9 a 1.4±0.5 b 0.6±0.5 b

Subgroup (c) 2.7±0.5 a 2.9±0.4 a 3±0 a

Subgroup (d) 0±0 c 0 ±0c 0±0 c

P-value 0.001* <0.001* <0.001*

Infl

am

ma

tory

cel

l co

un

t

Subgroup (a) 25±1.6 a 16.4±2 b 10.2±1.4 b

Subgroup (b) 25.5±2 a 16.6±1.8 b 10.4±1.7 b

Subgroup (c) 28.4±1.3 a 36.7±1.9 a 38.9±2.9 a

Subgroup (d) 2.5±0.9 c 2.3±1.2 c 2.1±0.9 c

P-value <0.001* <0.001* <0.001*

*: Significant at P ≤ 0.05, Different letters in the same column are statistically significantly different.

Tis

sue

in-g

row

th

Subgroups Group I Group II Group III

Subgroup (a) 0.6±0.5 a 1.3±0.8 b 1.9±0.4 b

Subgroup (b) 0.9±0.4 a 1.9±0.7 a 2.6±0.5 a

Subgroup (c) 0.3±0.5 b 0.9±0.4 b 0.9±0.4 c

P-value 0.099 0.001* <0.001*

Min

era

liza

tion

sco

re

Subgroup (a) 0.4±0.5 a 0.7±0.5 a 1.3±0.5 a

Subgroup (b) 0.6±0.5 a 0.9±0.4 a 1.4±0.5 a

Subgroup (c) 0±0 b 0±0 b 0±0 b

P-value 0.166 0.003* <0.001*

*: Significant at P ≤ 0.05, Different letters in the same column are statistically significantly different according to Mann-Whitney U test

Bon

e re

sorp

tion

Subgroups Group I Group II Group III

No % No % No %

Subgroup (a) 9 75 5 41.6 2 16.6

Subgroup (b) 9 75 5 41.6 2 16.6

Subgroup (c) 10 83.3 12 100 12 100

Subgroup (d) 0 0 0 0 0 0

P-value 0.007* 0.038* 0.002*

Ap

ical

closu

re

Subgroup (a) 0 0 4 33.3 6 50

Subgroup (b) 0 0 4 33.3 6 50

Subgroup (c) 0 0 0 0 0 0

Subgroup (d) 0 0 6 50 7 58.3

P-value NC** 0.295 0.098

Figure 2: (a) Photomicrograph of subgroup II (a) showing dense fibrous connective

tissue in-growth up to apical third of the canal and numerous blood vessels (H & E X

100). (b) Photomicrograph of subgroup III (b) showing tissue in-growth up to middle

third of the canal (H & E X 40). (c) Photomicrograph of subgroup III (b) showing

hard tissue deposition (H & E X100).

Final rinse using 17% EDTA

solution before revascularization

has a positive impact on tissue

interaction along dentinal walls

without modification of the cell

type.