ORIGINAL ARTICLE

Histologic analysis of pulpal revascularization

of autotransplanted immature teeth after
removal of the original pulp tissue
Inne Claus, DDS,a Wim Laureys, DDS,a Ria Cornelissen, DDS, PhD,b and Luc R. Dermaut, DDS, PhDc
Gent, Belgium

The survival rate of replanted and autotransplanted teeth is mainly affected by the reaction of the pulp. Pulpal
necrosis can cause periapical inflammation and inflammatory root resorption. The purpose of this study is to
learn more about the pulpal changes in autotransplanted immature teeth whose pulp tissue was removed
before transplantation. The experimental material consisted of 16 single-rooted teeth with open apices, from
a beagle dog (3 months of age). At day 0, 4 teeth were extracted, the pulpal tissues were removed, and the
teeth were then transplanted to their contralateral side. The same procedure was carried out on days 9, 16,
and 23, each time for 4 single-rooted teeth. Longitudinal paraffin sections were made for histologic
investigation. The results showed that, after 7 days, 2 of the 4 teeth had an ingrowth of new tissue over one
fourth of their length. After 14 days, all 4 teeth had ingrowth (ⱖ one fourth of the pulp chamber). At the 21-day
observation, more than half of the pulp chambers of all teeth were filled, and, after 30 days, there was total
ingrowth in 3 of the 4 teeth. This new tissue consisted of well-organized and well-vascularized connective
tissue. (Am J Orthod Dentofacial Orthop 2004;125:93-9)

tissue, Laureys et al8 investigated the revascularization

A
utotransplantation of teeth in humans is now
frequently carried out. Third molars with incom-
plete root formation are transplanted to sites of
agenetic premolars or cariously involved first molars.1,2 In
the case of avulsed central incisors, mandibular first
premolars are also transplanted to replace the incisors.
The survival rate of replanted or autotransplanted
teeth is often affected by the reaction of the pulp.3
These teeth should have open apices to maintain the
vitality of the teeth. Pulpal necrosis can cause periapi-
cal inflammation and inflammatory root resorption.4,5 It
is known from the studies by Skoglund and Tronstad6
and Skoglund7 that, as a rule, the original pulp of
autotransplanted immature and mature apicoectomized
teeth becomes necrotic after transplantation. Repair
occurs by ingrowth of well-vascularized cell-rich con-
nective tissue, but a reduction of cells and blood vessels
occurs after a period of time. Finally, the new tissue
resembles bone or cementum for most parts of the pulp
cavity; this leads to obliteration.
Because of the difficulty of cryopreserving pulp
From the University of Gent.
a
Instructor, Department of Orthodontics.
b
Associate professor, Department of Anatomy, Embryology and Histology.
c
Head, Department of Orthodontics.
Reprints requests to: Inne Claus, U.Z.—Tandheelkunde, De Pintelaan 185
B-9000, Gent, Belgium; e-mail, inneclaus@yahoo.com.
Submitted, October 2002; revised and accepted, February 2003.
0889-5406/$30.00
Copyright © 2004 by the American Association of Orthodontists.
doi:10.1016/S0889-5406(03)00619-X
of immature and mature apicoectomized teeth after
cryopreservation and autotransplantation. The original
pulp tissue was removed from these teeth before
transplantation. The penetration of cryoprotective
agents into the pulp chamber is poor. This problem can
be solved by removing the original pulp before cryo-
preservation. Laureys et al8 showed different amounts
of revascularization after removal of the original pulp
in immature teeth with open apices and in mature teeth
in which an open apex was created by an apicoectomy.
They also found that cryopreservation did not compro-
mise the amount of revascularization compared with
direct transplantation. Emphasis was put on “revascu-
larization” registered with microangiography. No infor-
mation about the histologic status in and around the
pulp chamber was provided. This information could
help to confirm or reject the hypothesis of Laureys et
al8 that pulp removal in combination with cryopreser-
vation could be an advisable clinical procedure.
The purpose of this study was to obtain, with
histologic techniques, a more detailed picture of the
properties of new pulp in immature teeth (without
cryopreservation) when the original pulp tissue was
removed before transplantation.
MATERIAL AND METHODS
The experimental material consisted of 16 single-
rooted teeth, incisors and first premolars, from a beagle
93
94 Claus et al American Journal of Orthodontics and Dentofacial Orthopedics
January 2004

Table. Number of experimental teeth for different

observation periods
Number of transplanted Observation
Day teeth period

0 2 maxillary first premolars 30 days

with open apices
2 mandibular first
premolars with open
apices
9 2 maxillary incisors with 21 days
open apices
2 mandibular incisors with
open apices
16 2 maxillary incisors with 14 days
open apices
2 mandibular incisors with
open apices
23 2 maxillary incisors with 7 days
open apices
2 mandibular incisors with
open apices
30 dog was killed

dog that was 3 months old at the beginning of the

experiment.9-12 The teeth had open apices at the time of
autotransplantation.
At day 0, the dog was anesthetized. Four teeth (first
premolars) were extracted, the pulpal tissues were
removed from the apical side with a nerve broach, and
the teeth were transplanted to their contralateral side.
No surgical enlargement of the alveolus was carried out
before the transplantation to prevent extra manipula-
tion. Due to the straight roots of these immature teeth,
no problems were experienced during transplantation.
This procedure was done by 2 investigators (I.C. and
W.L.) to minimize the extra-alveolar time. The trans-
planted teeth were fixed with a surgical suture to
maintain their physiologic mobility.
The same procedure was carried out after 9, 16, and
23 days, each time for 4 single-rooted teeth. The
weekly surgical interventions did not interfere with the
healing of the previously transplanted teeth because
there was no proximal contact between the different
transplanted teeth. The observation periods were at 7,
14, 21, and 30 days (Table).
The dog received soft food and oral penicillin
during the experiment.
At day 30, the dog was killed by an overdose of
barbiturate solution. The jaw sections, including the
teeth and the alveolar bone, were excised and fixed in
4% neutral buffered formaldehyde (pH 7.4) and decal-
cified in 25% EDTA. Longitudinal 5 ␮m-thick paraffin
sections were taken from the teeth in a buccolingual
direction and stained with hematoxylin and eosine. For
Fig 1. Schematic representation of ingrowth of new
tissue. , Immature tooth at start of experiment; d,
numbers of days posttransplantation (observation
period); , extent of revascularization; numbers, tooth
numbers; , empty area.
control purposes, the second premolars of each quad-
rant were also sectioned.
RESULTS
Normal dental pulp (control teeth) consists of con-
nective tissue, resembling primitive mesenchyma. It
contains stellate fibroblasts and a rich network of
capillaries, arterioles, and small veins. In connection
with the pulp tissue, odontoblasts form a pseudostrati-
fied layer of columnar cells. Internal or external root
resorption was not seen.
One week after the transplantation, the pulp cavity
of all 4 incisors (Fig 1) was partially filled with
remnants of blood clot (apical one third) originating
from the extraction trauma (Fig 2, A). Inflammatory
material, mainly lymphocytes and plasma cells, was
seen in the middle part of some teeth. This inflamma-
tory material became necrotic. Most of the odontoblas-
tic layer was absent due to the extirpation of the
American Journal of Orthodontics and Dentofacial Orthopedics
Volume 125, Number 1
Fig 2. Ingrowth of new tissue after 7 days; a, remnants of blood clot; b, well-vascularized cell-rich
connective tissue.
original pulp tissue during the experiment, although
some odontoblasts were still present.
No ingrowth of new tissue was seen in the 2
maxillary transplanted incisors, although, in the 2
mandibular transplanted incisors, 25% of the pulp
cavity was filled with new tissue: a cell-rich connective
tissue. The onset of formation of cell-rich connective
tissue can be observed in the apical part of the pulp
chamber (Fig 2, B).
Two weeks after transplantation, some ingrowth of
new tissue occurred in the apical part in 3 of 4
transplanted incisors, and, in 1 incisor, half of the pulp
cavity was filled with new tissue that contained many
spindle-shaped cells embedded in a collagen matrix
(Fig 3). This cell-rich connective tissue was highly
vascularized.
Three weeks postoperatively, half of the pulp cavity
was filled in the 2 maxillary transplanted incisors, while
75% of the pulp cavity was filled in the 2 mandibular
Claus et al 95
transplanted incisors (Fig 4). Necrotic material and
lymphocytes were still present coronally.
After approximately 4 weeks (30 days), 1 maxillary
transplanted premolar had an ingrowth of 75%, and
another was 80% filled. The 2 mandibular transplanted
premolars had completely filled pulp cavities.
The histologic picture (Figs 4 and 5) in both groups
(21 and 30 days postoperatively) was comparable.
Some remnants of the blood cloth were still present
coronally, but most of the pulp chamber was filled with
well-organized connective tissue. In the middle part,
this tissue contained spindle-shaped cells (Fig 5, C).
Inflammatory cells—lymphocytes—were located not
only coronally but also more apically in the connective
tissue.
The blood vessels can be differentiated into arterial
and venous types (Fig 5, A). In the apical part of the
pulp chamber, islands of bone with osteocytes and
osteoblasts were present in the connective tissue (Fig 5,
96 Claus et al
Fig 3. Ingrowth of new tissue after 14 days; a, inflammatory material, mainly lymphocytes and
plasmacells; b, new tissue consists of cell-rich vascularized connective tissue.
B). A cell-rich connective tissue surrounded the trans-
planted elements (Fig 5, D).
Two of the 16 transplanted teeth showed signs of
internal root resorption, and 3 had external resorption.
Inflammatory cells were located near the internal re-
sorption area. Most of these cells were lymphocytes,
but macrophages were also seen in small numbers. In
the areas of external resorption, no osteoclasts or
osteoblasts could be found. In 1 tooth, the external
resorption process had reached the pulp cavity. Internal
resorption was not necessarily accompanied by external
resorption.
Five teeth showed further root formation, but 4
did not. We did not have adequate sections of the
other 7 teeth to evaluate root formation. The teeth
with further root formation had a layer of odonto-
blasts along the dentinal wall. No obvious remnants
of Hertwig’s epithelial sheath were observed in the
studied sections.
American Journal of Orthodontics and Dentofacial Orthopedics
January 2004
DISCUSSION
Several studies8,13,14 have shown revascularization
in replanted and autotransplanted immature teeth whose
original pulp tissue was not removed. Skoglund et al13
found with microangiography that revascularization of
the pulp occurs mainly by ingrowth of new vessels.
According to Skoglund et al,13 anastomoses occur to
pre-existing vessels in the pulp of only a few teeth.
In our study, the original pulp tissue was removed
before transplantation of the experimental teeth. There-
fore, anastomoses to the original vessels could not be
formed. However, this did not seem to influence the
ingrowth of new tissue. These findings corresponded
well with a previous study of Laureys et al,8 who
showed revascularization through ingrowth of new
tissue by means of microangiography.
Seven days postoperatively, pulpal repair was seen
in the apical area. Twenty-one days postoperatively,
American Journal of Orthodontics and Dentofacial Orthopedics
Volume 125, Number 1
Fig 4. Ingrowth of new tissue after 21 days; further root formation is observed. Dentin is surrounded
by epithelial-like structure.
75% of the pulp chamber was filled, and, after 30 days,
complete repair was seen in 2 of 4 teeth. Thus, the
longer the observation period, the better the ingrowth of
new tissue. This finding was also observed by Skoglund
and Tronstad6 and Monsour,15 although the original
pulpal tissue was not removed in their studies.
There was a difference in the amount of ingrowth of
new tissue in the mandibular and maxillary transplanted
teeth. The success rate in the mandibular teeth was
higher. Better results were also obtained with transplan-
tation of premolars than with incisors: total ingrowth
can be attained sooner because the first premolars are
much shorter than the incisors. This is in contradiction
with the findings of Laureys et al,8 who concluded that
ingrowth in premolars seemed to occur more slowly
than in incisors. This could be explained by the fact
that, in their study, all transplanted premolars were
apicoectomized, while half of the transplanted incisors
were immature, and half were apicoectomized. In our
Claus et al 97
study, however, only immature teeth were transplanted.
Perhaps, while apicoectomizing the apices in the study
of Laureys et al,8 a difference in width of the new
foramina was created in the premolars compared with
the incisors. If, after apicoectomy, the foramina of the
premolars were smaller than those of the incisors,
ingrowth of new tissue could be attained sooner in the
incisors. This hypothetical explanation needs further
research.
Histologic findings of the new tissue reported by
Skoglund and Tronstad6 show a well-vascularized,
cell-rich connective tissue. Thirty days postoperatively,
a few teeth still had an odontoblastic layer, which had
formed tubular dentin and was present along the entire
length of the pulp chamber. The same results were
found in our study with the exception of the odonto-
blastic layer, which was mostly removed by extirpating
the original pulp.
Bone tissue with osteocytes and osteoblasts was
98 Claus et al American Journal of Orthodontics and Dentofacial Orthopedics
January 2004

Fig 5. Detail ingrowth of new tissue after 4 weeks: more organized and vascularized connective
tissue; a, arterial and venous blood vessels; b, islands of bone with osteocytes and osteoblasts; c,
connective tissue with spindle-shaped cells; d, cell-rich connective tissue surrounding elements.

also observed, although the amount of hard tissue was
very small and did not occupy large parts of the pulp
cavity. This is in accordance with the studies of
Skoglund and Tronstad,6 in which a cell-containing,
atubular hard tissue was seen at the root canal wall.
After 180 days of transplantation, they found a bone-
like tissue formed centrally in the pulp, and, in some
teeth, this tissue was connected to the alveolar bone
through the open apical foramen.
It is suggested that pulpal necrosis is the main cause
of periapical inflammation and inflammatory root re-
sorption.4,5 Because, in our study, the original pulp
tissue was removed before transplantation, no necrosis
occurred postoperatively, although 2 transplanted teeth
showed small areas of internal root resorption. Re-
moval of the blood clot accompanied by some inflam-
mation could explain this observation. Thus, even
without pulpal necrosis, internal root resorption might
occur. Three of 16 teeth showed external root resorp-
tion. This is much more favorable than the number of
teeth with external root resorption in the study of
Skoglund and Tronstad.6 They found that all trans-
planted teeth (65 single-rooted teeth) were affected
after 30 days.
There was a cell-rich connective tissue between the
alveolar bone and the teeth. This connective tissue was
not yet well organized. Andreasen et al16 demonstrated
periodontal healing radiographically and found partial
healing after 4 weeks. Most transplants showed com-
plete periodontal healing after 8 weeks. Andreasen et
al16 defined periodontal healing by “a newly formed
periodontal ligament space surrounding the entire root
periphery,” although no information was given about
the histologic properties of this periodontal ligament. In
this study, the observed tissue along the roots therefore
could be described as a possible periodontal ligament,
but our histologic sections did not enable us to define it
as such.
Some teeth showed signs of further root formation.
These teeth had a layer of odontoblasts along the
dentinal wall, forming dentin during the new root
development. Odontoblasts are formed as epithelial
American Journal of Orthodontics and Dentofacial Orthopedics
Volume 125, Number 1
cells of the external and internal dental epithelium
proliferate to form a double layer of cells known as
Hertwig’s epithelial root sheath.17 This typically dou-
ble layer of cells could not be found in our sections,
although the odontoblastic layer surrounded the end of
the root. The teeth with no further root formation did
not have such an odontoblastic layer, almost certainly
due to the extirpation of the pulp. It was never our
intention to concentrate on the histologic development
of the root. The histologic sections were not chosen
accordingly.
CONCLUSIONS
This study clearly shows that ingrowth of new
cell-rich and well-vascularized connective tissue occurs
in immature transplanted teeth after removing the
original pulp.
We thank Dr I. De Cock for anesthetizing the
animals, Dr B. De Smet, the director of the animal
laboratory, and G. Dermout for the illustrations.
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