228

Volume 45 Number 4 December 2012

Research Report

The increasing of fibroblast growth factor 2, osteocalcin, and 
osteoblast due to the induction of the combination of Aloe vera 
and 2% xenograft concelous bovine 

utari Kresnoadi
Department of Prosthodontics
Faculty of Dentistry, Universitas Airlangga
Surabaya - Indonesia

abstract

Background: To make a successfull denture prominent ridge is needed, preservation on tooth extraction socket is needed in order 
to prevent alveol bone resorption caused by revocation trauma. An innovative modification of the material empirically suspected to be 
able reduce inflammation caused by the revocation trauma is a combination of Aloe vera and xenograft concelous bovine (XCB) and 
Aloe vera is a biogenic stimulator and accelerating the growth of alveolar ridge bone after tooth extraction. Purpose: The research was 
aimed to determine of the increasing alveol bone formation by inducing the combination of Aloe vera and 2% xenograft concelous bovine. 
Methods: To address the problems, the combination of Aloe vera and xenograft concelous bovine was induced into the tooth extraction 
sockets of Cavia cabayas which devided on 8 groups. Groups control, filled with XCB, Aloe vera and Aloe vera and XCB combination, 
at 7 days and 30 days after extraction. Afterwards, immunohistochemical examination was conducted to examine the expressions of 
FGF-2 and osteocalcin, as the product of the growth of osteoblasts. results: There were significantly increases expression of FGF-2 
and osteocalcyn on group which filled with XCB, Aloe vera and combined Aloe vera and XCB. Conclusion: It may be concluded that 
the induction of the combination of Aloe vera and xenograft concelous bovine into the tooth sockets can enhance the growth expressions 
of FGF-2 and osteocalcin as the product of osteoblasts, thus, the growth of alveolar bone was increased.

Key words: Fibroblast growth factor 2, osteocalcin, osteoblast, alveolar bone, combination of Aloe vera and graft

abstrak

latar belakang: Untuk keberhasilan pembuatan gigitiruan diperlukan ridge yang prominent, maka diperlukan suatu preservasi 
soket pencabutan gigi untuk mencegah terjadinya resopsi tulang alveolar akibat trauma pencabutan. Suatu inovasi modifikasi bahan 
yang diduga secara empiris dapat mengurangi keradangan karena trauma pencabutan adalah berupa kombinasi Aloe vera dan xenograft 
concelous bovine (XCB). Aloe vera yang merupakan biogenik stimulator untuk merangsang dan mempercepat pertumbuhan tulang 
alveolar setelah pencabutan gigi. tujuan: untuk membuktikan bahwa induksi kombinasi Aloe vera dan xenograft concelous bovine 
2% pada soket pencabutan gigi, dapat meningkatkan pembentukan tulang alveolar. Metode: Pengisian kombinasi Aloe vera dan XCB 
pada soket pencabutan gigi Cavia cabaya yang dibagi dalam 8 kelompok. Kelompok: kontrol, Aloe vera, XCB, kombinasi Aloe vera 
dan XCB, masing-masing dalam pemeriksaan 7 dan 30 hari. Kemudian dilakukan pemeriksaan imunohistokimia ekspresi FGF-2 dan 
osteocalcyn sebagai produk pertumbuhan sel osteoblas. hasil: Terdapat peningkatan ekspresi FGF-2 dan osteocalcyn pada kelompok 
yang diisi XCB, Aloe vera dan kombinasi Aloe vera + XCB dibanding dengan kelompok kontrol. Kesimpulan: Induksi kombinasi Aloe 
vera dan xenograft concelous bovine 2% pada soket pencabutan gigi dapat, meningkatkan ekspresi FGF-2 dan osteocalcyn, sehingga 
dapat meningkatkan pertumbuhan tulang alveol.

Kata kunci: Fibroblast growth factor 2, osteocalcin, osteoblas, tulang alveolar, kombinasi Aloe vera dan graft 

Correspondence: Utari Kresnoadi, c/o: Departemen Prostodonsia, Fakultas Kedokteran Gigi Universitas Airlangga. Jl. Mayjend. Prof. 
Dr. Moestopo 47 Surabaya 60132, Indonesia. E-mail: ut.kres@yahoo.com 

 



229Kresnoadi: The increasing of fibroblast growth factor 2, osteocalcin, and osteoblast: The increasing of fibroblast growth factor 2, osteocalcin, and osteoblastThe increasing of fibroblast growth factor 2, osteocalcin, and osteoblast 

introduction

The success of denture manufacture requires supporting 
anatomical condition of oral cavity, especially prominent 
ridge. To achieve the good condition of ridge, after tooth 
extraction, the ridge must be prevented from resorption first. 
Some users of fixed denture and partially removable denture 
feel uncomfortable with the dentures due to sustained 
resorption of residual ridge after tooth extraction, and this 
situation may cause structural changes in jaw bone. Level 
of residual ridge resorption in each person, however, is not 
the same, so we need to conduct the prevention of residual 
ridge resorption. 1 

After tooth extraction, the alveolar ridge usually 
decreases about 50%, and this condition occurs in the 
period of 6-12 months. The decreasing even occurs several 
times due to residual ridge resorption that physiologically 
occurs after tooth extraction.2 Residual ridge resorption is 
a biomechanical disease caused by multi-factors, such as 
anatomic, metabolic, and mechanical determinants.3 After 
tooth extraction, there will usually be inflammation as rapid 
reaction in the area of extraction, and the sockets of the 
extraction will be filled with blood clot. Later, the epithelial 
tissue will be proliferated and migrated in one week, and the 
tissue will fix the sockets of the extraction quickly. 

The use of bovine graft to improve either damaged bone 
or augmentation is actually often conducted in the field 
of general surgery and oral surgery. However, the use of 
bovine graft still can not give satisfactory results. Therefore, 
we need an innovative material, biogenic stimulator, to 
stimulate graft that can accelerate bone growth. 

Aloe vera has been considered as biogenic stimulator 
and hormonal activator in healing wound. Aloe vera 
extract can be used to prevent scar tissue during incision, 
and when Aloe vera gel is used after surgery, the incision 
will heal quickly.4 Significant changes of the amount of 
collagen of the control and treatment groups indicate how 
Aloe vera can stimulate fibroblast cell growth and wound 
healing process.5 Thus, this research was aimed to prove 
whether the induction of the combination of Aloe vera 
and 2% xenograft concelous bovine (XCB) into tooth 
extraction sockets can increase the expression of FGF2 
and osteocalcin, which then may cause the increasing of 
alveolar bone growth. 

materials and methods

This research was an experimental research with 
randomized post-test control group design with the 
combination of Aloe vera and 2% XCB. The combination 
of Aloe vera and 2% XCB can indicate that each active 
substance contained is about 2%. XCB is the product 
of Bank of tissue of Dr. Soetomo Hospital Surabaya. 
Other materials used in this research consist of Aloe vera 
extracts, sterile distilled water, XCB, absolute alcohol, 70% 
alcohol, anti-FGF2 monoclonal antibodies, anti osteokalcin 

monoclonal antibodies, and reagents. Those materials were 
used for immunohistochemical examination, and reagents 
used for staining was Hematoxilin-eosin (HE). Instruments 
used in this research, moreover, were a set of tools for 
both immunohistochemical examination and preparation 
manufacture, such as micropipette, tip (yellow, white, blue), 
light microscopy, object glass, and cover glass. 

Animals used in this research, furthermore, were healthy 
and active male Cavia cobaya (marmots) with 300 -350 
grams of weight and in the age of 3 to 3.5 months. Those 
animals were then divided into 8 groups. Next, lower right 
incisor extraction was then conducted in all those groups. 
Firstly, Cavia cobaya were taken from their care center, 
and then anaesthetized with intra venous ketamine 0.2 
cc/300g BM.6 The tooth extraction process was conduced 
by using special pliers (needle holder). After the tooth 
extraction, the sockets were filled with materials as much 
as 0.1 cc, and then stitched. The materials were loaded as 
follows: group I: the sockets of tooth extraction were filled 
with polyetilene glycol (PEG) as the control group, and 
then examined after 7 days; group II: the sockets of tooth 
extraction were filled with PEG (as the control group), and 
then examined after 30 days; group III: the sockets of tooth 
extraction were filled with xenograft concelous bovine 
(XCB) and PEG, and then examined after 7 days; group 
IV: the sockets of tooth extraction were filled with XCB 
and PEG, and then examined after 30 days; group V: the 
sockets of tooth extraction were filled with Aloe vera and 
PEG, and then examined after 7 days; group VI: the sockets 
of tooth extraction were filled with Aloe vera and PEG, and 
then examined after 30 days; group VII: the sockets of tooth 
extraction were filled with Aloe vera, XCB, and PEG, and 
then examined after 7 days; and group VIII: the sockets of 
tooth extraction were filled with Aloe vera, XCB, and PEG, 
and then examined after 30 days. 

After 7 days and 30 days, after being decalcified with 
2% nitric acid, the jaws of those tested animals were cut 
to make preparations of block paraffin. Afterward, slides 
were made for staining with imunnohistochemistry and 
hematoxilin eosin (HE). Next, the calculation was conducted 
on FGF-2, osteocalcin, and immunohistochemical staining 
by using microscopic examination, and then the amount 
of osteoblasts and osteoclasts was calculated on HE 
staining.

The tested animals were taken care at Biochemical 
Laboratory of Medical Faculty, Universitas Airlangga, 
while the manufacture of tissue preparation was conducted 
at Histology Laboratory and Pathology Anatomi Laboratory 
of Dr. Soetomo Hospital. Moreover, the manufacture of 
Aloe vera extract was conducted at Physical Chemistry 
Laboratory of Pharmacy Faculty, Universitas Airlangga, 
while the manufacture of Aloe vera frezze dryed was 
conducted at Biology Laboratory of Science and Technology 
Faculty, Universitas Airlangga. And, immunohistochemical 
examination was conducted at Immunohistochemistry 
Laboratory, LPT Tropical Disease Center (TDC) of 
Universitas Airlangga, and at Biomedical Laboratory of 



230 Dent. J. (Maj. Ked. Gigi), Volume 45 Number 4 December 2012: 228–233

Universitas Brawijaya. The calculation results were then 
recorded and tabulated. Afterwards, the result data were 
analyzed by using ANOVA, and then multiple comparison 
of the data of those four groups was conducted.

results

The increasing of the expressions of FGF-2 and 
osteocalcin in the control group and in the treatment groups 
induced with XCB, Aloe vera, and XCB and Aloe vera on 
day 7 and day 30 can be seen in Figure 1. Based on Figure 
1, it can be indicated that the expressions of FGF2 and 
osteocalcin in those three treatment groups induced with 
XCB, Aloe vera, and Aloe vera + XCB were higher than 
those in the control group. However, the highest expressions 
of FGF-2 and osteocalcin were found in the group induced 
with Aloe vera + XCB on day 30. The expression of FGF-2 
in those four groups can be seen in Figure 2.

 Based on the Figures 2, it is known that the expressions 
of FGF-2 in the treatment groups induced with XCB (2b), 
Aloe vera (2c), and Aloe vera and XCB (2d), increased 
more as pointed by black arrows than those in the control 
group (2a). Meanwhile, the expressions of osteocalcin on 
day 30 of immunohistochemical examination can be seen 
in Figures 3a, b, c, d.

Based on Figures 3, it is known that the expressions of 
osteocalcin in the treatment groups induced with XCB (3b), 

Aloe vera (3c), and Aloe vera + XCB (3d) increased more as 
pointed by black arrows than those in the control group (3a). 
Based on ANOVA analysis, moreover, it is also known that 
there was significant difference of the expressions of FGF-
2, p <0.05, between the control group and the treatment 
groups induced with XCB, Aloe vera, and Aloe vera + 
XCB. Similarly, there was also significant difference of 
the expressions of osteocalcin, p<0.05, between the control 
group and the treatment groups induced with XCB, Aloe 
vera, and Aloe vera + XCB.

figure 1.  The average expressions of FGF-2 and osteocalcin 
in the control group, in the treatment groups induced 
with XCB+PEG, Aloe vera + PEG, and Aloe vera + 
XCB + PEG on day 7 and on day 30.

figure 2.  a) The expression of FGF-2 on day 30 in the control group; b) The expression of FGF-2 on day 30 in the group induced 
with XCB; c) The expression of FGF-2 on day 30 in the group induced with Aloe vera; d) The expression of FGF-2 on day 
30 in the group induced with Aloe vera dan XCB. Black arrows indicate the expressions of FGF-2.

FGF-2

Osteocalcyn

a b

c d



231Kresnoadi: The increasing of fibroblast growth factor 2, osteocalcin, and osteoblast: The increasing of fibroblast growth factor 2, osteocalcin, and osteoblastThe increasing of fibroblast growth factor 2, osteocalcin, and osteoblast 

Furthermore, based on Tukey HSD analysis, there 
was significant difference of the expressions of FGF-2, 
p<0.05, between the control group and the groups induced 
with XCB, Aloe vera, and Aloe vera + XCB. There was 
also significant difference of the expression of osteocalcin 
between the control group and the treatment groups induced 
with XCB, Aloe vera, and Aloe vera + XCB. However, there 
was no significant difference between the group induced 
with Aloe vera and the group induced with XCB. 

Next, the expressions of FGF-2 and osteocalcin on 
day 7 and on day 30 were analyzed by using t-test. The 

results then showed that there was significant difference of 
the expressions of FGF-2 and osteocalcin, t<0.05, started 
from day 7 to day 30. Finally, how the increasing of the 
expressions of FGF-2 and osteocalcin stimulated the growth 
of osteoblast can be seen in Figure 4.

Based on Figure 4, it is known that the growth of 
osteoblasts in the control group and in the treatment 
groups induced with XCB, Aloe vera, and Aloe vera + 
XCB increased from day 7 to day 30 (see red bar). And, 
the highest growth of osteoblasts occurred in the group 
induced with Aloe vera+ XCB on day 30.

discussion

Based on Figure 1, it is known that that the expressions 
of FGF2 and osteocalcin in the control group and in the 
treatment groups induced with XCB, Aloe vera, and Aloe 
vera + XCB increased started from day 7 to day 30 of the 
examination. The highest FGF-2 expression and osteocalcin 
were found in the group induced with Aloe vera + XCB 
started from day 7 to day 30 of the examination. It indicates 
that Aloe vera + xenograft concelous bovine can stimulate 
the growth of alveolar bone in the socket of tooth extraction. 
Healing process with graft induction can prevent alveolar 
bone from resorption prior to the formation of new alveolar 
bone.7 Thus, graft material used must be biocompatible, so 
it can inhibit bone resorption and stimulate osteogenesis. 

Figure 3.  a) The expression of osteocalcin on day 30 in the control group; b) The expression of osteocalcin on day 30 in the group induced 
with XCB; c) The expression of osteocalcin on day 30 in the group induced with Aloe vera; d) The expression of osteocalcin 
on day 30 in the group induced with Aloe vera and XCB. Black arrows indicates the expression of osteocalcin

figure 4.  The average and standard deviation of osteoblast cells 
in the control group and the treatment groups induced 
with XCB, Aloe vera, and Aloe vera + XCB after HE 
on day 7 and on day 30.

a b

c d

co
ntr

ol 
7

co
ntr

ol 
30

Osteoblast

Osteoblast



232 Dent. J. (Maj. Ked. Gigi), Volume 45 Number 4 December 2012: 228–233

This opinion is also supported by some experts stating 
that graft induced into tooth extraction sockets can cause 
osteoinduction, stimulating bone growth.8-11

The increasing of osteocalcin expression can also be 
caused by osteocalcin mostly produced by osteoblasts. 
There are actually collagen and non-collagen proteins in 
osteoblasts, which usually synthesize non-collagen protein 
in bone matrix, namely osteocalcin and osteonektin as 
much as 40–50% in bone.12 Fibroblast growth factor 
2, furthermore, is an important modulator for both the 
growth of bone as well as the differentiation, expression 
and regulation of osteoblast.11 In addition, FGF-2 is also 
an important signaling molecule for the regulation of bone 
development stages, thus, if FGF-2 increases, the amount 
of osteoblasts will also increase.13,14 

The expressions of FGF-2, osteocalcin, and osteoblast 
in the group induced with Aloe vera were higher than those 
in the group induced with XCB. It indicates that Aloe vera 
is an anti-inflammatory, antibacterial, anti-virus because 
it contains not only antthraquinones components, namely 
aloin, aloe emodin, and barbaloin, but also pure mannan 
carbohydrate, acetyl mannan glukomannan, as well as 
alkaline phosphatase and bradikinase enzymes. There are 
also healing hormones, auxin, gibberellin, and saponins, 
as well as proteins in Aloe vera that can enhance tissue 
and bone healing process so that Aloe vera can reduce 
inflammation caused by tooth extraction trauma, then can 
induce the healing process of the wound, and can finally 
stimulate osteoblast, as consequence, the growth of new 
bone will increase.4,15-22 This increasing process is also 
stimulated by Aloeride as polysaccharides derived from 
Aloe vera that has a high molecular weight and strong 
immunostimulator activity.23 

Thus, the group induced with Aloe vera and XCB had 
the highest expressions of FGF2, osteocalcin, osteoblasts, 
and fibroblasts started from day 7 to day 30 of examination. 
It indicates that Aloe vera and XCB in tooth extraction 
cases can accelerate wound healing process, can prevent 
inflammation, and then can induce new bone growth. It is 
because when Aloe vera is combined with XCB which is 
osteoinduction, it will accelerate healing process and new 
bone growth.2,8,9,10 

The increasing of osteoblast will not occur continuously, 
but at a certain time it will decrease. In a previous 
study, it is known that the growth of osteoblasts in tooth 
extraction socket filled with Aloe vera and XCB with 0.5% 
concentration of the active substance increased on day 30, 
but on histologic examination it is known that the growth 
of osteoblasts decreased on day 60.24 On day 12 after the 
tooth extraction, the formation of new bone occurred along 
the socket wall as well as in trabecular spaces around the 
tooth extraction area, and trabeculae bone was also woven at 
the periphery socket, osteoprogenitor cells, preosteoblasts, 
and osteoblasts. Periodontal ligament also moved into the 
middle of the socket, but not attached to the socket wall, and 
then got apoptosis. Collagen density, as a result, increased, 
and then was gradually replaced by bone. After this phase 

of bone resorption, freezing, granulation, and collagenase 
phases of the healing process can be bypassed, and then 
the socket of tooth extraction may regenerate.7 Finally, it 
may be concluded that the induction of the combination 
Aloe vera and 2% xenograft concelous bovine  into the 
socket of tooth extraction can increase the expressions of 
FGF2 and osteocalcin, so the growth of the alveolar bone 
can increase.

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