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The Effect of Hyaluronan on Bone and

Soft Tissue and Immune Response in
Wound Healing
Per-Erik Engstrm,* Xie-Qi Shi, Gunilla Tronje, Anders Larsson, Ulf Welander, Lars Frithiof, and
Gunilla Norhagen Engstrom*

Background: The aims of this study were to investigate the anti-inflammatory effect and the effect on
bone regeneration of hyaluronan in surgical and non-surgical groups.
Methods: In each of 15 individuals, 2 teeth with defects of similar character and magnitude in the upper
or lower jaw were chosen. There were at least 2 teeth between the test and the control sites. In the surgical group, a bioabsorbable membrane was used for both test and control sites, and hyaluronan was placed
in the intrabony pocket of the test site. In the non-surgical group, the periodontal pockets were scaled and
hyaluronan was administered 3 times with an interval of 1 week in the test pockets. Alveolar bone height
and bone healing patterns were analyzed using digital intraoral radiographs. Measurements of bone height
were performed in the original digital black-and-white radiographs to obtain quantitative data on bone gain
or loss. Bone healing patterns were studied with color-coded radiographs, using specially designed software
in a personal computer with subsequent combinations of radiographs. Gingival crevicular fluid immunoglobulin (Ig)G, C3, and prostaglandin E2 (PGE2) responses; periodontal probing depth; bleeding on probing; and
the presence of plaque were studied to evaluate the anti-inflammatory effect. Data were obtained at baseline before treatment, and at 2 weeks, and 1, 3, 6, and 12 months after treatment.
Results: For the surgical treatments, bone height was increased in the test group treated with hyaluronan (mean value 2.2%, corresponding to an average increase of approximately 0.5 mm) and reduced in
the control group (mean value 1.8%, corresponding to an average decrease of approximately 0.4 mm)
(P <0.05) after 12 months. For the non-surgical treatments, bone height was reduced by a mean value of
1.1% (corresponding to an average decrease of approximately 0.25 mm) in the test group treated with
hyaluronan and 3.3% (corresponding to an average decrease of approximately 0.75 mm) in the control
group after 12 months (N.S.). According to the digital color-coded radiographs, the test sites in the surgical and non-surgical groups showed apposition of bone minerals. Immune responses showed no differences
during the 12 months studied for the surgical and non-surgical sites. Mean periodontal probing depths were
reduced between 2.5 mm and 4.1 mm in the surgical and non-surgical groups.
Conclusions: The observed difference in bone height between test and control sites in the surgical group
after 12 months was less than 1 mm, which was only detectable on radiographs. No statistical difference
was found on radiographs in the non-surgical group, where a decrease in bone height was found for both
groups after scaling. Probing depth reduction after the surgical treatment, as well as after scaling and root
planing, was as expected. Hyaluronan in contact with bone and soft tissues had no influence on the immune
system in this study. Further studies are needed to determine the extent to which hyaluronan can lead to
clinically significant healing of periodontal lesions. J Periodontol 2001;72:1192-1200.
KEY WORDS
Follow-up studies; hyaluronan/therapeutic use; immune response; periodontitis/drug therapy;
radiography, dental, digital; wound healing.
* Division of Clinical Immunology, Department of Microbiology, Pathology and Immunology, Karolinska Institutet, Huddinge University Hospital, Stockholm,
Sweden.
Department of Periodontology, Institute of Odontology, Karolinska Institutet.
Department of Oral Radiology, Institute of Odontology, Karolinska Institutet.
Division of Clinical Chemistry, Department of Medical Sciences, University Hospital, Uppsala, Sweden.

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yaluronan (HA) is a linear polysaccharide with a

high molecular weight. It is found in all extracellular matrices and has the same structure in all
species. It has a unique influence on cell differentiation, motility, and cell adherence and has been suggested to have anti-inflammatory effects.1-3 If HA is
administered during surgery, scar formation is prevented.4-6 It influences and enhances tissue regeneration
through its ability to retain large amounts of water.7,8
Locally applied HA does not result in any phagocytosis or immunological reactions.9 Important indications
for the use of HA in humans are found in connection
with eye surgery, regeneration of the tympanic membrane, and local treatment of osteoarthrosis.10
Hyaluronan, heparin sulphate, dermatan sulphate,
and chondroitin-4-sulphate are 4 glycosaminoglycans
that have been identified in human gingival tissue.11
These compounds may contribute to the attachment
of the epithelium to the surface of the tooth under normal conditions, as well as after surgical treatment. The
glycosaminoglycans play a role in the defense mechanisms of the tissues adjacent to teeth. Periodontal tissue was chosen as an experimental model to study
the effect of HA on wound healing.
Treatment of periodontal diseases usually starts with
providing the patient with information about the disease, together with instructions on obtaining proper
oral hygiene; scaling, debridement, and root planing of
the teeth are then performed. These factors are
involved in the initial cause-related anti-inflammatory
and antimicrobial therapy. When necessary, surgical
procedures are performed after the initial phase. Surgical techniques make it possible to remove the
remaining infectious material from the teeth and to
eliminate tissues contaminated with bacteria and bacterial products, such as lipopolysaccharides and proteins, e.g., endotoxins and leukotoxin. Generally, healing results in a long epithelial attachment and probing
depth reduction.
Guided tissue regeneration (GTR) has been used
since the 1980s in the treatment of intrabony pockets
to create new bone, new cementum, and a new connective tissue attachment.12-14
The aims of this study were to investigate the antiinflammatory and bone regenerative effects of hyaluronan. The effects of hyaluronan are described. One
group consisted of patients receiving hygiene instruction and scaling and root planing, and the other receiving GTR surgery with a bioabsorbable membrane. Both
surgical and non-surgical test sites received hyaluronan during the wound healing period.
MATERIALS AND METHODS
Clinical Measurements
Fifteen subjects with a diagnosis of chronic periodontitis were included in the study. Two teeth were cho-

Engstrm, Shi, Tronje, et al.

sen in each patient, both in the same jaw. The selected

teeth displayed defects of similar character and magnitude, and there were at least 2 teeth between the
test and the control tooth. The test and control sites
were randomly selected. The exclusion criteria
included: having received antibiotics 1 month prior to
treatment or metronidazole (topical or oral) 6 months
prior to treatment; receiving treatment for allergy;
hypersensitivity to amalgam or other metals; previous
radiation treatment (head or neck); alcohol or drug
abuse; or other serious disease.
Periodontal pockets were measured with a manual
probeL marked in millimeter increments. Bleeding on
probing was recorded and the presence of plaque was
registered mesially, buccally, distally, and lingually.
Data were obtained at baseline before treatment,
and at 2 weeks, and 1, 3, 6, and 12 months after treatment.
This study was approved by the Ethical Committee
at Huddinge University Hospital and by the Medical
Products Agency, Uppsala, Sweden.
Study Drug
Hyaluronan was available in syringes containing 0.85
ml. The drug is composed of 14 mg sodium
hyaluronate (from Roosters comb); 8.5 mg sodium
chloride; 0.28 mg disodium hydrogen phosphate dihydrate; 0.04 mg sodium dihydrogen phosphate hydrate;
and water for injection per ml.
Surgical Group
Six individuals underwent surgery (4 males and 2
females; age range 30 and 69 years; mean 49). They
were selected from among patients with intrabony
pockets and pockets 6 mm deep.
After administration of local anesthesia, sulcular
incisions were made at the buccal and lingual sides and
full-thickness flaps were elevated. Debridement and
root preparation were carried out with hand instruments. In both test and control sites, mesial and/or
distal bone pockets, initially >3 mm deep and 3 mm
wide, were covered with a matrix barrier based on
bioabsorbable polylactic acid mixed with citric acid
ester,# according to the GTR technique.15 In addition,
hyaluronan was locally administered below the membrane on the test teeth. After surgery, patients were
instructed to rinse their mouths twice daily with 10 ml
of 0.2% chlorhexidine for 6 weeks.
Non-Surgical Group
Nine patients (4 males and 5 females; age range 29
to 61 years; mean 48) participated. Pockets were 5
mm deep and exhibited inflammation and radiographic
signs of bone loss. Treatment included scaling and
L UNC-15, Hu-Friedy Manufacturing Inc., Chicago, IL.
Healon GV, Pharmacia & Upjohn, Uppsala, Sweden.
# Guidor AB, Huddinge, Sweden.

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Volume 72 Number 9

root planing of both test and control teeth.

Hyaluronan was administered in the test
pockets 3 times at 1-week intervals.
Radiographic Examination
Digital intraoral radiographs** were taken,
with an x-ray unit operating at 70 kVp and
7 mA.
The examination technique was standardized to obtain radiographs as similar as
possible. Bite blocks were used to hold the
sensor of the digital system. Impressions
were made at the first examination and saved
for follow-up control examinations. Radiographs were exposed immediately before
treatment and at 1, 3, 6, and 12 months after
treatment. All radiographs were saved and
transferred to a station where sequential
radiographs were registered so that the anatomical structures coincided, as previously
Figure 1.
described.16,17
Examples of color-coded radiographs combined from baseline and the 2-month
examination. A) Test site, where bone gain is observed as green. B) Control site, where
Using specially designed personal combone loss is observed as magenta.The small magenta-colored areas on the distal side
puter software, subsequent combinations of
of 35 and the mesial side of 36 indicate removed calculus.The green lines along the
radiographs were color-coded. The colorteeth and the magenta shading of 36 are due to a slightly imperfect registering of the
coded images were made by combining 2
2 combined radiographs.
radiographs; i.e., first and second; second
and third, etc. The first radiograph of each
Table 1.
pair was displayed twice. Then, the radiograph was
transformed into monochromatic images in red and
Scale Used to Evaluate Color-Coded
blue. The second radiograph was transformed into
Radiographs
green. In the color-coded image, the 3 monochromatic
radiographs were combined. Changes in bone struc3
Marked bone gain
ture over time were either exhibited in green or
2
Moderate bone gain
magenta, where green indicated bone gain and
magenta bone loss (Fig. 1). The presence of both
1
Bone gain dominates over loss
green and magenta indicated that both bone gain and
0
No change
loss were simultaneously present.
Evaluation of the series of color-coded radiographs
1
Bone loss dominates over gain
was performed on the successive combinations. Three
2
Moderate bone loss
radiologists performed the evaluation together and
arrived at a consensus. The evaluators had no infor3
Marked bone loss
mation regarding the treatment performed or which
side was the test or control site. The scores shown in
Statistical analyses were performed employing paired
Table 1 were applied. The non-parametric data thus
t tests.
obtained subjectively from the radiographs were used
to plot graphs indicating the successive development
Gingival Crevicular Fluid
of each case.
Gingival crevicular fluid (GCF) was collected, using
Measurements were performed in the original
filter strips, and measured. The filter strips were kept
black-and-white radiographs to obtain quantitative
in 50 l phosphate buffered saline (PBS) with 0.05%
data on bone gain or loss. From the apices, 2 disTween 20L L containing 0.02% NaN3. The samples were
tances were registered, one to the cemento-enamel
frozen until analyzed.
junction and one to the deepest point of the periodontal pocket. The ratio between these distances
** Regam Medical Systems Int. AB, Sundsvall, Sweden.
was calculated. For subsequent examinations, the dif Oralix DC, Gendex Corp., Monza, Italy.
Sun Microsystems, Inc., Palo Alto, CA.
ferences between the ratios were calculated and con Periotron 6000, Pro-Flow, Inc., Amityville, NY.
verted into percentages indicating bone gain or loss.
LL Polysorbatum, Apoteksbolaget, Stockholm, Sweden.
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J Periodontol September 2001

Human IgG Enzyme-Linked Immunosorbent Assay

(ELISA)
Flat-bottomed microelisa plates were coated with
100 l of 5 g affinity purified chicken anti-human
IgG/ml## in PBS (0.02 M NaH2PO4, 0.15 M NaCl, pH
7.2) overnight at 4C. The plates were then blocked
with 150 l of 100 g unspecific chicken IgY/ml##
overnight at 4C. The plates were washed 4 times with
PBS-Tween (0.02 M NaH2PO4, 0.15 M NaCl, 0.05%
Tween 20, pH 7.2) in a washing machine.*** One hundred (100) l samples and normal human serum at
various dilutions in PBS-Tween were added to the plate.
The plates were incubated for 2 hours at room temperature on an orbital shaker and the plates were then
washed 4 times with PBS-Tween. One hundred (100)
l of chicken anti-human IgG-HRP diluted 11,000 in
PBS-Tween were added; the plates were incubated for
2 hours at room temperature on an orbital shaker and
then washed 4 times with PBS-Tween. One hundred
(100) l of K-Blue substrate was added and the plates
were incubated for 5 minutes at room temperature. The
reaction was stopped with 50 l of 1.8 M H2SO4 and
the plates were read. The IgG concentrations in the
samples were calculated from the standard curve.
Human C3 ELISA
Flat-bottomed microelisa plates were coated with 100
l of 4 g affinity purified chicken anti-human IgG/ml##
in PBS overnight at 4C. The plates were then blocked
with 150 l of 100 g unspecific chicken IgY/ml##
overnight at 4C. The plates were washed 4 times with
PBS-Tween in a washing machine. One hundred (100)
l samples and purified human C3 at various dilutions
in PBS-Tween were added to the plate. The plates were
incubated for 2 hours at room temperature on an orbital
shaker, and the plates were then washed 4 times with
PBS-Tween. One hundred (100) l of chicken antihuman C3-biotin## diluted 1:2,000 in PBS-Tween was
added; the plates were incubated for 1 hour at room
temperature on an orbital shaker and then washed 4
times with PBS-Tween. One hundred (100) l of streptavidin-horseradish peroxidase diluted 1:2,000 in
PBS-Tween was added; the plates were incubated for
1 hour at room temperature on an orbital shaker and
then washed 4 times with PBS-Tween. One hundred
(100) l of K-Blue substrate was added and the plates
were incubated for 5 minutes at room temperature. The
reaction was stopped with 50 l of 1.8 M H2SO4 and
the plates were read. The C3 concentrations in the
samples were calculated from the standard curve.
Prostaglandin E2 ELISA
Prostaglandin E2 was analyzed via ELISA.LLL
Microbiological Examination
Plaque samples were collected with sterile absorbent
paper points at the same time body fluids were col-

Table 2.

Mean Values of Differences in Percentage

of Bone Height Between Baseline and 1, 3,
6, and 12 Months
Month
Group

12

Surgical treatment (n = 6)
Test sites
1.1%
Control sites
0.1%

3.2%
0.1%

1.6%
0.1%

2.2%
1.8%*

Non-surgical treatment (n = 9)
Test sites
0.9%
Control sites
0.5%

0.3%
1.1%

1.7%
2.9%

1.1%
3.3%

* P <0.05, comparison between test and control sites after 12 months; all
other differences were not significant.

lected. The paper points were transported in VMG III

medium.18
The presence of Actinobacillus actinomycetemcomitans was determined on tryptic soy agar containing horse serum and supplemented with 75 g/ml
bacitracin and 5 g/ml vancomycin.19 The plates were
incubated at 37C in an atmosphere of 5% to 10% CO2
for 3 or 4 days. A. actinomycetemcomitans was identified by its morphology on selective medium and positive catalase reaction. The detection limit was 10 bacteria/ml.
The presence of black-pigmented species was determined on Brucella-blood agar with 0.1 ml menadione
and 5% horse serum cultivated for 1 week. The total
number of colony-forming units was counted and the
presence of Prevotella species and Porphyromonas gingivalis was detected by long-wave UV-light fluorescence. The detection limit was 10 bacteria/ml.
RESULTS
Radiographic Evaluation
After 12 months, the bone height had increased at surgical test sites treated with hyaluronan (mean value
2.2%, corresponding to an average increase of approximately 0.5 mm) and decreased at surgical control
sites (mean value 1.8%, corresponding to an average
decrease of approximately 0.4 mm) (P <0.05) (Table
2). At non-surgical test sites treated with hyaluronan,
bone height had decreased after 12 months by a mean
value of 1.1% (corresponding to an average decrease
of approximately 0.25 mm); at control sites, the corresponding value was 3.3% (corresponding to an aver F96 Polysorp, NUNC, Roskilde, Denmark.
## Immunsystem, Uppsala, Sweden.
*** Denley Instruments Ltd., Billingshurst, UK.
Neogen, Lexington, KY.
SpectraMax, Molecular Devices, Sunnyvale, CA.
PRN1231, Amersham Pharmacia Biotech, Uppsala, Sweden.
LLL Cayman Chemical Corp., Ann Arbor, MI.

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age decrease of approximately 0.75 mm). The data

from the 1-, 3-, 6-, and 12-month evaluations are
shown in Table 2.
Non-parametric data subjectively obtained from radiographs were used to plot graphs indicating the successive development of each case. Bone healing patterns in test and control groups were studied using
digital color-coded radiographs (Fig. 2). Surgical test
and control sites showed more pronounced negative
values during the first 6 months as compared to nonsurgical test and control sites.
Clinical Evaluation
Surgical group. After 6 months, mean probing depths
were reduced from 7.8 mm to 3.7 mm at test sites
treated with hyaluronan, and from 7.3 mm to 4.3 mm
at control sites (Table 3). Between 6 and 12 months,
mean probing depths were basically stable (Table 3).
Bleeding on probing and the presence of plaque were
found to be 25% in connection with both test and

Volume 72 Number 9

control teeth on mesial, buccal, distal, and lingual

sides as shown in Tables 4 and 5.
Non-surgical group. After 6 months, mean probing
depths were reduced from 6.4 mm to 3.9 mm at
test sites treated with hyaluronan, and from 6.8 mm
to 4.2 mm at control sites (Table 3). From baseline
to 12 months, the mean probing depths at test and
control sites were reduced by 2.5 mm and 3.1
mm, respectively (Table 3).
Bleeding on probing and the presence of plaque
were found to be 25% for both test and control teeth
on mesial, buccal, distal, and lingual sides, as shown
in Tables 4 and 5.
IgG in Gingival Crevicular Fluid
During the 12-month period, the mean IgG levels in
GCF varied between 0.9 and 3.0 g/l at test sites and
between 1.3 and 3.3 g/l at control sites in the surgical group (Table 6). Corresponding values for the
non-surgical group varied between 1.5 and 2.8 g/l

Figure 2.
Non-parametric plots of the individual development of all cases. On the y-axis, the subjectively evaluated change in successive pairs of radiographs is
plotted.The values indicate the sum of the scores according to Table 1. A) Test sites of the surgical group, initial bone loss occurs followed by gain. B)
Control sites of the surgical group, bone loss predominates. C) Test sites of the non-surgical group, bone gain predominates. D) Control sites of the nonsurgical group; the range covers both bone gain and loss. No trend can be detected.

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Table 3.

Table 5.

Mean ( SD) Probing Depth (in mm) at

Baseline and 6 and 12 Months

Number of Teeth With Plaque Index 25%

at Baseline and at 0.5, 1, 3, 6, and 12
Months

Month
Group

Baseline

Month

12
Group

Surgical treatment (n = 6)
Test sites
7.8 1.1
Control sites
7.3 0.9

3.7 0.9
4.3 1.2

3.8 0.7
4.3 1.4

Non-surgical treatment (n = 9)
Test sites
6.4 1.3
Control sites
6.8 1.5

3.9 1.2
4.2 1.4

3.9 1.4
3.7 1.5

Table 4.

Number of Teeth With Bleeding on Probing

25% at Baseline and at 0.5, 1, 3, 6, and
12 Months
Month
Group

Baseline

0.5

12

Surgical treatment (n = 6)
Test sites
0
Control sites
1

1
3

1
2

3
3

4
3

5
4

Non-surgical treatment (n = 9)
Test sites
3
Control sites
4

5
4

7
6

3
4

7
6

5
6

at test sites and between 2.1 and 2.7 g/l at control

sites (Table 6).
Complement Factor 3 in Gingival Crevicular Fluid
During the 12 months studied, the mean C3 levels in
gingival crevicular fluid varied between 4.2 and 44.8
ng/l at test sites and between 7.1 and 42.4 ng/l at
control sites of the surgical group (Table 7). Corresponding values for the non-surgical group varied
between 16.4 and 33.6 ng/l at test sites, and between
12.1 and 69.6 ng/l at control sites (Table 7).
Prostaglandin E2 in Gingival Crevicular Fluid
During the 12 months studied, the mean PGE2 levels
in GCF varied between 2.0 and 12.0 pg/l at test sites,
and between 4.6 and 8.1 pg/l at control sites of the
surgical group (Table 8).
Microflora
The occurrences of Prevotella species, A. actinomycetemcomitans, and P. gingivalis are shown in Table
9. In the surgical group, 2 out of 6 individuals harbored A. actinomycetemcomitans in periodontal pockets, and in the non-surgical group, A. actinomycetem-

Baseline

0.5

12

Surgical treatment (n = 6)
Test sites
5
Control sites
6

4
5

5
5

5
5

5
5

6
6

Non-surgical treatment (n = 9)
Test sites
5
Control sites
5

8
8

6
6

8
8

7
6

8
8

comitans was present in periodontal pockets in 1 out

of 9 individuals (Table 9) during the 12 months studied. P. gingivalis appeared in one individual at baseline in the surgical group and was not detected later.
In the non-surgical group, 1 individual harbored P. gingivalis 3 months after baseline, which was not detected
later. Prevotella species were detected in several
patients.
DISCUSSION
This randomized study was performed to investigate
the role of hyaluronan in wound healing. For this purpose, radiological, immunological, microbiological, and
clinical evaluations of surgical and non-surgical periodontal treatments were performed.
All forms of periodontal disease are considered
inflammatory and triggered by microorganisms on the
tooth surfaces and in periodontal pockets.20 Together
with clinical findings, periodontal lesions are diagnosed,
using radiography, which demonstrates the possible
presence of horizontal and/or vertical bone destruction.
One problem in treating advanced periodontal disease
is deep gingival pockets and horizontal and/or vertical
bony defects. In the Stockholm area of Sweden, about
13% of the population between 31 and 40 years of age
are considered to have periodontitis, with pathological
probing depths that need treatment.21
Conventional periodontal treatment consists of providing information to the patient about periodontal disease, instructions concerning oral hygiene, and professional scaling. Should there be no improvement,
surgery is indicated to gain access to the pocket area
for proper debridement; to eliminate the pathological
tissue; and to reconstruct bone, cementum, periodontal ligament, and gingiva. Another strategy is to use
antibiotics either during the scaling or surgery phase.
Combinations of different antibiotics, such as metronidazole and tetracycline, may be administered generally or locally to reduce the specific anaerobic
microflora (P. gingivalis, Prevotella intermedia) or
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ament, cementum, and alveolar bone

regeneration is increased. The memMean ( SD) Gingival Crevicular Fluid Levels of IgG in
brane also prevents other structures,
g/l at Baseline and at 0.5, 1, 3, 6, and 12 Months
such as epithelium, from proliferating
into the wound. The original method has
Month
been modified and is used in the surgical treatment of bone pockets.14 In some
Group
Baseline
0.5
1
3
6
12
cases, regeneration of supporting tissue
Surgical treatment (n = 6)
occurs. In this study, we used this surgiTest sites
1.8 2.0 2.8 2.4* 1.6 3.1* 3.0 2.3* 0.9 0.6 3.0 3.4
cal technique with a bioabsorbable memControl sites 2.3 1.9 3.3 4.2 1.6 1.8 1.3 0.9* 3.3 2.8 1.9 2.4
brane to cover the administered hyaluronan.
Non-surgical treatment (n = 9)

Wound healing is complex, involving

Test sites
2.8 2.1 2.0 1.3 1.8 2.3 2.8 2.7
1.5 1.5 1.7 1.7
Control sites 2.7 2.2 2.4 1.8 2.7 2.0 2.4 1.9
2.1 2.2 2.3 2.3
interference by microorganisms, inflammatory components, and mechanical
* N = 5.
disturbances. The oral cavity is an excel N = 8.
lent model for studying
Table 7.
wound healing, because of
the ease of collecting samMean ( SD) Gingival Crevicular Fluid Levels of C3 in ng/l at
ples and performing radiBaseline and at 0.5, 1, 3, 6, and 12 Months
ographic and clinical examinations.
Month
Hyaluronan in contact with
bone and soft tissues did not
Group
Baseline
0.5
1
3
6
12
influence the immune reSurgical treatment (n = 6)
sponse during the 12 months
Test sites
10.5 9.7
9.3 5.5* 18.2 22.0 17.4 12.4
4.2 3.2 44.8 72.1*
studied. The anti-inflammaControl sites 20.0 19.0 25.1 28.1
7.1 5.0
29.2 34.6
8.8 6.7 42.4 44.2*
tory effect of HA as proposed
Non-surgical treatment (n = 8)
previously1-3 could not be

Test sites
33.6 68.4 16.4 13.8 18.4 25.7 29.5 67.3 25.8 31.8 29.9 27.6
verified in this study in relaControl sites 53.2 78.3 30.6 39.0 69.6 63.7 22.7 38.1 12.1 6.2 19.1 11.6
tion to the IgG, C3 and PGE2
responses.
* N = 4.
N = 7.
Immunoglobulin G levels
N = 6.
in GCF from individuals with
periodontitis were comparaTable 8.
ble to those in earlier reports.23 Prostaglandin
Mean ( SD) Gingival Crevicular Fluid Levels of PGE2
E2 was in the same concentration range as
in pg/l at Baseline and at 0.5, 1, 3, 6, and 12 Months in a study concerning elevated PGE2 levels
in GCF from individuals with Downs synin Surgically Treated Group
drome.24 During the study period, there was
no difference in the acute-phase protein C3
Month
Sites
levels in surgical and non-surgical test or
(n = 6)
Baseline
0.5
1
3
6
12
control groups. This may be due to the fact
that after 2 weeks or more, the inflammatory
Test
4.6 2.5 9.6 3.5* 12.0 14.9 9.1 6.9 2.0 2.5 5.7 6.0*
response in the samples had receded.
Control 8.1 8.0 6.3 5.2
6.6 4.9 4.6 2.2 5.3 6.3 5.1 3.0*
In all groups, the microflora (Prevotella
species,
P. gingivalis, or A. actinomycetem* N = 4.
comitans) showed a stable microecology.
facultative anaerobic microorganisms (A. actinoBone height measurements in original black-andmycetemcomitans) which are often found in cases of
white digital radiographs of the surgical patients
refractory periodontitis.22
demonstrated a decrease at the control examination
Regeneration of the periodontal supporting tissue is
performed 1 month after surgery. Color-coded radipossible under certain conditions through GTR proceographs showed a decrease in mineral contents condures, where the natural ability to regenerate is
firming this finding. No quantitative decrease could be
mechanically guided by a membrane.12,13 The memregistered at control sites. A decrease in mineral conbrane is placed so that the possibility of periodontal ligtents 1 month after periodontal flap surgery has been
Table 6.

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Engstrm, Shi, Tronje, et al.

J Periodontol September 2001

Table 9.

Microorganisms in Test and Control Sites

at Baseline and at 0.5, 1, 3, 6, and 12
Months
Month
Group
Surgical treatment (n = 6)
Test sites
A. actinomycetemcomitans
P. gingivalis
Prevotella species
Control sites
A. actinomycetemcomitans
P. gingivalis
Prevotella species
Non-surgical treatment (n = 9)
Test sites
A. actinomycetemcomitans
P. gingivalis
Prevotella species
Control sites (n = 9)
A. actinomycetemcomitans
P. gingivalis
Prevotella species

Baseline

2
1
4

0.5

12

1
1
3

1
1
3

1
1
4

2
1
3

demonstrated using 125I-radiography, followed by an

increase during the next 6 months.25 According to the
color-coded images, the test sites in this study demonstrated a similar pattern. In the Bergstrm and Henrikson study,25 bone was in contact with HA through
the gingiva, which normally contains HA. In our study,
a drug containing HA was administered to the bone
covered with a bioabsorbable membrane. In the control sites in our study, the bone was in contact only with
a bioabsorbable membrane and a different healing pattern may have occurred. Hyaluronan creates a local
environment with a water binding effect, which may
give an opportunity for osteoblasts to produce a matrix
for wound healing.
The increased bone height in the surgical group
recorded at the examination 12 months after surgery
may be explained by the osteoinductive properties of
HA previously reported.26 It has been proposed that an
accelerated wound healing in bone matrix will occur
due to stimulation of angiogenesis by HA.27 Another
important effect of HA and the HA receptor is the locomotion of cells, as shown for transforming growth factor- stimulated fibrosarcoma cells.28 Hyaluronan is
also involved in the process of dental implant osseointegration.29
The differences in bone height between test and
control sites in the surgical group 12 months after
treatment was less than 1 mm, which was detectable

only on radiographs. No statistical difference was found

clinically or on radiographs in the non-surgical group,
where a decrease in bone height was found for both
test and control groups after scaling. Probing depth
reduction after the surgical treatment,30 as well as after
scaling and root planing, was as expected. Hyaluronan
in contact with bone and soft tissues did not influence
the immune system in this study.
ACKNOWLEDGMENTS
The authors are grateful for the microbiological analysis performed by the Division of Clinical and Oral Bacteriology, Department of Microbiology, Pathology and
Immunology, Karolinska Institutet, Huddinge University Hospital, Stockholm, Sweden. We are also grateful for the contributions of Pr-Erik Wejker, Srgrden,
Morgongva, Sweden. This work was supported by
funds from Karolinska Institutet; Pharmacia & Upjohn,
Uppsala; and Guidor, Huddinge, Sweden.
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Send reprint requests to: Dr. Per-Erik Engstrm, Division of
Clinical Immunology, F 79, Department of Microbiology,
Pathology and Immunology, Karolinska Institutet, Huddinge
University Hospital, SE-141 86 Stockholm, Sweden.
Accepted for publication March 8, 2001.