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Received: 26 November 2018    Revised: 2 February 2019    Accepted: 11 March 2019

DOI: 10.1111/jocd.12950

ORIGINAL CONTRIBUTION

Biostimulatory effects of polydioxanone, poly‐d, l lactic acid,

and polycaprolactone fillers in mouse model

Tae‐Rin Kwon PhD1  | Sung Won Han MS1,2 | In Kwon Yeo MD1,2 |

Jong Hwan Kim MS1,2 | Jae Min Kim MD1,2 | Ji‐Yeon Hong MD1,2  |
Byung‐Chul Lee MS  | Sung‐Eun Lee MS  | Ho Sang Moon PhD  | Han Jin Kwon PhD3 |
1 1 3

Beom Joon Kim MD1,2

1
Department of Dermatology, College of
Medicine, Chung‐Ang University, Seoul, Summary
Korea Background: Numerous fillers are increasingly used for augmentation of volume loss
2
Department of Medicine, Graduate
and relaxation of facial wrinkles. Collagen stimulators are the latest next‐generation
School, Chung‐Ang University, Seoul, Korea
3
Research Center, UltraV Co., Ltd., Seoul,
dermal fillers that can induce neocollagenesis. To investigate biophysical characteris‐
Korea tics, safety, and efficacy of newly developed polydioxanone (PDO) filler in compari‐

Correspondence
Beom Joon Kim, Department of
Dermatology, Chung‐Ang University
Hospital, Seoul, Republic of Korea
Email: beomjoon@unitel.co.kr
son with poly‐l lactic acid (PLLA) and polycaprolactone (PCL) fillers.
Methods: In vitro assay, morphology of particles, and rheological property of fillers
were measured. A total of 24 female hairless mice (SKH1‐Hrhr) were randomly divided
into three groups and injected with PDO, PLLA, or PCL fillers. Durability of fillers was
assessed at 0, 3 days, and 1, 4, 8, 12 weeks after injection using folliscope and PRIMOS.
To determine biocompatibility and neocollagenesis, histologic evaluation was per‐
formed at 1, 4, 8, and 12 weeks after injection. Efficacy was also evaluated based on
skin surface roughness changes using PRIMOS in a hairless mouse photoaging model.
Results: In the particle morphology test, PDO microspheres had an irregular surface and
were spherical and uniformly sized. PDO filler demonstrated similar neocollagenesis and
inflammatory response to other collagen stimulators. PDO filler showed better biodeg‐
radability than PLLA and PCL fillers. In the hairless mouse photoaging model, there was
a statistically significant decrease in skin surface roughness after PDO filler injection.
Conclusions: Our data suggest that newly developed collagen stimulating PDO filler
might be a safe and effective option for correction of volume loss and rejuvenation of
photoaging skin.

KEYWORDS
neocollagenesis, polycaprolactone, poly‐d, l lactic acid, polydioxanone

1 |  I NTRO D U C TI O N
Skin aging can be divided into intrinsic aging due to genetic factors
of individuals as they age and extrinsic aging caused by ultraviolet
Kwon and Han are contributed equally to this study.
rays, irritation by external substances, and other oxidizing stress.1
The main cause of extrinsic aging is repeated ultraviolet exposure.
Ultraviolet rays produce tumor necrosis factor‐α, tumor necrosis
factor‐γ, nitric oxide, and interleukins that inhibit collagen syn‐
thesis and increase the expression of enzymes that break down

J Cosmet Dermatol. 2019;1–7. © 2019 Wiley Periodicals, Inc. |  1

wileyonlinelibrary.com/journal/jocd  
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2       KWON et al.
collagen. Such characteristic changes exhibit clinical features such
as early formation of wrinkles, skin dryness, discoloration, and
telangiectasia.
Dermal fillers have been used to fill rhytides and augment soft
tissue volume loss due to disease or skin aging. Fillers can be di‐
vided into biodegradable (such as hyaluronic acid), biodegradable
collagen stimulator (such as calcium hydroxylapatite [CaHA], po‐
ly‐l lactic acid (PLLA), polycaprolactone [PCL]), and nonbiodegrad‐
able products (such as silicone and polymethylmethacrylate). 2,3
Biodegradable collagen stimulators are the latest next‐generation
dermal fillers that stimulate surrounding skin tissue to synthesize
new collagen.4
Polydioxanone (PDO) is a biodegradable with ester linked poly‐
mers. Ester is a bond with polar nature‐less stable‐hence more
reactive that can be broken down by hydrolysis, resulting in (2‐
5
Hydroxyethoxy)acetic acid PDO has been used mainly in laparot‐
omy sutures and esophageal stents. Recently, PDO is increasingly
used for wrinkle reduction with multiple applications of single PDO
filament for various areas of the face. Although previous reports
have demonstrated the efficacy of PDO filament, filler made of PDO
has not been developed for volume augmentation or anti‐wrinkle
purposes. The objective of the present study was to evaluate bio‐
physical characteristics, safety and neocollagenesis of newly devel‐
oped PDO filler in comparison with PLLA and PCL fillers. Moreover,
we investigated efficacy of PDO filler in a hairless mouse photoaging
model.
2 |  M ATE R I A L S A N D M E TH O DS
2.1 | Fillers
Polydioxanone (Trade name: ULTRACOL. Ultra V Co., Ltd., Seoul,
Korea), PLLA (Sculptra®, Galderma Laboratories, Nestlé SA,
®
Switzerland), and PCL (Ellansé‐M , Sinclair Pharma, Irvine, CA)‐
based fillers were tested. Newly manufactured PDO filler was
composed of PDO and sodium carboxymethyl‐cellulose (CMC).
Sculptra® containing PLLA was approved by the US Food and Drug
Administration (FDA) for treatment of HIV‐related facial lipoatrophy
in 2004 and for voluminization of aging face in 2009. Ellansé‐M®
made of PCL has not received approval by FDA yet.
2.2 | Particle morphology
Three different kinds of fillers were washed with 30% ethanol and
then desiccated in an oven for 48 hours. Filler particle morphology
was evaluated by field emission scanning electron microscopy (FE‐
SEM; LEO SUPRA 55, Carl Zeiss, Germany).
2.3 | Rheological measurements
Rheological characterization was performed twice using a Kinexus
pro Rheometer (Malvern Instruments Ltd., Worcestershire, UK). The
test was conducted by placing a sample between parallel plates,
vibrating and rotating parallel plates, and measuring resistant force
and loss of force in response to applied forces. Measurement condi‐
tion was configured to Geometry: PU20 S0163SS, Oscillation mode,
Shear strain: 1.5%, 0.1‐10 Hz at 1 Hz Frequency, Gap: 0.5 mm, and
Temperature: 25°C. Viscoelastic response was measured by a fre‐
quency sweep to determine G' (storage modulus; elastic modulus),
G'' (loss modulus; viscous modulus), G* (complex modulus), ŋ* (com‐
plex viscosity), and δ (phase angle).
2.4 | Preparation of animals
All animal experiments were approved by Institutional Animal Care
and Use Committee of Chung‐Ang University College of Medicine
(approval No. 16‐00069). Seven‐week‐old female hairless mice
(SKH1‐Hrhr; Saeron Bio Inc, Seoul, Korea) were raised in an inde‐
pendent space maintained at environmental conditions with con‐
trolled temperature (24 ± 2°C), relative humidity (50 ± 10%), and
light (12 hours light/12 hours darkness, without any ultraviolet emis‐
sion). There was a stabilization period of at least 1 week for acclima‐
tization prior to experiment.
2.5 | Durability, biocompatibility, and
neocollagenesis
A total of 24 mice were randomly divided into three groups: Group
1, PDO filler; Group 2, PLLA filler; and Group 3, PCL filler. Filler
(100 µl) was injected into the central dorsal skin of hairless mice
(1 cm from the tail) after anesthetization with zoletil and rompun.
To evaluate durability of filler, at 0, 3 days and 1, 4, 8, 12 weeks
after injection, folliscope (Lead M, Seoul, Korea) and PRIMOS (GFM,
Teltow, Germany) three‐dimensional (3D) images were obtained. For
biocompatibility test, histologic evaluation was performed at 1, 4, 8,
and 12 weeks after injection. Skin biopsies were fixed in 10% phos‐
phate‐buffered formaldehyde, embedded in paraffin, and processed
for routine histology. Sections were prepared at a thickness of 5 μm
and stained with hematoxylin‐eosin (H&E) and Masson trichrome for
collagen analysis.
2.6 | Efficacy in a hairless mouse photoaging model
Dorsal skins of hairless mice were irradiated with a UV‐emitting
system Bio‐Spectra (Vilber Lourmat, Marne La Vallée, France) three
times a week. The SKH‐1 hairless mice were irradiated using a UVA
or B‐emitting system from Biospectra (Vilber Lourmat, Marne La
Vallée, France), which has been designed for UV irradiation of test
animals and can be programed to deliver specific UV radiation dos‐
ages in energy (Figure 1). A total of 32 mice were randomly divided
into four groups. After anesthetization, Group 1, 2, 3, and 4 were
injected with 100 µl of phosphate‐buffered saline (PBS), PDO filler,
PLLA filler, and PCL filler, respectively, into UV‐induced wrinkle for‐
mation site on the back of mouse at 4 weeks after UV irradiation. A
second filler injection (100 µl) was conducted at 8 weeks after UV
irradiation. Assessments of cutaneous roughness were performed
KWON et al.
F I G U R E 1   Study design using a
hairless photoaging mouse model
F I G U R E 2   Particle morphology of
fillers
using PRIMOS at 0 day (before injection) and 5, 8, 12 weeks after
the first injection. Traditional H&E staining was performed to evalu‐
ate epidermal thickness. Special staining with Masson trichrome and
immunohistochemistry (IHC) analysis using anti‐collagen antibodies
to types I collagen was also performed. Schematic study design in a
hairless mouse photoaging model is shown in Figure 1.
2.7 | Statistical analysis
Statistical comparisons were performed using a one‐way analysis of
variance (ANOVA) followed by the Tukey honest significant differ‐
ences test. When P value was <0.05, it was considered statistically
significant.
3 | R E S U LT S
3.1 | In vitro test
Particles of PDO filler had an irregular surface, uniformly sized, and
spherical shaped. PLLA microspheres were very rough, nonuniform
sized, and flat with pointed shape. PCL microspheres had smooth
and uniformly sized spherical particles (Figure 2). Results of rheo‐
logical property of fillers are shown in Table 1. Measured G′, G″, and
η* values of PCL filler were the highest, followed by those of PDO
filler and PLLA filler. Based on Tan δ value, PDO and PCL fillers were
mainly elastic (tan δ < 1) while PLLA filler was mainly viscous (tan
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δ > 1). However, values of PLLA filler were not consistent because
PLLA particles did not dissolve in distilled water homogeneously.
They quickly separated into layers.
3.2 | In vivo animal test
Gross extracted fillers were observed inside and outside of the
mouse skin using a folliscope. On 3D images and topography using
PRIMOS, calculated volumes of PDO and PLLA fillers were gradually
decreased over time. At 4 weeks after injection, we failed to iden‐
tify injected PDO filler on the skin surface. The volume of PCL was
the maximum at one week after injection and decreased over time
(Figure 3, Figure 4).
Infiltration of inflammation‐related cells was observed at 1 week
after injection. It showed improvement from 4 weeks after injection
in all groups. However, there was no fibrosis or severe foreign body
reaction. PDO group showed similar neocollagenesis and inflamma‐
tory response to PLLA and PCL group. A time comparison of these
experimental groups showed that, in PDO filler group for which fol‐
liscope and PRIMOS measurements were impossible, there were a
small number of particles and CMC components at 8 and 12 weeks
after injection (Figure 5).
In a hairless mouse photoaging model, skin roughness was
evaluated using values of Ra and Rz. Results of roughness analysis
showed that roughness values were increased at baseline (0 day),
indicating successful induction of wrinkles due to photoaging. PDO
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4       KWON et al.

TA B L E 1   Rheological properties of fillers collagen stimulatory filler materials such as CaHA, PLLA, and PCL
have emerged.6,7
G′ (Pa) G″ (Pa) η* (Pa s) δ (°)
  (1Hz) (1Hz) (1Hz) (1Hz) Tan δ Polydioxanone is a colorless, crystalline, and absorbable polymer
that is particularly used in the preparation of surgical sutures. PDO
PDO 230.1 48.64 37.53 11.93 0.2114
has good flexibility and tensile strength with suitable biocompatibil‐
PLLA 0.23 0.25 0.05 46.89 1.0870
ity and minimal inflammation compared to other suture materials.8,9
PCL 552.4 447 113.4 38.98 0.8091
In recent years, PDO thread has been increasingly used for lifting
G′, elastic property; G″, viscous property; η*, overall viscoelastic and tightening neck and facial wrinkles, including malar, nasolabial,
property or hardness; δ, phase angle; Tan δ, the ratio between viscous
and marionette folds. However, patients frequently complain of mi‐
and elastic property (G''/G').
gration or total extrusion of the thread, skin dimpling, and rippling.9
On the other hand, filler has advantages of ease of handling, short
and PLLA groups exhibited reduction in both Ra and Rz values at 5, procedure time, and precise volume augmentation. Patients might
8, and 12 weeks after injection in comparison with baseline (0 day). feel that PDO filler is softer and more natural in appearance than
Their decreases were statistically significant compared to PBS group PDO thread.
(Figure 6). The immediate volumizing effect of PCL might have af‐ In the present study, microspheres of PDO filler showed relative
fected roughness evaluation and undermined the accuracy of its uniformly size and spherical shape shown to be suitable as dermal
analysis compared to PDO and PLLA. fillers. Fast degradation of PDO filler might be due to the viscous
Histopathologic study results showed that PBS group exhibited property of the PDO filler. In addition, the value G′ of PDO filler was
epidermal dyskeratosis or hyperkeratosis, with intensifying deep lower than that of PCL filler, indicating its highly deformable prop‐
wrinkles reaching the dermis. Filler injection groups demonstrated erty. PDO biodegradation occurs via slow hydrolysis to 2‐hydroxye‐
slow increases in epidermal thickness and gradual depositions of thoxyacetic monomer, most of which is excreted in urine while the
dermal collagen fibers compared to PBS group (Figure 7). On IHC rest is eliminated by digestion or exhaled as CO2.10 Different rheo‐
staining, the presence of type I collagen deposition around filler mi‐ logical characteristics of fillers might provide us scientific rationale
crospheres was observed in filler injection groups (Figure 8). to select appropriate products. However, rheological differences of
collagen stimulators are not correlated with ultimate clinical out‐
come. Their clinical effects do not depend on elasticity or viscosity
4 |  D I S CU S S I O N at the time of injection.11 Instead, they depend on neocollagenesis
over several weeks to months after injection.12
Cutaneous aging is a process caused by intrinsic and extrinsic factors, Biostimulation indicates that infiltration into the dermis of a ma‐
eventually leading to skin volume loss and wrinkles. Currently, filler terial can leads to production of new collagen and connective tis‐
injection for skin rejuvenation has become a standard therapeutic sue, resulting in improvement of elasticity, roughness, and turgor of
method as a noninvasive aesthetic treatment. Although hyaluronic skin.13 The present study shows that PDO filler can stimulate the
acid is the most commonly used filler material, new biodegradable synthesis of new collagen. Furthermore, at 12 weeks after injection,

F I G U R E 3   Skin surface changes outside hairless mice using images acquired with a folliscope (original magnification x15)
KWON et al. |
      5

PDO particles remained well in their original site, indicating its

long‐lasting effect. After injection of PDO filler, CMC gel carrier is
gradually absorbed by macrophages over several weeks. PDO micro‐
spheres stimulate neocollagenesis to replace the volume of resorbed
CMC gel carrier.3,14 After type I collagen is generated by biostimu‐
latory effect, it becomes progressively predominant over collagen
type III.13
The characteristic of PDO based filler is that it has biostimulatory
effect instead of an immediate direct filling effect. Although other
fillers show greater postprocedure improvement than collagen stim‐
ulatory fillers, such improvement will diminish over time. Collagen
stimulatory fillers initially show minor improvement in wrinkles.
However, over time, such fillers induce the synthesis or regenera‐
tion of collagen and other connective tissues to create spaces and
scaffolds for fibroblasts or vascular cells to enter, resulting in greater
effectiveness in later stages than other fillers. Therefore, PDO filler
is suitable for patients desiring for gradual improvement. It also
might have advantages over hyaluronic acid‐ or CaHA‐ based fillers
because of its stability and duration of results. 2 PDO can be used for

F I G U R E 5   Histologic response to different fillers after injection

F I G U R E 4   Calculated volume changes of implanted filler using based on (A) H&E and (B) Masson trichrome staining (red arrows:
PRIMOS 3D images collagen fibers, original magnification x400)

F I G U R E 6   Skin surface roughness in photoaging model using values of (A) Ra (average value of all peaks and valleys in the direction of
the average line at sampling length) and (B) Rz (10‐point average roughness, average value Ra of five highest peaks and lowest valleys) (*:
P < 0.05; **: P < 0.01; ***: P < 0.001)
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6       KWON et al.
volume augmentation in the nasolabial fold, chin, jawline, and buccal
hollows. It should not be used in the eyelid or lip owing to the lack of
subcutaneous fat in such places, similar to other biostimulatory fill‐
ers.15,16 PDO filler should be injected into the dermal‐subcutaneous
junction or preperiosteal tissues with care being taken to avoid high‐
pressure gradients. Fanning, linear threading, or crosshatching grid
techniques are recommended. Formation of papules or nodules may
be technical errors, such as superficial or bolus injections. Proper
injection technique and understanding of the anatomy are important
to successful results with minimal complications.
F I G U R E 7   Changes of (A) epidermal
thickness (original magnification x100)
and (B) dermal collagen fibers (original
magnification x100, x200)
To the best of our knowledge, this is the first study that demon‐
strates in vitro and in vivo safety, and efficacy of PDO based der‐
mal filler. Moreover, there is a paucity of data regarding the efficacy
of collagen stimulatory filler in a hairless mouse photoaging model.
Our study indicates that newly developed PDO filler showed sim‐
ilar neocollagenesis and inflammatory response to PLLA and PCL
filler. PDO filler has better biodegradability compared to other fill‐
ers. There was a significant decrease in skin surface roughness after
PDO filler injection. Thus, PDO filler might be an attractive option
for correction of volume loss and rejuvenation of photoaging skin.
KWON et al.
F I G U R E 8   Type I collagen deposition
around filler microspheres based on IHC
staining (original magnification x200)
Further long‐term human study is needed to determine the safety
and efficacy of PDO filler.
AC K N OW L E D G M E N T S
We thank Ultra V Co., Ltd. for providing the PDO fillers.
CONFLICT OF INTEREST
Authors have no potential conflicts of interest to disclose.
ORCID
Tae‐Rin Kwon  https://orcid.org/0000-0002-9892-7714
Ji‐Yeon Hong  https://orcid.org/0000-0002-5632-8449
Beom Joon Kim  https://orcid.org/0000-0003-2320-7621
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How to cite this article: Kwon T‐R, Han SW, Yeo IK, et al.
Biostimulatory effects of polydioxanone, poly‐d, l lactic acid,
and polycaprolactone fillers in mouse model. J Cosmet
Dermatol. 2019;00:1–7. https://doi.org/10.1111/jocd.12950