Minimally invasive skin rejuvenation with Erbium: YAG laser ... - CMS

Lasers in Surgery and Medicine
Minimally Invasive Skin Rejuvenation With Erbium:
YAG Laser Used in Thermal Mode
Karin Kunzi-Rapp, MD, 1,2 * Christine C. Dierickx, MD, 3,4 * Bernard Cambier, MD, 5 and
Michael Drosner, MD, PhD 6,7
1
Institute for Laser Technologies in Medicine and Metrology, University of Ulm, Germany
2
Department of Dermatology and Allergology, University of Ulm, Germany
3
Laser Skin Center, Boom, Belgium
4
Wellman Laboratories of Photomedicine, Havard Medical School, Boston
5
Laser Clinic, Gent, Belgium
6
Cutaris Zentrum für Haut, Venen und Lasermedizin, Munich, Germany
7
Department of Dermatology and Allergology, Technical University of Munich, Germany
Background and Objectives: To evaluate the efficacy
and safety of a thermal mode Erbium:YAG laser several invivo
morphological as well as clinical changes were
monitored in a multi-center investigation.
Study Design/Materials and Methods: An Erbium:YAG
laser was used at a thermal mode with sub-ablative
fluences of 2.1 and 3.1 J/cm 2 with parallel air cooling to
treat either periorbital, perioral rhytides or patients with
post-traumatic or acne scars. Two treatments were applied
2 months apart, with follow-up at 1, 3, 6, and 12 months
post-treatment. Photographs were taken before and at each
follow-up visit and evaluated by three blinded independent
reviewers. Histology and immunohistochemistry for procollagen
expression were investigated. Optical coherence
tomography (OCT) was performed before, and at 4, 14, and
28 days after single pass treatment with Erbium:YAG
thermal pulses.
Results: The improvement of rhytides at 1–3 months
follow-up was graded as excellent in 19%, good in 19%, fair
in 31%, and no improvement in 31%. At the 6- to 12-month
follow-up, the improvement was excellent in 40%, good in
40%, fair in 20%, and no improvement in 0%. The
improvement of scars at 3–6 months follow-up was graded
as excellent in 50%, good in 25%, fair in 25%, and no
improvement in 0%. Intra- and post-operative discomfort
was described as mild by the patients. OCT, histological
sections and immunohistochemistry demonstrated production
of new collagen bundles.
Conclusions: Thermal Erbium:YAG pulses can induce
collagen neogenesis, as proved by temperature elevation
and morphological changes in the upper dermis. This leads
clinically to visible and long lasting reduction of wrinkles
and scars after applying multiple passes with minimal sideeffects.
Lasers Surg. Med.
ß 2006 Wiley-Liss, Inc.
Key words: Er:YAG; thermal pulses; collagen neogenesis;
histology; immunohistochemistry; minimally invasive
resurfacing; optical coherence tomography (OCT);
scar reduction; temperature recording; wrinkle reduction
ß 2006 Wiley-Liss, Inc.
INTRODUCTION
Patients and physicians look for less invasive techniques
to improve rhytides and scars. For many years, ablative
lasers (CO2 and more recently Er:YAG lasers) have been
used successfully for the treatment of wrinkles and scars
[1–5], but their use is limited by the pain and down time for
the patient and the relatively high risk of side effects and
complications for the physician [6]. Therefore, the market
of non-ablative techniques is growing fast and all kinds of
different methods are available that claim to be efficient for
reduction of wrinkles and scars. But after critical review
and assessment of current literature in terms of their
efficacy, all these non-ablative methods are not a comparable
alternative to the ablative skin resurfacing [7–10].
Both physician and patient should be willing to accept
subtle, incremental, and gradual improvements. Also, a
relatively high proportion of non-responders limit the
clinical success rate [11]. Additionally these methods often
need multiple treatments, are time consuming, and sometimes
painful which makes them less attractive than
originally intended. Finally, the non-ablative rejuvenation
treatment modalities are based on newly developed
technologies. Therefore, these new devices with modest
efficacy are often expensive for the practitioner.
Compared to the CO2 laser, the main advantage of the
Er:YAG laser for skin resurfacing, is precise ablation with
limited residual thermal damage (RTD). This results in
faster reepithelialization and an improved side effect
profile. On the other hand, immediate collagen shrinkage
and delayed new collagen formation is reduced together
with poor intra-operative hemostasis.
Karin Kunzi-Rapp and Christine C. Dierickx contributed
equally to the work.
*Correspondence to: Dr. Karin Kunzi-Rapp, Institute for Laser
Technologies and Metrology in Medicine, University of Ulm,
Helmholtzstr. 12, D-89081 Ulm, Germany.
E-mail: karin.rapp@ilm.uni-ulm.de
Accepted 15 June 2006
Published online in Wiley InterScience
(www.interscience.wiley.com).
DOI 10.1002/lsm.20380

2 KUNZI-RAPP ET AL.
In an attempt to overcome the limitations of the shortpulsed,
ablative Er:YAG lasers, modulated (short- and
long-pulsed) Er:YAG systems were introduced. With the
addition of significant coagulative properties, modulated
Er:YAG systems combine precise control of ablation with
the ability to improve hemostasis and induce dermal
collagen formation by means of thermal injury.
However, compared to non-ablative skin rejuvenation
techniques, conventional CO2 and erbium resurfacing
techniques cause thermo-mechanical tissue ablation with
the implication of visible and longer healing times. This has
caused that traditional CO2 and erbium resurfacing
techniques are nowadays largely abandoned, but has
stimulated the search for techniques to deliver energy with
these lasers, deeper in the skin without the unwanted
ablation.
The main effect of CO 2 and Er:YAG laser resurfacing is
the stimulation of new collagen growth in the dermis.
Histologically, it has been shown that these new collagens
replace the elastotic collagen of the connective tissue
matrix associated with wrinkles and photodamaged skin
in the upper dermis. Because water is the target chromophore
of these lasers, they ablate the full epidermis before
the laser energy affects the papillary dermis by heat
diffusion.
Theoretically, animal and clinical studies have shown
that it is indeed possible to deliver CO 2 or erbium laser
energy deeper in the skin without causing unwanted
epidermal ablation [12–20]. Ross et al. 1999 [12] described
the effect of stacking pulses of a CO 2-laser causing a less
distinct line between denatured and intact collagen and an
increased depth of the RTD. Using heat transfer models as
well as animal models, it could be shown, that an erbium
laser pulse can achieve greater RTD by lengthening the
pulse width [13–15].
The goal of our study was to determine whether an
Er:YAG laser used with a sequence of sub-ablative pulse
fluences below the ablation threshold, a so called thermal
mode, could produce enough dermal injury without complete
epidermal ablation to cause new collagen synthesis.
To evaluate the efficacy and safety for rejuvenation of a
laser system already established in the dermatological
practice the thermal mode of an Er:YAG laser was used for
the treatment of facial wrinkles. In a first preliminary
study we used a sub-ablative setting. A group of 29
volunteers were treated twice at a 6-month interval at 33
areas (periorbital 19, upper lips 11, lower lips/chin 2, cheeks
1) with a single pass of Er:YAG thermal laser pulses at subablative
fluences of 3.1–4.2 J/cm 2 , using parallel cooling
with cold air. The clinical efficacy was evaluated by
comparing pre- and post-photographs taken at baseline,
1, 3, 6, and 12 months after the two treatments. Although
the wrinkles were improved transitory at months 1 and 3,
the 6 and 12 months follow-up photographs revealed no
improvement, evaluated by the volunteers themselves,
by the physicians or by uninvolved, blinded interpreters
(Fig. 1).
As some individuals showed good clinical responses [21]
and some investigations reported histological changes
Fig. 1. Periorbital wrinkles treated with a single pass of
Erbium:YAG thermal laser pulses at sub-ablative fluences of
2.4 J/cm 2 , parallel cooling with cold air, (A) before, (B) 7 months
after treatment with mild structure changes.
[22–24], a second study was initiated. In this study, the
efficacy of multiple passes of Er:YAG thermal laser pulses
at fluences below the ablation threshold was evaluated by
clinical means, by histology and immunohistochemistry, by
optical coherence tomography (OCT) and by in vitro/in vivo
temperature measurements.
MATERIALS AND METHODS
Laser
Two variable pulsed Er:YAG lasers (SupErb XL and
BURANE XL, WaveLight Laser Technologie AG, Erlangen,
Germany) were used. We used the laser in a special thermal
mode consisting of a sequence of 9–11 short pulses each
with a fluence below the ablation threshold with an overall
pulse duration of 200–270 milliseconds and a total energy
density of 2.1–3.1 J/cm 2 . The parameters of this pulse

sequence were determined by temperature calculations as
well as Monte–Carlo simulations in order to optimize
heat penetration by conduction. Sub-ablative thermal
Er:YAG laser pulses heat the stratum corneum and the
epidermis due to absorption by the water content and
cause temperature increase of the upper dermis by heat
conduction.
Histology
The tissue samples were fixed in 4% freshly prepared
paraformaldehyde, paraffin embedded, cut in 3 mm sections
and stained with hematoxylin and eosin for histopathology.
A specialized procedure to pronounce collagen fibers was
done by Alcian blue staining.
Immunohistochemistry
Paraffin sections were mounted on poly-L-lysine coated
slides. After deparaffinizing and rehydrating antigen
retrieval of sections was done by protein kinase K at 378C
for 10 minutes. Non-specific binding sites were blocked
with goat serum. Human pro-collagen Type I C-peptide
mouse monoclonal antibody (TaKaRa, Bio Europe S.A.,
Gennevillier, France) was applied according to the manufacturer
protocol. The highly sensitive Histostain-Plus
streptavidin peroxidase staining procedure (Zymed
Laboratories, Inc., San Francisco, CA) was used with
DAB chromogen staining. At optimal color development
sections were immersed in sterile water, counterstained
with Mayer’s hematoxylin and covered.
Quantitative assessment of pro-collagen expression.
To determine pro-collagen expression, positive staining
fibroblasts were evaluated at 100 magnification using
an optical microscope (Axiophot, Carl Zeiss, Jena, Germany)
with a CCD camera (Sony MC-3249) and calculated
as percentage of the total number of fibroblasts by an
imaging analysis software (OPTIMAS, MediaCypernetics,
Silverspring MD). The numbers in Figure 5 represent
averaged values of four counts.
Optical Coherence Tomography (OCT)
OCT (ISIS Optronics GmbH, Mannheim, Germany) was
performed before, and at 4, 14, and 28 days after single pass
treatment of the outer forearm in two volunteers with
Er:YAG thermal pulses (fluence 4.2 J/cm 2 , spot size 5 mm),
parallel cooling with cold air.
Temperature Recording
For temperature measurements we used a digital
temperature probe (80 TK thermocouple module, Fluke
Cooperation, Everett WA).
Patient Selection
Patient selection for the morphological (biometrical)
study. Tissue biopsies were taken from seven
volunteers scheduled for blepharoplasty or abdominoplasty
with Fitzpatrick skin types 2–3 after informed written
consent was obtained before, 2 and 4 weeks after treatment
with Er:YAG thermal pulses at the same body side.
MINIMALLY INVASIVE SKIN REJUVENATION WITH ER:YAG 3
Patient selection for the clinical study. Subjects
with wrinkles or scars were included in the clinical
study. The first group consisted of 20 female subjects with
peri-orbital, peri-oral, or wrinkles on the cheeks. The
Fitzpatrick phototypes ranged from I to III and the ages
varied from 38 to 78 years (mean 55 years).
A second group included 12 patients with scars. Six
female and six male patients with phototypes I–III and
ages ranging from 12 to 39 years (mean 29 years) were
treated. The scars were either post-traumatic in nature and
located on the face and extremities or were atrophic facial
scars due to acne.
Treatment Protocol
All patients were treated with a 2,940 nm, variable pulse
Erbium:YAG laser (SupErb XL or BURANE XL, Wave-
Light Laser Technologie AG) in the thermal mode program.
Thermal mode pulse sequences (total pulse duration 200–
270 milliseconds) at sub-ablative fluences were used at
total energy density of 2.1–3.1 J/cm 2 for a 5 mm spot size
and at a frequency of 3 Hz. All procedures were performed
without local anesthesia. Eyes were protected with eye
shields. Parallel to the application of thermal laser pulses
cooling was provided with a constant flow of cold air level 1
(Cryo 5 skin cooling system, Zimmer MedizinSystems, Neu-
Ulm, Germany). No pre-cooling was performed. At each
treatment, the entire treatment area was irradiated with
3–5 consecutive laser passes with minimal overlap. No
wiping was performed between the passes. Additional
passes over the wrinkles or scars were given until the
clinical endpoint of skin whitening was obtained (Fig. 2).
The number of passes needed to reach the skin whitening
depended on the humidity of the skin and coincidences with
thermally induced necrosis of the stratum corneum and the
epidermis. Normally, more than five passes were necessary.
All treatment sites received two treatments with an
interval of 2 months. Post-exposure skin care consisted of
Fig. 2. Clinical endpoint (whitening of the epidermis) for the
Er:YAG treatment of wrinkles with multiple passes of Er:YAG
thermal laser pulses at sub-ablative fluences of 2.1 J/cm 2 and
parallel cooling with cold air.

4 KUNZI-RAPP ET AL.
an antibiotic ointment for reepithelialization and moistening
purposes. Sun blocks were recommended once healing
was complete. Patients came in for follow-up at 1, 3, 6, and
12 months post-treatment. Photographs were taken at each
follow-up visit and side effects were noted. Pre-and postoperative
photographs were captured with a digital Sony
camera (Cyber-shot 3.3 megapixels, DSC-F505V, Sony
Electronics) and were compared to evaluate treatment
response. Assessments were done by three blinded independent
reviewers, using a five-point improvement scale
(Table 1).
RESULTS
Biometrical Results
Histology. Examination of the biopsies taken 2 weeks
after treatment under light microscopy H&E staining
revealed a hypertrophic epidermis, caused by a thickened
stratum spinosum. In the papillary dermis we found
vasodilatation and a mild perivascular infiltration of
inflammatory cells. Four weeks after treatment the
epidermis flattened and the upper dermis demonstrated
more tightly packed collagen bundles with parallel orientation
to the skin surface (Fig. 3).
Structural evaluation of the treated samples compared to
non-treated samples showed a decrease in clumping of
collagen bundles and an increased amount of thin collagen
fibers with regular orientation in the upper dermis
extending from the basement membrane zone.
Immunohistochemistry. Immunhistological staining
4 weeks after treatment (Fig. 4) showed a marked increase
of pro-collagen expression in dermal fibroblasts till a depth
of about 320 mm. When we compared different treatment
modalities, mild ablation with two passes at a fluence of 5 J/
cm 2 as well as ablation followed by one pass of the
combination mode (ablative pulses in combination with
the thermal pulses) induced pro-collagen expression in
40.1% of the fibroblasts. Two passes of thermal pulses alone
showed a pro-collagen expression in 25.5%. In untreated
skin 5.6% of the fibroblasts were activated (Fig. 5).
Optical coherence tomography. OCT is a noninvasive
technique for high resolution imaging in the
tissue. It is based on the same concept like ultrasound:
waves are backscattered by tissue inhomogenities and
analyzed over time of flight to obtain spatial resolved
resolution. Before laser treatment OCT pictures of normal
skin clearly showed the epidermal–dermal junction separated
by a small band with a low scattering structure. In the
upper dermis clearly demarcated homogeneous dark
structures indicated the blood vessels. Four days after
thermal Er:YAG laser pulses crusting and edema were still
TABLE 1. Grading Scale
Worse 0
No improvement 1
Fair improvement 2
Good improvement 3
Excellent improvement 4
obvious. After 2 weeks OCT images could still detect
inflammatory cells in the tissue marked by blurred skin
structures. Four weeks after treatment dense reflecting
structures in the upper dermis were indicating an increase
of collagen fibers (Fig. 6).
Fig. 3. Histological sections H&E stained before and 4 weeks
after Er:YAG thermal laser pulses (original magnification
100 ): (A) before Er:YAG laser treatment and (B) 4 weeks after
Er:YAG thermal laser pulses; the epidermis flattened and in
the upper dermis (*) new collagen bundles and less elastosis
became obvious. [Figure can be viewed in color online via
www.interscience.wiley.com.]

Fig. 4. Immunhistochemical sections stained with mAb
against pro-collagen: (A) before and (B) 4 weeks after single
pass treatment with Er:YAG thermal laser pulses at subablative
fluences of 3.5 J/cm 2 , parallel cooling with cold air. The
brown staining indicates the pro-collagen expression of dermal
fibroblasts (original magnification 100 ).
Temperature recording. The basic approach of the
thermal pulses was to heat up the upper dermis to a
sublethal temperature and maintain this temperature for a
time period up to 300 milliseconds to induce collagen
remodeling. Monte–Carlo simulations as well as heat
transfer calculations resulted in temperature profiles at
different depths of the skin. These simulations resulted in a
pulse sequence consisting of short pulses with different
pulse energies below the ablation threshold. The calculations
determined the temperature increase till a maximum
of about 608C in the depth of the upper dermis (150 mm) was
reached. This temperature was maintained for more than
30 milliseconds. Temperature profiles were verified by
temperature measurements with a digital temperature
probe in ex vivo human skin samples. Different programs of
thermal pulses showed characteristic temperature elevations
in the epidermis shown in Figure 7.
MINIMALLY INVASIVE SKIN REJUVENATION WITH ER:YAG 5
% positive cells
45
40
35
30
25
20
15
10
5
0
25,5
In one volunteer, the temperature profile was measured
in vivo by placing the temperature probe into the dermis of
the forearm in a depth of 335 mm. The depth of this probe
was controlled by OCT. After application of one thermal
Er:YAG laser pulse sequence temperature profile was
recorded. In this depth the temperature increase was about
38C after a single thermal pulse sequence with a fluence of
3.5 J/cm 2 and a spot size of 5 mm. It took about 40 seconds to
reach the initial skin temperature (Fig. 7c). If cooling was
performed during repetitive pulses the time to reach the
value of the initial temperature was reduced to about
1 second (picture not shown).
Clinical Results
Average post-operative follow-up was 6–12 months for
the wrinkle group and 3–6 months for the scar group. The
wrinkle reduction or scar improvement was significant in
most cases and improved over time. Improvement of
rhytides at 1–3 months follow-up was graded as excellent
in 19%, good in 19%, fair in 31%, and no improvement in
31%. At the 6–12 months follow-up, the improvement was
excellent in 40%, good in 40%, fair in 20%, and no
improvement in 0% (Figs. 8 and 9). Improvement of scars
at 3–6 months follow-up was graded as excellent in 50%,
good in 25%, fair in 25%, and no improvement in 0%
(Figs. 10 and 11).
Intra- and post-operative discomfort was described as
mild by the patient. Superficial crusting occurred and
reepithelialization was complete in 3–4 days. Mild postoperative
erythema persisted for 3–4 weeks. In one patient
with a history of herpes simplex, a reactivation of latent
labial herpes simplex virus infection occurred after the first
laser treatment. When acyclovir was given prophylactically
40,1
TP 3.5 J/cm² ablation 5 J/cm² control
Fig. 5. Quantitative assessment of pro-collagen expression
calculated as a percentage of total number of fibroblasts (the
numbers represent averaged values of four counts). Biopsies
taken 4 weeks after single pass treatment with Er:YAG laser:
TP 3.5 J/cm 2 : thermal pulses at sub-ablative fluences of 3.5 J/
cm 2 ; ablation 5 J/cm 2 : ablative Er:YAG laser pulses at ablative
fluences of 5 J/cm 2 ; control: untreated skin in the same
individual of the same location (either eyelid or abdominal
skin).
5,6

6 KUNZI-RAPP ET AL.
Fig. 6. Optical coherence tomography (OCT) before and after
single pass treatment with Er:YAG thermal laser pulses at
sub-ablative fluences of 4.2 J/cm 2 , parallel cooling with cold air:
(A) before laser treatment OCT pictures clearly showed the
epidermal–dermal junction separated by a small band with a
low scattering structure (dotted line). In the upper dermis
clearly demarcated homogeneous dark structures indicated
the blood vessels *. B: Four days after Er:YAG thermal laser
before the second treatment, another outbreak could be
prevented. No hyper- or hypopigmentation, textural
changes or scarring were observed in this patient group
with skin types I–III. Average post-operative follow-up was
6–12 months for the wrinkle group and 3–6 months for the
scar group.
DISCUSSION
CO2 or erbium resurfacing is a well-established method
to treat facial rhytides associated with photoaging [1–5].
These techniques completely disrupt or remove the
epidermis. Subsequent loss of barrier function results in
discomfort, edema, transudation, and focal crusting.
Epidermal loss also increases the risk of infection, pigmentary
changes and scarring [6].
Non-ablative skin rejuvenation, which does not remove
the epidermis, was specifically developed as a bettertolerated
alternative to ablative laser resurfacing. The
objective is to achieve selective, heat-induced denaturation
of dermal collagen that leads to subsequent new collagen
deposition with as little damage to the epidermis as
pulses crusting and edema were still obvious (bars indicate
thickness of the crusts). C: After 2 weeks OCT images could
still detect inflammatory cells in the tissue marked by blurred
skin structures. The epidermal–dermal junction is not
demarcated. D: Four weeks after treatment dense reflecting
structures in the upper dermis were indicating an increase of
collagen fibers. [Figure can be viewed in color online via www.
interscience.wiley.com.]
possible. The main problem with non-ablative skin rejuvenation,
however, is poor and/or unpredictable efficacy
compared with ablative treatments [7–11,25,26].
Microscopic changes associated with wrinkles occur
primarily in the dermis [27–30]. Wrinkle reduction, by
means of thermal damage to the dermis, is based on the
induction of synthesis of new collagen and other components
of extracellular matrix [13,25]. In this study, a
thermal mode Er:YAG laser was used to examine the effect
on wrinkle/scar reduction by dermal heating.
At commonly utilized ablative Er:YAG parameters, the
zone of RTD typically does not exceed 50 mm [33,34]. The
amount of thermal damage is dependent on the repetition
frequency, while the laser fluence is of much less importance
for the depth of necrosis [35,36]. The thermal pulse
structure in this study was composed of a sequence of short
Er:YAG pulses (200–270 milliseconds) below the ablation
threshold. It was developed in such a way to increase the
temperature in the upper dermis to about 608C in order to
induce collagen denaturation. In our ex/in vivo studies we
could confirm the temperature increase to 608C only for the

A
temperature / °C
B
temperature / °C
C
temperature / °C
80
60
40
20
0
0 10 20
time / s
30 40
100
80
60
40
20
0
0 5 10 15 20 25 30
time / s
33
32
31
30
29
28
27
0 20 40 60 80
time / s
Fig. 7. Temperature measurements with a digital temperature
probe: (A) at the epidermis in ex vivo skin samples after a
single Er:YAG thermal laser pulse (fluence 3.5 J/cm 2 , spot size
5 mm); (B) at the epidermis in ex vivo skin samples after a
single Er:YAG thermal laser pulse with a fluence of 4.2 J/cm 2 ;
(C) in vivo by placing a temperature probe into the dermis of
the forearm at a depth of 330 mm (OCT controlled) after a single
Er:YAG thermal laser pulse (fluence 3.5 J/cm 2 , spot size 5 mm);
the temperature profile was recorded for 70 seconds. It took
about 40 seconds to reach the initial skin temperature.
layers near the basal membrane zone. In deeper dermal
layers the temperature increase was only very mild after
application of only one pulse. However, in our clinical
study, we used multiple passes over the wrinkles. This most
likely resulted in a thermal build up by heat conduction
because the thermal relaxation of the treated tissue is very
slow (Fig. 7). After desiccation of the tissue the main
chromophore for the Er:YAG laser radiation deprived. As a
result, the optical penetration depth was enlarged, resulting
in further diminished ablation efficiency, enhanced
deposition of heat, and increased the zone of thermal injury.
MINIMALLY INVASIVE SKIN REJUVENATION WITH ER:YAG 7
% Patients
50
45
40
35
30
25
20
15
10
5
0
none slight moderate dramatic
1-3 months 6-12 months
Fig. 8. Wrinkle improvement in 20 female subjects with
periorbital, perioral, or wrinkles on the cheeks treated with
multiple passes of Er:YAG thermal laser pulses at sub-ablative
fluences of 2.1 J/cm 2 , parallel cooling with cold air. Assessments
were done on photographs taken at 1–3 months or 6–
12 months follow-up by three blinded independent reviewers,
using a five-point improvement scale (Table 1).
This finding can be understood by recalling that the
threshold fluence for skin ablation with the Er:YAG laser
is between 0.5 and 1 J/cm 2 [14,33,35]. At pulse fluences
below these values, the coagulation depth increases
linearly with the applied fluence [36]. Longer pulse
durations increase the ablation threshold [15]. A higher
ablation threshold simply enables a larger heat deposition
into the tissue and the elevated temperature persists for a
longer time due to the lower temperature gradients at these
depths.
While non-ablative skin rejuvenation with intense
pulsed light sources, visible or near-infrared light sources
or radiofrequency cause no epidermal damage at all, a
thermal mode Er:YAG laser damages the epidermis but
does not remove it [10].
After two passes of thermal pulses clinical as well as OCT
data revealed limited thermal damage of the epidermis. In
an in vivo skin model we could show 4 days after laser
treatment mostly upper epidermis was injured but basal
Fig. 9. Wrinkle improvement at 12 months follow-up in a 56year-old
woman treated with multiple passes of Er:YAG
thermal laser pulses at sub-ablative fluences of 2.1 J/cm 2 ,
parallel cooling with cold air.

8 KUNZI-RAPP ET AL.
% Patients
60
50
40
30
20
10
0
no slight moderate dramatic
Fig. 10. Scar improvement in 12 patients (6 female, 6 male)
with post-traumatic or acne scars located on face and
extremities treated with multiple passes of Er:YAG thermal
laser pulses at sub-ablative fluences of 2.1 J/cm 2 , parallel
cooling with cold air. Assessments were done on photographs
taken at 3–6 months follow-up by three blinded independent
reviewers, using a five-point improvement scale (Table 1).
keratinocytes were preserved [22,23]. The damaged epidermis
was not removed and acted as a wound dressing.
After reepithelialization, a hypertrophic epidermis as well
as inflammatory cells in the upper dermis persisted for
more than 2 weeks. This indicated that the reparative
phase was not finished at that time. Immunhistochemical
findings still showed activated fibroblasts with pro-collagen
1 expression in the upper dermis 4 weeks after treatment.
The number of these fibroblasts was significantly higher
than the basal expression of pro-collagen 1 in untreated
skin, but it was not as high as the expression after a mild
ablative treatment. Because the temperature increase in
the upper dermis in ablative resurfacing is less than after
sub-ablative thermal pulses the effect seems not only to be
based on the temperature. Fatemi et al. [37] focused on the
early histological changes after non-ablative laser treatment
with a 1,320 nm Nd:YAG laser. They concluded that
immediate vascular damage, recruitment of inflammatory
cells, and release of mediators may be responsible for
the clinical improvements associated with non-ablative
Fig. 11. Scar improvement at 6 months follow-up in a 49-yearold
woman treated with multiple passes of Er:YAG thermal
laser pulses at sub-ablative fluences of 2.1 J/cm 2 , parallel
cooling with cold air. [Figure can be viewed in color online via
www.interscience.wiley.com.]
resurfacing. Recently Drnovsˇek-Olup et al. [38] found a
sub-epidermal regeneration zone to a depth of about 120–
240 mm after treatment with non-ablative Er:YAG laser
fluences of 1.5 and 1.75 J/cm 2 . This zone consisted of
edematous tissue with stellate appearing cells with
immunhistochemically positive staining to smooth muscle
actin monoclonal antibody. These activated fibroblasts did
undergo an epidermal–mesenchymal transition (EMT)
which indicated the proliferative phase of wound healing
with production of extracellular matrix components [39].
The last phase of wound healing is characterized by
remodeling of the granulation tissue. In this stage collagen
3 is replaced by collagen 1 and proteoglycans are synthesized.
This neocollagen brings an indispensable support to
the dermis and fills the wrinkle. Furthermore myofibroblasts
in wound healing are responsible for wound
contraction and fibroplasia [40,41].
The purpose of our clinical study was to determine the in
vivo response to dermal injury without complete epidermal
ablation. We therefore used a 2,940 nm Er:YAG laser in a
thermal mode with sub-ablative settings: fluence of 2.1–
3.1 J/cm 2 , 200–270 milliseconds, slight overlapping, a spot
size of 5 mm, a repetition rate of 3 Hz and multiple pulse
stacking mode. These laser settings were chosen on the
base of results of theoretical studies of repetitive Er:YAG
treatments [14,36].
We showed that the thermal damage pattern included
limited epidermal damage, but no epidermal removal. A
rapid repair response was initiated with complete recovery
of the epidermis within 3–4 days with minimal discomfort.
Since the dermis was uniformly denatured by the multiple
pulse stacking technique, we introduced a way to reproducibly
induce denaturation of the treated tissue area
and thus may allow to achieve more reproducible therapeutic
results: in the wrinkle group up to 80% of patients
obtained moderate to dramatic improvement of wrinkles
while 75% of patients obtained the same results in the scar
group.
Although clinically observable results were present in
most of the patients, overall patient satisfaction was low.
The in vivo response to tissue damage consists namely of
three consecutive phases: an inflammatory phase, a
proliferation phase, and a remodeling phase [31,32]. This
explains why clinically observable results were only seen
between 3 and 6 months after initial treatment, with
further improvement between 6 and 12 months. This
relatively long period before clinically observable results
were seen, may lead to low patient satisfaction grade.
Additional topical or other non-invasive treatment modalities
might help to speed up the results and increase the
patient satisfaction.
CONCLUSIONS
Thermal mode Er:YAG laser pulses can induce collagen
neogenesis, as proved by temperature elevation and
morphological changes in the upper dermis, which leads
clinically to visible and long lasting reduction of wrinkles
and scars after applying multiple passes with minimal
side-effects.

ACKNOWLEDGMENTS
The authors thank Detlev Russ, Institute for Laser
Technologies in Medicine and Metrology at the University
of Ulm for providing the theoretical data and for the
performance of the temperature measurements.
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