Characterization of the canine skin barrier restoration following acute disruption by tape stripping

Characterization of the canine skin barrier restorationfollowing acute disruption by tape stripping

Emilie Videmont*, Claire Mariani†, Stephanie

Vidal† and Didier Pin*‡

*Unite Dermatologie and

‡UPSP ICE (Interactions Cellule-Environnement), VetAgro Sup Cam-

pus Veterinaire de Lyon, Marcy l’Etoile F-69280, France

†Royal Canin SAS, Aimargues F-30470, France

Correspondence: Emilie Videmont, Unite Dermatologie, VetAgro

Sup Campus Veterinaire de Lyon, Marcy l’Etoile F-69280, France.

E-mail: [email protected]

Sources of Funding

The laboratory Royal Canin has partially funded the study.

Conflict of Interest

Claire Mariani and Stephanie Vidal are employees of

Royal Canin.

Abstract

The stratum corneum (SC) forms the main part of

the permeability barrier of the skin. In mice and in

humans, cutaneous barrier disruption can be gener-

ated by removing the SC with tape stripping (TS) and

the skin barrier function can be assessed by measure-

ment of transepidermal water loss (TEWL). The aim

of the present study was to characterize the skin bar-

rier restoration in the dog following mechanical dis-

ruption and to analyse the correlation between the

skin barrier recovery and TEWL measurement. Thirty

sequential TS were performed on 12 sites on four

healthy beagle dogs. The number of TS was chosen

to ensure a sufficient barrier disruption with a slow

recovery. Skin repair was assessed for 72 h by clinical

and histological examinations, and TEWL measure-

ments. The results showed that performing 30 TS

was adequate to disrupt the skin barrier in the dog.

The homeostatic repair response, initiated in the

skin, was rapid and characterized by complete

restoration of the SC within 72 h, accompanied by

greater basal cell proliferation, and dermal eosinophi-

lic inflammation. TEWL was significantly increased

by complete removal of the SC but recovered along

with restoration of the SC (Scheffe test, P £ 0.05).

Characterization of a canine model of barrier disrup-

tion and restoration and assessment of the skin bar-

rier function by TEWL measurements could help

better understand the events implied in skin barrier

function. Development of this canine model is also

necessary for future studies on the effects of treat-

ments aimed at restoring the skin barrier.

Accepted 4 August 2011

Introduction

The barrier function of the skin is primarily controlled by the

stratum corneum (SC), the outermost layer of the epider-

mis.1,2 One of the most important functions of the SC is to

form a permeability barrier that allows survival in a poten-

tially desiccating external environment. The barrier to

water permeation is not absolute, and normal movement

of water through the SC into the atmosphere is called

‘transepidermal water loss’ (TEWL), an insensible water

loss. The measurement of TEWL is widely used in humans

and laboratory animals to assess the skin barrier function in

a noninvasive manner. Increases in TEWL are consistent

with skin barrier impairment.3 Removal of the SC by

repeated applications of adhesive tapes is known as tape

stripping (TS).4,5 This mechanical disruption of the barrier

triggers a rapid homeostatic response to restore skin bar-

rier function.6 Barrier restoration following TS has been

studied extensively in murine models. Tape stripping and

measurement of TEWL are used to test the effect of topical

treatments aimed at restoring the skin barrier function.7,8

In recent years, TEWL measurement has been per-

formed in dogs, and the repeatability of the method varies

from study to study.9–12 The TEWL has been reported to

be affected by breed,10 age,13 anatomical site,14 and hair

clipping.14,15 Current studies indicate that TEWL measure-

ment is a suitable method to assess the barrier function in

dogs. Reports concerning the use of TS in dogs are limited.

A study by Shimada et al.16 described the disruption of the

skin barrier in the dog by TS, correlating skin barrier func-

tion with TEWL measurements. In a recent study, Olivry et

al.17 reported that performing TS in atopic dogs removed

the SC and facilitated experimental sensitization to mite

allergens. Finally, Ohmori et al.18 evaluated, with TEWL

measurements, the effect of ultrapure soft water on the

skin barrier restoration after experimental disruption by TS.

Data concerning the skin barrier restoration after mechani-

cal disruption are, to the best of the knowledge of the

authors, lacking with respect to the dog.

The aim of the present study was to characterize in the

dog the response of the skin following acute barrier dis-

ruption by TS and to compare this response with those

observed in humans and mice using clinical, histological

and TEWL measurements. Development of a canine

model of skin barrier disruption and restoration is neces-

sary for future studies on the effect of treatments aimed

at restoring the skin barrier.

Materials and methods

All the procedures used in this study were approved by the Institu-

tional Animal Care and Use Committee of the National Veterinary

School of Lyon, France.

ª 2011 The Authors. Veterinary Dermatology

ª 2011 ESVD and ACVD, Veterinary Dermatology, 23, 103–e23. 103

DOI: 10.1111/j.1365-3164.2011.01019.x

AnimalsFour 2-year-old, neutered male beagle dogs were used in the study.

Prior to the study, dogs were examined to ensure they were healthy

and had no history or evidence of skin lesions. None of the dogs had

received systemic or topical therapies for 3 months prior to the start

of the study. The dogs belonged to a research colony housed indoors

in a temperature- and humidity-controlled facility (25–28�C, relative

humidity of 40–60%). The dogs were housed in individual cement

runs cleaned twice daily. The dogs were fed maintenance dry food

for 12 weeks prior to testing and tap water ad libitum. Before the

start of the experiment, the dogs were acclimated to their environ-

ment for 1 week.

Tape strippingTape stripping of the SC for barrier disruption was performed as fol-

lows. A commercial adhesive tape was used (Transparent Tape

Scotch�, 19 mm; 3M France, Cergy-Pontoise, France). At each test

site, a new piece of tape was applied and compressed with finger

pressure for 10 s before being removed in one swift motion.

Experimental procedureThe lateral thorax of the dog was selected as the test site because it

provided adequate surface area for TS, skin biopsy and a horizontal

orientation for the evaporimeter probe. In addition, TS in this area trig-

gered little response by the dog and was less likely to be licked. The

hair was clipped using an electric hair clipper 24 h before the start of

the study in order to minimize any effect of recent hair clipping. The

same investigator performed all of the procedures and measure-

ments in order to minimize interindividual variations. A total of 12

sites were evaluated, six on each lateral thorax. Sites were approxi-

mately 4 cm2 and 3 cm apart. In a preliminary study, we performed

increasing numbers (from five to 50) of TSs in two healthy beagle

dogs and measured TEWL. Performing 30 TSs led to an eight- to

ninefold increase in TEWL over the basal value. In mice, Oudshoorn

et al.7 have shown that a mechanical disruption of the skin barrier

leading to an eight- to ninefold increase in TEWL over the basal value

allowed the creation of an accurate model to study the skin barrier

restoration. These previous findings led us to perform 30 consecutive

TSs on each site on day 1 in the present study.

Clinical observationsEach site was monitored for changes in skin colour, crusts, scaling

and ⁄ or exudation and inflammation before, immediately after and 18,

24, 48 and 72 h after performing TS.

Histological examinationSkin biopsy specimens were collected using local anaesthesia [sub-

cutaneous injection of lidocaine (Xylovet; Ceva, Libourne, France)]

and a 6 mm skin biopsy punch. Specimens were collected from

normal skin and test sites at 1, 6, 18, 24, 48 and 72 h after TS. A

normal skin sample was collected from one of two control sites

located 3 cm below the first tape-stripped site (the other control site

was used to measure TEWL). Skin biopsy specimens were fixed in

10% neutral buffered formalin, embedded in paraffin, routinely

processed in the laboratory of two of the authors (EV, DP), sectioned

at 4 lm thickness and stained with haematoxylin and eosin. The fol-

lowing histological changes were noted: stratum corneum thickness,

epidermal hyperplasia, modification of the different epidermal layers

(i.e. hyper- or hypogranulosis), dermal inflammation and cellular infil-

tration. Eosinophils were visualized and counted using Luna’s stain

for eosinophils. All the skin biopsies were examined by two of the

authors (EV, DP), blinded to the samples.

ImmunohistochemistryImmunohistochemistry was performed on formalin-fixed, paraffin-

embedded tissues. The antibodies used included anti-Ki-67 antigen

(1:25 dilution, MIB-1, mouse monoclonal; DakoCytomation, Glostrup,

Denmark) labelling proliferative cells, anti-myeloid ⁄ histiocyte antigen

(1:50 dilution, MAC 387, mouse monoclonal; DakoCytomation) label-

ling macrophages, anti-CD3 antigen (no dilution, CD3-12, rabbit

monoclonal; AbCys, Paris, France) labelling T cells, anti-CD21 antigen

(1:50 dilution, BL13, mouse monoclonal; Immunotech, Marseille,

France), anti BLA 36 antigen (1:25 dilution, A27.42, mouse monoclo-

nal; AbCys) and anti-CD79a antigen (1:25 dilution, HM 57, mouse

monoclonal; DakoCytomation) labelling B cells, and anti-CD208 ⁄DC-LAMP antigen (1:100 dilution, 1010E1.01, rat monoclonal;

Dendritics, Lyon, France) labelling dendritic cells.

Briefly, antigen retrieval was performed by heat treatment in

citrate buffer at pH 6 for 40 min. Endogenous peroxidase activity

was blocked with hydrogen peroxide for 10 min at room temperature

and then rinsed in phosphate-buffered saline for 10 min. For

MAC 387, sections were treated with 0.1% trypsin at 37�C for

20 min. To reduce nonspecific binding, slides were treated with a

blocking reagent (Super Block; ScyTek Laboratories, Logan, UT,

USA) for 30 min at room temperature before application of the pri-

mary antibody for 1 h. After rinsing, slides were incubated for 10 min

with biotinylated goat anti-polyvalent antibody (UltraTek Anti-Polyva-

lent; ScyTek Laboratories) followed by 10 min incubation with strep-

tavidin–peroxidase (UltraTek HRP; ScyTek Laboratories). Peroxidase

activity was detected with a peroxidase substrate kit (Vector� Nov-

aREDTM substrate kit; Vector Laboratories, Burlingame, CA, USA).

Slides were counterstained lightly with Mayer’s haematoxylin for

2 min, before rinsing, dehydrating, clearing and mounting with cover-

slips. Tissue sections treated with phosphate-buffered saline instead

of the specific antibody were used as negative controls. The Ki-67

labelling index (KI) was calculated as the percentage of positive nuclei

divided by the total number of keratinocytes examined. At least 500

keratinocytes per section were examined in 10 randomly selected

fields using light microscopy (·400 magnification).

Measurements of TEWLThe TEWL was measured before and at 1, 3, 24, 48 and 72 h after

TS. Basal TEWL was measured on the second control site (lateral

thorax, located 3 cm below the first tape-stripped site). To assess

day-to-day variations, TEWL was also measured on this control site

24, 48 and 72 h after the start of the study. The TEWL was measured

using a closed-chamber evaporimeter (VapoMeter�; Delfin Technolo-

gies Ltd, Kuopio, Finland). This device has a digital screen providing

readings for TEWL (in grams per square metre per hour) from the

skin and a measurement of relative humidity and temperature in the

chamber before skin contact. The VapoMeter� was activated by

pressing a single button, after which it was placed directly onto the

skin, perpendicular to the surface. As the probe was 1 cm in diame-

ter and the size of the test area was 2 cm, the probe was positioned

in the centre of the test area for the measurement. The pressure

applied to the probe was just enough to prevent leakage of air

between the probe and the skin. The device automatically deter-

mined the measurement duration and signalled completion with an

audible tone. All TEWL measurements were done in triplicate, and

the mean of the readings was used for analysis. All measurements

were performed in air-conditioned rooms where the temperature and

humidity were the same as the facility in which the dogs lived. Prior

to each measurement, the dogs were acclimated to experimental

conditions for 15–20 min. Dogs were not allowed to exercise during

the hour prior to the measurements. The percentage of barrier recov-

ery was calculated using the following formula:

1� ½ðTEWL at indicated time point� TEWL of control siteÞ=ðTEWL immediately after tape stripping� TEWL of control siteÞ�

The TEWL at the control site represented the TEWL values measured

at the control site before the start of the tape-stripping procedure.

Statistical analysisAll data were analysed using the Statistical Analysis Systems Institute

package (Statistical Analysis System, SAS user’s guide, version 8;

SAS Institute Inc., Cary, NC, USA, 1999).Transepithelial water loss

values were normally distributed and analysed by ANOVA using the

mixed procedures of SAS. The statistical model included the fixed


104 ª 2011 ESVD and ACVD, Veterinary Dermatology, 23, 103–e23.

Videmont et al.

effect of the time-related tape-stripping execution with seven levels

(before TS, immediately after TS, 1, 3, 24, 48 and 72 h after TS). The

various sites on the same dog were defined as a random term to com-

pare TEWL values of each site per dog through the time ⁄ tape strip-

ping. Least squares (LS) means statement was used to fit data to this

statistical mixed model. Differences between LS means were deter-

mined using the Scheffe test, with a significance level of P £ 0.05.

The TEWL values are expressed in the text as LS means ± SEM.

Results

Clinical assessment

Tape stripping induced erythema, shininess and loss of

skin texture (i.e. ‘cobblestones’). There was no exudation

or scaling (Figure 1a). Inflammation gradually decreased

during the study (Figure 1b–d). Very small crusts were

present on the disrupted skin 48 h post-TS procedure

(i.e. during recovery; Figure 1c). The skin had a normal

appearance after 72 h, and hair had partly regrown

(Figure 1d).

Histological findings

Tape stripping completely removed the SC in all dogs,

with minimal alteration of the underlying viable epider-

mis (Figure 2a). Six hours after TS, multifocal small

areas (100–150 lm in length) of epidermal necrosis

were present. These areas were subsequently covered

by small crusts that gradually disappeared within 48 h.

After a 48 h recovery period, some uneven, elongated,

pointed rete ridges were observed. Regular hyperplasia

was present after 72 h. The intensity of hyperplasia var-

ied from moderate to severe. Cell layers of corneocytes

gradually reappeared; after 48 h, one or two corneocyte

layers were observed, and a complete SC was observed

after 72 h (Figure 2b). The number of cell layers of

corneocytes varied from three to more than six. Dermal

inflammation, dermal oedema and congestion, extrava-

sation of erythrocytes from dermal vessels and dermal

infiltrate appeared as early as 1 h after TS. Inflammation

increased to its peak intensity between 18 and 24 h

after TS and subsequently decreased over time. The

infiltrate was sparse 1 h after TS and it was moderate

at its peak, perivascular to diffuse in distribution and

extended from superficial to mid-dermis. Eosinophils

represented more than 90% of the infiltrate. Luna’s

stain confirmed the predominance of eosinophils (Fig-

ure 2c). After the peak, the infiltrate gradually decreased

and was concentrated around vessels in the superficial

dermis. After a 48 h recovery period, mononuclear cells

were intermingled with eosinophils (Figure 2d). Seventy-

two hours after TS, the infiltrate was mild and

composed predominantly of these mononuclear cells.

Concerning histopathological findings, minor differences

were observed between the dogs and consisted mainly

of differences in the intensity of the reaction following

disruption.

Immunohistochemical findings

The dermal infiltrate was composed of some MAC 387-

positive cells and a majority of BLA 36-positive cells

(Figure 3a). Some BLA 36-positive cells were observed

1 h after TS, and their number significantly increased

from 18 to 48 h of recovery. These BLA 36-positive

cells often had projections expected with dendritic cells

(Figure 3a).

(a) (b)

(c) (d)

Figure 1. Macroscopic aspect of disrupted skin immediately after tape stripping (TS; a) and at 24 (b), 48 (c) and 72 h after TS (d). Twelve test sites

of 4 cm2 were delimited with a dermatological pen to permit the application of 2 · 2 cm pieces of tape to the same site. Immediately after TS, the

skin was very erythematous and shiny. Inflammation decreased gradually, and the skin had a normal appearance after 72 h of recovery.



Canine skin barrier restoration

No significant staining differences were observed in

any specimen using antibodies against DC-LAMP, CD3,

CD21 and CD79a.

The KI-67 labelling index was stable until 48 h following

TS, with an index close to that observed in normal skin

and ranging from 5 to 15% (Figure 3b). At 48 h of recov-

ery, the KI ranged from 70 to 90% (Figure 3c), with

nuclear staining regularly distributed in the majority of

both the basal and the spinous layers. The KI decreased

markedly after 72 h and ranged from 20 to 30%.

Measurements of TEWL

Before disruption of the skin barrier, the TEWL values

were homogeneous between the four dogs (Figure 4a),

ranging from 11.76 to 12.8 g ⁄ m2 ⁄ h, with a mean of

12.26 g ⁄ m2 ⁄ h. When TEWL was measured at a control

site before TS and 24, 48 and 72 h after the start of the

study, no significant day-to-day variations were observed.

The four TEWL values of the control site ranged from

12.1 to 13.6 (SEM 0.58) for dog 1, from 10.8 to 12.1

(SEM 0.50) for dog 2, from 11.8 to 13.3 (SEM 0.66) for

dog 3 and from 11.4 to 13.1 (SEM 0.66) for the remain-

der. Tape stripping led to a marked increase in TEWL,

with a mean value of 104.88 g ⁄ m2 ⁄ h (range 96.87–

115.5 g ⁄ m2 ⁄ h) immediately after TS. The TEWL value

gradually decreased over time. Variations were observed

between dogs. In some of them, a linear decrease was

observed, whereas in others some occasional increases

in the TEWL value were noted. Statistical differences

were significant between the TEWL value of undisrupted

skin and the TEWL value immediately and 1, 3 and 24 h

after TS (Scheffe test, P £ 0.05). During the recovery

phase, the statistical difference was also significant for all

pairwise analyses (P £ 0.05), except between TEWL

values measured immediately and 1 h after TS (P =

0.9270) and TEWL values measured 48 and 72 h after TS

(P = 0.9962). After 72 h of recovery, TEWL was still

slightly elevated (mean 19.94 g ⁄ m2 ⁄ h; range 16.89–

22.9 g ⁄ m2 ⁄ h) in comparison with values of undisrupted

skin, but the difference was not significant (P = 0.4080;

(a) (c)

(b)

(d)

Figure 2. Pathological changes in the skin following TS. (a) Skin 1 h after TS. (b) Skin 72 h after TS. (c) Skin 18 h after TS. (d) Skin 48 h after TS.

Tape stripping completely removed the stratum corneum (SC; a). Corneocytes reappeared after 48 h, and a SC, ranging from two to four cell

layers, was observed after 72 h (b). The intensity of inflammatory infiltrate was maximal 18 h after TS, and it was composed mainly of eosinophils

(see arrows), as confirmed with Luna’s stain (c). Forty-eight hours after TS, the infiltrate was mild and composed predominantly of mononuclear

cells (d). Haematoxylin and eosin, except Luna’s stain in (c). Scale bars represent 20 lm.



Videmont et al.

Figure 4a). After TS, the recovery of the TEWL was very

fast; a recovery higher than 50% was observed within

24 h in three of the four dogs. After TS, a mean recovery

of 93.25% of the TEWL was observed within 72 h (SEM

1.94) (Figure 4b).

Discussion

The present study showed that performing 30 TSs was

accurate to alter the skin barrier in dogs and obtain a rela-

tively slow barrier restoration, with minimal alteration of

the viable epidermis. Indeed, restoration of the stratum

corneum, based on histological assessment, was com-

pleted in 72 h, and epidermal necrosis due to mechanical

trauma was minimal and disappeared rapidly. These

results are consistent with those observed in murine

models after the same intensity of skin barrier disruption.7

To the best of our knowledge, this study represents

the first investigation of the homeostatic response,

assessed by sequential histological examinations, occur-

ring after mechanical disruption of the skin barrier in the

dog. In the present study, removal of the SC by TS trig-

gered an early inflammatory response. Interestingly,

eosinophils, as confirmed by Luna’s stain, represented

more than 90% of the inflammatory infiltrate observed

6 h after TS. In a recent study in a murine model, eosin-

ophils also largely dominated in tape-stripped skin, in the

absence of any allergic or parasitic context.19 In another

study in mice, it was shown that TS polarized skin den-

dritic cells to elicit a T-helper 2 response20 that was

responsible for the influx of eosinophils. In further studies

in dogs, it would be interesting to study cytokine mRNA

levels by quantitative PCR after TS in order to compare

results in dogs with those observed in mice and confirm

the overexpression of T-helper 2 cytokines in the dog. In

the present study, eosinophilic infiltration was followed

by infiltration with a majority of BLA 36-positive mononu-

clear cells. BLA 36 is a plasma membrane glycoprotein

expressed on B-cell progenitors and B cells in humans

and in animals. In the dog, the antibody anti-BLA 36

seems also to recognize dendritic cells.21,22 In the pres-

(a)

(b)

(c)

Figure 3. Immunoreactivity to BLA 36 antibody (a) and to Ki-67 anti-

body (b, c). (a, c) Skin 48 h after TS. (b) Skin 1 h after TS. BLA 36-

positive cells appeared 18 h after TS and their number was elevated

until 48 h. In comparison with skin 1 h after TS (b), the majority of

basal and suprabasal cells expressed Ki-67 antigen 48 h after TS (c).

Immunohistochemistry. Scale bars represent 50 lm.

120

140

0

20

40

60

80

100

160

Undisruptedskin

Immediatelya er TS

1 h a er TS 3 h a er TS 24 h a er TS48 h a er TS72 h a er TS

TEW

Lg/

m2 /h

40

50

60

70

80

90

100

Reco

very

(%)

0

10

20

30

7248240Time (h)

(a)

(b)

Figure 4. (a) Variation of transepidermal water loss (TEWL) follow-

ing TS in the four dogs. The TEWL values are represented as least

square means ± SEM. Statistical differences between least square

means were determined using the Scheffe test, with a significant

level of P £ 0.05. Significant statistical difference existed between

undisrupted and disrupted skin immediately after and 1, 3 and 24 h

after TS. The difference was not significant at 48 and 72 h of recov-

ery. (b) Recovery rates of TEWL after TS in the four dogs. Values are

means ± SEM. After TS, the recovery of the TEWL was very fast; a

recovery higher than 50% was observed within 24 h in three of four

dogs. A mean recovery of 93.25% of the TEWL was observed within

72 h (SEM 1.94).




ent study, BLA 36-positive cells had a dendritic morphol-

ogy and they were negative for other B-cell markers,

CD21 and CD79a. Immunostaining with antibody against

dendritic cell-lysosomal-associated-membrane protein

(DC-LAMP) was unfortunately unable to confirm the den-

dritic origin of these BLA 36-positive cells.

Finally, histological examination revealed that TS stimu-

lated proliferation of epidermal cells and proliferative activ-

ity, assessed by the evaluation of Ki-67 index, which was

maximal 48 h following TS. This intense proliferative activ-

ity of epidermal cells is consistent with features observed

in humans5 and mice23 and probably results from the influ-

ence of cytokines.24 In the absence of further TS, the pro-

liferation rate rapidly returned to baseline value.

In humans, measurement of TEWL is widely used to

monitor changes in the cutaneous barrier function nonin-

vasively.25 Both open-chamber and closed-chamber

devices accurately measure TEWL.26,27 In comparison

with human medicine, data concerning TEWL measure-

ments in the dog are recent, and the most suitable device

in the dog varies from study to study.10,12 In the present

study, measuring TEWL with a closed-chamber device

showed that TEWL measurements were repeatable in a

group of dogs of the same breed, the same age, fed the

same diet and housed in the same controlled environ-

ment. Lau-Gillard et al.10 showed that TEWL measure-

ments varied from site to site, day to day and dog to dog.

Using an experimental canine model in which confound-

ing factors are controlled can limit these undesirable vari-

ations. The results of the present study contrast with

those of Lau-Gillard et al.10 concerning day-to-day varia-

tions, and significant variations were not observed. Wat-

son et al.11 showed that training dogs to stand still for the

duration of a measurement significantly reduced day-to-

day variations in TEWL readings. We used laboratory

dogs that were docile and accustomed to being handled.

In the present study, the increase of TEWL was

important immediately after TS and, despite the rela-

tively small number of dogs, the mean TEWL value

was significantly higher after TS than in the nonstripped

area, confirming the correlation between TEWL and bar-

rier function. This finding is in agreement with previous

reports in humans, mice28 and dogs.16,18 In murine

models, three phases of barrier recovery with distinct

metabolic activities occur after acute barrier disruption.

The first one is characterized by secretion of a pre-

formed pool of lamellar bodies and leads to a rapid

decrease of the TEWL value.29 The second phase is

defined by increased lipid synthesis and by accelerated

lamellar body formation and secretion,30,31 leading to a

slower decrease of TEWL value. During the third phase,

increased keratinocyte proliferation32 and differentiation

complete the skin barrier recovery and the return of the

TEWL value to normal.6

In the present study, TEWL evolution presented the

same type of pattern of recovery; a rapid decrease was

observed within 24 h of recovery in all of the dogs and

was followed by a slower decrease. Seventy-two hours

after TS, the TEWL value was still slightly higher than that

of undisrupted skin, but the difference was not signifi-

cant. In further studies in dogs, it would be interesting to

perform electron microscopy in order to study lipid

metabolism following barrier disruption and to compare it

with findings from humans and in mice. In the present

study, findings similar to those in mice with respect to a

difference between the TEWL value and reconstitution of

SC assessed by histological examination were observed.

A recovery of the TEWL value higher than 50% within

24 h was present in all the dogs even though corneocyte

layers had not yet reappeared. Seventy-two hours after

TS, the TEWL value was still slightly higher than that

of undisrupted skin even though a complete SC was

observed on histological examination. However, the

decrease of TEWL paralleled the global reappearance of

the SC and the restoration of skin barrier function. The

decrease of TEWL preceding the reappearance of SC

could be explained by the early release of the contents of

the lamellar bodies from the outer stratum granulosum

cells, leading to the rapid reconstitution of a water-

impermeable membrane.33

In humans, recovery of the skin barrier is influenced by

various parameters.34 In the present study, variations

were limited by using dogs of the same breed, age and

sex, housed in the same conditions and by choosing a dis-

tinct anatomical location. Variability between the dogs

within the recovery period following mechanical disrup-

tion was observed and was probably due to variability in

inflammatory response between the dogs. Despite the

existence of variations between the dogs, the barrier

recovery pattern based on TEWL measurement was the

same in all the dogs.

In conclusion, we have shown that an acute barrier dis-

ruption could be generated in healthy dogs by TS.

Mechanical disruption of the skin barrier induces a rapid

reconstitution of the SC and a marked eosinophilic infiltra-

tion followed by mononuclear cell infiltration into tape-

stripped canine skin. This very good correlation between

the degree of barrier disruption assessed clinically and

histologically, the TEWL and their parallel patterns of

recovery show that TEWL measurement is a valuable

noninvasive method to assess the barrier function of the

skin in the dog. This standardized, repeatable canine

model of acute skin barrier disruption and restoration

could help in elucidation of the pathogenesis of dermato-

ses with abnormal barrier function, such as atopic derma-

titis. This model could also be used to study treatment

modalities aimed at restoring skin barrier function. Even

though there was barrier disruption by TS in the present

study, the recovery was still very rapid, and the develop-

ment of a chronic model of skin barrier disruption would

be necessary to study chronic inflammatory dermatoses

and their treatment.

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Resume Le stratum corneum (SC) forme la partie principale de la barriere cutanee permeable. Chez

l’homme et la souris, une rupture de la barriere cutanee peut etre generee par retrait du SC a l’aide de cello-

phane adhesive (TS), et la fonction de barriere cutanee peut etre evaluee par mesure de la perte hydrique

transepidermique (TEWL). L’objectif de cette etude est de determiner, a la suite d’une rupture mecanique,

la restauration de la barriere cutanee chez le chien et d’analyser la correlation entre cette restauration et la

mesure du TEWL. Trente TS sequentielles ont ete realisees sur 12 sites sur quatre beagles sains. Le

nombre de TS a ete choisi pour permettre une rupture suffisante de la barriere cutanee avec une lente res-

tauration. La reparation cutanee a ete evaluee pendant 72 heures par des examens cliniques et histologi-

ques et des mesures de TEWL. Les resultats ont montre que de realiser 30 TS permettait de rompre la

barriere cutanee chez le chien. La reparation homeostatique, initiee dans la peau, etait rapide et caracteri-

see par une restauration complete du SC dans les 72 heures, accompagnee par une proliferation accrue

des cellules basales et une inflammation eosinophilique dermique. La TEWL etait significativement aug-

mentee avec un retrait complet du SC mais retournait a la normale avec la restauration du SC (Scheffe test,

P £ 0.05). La caracterisation d’un modele canin de rupture et de restauration de la fonction de barriere cuta-

nee par les mesures de TEWL pourrait permettre de mieux comprendre les evenements impliques dans la

fonction de barriere cutanee. Le developpement de ce modele canin est egalement necessaire pour de

futures etudes sur les effets des traitements sur la restauration de la barriere cutanee.




Resumen El estrato corneo (SC) forma la parte principal de la barrera de permeabilidad de la piel. En

ratones y en seres humanos, se puede crear una alteracion de la funcion de barrera cutanea quitando el SC

con cinta adhesiva (TS) y la funcion de la barrera de la piel se puede determinar por la medida de la perdida

de agua transepidermal (TEWL). El proposito de este estudio fue caracterizar la regeneracion de la funcion

de barrera de la piel en el perro tras la alteracion mecanica y analizar la correlacion entre la recuperacion de

la funcion de barrera de la piel y la medida de TEWL. Se realizaron treinta TS secuenciales en 12 sitios en

cuatro perros sanos de raza Beagle. El numero de TS fue elegido para asegurar una suficiente alteracion

mecanica de la barrera con una recuperacion lenta. La reparacion de la piel fue determinada a las 72 horas

mediante examenes clınicos e histologicos, y la medidas de TEWL. Los resultados demostraron que reali-

zar 30 TS fue adecuado para alterar la funcion de barrera la piel en el perro. La respuesta homeostatica de

reparacion, iniciada en la piel, fue rapida y caracterizada por la restauracion completa del SC en el plazo de

72 horas, acompanada por la mayor proliferacion de celulas basales, y la presencia de inflamacion eosinofi-

lica dermica. TEWL aumento perceptiblemente con la retirada del SC pero se recupero a la par con la res-

tauracion del SC (prueba de Scheffe, P £ 0.05). La caracterizacion de un modelo canino de alteracion y

restauracion de la barrera y la valoracion de la funcion de barrera de la piel midiendo TEWL podıa contribuir

a entender mejor los acontecimientos implicados en la funcion de la barrera de la piel. El desarrollo de este

modelo canino es tambien necesario para los estudios futuros en los efectos de los tratamientos dirigidos

a restaurar la barrera de la piel.

Zusammenfassung Das Stratum corneum (SC) stellt den wichtigsten Teil der Durchlassigkeitsbarriere

der Haut dar. Bei Mausen und Menschen kann eine Storung der Hautbarriere durch das Entfernen des SC

mittels Tape Stripping (TS) simuliert werden und die Funktion der Hautbarriere kann durch Messung des

transepidermalen Wasserverlustes (TEWL) beurteilt werden. Das Ziel der gegenwartigen Studie war es,

die Sanierung der Hautbarriere nach einer mechanischen Storung beim Hund zu beschreiben und die Korre-

lation zwischen der Erholung der Hautbarriere und der TEWL Messung zu analysieren. Es wurden dreißig

aufeinander folgende TS an 12 Hautstellen von vier gesunden Beagles durchgefuhrt. Die Anzahl der TS

wurde so ausgewahlt, um eine ausreichende Storung der Barriere mit einer langsamen Regenerierung zu

gewahrleisten. Die Wiederherstellung der Haut wurde uber 72 Stunden mittels klinischen und histolo-

gischen Untersuchungen, sowie mittels TEWL Messungen beurteilt. Die Ergebnisse zeigten, dass die

Durchfuhrung von 30 TS ausreichte, um die Hautbarriere beim Hund zu storen. Die Wiederherstellung der

Homoostase, die durch die Haut initiiert wurde, war schnell und durch eine komplette Reparatur der SC

innerhalb von 72 Stunden gekennzeichnet. Der Vorgang war von einer großeren Basalzellproliferation und

dermaler eosinophiler Entzundung begleitet. Der TEWL war durch die vollige Entfernung des SC signifikant

erhoht, erholte sich aber parallel zur Wiederherstellung des SC (Scheffe Test, P £ 0.05). Die Darstellung

eines Hundemodells fur die Storung der Hautbarriere, sowie deren Wiederherstellung und Beurteilung der

Hautbarrierenfunktion mittels TEWL Messungen konnte dabei helfen, die Vorgange, die mit der Funktion

der Hautbarriere zusammenhangen, besser zu verstehen. Die Entwicklung dieses Hundemodells ist auch

fur zukunftige Studien uber Auswirkungen von Therapien, die auf die Reparatur der Hautbarriere abzielen,

wichtig.

Videmont et al.


e23 ª 2011 ESVD and ACVD, Veterinary Dermatology, 23, 103–e23.

Documents

Characterization of the canine skin barrier restoration following acute disruption by tape stripping