Ultrastructural Pathology, 2014; 38(5): 344–357 ! Informa Healthcare USA, Inc. ISSN: 0191-3123 print / 1521-0758 online DOI: 10.3109/01913123.2014.911788
ORIGINAL ARTICLE
Immunolocalization of Androgen Receptor and Estrogen Receptors in Skin Tags* Ola Ahmed Bakry, MD1, Rehab Monir Samaka, MD2, Mohamed Abdel Moneim Shoeib, MD1, and Aziza Maher, MSc1 1
Department of Dermatology, Andrology and STDs and 2Department of Pathology, Faculty of Medicine, Menoufiya University, Menoufiya Governorate, Egypt
ABSTRACT Skin tags (STs) are benign connective tissue tumors of the dermis. Several clinical observations suggested the involvement of sex steroids in their development. This study aimed at investigating the possible role of androgen receptor (AR) and estrogen receptors (ERs) in STs pathogenesis through their immunohistochemical (IHC) localization in skin biopsies of this disease and to correlate their expression with different clinical and histopathological parameters. Using IHC techniques, we examined 62 cases with STs and 30 gender- and age-matched, healthy subjects, representing the control group. ERa, ERb, and AR were upregulated in STs compared to normal skin in epidermis and dermis (p5.001 for all). Higher AR H score was significantly associated with axillary STs (p = .02), skin colored tags (p = .03), acanthosis, and papillomatosis (p = .04 for both). Higher ERa H score was significantly associated with hyperpigmented tags (p5.001) and positive family history (p = .003). Higher ERb H score was significantly associated with female gender and obesity (p = .004 for both). Higher ERa and AR H scores were significantly associated with loose collagen arrangement (p = .02, p = .004, respectively). Higher AR, ERa, and ERb H scores were significantly associated with the presence of mast cells (p = .01, p = .04, p = .002, respectively) and dilated blood vessels (p = .006, p = .04, p = .04, respectively). In conclusion, AR and ERs may share in STs pathogenesis through their effect on keratinocytes, fibroblasts, and mast cells. Keywords: Skin tags, androgen receptor, estrogen receptors, pathogenesis
INTRODUCTION
and serves as a transcriptional factor to regulate the expression of its target genes. In addition to androgendependent activation, AR function can also be modulated by interaction with various co-regulators through their binding to the AR N-terminal domain or ligand-binding domain [3]. Estrogen receptors (ERs) are a group of proteins found inside cells. Two classes of ERs exist: ER, which is a member of the nuclear hormone family of intracellular receptors, and G protein-coupled ER, which is a member of the rhodopsin-like family of G protein-coupled receptors [4]. Once activated by estrogen, ER is able to translocate into the nucleus and bind to DNA to regulate the activity of different genes. However, it also has additional functions independent of DNA binding [5].
Skin tags (STs) are small, soft, common, benign, usually pedunculated neoplasm. It is usually skin colored or hyperpigmented, and it may appear as surface nodules or papillomas on healthy skin. Most STs vary in size from 2 to 5 mm in diameter, although larger lesions up to 5 cm in diameter are sometimes evident. They mainly consist of loose fibrous tissue and occur on the neck and axillae, but any skin fold, including the groin, may be affected [1]. Androgen receptor (AR) belongs to the nuclear receptor superfamily and contains a ligand-binding domain, a DNA-binding domain, and N-terminal regulatory domain [2]. Upon binding to its ligand, AR translocates from the cytosol into the nucleus
*This manuscript has not been published elsewhere and it has not been submitted simultaneously for publication elsewhere. Received 4 March 2014; Revised 22 March 2014; Accepted 31 March 2014; Published online 15 May 2014 Correspondence: Ola Ahmed Bakry, Department of Dermatology, Andrology and STDs, Menoufiya University, Shibeen El Koom, 32817 Menoufiya Governorate, Egypt. Tel: +20 1065190509. Fax: +20 482318574. E-mail: [email protected]
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AR and ERs in skin tags There are two different forms of ERs, usually referred to as a and b, each is encoded by a separate gene (ESR1 and ESR2, respectively). Hormoneactivated ERs form dimers, and, since the two forms are co-expressed in many cell types, the receptors may form ERa (aa) or ERb (bb) homodimers or ERab (ab) heterodimers [6]. AR is present in epidermal and follicular keratinocytes, sebocytes, sweat gland cells, dermal papilla cells, dermal fibroblasts, endothelial cells, and genital melanocytes [7]. ERa immunoreactivity was found in keratinocytes and sebaceous glands, particularly in partially differentiated sebocytes [8]. ERb was found to be highly expressed in the epidermis, sebaceous glands, hair follicles, eccrine sweat gland cells [9,10], and melanocytes [11]. Both AR and ERs are known to contribute to skin homeostasis and cellular proliferation [8,10]. However, their role in proliferative skin disorders is not extensively studied. In the current study, we aimed to investigate the possible role of AR and ERs in STs pathogenesis through their immunohistochemical (IHC) localization in skin biopsies of this disease entity. Furthermore, we aimed to correlate the expression of them with different clinical and histopathological parameters of studied cases.
MATERIALS AND METHODS Studied population This prospective case–control study was carried out on 92 subjects; 62 cases with STs and 30 age and gender-matched normal subjects as a control group. Cases were selected from the Dermatology Outpatient Clinic, Menoufiya University Hospital spanning the period from January 2011 to December 2012. Normal skin specimens were obtained from subjects attending the Plastic Surgery Department, Menoufiya University Hospital. A written consent from approved by The Committee of Human Rights in Research in Menoufiya University was obtained from every participant before the study initiation. This was in accordance with the Helsinki Declaration of 1975 (revised in 2000). All studied patients were subjected to: (1) Complete history taking, general and dermatological examination. (2) Body mass index (BMI) calculation: Values equal to or greater than 30 kg/m2 denoted obesity [12]. (3) Fasting blood glucose and postprandial blood glucose estimation: Evaluation was done according to guidelines of the American Diabetic Association [13]. !
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Biopsies Skin biopsy samples were taken under 2% lignocaine local anesthesia from every case and control subject. All specimens were fixed in 10% neutral-buffered formalin and subjected to routine tissue processing that ended with paraffin-embedded blocks.
Histopathological assessment Hematoxylin and eosin (H and E)-stained sections H- and E-stained sections were evaluated for detecting both epidermal and dermal histopathological changes. Giemsa-stained sections For mast cell count in 10 randomly selected high power fields. Mallory’s phosphotungstic and hematoxylin (PTAH)-stained sections For demonstration of mast cells and collagen bundles. IHC staining Five-micrometer-thick sections were cut from the paraffin-embedded blocks with subsequent steps of deparaffinization using xylene and then rehydrated in decreasing concentrations of ethanol. Antigen retrieval was performed by boiling in 10 mL citrate buffer (pH 6.0) for 20 min, followed by cooling at room temperature. The slides were incubated overnight at room temperature with antibodies demonstrated in Table 1. IHC staining was performed using the universal cytomation Labelled stretavidin–Biotin-2 system, Horseradish Peroxidase (LSAB-2 System, HRP Kit, Cat. # No. k0679; Dako, Copenhagen, Denmark). The reaction was visualized by an appropriate substrate/chromogen (diaminobenzidine) reagent with Mayer’s hematoxylin as a counter stain.
Interpretation of the immunostaining results Positive immunostaining for AR and ER was identified by brownish nuclear discoloration [14,15]. For each marker, the following tools of assessment were used in both epidermis and dermis: (1) Positivity: For AR: was done according to Marocchio et al. [16]. For ERs: was done according to the American Society of Clinical Oncology/College of American Pathologists guideline recommendations for IHC testing of ERs [17]. (2) Intensity: mild, moderate and strong.
346 O. A. Bakry et al. TABLE 1. AR and ERs antibodies used for IHC staining procedure. Antibody
Clonality
Company
AR
Mouse monoclonal antibody (Clone AR441)
ERa
Mouse monoclonal antibody (Clone 1D5)
Thermo Fisher Scientific Anatomical Pathology (Fremont, CA) Dako, Copenhagen, Denmark
ERb
Rabbit polyclonal antibody (Clone 1D5)
Thermo Fisher Scientific Anatomical Pathology (Fremont, CA)
(3) Distribution of epidermal immunostaining was assessed: Basal Suprabasal Whole thickness Histo-score (H-score) was applied to evaluate the positive cases, where both the intensity and the percentage of positivity were considered using the following formula: H score = +3 (strong intensity) % + 2 (moderate intensity) % + 1 (mild intensity) %. The score ranges between 0 and 300 [18].
Statistical analysis Data were statistically analyzed using a personal computer with ‘‘(SPSS) version 11’’ (SPSS Inc., Chicago, IL) program. Chi-square test was used to study the association between qualitative variables. Mann–Whitney test was used for comparison between quantitative variables. Student’s t test was used for comparison between two groups having quantitative variables. Pearson’s coefficient (r) was used to measure the correlation between two quantitative variables. Differences were considered statistically significant with p5.05.
RESULTS Clinical data of studied groups Cases included 14 males (22.6%) and 48 females (77.4%). Their ages ranged from 16 to 62 years. The lesions were located on abdomen (2 cases, 3.2%), axillae (14 cases, 22.6%), back (10 cases, 16.1%), and neck (36 cases, 58.1%). Lesions were multiple (44 cases, 71.0%) or single (18 cases, 29.0%). The color varied from skin colored (38 cases, 61.3%) to hyperpigmented (24 cases, 9.6%). Family history was positive in 14 cases (22.6%). Ten cases (16.1%) have diabetes and 36 cases (58.1%) were obese. Control group included 6 males (20%) and 24 females (80%). Their ages ranged from 18 to 60 years.
Code
Status
Dilution Positive control
Cat. # Code MS-443P0
0.1 mL concentrated 1:25
Prostatic carcinoma
Cat. # Code M7047, IR657 Cat. # Code RB-10658-P1
1 mL concentrated
1:50
Breast carcinoma
1 mL concentrated
1:50
Breast carcinoma
Histopathological findings of H and E, Giemsa- and PTAH-stained sections of STs cases Epidermal changes Acanthosis was present in 48 cases (77.4%). Hyperkeratosis was present in all cases (100%). Papillomatosis was present in 48 cases (77.4%). Dermal changes Dilated blood vessels were present in 44 cases (71%). Mast cells were present in 50 cases (80.6%) with mean ± SD of 5.1 ± 4.1 cells. Collagen arrangement was haphazard in 24 cases (38.7%) and loose in 38 cases (61.3%) (Figures 1 and 2).
AR expression in studied groups (A) Control group: Epidermal and dermal immunoreactivity were demonstrated in Table 2 and Figure 3. (B) STs group: Epidermal and dermal immunoreactivity were demonstrated in Table 2 and Figure 4.
Comparison of AR expression in studied cases and controls Cases showed significantly enhanced expression of AR as the whole thickness positivity was higher than controls (p5.001). Strong epidermal intensity of AR (p5.001), nuclear pattern (p5.001), and higher epidermal H score (p5.001) were significantly associated with cases compared to control group. Significant differences were noticed also regarding dermal intensity and dermal H-score (p5.001 for both) (Table 2).
ERs expression in studied groups I-ERa (A) Control group: Epidermal and dermal immunoreactivity were demonstrated in Table 3 and Figure 5. Ultrastructural Pathology
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FIGURE 1. Section of a skin tag showed scattered mast cells with deep violet color in dermis (circles). Insert: zooming view for mast cell with cytoplasmic purple granules (Giemsa stain 400 for A, B).
FIGURE 2. Section of a skin tag showed (A) mast cell (circle) and haphazardly arranged collagen bundles (H&E 400) (B) Scattered mast cells (circles) and haphazardly arranged collagen bundles with red brown discoloration. Insert: Zooming view for a mast cell (PTAH stain 400).
(B) STs group: Epidermal and dermal immunoreactivity were demonstrated in Table 3 and Figure 6.
Comparison of ERa expression in studied cases and controls There was significant difference between cases and controls regarding epidermal intensity of ERa !
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(p5.001), epidermal H score (p5.001) and epidermal localization (p = .003). A significant difference was found between cases and controls regarding dermal H-score (p5.001) and dermal intensity (p = .002) (Table 3). II-ERb: (A) Control group: Epidermal and dermal immunoreactivity were demonstrated in Table 3 and Figure 7.
348 O. A. Bakry et al. TABLE 2. AR expression in studied groups. Variable Epidermal expression Present Absent Epidermal localization Basal Suprabasal Whole thickness Epidermal intensity Mild Moderate Strong Epidermal pattern Nuclear Nucleo-cytoplasmic Epidermal H-score Mean ± SD Dermal expression Present Absent Dermal intensity Mild Moderate Strong Dermal pattern Nuclear Nucleo-cytoplasmic Dermal H-score Mean ± SD
STs group No = 62
Control group No = 30
No 40 22
% 64.5 35.5
No 21 9
% 70 30
0 2 38
0 5 95
8 0 13
38.1 0 61.9
12 20 8
30 50 20
17 4 0
80.9 19.1 0
34 6
85 15
0 21
(B) STs Group: Epidermal and dermal immunoreactivity were demonstrated in Table 3 and Figure 8. p Value .60
5.001**
129.0 ± 59.0
5.001**
0 100
5.001**
68.1 ± 10.4
5.001**
38 24
61.3 38.7
21 9
70 30
.41
26 4 8
68.4 10.5 21.1
20 1 0
95.2 4.8 0
.04*
36 2
94.7 5.3
18 3
85.7 14.3
.23
64.7 ± 37.1
34.8 ± 6.0
Note. *Significant and **highly significant.
5.001**
Comparison of ERb expression in studied cases and controls There was significant difference between cases and controls regarding epidermal intensity of ERb (p = .007), epidermal H score, epidermal pattern and epidermal localization (p5.001 for all). A significant difference was found between cases and controls regarding dermal intensity (p = .001) (Table 3).
Association between AR H scores and clinical and histopathological data of studied cases Higher epidermal AR H score was significantly associated with axillary STs (p = .02), hyperpigmented tags (p = .03), the presence of acanthosis, the presence of papillomatosis (p = .04 for both), and loose collagen arrangement (p5.001) (Table 4). Higher dermal AR H score was significantly associated with the presence of papillomatosis (p = .002), dilated blood vessels (p = .006), the presence of mast cells (p = .01), and loose collagen arrangement (p = .02) (Table 4).
FIGURE 3. Normal skin showing (A) moderate nuclear AR expression in the whole thickness of the epidermis, (B) mild nuclear AR expression of epidermal cells and sebocytes (C) moderate nucleo-cytoplasmic AR expression in the whole thickness of the epidermis, hair follicles and in few basal cells of sebaceous glands (IHC 100 for A, 200 for B and 400 for C). Ultrastructural Pathology
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FIGURE 4. A case of ST showing (A) moderate AR expression in the whole thickness of epidermis and mild expression in dermal stroma (IHC 100). (B) Higher power view showing moderate nuclear AR expression in the whole thickness of epidermis and mild nuclear expression in dermal stroma (IHC 200). (C) Strong nucleo-cytoplasmic AR expression in epidermis and mild nuclear expression in dermal stroma (IHC 400). (D) Mild nucleo-cytoplasmic AR expression in dermal stromal cells (IHC 100 for A, 200 for B and 400 for both C and D).
Association between ERa H scores and clinical and histopathological data of studied cases Higher epidermal ERa H score was significantly associated with hyperpigmented tags (p5.001), positive family history (p = .003), and loose collagen arrangement (p = .004). Higher dermal ER H score was significantly associated with positive family history (p = .04), the presence of dilated blood vessels and the presence of mast cells (p = .04 for both) (Table 5).
Association between ERb H scores and clinical and histopathological data of studied cases Higher epidermal ERb H score was significantly associated with female gender (p = .004). Higher dermal ERb H score was significantly associated with obesity (p = .004), the presence of mast cells (p = .002), and dilated blood vessels (p = .04) (Table 6).
Correlation between AR, ERa and ERb H scores in studied cases Significant positive correlation was noted between AR and ERa H scores and between AR and ERb H scores !
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in epidermis (r = .52, p = .04 and r = .4, p = .02, respectively) and in dermis (r = .35, p = .03, r = .37, p = .01 respectively) of studied cases (data not shown in tables).
DISCUSSION STs manifest exclusively in postpubertal life [19]. Some studies stated that STs predominate in women with two peaks: one in pregnancy and generally regresses in puerperium and the other peak occurs in menopause [20]. These phenomena raise attention of hormonal involvement in their pathogenesis [21]. The increased estrogen during pregnancy and the relative increase in androgen following menopause [22] may provide evidence about the sharing of these sex steroids in pathogenesis. In addition, STs are more common in obese males in whom estrogen levels in blood increase in a direct proportion with BMI [23]. AR and ERs are expressed in various types of skin cells [7 11]. However, their role in proliferative skin disorders is not extensively studied. In the present study, ERs were upregulated in STs compared with normal skin. This went with El Safoury et al. [24] and may provide evidence about the involvement of ERs in STs pathogenesis. This role was explained by Verdier-Sevrain et al. [25] on the cellular scale; keratinocytes express both ERs
350 O. A. Bakry et al. TABLE 3. ERs expression in studied groups. ERa expression
Variable Epidermal expression Present Absent Epidermal localization Basal Suprabasal Whole thickness Epidermal intensity Mild Moderate Strong Epidermal pattern Nuclear Nucleo-cytoplasmic Epidermal H-score Mean ± SD Dermal expression Present Absent Dermal intensity Mild Moderate Strong Dermal pattern Nuclear Nucleo-cytoplasmic Dermal H-score Mean ± SD
ERb expression
STs group No = 62
Control group No = 30
STs group No = 62
Control group No = 30
No (%)
No (%)
p Value
No (%)
No (%)
p Value
46 (74.2) 16 (25.8)
19 (63.3) 11 (36.7)
.28
62 (100) 0 (0)
30 (100) 0 (0)
–
8 (17.4) 6 (13.0) 32 (69.6)
11 (57.9) 0 (0) 8 (42.1)
.003**
0 (0) 0 (0) 62 (100)
9 (30) 0 (0) 21 (70)
5.001**
14 (30.4) 22 (47.8) 10 (21.8)
19 (100) 0 (0) 0 (0)
5.001**
18 (29) 38 (61.3) 6 (9.7)
19 (63.3) 11 (36.7) 0 (0)
.007**
44 (95.7) 2 (4.3)
15 (100) 0 (0)
.41
24 (38.7) 38 (61.3)
0 (0) 30 (100)
5.001**
123.0 ± 51.6
51.6 ± 6.5
70.8 ± 29.5
5.001**
5.001**
143.3 ± 38.6
38 (61.3) 24 (38.7)
19 (63.3) 11 (36.7)
.85
62 (100) 0 (0)
30 (100) 0 (0)
–
21 (55.3) 13 (34.2) 4 (10.5)
19 (100) 0 (0) 0 (0)
.002**
38 (61.3) 18 (29) 6 (9.7)
26 (86.7) 4 (13.3) 0 (0)
.001**
35 (92.1) 3 (7.9)
16 (84.2) 3 (15.8)
.36
35 (56.4) 27 (43.6)
26 (86.7) 4 (13.3)
.36
5.001**
66.0 ± 24.4
57.2 ± 11.9
.06
66.1 ± 28.2
45.0 ± 6.7
Note. **Highly significant.
(ERa and ERb). Estradiol binds to keratinocytes’ ERa and ERb with high affinity and in its normal physiological range induces keratinocyte proliferation. In addition, 17b-estradiol has been shown to increase fibroblast proliferation in human skin [26]. In the current study, AR was upregulated in STs compared with normal skin. This went with EL safoury et al. [24]. Choudhry et al. [27] postulated that, although keratinocytes are known to express AR and are target cells for androgen action, the effect of androgens on keratinocytes’ activity remains to be clarified. Pelletier and Ren [10] concluded that, the presence of AR-expressing keratinocytes within the pilosebaceous ducts supported the hypothesis that androgens can directly influence direct keratinization occurring in acne process. In addition, Tadokoro et al. [28] stated that AR can modulate proliferation and differentiation of epithelial cells of the epidermis and adnexa, functional activity of dermal fibroblasts and fibrocytes. The significant association between higher epidermal AR H score and the presence of acanthosis and papillomatosis in studied cases may support the role of AR in epithelial proliferation and STs’ pathogenesis. Further studies at the molecular level and on
larger scale of cases are warranted for firmer conclusion. The upregulation of AR and ERs in STs and their possible role in pathogenesis went with the conclusion of Thornton et al. [8] that tissues traditionally thought to be responsive to one class of steroids, contain receptors for other classes, and there is growing evidence that steroid receptors can crosstalk with one another. This was underscored by the significant positive correlation between AR and ERs H scores in epidermis and dermis, detected in the current work. In the current study, higher dermal AR and ERs H scores was significantly associated with the presence of mast cells. Similar observation was reported by Zaher et al. [29]. Mast cells express high-affinity ERs and 17b-estradiol augments the secretion of histamine and serotonin [30]. Binding of estradiol to ERa, initiates a rapid onset, progressive influx of extracellular Ca2, inducing exocytosis of mast cell products [31]. Mast cell mediators are capable of inducing keratinocyte and fibroblast proliferation with epidermal hyperplasia and increased collagen synthesis, contributing to the main pathologic abnormalities observed in STs [29]. Therefore, a question arises; Ultrastructural Pathology
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FIGURE 5. Expression of ERa in normal skin (A) moderate nucleo-cytoplasmic expression in the whole thickness of the epidermis, hair follicle, few stromal cells and blood vessel endothelium, (B) mild nucleo-cytoplasmic expression in basal and differentiated cells of sebaceous glands (C) mild to moderate nuclear expression in blood vessel endothelium and stromal cells (IHC 200 for A and 400 for B, C).
FIGURE 6. Expression of ERa in STs showing (A) strong nuclo-cytoplasmic expression in the whole thickness of epidermal and moderate nuclo-cytoplasmic expression in dermal stroma, (B) moderate nuclo-cytoplasmic expression in the whole thickness of epidermal and (C) moderate nuclo-cytoplasmic expression in suprabasal layers of epidermal, strong nuclear expression in basal cell layer and strong nuclo-cytoplasmic in dermal stromal cells (H&E 200 for A and 400 for B and C).
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FIGURE 7. Expression of ERb in normal skin showing (A) moderate nucleo-cytoplasmic expression in the whole thickness of the epidermis, few stromal cells and blood vessels, (B) mild to moderate nucleo-cytoplasmic expression in outer and inner root sheath of hair follicles and fibroblastic stromal cells (C) mild to moderate nuclear expression in blood vessels and stromal cells in between (D) mild nucleo-cytoplasmic expression in basal and differentiated cells of sebaceous glands (IHC 100 for A, 200 for B and 400 for C, D).
FIGURE 8. Expression of ERb in STs (A) panoramic view exhibiting strong nucleo-cytoplasmic expression in the whole thickness of epidermis and mild nucleo-cytoplasmic expression in scattered dermal stromal cells and blood vessels (B) closer view of the previous figure illustrating epidermal and dermal expression (C) mild nucleo-cytoplasmic expression in dermal stromal cells and blood vessels (IHC 100 for A, 200 for B and 400 for C).
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TABLE 4. Association between AR H scores and clinical and histopathological data of studied cases (values were expressed in mean ± SD).
Clinical variables Gender Male Female Site Abdomen Axilla Back Neck Obesity Present Absent Diabetes mellitus Present Absent Family history Positive Negative Color Skin colored Hyperpigmented Histopathological variables Acanthosis Present Absent Papillomatosis Present Absent Dilated blood vessels Present Absent Mast cells Present Absent Collagen arrangement Haphazard Loose
Epidermal H score
p Value
Dermal H score
p Value
162.5 ± 63.8 120.6 ± 55.7
.07
72.0 ± 31.55 62.14 ± 39.09
.48
140.0 ± 0.0 185.0 ± 40.4 107.5 ± 30.6 115.5 ± 63.6
.02*
70.0 ± 0.0 85.0 ± 46.9 50.0 ± 17.9 60.9 ± 37.2
.32
135.6 ± 60.0 123.6 ± 59.1
.48
67.5 ± 38.2 62.7 ± 37.1
.79
150 ± 45.7 122 ± 61.9
.11
.15
103.3 ± 53.9 133.5 ± 59.5
.36
40.0 ± 11.5 67.7 ± 38.1 62.3 ± 36.5 70.0 ± 39.5
135.0 ± 5.8 112.3 ± 53.1
.03*
50.0 ± 17.9 67.5 ± 39.3
.42
140.0 ± 60.1 96.0 ± 42.9
.04*
72.5 ± 39.4 51.4 ± 29.6
.09
138.1 ± 58.6 92.5 ± 48.0
.04*
75.7 ± 35.6 34.0 ± 20.7
.002**
133.8 ± 50.3 110.0 ± 87.8
.12
74.3 ± 38.6 38.0 ± 10.3
.006**
136.7 ± 65.9 127.7 ± 58.7
.94
73.6 ± 38.2 40.0 ± 18.9
.01*
90.0 ± 61.0 193.3 ± 45.9
5.001**
46.7 ± 32.3 96.7 ± 37.2
.02*
.53
Note. *Significant and **highly significant.
could antihistamines and mast cell stabilizers be beneficial in treatment and/or prevention of STs, associated with large number of mast cells? The answer requires future clinical trials. Mast cells stimulated by friction can start STs formation through its interaction with fibroblasts and keratinocytes. However, the finding that STs are not a common association in mastocytosis indicates that mast cells without a precipitating factor can not initiate STs [32]. Contrary to our findings, Di Matteo et al. [30] found that blocking AR with sex hormone antagonists in the testis of frog causes a significant increase in mast cell count. In addition, it was found that testosterone has an inhibitory effect on mast cell secretion of histamine and serotonin in rats [33]. Further investigation is needed to prove or deny current observation. In the current study, higher ERa and AR H scores were significantly associated with cases showing loosely arranged collagen fibers. This finding was !
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not discussed before in similar studies. Estrogen targets human skin fibroblasts and has been shown to increase fibroblast proliferation in human skin [26]. However, the association between ERa and AR upregulation and loose collagen arrangement requires further investigation. In the current study, higher epidermal ERa H score was significantly associated with positive family history of STs in studied cases. Similar observation was reported before in breast cancer cases and was explained on genetic basis [34,35]. Whether this is applicable for STs or not, requires further studies on larger scales including cases and their relatives for firmer conclusion. In the current work, higher epidermal AR H scores were significantly associated with axillary STs; a finding that was not reported before. Whether this finding denotes regional differences in AR density in different body sites or not requires further research. STs are related to obesity [36], and a hormonal mechanism has been suggested in obese females,
354 O. A. Bakry et al. TABLE 5. Association between epidermal ERa H scores and the clinical and histopathological data of ST cases (values were expressed in mean ± SD).
Clinical variables Gender Male Female Site Abdomen Axilla Back Neck Obesity Present Absent Diabetes mellitus Present Absent Family history Positive Negative Color Skin colored Hyperpigmented Histopathological variables Acanthosis Present Absent Papillomatosis Present Absent Dilated blood vessels Present Absent Mast cells Present Absent Collagen arrangement Haphazard Loose
Epidermal H score
p Value
Dermal H score
p Value
120.0 ± 42.6 124.1 ± 54.3
.81
58.0 ± 33.59 68.9 ± 26.2
.30
180.0 ± 0.0 136.7 ± 45.0 100 ± 32.2 121.3 ± 54.2
.25
80.0 ± 0.0 78.8 ± 8.3 61.7 ± 7.5 61.4 ± 35.5
.43
135.0 ± 74.6 120.5 ± 45.6
.47
45.0 ± 17.3 68.5 ± 28.4
.09
125.7 ± 58.8 118.9 ± 37.1
.79
68.2 ± 34.6 63.1 ± 16.6
.91
134.4 ± 50.4 82.0 ± 27.8
.003**
69.7 ± 28.9 46.7 ± 13.7
.04*
150.0 ± 46.2 168.3 ± 55.1
5.001**
60.0 ± 20.9 62.5 ± 9.6
.11
127.8 ± 50.9 106 ± 50.6
.24
66.25 ± 26.34 65.71 ± 32.28
.96
123.7 ± 52.1 120.0 ± 48.7
.91
66.8 ± 22.8 64.0 ± 41.0
.35
128.1 ± 54.4 111.4 ± 42.0
.44
69.3 ± 20.2 57 ± 43.9
.04*
127.4 ± 54.2 102.5 ± 25.5
.41
71.1 ± 29.7 52.0 ± 18.1
.04*
66 ± 19.6 99.2 ± 35.0
.004**
78 ± 27.8 86.7 ± 50.9
.82
Note. *Significant and **highly significant.
where there is increased peripheral aromatization of androgens to estrogens [37]. Similar hormonal imbalance exists in obese males with hyperestrenemia [23] as obese men have elevated levels of estrone, both free and total, as well as estradiol [38]. In the current study, higher ERb H score was significantly associated with obese cases. However, no significant difference in H score values of ERa and AR between obese and non obese cases was noted. This difference in receptor immunoreactivity between ERb on one hand and ERa and AR on the other hand regarding relation to obesity, may be explained on basis of ERb predominance in human skin more than ERa [8,39]. However, further studies are warranted. El Safory et al. [24] did not find an association between AR and ERs expression and BMI of their examined cases. Further studies including larger number of cases with different BMI are needed. STs were reported to have a probable association with diabetes mellitus where they may be a skin
marker for underlying impaired carbohydrate metabolism and patients with STs should be evaluated for the presence of diabetes mellitus [40,41]. In the present work, no significant difference in ERs and AR H scores was found between diabetic and non diabetic cases. Testosterone levels were reported to be lower in diabetic men [42]. However; the effect of diabetes on estrogen, AR or ERs was not discussed before. In the present work, ERa, ERb immunoreactivity was detected in vascular endothelium both in normal skin and STs but AR immunoreactivity in vascular endothelium was positive in STs and negative in normal skin sections (as demonstrated in histopathological figures). Pelletier and Ren [10] reported positive endothelial immunoreactivity of ERb and negative immunoreactivity of AR and ERa in their study on normal skin specimens. More studies involving different body sites are recommended to prove or deny current observations. In addition, the current work showed that higher ERs and AR H Ultrastructural Pathology
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TABLE 6. Association between ERb H scores and the clinical and histopathological data of ST cases (values were expressed in mean ± SD). Variable Clinical variables Gender Male Female Site Abdomen Axilla Back Neck Obesity Present Absent Diabetes mellitus Present Absent Family history Positive Negative Color Skin colored Hyperpigmented Histopathological variables Acanthosis Present Absent Papillomatosis Present Absent Dilated blood vessels Present Absent Mast cell Present Absent Collagen arrangement Haphazard Loose
Epidermal H score
p Value
Dermal H score
p Value
136.3 ± 28.6 171.7 ± 58.3
.004**
62.9 ± 23.6 78.3 ± 24.4
.40
170.0 ± 0.0 132.9 ± 28.7 157.5 ± 68.6 142.8 ± 33.6
.39
90.0 ± 0.0 58.6 ± 19.6 67.5 ± 33.3 143.3 ± 38.6
.35
163.8 ± 37.6 116.5 ± 18.1
.31
73.8 ± 22.7 55.8 ± 23.0
.004**
167.5 ± 33.3 139.6 ± 38.2
.06
77.5 ± 13.9 64.2 ± 25.2
.15
140.0 ± 13.6 144.4 ± 43.5
.56
71.4 ± 10.3 64.4 ± 27.1
.15
143.8 ± 51.6 143.2 ± 33.3
.96
56.3 ± 26.3 69.6 ± 22.9
.06
146.7 ± 41.4 130.0 ± 20.0
.05
68.3 ± 25.7 56.7 ± 15.6
.14
146.8 ± 40.4 126.0 ± 21.7
.12
67.2 ± 25.8 60.0 ± 14.9
.24
146.9 ± 47.6 140.6 ± 30.4
.55
73.9 ± 24.8 60.0 ± 22.6
.04*
143.6 ± 43.6 142.5 ± 19.8
.89
71.4 ± 23.1 51.3 ± 22.2
.002**
120.0 ± 0.0 144.1 ± 38.9
.39
50.0 ± 0.0 66.6 ± 24.6
0.28
Note. *Significant and **highly significant.
scores were significantly associated with cases showing dilated blood vessels. Estrogen is known to have a vasodilator effect on vessel wall through the release of prostacyclin and nitric oxide by a non-genomic action [43]. The effect of androgen on vessel wall is poorly investigated with few studies reported vasodilator effect [44,45]. Further research is needed for more clarification of androgens and AR vascular role. In the current work, higher ERa H score was significantly associated with cases having hyperpigmented lesions. In vitro studies have shown that cultured human melanocytes express ERa and ERb, and estradiol increases the levels of tyrosinase, tyrosinase-related-protein 1 and tyrosinase-relatedprotein 2, the enzymes involved in human eumelanogenesis within normal human melanocytes [46,47]. However, the association between lower AR H score and hyperpigmented lesions, detected in the current work, remains unexplained. ERs and AR are known to exhibit nuclear immunoreactivity [14,15]. In our study, nucleo-cytoplasmic !
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pattern of both ERs and AR was detected in examined cases and control subjects. Fu and Simoncini [48] suggested that androgens and other sex steroid hormones may affect cells through mechanisms that do not involve gene transcription/expression, but through activation of cytoplasmic signaling pathways that affect cell survival and/or modulate other growth factor signaling [49]. Parker [50] concluded that ERs can move between the nucleus and the cytoplasm, but under normal conditions they are predominantly located in the nucleus. Pelletier and Ren [10] detected nucleo-cytoplasmic ERb immunoreactivity in ducts of eccrine sweat glands. Heemers and Tindall [2] concluded that, AR, in its inactive form, is located in the cytoplasm of the cell in complex with heat-shock proteins. Upon ligand binding, AR homodimerizes and translocates to the nucleus where it binds to the specific sequences on the DNA known as androgen response elements. It may also recruit cofactors that either enhance or reduce AR transactivation [51]. In the current study, there was
356 O. A. Bakry et al. significant association between nuclear pattern of AR and ERb with STs when compared to control. In addition, nuclear pattern appears more in STs regarding dermal AR and ERa although the difference between cases and controls was insignificant. These observations may provide evidence about the role of AR and ERs in STs pathogenesis through their presence in their active form. In conclusion, AR and ERs may share in STs pathogenesis through their effects on keratinocytes, fibroblasts and mast cells. This may open new avenue for treatment and prevention in susceptible individuals. Further studies on larger scales are warranted to investigate their exact molecular mechanisms in tag development.
DECLARATION OF INTEREST The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper. Samples were taken after obtaining written consent from the patients and control subjects.
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12. Tsai MC, Chen W, Cheng Y, et al. Higher body mass index is a significant risk factor for acne formation in schoolchildren. Eur J Dermatol 2006;163: 251–3. 13. American Diabetes Association. Report of the expert committee on the diagnosis and classification of diabetes mellitus. Diabetes Care 1997;20: 1183–97. 14. Hibberts NA, Howell AE, Randall VA. Balding hair follicle dermal papilla cells contain higher levels of androgen receptors than those from non-balding scalp. J Endocrinol 1998;133: 141–7. 15. Taylor Ah, Al-Azzawi F. Immunolocalization of estrogen receptor beta in human tissues. J Mol Endocrinol 2000;24: 145–55. 16. Marocchio LS, Giudice F, Correˆa L, et al. Estrogens and androgen receptors in oral squamous cell carcinoma. Acta Odontol Scand 2013;71: 1513–19. 17. Hammond ME, Hayes DF, Dowsett M, et al. American Society of Clinical Oncology/College of American Pathologists Guideline Recommendations for immunohistochemical testing of estrogen and progesterone receptors in breast cancer. Arch Pathol Lab Med 2010;134: 907–22. 18. Smyth J, Gourley C, Walker G. Antiestrogen therapy is active in selected ovarian cancer cases: The use of letrozole in estrogen receptor – Positive patients. Clin Cancer Res 2007;13: 3617–22. 19. Banik R, Lubach D. Skin tags. Localization and frequencies according to sex and age. Dermatologica 1987;174: 180–3. 20. Graham-Brown RAC. Skin tags. In Rook A, Wilkinson DS, Ebling FJ, eds. Text book of dermatology. London, UK: Oxford Blackwell Scientific Publications, 2004:701–30. 21. Errickson CV, Matus NR. Skin disorders of pregnancy. Am Fam Physician 1994;49: 605–10. 22. Tchernof A, Despre´s JP. Sex steroid hormones, sex hormone-binding globulin, and obesity in men and women. Horm Metab Res 2000;32: 526–36. 23. Be´langer C, Hould FS, Lebel S, et al. Omental and subcutaneous adipose tissue steroid levels in obese men. Steroids 2006;71: 674–82. 24. El Safoury O, Rashid L, Ibrahim M. A study of androgen and estrogen receptors a, b in skin tags. Indian J Dermatol 2010;55: 20–4. 25. Verdier-Sevrain S, Yaar M, Cantatore J, et al. Estradiol induces proliferation of keratinocytes via receptormediated mechanisms. FASEB J 2004;18: 1252–4. 26. Tomaszewski J, Adamiak A, Skorupski P, et al. Effect of 17 aˆ estradiol and phytoestrogen daidzein on the proliferation of pubocervical fascia and skin fibroblasts derived from women suffering from stress urinary incontinence. Ginekol Pol 2003;74: 1410–14. 27. Choudhry R, Hodgins MB, Van der Kwast TK, et al. Localization of androgen receptors in human skin by immunohistochemistry: Implications for the hormonal regulation of hair growth, sebaceous glands and sweet glands. J Endocrinol 1992;133: 467–75. 28. Tadokoro T, Itami S, Hosokawa K, et al. Human genital melanocytes as androgen target cells. J Invest Dermatol 1997;109: 513–17. 29. Zaher H, El Safoury OS, El Komy MM, et al. Study of mast cell count in skin tags. Indian J Dermatol 2007;52: 184–7. 30. Di Matteo L, Vitieello II, Minucci S. Effect of sex steroid hormones and antagonists on mast cell number in the testis of frog. Rana esculenta Zygot 2000;8: 225–34. 31. Zaitsu M, Narita S, Lambert KC, et al. Estradiol activates mast cells via a non-genomic estrogen receptor-a and calcium influx. Mol Immunol 2007;44: 1977–85. 32. El Safoury O, Fawzi M, Abdel Hay RM, et al. The possible role of trauma in skin tags through release of mast cell mediators. Indian J Dermatol 2011;56: 641–6. Ultrastructural Pathology
AR and ERs in skin tags 33. Vliagoftis H, Dimitriadou V, Boucher W, et al. Estradiol augments while tamoxifen inhibits rat mast cell secretion. Int Arch Allergy Immunol 1992;98: 398–409. 34. Zuppan P, Hall JM, Lee MK, et al. Possible linkage of the estrogen receptor gene to breast cancer in a family with late onset disease. Am J Hum Genet 1991;48: 1065–8. 35. Thomas A, Lawrence H, John D, et al. Effect of family history, body-fat distribution, and reproductive factors on the risk of postmenopausal breast cancer. N Engl J Med 1992;326: 1323–9. 36. Bhargava P, Mathur SK, Mathur DK, et al. Acrochordon, diabetes and associations. Indian J Dermal Veneriol Leprol 1996;62: 226–8. 37. Diamanti-Kandarakis E, Bergiele A. The influence of obesity on hyperandrogenism and infertility in the female. Obes Rev 2001;2: 231–7. 38. Zumoff B. Hormonal abnormalities in obesity. Acta Med Scand 1988;723: 153–60. 39. Pelletier G. Localization of androgen and estrogen receptors in rat and primate tissues. Histol Histopathol 2000;15: 1261–70. 40. Demir S, Demir Y. Acrochordon and impaired carbohydrate metabolism. Acta Diabetol 2002;39: 57–9. 41. Rasi A, Soltani-Arabshahi R, Shahbazi N. Skin tag as a cutaneous marker for impaired carbohydrate metabolism: A case–control study. In J Dermatol 2007;46: 1155–9.
42. Allan CA. Sex steroids and glucose metabolism. Asian J Androl 2014;16: 232–8. 43. Mendelsohn ME, Karas RH. Estrogen and the blood vessel wall. Curr Opin Cardiol 1994;9: 619–26. 44. Yildiz O, Seyrek M. Vasodilating mechanisms of testosterone. Exp Clin Endocrinol Diabetes 2007;115: 1–6. 45. Lu Y, Fu Y, Ge Y, et al. The vasodilatory effect of testosterone on renal afferent arterioles. Gend Med 2012;9: 103–11. 46. Im S, Eun-So L, Kim W, et al. Donor specific response of estrogen and progesterone on cultured human melanocytes. Korean Med Sci 2002;17: 58–64. 47. Kippenherger S, Loitsch S, Solano F, et al. Quantification of tyrosinase, TRP-1 and TRP-2 transcripts in human melanocytes by reverse transcriptase-competitive multiplex PCR-regulation by steroid hormones. J Invest Dermatol 1998;110: 364–7. 48. Fu XD, Simoncini T. Extra-nuclear signaling of estrogen receptors. IUBMB Life 2008;60: 502–10. 49. Simoncini T, Genazzani AR. Non-genomic actions of sex steroid hormones. Eur J Endocrinol 2003;148: 281–92. 50. Parker MG. Structure and function of estrogen receptors. Vitam Horm 1995;51: 267–87. 51. Popovscaia M, Baumgartner C, Handler M, et al. Identifying biologically meaningful groups of genes in the androgen receptor network. Biomed Technol 2013 Sep 7. [Epub ahead of print]. doi: 10.1515/bmt-2013-4307.
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ORIGINAL ARTICLE
Immunolocalization of Androgen Receptor and Estrogen Receptors in Skin Tags* Ola Ahmed Bakry, MD1, Rehab Monir Samaka, MD2, Mohamed Abdel Moneim Shoeib, MD1, and Aziza Maher, MSc1 1
Department of Dermatology, Andrology and STDs and 2Department of Pathology, Faculty of Medicine, Menoufiya University, Menoufiya Governorate, Egypt
ABSTRACT Skin tags (STs) are benign connective tissue tumors of the dermis. Several clinical observations suggested the involvement of sex steroids in their development. This study aimed at investigating the possible role of androgen receptor (AR) and estrogen receptors (ERs) in STs pathogenesis through their immunohistochemical (IHC) localization in skin biopsies of this disease and to correlate their expression with different clinical and histopathological parameters. Using IHC techniques, we examined 62 cases with STs and 30 gender- and age-matched, healthy subjects, representing the control group. ERa, ERb, and AR were upregulated in STs compared to normal skin in epidermis and dermis (p5.001 for all). Higher AR H score was significantly associated with axillary STs (p = .02), skin colored tags (p = .03), acanthosis, and papillomatosis (p = .04 for both). Higher ERa H score was significantly associated with hyperpigmented tags (p5.001) and positive family history (p = .003). Higher ERb H score was significantly associated with female gender and obesity (p = .004 for both). Higher ERa and AR H scores were significantly associated with loose collagen arrangement (p = .02, p = .004, respectively). Higher AR, ERa, and ERb H scores were significantly associated with the presence of mast cells (p = .01, p = .04, p = .002, respectively) and dilated blood vessels (p = .006, p = .04, p = .04, respectively). In conclusion, AR and ERs may share in STs pathogenesis through their effect on keratinocytes, fibroblasts, and mast cells. Keywords: Skin tags, androgen receptor, estrogen receptors, pathogenesis
INTRODUCTION
and serves as a transcriptional factor to regulate the expression of its target genes. In addition to androgendependent activation, AR function can also be modulated by interaction with various co-regulators through their binding to the AR N-terminal domain or ligand-binding domain [3]. Estrogen receptors (ERs) are a group of proteins found inside cells. Two classes of ERs exist: ER, which is a member of the nuclear hormone family of intracellular receptors, and G protein-coupled ER, which is a member of the rhodopsin-like family of G protein-coupled receptors [4]. Once activated by estrogen, ER is able to translocate into the nucleus and bind to DNA to regulate the activity of different genes. However, it also has additional functions independent of DNA binding [5].
Skin tags (STs) are small, soft, common, benign, usually pedunculated neoplasm. It is usually skin colored or hyperpigmented, and it may appear as surface nodules or papillomas on healthy skin. Most STs vary in size from 2 to 5 mm in diameter, although larger lesions up to 5 cm in diameter are sometimes evident. They mainly consist of loose fibrous tissue and occur on the neck and axillae, but any skin fold, including the groin, may be affected [1]. Androgen receptor (AR) belongs to the nuclear receptor superfamily and contains a ligand-binding domain, a DNA-binding domain, and N-terminal regulatory domain [2]. Upon binding to its ligand, AR translocates from the cytosol into the nucleus
*This manuscript has not been published elsewhere and it has not been submitted simultaneously for publication elsewhere. Received 4 March 2014; Revised 22 March 2014; Accepted 31 March 2014; Published online 15 May 2014 Correspondence: Ola Ahmed Bakry, Department of Dermatology, Andrology and STDs, Menoufiya University, Shibeen El Koom, 32817 Menoufiya Governorate, Egypt. Tel: +20 1065190509. Fax: +20 482318574. E-mail: [email protected]
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AR and ERs in skin tags There are two different forms of ERs, usually referred to as a and b, each is encoded by a separate gene (ESR1 and ESR2, respectively). Hormoneactivated ERs form dimers, and, since the two forms are co-expressed in many cell types, the receptors may form ERa (aa) or ERb (bb) homodimers or ERab (ab) heterodimers [6]. AR is present in epidermal and follicular keratinocytes, sebocytes, sweat gland cells, dermal papilla cells, dermal fibroblasts, endothelial cells, and genital melanocytes [7]. ERa immunoreactivity was found in keratinocytes and sebaceous glands, particularly in partially differentiated sebocytes [8]. ERb was found to be highly expressed in the epidermis, sebaceous glands, hair follicles, eccrine sweat gland cells [9,10], and melanocytes [11]. Both AR and ERs are known to contribute to skin homeostasis and cellular proliferation [8,10]. However, their role in proliferative skin disorders is not extensively studied. In the current study, we aimed to investigate the possible role of AR and ERs in STs pathogenesis through their immunohistochemical (IHC) localization in skin biopsies of this disease entity. Furthermore, we aimed to correlate the expression of them with different clinical and histopathological parameters of studied cases.
MATERIALS AND METHODS Studied population This prospective case–control study was carried out on 92 subjects; 62 cases with STs and 30 age and gender-matched normal subjects as a control group. Cases were selected from the Dermatology Outpatient Clinic, Menoufiya University Hospital spanning the period from January 2011 to December 2012. Normal skin specimens were obtained from subjects attending the Plastic Surgery Department, Menoufiya University Hospital. A written consent from approved by The Committee of Human Rights in Research in Menoufiya University was obtained from every participant before the study initiation. This was in accordance with the Helsinki Declaration of 1975 (revised in 2000). All studied patients were subjected to: (1) Complete history taking, general and dermatological examination. (2) Body mass index (BMI) calculation: Values equal to or greater than 30 kg/m2 denoted obesity [12]. (3) Fasting blood glucose and postprandial blood glucose estimation: Evaluation was done according to guidelines of the American Diabetic Association [13]. !
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Biopsies Skin biopsy samples were taken under 2% lignocaine local anesthesia from every case and control subject. All specimens were fixed in 10% neutral-buffered formalin and subjected to routine tissue processing that ended with paraffin-embedded blocks.
Histopathological assessment Hematoxylin and eosin (H and E)-stained sections H- and E-stained sections were evaluated for detecting both epidermal and dermal histopathological changes. Giemsa-stained sections For mast cell count in 10 randomly selected high power fields. Mallory’s phosphotungstic and hematoxylin (PTAH)-stained sections For demonstration of mast cells and collagen bundles. IHC staining Five-micrometer-thick sections were cut from the paraffin-embedded blocks with subsequent steps of deparaffinization using xylene and then rehydrated in decreasing concentrations of ethanol. Antigen retrieval was performed by boiling in 10 mL citrate buffer (pH 6.0) for 20 min, followed by cooling at room temperature. The slides were incubated overnight at room temperature with antibodies demonstrated in Table 1. IHC staining was performed using the universal cytomation Labelled stretavidin–Biotin-2 system, Horseradish Peroxidase (LSAB-2 System, HRP Kit, Cat. # No. k0679; Dako, Copenhagen, Denmark). The reaction was visualized by an appropriate substrate/chromogen (diaminobenzidine) reagent with Mayer’s hematoxylin as a counter stain.
Interpretation of the immunostaining results Positive immunostaining for AR and ER was identified by brownish nuclear discoloration [14,15]. For each marker, the following tools of assessment were used in both epidermis and dermis: (1) Positivity: For AR: was done according to Marocchio et al. [16]. For ERs: was done according to the American Society of Clinical Oncology/College of American Pathologists guideline recommendations for IHC testing of ERs [17]. (2) Intensity: mild, moderate and strong.
346 O. A. Bakry et al. TABLE 1. AR and ERs antibodies used for IHC staining procedure. Antibody
Clonality
Company
AR
Mouse monoclonal antibody (Clone AR441)
ERa
Mouse monoclonal antibody (Clone 1D5)
Thermo Fisher Scientific Anatomical Pathology (Fremont, CA) Dako, Copenhagen, Denmark
ERb
Rabbit polyclonal antibody (Clone 1D5)
Thermo Fisher Scientific Anatomical Pathology (Fremont, CA)
(3) Distribution of epidermal immunostaining was assessed: Basal Suprabasal Whole thickness Histo-score (H-score) was applied to evaluate the positive cases, where both the intensity and the percentage of positivity were considered using the following formula: H score = +3 (strong intensity) % + 2 (moderate intensity) % + 1 (mild intensity) %. The score ranges between 0 and 300 [18].
Statistical analysis Data were statistically analyzed using a personal computer with ‘‘(SPSS) version 11’’ (SPSS Inc., Chicago, IL) program. Chi-square test was used to study the association between qualitative variables. Mann–Whitney test was used for comparison between quantitative variables. Student’s t test was used for comparison between two groups having quantitative variables. Pearson’s coefficient (r) was used to measure the correlation between two quantitative variables. Differences were considered statistically significant with p5.05.
RESULTS Clinical data of studied groups Cases included 14 males (22.6%) and 48 females (77.4%). Their ages ranged from 16 to 62 years. The lesions were located on abdomen (2 cases, 3.2%), axillae (14 cases, 22.6%), back (10 cases, 16.1%), and neck (36 cases, 58.1%). Lesions were multiple (44 cases, 71.0%) or single (18 cases, 29.0%). The color varied from skin colored (38 cases, 61.3%) to hyperpigmented (24 cases, 9.6%). Family history was positive in 14 cases (22.6%). Ten cases (16.1%) have diabetes and 36 cases (58.1%) were obese. Control group included 6 males (20%) and 24 females (80%). Their ages ranged from 18 to 60 years.
Code
Status
Dilution Positive control
Cat. # Code MS-443P0
0.1 mL concentrated 1:25
Prostatic carcinoma
Cat. # Code M7047, IR657 Cat. # Code RB-10658-P1
1 mL concentrated
1:50
Breast carcinoma
1 mL concentrated
1:50
Breast carcinoma
Histopathological findings of H and E, Giemsa- and PTAH-stained sections of STs cases Epidermal changes Acanthosis was present in 48 cases (77.4%). Hyperkeratosis was present in all cases (100%). Papillomatosis was present in 48 cases (77.4%). Dermal changes Dilated blood vessels were present in 44 cases (71%). Mast cells were present in 50 cases (80.6%) with mean ± SD of 5.1 ± 4.1 cells. Collagen arrangement was haphazard in 24 cases (38.7%) and loose in 38 cases (61.3%) (Figures 1 and 2).
AR expression in studied groups (A) Control group: Epidermal and dermal immunoreactivity were demonstrated in Table 2 and Figure 3. (B) STs group: Epidermal and dermal immunoreactivity were demonstrated in Table 2 and Figure 4.
Comparison of AR expression in studied cases and controls Cases showed significantly enhanced expression of AR as the whole thickness positivity was higher than controls (p5.001). Strong epidermal intensity of AR (p5.001), nuclear pattern (p5.001), and higher epidermal H score (p5.001) were significantly associated with cases compared to control group. Significant differences were noticed also regarding dermal intensity and dermal H-score (p5.001 for both) (Table 2).
ERs expression in studied groups I-ERa (A) Control group: Epidermal and dermal immunoreactivity were demonstrated in Table 3 and Figure 5. Ultrastructural Pathology
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FIGURE 1. Section of a skin tag showed scattered mast cells with deep violet color in dermis (circles). Insert: zooming view for mast cell with cytoplasmic purple granules (Giemsa stain 400 for A, B).
FIGURE 2. Section of a skin tag showed (A) mast cell (circle) and haphazardly arranged collagen bundles (H&E 400) (B) Scattered mast cells (circles) and haphazardly arranged collagen bundles with red brown discoloration. Insert: Zooming view for a mast cell (PTAH stain 400).
(B) STs group: Epidermal and dermal immunoreactivity were demonstrated in Table 3 and Figure 6.
Comparison of ERa expression in studied cases and controls There was significant difference between cases and controls regarding epidermal intensity of ERa !
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(p5.001), epidermal H score (p5.001) and epidermal localization (p = .003). A significant difference was found between cases and controls regarding dermal H-score (p5.001) and dermal intensity (p = .002) (Table 3). II-ERb: (A) Control group: Epidermal and dermal immunoreactivity were demonstrated in Table 3 and Figure 7.
348 O. A. Bakry et al. TABLE 2. AR expression in studied groups. Variable Epidermal expression Present Absent Epidermal localization Basal Suprabasal Whole thickness Epidermal intensity Mild Moderate Strong Epidermal pattern Nuclear Nucleo-cytoplasmic Epidermal H-score Mean ± SD Dermal expression Present Absent Dermal intensity Mild Moderate Strong Dermal pattern Nuclear Nucleo-cytoplasmic Dermal H-score Mean ± SD
STs group No = 62
Control group No = 30
No 40 22
% 64.5 35.5
No 21 9
% 70 30
0 2 38
0 5 95
8 0 13
38.1 0 61.9
12 20 8
30 50 20
17 4 0
80.9 19.1 0
34 6
85 15
0 21
(B) STs Group: Epidermal and dermal immunoreactivity were demonstrated in Table 3 and Figure 8. p Value .60
5.001**
129.0 ± 59.0
5.001**
0 100
5.001**
68.1 ± 10.4
5.001**
38 24
61.3 38.7
21 9
70 30
.41
26 4 8
68.4 10.5 21.1
20 1 0
95.2 4.8 0
.04*
36 2
94.7 5.3
18 3
85.7 14.3
.23
64.7 ± 37.1
34.8 ± 6.0
Note. *Significant and **highly significant.
5.001**
Comparison of ERb expression in studied cases and controls There was significant difference between cases and controls regarding epidermal intensity of ERb (p = .007), epidermal H score, epidermal pattern and epidermal localization (p5.001 for all). A significant difference was found between cases and controls regarding dermal intensity (p = .001) (Table 3).
Association between AR H scores and clinical and histopathological data of studied cases Higher epidermal AR H score was significantly associated with axillary STs (p = .02), hyperpigmented tags (p = .03), the presence of acanthosis, the presence of papillomatosis (p = .04 for both), and loose collagen arrangement (p5.001) (Table 4). Higher dermal AR H score was significantly associated with the presence of papillomatosis (p = .002), dilated blood vessels (p = .006), the presence of mast cells (p = .01), and loose collagen arrangement (p = .02) (Table 4).
FIGURE 3. Normal skin showing (A) moderate nuclear AR expression in the whole thickness of the epidermis, (B) mild nuclear AR expression of epidermal cells and sebocytes (C) moderate nucleo-cytoplasmic AR expression in the whole thickness of the epidermis, hair follicles and in few basal cells of sebaceous glands (IHC 100 for A, 200 for B and 400 for C). Ultrastructural Pathology
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FIGURE 4. A case of ST showing (A) moderate AR expression in the whole thickness of epidermis and mild expression in dermal stroma (IHC 100). (B) Higher power view showing moderate nuclear AR expression in the whole thickness of epidermis and mild nuclear expression in dermal stroma (IHC 200). (C) Strong nucleo-cytoplasmic AR expression in epidermis and mild nuclear expression in dermal stroma (IHC 400). (D) Mild nucleo-cytoplasmic AR expression in dermal stromal cells (IHC 100 for A, 200 for B and 400 for both C and D).
Association between ERa H scores and clinical and histopathological data of studied cases Higher epidermal ERa H score was significantly associated with hyperpigmented tags (p5.001), positive family history (p = .003), and loose collagen arrangement (p = .004). Higher dermal ER H score was significantly associated with positive family history (p = .04), the presence of dilated blood vessels and the presence of mast cells (p = .04 for both) (Table 5).
Association between ERb H scores and clinical and histopathological data of studied cases Higher epidermal ERb H score was significantly associated with female gender (p = .004). Higher dermal ERb H score was significantly associated with obesity (p = .004), the presence of mast cells (p = .002), and dilated blood vessels (p = .04) (Table 6).
Correlation between AR, ERa and ERb H scores in studied cases Significant positive correlation was noted between AR and ERa H scores and between AR and ERb H scores !
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in epidermis (r = .52, p = .04 and r = .4, p = .02, respectively) and in dermis (r = .35, p = .03, r = .37, p = .01 respectively) of studied cases (data not shown in tables).
DISCUSSION STs manifest exclusively in postpubertal life [19]. Some studies stated that STs predominate in women with two peaks: one in pregnancy and generally regresses in puerperium and the other peak occurs in menopause [20]. These phenomena raise attention of hormonal involvement in their pathogenesis [21]. The increased estrogen during pregnancy and the relative increase in androgen following menopause [22] may provide evidence about the sharing of these sex steroids in pathogenesis. In addition, STs are more common in obese males in whom estrogen levels in blood increase in a direct proportion with BMI [23]. AR and ERs are expressed in various types of skin cells [7 11]. However, their role in proliferative skin disorders is not extensively studied. In the present study, ERs were upregulated in STs compared with normal skin. This went with El Safoury et al. [24] and may provide evidence about the involvement of ERs in STs pathogenesis. This role was explained by Verdier-Sevrain et al. [25] on the cellular scale; keratinocytes express both ERs
350 O. A. Bakry et al. TABLE 3. ERs expression in studied groups. ERa expression
Variable Epidermal expression Present Absent Epidermal localization Basal Suprabasal Whole thickness Epidermal intensity Mild Moderate Strong Epidermal pattern Nuclear Nucleo-cytoplasmic Epidermal H-score Mean ± SD Dermal expression Present Absent Dermal intensity Mild Moderate Strong Dermal pattern Nuclear Nucleo-cytoplasmic Dermal H-score Mean ± SD
ERb expression
STs group No = 62
Control group No = 30
STs group No = 62
Control group No = 30
No (%)
No (%)
p Value
No (%)
No (%)
p Value
46 (74.2) 16 (25.8)
19 (63.3) 11 (36.7)
.28
62 (100) 0 (0)
30 (100) 0 (0)
–
8 (17.4) 6 (13.0) 32 (69.6)
11 (57.9) 0 (0) 8 (42.1)
.003**
0 (0) 0 (0) 62 (100)
9 (30) 0 (0) 21 (70)
5.001**
14 (30.4) 22 (47.8) 10 (21.8)
19 (100) 0 (0) 0 (0)
5.001**
18 (29) 38 (61.3) 6 (9.7)
19 (63.3) 11 (36.7) 0 (0)
.007**
44 (95.7) 2 (4.3)
15 (100) 0 (0)
.41
24 (38.7) 38 (61.3)
0 (0) 30 (100)
5.001**
123.0 ± 51.6
51.6 ± 6.5
70.8 ± 29.5
5.001**
5.001**
143.3 ± 38.6
38 (61.3) 24 (38.7)
19 (63.3) 11 (36.7)
.85
62 (100) 0 (0)
30 (100) 0 (0)
–
21 (55.3) 13 (34.2) 4 (10.5)
19 (100) 0 (0) 0 (0)
.002**
38 (61.3) 18 (29) 6 (9.7)
26 (86.7) 4 (13.3) 0 (0)
.001**
35 (92.1) 3 (7.9)
16 (84.2) 3 (15.8)
.36
35 (56.4) 27 (43.6)
26 (86.7) 4 (13.3)
.36
5.001**
66.0 ± 24.4
57.2 ± 11.9
.06
66.1 ± 28.2
45.0 ± 6.7
Note. **Highly significant.
(ERa and ERb). Estradiol binds to keratinocytes’ ERa and ERb with high affinity and in its normal physiological range induces keratinocyte proliferation. In addition, 17b-estradiol has been shown to increase fibroblast proliferation in human skin [26]. In the current study, AR was upregulated in STs compared with normal skin. This went with EL safoury et al. [24]. Choudhry et al. [27] postulated that, although keratinocytes are known to express AR and are target cells for androgen action, the effect of androgens on keratinocytes’ activity remains to be clarified. Pelletier and Ren [10] concluded that, the presence of AR-expressing keratinocytes within the pilosebaceous ducts supported the hypothesis that androgens can directly influence direct keratinization occurring in acne process. In addition, Tadokoro et al. [28] stated that AR can modulate proliferation and differentiation of epithelial cells of the epidermis and adnexa, functional activity of dermal fibroblasts and fibrocytes. The significant association between higher epidermal AR H score and the presence of acanthosis and papillomatosis in studied cases may support the role of AR in epithelial proliferation and STs’ pathogenesis. Further studies at the molecular level and on
larger scale of cases are warranted for firmer conclusion. The upregulation of AR and ERs in STs and their possible role in pathogenesis went with the conclusion of Thornton et al. [8] that tissues traditionally thought to be responsive to one class of steroids, contain receptors for other classes, and there is growing evidence that steroid receptors can crosstalk with one another. This was underscored by the significant positive correlation between AR and ERs H scores in epidermis and dermis, detected in the current work. In the current study, higher dermal AR and ERs H scores was significantly associated with the presence of mast cells. Similar observation was reported by Zaher et al. [29]. Mast cells express high-affinity ERs and 17b-estradiol augments the secretion of histamine and serotonin [30]. Binding of estradiol to ERa, initiates a rapid onset, progressive influx of extracellular Ca2, inducing exocytosis of mast cell products [31]. Mast cell mediators are capable of inducing keratinocyte and fibroblast proliferation with epidermal hyperplasia and increased collagen synthesis, contributing to the main pathologic abnormalities observed in STs [29]. Therefore, a question arises; Ultrastructural Pathology
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351
FIGURE 5. Expression of ERa in normal skin (A) moderate nucleo-cytoplasmic expression in the whole thickness of the epidermis, hair follicle, few stromal cells and blood vessel endothelium, (B) mild nucleo-cytoplasmic expression in basal and differentiated cells of sebaceous glands (C) mild to moderate nuclear expression in blood vessel endothelium and stromal cells (IHC 200 for A and 400 for B, C).
FIGURE 6. Expression of ERa in STs showing (A) strong nuclo-cytoplasmic expression in the whole thickness of epidermal and moderate nuclo-cytoplasmic expression in dermal stroma, (B) moderate nuclo-cytoplasmic expression in the whole thickness of epidermal and (C) moderate nuclo-cytoplasmic expression in suprabasal layers of epidermal, strong nuclear expression in basal cell layer and strong nuclo-cytoplasmic in dermal stromal cells (H&E 200 for A and 400 for B and C).
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352 O. A. Bakry et al.
FIGURE 7. Expression of ERb in normal skin showing (A) moderate nucleo-cytoplasmic expression in the whole thickness of the epidermis, few stromal cells and blood vessels, (B) mild to moderate nucleo-cytoplasmic expression in outer and inner root sheath of hair follicles and fibroblastic stromal cells (C) mild to moderate nuclear expression in blood vessels and stromal cells in between (D) mild nucleo-cytoplasmic expression in basal and differentiated cells of sebaceous glands (IHC 100 for A, 200 for B and 400 for C, D).
FIGURE 8. Expression of ERb in STs (A) panoramic view exhibiting strong nucleo-cytoplasmic expression in the whole thickness of epidermis and mild nucleo-cytoplasmic expression in scattered dermal stromal cells and blood vessels (B) closer view of the previous figure illustrating epidermal and dermal expression (C) mild nucleo-cytoplasmic expression in dermal stromal cells and blood vessels (IHC 100 for A, 200 for B and 400 for C).
Ultrastructural Pathology
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353
TABLE 4. Association between AR H scores and clinical and histopathological data of studied cases (values were expressed in mean ± SD).
Clinical variables Gender Male Female Site Abdomen Axilla Back Neck Obesity Present Absent Diabetes mellitus Present Absent Family history Positive Negative Color Skin colored Hyperpigmented Histopathological variables Acanthosis Present Absent Papillomatosis Present Absent Dilated blood vessels Present Absent Mast cells Present Absent Collagen arrangement Haphazard Loose
Epidermal H score
p Value
Dermal H score
p Value
162.5 ± 63.8 120.6 ± 55.7
.07
72.0 ± 31.55 62.14 ± 39.09
.48
140.0 ± 0.0 185.0 ± 40.4 107.5 ± 30.6 115.5 ± 63.6
.02*
70.0 ± 0.0 85.0 ± 46.9 50.0 ± 17.9 60.9 ± 37.2
.32
135.6 ± 60.0 123.6 ± 59.1
.48
67.5 ± 38.2 62.7 ± 37.1
.79
150 ± 45.7 122 ± 61.9
.11
.15
103.3 ± 53.9 133.5 ± 59.5
.36
40.0 ± 11.5 67.7 ± 38.1 62.3 ± 36.5 70.0 ± 39.5
135.0 ± 5.8 112.3 ± 53.1
.03*
50.0 ± 17.9 67.5 ± 39.3
.42
140.0 ± 60.1 96.0 ± 42.9
.04*
72.5 ± 39.4 51.4 ± 29.6
.09
138.1 ± 58.6 92.5 ± 48.0
.04*
75.7 ± 35.6 34.0 ± 20.7
.002**
133.8 ± 50.3 110.0 ± 87.8
.12
74.3 ± 38.6 38.0 ± 10.3
.006**
136.7 ± 65.9 127.7 ± 58.7
.94
73.6 ± 38.2 40.0 ± 18.9
.01*
90.0 ± 61.0 193.3 ± 45.9
5.001**
46.7 ± 32.3 96.7 ± 37.2
.02*
.53
Note. *Significant and **highly significant.
could antihistamines and mast cell stabilizers be beneficial in treatment and/or prevention of STs, associated with large number of mast cells? The answer requires future clinical trials. Mast cells stimulated by friction can start STs formation through its interaction with fibroblasts and keratinocytes. However, the finding that STs are not a common association in mastocytosis indicates that mast cells without a precipitating factor can not initiate STs [32]. Contrary to our findings, Di Matteo et al. [30] found that blocking AR with sex hormone antagonists in the testis of frog causes a significant increase in mast cell count. In addition, it was found that testosterone has an inhibitory effect on mast cell secretion of histamine and serotonin in rats [33]. Further investigation is needed to prove or deny current observation. In the current study, higher ERa and AR H scores were significantly associated with cases showing loosely arranged collagen fibers. This finding was !
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not discussed before in similar studies. Estrogen targets human skin fibroblasts and has been shown to increase fibroblast proliferation in human skin [26]. However, the association between ERa and AR upregulation and loose collagen arrangement requires further investigation. In the current study, higher epidermal ERa H score was significantly associated with positive family history of STs in studied cases. Similar observation was reported before in breast cancer cases and was explained on genetic basis [34,35]. Whether this is applicable for STs or not, requires further studies on larger scales including cases and their relatives for firmer conclusion. In the current work, higher epidermal AR H scores were significantly associated with axillary STs; a finding that was not reported before. Whether this finding denotes regional differences in AR density in different body sites or not requires further research. STs are related to obesity [36], and a hormonal mechanism has been suggested in obese females,
354 O. A. Bakry et al. TABLE 5. Association between epidermal ERa H scores and the clinical and histopathological data of ST cases (values were expressed in mean ± SD).
Clinical variables Gender Male Female Site Abdomen Axilla Back Neck Obesity Present Absent Diabetes mellitus Present Absent Family history Positive Negative Color Skin colored Hyperpigmented Histopathological variables Acanthosis Present Absent Papillomatosis Present Absent Dilated blood vessels Present Absent Mast cells Present Absent Collagen arrangement Haphazard Loose
Epidermal H score
p Value
Dermal H score
p Value
120.0 ± 42.6 124.1 ± 54.3
.81
58.0 ± 33.59 68.9 ± 26.2
.30
180.0 ± 0.0 136.7 ± 45.0 100 ± 32.2 121.3 ± 54.2
.25
80.0 ± 0.0 78.8 ± 8.3 61.7 ± 7.5 61.4 ± 35.5
.43
135.0 ± 74.6 120.5 ± 45.6
.47
45.0 ± 17.3 68.5 ± 28.4
.09
125.7 ± 58.8 118.9 ± 37.1
.79
68.2 ± 34.6 63.1 ± 16.6
.91
134.4 ± 50.4 82.0 ± 27.8
.003**
69.7 ± 28.9 46.7 ± 13.7
.04*
150.0 ± 46.2 168.3 ± 55.1
5.001**
60.0 ± 20.9 62.5 ± 9.6
.11
127.8 ± 50.9 106 ± 50.6
.24
66.25 ± 26.34 65.71 ± 32.28
.96
123.7 ± 52.1 120.0 ± 48.7
.91
66.8 ± 22.8 64.0 ± 41.0
.35
128.1 ± 54.4 111.4 ± 42.0
.44
69.3 ± 20.2 57 ± 43.9
.04*
127.4 ± 54.2 102.5 ± 25.5
.41
71.1 ± 29.7 52.0 ± 18.1
.04*
66 ± 19.6 99.2 ± 35.0
.004**
78 ± 27.8 86.7 ± 50.9
.82
Note. *Significant and **highly significant.
where there is increased peripheral aromatization of androgens to estrogens [37]. Similar hormonal imbalance exists in obese males with hyperestrenemia [23] as obese men have elevated levels of estrone, both free and total, as well as estradiol [38]. In the current study, higher ERb H score was significantly associated with obese cases. However, no significant difference in H score values of ERa and AR between obese and non obese cases was noted. This difference in receptor immunoreactivity between ERb on one hand and ERa and AR on the other hand regarding relation to obesity, may be explained on basis of ERb predominance in human skin more than ERa [8,39]. However, further studies are warranted. El Safory et al. [24] did not find an association between AR and ERs expression and BMI of their examined cases. Further studies including larger number of cases with different BMI are needed. STs were reported to have a probable association with diabetes mellitus where they may be a skin
marker for underlying impaired carbohydrate metabolism and patients with STs should be evaluated for the presence of diabetes mellitus [40,41]. In the present work, no significant difference in ERs and AR H scores was found between diabetic and non diabetic cases. Testosterone levels were reported to be lower in diabetic men [42]. However; the effect of diabetes on estrogen, AR or ERs was not discussed before. In the present work, ERa, ERb immunoreactivity was detected in vascular endothelium both in normal skin and STs but AR immunoreactivity in vascular endothelium was positive in STs and negative in normal skin sections (as demonstrated in histopathological figures). Pelletier and Ren [10] reported positive endothelial immunoreactivity of ERb and negative immunoreactivity of AR and ERa in their study on normal skin specimens. More studies involving different body sites are recommended to prove or deny current observations. In addition, the current work showed that higher ERs and AR H Ultrastructural Pathology
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TABLE 6. Association between ERb H scores and the clinical and histopathological data of ST cases (values were expressed in mean ± SD). Variable Clinical variables Gender Male Female Site Abdomen Axilla Back Neck Obesity Present Absent Diabetes mellitus Present Absent Family history Positive Negative Color Skin colored Hyperpigmented Histopathological variables Acanthosis Present Absent Papillomatosis Present Absent Dilated blood vessels Present Absent Mast cell Present Absent Collagen arrangement Haphazard Loose
Epidermal H score
p Value
Dermal H score
p Value
136.3 ± 28.6 171.7 ± 58.3
.004**
62.9 ± 23.6 78.3 ± 24.4
.40
170.0 ± 0.0 132.9 ± 28.7 157.5 ± 68.6 142.8 ± 33.6
.39
90.0 ± 0.0 58.6 ± 19.6 67.5 ± 33.3 143.3 ± 38.6
.35
163.8 ± 37.6 116.5 ± 18.1
.31
73.8 ± 22.7 55.8 ± 23.0
.004**
167.5 ± 33.3 139.6 ± 38.2
.06
77.5 ± 13.9 64.2 ± 25.2
.15
140.0 ± 13.6 144.4 ± 43.5
.56
71.4 ± 10.3 64.4 ± 27.1
.15
143.8 ± 51.6 143.2 ± 33.3
.96
56.3 ± 26.3 69.6 ± 22.9
.06
146.7 ± 41.4 130.0 ± 20.0
.05
68.3 ± 25.7 56.7 ± 15.6
.14
146.8 ± 40.4 126.0 ± 21.7
.12
67.2 ± 25.8 60.0 ± 14.9
.24
146.9 ± 47.6 140.6 ± 30.4
.55
73.9 ± 24.8 60.0 ± 22.6
.04*
143.6 ± 43.6 142.5 ± 19.8
.89
71.4 ± 23.1 51.3 ± 22.2
.002**
120.0 ± 0.0 144.1 ± 38.9
.39
50.0 ± 0.0 66.6 ± 24.6
0.28
Note. *Significant and **highly significant.
scores were significantly associated with cases showing dilated blood vessels. Estrogen is known to have a vasodilator effect on vessel wall through the release of prostacyclin and nitric oxide by a non-genomic action [43]. The effect of androgen on vessel wall is poorly investigated with few studies reported vasodilator effect [44,45]. Further research is needed for more clarification of androgens and AR vascular role. In the current work, higher ERa H score was significantly associated with cases having hyperpigmented lesions. In vitro studies have shown that cultured human melanocytes express ERa and ERb, and estradiol increases the levels of tyrosinase, tyrosinase-related-protein 1 and tyrosinase-relatedprotein 2, the enzymes involved in human eumelanogenesis within normal human melanocytes [46,47]. However, the association between lower AR H score and hyperpigmented lesions, detected in the current work, remains unexplained. ERs and AR are known to exhibit nuclear immunoreactivity [14,15]. In our study, nucleo-cytoplasmic !
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pattern of both ERs and AR was detected in examined cases and control subjects. Fu and Simoncini [48] suggested that androgens and other sex steroid hormones may affect cells through mechanisms that do not involve gene transcription/expression, but through activation of cytoplasmic signaling pathways that affect cell survival and/or modulate other growth factor signaling [49]. Parker [50] concluded that ERs can move between the nucleus and the cytoplasm, but under normal conditions they are predominantly located in the nucleus. Pelletier and Ren [10] detected nucleo-cytoplasmic ERb immunoreactivity in ducts of eccrine sweat glands. Heemers and Tindall [2] concluded that, AR, in its inactive form, is located in the cytoplasm of the cell in complex with heat-shock proteins. Upon ligand binding, AR homodimerizes and translocates to the nucleus where it binds to the specific sequences on the DNA known as androgen response elements. It may also recruit cofactors that either enhance or reduce AR transactivation [51]. In the current study, there was
356 O. A. Bakry et al. significant association between nuclear pattern of AR and ERb with STs when compared to control. In addition, nuclear pattern appears more in STs regarding dermal AR and ERa although the difference between cases and controls was insignificant. These observations may provide evidence about the role of AR and ERs in STs pathogenesis through their presence in their active form. In conclusion, AR and ERs may share in STs pathogenesis through their effects on keratinocytes, fibroblasts and mast cells. This may open new avenue for treatment and prevention in susceptible individuals. Further studies on larger scales are warranted to investigate their exact molecular mechanisms in tag development.
DECLARATION OF INTEREST The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper. Samples were taken after obtaining written consent from the patients and control subjects.
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Notice of Correction: A misspelling of one author’s name has been corrected following initial online publication.
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