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Keywords:

  • ectodermal dysplasia;
  • skin;
  • hair;
  • biopsy;
  • histopathology;
  • electron microscopy

Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

Ankyloblepharon-Ectodermal defects-Cleft lip/palate (AEC) syndrome is a rare disorder of hair, skin, nails, and dentition caused by mutations in the p63 gene. Pathologic changes of skin and hair in AEC syndrome have previously been described in isolated case reports. Biopsies of normal and lesional skin from 19 patients with AEC syndrome were examined by light microscopy. Hair samples from 18 patients were examined by light and scanning electron microscopy. Histopathologic changes identified within the skin biopsies from clinically unaffected skin include mild atrophy, focal orthokeratosis, and mild superficial perivascular lymphocytic dermatitis. Scattered melanophages in the superficial and deep dermis likely reflect post-inflammatory change. One patient with a unilateral eruption of monomorphic papulopustules on the chest and shoulder demonstrated an acneiform intraepidermal pustule. Examination of the hair shafts revealed atrophy and loss of melanin pigment in some of the patients. Structural abnormalities included pili torti, pili trianguli et canaliculi, and irregular indentation and shallow grooves. Skin and hair findings in AEC syndrome were found to be generally similar to those described in other ectodermal dysplasia syndromes and corroborates the few prior descriptions in AEC syndrome specifically. © 2009 Wiley-Liss, Inc.


INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

The ectodermal dysplasias are a large group of rare disorders resulting from abnormal embryonic development of the ectoderm and abnormal interactions of the ectoderm with the mesoderm and endoderm [Freire-Maia, 1971; Solomon and Keuer, 1980; Pinheiro and Freire-Maia, 1994; Priolo and Laguna, 2001; Itin et al., 2003; Lamartine, 2003; Itin and Fistarol, 2004].

Clinical manifestations are variable, but they are generally unified by the presence of abnormalities of the skin, nails, hair, teeth, and/or sweat glands. Dermatologic findings in these disorders may include adermatoglyphia, reticular pigmentation or hypopigmentation, atopic dermatitis, scaling, telangiectasias, atrophy, and palmoplantar hyperkeratosis. Alopecia and hypotrichosis are components of many ectodermal dysplasias, often associated with brittle or uncombable hair [AEC, 2008].

Due to the rarity of AEC syndrome, descriptions of the skin and hair abnormalities have been limited to predominantly clinical descriptions in a few individual patients and families. Skin biopsies and hair samples have been reported in very few of these patients.

Prior reports of light microscopic findings of AEC hairs have shown miniature hair bulbs. Previously reported scanning electron microscopy findings of hair samples documented abnormal shafts with longitudinal fluting and twisting as seen in pili torti, a scalloped appearance, as well as longitudinal, sigmoid-shaped, or transverse greenstick fractures (trichoclasis) [Hay and Wells, 1976; Greene et al., 1987; Rowan, 1996]. Systematic study of the pathologic changes of skin and hair has not previously been possible due to the rarity of this condition.

METHODS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

An international conference on AEC syndrome was convened at Texas Children's Hospital (Baylor College of Medicine, Houston, TX, USA) in November 2006. During this research conference, AEC-affected patients were invited to provide skin and hair samples for pathology studies. Twenty-one participants and relatives were consented for this component of the study. Four-mm punch biopsies of skin were obtained from normal-appearing skin in most cases, and in a few cases from regions with pigmentary, hyperkeratotic, or inflammatory lesions. The skin biopsies were fixed in 10% neutral buffered formalin. After standard processing, a representative hematoxylin & eosin (H&E)-stained section of each biopsy was examined and the histopathologic changes tabulated. Leveled sections or horizontal sections of dermis were not performed. Scalp hair samples were also obtained from each patient with consent. Several hair strands from each sample were mounted on glass slides and examined by light microscopy using transmitted and polarized light. Additional hair shaft material from each sample was also examined by scanning electron microscopy (EM) using standard methods. The study was conducted after institutional research board (IRB) approval and with informed consent from participants. Funding for this conference was provided by the National Foundation for Ectodermal Dysplasias.

RESULTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

Skin punch biopsies and scalp hair shaft samples were obtained from 20 participants, including 17 patients with samples of both skin and hair, 2 patients with skin biopsy only, and 1 patient with hair samples only. One patient had skin biopsies performed at two sites, yielding a total of 20 punch biopsies and 18 hair samples. Patient demographics included 8 male and 12 female patients with an age range of 4 months to 59 years. The patients providing skin and/or hair samples included 4 infants (age <24 months), 11 children and adolescents (age 2–17 years), and 5 adults (18 years or older). Patient ethnicities included 16 Caucasian, 1 Hispanic, and 3 African-Americans. There were two sib sets (AEC 3, 10; AEC2, 12, 14) and 4 parent–child pairs.

Skin biopsies were obtained from the arm (9), thigh (4), leg (1), back (4), shoulder (1), and an unspecified site (1). While some patients showed scalp erosions and dermatitis clinically, no patients were biopsied at this site. Pathologic findings identified by light microscopy of the skin biopsies are summarized in Table I. Three biopsies were obtained at sites of clinically apparent abnormality including one patient with atrophy, telangiectasia, and hypopigmentation on the back, one patient with reticular hyperpigmentation and hypopigmentation on the back, and one patient with a pustule on the shoulder. The remaining 17 biopsies were obtained from “normal” skin without clinically apparent lesions. The most common findings were mild hyperkeratosis (47.3%; 9/19 patients), mild papillomatosis (15.8%; 3/19), epidermal atrophy (42.1%; 8/19), variable basilar pigmentation and/or pigment incontinence (78.9%; 15/19), and prominence of the superficial perivascular plexus with a minimal to mild perivascular lymphocyte infiltrates (89.5%; 17/19) (Fig. 1). Skin appendages were noted in some cases, but were generally inconspicuous. Representative sections of skin in these 19 patients showed hair follicles in 6 cases (31.6%) and eccrine glands and/or ducts in 15 cases (78.9%). These structures were absent in 3 of 19 patients (15.8%).

Table I. Summary of Skin Histopathologic Changes in AEC Syndrome
Histopathologic findingNumber, n = 19Percentage (%)
Mild hyperkeratosis947
Epidermal atrophy, mild842
Mild papillomatosis316
Irregularity and bridging of rete632
Prominent superficial vascular plexus1158
Superficial perivascular lymphocytes, minimal/mild1789
Focal exocytosis of lymphocytes (interface dermatitis)632
Variable basilar pigmentation211
Dermal melanophages (pigment incontinence)1579
Rare dyskeratotic cell15
Pustule15
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Figure 1. Histopathology of skin in AEC syndrome. Low power demonstrates variable epidermal architecture, with some biopsies showing atrophy and others with mild papillomatosis and mild orthokeratosis (A, thigh, AEC9 10×). The epidermis typically showed an irregular blunted rete ridge pattern with short “digitate” rete in many cases. The majority of biopsies showed minimal to mild lymphocyte infiltrates associated with a prominent reactive superficial vascular plexus and scattered melanophages in the superficial dermis (B, AEC21 20×). Variable basilar melanin pigment is also noted in this African-American child.

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A variety of pathologic findings were identified by light microscopy and scanning electron microscopy in the hair samples (Table II). Most of the hair shafts were thin and atrophic-appearing; although, there was slight variation in width of the hair shafts, even in individual patients. One-third of patients had hair of two different colors (6/18); that is, some pigmented hair shafts and some with near-absent pigment. There was also variable pigmentation within individual hair shafts, including variable discontinuous medullation, areas of coarse clumped central pigment, pili annulati, and pigmented hairs with focal areas devoid of pigment (Fig. 2). All cases (100%, 18/18) showed irregularity of hair shaft structure, including pili torti, bent shafts, and at least focal grooved contours. Pili torti were noted by light and/or scanning electron microscopy in 4/18 cases (22.2%) (Fig. 3). Scanning electron microscopy showed more gradual non-periodic twists in an additional 7/18 cases (39%), yielding a total of 61.1% of cases with twisting of the hair shaft. Many hairs were flattened to ovoid and some were angulated to reniform (pili trianguli). All cases had at least irregular shallow grooves and irregular indentation, and 12 cases (66.7%) had distinct deep linear grooves (pili canaliculi). Although polarization of hair in most cases showed areas of light and dark mottling reflecting the irregular undulating surface, the alternating light and dark bands of trichothiodystrophy were not demonstrated. Non-specific cuticular weathering, partial hair fracture (trichoclasis), and trichorrhexis nodosa were seen occasionally. No cases showed trichorrhexis invaginata, trichoschisis, or monilothrix.

Table II. Summary of Scalp Hair Pigment and Structure in AEC Syndrome, by Light Microscopy and Scanning Electron Microscopy
 Number, n = 18 patientsPercentage (%)
Pigmentation
 Amount of pigment
  Bicolored (pale and pigmented)633
  Pale/sparse to absent pigment211
  Light uniform pigment317
  Dark uniform pigment739
 Pattern of pigment distribution
  Continuous medulla633
  Discontinuous medulla/aggregated pigment1794
  Non-medullated528
  Pili annulati16
Structure
 Thin, atrophic18100
 Torsion1161
  Pili torti (regular periodic torsion)422
  Gradual twisting only739
 Grooves18100
  Pili canaliculi (deep grooves)1267
  Shallow linear grooves only633
 Pili trianguli633
 Bent shaft/kinking1056
 Cuticle weathering16
 Trichoptilosis16
 Trichoclasis528
 Trichorrhexis nodosa16
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Figure 2. Hair shaft pigment and structure: Light microscopy. The hair shafts were generally thin with variable pigment and non-specific structural abnormalities. Some patients showed bicolored hair (A, AEC15 4×). Pigment alterations included alternating dark and light bands of pili annulati (B, AEC13 4×) and focal segments devoid of pigment. Structural defects included surface grooves, gentle non-periodic twisting of the hair shafts, and other irregular bending of the hair shaft (C, AEC10 10×). Polarized light accentuates the irregularities in the hair shaft structure, including pili torti (upper right) and variable light and dark mottling (lower left) (D, AEC4 10×).

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Figure 3. Hair shaft structure. Scanning electron microscopy demonstrates longitudinal linear grooves (A, AEC15) and reniform shape of the hair shaft on cross-section, typical of pili trianguli et canaliculi. Other structural defects included pili torti (B, AEC12) and bent or kinked hair shafts.

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DISCUSSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

Skin Pathology

Due to the rarity of AEC syndrome, pathologic changes of skin in this syndrome are inadequately described and limited to individual case reports or small series. In one such report, skin biopsies from a parent and child with AEC syndrome showed mild to moderate papillomatosis with basal layer hyperpigmentation and epidermal digitate budding (reticulate acropigmentation, Kitamura-like), with a lesser degree in the mother relative to her 2-year-old and very similar to that seen in our patients [Drut et al., 2002]. The erosive scalp dermatitis characteristic of AEC syndrome leads to scarring alopecia and corresponds microscopically to perifollicular and intrafollicular neutrophil infiltrates, focal follicle destruction with lymphoplasmacytic infiltrates, “naked” hair shafts, fibrous follicle tracts, and reduced terminal hair density [Park et al., 2005]. In this syndrome, it has been suggested that immaturity of the epidermis and appendages on the scalp may predispose to fragility, inflammation and infection [Cliff and Holden, 1997]. Reports of follicular and eccrine structures in AEC patients have been variable, with an initial description indicating “almost complete absence of epidermal appendages” and later reports indicating normal-appearing eccrine structures [Hay and Wells, 1976; Fosko et al., 1992].

In our patients, the skin biopsies were obtained predominantly from random skin sites (arm, thigh, leg, back) with a discrete lesion biopsied in only one case (acneiform pustule, AEC19). The scalp dermatitis and palmoplantar keratotic lesions were not biopsied. Many of our patients had irregular reticulate pigmentation abnormalities, which are reflected in the random biopsies by variable basilar pigmentation and scattered melanophages in the superficial dermis. Given that most of these biopsies also showed minimal to mild perivascular lymphocytes and rare exocytosis of lymphocytes, the melanin incontinence may be a post-inflammatory phenomenon. Eccrine glands and hair follicles were present in a minority of the skin biopsy sections, suggesting paucity of these structures overall; although, systematic quantitation was not performed. In those cases with skin appendages, no morphologic abnormalities of the hair follicles, eccrine ducts or glands were identified.

Hair Pathology

Little is known about the hair pigmentation and structure in AEC patients. Hypopigmentation of terminal hairs is a common finding in reports of both AEC syndrome and Rapp–Hodgkin syndrome, which are now considered to represent varying clinical manifestations of the same condition [Hay and Wells, 1976; Greene et al., 1987; Fosko et al., 1992; Zenteno et al., 1999; Park et al., 2005]. Pili torti and pili canaliculi have been reported in AEC and Rapp–Hodgkin syndrome (RHS) patients [Silengo et al., 1982; Salinas and Montes, 1988; Walpole and Goldblatt, 1991; Camacho et al., 1993; Sahin et al., 2004; Park et al., 2005; Steele et al., 2005], as well as other ectodermal dysplasia syndromes [Micali et al., 1990; Trüeb et al., 1994, 1995; Hicks et al., 2001]. In one review of AEC syndrome, 4 of 12 patients reportedly had pili torti et canaliculi [Fosko et al., 1992]. Longitudinal grooves and flattening of hair surfaces have also been noted in hypohidrotic ectodermal dysplasia and ectrodactyly ectodermal dysplasia-clefting (EEC) syndrome [Hicks et al., 2001; Park et al., 2005]. Scanning electron microscopy of hair shafts in AEC syndrome has also shown defective cuticles.

In the current cases, light microscopy and scanning electron microscopy of hair from AEC patients showed a number of changes in common with uncombable hair syndrome (pili trianguli et canaliculi) [Hicks et al., 2001]. In contrast to the round or ovoid shape of normal hair on cross-sections, the hair from patients with AEC syndrome frequently showed flattening with angulation, triangular shape, deep grooves (canaliculi) resulting in a reniform shape or undulating contours, shallow grooves, and irregular surface indentations. Most cases showed at least occasional partial twists with a few of these showing several tight twists (pili torti). The surface irregularity, flattening, and twists in the hair shaft were accentuated by examination under polarized light.

In addition to these structural abnormalities, the hair shafts showed variable pigmentation, with many showing pallor and near absent pigment. Several patients had bicolored hair with some hairs showing retained pigment and some pale hairs nearly devoid of pigment. One case showed an area of pili annulati composed of alternating bands of depigmentation and aggregated dark pigment. Others showed less conspicuous aggregation or clumping of central pigment. However, no pathognomonic hair shaft alterations were identified.

Role of p63 in AEC syndrome

Five syndromes are caused by mutations in the p63 gene: AEC, ectrodactyly-ectodermal dysplasia-clefting (EEC), Acro-dermato-ungual-lacrimal-tooth (ADULT), Rapp–Hodgkin (RHS), and limb-mammary syndrome (LMS). p63 mutations have also been described in two non-syndromic conditions: split-hand/split-foot malformation (SHFM) and non-syndromic cleft lip [Brunner et al., 2002a,b; van Bokhoven and Brunner, 2002; Rinne et al., 2007]. AEC syndrome typically results from heterozygous missense mutations in the sterile alpha motif (SAM) domain [McGrath et al., 2001]. The SAM domain contains protein–protein interaction modules of over 40 proteins involved in developmental regulation, and it is thought that the p63 SAM domain determines interaction of TP63 protein with other proteins involved in transcriptional regulation and skin appendage development [McGrath et al., 2001; Barbieri and Pietenpol, 2006; Mikkola, 2007]. Immunohistochemistry using antibodies to p63 has been used to study expression in skin biopsies from AEC patients [McGrath et al., 2001; Drut et al., 2002]. In these cases, p63 shows abnormally increased expression in the basal layer and also in the suprabasilar keratinocyte nuclei. Studies of keratins and other structural skin proteins in AEC patients have shown a normal staining pattern with keratins 10 and 14, but aberrant nuclear and cytoplasmic expression of anti-filaggrin, which is normally expressed only in the cytoplasm [McGrath et al., 2001]. No increase in apoptosis has been observed by TUNEL staining. TP63 protein is likely essential not only for embryologic commitment of immature ectoderm to specific epidermal lineages, but also for the maintenance of the epidermal stem cell population necessary for epithelial morphogenesis and renewal [Morasso and Radoja, 2005]. Precisely how the interactions of the abnormal TP63 protein and other regulatory proteins in the skin translate into the clinical phenotypes of skin fragility, pigmentation, and hyperkeratosis remains unknown; although, p63 regulation of involucrin and loricrin in the skin may play a role [Barbieri and Pietenpol, 2006].

The role of p63 in producing the abnormally sparse and wiry hair in AEC syndrome is also unclear. In general, pili trianguli et canaliculi are thought to result from focal asymmetry of hair bulbs with lateral loss of hair matrix, resulting in a linear groove along the hair shaft. The inner root sheath is of uniform thickness, conforming to the contour of the triangular or grooved hair shaft, while the outer root sheath fills in the focal defect of the hair shaft, resulting in round hair follicles on cross-section. These morphologic changes are the converse of the normal relationship of the inner and outer root sheaths by which the inner root sheath compensates for irregularities in the hair surface and the outer root sheath is a fixed cylindrical structure. The variability in the outer root sheath in pili trianguli et canaliculi has been suggested to reflect a defect in the keratinization process [Hicks, 2001].

While there were no pathognomonic findings of the skin and hair identified in our study, there were several interesting changes noted as characteristic and consistent among subjects. The histopathologic changes in the skin may be due to the underlying mutation but may also be secondary to inflammation related to the disrupted gene product and its effects. The structural and pigmentary changes of the hair may be due to similar phenomena. Regardless of the cause, these pathologic features of the skin and hair are noteworthy as they are described in the largest cohort of patients affected by AEC syndrome to date.

Acknowledgements

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES

The authors wish to thank Mr. Jim Barrish and Mr. Ralph Nichols for their technical expertise in performing the scanning electron microscopy for this study. Thank you also to the patients who participated and to the National Foundation for Ectodermal Dysplasias for support of this symposium.

REFERENCES

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. REFERENCES