ARTICLE
Auteur(s) : Monica Pascucci1, Patrizia
Posteraro2, Cristina Pedicelli3, Alessia
Provini3, Luigi Auricchio4, Mauro
Paradisi3, Daniele
Castiglia1
1Laboratory of Molecular and Cell Biology, IDI-IRCCS,
via Monti di Creta 104, 00167 Rome, ItalyFax (+39) (06) 6646
4705
2Laboratory of Clinical Pathology and Microbiology,
Ospedale S.Carlo, Rome, Italy
3VII Dermatology Division, IDI-IRCCS, Rome, Italy
4Department of Systematic Pathology, Dermatology
Division, University “Federico II” of Naples, Naples, Italy
accepté le 13 Juillet 2006
Epidermolysis bullosa simplex with mottled pigmentation (EBS-MP,
OMIM 131960), a rare cutaneous genetic disorder, was first
described by Fisher and Gedde-Dahl [1]. This condition can be
distinguished from the other forms of EBS by the progressive
appearance of reticulated hyperpigmentation, especially of the
neck, upper trunk and extremities, beginning during
infancy.Blistering lesions affecting predominantly the extremities
appear at birth or in early childhood, heal without scarring and
milia formation and show a tendency to improve with age. Another
regular feature is the development of small warty palmoplantar
keratoses measuring 2-5 mm in diameter. Skin atrophy and nail
dystrophy have also been reported in some patients
[2].Histologically, intraepidermal cleavage and keratinocyte
degeneration are constant features of EBS-MP. An increased number
of melanosomes within basal cells is also demonstrated by
ultrastructural analysis of hyperpigmented areas [3].EBS-MP is
inherited in an autosomal dominant pattern and the disease-causing
mutation, P25L, has been found in the non-helical domain V1 of
keratin 5 [4]. Mutations affecting the central rod domains of
keratins 5 and 14 have been shown to interfere with normal
intermediate filament (IF) formation and to cause EBS Koebner and
Weber-Cockayne types, whereas the most severe form of EBS
Dowling-Meara has been associated to mutations in the end terminal
rod domains of keratins 5 and 14.
Case report
The proband was a 3-year-old Italian male, born of
non-consanguineous and healthy parents. Another sibling was
unaffected and there was no family history of skin or genetic
disease.
The patient had been suffering from skin blisters and erosions
since birth: the lesions occurred mainly over the hands and feet
and worsened during summer months. By the age of 2 years the
blisters were localized exclusively over the hands and feet and a
reticulated hyperpigmentation, especially involving the upper
extremities, had appeared. On physical examination, the patient
displayed a mosaic pattern of dark and light pigmented spots,
measuring 2 to 5 mm in diameter, distributed over the trunk
and distal extremities where they appeared more pronounced. A few
blisters over the hands and feet were observed ( (figure 1 )A, B). The
patient also presented cone-shaped incisors, attenuated
dermatoglyphics on the hands, and erosive blepharitis of the left
eye.
On the basis of clinical features, EBS-MP and Kindler syndrome
(KS) were considered. A skin biopsy and blood samples were obtained
from the patient and his parents after written informed consent.
Ultrastructural analysis of a skin biopsy from a pigmented lesion
showed numerous mature melanosomes and melanosome complexes around
the nuclei of keratinocytes in the basal and suprabasal epidermal
layers. Several melanophages filled with clusters of mature
melanosomes were observed in the papillary dermis ( (figure 1C) ). No basement
membrane duplications were noted. On the basis of the
ultrastructural findings, we decided first to carry out a mutation
search in the keratin 5 gene (KRT5), by targeting the P25L
mutation.
The exon 1 of KRT5 was amplified as previously described [5]
from the proband and family members’ DNA, and the corresponding PCR
products were directly sequenced (ABI Prism 377, Applied
Biosystems). Sequence analysis of exon 1 from the proband DNA
revealed a heterozygous C to T transition at nucleotide 74 that
converts a proline to leucine residue ( (figure 1D) ). The P25L
mutation was confirmed on the patient DNA by allele specific
amplification (ASA) analysis using the primer pairs and PCR
conditions previously described [5]. The mutation was not detected
in the parents’ peripheral blood DNA, indicating that P25L occurred
as a de novo event, probably originating in the germline cells (
(figure 1E) ).
Paternity was therefore confirmed by comparison of five informative
polymorphic markers (HLA DQα, D7S460, ACTBP2, D1S80, HUMTH01)
[6].
Discussion
Our case represents the first Italian report of EBS-MP due to the
P25L mutation in KRT5 and extends the limited number (only eleven
unrelated families) of cases described so far in which this
mutation has been found [1, 4, 5, 7-11]. Four reports, including
ours, were European sporadic cases; the mutation resides in a CpG
site, within the codon 25, that is known to cause increased
susceptibility to C→T transition, hence providing an explanation
for this recurrent sequence variation [5, 8, 10]. Identification of
the same mutation in an EBS-MP family of different ethnic
background [9] adds further evidence that codon 25 represents a
“hotspot” for mutation in the KRT5 gene. Recently, a Japanese
family with EBS-MP has been shown to carry a frame shift mutation
(1649delG) in the tail domain (V2) of keratin 5, indicating that
this peculiar phenotype could also result from a different mutation
outside the V1 domain of keratin 5 [12]. This finding suggests that
the presence of the 1649delG mutation in exon 9 of KRT5 should
always be checked in the absence of the P25L mutation in patients
affected with EBS-MP.
The molecular mechanism leading to the unusual phenotype of
EBS-MP has been investigated [4, 7]. There is in vitro evidence of
a role of the amino-terminal end of keratin 5 in which the P25L
occurs, in the elongation and lateral alignment of K5/K14
heterodimers leading to the assembly of higher-order IF [4, 13].
The identification of the 1649delG mutation in the V2 domain of K5
in a family with EBS-MP further emphasises the role of nonhelical
head/tail domains in this process. In addition, a direct
interaction between a conserved 18-amino acid residue stretch in
the K5 head and the C-terminal end of desmoplakin 1 has been
reported, suggesting that the P25L mutation could also alter the
binding between keratin IF and desmosomes [14]. With regard to the
EBS-associated pigmentation, it has been suggested that
perturbation in the keratin IF assembly may influence either the
longevity of melanin granules in basal cells or, alternatively, the
efficiency of melanosome transfer to melanocytes [4, 7]. This could
result in the anomalous skin pigmentation that represents the
peculiar feature of EBS-MP.
Concerning the differential diagnosis, there are several
similarities between EBS-MP and KS, a disorder known to result from
recessive mutations in the KIND1 gene encoding for kindlin-1, a
novel keratinocyte focal contact protein [15]. EBS-MP patients can
show some clinical, immunological and ultrastructural features also
consistent with KS, i.e. the appearance of reticular
hyperpigmentation and skin atrophy, cleavage through the basal cell
cytoplasm and labeling of hemidesmosomal proteins at the blister
floor in immunomapping analysis. Misinterpretation of the diagnosis
in a patient with KS, who had been initially considered as having
EBS-MP, has been reported [16, 17]. A parallel between these two
disorders is provided by the function of the proteins involved,
both participating in the linkage between the cytoskeleton and the
cell membrane: kindlin-1 connects the actin cytoskeleton to the
focal contacts on the keratinocyte cell membrane, and keratin 5
forms, together with keratin 14, the keratin filaments, which are
attached to the cell membrane via the hemidesmosomes. In the case
presented here, KS was also considered in view of the apparent
recessive mode of inheritance, of the presence of pigmentary
changes and the exclusive acral localization of the blisters.
Although re-duplication of the basement membrane, a unique
ultrastructural finding in KS [18], was not observed in our case, a
definite diagnosis of EBS-MP was only established following genetic
analysis which revealed the de novo occurrence of the causative
mutation in KRT5. These findings illustrate the usefulness of
molecular analysis, in addition to clinical, immunological and
ultrastructural examinations, for a correct diagnosis of rare
genodermatoses.
Acknowledgements
The authors thank R. Fusari and A. Bucci for skilful technical
assistance, and the SIM (Servizio Immagini Mediche) for artwork.
This work was supported by grants from the Italian Ministry of
Health and Istituto Superiore di Sanità (no. 526/A29).
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