ARTICLE
Lichen planus-like keratosis (LPLK) usually consists of a solitary, nonpruritic
papule or slightly indurated plaque [1] on the trunk and upper extremities
of adults older than the fifth decade [2], and is considered to be the
inflammatory stage of regressing solar lentigines and reticulated seborrheic
keratosis [3, 4]. The main histological feature of LPLK is a band-like
lymphocytic infiltrate at the dermal-epidermal junction with necrotic
keratinocytes [1]. Some authors have stressed the presence of parakeratosis
[4, 5], plasma cells, and eosinophils, and also the association with solar
lentigo for the histological diagnosis of LPLK [4]. However, since all
cases do not always show the above findings, the clinical history of a
solitary, nonpruritic lesion is crucial for distinguishing LPLK from lichen
planus [1]. In this report, we describe a rare and interesting case of
LPLK with an autoantibody to the epidermal side of the basement membrane
zone (BMZ) demonstrated by immunofluorescence studies.
Case report
A-67-year-old man who had a solitary dark brown macule on the left breast
without pain and itching for more than 6 years was admitted to our Department
on April 1, 1998. He had not received any drugs except aspirin for the
common cold about 2 years before. The eruption was a slightly elevated
and round macule with a diameter of 17 mm with a small erosion (Fig.
1). No involvement was observed on the skin elsewhere or on mucous
membranes and Nikolsky's sign was negative. Laboratory data were all within
normal limits. The macule was removed by simple resection. Histological
findings showed hypergranulosis, irregular acanthosis, liquefaction degeneration
of basal cells, band-like infiltration of lymphocytes in the subepidermal
portion, and a cleft at the basement membrane zone (Fig.
2A) showing a subepidermal blister (Fig.
2B). The eruption has not recurred by November 19, 1998.
Immunofluorescence study
Biopsy samples for direct immunofluorescence (DIF) were obtained from
the perilesional skin which appeared to be clinically normal. Prior to
the biopsy, the patient was informed about the aim and the methods of
this study including the extra biopsy from the perilesional skin and gave
written consent.
Normal skin samples for indirect immunofluorescence study (IDIF) were
from the normal skin adjacent to the nevus cell nevus of other patients.
Tissue specimens were embedded in OCT compound and stored at 80°
C. In DIF, cryostat 4-µm sections were rinsed with PBS and then reacted
for 45 min with fluorescence-conjugated goat antibodies against human
IgG, IgA, IgM, C3 and C1q. In IDIF, cryosections on the slides were rinsed
with PBS at room temperature and reacted with various dilutions of the
patient's serum or normal control serum for 45 min. After washing with
PBS, the sections were incubated for 45 min at room temperature with fluorescence-conjugated
goat anti-human IgG, IgA, IgM. The sections for IDIF or DIF were washed
with PBS and covered with the glass coverslips. Observation was carried
out using a fluorescence microscope, Nikon FX (Nikon, Tokyo, Japan). Direct
immunofluorescence of perilesional skin showed a linear deposition of
IgG, but not IgA and IgM, C3 and C1q at the basement membrane zone (BMZ)
(Fig. 3A). Indirect
immunofluorescence findings of serum obtained on April 1, 1998 using normal
skin as a substrate, showed a linear deposition of IgG at BMZ (Fig.
3B), and the antibody titer was at 1:80. To study the localization
of immunoreactants in detail, we performed IDIF study using normal tissue
specimens incubated in 1 M NaCl solution for 48 hrs at 4° C to separate
the epidermis and dermis. The positive staining was observed on the epidermal
side of skin split by 1 M NaCl. Indirect immunofluorescence study
using the patient's serum on November 19, 1998 showed negative. Two other
sera obtained from two patients with LPLK revealed negative staining by
indirect immunofluorescence.
Immunoblot analysis
Extracts from normal human epidermis and normal human dermis were fractionated
by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE)
using 6% gel according to the standard method [6, 7]. Separated proteins
were electrophoretically transferred onto nitrocellulose membrane. After
blocking with 5% skimmed milk in 0.2% Tween-20 in phosphate buffered saline
(PBS), the membrane was incubated with 1:100 dilution of the patient's
serum overnight at 4° C. Specific reactions by the serum were detected
with enhanced chemiluminescence (ECL) Western blotting detection reagents
(Amersham International, Buckinghamshire, UK).
The patient's serum reacted with a polypeptide with a molecular weight
of 230 kDa in epidermal extracts (Fig.
4), but not with 180 kDa polypeptide even in 1:10 diluted patient's
serum, whereas the serum from other patients with typical bullous pemphigoid
reacted with either 180 kDa or 230 kDa polypeptides. The sera did not
react with the dermal extracts (Fig.
4).
Discussion
The current case can be diagnosed as LPLK with antibody to 230 kDa bullous
pemphigoid antigen (BPAG1) from clinical and histological findings, immunofluorescence
studies and immunoblot analysis. Although the blistering was not clinically
observable, the blister was histologically demonstrated at the BMZ. However,
several other possible diagnoses such as a localized type of lichen planus
pemphigoides (LPP), a localized type of bullous lichen planus, and the
coexistence of LPLK and localized BP, can not be ruled out. The case reported
here is not bullous lichen planus, because plural skin lesions commonly
exist and direct and indirect immunofluorescence is negative in bullous
lichen planus except in some cases [8].
LPP is characterized by bullae arising on lichen planus papules and
on clinically uninvolved skin, subepidermal bullae in histopathology,
and linear deposits of IgG and C3 along the BMZ on direct immunofluorescence
of peribullous skin [9]. Recent immunoblotting data revealed that antibodies
to BPAG1 and/or BPAG2 were detected in the patients' sera [9-12], and
also the antibody to 200 kDa antigen [12]. The most appealing hypothesis
to explain LPP compatible with the clinical and biological findings was
proposed by Stingl and Holubar [13]. Deposition of immunoreactants at
the dermoepidermal junction could result from the damage of the BMZ through
the lichenoid inflammatory process, unmasking previously unexposed antigens
or creating new antigens, which can result in the generation of circulating
antibodies. The bullae would be formed as a secondary phenomenon, as happens
in bullous lupus erythematosus. The bound and circulating antibodies to
BMZ have also been found in cases of apparently non-bullous lichen planus
[14, 15].
In our case, the fact that the antibody to BMZ was negative 7 months
after the resection of the lesion suggests that the antibody to BPAG1
is generated because the basal cells are continuously injured.
Immunoblot analysis revealed the presence of
antibody directed to 230 kDa polypeptide which is considered to be 230
kDa BPAG1 in the patient's serum. Bullous pemphigoid is an autoimmune
blistering disease which is characterized by the presence of circulating
autoantibodies to two major hemidesmosomal molecules, BPAG1 and BPAG2
[16-19]. Recent molecular genetic studies on these antigens demonstrated
that BPAG1 is an intracellular plaque protein of hemidesmosome, which
is a member of the plectin family [20-22], and that BPAG2 is a transmembrane
collagenous protein [23, 24]. It is now widely accepted that antibody
to BPAG2 is the most possible pathogenic agent for BP, because a blistering
animal model that mimics BP has been generated by the injection of antibody
against noncollagenous domain (NC16a) of BPAG2 to mouse [25]. However,
antibodies to BPAG2 are present at lower levels than those to BPAG1 in
BP sera [26, 27].
Localized bullous pemphigoid is a rare variant of BP, which shows tense
bullae at limited sites [28]. Localized BP sera react with BPAG1 [29-31]
in epidermal extracts more frequently compared with BPAG2 [32]. Antibody
to BPAG2 is detected less frequently in localized BP than in generalized
BP. These results suggest that antibody to BPAG1 is more crucially related
to localized BP than to generalized BP [30]. The mechanism by which antibody
to BPAG1, which is an intracellular protein, forms the blister, is unknown.
When the basal keratinocytes are injured, the BPAG1, which may possess
a higher antigenicity because it is an intracellular protein, is exposed
to the immune system in a large quantities, so that a sufficient immune
complex may be generated to cause the inflammation and blister formation,
as suggested in case of generalized BP [33, 34]. Although there was only
one lesion in the present case, it is likely that the blister may have
been formed by a similar mechanism as a secondary phenomenon because the
lesion had existed for at least 6 years.
Immunoblot analysis using the patient's serum only showed the presence
of the antibody to BPAG1, but not BPAG2. Since we could not check the
reactivity of the serum to the recombinant protein of BPAG2 NC 16a domain,
we cannot completely deny the possibility that the patient's serum does
not contain enough antibody titer to detect BPAG2 by immunoblot using
epidermal extracts. This possibility is thought, however, to be very unlikely,
because there would have been generalized blisters, if such an active
antibody against BPAG2 NC 16a domain were formed in the patient's sera.
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