|Year : 2016 | Volume
| Issue : 3 | Page : 288-294
|A clinicopathological study of pemphigus in Eastern India with special reference to direct immunofluorescence
Joyeeta Chowdhury1, Pijush Kanti Datta2, Satyendra Nath Chowdhury3, Nilay Kanti Das3
1 Department of Dermatology, Nil Ratan Sircar Medical College and Hospital, Kolkata, West Bengal; Department of Dermatology, Medical College and Hospital, Kolkata, West Bengal, India
2 Department of Dermatology, Mata Gujri Memorial Medical College, Kishanganj, Bihar, India
3 Department of Dermatology, Medical College and Hospital, Kolkata, West Bengal, India
|Date of Web Publication||13-May-2016|
Dr. Joyeeta Chowdhury
Department of Dermatology, Nil Ratan Sircar Medical College and Hospital, Kolkata, West Bengal
Source of Support: None, Conflict of Interest: None
| Abstract|| |
Background: Pemphigus is a group of chronic autoimmune vesico-bullous disorders in which the epidermis and the basement membrane zone are the focus of attack resulting in cutaneous and mucosal blister formation. Direct immunofluorescence (DIF) test is a very sensitive test for the diagnosis Aim: To study the clinico histopathological patterns of pemphigus in eastern India. The study also aims to correlate DIF with clinical and histologic findings as well as severity of skin involvement [scoring systems]. Materials and Methods: Total 41 patients were studied over a period of 1 year in the Post-graduate centre of Dermatology in Eastern India. DIF, histopathology and clinical data were correlated. Results: In our study Pemphigus vulgaris (PV) was the predominant type with 32 cases followed by 8 cases of pemphigus foliaceus (PF) and a single case of IgA pemphigus. Mean age at presentation was late middle age. Majority of the patients, 26 (63.41%) initially had cutaneous involvement followed by mucosal involvement. In this study group 36 (87.80%) patients showed acantholytic cells on histopathological examination. Most patients of PV showed suprabasal blister 20 (62.50%) followed by intraspinous 5 (15.62%) and subcorneal 5 (15.62%) blister. In majority 28 (87.50%) of the PV patients IgG and C3 antibodies were deposited throughout the epidermis. The strength of antibody positivity was strong in most of the patients (71.87%). In cases of PF mostly IgG 6 (75%) antibodies were deposited in the upper epidermis. DIF intensity had poor correlation with disease activity/severity except in PF. Conclusion: Almost 85.36% cases of pemphigus were diagnosed clinicopathologically. But 6 cases couldn't be diagnosed accurately on clinicopathological basis and in them DIF was confirmatory. Two cases of pure mucosal PV and 1 case of IgA pemphigus was confirmed by DIF. Two cases of bullous pemphigoid clinico-histologically mimicking PV were also excluded by DIF. So it appears from our study that DIF is confirmatory for diagnosis of pemphigus in all cases.
Keywords: Direct immunofluorescence, Mahajan's score, pemphigus area and activity score, pemphigus foliaceus, pemphigus vulgaris
|How to cite this article:|
Chowdhury J, Datta PK, Chowdhury SN, Das NK. A clinicopathological study of pemphigus in Eastern India with special reference to direct immunofluorescence. Indian J Dermatol 2016;61:288-94
|How to cite this URL:|
Chowdhury J, Datta PK, Chowdhury SN, Das NK. A clinicopathological study of pemphigus in Eastern India with special reference to direct immunofluorescence. Indian J Dermatol [serial online] 2016 [cited 2021 Nov 28];61:288-94. Available from: https://www.e-ijd.org/text.asp?2016/61/3/288/182422
What was known?
- Pemphigus vulgaris (PV) is a disease of middle age
- Mucosa is involved initially in most of the cases PV
- There is a correlation of indirect immunofluorescence (IIF) with severity of involvement, and a recent report has also documented that IIF is also unreliable in situations.
| Introduction|| |
Pemphigus is a rare group of blistering autoimmune diseases affecting the skin and mucous membranes, where autoantibodies are directed against the cell surface of keratinocytes. The loss of cell–cell adhesion of keratinocytes occurs through a process called acantholysis. The term “pemphigus vulgaris” (PV) was coined by Boissier de Sauvages in 1750. The incidence of pemphigus is very low varying from 0.5 to 3.2 cases per 1,00,000 population per year.. It is most common in the fourth and fifth decades and equally affects males and females.. Pemphigus is classified into 5 five types, namely  PV and its variant pemphigus vegetans, pemphigus foliaceus (PF) and its variant pemphigus erythematosus, its endemic variant fogo selvagem, drug-induced and drug-triggered pemphigus, IgA pemphigus and paraneoplastic pemphigus. PV is characterized by oral blister and erosions in 50–70% of the cases along with flaccid skin blisters, especially over face, scalp, axillae, sternum, groin, and pressure points. Here, blister appears in the suprabasal zone.
PF occurs in middle-aged persons and presents with flaccid bullae over an erythematous base or as scaly crusted erosions without any blister formation. Here, the cleft is in the superficial epidermis, especially in the granular layer.
The target antigens in pemphigus are against the desmosomal components that lead to acantholysis. Mainly IgG and C3 antibodies are deposited in the pemphigus group except in IgA pemphigus  The diagnosis of pemphigus should be prompt as clinically it may be difficult to differentiate the different variants leading to delay or wrong treatment thus raising the mortality and morbidity rates. Direct immunofluorescence (DIF) test is a very sensitive test for the diagnosis. Indirect immunofluorescence (IIF) is widely used for predicting the disease severity, but a recent report has suggested it is unreliable at situations.
A lot of studies have been carried out on the clinico-histopathological correlation with DIF in different parts of India, but there is paucity of such studies in Eastern India. The study also explored the possibility of using the DIF intensity for predicting the disease activity.
| Materials and Methods|| |
The study was carried out among patients attending the Dermatology Department of a tertiary care center in Eastern India over a period of 1 year from May 2010 to April 2011. The study was approved by the Institutional Ethics Committee, and informed consent was taken in all patients. Forty-six clinically suspected cases of pemphigus were screened. Patients with flaccid blisters especially over face, scalp, axillae, groin, and pressure points with mucosal erosions, and positive Nikolsky sign were clinically suspected with PV. Disease activity/severity was judged using the pemphigus area and activity score (PAAS), Mahajan's score, and body surface area (BSA) involvement.
All the cases were subjected to histopathology and DIF, and final diagnosis was reached on the basis of clinical, histopathological, and immunofluorescence patterns. DIF intensity was correlated with the disease activity scores.
A detailed history, clinical profile, histological, and immunofluorescence pattern were collected in a predesigned and pretested case record form. Chi-square test, Fisher's exact tests, Spearman's correlation coefficient, inter-rater agreement (κ) test were used as applicable. Medcal ® version 10.2.0.0 (http://www.medcalc.be) was used for statistical analysis, and P < 60; 0.05 was considered statistically significant.
| Results|| |
Among 46 screened patients, three patients did not give consent, and two patients were finally diagnosed as bullous pemphigoid by DIF and were excluded from the final analysis. A total of 41 patients of pemphigus were evaluated. Of the 41 cases of pemphigus evaluated, PV was the predominant type with 32 cases (78%) followed by 8 cases (19.5%) of PF and a single case of IgA pemphigus. There was no case of paraneoplastic pemphigus or drug-induced pemphigus. There was an overall female preponderance (male:female ratio of 14:27) with age at presentation found to range from 16 to 76 years (mean 49.48 ± 16.51 years). Of the 41 cases of PV, 6 cases (18.75%) were between 21 and 40 years, 15 cases (46.87%) between 41 and 60 years, 9 cases (28.12%) were above 60 years, and 2 cases (6%) were below 20 years of age. Of the 8 cases of PF, 2 patients (25%) each were in the age group of 21 and 40 and 41–60 years, whereas 3 (37.5%) patients were above 60 years. Mean age at presentation suggests it to be a disease of the late middle age.
Duration of the illness at presentation in pemphigus group ranged from 0.16 to 108 months (mean ± standard deviation = 8.21 ± 17.96) [Table 1]. Majority of the patients 36 (87.80%) had symmetrical distribution of the lesions. Nineteen (46.3%) patients of this study had both skin and mucosal involvement. There were 2 patients (4.9%) who had only mucosal involvement and both of them were clinically diagnosed as PV. Majority of the patients, 26 (63.41%) started with cutaneous involvement mucosal lesions were present in 63.41% (26 patients) of all cases. Of them, 20 (48.78%) had oral mucosal and 6 (14.63%) had involvement of nasal and genital mucosa.
The extent of skin involvement was mainly divided into 3 groups (BSA <10%, 11–30%, and > 30%). Most of the patients, 28 (68.3%) had a widespread skin involvement (BSA >30%). Nikolsky's sign in this study was found to be positive in almost 90.24% cases. Systemic involvement, especially of the gastrointestinal tract and genitourinary system, was found in 14 (34.14%) patients. Histopathological picture showed variable findings. In this study group, 36 (87.80%) patients showed acantholytic cells on histopathological examination. This was statistically significant, P = 0.024 [Table 2]. Dermal inflammation, consisting mostly of neutrophils, was present in 95.12% patients of pemphigus [Table 3].
Seventeen (53.12%) of PV patients had both skin and mucosal involvement. Eighteen (56.25%) of PV patients had lesions in oral mucosa [Table 1]. Among 32 patients of PV, in 19 (59.37%) patients the disease started with cutaneous lesions followed by mucosal involvement later; in the rest, cutaneous involvement was noted following mucosal involvement. Majority of the PV patients 27 (65.9%) presented with flaccid vesicles and bullae followed by only vesicles (19.5%), and concomitant pustules (4.9%). Nikolsky's sign was in 29 (90.62%) cases. Systemic involvement was seen in 13 (40.62%) cases of PV. Eighteen (56.25%) of PV patients had lesions in oral mucosa [Table 1]. Most of the patients of PV showed suprabasal blister 20 (62.50%) followed by intraspinous 5 (15.62%) and subcorneal 5 (15.62%) blister. Row of tombstone appearance of basal layer was present in 17 (53.12%) of PV patients. In majority of the PV patients, IgG + C3 antibodies were deposited throughout the epidermis 28 (87.50%) followed by deposition in lower epidermis 3 (9.3%). The strength of antibody positivity was strong in most of the patients [23 (71.87%)].
Two (25%) of PF patients had both skin and mucosal involvement. Two (25%) of PF patients had lesions in oral mucosa [Table 1]. More number of PF patients (87.50%) showed extensive skin involvement compared to PV patients (65.62%) [Table 1]. In PF, blisters arising on erythematous skin were seen in 2 cases (25%), crusted lesions and erosions in 6 cases (75%). Nikolsky's sign was positive in all the cases of PF. Systemic involvement (renal failure) was found in one (12.50%) patient who succumbed within 1 week of admission. Two (25%) of PF patients had lesions in oral mucosa [Table 1]. In PF, 3 (37.5%) patients showed subcorneal vesicle. Dyskeratosis was found in one patient of PF. In cases of PF mostly IgG 6 (75%) antibodies were deposited in the upper epidermis. The strength of deposit (DIF intensity) was strong in 7 (87.5%) of the cases [Table 4]. DIF intensity was correlated with the different activity/severity scoring systems used in pemphigus (PAAS, Mahajan's score, and BSA) and it was noted that there was poor correlation between the different scores and DIF intensity except for BSA score in PF (r = 0.659, P = 0.07) [Table 5]. Inter-rater agreement (kappa test) between Mahajan's score and PAAS revealed a poor agreement (κ = 0.188) both for PV (κ = 0.198) or PF (κ = 0.06).
|Table 4: Direct immunofluorescent findings in patients with pemphigus and its variants|
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|Table 5: Correlation of direct immunofluorescence intensity with clinical parameters|
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From this study, it is seen that almost 85.36% (35 of 41) cases of pemphigus were diagnosed clinicopathologically. However, six cases could not be diagnosed accurately on clinicopathological basis and in them DIF was confirmatory. Two cases of pure mucosal PV [Figure 1]a,[Figure 1]b,[Figure 1]c and 1 case of IgA pemphigus [Figure 2]a,[Figure 2]b,[Figure 2]c were confirmed by DIF. Two cases of bullous pemphigoid [Figure 3]a,[Figure 3]b,[Figure 3]c,[Figure 3]d were considered as PV on the basis of clinico-histopathology, but they were correctly diagnosed by DIF. Hence, it appears from our study that DIF is confirmatory for the diagnosis of pemphigus in 100% cases [Table 6].
|Figure 1: (a) Pure mucosal pemphigus vulgaris. (b) Inconclusive histology of mucosal pemphigus. (c) Direct immunofluorescence showing IgG deposition in epidermis|
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|Figure 2: (a) Flaccid vesicles over upper and lower limbs. (b) Histopathology showing intra-epidermal blister in × 400. (c) Direct immnunofluorescence showing IgA deposits in epidermis|
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|Figure 3: (a) Multiple flaccid and tense bullae in a generalized pattern. (b) Mucosal erosions in the same patient. (c) Histopathological examination from bulla in × 400 showing intra-epidermal split. (d) Direct immunofluorescence showing linear IgG deposit in basement membrane zone|
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|Table 6: Comparison of histopathological and direct immunofluorescence finding in patients with pemphigus and its variants|
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A 50-year-old female with history of buccal ulceration for 9 years was clinically diagnosed as oral pemphigus, but histology was not supportive. DIF showed intra-epidermal IgG deposition in fish net pattern. The limitation is added: IIF to assess anti-desmoglein titer could not be done.
An 18-year-old girl with painful oral and labial ulceration for 7 days was clinically diagnosed with herpes simplex or adverse drug reaction. Did not heal after a course of acyclovir and stopping of all other drugs. Histology was inconclusive. However, DIF revealed it to be PV [Figure 1]a,[Figure 1]b,[Figure 1]c.
A 45-year-old male presented with flaccid bullous eruption over both upper and lower limbs only for 15 days. Clinically, a phototoxic bullous eruption and PV were kept as differential diagnosis. Histopathology revealed only an intraepidermal blister. DIF showed IgA deposit in epidermis [Figure 2]a,[Figure 2]b,[Figure 2]c.
A 42-year-old female presented with flaccid vesicles on her upper limbs only for 10 days. Histopathology showed subcorneal vesicle. DIF showed IgG deposition in epidermis.
A 39-year-old male presented with scaly erythematous lesion on his cheeks for last 2 months with no mucosal involvement. Clinically it was diagnosed as lupus with irritant contact dermatitis. Histopathology revealed inflammatory infiltrate in dermis. DIF showed IgG deposit in upper epidermis without any other deposits was finally diagnosed as PF [Figure 4]a,[Figure 4]b,[Figure 4]c.
|Figure 4: (a) Erythematous scaly lesions over cheeks. (b) Histopathology showing inflammatory infiltrate in dermis in × 400. (c) Direct immunofluorescence showed IgG deposit in upper epidermis|
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| Discussion|| |
Age at presentation in pemphigus was found to range from 16 to 76 years (mean 49.48 ± 16.51 years). Most of the patients were between 41 and 60 years of age and almost 29.26% cases were above 60 years. Thus, the age of onset of our study group was a decade older than studies done at Libya, and the trend is shifting towards older age group.,, Furthermore, there were four patients of PV, who were above 75 years age. In a recent study by Ingen-Housz-Oro et al ., immunobullous disorders have been reported in elderly patients in the age group ranging from 70 to 96 years with a high mortality rate within 12 months of onset.
In our study, female predominance was noticed. Female: male ratio was 1.9:1. In PF group, the sex ratio was equal. However, in PV females outnumbered males (male:female = 1:2.2). This was different in comparison to a study done by Wu et al ., where sex ratio was equal and also from some Indian studies where the males predominated over females. It was similar to a study done in Libya where the male:female ratio was 1:4.7.
Of the 41 cases of pemphigus studied, PV was the predominant type with 32 cases (78%) followed by 8 cases (19.51%) of PF and 1 case (2.4%) of IgA pemphigus. This was in accordance with studies elsewhere, for example, by Arya et al ., where 43 of 70 (61.42%) cases were PV and 25 (35.71%) were PF. Most of the other studies also revealed similar results.,, The most probable reason for this may be that PV being a more morbid condition is being referred more to hospital than PF.
PV showed initial lesions involving mucous membranes in 40.62% cases. Mucosal involvement at some point of the disease was seen in 26 (63.41%) patients. Out of them, 2 (6.25%) had only mucosal involvement and 13 had only cutaneous involvement throughout the course of their disease. This change in the evolution of the disease of PV may represent a paradigm shift in the clinical manifestation of pemphigus in our study population. This necessitates further evaluation of anti-Dsg1 and anti-Dsg3 by IIF. It can also be hypothesized that more common initial cutaneous involvement (instead of mucosal involvement) may be the result of mild mucosal erosions going unnoticed. This was different from results of the study done by Valand and Krishna, where more than 53% patients had mucosal involvement initially. It was also different from the study done by Rao et al ., where there was mucosal involvement in 67% cases. On the other hand, in PF, 25% of PF patients had both mucosal and skin involvement and 75% cases had only skin and involvement. Distribution of skin lesions showed common sites of involvement as noted in previous studies on PV. Of the 32 cases of PV, 20 cases (62.5%) showed intra-epidermal suprabasal vesicles and 5 cases (15.62%) showed mid-epidermal vesicles. Mid-epidermal vesicles were seen in old bulla, due to regeneration of the cells from the floor of the bulla. Acantholysis was seen in 29, i.e., 90.62% cases. In our study, histopathology of patients with pure mucosal cases revealed no blister. Dyskeratosis, basal layer budding, and pseudoepitheliomatous proliferation were not seen in any of the cases. An inflammatory infiltrate was present in the bulla cavity in 23 cases (53.5%). A row of tombstone appearance was seen in 17 (53.12%) cases. Eosinophilic spongiosis was seen in 1 case. Dermal inflammation with chronic inflammatory cells was present in 31 (96.87%) cases. Four cases (12.5%) could not be diagnosed on the basis of the above histological features (including two pure mucosal). This was similar to the study of pemphigus group by Kabir et al .
Of the 8 cases of PF studied, 7 cases (87.50%) showed acantholysis. Subcorneal bulla was seen in 3 cases (37.50%). Basal layer budding, and pseudoepitheliomatous proliferation were not seen in any of the cases. An inflammatory infiltrate was present in the bulla cavity in four cases. Eosinophilic spongiosis was seen in three cases (6.9%). In two cases (18.6%), the epidermis was lost during the process of preparation of sample for hematoxylin and eosin staining.
DIF study from perilesional skin showed mainly IgG and C3 deposits. In our study, 19 of 32 patients of pemphigus had intra-epidermal deposition of intercellular IgG and C3 both, 13 had only IgG deposition. Six cases of PF showed IgG deposition.
In PV, 28 of 32 (87.5%) cases showed antibody deposit throughout the epidermis, 9.7% cases in the lower epidermis. In 71.87% cases, the strength of deposit was moderate. In PF subgroup, 75% had antibody deposit in the upper epidermis and the strength of deposit was strong in 87.5%. The two cases of suspected oral PV are who had not showed suggestive findings in histopathology, showed intra-epidermal IgG deposits. The single case of IgA pemphigus was diagnosed by immunofluorescence and it showed moderately strong deposit of IgA antibody in upper epidermis. This was in accordance with the study conducted by Kabir et al . Two cases of bullous pemphigoid were excluded by DIF.
| Conclusion|| |
DIF intensity was found to have poor correlation with the clinical severity/activity scores; thus, it cannot be used as a biomarker for assessing the activity/severity of the disease. However, in PF, the BSA has got good correlation with DIF intensity suggesting that DIF intensity can only be used to predict the extent of skin involvement. This observation needs to be validated by cohort study though.
The study also revealed poor inter-rater agreement between the two mostly used scores for pemphigus, emphasizing that these two scores explore different aspect/parameters of the same disease and they are complementary to each other.
Declaration of patient consent
The authors certify that they have obtained all appropriate patient consent forms. In the form the patient(s) has/have given his/her/their consent for his/her/their images and other clinical information to be reported in the journal. The patients understand that their names and initials will not be published and due efforts will be made to conceal their identity, but anonymity cannot be guaranteed.
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Conflicts of interest
There are no conflicts of interest.
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What is new?
- Age distribution trend in pemphigus vulgaris (PV) is changing toward older age group
- In most cases of PV, initial cutaneous involvement was noted
- Mucosal involvement is found in cases of pemphigus foliaceus (PF) too
- DIF intensity has got no correlation with disease activity excepting for PF.
[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5], [Table 6]
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