Dietary survey and patch testing for tannic acid and allyl compound in Pemphigus vulgaris and Bullous pemphigoid patients

Authors: MM Dhandha, N Burkemper, R King, MY Hurley

Corresponding Author:

Maulik Manharlal Dhandha

maulikdhandha@gmail.com

Abstract:

Pemphigus Vulgaris (PV) and Bullous Pemphigoid (BP) are rare autoimmune skin blistering disorders that can involve mucous membranes. Previous studies have shown a possible association of diet containing allyl compounds (garlic, leek, onion etc.) with acantholysis. Similarly it has been shown that tannic acid containing food products (cassava, red chilies, tea etc.) can also provoke cytotoxic & acantholytic effects. Our study explores whether type IV hypersensitivity reaction to diet containing diallyl disulfide or tannic acid is involved in the pathogenesis of these disorders. Our survey study of 61 patients with 20 responses – 10 PV and 10 BP showed significantly more frequent intake of tannic acid based diet in PV compared to BP patients prior to diagnosis. No such difference in allyl compounds containing diet was observed. Patients with PV reported significantly decreased frequency of allyl compounds based diet post vs pre-diagnosis. No such difference in BP was observed. 4 patients from the survey (2 PV and 2 BP) completed patch testing for - Tannic Acid – diluted at 0.25% in distilled water and Di-allyl-disulfide – 1% pet. No reaction to either compounds was noted in the 4 patients tested. Our study shows that of the limited number of patients tested, type IV hypersensitivity to allyl compounds or tannic acid does not impact PV or BP patients. Future studies with larger patient population may provide further insight into the pathogenesis of PV or BP

Keywords:

Pemphigus vulgaris, bullous pemphigoid, ally compound, tannic acid, type IV hypersensitivity, patch testing, diet

Introduction:

Pemphigus vulgaris (PV) and Bullous pemphigoid (BP) are rare autoimmune skin blistering disorders that can involve mucous membranes [1]. Susceptibility to autoimmune diseases is multifactorial. Several genetic and environmental etiologic factors have been postulated. Especially in pemphigus the variability in the incidence as well as age of onset of the disease cannot be explained by genetic factors alone and hence environmental factors are implied [1]. Food could thus be a potential environmental trigger [2-9]. Previous studies have shown possible association of diet containing allium group compounds (garlic, leek, onion etc.) with acantholysis in in-vitro models [2,3]. This reaction is more prominent in samples taken from a donor with pemphigus prone antigen (HLA-DR4) [3]. Similarly it has been shown that Tannic acid containing food products (like cassava, red chilies, tea etc.) can provoke cytotoxic and acantholytic effects [3]. No such dietary associations have been shown for BP [10]. Other compounds reported to induce pemphigus include – phenols [11].

Food products in the allium group (garlic, leek and onion) contain thiol compounds (allylmercaptan, allylmethylsulfide and allyl sulfide) as well as diallyl disulfide. It is well known that thiol group containing drugs can induce pemphigus [11,12]. Proposed mechanisms include: inhibition of enzymes that aggregate keratinocytes [13], activation of enzymes like plasminogen activator which disaggregate keratinocytes [13], disturbance to cell adhesion by formation of thiol-cysteine bonds instead of cysteine – cysteine bonds and/or formation of neo-antigen by an immunological reaction [13]. In addition, diallyl disulfide (a component of garlic) is also known allergen to cause garlic allergy [14].

Tannins (found in cassava, red chilies, tea etc.) can affect the pathogenesis of pemphigus vulgaris in several possible ways including but not limited to – precipitating interaction with proteins, interaction with medications, release of neutrophilic chemotactic factor, alveolar macrophage stimulation, platelet activation, enzyme inhibition and major ion deprivation [4].

In contrast, nicotine has anti-acantholytic effects. Thus, smoking is not contraindicated in pemphigus [4,10]. Furthermore, nicotinamide is frequently used for treatment of bullous pemphigoid [15].

PV and BP both involve autoantibodies that bind to antigens on the patient’s own cell surfaces. In the case of PV, the IgG autoantibodies bind to desmosomal antigens desmoglein 1 and desmoglein 3 on the cell surface of keratinocytes resulting in acantholysis and intra-epithelial blisters [16-19]. BP involves IgG autoantibodies that bind to BP 230 (BP Ag1) and BP 180 (BP Ag2) in the skin basement membrane which activates the complement system and inflammatory mediators resulting in sub-epidermal blistering [20-29]. Patch testing studies type IV hypersensitivity reactions – allergic contact hypersensitivity. CD8+ helper T cells recognize low molecular weight allergens complexed with host proteins via haptenization on the surface of antigen presenting cells and induce an immune response [30]. It would however be difficult to know if Allergic Contact Dermatitis leads to skin breakdown and hence exposing desmosomal and/or hemidesmosomal protein antigens to immune system leading to autoimmune diathesis or autoimmune diathesis is causing allergen sensitivity in these immunobullous disorders. The purpose of our study was to explore whether delayed type hypersensitivity reaction to diallyl di-sulfide (allergen in garlic) and tannic acid could be involved in pathogenesis of PV or BP.

For this, we designed a two step study – 1) Survey study to identify dietary and smoking habits of patients with pemphigus vulgaris or bullous pemphigoid, 2) Patch testing for tannic acid and allyl compound in pemphigus vulgaris and bullous pemphigoid patients.

Patient population

Both the survey study and the patch test study were approved by the Saint Louis University Institutional Review Board. The survey population consisted of patients from a tertiary care center practice – Department of Dermatology at Saint Louis University. The survey included patients seen at Saint Louis University Department of Dermatology from 2009 to 2014. Using the software IDX for patient scheduling, search terms “Pemphigus”, “Pemphigus Vulgaris”, “Bullous” and “Bullous Pemphigoid” were used to identify patients for the study. 241 patients were identified. Review of electronic health records for these patients was then done. 180 patients were excluded due to other diagnosis including pemphigus foliaceous, mucous membrane pemphigoid, cicatricial pemphigoid, paraneoplastic pemphigus, IgA pemphigus. Nursing home patients and patients with dementia were also excluded due to difficulty of contacting patients for the survey. Patients with no confirmatory diagnosis were excluded from the study as well. Of the remaining 61 patients, 59 were contacted by phone call at least twice if no reply the first time and two patients were asked about taking the survey at the end of a clinic visit. A total of 20 patients were surveyed – 10 with pemphigus vulgaris and 10 with bullous pemphigoid.

Methods:

Survey Study

The following survey questions were asked by phone or at the end of clinic visit (Table 1)

Patch Study

Of the twenty patients surveyed, six patients agreed to undergo patch study. Four (two with diagnosis of PV and other two with BP) completed the study. Patch testing was done over the course of five days (Monday through Friday). Written consent after explaining the risks, benefits and concerns related to the study was obtained from patients prior to the study. The following compounds were tested – 1) Tannic acid – diluted at 0.25% in distilled water – Sigma Aldrich (to avoid severe reaction), 2) Di-allyl-sulfide 1% pet (from Dormer laboratories). These compounds were first poured in Finn chambers (Allerderm laboratories) and were then adhered to Scanpor tape (Allerderm laboratories). These patches were then applied to the patient’s back and then reinforced with more Scanpor tape. A map of where the compounds were placed was created. Patients were then sent home with instructions to keep the back dry and the patches secured until the second visit at 48 hours. Patients were told to avoid excessive sweating or heavy exercise, to avoid loosening and detachment of the patches. Patients were subsequently seen in clinic at 48 hours on Wednesday. Contact patches were removed and sites checked for reaction. Any positive reaction was scored according to the International Grading System [31] (Table 2). The patients were then asked to keep the back dry until the 72 hour reading on Friday. On Friday, positive reactions if any were graded according to the standard system mentioned before.

Statistics

Correlational and descriptive analysis were done using Vassar stats software. P value was significant at < 0.05.

Results:

Our findings show that PV patients ate diet containing tannic acid significantly more frequently compared to BP patients prior to diagnosis (p = 0.0082, t=2.87, df=18) (figure 1). No significant difference was found between the PV and BP patients after the diagnosis ((p=0.2733, t=1.13, df=18). In addition, no difference in the diet containing tannic acid was found before compared to after diagnosis of PV (p=0.1122, t=-1.67, df=18) or BP (p=1.0, t=0, df=18).

No significant difference between the PV and BP groups in allyl compounds containing diet before diagnosis of PV or BP (p=0.5691, t=0.58, df=18) was found. Similarly, no significant difference was noted between PV and BP group post diagnosis in diet containing allyl compounds (p=0.0778, t=-1.87, df=18). No significant difference in the diet containing allyl compounds was found before compared to after diagnosis of BP (p=0.7452, t=0.33, df=18) groups. We however noted significantly decreased dietary intake of diet containing allyl compounds ((p=0.0336, t=-0.23, df=18) after diagnosis of PV. This may be more from subjective reporting due to pain from mucosal erosions/ulcerations (figure 2)

About 50% of BP patients were “likely” to change diet if studies showed dietary associations to disease flares. 30% were somewhat likely and 20% were unlikely to change diet. On the contrary in PV patients about 80% were likely to change diet, 20% were somewhat likely and 0% were unlikely to change the diet. The difference may be due to age difference in PV or BP patient populations (figure 3).

Smoking

About 60% of PV patients reported smoking before the diagnosis of PV and another 40% reported rare/no use. In BP, 40% reported smoking cigarettes before the diagnosis and another 40% reported rare/no use. Data from smoking was limited because of recall bias and majority of patients not recalling timeline for smoking in relation to onset of PV or BP.

Medications

Patients had a variety of medical treatments for PV or BP. Systemic medications for PV included rituximab as well as prednisone, mycophenolate mofetil, azathioprine, methotrexate. Of the six patients on rituximab – all six reported in remission after 1-3 infusions. Topical medications for PV included clobetasol.

For BP systemic medications included prednisone, mycophenolate mofetil, azathioprine, niacinamide, methotrexate. Topical medications included clobetasol, fluocinonide, betamethasone

Of the four patients that were patch tested, all had negative reaction at 48 hours or 96 hours to tannic acid or di-allyl-disulfide compounds.

Conclusion and discussion:

Pemphigus vulgaris and bullous pemphigoid are hetergenous disorders for which the pathogenesis is not completely understood. Environmental etiologic factors like medications [11,12] and food products can play a role in induction of pemphigus [2-9]. Several medications have been implicated in the etiology of bullous pemphigoid [11,12], however no dietary associations have been studied.

From the survey study we show that patients with PV reported significantly more frequent intake of tannic acid in their diet compared to BP patients prior to diagnosis. Patients with PV reported significantly decreased frequency of allyl containing diet post diagnosis when compared to pre-diagnosis. Smoking patterns of PV and BP could not be elucidated from the limited data in our study. In our patch study, of the limited number of patients tested, type IV hypersensitivity to allyl compounds or tannic acid was not identified in PV or BP patients.

Some of the limitations of the study include 1) Small patient population, 2) Allergen diallyl disulfide is an oxidized form of sulfur. Although it is part of the Allium group of compounds, it is not a thiol compound. Repeat patch testing with another allyl compound containing a thiol/sulfhydryl group could yield different results. Future studies could provide further insights into the immune-pathogenesis of these immune-bullous disorders and the role, if any that delayed type hypersensitivity plays.

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Figure legends:

PV patients ate diet containing tannic acid significantly more frequently compared to BP patients prior to diagnosis
PV Patients had significantly decreased dietary intake of diet containing Allyl Compounds after the diagnosis
PV patients were more likely to change lifestyle compared to BP patients.

Table 1

Survey question	Answer choices (select one)
1) Which immunobullous disease have you been diagnosed with?	pemphigus vulgaris/bullous pemphigoid
2) Did you eat diet containing tannic acid (like cassava, red chilies, tea etc) before your diagnosis of PV or BP?	Daily/ Few days a week/ Weekly/ Monthly/ Very infrequent or none at all
3) Did you eat diet containing allyl compounds (like garlic, leek, onion) before your diagnosis of PV or BP	Daily/ Few days a week/ Weekly/ Monthly/ Very infrequent or none at all
4) Did you change your diet after your diagnosis of PV or BP to exclude (or decrease eating) these tannic acid containing food items. Please mark how often you eat these after the diagnosis?	Daily/ Few days a week/ Weekly/ Monthly/ Very infrequent or none at all
5) Did you change your diet after your diagnosis of PV or BP to exclude (or decrease eating) these allyl compounds containing food items. Please mark how often you eat these after the diagnosis?	Daily/ Few days a week/ Weekly/ Monthly/ Very infrequent or none at all
6) Would you consider making a dietary change to your lifestyle if indicated for your disease? (On a scale of 1 to 5 – 1 being most likely, and 5 being least likely)	Most likely/ likely/ somewhat likely/ less likely/ least likely or unlikely
7) Did you smoke before your diagnosis of PV or BP?	> 10 cigarettes a day/ 1-10 cigarettes a day/ Few days a week/ Occasionally/ Never
8) Did you start smoking after your diagnosis of PV or BP?	> 10 cigarettes a day/ 1-10 cigarettes a day/ Few days a week/ Occasionally/ Never
What is your current treatment for BP or PV?	Steroids/Immunomodulators – methotrexate, mycophenolate mofetil, cyclosporine, azathioprine/ Biologics/ IVIG/ Other – Please specify
What did your past treatment for BP or PV includes?	Steroids/Immunomodulators – methotrexate, mycophenolate mofetil, cyclosporine, azathioprine/ Biologics/ IVIG/ Other – Please specify
Would you be willing to participate in a patch test study to assess for presence of contact allergy to Allyl and/or Tannic Acid compounds?	Yes/ No

Table 2

INTERNATIONAL GRADING SYSTEM FOR PATCH TESTS
+/-	Doubtful reaction, faint macular erythema
+	Weak, non-vesicular reaction with erythema, infiltration and papules
++	Strong, vesicular reaction with erythema, infiltration and papules
+++	Spreading bullous reaction
-	Negative reaction
IR1	Irritant reaction

Figure 1

Figure 2

Figure 3

Preprint

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Dietary survey and patch testing for tannic acid and allyl compound in Pemphigus vulgaris and Bullous pemphigoid patients