- Email: [email protected]
Immunoglobulins G, M, A, D and E in Behcet's syndrome
237
Clinica Chimica Acta, 120 ( 1982) 237-242 Elsevier Biomedical Press
CCA 207 1
~~unog~obu~ins c. Scully u University
G, M, A, D and E in Behcet’s syndrome a**, P. Boyle b and P.L. Yap c
Department of Oral Medicine and Pathology, Glasgow Dental Hospital, G(usgow (UK), h West of
Scotland Cancer Surveillance
Unit, Ruchill Hospital, Glasgow {UK) and c Edinburgh and South-East
Scotland Regional Blood Transfusion Service, Royal Infirmary,
Edinburgh (UK)
(Received August IOth, revision November 30th‘ 1981)
summaly Serum IgG, IgM, IgA, IgD and IgE concentrations were examined in 26 patients with Behcet’s syndrome, 70 patients with recurrent aphthous stomatitis and 56 healthy controls. IgA concentrations, but not IgG, IgM, IgD or IgE, were significantly raised in Behcet’s syndrome compared with controls. Serum IgD and IgE concentrations but not IgA, IgG or IgM were significantly greater in recurrent aphthous stomatitis than in controls.
Introduction Behcet’s syndrome (BS) is an uncommon systemic disease character&d mainly by recurrent aphthous stomatitis, genital ulceration, uveitis and various systemic lesions, but there are no laboratory findings that unequivocally distinguish BS from recurrent aphthous stomatitis (R4S) occurring alone. The aetiology of BS and R4S is unknown, although for both the evidence suggests an immunopathogenesis [l-4]. Histopathological examination of the early lesions of RAS [5] and of the mucosal[5] and cutaneous [6-S] lesions of BS shows that, in addition to a lymphocytic and monocytic infiltrate, there is an increase in mast cells, and these have been implicated in the i~~opathogenesis of BS [S-10]. It has been suggested that IgE may be involved in the immunopathogenesis of BS [9]. Lymphocytes bearing IgD and IgE have been identified in the oral lesions and peripheral blood of patients with IXAS [ 1l] but have not been studied in BS. Serum IgE ~ncentrations have been examined in a small group of patients with BS [9]; serum levels of IgD do not appear to have been examined, and studies of the serum concentrations of other immunoglobulins have been somewhat equivocal; serum IgG * Correspondence to Dr. C. Scufly, Immunology Unit, Department of Oral Medicine and Pathology, Glasgow Dental Hospital. Glasgow, G2 352, UK. 0009-898 l/82/~-~/$02.75
@ 1982 Elsevier Biomedical Press
and IgM levels are usually unchanged in BS with increased IgA (12j. We have therefore estimated serum concentrations of IgG, IgM, IgA, IgD and IgE in patients with Behcet’s syndrome and recurrent aphthous stomatitis.
Materials and methods The study group of 96 adult Caucasians consisted of 26 patients with Behcet’s syndrome (median age 32 years: range 20-52: 14 males) diagnosed on the basis of recurrent oral and genital ulceration together with ocular, cutaneous, arthritic or neurological manifestations [3] and 70 patients with recurrent aphthous stomatitis (median age 25 years: range 15-37: 30 males). The control group consisted of 56 healthy subjects with no oral mucosal disease (median age 28 years: range 19-46: 25 males). Patients and controls were matched as closely as possible for a history of allergies. No patient had received systemic medication within the previous three months but most were using topical oral applications of chlorhexidine, triamcinolone or hydrocortisone.
Serum immunoglobulin estimations Sera were separated after clotting at 20°C for 30 min and overnight at 4*C, and stored in aliquots at -20°C. Serum IgG, IgM, IgA and IgE concentrations were measured in all subjects but IgD estimation could only be carried out in 39 of the controls and in 24 of those with BS. All immunoglobulins were assayed by immunodiffusion techniques, except IgE which was assayed by radio-immunoassay. Serum IgG, IgM and IgA concentrations were measured in duplicate by single radial immunodiffusion as previously described [ 121, and expressed as mg/ 100 ml (mean k standard deviation). Serum IgD concentrations were measured in duplicate by a single radial immunodiffusion technique using sheep anti S antiserum (Scottish Antibody Production Unit, Law Hospital; Carluke) and an IgD myeloma standard previously calibrated against the World Health Organisation IgD standard, and expressed as lug/l. Serum IgE concentrations were measured in duplicate by a solid phase radioimmunoassay (Phadebas IgE PRIST, Pharmacia, Sweden), and expressed as kU/l.
statistical analysis Results were analysed by ~skal-Wallis analysis of variance. IgG, IgM and IgA results were then analysed by the Dunnett test but the Mann-W~tney test was used to analyse IgD and IgE results because of the non-parametric distribution of data. Results Serum IgG, IgM and IgA concentrations are shown in TableI. Serum IgA concentrations in Behcet’s syndrome (BS) were significantly greater than in controls (p-c 0.05) but not greater than levels in recurrent aphthous stomatitis (RAS). No significant differences could be established between any groups either for serum IgG or for IgM concentrations.
239
TABLE SERUM
kG
I CONCENTRATIONS
OF MAIOR
IMMUNOGLOBULINS
IN BEHCET’S
Behcet’s syndrome
Controls
Recurrent
1690 * 428
1443*205
16611214
aphthous
IgM
1022
46
1122
53
108-c
42
IgA
386*114**
242k
63
298*
12
* Mean k SD in mg/ 100 ml. ** Significantly different from controls
SYNDROME
*
stomatitis
(p
0.20
. .
.
. : .
f . Aa
.
8
.
BS
c
Fig. I. Serum concentrations controls (C).
:
!
RAS
of IgD in Behcet’s syndrome
(BS), recurrent
aphthous
stomatitis
@AS) and
240
..
450
400
‘gE
.
KU/l
. .
I . .
z .
ia
w
. I
.
H q
00. .
cm
..
BS
C
..
c Fig. 2. Serum concentrations control5 (C).
of IgE in Behcet’s syndrome
(BS). recurrent
aphthous
stomatitis
(RAS) and
Serum IgD concentrations are shown in Fig. 1. Serum IgD levels although raised, were not significantly different in BS (median 0.014 pg/l) from controls (0.006 fig/l). Serum IgD concentrations in RAS (0.016 pg/l) were however, significantly greater than in controls ( p < 0.05). Serum IgE concentrations are shown in Fig. 2. Serum IgE levels in BS (median 18.8 kU/l) were not significantly different from controls (15.2 kU/l). Serum IgE concentrations in RAS (30.0 kU/l) were significantly greater than in either controls (p -L 0.01) or BS (p C 0.05). No correlation could be demonstrated between the serum IgD and IgE levels in any of the groups. Discussion The results indicate a significant rise in serum concentrations of IgA, insignificant rises in IgG, IgM, IgD and IgE in patients with Behcet’s syndrome
with (BS)
241
compared with healthy controls. In recurrent aphthous stomatitis (RAS), serum con~ntrations of both IgD and IgE were si~ficantly greater than in controls, but IgC, IgM and IgA concentrations did not significantly differ from levels in controls. There were no significant differences in the concentrations of any immunoglobulins between RAS and BS. These findings confirm previous reports with regard to IgG, IgM and IgA [12] and extend the immunological findings in Behcet’s syndrome to demonstrate normal serum concentrations of IgD and IgE. Although IgD immunocytes may contribute significantly to the secretory immune system of the upper gastrointestinal tract [13] and have been identified in the lesions of RAS [ 111, they have not been studied in BS. In the present study serum IgD levels were raised in RAS, and it is possible that IgD plays a role in the immunopathogenesis of BAS, perhaps by an immunoregulatory mechanism [14]. However, although IgD is increased in some viral infections [x5], and a viral aetiology has been implicated in BS [16], serum IgD was not increased in our patients with BS to a significant extent. Previous studies in small numbers of patients with BS have shown increased serum IgE levels and it was suggested that IgE is involve in a mast cell response in the immunopathogenesis of the lesions (91. However, the present study failed to confirm a significant increase in IgE concentrations in BS over controls and certainly the slight rise in serum IgE levels in BS shown in this study is not ~mpatible with a major atopic~~er~c component in BS 1171.Indeed, there is little, if any, evidence for atopy or allergy in BS except for the possible association with the ingestion of walnut [ 181. Increased serum concentrations of IgE were found in RAS. IgE-bearing lymphocytes have been identified in RAS lesions, and are present in increased numbers in the peripheral blood of patients with RAS [I 11. Furthermore, increased mast cell concentrations may be observed in the lesions of RAS [5] with frequent mast cell degranulation [ 191, a process often mediated by IgE antibodies. Some patients categorically state that their BAS is directly related to the ingestion of certain foods [ZO] but, although an increased frequency of allergic disorders is seen in RAS [21], the evidence for food allergy is not good in most cases [22]. The IgE concentrations seen here in RAS fall well below the levels usually associated with allergic disease fl7] and it is possible that the increases in serum IgE in RAS might simply reflect a non-specific phenomenon, as seen in some skin diseases [23]: The present study shows that serum concentrations of IgA but not IgG, IgM, IgD or IgE, were increased in Behcet’s syndrome. In contrast, serum IgD and IgE concentrations were increased in recurrent aphthous stomatitis. IgA, IgD and IgE may all be synthesised predominantly in secretory sites [24,25] and the present finding of raised serum levels may be relevant in view of the previously reported dissociation of secretory and systemic immune responses in BS 1121. Acknowledgements
We thank Dr. P. Atkinson (Pharmacia Ltd., UK) for gifts of IgE PRIST kits and colleagues for referring their patients or supplying sera.
242
References I Rogers RS. Recurrent aphthous stomatitis: clinical characteristics and evidence for an immunopathogenesis. J Invest Dermatol 1977; 69: 4999507. 2 Dilsen N, Konice M, Ovul C. Behcet’s disease. Amsterdam: Excerpta Medica. 1979: International Congress Series 467. 3 Lehner T. Barnes CG. Behcet’s Syndrome. London: Academic Press, 1979. 4 Aphthous stomatitis-Behcet’s Syndrome Symposium. J Oral Path01 1978; 7: 341-440. 5 Lehner T. Pathology of recurrent oral ulceration and oral ulceration in Behcet’s syndrome. Light. electron and fluorescence microscopy. J Oral Path01 1969: 97: 48 l-494. 6 Nazzaro P. Cutaneous manifestations of Behcet’s disease: clinical and histological findings. Proceedings of International Symposium on Behcet’s Disease. Basel: Karger. 1966; 15. 7 Haim S, Sobel JD, Friedman-Bimbaum R, Lichtig C. Histological and direct immunofluorescence study of cutaneous hyperreactivity in Behcet’s disease. Br J Dermatol 1976: 95: 63 I-636. 8 Lichtig C, Haim S, Hammel I, Friedman-Bimbaum R. The quantification and significance of mast cells in lesions of Behcet’s disease. Br J Dermatol 1980; 102: 255-259. 9 Haim S. The pathogenesis of lesions in Behcet’s disease. Dermatologica 1979: 158: 3 l-37. IO Haim S, Gilhar A. Clinical and laboratory criteria for the diagnosis of Behcet’s disease. Br J Dermatol 1980: 102: 361-363. I I Bays RA, Hamerlinck F, Cormane RH. Immunoglobulin-bearing lymphocytes and polymorphonuclear leucocytes in recurrent aphthous ulcers in man. Arch Oral Biol 1977: 22: 147- 153. 12 Scully C, Lehner T, Harfitt F. Serum salivary and lacrimal immunoglobulins in Behcet’s syndrome and recurrent oral ulcers. In: Lehner T. Barnes CG. eds., Behcet’s syndrome. London: Academic Press, 1979: 77-90. 13 Brandtzaeg P, Surjan L, Berdal P. Immunoglobulin systems of human tonsils. 1. Control subjects of various ages. Quantification of Ig-producing cells, tonsillar morphology and serum immunoglobulin levels. Clin Exp Immunol 1978; 3 I : 367-38 I. 14 Parkhouse RME, Cooper MD. A model for the differentiation of B lymphocytes with implications for the biological role of IgD. Immunol Rev 1977; 37: 105- 126. 15 Rostenberg I, Penaloza R. Serum IgG and IgD levels in some infectious and non infectious diseases. Clin Chim Acta 1978; 85: 319-321. 16 Denman AM, Fialkow PJ, Pelton BK, Salo A, Appleford DJ, Gilchrist C. Lymphocyte abnormalities in Behcet’s syndrome. Clin Exp Immunol 1980; 42: 175- 185. 17 Johansson SGO. Raised levels of a new immunoglobulin class (IgND) in asthma. Lancet 1967; 2: 951-953, 18 Marquardt JL, Snyderman R, Oppenheim JJ. Depression of lymphocyte transformation and exacerbation of Behcet’s syndrome by ingestion of English Walnuts. Cell Immunal 1973: 9: 263-272. 19 Dolby AE, Allison RT. Quantitative changes in the mast cell population in Mikulicz’s recurrent oral aphthae. J Dent Res 1969; 48: 90-903. 20 Finn R, Cohen HN. Food allergy: fact or fiction. Lancet 1978; I: 426-428. 21 Spouge JD, Diamond HF. Hypersensitivity reactions in mucous membranes. Oral Surg 1963; 16: 412-421. 22 Barbash R. Kutscher AH, Zegarelli EC, Silvers HF. Recurrent ulcerative (aphthous) stomatitis: intradermal food test studies. J Allergy 1958; 29: 442-445. 23 O’Laughlin S, Diaz-Perez JL, Gleich GJ. Serum IgE in dermatitis and dermatosis. An analysis of 497 cases. Arch Dermatol 1977; 113: 309-315. 24 Nakajima S, Gillespie DN, Gleich GJ. Differences between IgA and IgE as secretory proteins. Clin Exp Immunol 1975; 21: 306310. 25 Brandtzaeg P, Gjeruldsen ST, Korsrud F, Baklien K, Berdal P, Ek J. The human secretory immune system shows striking heterogeneity with regard to involvement of J chain positive IgD immunocytes. J Immunol 1979; 122: 503-510.
Clinica Chimica Acta, 120 ( 1982) 237-242 Elsevier Biomedical Press
CCA 207 1
~~unog~obu~ins c. Scully u University
G, M, A, D and E in Behcet’s syndrome a**, P. Boyle b and P.L. Yap c
Department of Oral Medicine and Pathology, Glasgow Dental Hospital, G(usgow (UK), h West of
Scotland Cancer Surveillance
Unit, Ruchill Hospital, Glasgow {UK) and c Edinburgh and South-East
Scotland Regional Blood Transfusion Service, Royal Infirmary,
Edinburgh (UK)
(Received August IOth, revision November 30th‘ 1981)
summaly Serum IgG, IgM, IgA, IgD and IgE concentrations were examined in 26 patients with Behcet’s syndrome, 70 patients with recurrent aphthous stomatitis and 56 healthy controls. IgA concentrations, but not IgG, IgM, IgD or IgE, were significantly raised in Behcet’s syndrome compared with controls. Serum IgD and IgE concentrations but not IgA, IgG or IgM were significantly greater in recurrent aphthous stomatitis than in controls.
Introduction Behcet’s syndrome (BS) is an uncommon systemic disease character&d mainly by recurrent aphthous stomatitis, genital ulceration, uveitis and various systemic lesions, but there are no laboratory findings that unequivocally distinguish BS from recurrent aphthous stomatitis (R4S) occurring alone. The aetiology of BS and R4S is unknown, although for both the evidence suggests an immunopathogenesis [l-4]. Histopathological examination of the early lesions of RAS [5] and of the mucosal[5] and cutaneous [6-S] lesions of BS shows that, in addition to a lymphocytic and monocytic infiltrate, there is an increase in mast cells, and these have been implicated in the i~~opathogenesis of BS [S-10]. It has been suggested that IgE may be involved in the immunopathogenesis of BS [9]. Lymphocytes bearing IgD and IgE have been identified in the oral lesions and peripheral blood of patients with IXAS [ 1l] but have not been studied in BS. Serum IgE ~ncentrations have been examined in a small group of patients with BS [9]; serum levels of IgD do not appear to have been examined, and studies of the serum concentrations of other immunoglobulins have been somewhat equivocal; serum IgG * Correspondence to Dr. C. Scufly, Immunology Unit, Department of Oral Medicine and Pathology, Glasgow Dental Hospital. Glasgow, G2 352, UK. 0009-898 l/82/~-~/$02.75
@ 1982 Elsevier Biomedical Press
and IgM levels are usually unchanged in BS with increased IgA (12j. We have therefore estimated serum concentrations of IgG, IgM, IgA, IgD and IgE in patients with Behcet’s syndrome and recurrent aphthous stomatitis.
Materials and methods The study group of 96 adult Caucasians consisted of 26 patients with Behcet’s syndrome (median age 32 years: range 20-52: 14 males) diagnosed on the basis of recurrent oral and genital ulceration together with ocular, cutaneous, arthritic or neurological manifestations [3] and 70 patients with recurrent aphthous stomatitis (median age 25 years: range 15-37: 30 males). The control group consisted of 56 healthy subjects with no oral mucosal disease (median age 28 years: range 19-46: 25 males). Patients and controls were matched as closely as possible for a history of allergies. No patient had received systemic medication within the previous three months but most were using topical oral applications of chlorhexidine, triamcinolone or hydrocortisone.
Serum immunoglobulin estimations Sera were separated after clotting at 20°C for 30 min and overnight at 4*C, and stored in aliquots at -20°C. Serum IgG, IgM, IgA and IgE concentrations were measured in all subjects but IgD estimation could only be carried out in 39 of the controls and in 24 of those with BS. All immunoglobulins were assayed by immunodiffusion techniques, except IgE which was assayed by radio-immunoassay. Serum IgG, IgM and IgA concentrations were measured in duplicate by single radial immunodiffusion as previously described [ 121, and expressed as mg/ 100 ml (mean k standard deviation). Serum IgD concentrations were measured in duplicate by a single radial immunodiffusion technique using sheep anti S antiserum (Scottish Antibody Production Unit, Law Hospital; Carluke) and an IgD myeloma standard previously calibrated against the World Health Organisation IgD standard, and expressed as lug/l. Serum IgE concentrations were measured in duplicate by a solid phase radioimmunoassay (Phadebas IgE PRIST, Pharmacia, Sweden), and expressed as kU/l.
statistical analysis Results were analysed by ~skal-Wallis analysis of variance. IgG, IgM and IgA results were then analysed by the Dunnett test but the Mann-W~tney test was used to analyse IgD and IgE results because of the non-parametric distribution of data. Results Serum IgG, IgM and IgA concentrations are shown in TableI. Serum IgA concentrations in Behcet’s syndrome (BS) were significantly greater than in controls (p-c 0.05) but not greater than levels in recurrent aphthous stomatitis (RAS). No significant differences could be established between any groups either for serum IgG or for IgM concentrations.
239
TABLE SERUM
kG
I CONCENTRATIONS
OF MAIOR
IMMUNOGLOBULINS
IN BEHCET’S
Behcet’s syndrome
Controls
Recurrent
1690 * 428
1443*205
16611214
aphthous
IgM
1022
46
1122
53
108-c
42
IgA
386*114**
242k
63
298*
12
* Mean k SD in mg/ 100 ml. ** Significantly different from controls
SYNDROME
*
stomatitis
(p
0.20
. .
.
. : .
f . Aa
.
8
.
BS
c
Fig. I. Serum concentrations controls (C).
:
!
RAS
of IgD in Behcet’s syndrome
(BS), recurrent
aphthous
stomatitis
@AS) and
240
..
450
400
‘gE
.
KU/l
. .
I . .
z .
ia
w
. I
.
H q
00. .
cm
..
BS
C
..
c Fig. 2. Serum concentrations control5 (C).
of IgE in Behcet’s syndrome
(BS). recurrent
aphthous
stomatitis
(RAS) and
Serum IgD concentrations are shown in Fig. 1. Serum IgD levels although raised, were not significantly different in BS (median 0.014 pg/l) from controls (0.006 fig/l). Serum IgD concentrations in RAS (0.016 pg/l) were however, significantly greater than in controls ( p < 0.05). Serum IgE concentrations are shown in Fig. 2. Serum IgE levels in BS (median 18.8 kU/l) were not significantly different from controls (15.2 kU/l). Serum IgE concentrations in RAS (30.0 kU/l) were significantly greater than in either controls (p -L 0.01) or BS (p C 0.05). No correlation could be demonstrated between the serum IgD and IgE levels in any of the groups. Discussion The results indicate a significant rise in serum concentrations of IgA, insignificant rises in IgG, IgM, IgD and IgE in patients with Behcet’s syndrome
with (BS)
241
compared with healthy controls. In recurrent aphthous stomatitis (RAS), serum con~ntrations of both IgD and IgE were si~ficantly greater than in controls, but IgC, IgM and IgA concentrations did not significantly differ from levels in controls. There were no significant differences in the concentrations of any immunoglobulins between RAS and BS. These findings confirm previous reports with regard to IgG, IgM and IgA [12] and extend the immunological findings in Behcet’s syndrome to demonstrate normal serum concentrations of IgD and IgE. Although IgD immunocytes may contribute significantly to the secretory immune system of the upper gastrointestinal tract [13] and have been identified in the lesions of RAS [ 111, they have not been studied in BS. In the present study serum IgD levels were raised in RAS, and it is possible that IgD plays a role in the immunopathogenesis of BAS, perhaps by an immunoregulatory mechanism [14]. However, although IgD is increased in some viral infections [x5], and a viral aetiology has been implicated in BS [16], serum IgD was not increased in our patients with BS to a significant extent. Previous studies in small numbers of patients with BS have shown increased serum IgE levels and it was suggested that IgE is involve in a mast cell response in the immunopathogenesis of the lesions (91. However, the present study failed to confirm a significant increase in IgE concentrations in BS over controls and certainly the slight rise in serum IgE levels in BS shown in this study is not ~mpatible with a major atopic~~er~c component in BS 1171.Indeed, there is little, if any, evidence for atopy or allergy in BS except for the possible association with the ingestion of walnut [ 181. Increased serum concentrations of IgE were found in RAS. IgE-bearing lymphocytes have been identified in RAS lesions, and are present in increased numbers in the peripheral blood of patients with RAS [I 11. Furthermore, increased mast cell concentrations may be observed in the lesions of RAS [5] with frequent mast cell degranulation [ 191, a process often mediated by IgE antibodies. Some patients categorically state that their BAS is directly related to the ingestion of certain foods [ZO] but, although an increased frequency of allergic disorders is seen in RAS [21], the evidence for food allergy is not good in most cases [22]. The IgE concentrations seen here in RAS fall well below the levels usually associated with allergic disease fl7] and it is possible that the increases in serum IgE in RAS might simply reflect a non-specific phenomenon, as seen in some skin diseases [23]: The present study shows that serum concentrations of IgA but not IgG, IgM, IgD or IgE, were increased in Behcet’s syndrome. In contrast, serum IgD and IgE concentrations were increased in recurrent aphthous stomatitis. IgA, IgD and IgE may all be synthesised predominantly in secretory sites [24,25] and the present finding of raised serum levels may be relevant in view of the previously reported dissociation of secretory and systemic immune responses in BS 1121. Acknowledgements
We thank Dr. P. Atkinson (Pharmacia Ltd., UK) for gifts of IgE PRIST kits and colleagues for referring their patients or supplying sera.
242
References I Rogers RS. Recurrent aphthous stomatitis: clinical characteristics and evidence for an immunopathogenesis. J Invest Dermatol 1977; 69: 4999507. 2 Dilsen N, Konice M, Ovul C. Behcet’s disease. Amsterdam: Excerpta Medica. 1979: International Congress Series 467. 3 Lehner T. Barnes CG. Behcet’s Syndrome. London: Academic Press, 1979. 4 Aphthous stomatitis-Behcet’s Syndrome Symposium. J Oral Path01 1978; 7: 341-440. 5 Lehner T. Pathology of recurrent oral ulceration and oral ulceration in Behcet’s syndrome. Light. electron and fluorescence microscopy. J Oral Path01 1969: 97: 48 l-494. 6 Nazzaro P. Cutaneous manifestations of Behcet’s disease: clinical and histological findings. Proceedings of International Symposium on Behcet’s Disease. Basel: Karger. 1966; 15. 7 Haim S, Sobel JD, Friedman-Bimbaum R, Lichtig C. Histological and direct immunofluorescence study of cutaneous hyperreactivity in Behcet’s disease. Br J Dermatol 1976: 95: 63 I-636. 8 Lichtig C, Haim S, Hammel I, Friedman-Bimbaum R. The quantification and significance of mast cells in lesions of Behcet’s disease. Br J Dermatol 1980; 102: 255-259. 9 Haim S. The pathogenesis of lesions in Behcet’s disease. Dermatologica 1979: 158: 3 l-37. IO Haim S, Gilhar A. Clinical and laboratory criteria for the diagnosis of Behcet’s disease. Br J Dermatol 1980: 102: 361-363. I I Bays RA, Hamerlinck F, Cormane RH. Immunoglobulin-bearing lymphocytes and polymorphonuclear leucocytes in recurrent aphthous ulcers in man. Arch Oral Biol 1977: 22: 147- 153. 12 Scully C, Lehner T, Harfitt F. Serum salivary and lacrimal immunoglobulins in Behcet’s syndrome and recurrent oral ulcers. In: Lehner T. Barnes CG. eds., Behcet’s syndrome. London: Academic Press, 1979: 77-90. 13 Brandtzaeg P, Surjan L, Berdal P. Immunoglobulin systems of human tonsils. 1. Control subjects of various ages. Quantification of Ig-producing cells, tonsillar morphology and serum immunoglobulin levels. Clin Exp Immunol 1978; 3 I : 367-38 I. 14 Parkhouse RME, Cooper MD. A model for the differentiation of B lymphocytes with implications for the biological role of IgD. Immunol Rev 1977; 37: 105- 126. 15 Rostenberg I, Penaloza R. Serum IgG and IgD levels in some infectious and non infectious diseases. Clin Chim Acta 1978; 85: 319-321. 16 Denman AM, Fialkow PJ, Pelton BK, Salo A, Appleford DJ, Gilchrist C. Lymphocyte abnormalities in Behcet’s syndrome. Clin Exp Immunol 1980; 42: 175- 185. 17 Johansson SGO. Raised levels of a new immunoglobulin class (IgND) in asthma. Lancet 1967; 2: 951-953, 18 Marquardt JL, Snyderman R, Oppenheim JJ. Depression of lymphocyte transformation and exacerbation of Behcet’s syndrome by ingestion of English Walnuts. Cell Immunal 1973: 9: 263-272. 19 Dolby AE, Allison RT. Quantitative changes in the mast cell population in Mikulicz’s recurrent oral aphthae. J Dent Res 1969; 48: 90-903. 20 Finn R, Cohen HN. Food allergy: fact or fiction. Lancet 1978; I: 426-428. 21 Spouge JD, Diamond HF. Hypersensitivity reactions in mucous membranes. Oral Surg 1963; 16: 412-421. 22 Barbash R. Kutscher AH, Zegarelli EC, Silvers HF. Recurrent ulcerative (aphthous) stomatitis: intradermal food test studies. J Allergy 1958; 29: 442-445. 23 O’Laughlin S, Diaz-Perez JL, Gleich GJ. Serum IgE in dermatitis and dermatosis. An analysis of 497 cases. Arch Dermatol 1977; 113: 309-315. 24 Nakajima S, Gillespie DN, Gleich GJ. Differences between IgA and IgE as secretory proteins. Clin Exp Immunol 1975; 21: 306310. 25 Brandtzaeg P, Gjeruldsen ST, Korsrud F, Baklien K, Berdal P, Ek J. The human secretory immune system shows striking heterogeneity with regard to involvement of J chain positive IgD immunocytes. J Immunol 1979; 122: 503-510.