Autoantibodies against desmoglein 2 are not pathogenic in pemphigus

Miguel, Marcela Calixto Brandão; Julio, Tamiris Amanda; Vernal, Sebastian; Paula, Natália Aparecida de; Lieber, Andre; Roselino, Ana Maria

doi:10.1016/j.abd.2021.06.004

Marcela Calixto Brandão Miguel

Division of Dermatology, Department of Internal Medicine, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil

https://orcid.org/0000-0001-8820-016X

Tamiris Amanda Julio

Division of Dermatology, Department of Internal Medicine, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil

https://orcid.org/0000-0002-0536-9726

Sebastian Vernal

Division of Dermatology, Department of Internal Medicine, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil

https://orcid.org/0000-0001-8589-0089

Natália Aparecida de Paula

Division of Dermatology, Department of Internal Medicine, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil

Laboratory of the Sector of Dermatology, Division of Dermatology, Department of Internal Medicine, Hospital das Clínicas, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil

https://orcid.org/0000-0002-3744-8947

Andre Lieber

Division of Medical Genetics, Department of Pathology, University of Washington, Seattle, WA, USA

https://orcid.org/0000-0001-6742-6266

Ana Maria Roselino ^**Corresponding author. E-mail:amfrosel@fmrp.usp.br (A.M. Roselino).

Division of Dermatology, Department of Internal Medicine, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil

Laboratory of the Sector of Dermatology, Division of Dermatology, Department of Internal Medicine, Hospital das Clínicas, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil

https://orcid.org/0000-0002-2709-1825

*Corresponding author. E-mail:amfrosel@fmrp.usp.br (A.M. Roselino).

☆
Study conducted at the Hospital das Clínicas, Division of Dermatology, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil.

Authors’ contributions

Marcela Calixto Brandão Miguel: Contributed with the collection, analysis and interpretation of data; drafting and editing of the manuscript; statistical analysis; critical review of the literature.

Tamiris Amanda Julio: Contributed with the collection, analysis and interpretation of data; critical review of the manuscript; approval of the final version of the manuscript.

Sebastian Vernal: Contributed with the data analysis; critical review of the manuscript; approval of the final version of the manuscript.

Natália Aparecida de Paula: Contributed with the collection, analysis and interpretation of data; critical review of the manuscript; approval of the final version of the manuscript.

Andre Lieber: Contributed with the planning of the study, critical review of the manuscript; approval of the final version of the manuscript.

Ana Maria Roselino: Contributed with the design and planning of the study; analysis and interpretation of data; critical review of the literature; critical review of the manuscript and effective participation in research orientation.

Conflicts of interest

None declared.

Figures (5)
Tables (2)

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Figure 1
IgG antibodies against Dsg1, Dsg2 and Dsg3 in the studied groups. A, Anti-Dsg2 by in-house ELISA. PF, PV and FPV had higher titers compared to controls (CTL; p < 0.0001); PF and FPV showed higher titers than FPF (p = 0.0046 and p = 0.0004, respectively). B, Anti-Dsg1 by commercial ELISA. PF had higher titers compared to all groups (p < 0.0001); PV, higher titers than FPF, FPV and CTL (p < 0.0001). C, Anti-Dsg3 by commercial ELISA. PV had higher titers than all groups (p < 0.0001). The horizontal line in each column indicates the median; the dotted line, the cut-off value. PF, Pemphigus Foliaceus; PV, Pemphigus Vulgaris; FPF, Family members of PF, FPV, Family members of PV. Statistical analysis: Kruskal-Wallis followed by Dunn’s test. D, Spearman’s correlation between anti-Dsg2 and anti-Dsg1 titers in PF. E, Spearman’s correlation between anti-Dsg2 and anti-Dsg3 titers in PV.

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Figure 2
Immunohistochemistry panel with Dsg1, Dsg2 and Dsg3 expressions in skin samples (original magnification: ×40). (A), Abdominoplasty control skin sample showing typical distribution of Dsg1, Dsg2 and Dsg3 in the epidermis. (B), PF - intact skin sample showing increased expressions of Dsg2 and Dsg3 in the entire epidermis and PF - lesional skin sample showing increased expressions of Dsg2 and Dsg3 on the roof and floor of the acantholytic bulla. (C), PV-intact skin sample showing increased expression of Dsg2 and Dsg3 in the entire epidermis and PV-lesional skin sample showing increased expression of Dsg2 and Dsg3 on the roof and floor of the bulla and in the acantholytic cells.

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Figure 3
Immunohistochemistry panel with Dsg1, Dsg2 and Dsg3 expressions in mucosal samples. (A), PF-intact mucosa showing the typical distribution of Dsg1, Dsg2 and Dsg3 in the epithelium (original magnification: ×63). (B), PV-lesional mucosa showing Dsg2 and Dsg3 expression surrounding the acantholytic bulla (original amplification: ×100 and ×40).

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Figure 4
Immunohistochemistry panel with Dsg1, Dsg2 and Dsg3 expressions in skin and mucosa samples from patients with PF and PV. (A), Dsg1 in intact and lesional skin samples from patient PF2 showing conspicuous intercellular and intracytoplasmic granules (Dsg1 internalization), widely distributed in lesional skin (original magnification: ×100). (B), Dsg2 comparing intact and lesional skin sample from patient PF2, confirming the absence of granules (original magnification: ×100). (C), Dsg3 comparing intact mucosa sample from patient PF2 and lesional mucosa from patient PV3, showing fine intercellular and intracytoplasmic granules (Dsg3 internalization) in the lesional mucosa (original magnification: ×100).

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Figure 5
Relative gene expressions (×10-4) of Dsg1, Dsg2 and Dsg3 in skin and mucosal samples. (A), Intact skin (IS) lesional skin (LS) samples of the PF group compared to the facelift (FL) control sample showing predominance of Dsg3 expression in comparison to Dsg1 and Dsg2. (B), IS and LS samples of the PV and FL groups. The expressions of Dsg1, Dsg2 and Dsg3 were lower than in the FL sample. (C), Lesional mucosa (LM) sample of the PV group in comparison to intact mucosa (IM) sample of the PF group showing predominance of Dsg3 expression in comparison to Dsg1 and Dsg2. Dsg2 expression is increased in mucosal samples. (D), Comparison of Dsg2 expression in all PF and PV samples. Relative gene expressions of Dsg1, Dsg2, Dsg3 were calculated using 2^(-ΔcT) and using 18S rRNA as the reference gene.

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Table 1
Demographic and clinical data, and anti-desmoglein (Dsg) ELISA results comparing patients with PF and PV, relatives of PF and PV (FPF and FPV) and Controls (CTL).

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Table 2
Demographic, clinical and anti-Dsg 1, 2 and 3 antibody titers by ELISA of patients with PF and PV submitted to skin and/or mucosa biopsy.

	PF	PV	FPF	FPV	Controls	p
	n = 202	n = 131	n = 50	n = 57	n = 114
Median age in years (25^th/75^th quartiles)	35 (21/54)^a	48 (35/62)^b	40 (29/49)^c	30 (19/45)^d	59 (45/71)^e	0.0001^a.b
						> 0.9999^a.c
						< 0.0001^a.e
						0.0001^b.d
						0.0068^b.e

	n (%)	n (%)	n (%)	n (%)	n (%)
	202	131	50	57	114
Sex						0.0033^a.b
Sex						0.0027^b.e
Female	102 (50.2)^a	88 (67.7)^b	36 (72.0)^c	40 (70.2)^d	55 (48.2)^e	0.8150^a.e
Female	102 (50.2)^a	88 (67.7)^b	36 (72.0)^c	40 (70.2)^d	55 (48.2)^e	1.0000^c.d
Male	101 (49.8)	42 (32.3)	14 (28.0)	17 (29.8)	59 (51.8)	0.0022^c.e
Male	101 (49.8)	42 (32.3)	14 (28.0)	17 (29.8)	59 (51.8)	0.0088^d.e

Disease duration (years)^* * Some data not compiled.	n = 199	n = 130
< 1	45 (22.6)	31 (23.8)	NA	NA	NA	0.8967
1 to -5	87 (43.7)	86 (66.2)	NA	NA	NA	0.0296
6 to -10	32 (16.1)	9 (6.9)	NA	NA	NA	0.0390
11 to 15	11 (5.5)	3 (2.3)	NA	NA	NA	0.2630
> 15	24 (12.1)	1 (0.8)	NA	NA	NA	0.0001

Clinical form (treatment-naïve patients)^* * Some data not compiled.	n = 68	n = 26
Exclusively mucosal	NA	8 (30.8)^b	NA	NA	NA	^a = 0.9636
Exclusively mucosal	NA	8 (30.8)^b	NA	NA	NA	^b = 0.3909
Muco-cutaneous	NA	18 (69.2)^b	NA	NA	NA
Localized	34 (50.0)^a	NA	NA	NA	NA
Generalized	34 (50.0)^a	NA	NA	NA	NA

Treatment on the date of serum collection^* * Some data not compiled.	n = 186	n = 127
Treatment-naïve	43 (23.1)^a	20 (15.8)^b	NA	NA	NA	^a < 0.0001
No treatment with active lesions	34 (18.3)^a	11 (8.7)^b	NA	NA	NA	^b < 0.0001
Undergoing treatment without active lesions	16 (8.6)^a	24 (18.8)^b	NA	NA	NA
Undergoing treatment with active lesions	80 (43.0)^a	65 (51.2)^b	NA	NA	NA
In remission	13 (7.0)^a	7 (5.5)^b	NA	NA	NA

Anti-Dsg2	n = 202	n = 131	n = 50	n = 57	n = 114	< 0.0001^a.e
Above the cutoff	129 (63.9)^a	87 (66.4)^b	19 (38.0)^c	42 (73.7)^d	31 (27.2)^e	< 0.0001^b.e
Above the cutoff	129 (63.9)^a	87 (66.4)^b	19 (38.0)^c	42 (73.7)^d	31 (27.2)^e	0.0012^a.c
Below the cutoff	73 (36.1)	44 (33.6)	31 (62.0)	15 (26.3)	83 (72.8)	< 0.0001^c.d
Below the cutoff	73 (36.1)	44 (33.6)	31 (62.0)	15 (26.3)	83 (72.8)	< 0.0001^d.e
Anti-Dsg1						< 0.0001^a.b
Anti-Dsg1						< 0.0001^a.e
Above the cutoff	179 (88.2)^a	67 (51.5)^b	6 (12.0)^c	1 (1.8)^d	4 (3.5)^e	< 0.0001^a.c
Above the cutoff	179 (88.2)^a	67 (51.5)^b	6 (12.0)^c	1 (1.8)^d	4 (3.5)^e	< 0.0001^b.d
Below the cutoff	23 (11.8)	64 (48.5)	44 (88.0)	56 (98.2)	110 (96.5)	< 0.0001^b.e
Below the cutoff	23 (11.8)	64 (48.5)	44 (88.0)	56 (98.2)	110 (96.5)	0.0325^c.d
Anti-Dsg3
Above the cutoff	9 (4.5)^a	102 (78.5)^b	4 (8.0)^c	0 (0.0)^d	3 (2.6)^e	< 0.0001^a.b
Above the cutoff	9 (4.5)^a	102 (78.5)^b	4 (8.0)^c	0 (0.0)^d	3 (2.6)^e	< 0.0001^b.e
Below the cutoff	193 (95.5)	29 (21.5)	46 (92.0)	57 (100.0)	111 (97.4)	< 0.0001^b.d
Below the cutoff	193 (95.5)	29 (21.5)	46 (92.0)	57 (100.0)	111 (97.4)	0.0446^c.d

	Age	Sex	Disease duration (months)	Clinical form	Treatment on biopsy date	ANTI-DSG1 (U/ML)^a a Anti-Dsg1 and 3 cut-off = < 20 U/mL (ELISA, MBL, Japan).	ANTI-DSG2 (OD)^b b Anti-Dsg2 cut-off = 0.1365 DO (in-house ELISA).	ANTI-DSG3 (U/ML)^a a Anti-Dsg1 and 3 cut-off = < 20 U/mL (ELISA, MBL, Japan).
PF1	23	Male	48	Generalized	In treatment for 30 days	99.80	0.088	1.93
PF2	18	Male	6	Generalized	Treatment naïve	193.08	0.223	4.15
PF3	25	Male	36	Localized	Treatment naïve	237.90	0.894	8.70
PV1	61	Male	24	Muco-cutaneous	No treatment for more than 60 days	3.87	0.211	172.78
PV2	50	Male	4	Muco-cutaneous	Treatment naïve	42.16	0.213	166.91
PV3	68	Male	6	Muco-cutaneous	Treatment naïve	167.62	0.311	163.80

[1] *Corresponding author. E-mail:amfrosel@fmrp.usp.br (A.M. Roselino).

Brasil

Brasil

Autoantibodies against desmoglein 2 are not pathogenic in pemphigus^☆ ☆ Study conducted at the Hospital das Clínicas, Division of Dermatology, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil.

Abstract

Background

Objective

Methods

Results

Study limitations

Conclusion

Brasil

Brasil

Autoantibodies against desmoglein 2 are not pathogenic in pemphigus☆ ☆ Study conducted at the Hospital das Clínicas, Division of Dermatology, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil.

Abstract

Background

Objective

Methods

Results

Study limitations

Conclusion

Autoantibodies against desmoglein 2 are not pathogenic in pemphigus^☆ ☆ Study conducted at the Hospital das Clínicas, Division of Dermatology, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil.