Desmoglein Autoantibodies and Disease Severity in Pemphigus Patients – Correlations and Discrepancies

Desmoglein


INTRODUCTION
Pemphigus comprises a group of rare potentially life-threatening autoimmune bullous diseases.Pemphigus vulgaris (PV) is the most common form.Mucosal lesions are present in almost all patients, with or without cutaneous bullae or erosions. [1,2][5] Dsg1 and Dsg3 are cadherins, which are calcium-dependent transmembrane glycoproteins of the epidermal or epithelial desmosomes.They are part of the maculae adherentes and allow the cell-to-cell adhesion in the epidermis and the epithelium. [6]olia Medica I 2023 I Vol.65 I No. 6

AIM
The aim of this study was to assess the correlation between the levels of anti-desmoglein-1 and anti-desmoglein-3 autoantibodies and disease severity in pemphigus patients.

Patients
This prospective study was conducted from March 2020 to November 2022.Thirty-eight patients with pemphigus were included based on the following criteria: (1) clinical diagnosis, (2) histopathological picture revealing intraepidermal acantholysis, and (3) deposition of IgG and complement component 3 on keratinocyte surface established by direct immunofluorescence assay.[9] A control group of 38 people (17 patients with bullous pemphigoid, two patients with psoriasis, five pregnant woman, and 14 healthy blood donors) served as controls.Controls were sex and age matched.The study was approved by the Ethics Committee of the Medical University of Plovdiv and conducted in accordance with the Declaration of Helsinki.Informed consent was obtained from all individual participants included in the study.

Anti-desmoglein-1 and antidesmoglein-3 enzyme-linked immunosorbent assay (ELISA)
Anti-Dsg1 and anti-Dsg3 antibodies (Abs) were evaluated using the EUROIMMUN anti-desmoglein-1 and anti-desmoglein-3 ELISA (IgG) test kit according to the manufacturer's instructions.Sera from pemphigus patients and controls were collected and stored at -80°C until assayed.The patient sera were diluted 1:101 in sample buffer.A value exceeding or equal to 20 RU/ml was deemed to be positive, and a value <20 RU/ml was interpreted as negative (EUROIMMUN Medizinische Labordiagnostika, Lübeck, Germany).

Statistical analysis
Continuous variables are given as means and standard deviations (±SD), and category variables as percentages.The Spearman correlation test was used to compare disease severity and autoantibody levels.Sensitivity and specificity of ELISA were analyzed with the McNemar test.A receiver operating characteristic (ROC) curve was calculated to determine a cutoff value for anti-Dsg1 and anti-Dsg3 Abs.Statistical analyses were performed with data analysis software IBM -SPSS v. 23.Statistical significance was considered significant at p<0.05.

DISCUSSION
Twenty (52.6%) of the 38 pemphigus patients in the current study newly diagnosed, 15 (39.5%) were receiving therapy, and three (7.9%) had ceased their medication on their own volition at the time of the ELISA testing.A significant correlation between the PDAI scores and the ELISA tests results was revealed.Anti-Dsg1 Abs showed stronger correlation with the disease severity compared with anti-Dsg3 Abs.[12] The correlation between the antibodies and PDAI in the present study was most significant in patients with mucocutaneous PV.Greek researchers concluded that patients with mucocutaneous phenotypes had a total PDAI score strongly correlated with the levels of anti-Dsg1 Abs and less strongly with the anti-Dsg3 Ab levels. [13]n the current study, a discrepancy was seen in a patient with severe relapse of the disease with a PDAI score of 86 and negative anti-Dsg1 Abs (16.581RU/ml) and anti-Dsg3 Abs (<2.000RU/ml).Another incongruity occurred in a case of mild pemphigus (PDAI=13) and highly positive levels of both autoantibodies.No significant correlation was reported between anti-Dsg1 and anti-Dsg3 ELISA Abs and disease severity in some patients. [14]It was published that anti-Dsg1 ELISA tests can be considered a predictive tool for the occurrence of cutaneous relapses, whereas anti-Dsg3 Ab ELISA values do not necessarily correspond to mucosal involvement. [15]A study on 72 patients with PV established that a single blood sample was enough to detect a significant correlation between disease severity and levels of both anti-Dsg1 and anti-Dsg3 Abs (p<0.001,p=0.020). [16]Chinese authors demonstrated that there was no correlation between conventional anti-Dsg3 Ab ELISA values and the PDAI score, whereas a positive correlation between conformational anti-Dsg3 ELISA values and activities of cutaneous lesions was found in 29 PV patients. [17]esearchers clarified that anti-desmoglein Abs significantly increased with worsening of the disease, whereas indirect immunofluorescence titers did not show correlation with disease activity. [18]According to Cheng et al., only appropriate dilutions in ELISA, such as 1:800 and 1:1600 or above, can provide true serological information about disease activity. [19]his research established a case of newly diagnosed mucosal PV with negative anti-Dsg3 Ab (10.911RU/ml) and anti-Dsg1 Ab levels (16.776RU/ml).Balighi et al. also found initial negative anti-Dsg1/anti-Dsg3 Ab levels in eight female pemphigus patients. [20]n explanation for these contradictory ELISA results could be the pathogenic non-desmoglein Abs, such as desmocollins, plakophilin3, thyroid peroxide antibodies, anti-mitochondrial proteins, cholinergic receptors, e-cadherin, and plakoglobin. [21]Negative levels of anti-Dsg1 and anti-Dsg3 Abs were reported in one of 38 pemphigus cases. [22]In this study, one pregnant female patient with pemphigus herpetiformis had high levels of anti-Dsg1 and anti-Dsg3 Abs (>200.000RU/ml).Antibodies from patients with PF are targeted primarily against Dsg1. [23]It was established that exfoliative toxin A, which causes bullous impetigo and staphylococcal scalded skin syndrome, cleaves Dsg1, but does not target Dsg3. [24,25]The incongruity between autoantibody profile and clinical phenotype in PF patients could be due to non-pathogenic anti-Dsg3 Abs. [14][28] In particular, autoantibody levels should not be interpreted regardless of the clinical findings.
The study's limitation is the insufficient number of cases included in it.

CONCLUSIONS
Anti-desmoglein-1 and anti-desmoglein-3 antibody levels measured by ELISA can be associated with the disease activity and severity in patients with pemphigus, but there are exceptions.

Figure 1 .
Figure 1.ROC and AUC of anti-Dsg1 Abs for determining the optimal cut-off value.

Folia
Medica I 2023 I Vol.65 I No. 6

Table 1 .
Coordinates of the anti-Dsg1 Abs ROC

Table 2 .
Coordinates of the anti-Dsg3 Abs ROC