INTRODUCTION

Pemphigus vulgaris (PV) is a rare and potentially fatal autoimmune disease characterized by blistering and painful erosive lesions on the mucous membranes or skin ^{1 , 2} . Autoantibodies are mainly directed against desmoglein (Dsg) 1 and Dsg3 and are the key factors in the pathogenesis of PV, leading to acantholysis and intraepidermal blister formation ^{1 , 2} . Blister formation can be extensive and even fatal if left untreated. Thus, prompt initiation of immunosuppressive and maintenance therapies is mandatory. Corticosteroids represent the first line of treatment, though rituximab, a monoclonal antibody directed against CD20+ B cells, has also been approved for treating moderate-to-severe PV ^{2 , 3} . However, the required duration of treatment for each patient is not clear. Therefore, planning an accurate strategy with an assessment of immunologic remission in a proper clinical setting is required ⁴ .

The level of autoantibodies appears to be an important marker of immunologic remission. Anti-Dsg1/3 antibodies can assess the inflammatory status in PV, reflect the disease activity, and even predict the prognosis ^{5 - 7} . However, these serological tests are costly and not always available. Therefore, searching for other helpful markers that indicate PV characteristics is valuable.

Recently, the neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), and mean platelet volume (MPV) have been introduced as novel biomarkers of inflammation ^{8 - 10} . Elevated levels of these hematological markers are associated with disease severity in various autoimmune diseases such as systemic lupus erythematosus (SLE) ¹¹ , rheumatoid arthritis (RA) ¹² , and psoriasis ¹³ . As these parameters are easily obtained from a routine hematology test and can be assessed inexpensively and rapidly, we aimed to evaluate their possible associations with PV characteristics, such as disease severity and serological autoantibody levels.

METHODS

Participants

From September 2018 to October 2019, 33 newly diagnosed PV patients were included in this cross-sectional study at Shohada-e Tajrish and Loghman Hakim hospitals (Tehran, Iran), with 33 age and sex-matched healthy subjects as controls. The patients were diagnosed based on clinical examination, histopathology, direct immunofluorescence showing intraepithelial immunoglobin deposition, and detectable levels of autoantibodies against Dsg1 or Dsg3. All participants were >18 years of age and newly diagnosed cases who had not taken prednisolone or other immunosuppressive drugs. Exclusion criteria included a history of other chronic inflammatory diseases such as cardiovascular, hematologic, chronic liver or kidney diseases, and autoimmune or neoplastic disorders.

Ethical considerations

The study protocol was approved by the Shahid Beheshti University of Medical Sciences Ethics Committee (IR.SBMU.SRC.REC.1397.011). All participants signed an informed consent form.

Clinical and laboratory assessments

Demographic, clinical, and laboratory data were collected from patients and controls. Age, sex, type of pemphigus involvement, time of diagnosis, and severity of the disease based on the Autoimmune Bullous Skin Disorder Intensity Score (ABSIS) were recorded. This score is composed of cutaneous and oral scores and ranges from 0 to 206 ¹⁴ .

For each patient, a complete blood count (CBC) was obtained, which included the mean platelet volume (MPV) and counts of white blood cells (WBC), neutrophils, lymphocytes, and platelets. The NLR was calculated by dividing the neutrophil count by the lymphocyte count, and PLR was obtained by dividing the platelet count by the lymphocyte count. Anti-Dsg3 and anti-Dsg1 levels were measured using commercial enzyme-linked immunosorbent assays (ELISA) for all the patients to confirm the diagnosis.

Statistical analysis

Continuous variables are reported as mean ± standard deviation (SD) or median (interquartile range), and categorical variables as numbers and percentages. To check the data distribution, the Kolmogorov-Smirnov test was used. For continuous variables, the difference between groups was determined by the student t-test or Mann-Whitney U-test for variables with or without normal distributions, respectively. The categorical variables were compared using the x2 test. Spearman's correlation test evaluated associations between laboratory findings and ABSIS. All statistical analyses were performed using SPSS version 24 (SPSS, Chicago, IL, USA), and a two-sided P-value below 0.05 was considered significant.

RESULTS

A total of 33 new cases of PV and 33 age and sex-matched healthy controls completed this study. The general characteristics of the study participants are described in Table 1.

The median (interquartile range) NLR and PLR values in patients were 2.50 (1.94–6.59) and 90.30 (71.60–196.80), respectively, compared with 1.69 (1.45–2.30) and 56.00 (50.00–85.00) in controls. The NLR and PLR were significantly higher in patients than in controls (P<0.001 for both). However, the two groups had no significant difference in MPV (P=0.89) (Table 2). There was no correlation between either disease severity (ABSIS) or anti-Dsg1/3 levels and the studied hematological markers (NLR, PLR, and MPV). As expected, the anti-Dsg1 and anti-Dsg3 levels were positively correlated with disease severity (ABSIS) (r=0.43, P=0.01; and r=0.33, P=0.05, respectively).

DISCUSSION

The present study is the first prospective study evaluating the NLR and PLR in patients with pemphigus vulgaris (PV) that also focused on assessing the relationship of these factors with anti-Dsgs levels and disease severity. Our findings showed that NLR and PLR were higher in PV patients compared with the control group, though these markers did not correlate with the anti-Dsg1/3 levels or ABSIS scores.

Several studies have linked elevated NLR and PLR values with chronic inflammatory diseases and malignancies ^{8 , 15} . Also, these new inflammatory markers are useful prognostic factors and guide patients' follow-up in some diseases. In a study of patients with breast cancer, Ozyalvacli et al. showed that NLR was higher when compared with patients with benign proliferative breast disease as the control group ¹⁶ . Several meta-analyses have shown an association between a high NLR and worse long-term outcomes after treatment of various types of cancers ⁸ . In a cohort study of 437 women with breast cancer, Azab et al. showed that a high pretreatment NLR was a significant risk factor for increased mortality, regardless of the chemotherapy regimen ¹⁷ . In another study of patients with gastric cancer, Lee et al. showed that NLR and PLR were independent prognostic factors considering the overall survival of the patients ¹⁸ .

Additional information on the diagnostic and predictive value of blood cell ratios can be derived from autoimmune disease cohorts ¹⁵ . In a meta-analysis including 1246 SLE patients and 976 healthy controls, NLR and PLR were significantly higher in the SLE patients, and higher levels of PLR and NLR were found to reflect lupus activity ¹⁹ . In another study involving 154 SLE patients and 151 healthy controls, both PLR and NLR values were again higher in the patients ¹¹ . In the same study, patients with nephritis had higher NLR and PLR levels than those without nephritis. However, multiple regression analysis revealed that only a high NLR, not PLR, was predictive of lupus nephritis ¹¹ . An analysis of the blood cell ratio of patients with psoriasis and psoriatic arthritis also demonstrated that NLR and PLR were significantly higher in patients with psoriasis but were unrelated to the disease's severity. Nevertheless, both NLR and PLR were strong predictors for the presence of arthritis among psoriasis patients ^{20 , 21} .

To our knowledge, only one study highlights the role of NLR and PLR in PV patients. Hayta et al. reported that the NLR and PLR of PV patients were higher than controls but were unrelated to the disease severity. Their results are in agreement with our findings. Hayta et al. also showed that the MPV in patients with PV decreased in remission and increased with relapse. They concluded that the MPV index can be used as a marker of relapse in patients with PV ²² . Current literature data suggest that MPV can be used as a new marker of inflammation and provides information about the course and prognosis of some inflammatory and neoplastic conditions; however, this is still controversial ²³ . Yazici et al. showed that the MPV is significantly higher in patients with rheumatoid arthritis (RA) and is correlated with inflammatory markers (ESR and CRP) and also disease activity (PAS-28 score in RA patients) ²⁴ . Nevertheless, Sahin et al. showed that MPV inversely correlates with ESR, CRP, and DAS-28 scores in RA patients ²⁵ . In most neoplastic diseases, such as colorectal and gastric cancers, higher levels of MPV have been reported; in some other cancers, such as non-small cell lung cancer and cervical cancer, a decreased level may be found ²³ . In the current study, we did not find any difference in MPV levels between patients and controls, and there was no relationship between this parameter and anti-Dsg levels or disease severity.

The present study was limited by its relatively small population due to the rareness of PV and the inclusion of only new cases who had not received any treatment, such as steroids, cytotoxic drugs, and rituximab, which have known effects on hematological parameters ²⁶ . We did not explore the NLR or PLR changes in the remission phase of PV because of the influence of the aforementioned drugs on hematologic parameters.

CONCLUSION

Our study revealed that NLR and PLR are elevated in patients with PV but do not correlate with disease activity (evaluated by the ABSIS) or anti-Dsg1/3 levels. These laboratory parameters can be considered inflammatory markers of PV but cannot predict the disease activity.

Authors contributions

F.G. and N.M. designed and directed the project; Z.D. collected data; N.M. analyzed data and wrote the article. All authors discussed the results and commented on the manuscript.

Funding source

A grant from the Skin Research Center, Shahid Beheshti University of Medical Sciences, supported this project.

Acknowledgment

This residency study was supported by the Skin Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran, and this manuscript was based on the residency thesis of Dr Zohreh Rakhshan. The authors would like to thank the Clinical Research Development Unit (CRDU) of Loghman Hakim Hospital, Shahid Beheshti University of Medical Sciences, Tehran, Iran, for their support, cooperation, and assistance throughout the study period.

Conflict of interest

None declared.

References

1. Kasperkiewicz M, Ellebrecht CT, Takahashi H, et al. Pemphigus. Nat Rev Dis Primers. 2017; 3(1):1-18.
2. Joly P, Maho-Vaillant M, Prost-Squarcioni C, et al. First-line rituximab combined with short-term prednisone versus prednisone alone for the treatment of pemphigus (Ritux 3): a prospective, multicentre, parallel-group, open-label randomized trial. The Lancet. 2017; 389(10083):2031-40.
3. Tavakolpour S, Mahmoudi H, Balighi K, et al. Sixteen-year history of rituximab therapy for 1085 pemphigus vulgaris patients: a systematic review. Int Immunopharmacol. 2018; 54:131-8.
4. Murrell DF, Peña S, Joly P, et al. Diagnosis and management of pemphigus: recommendations by an international panel of experts. J Am Acad Dermatol. 2020; 82(3):575-85.
5. Kumar B, Arora S, Kumaran MS, et al. Study of desmoglein 1 and 3 antibody levels in relation to disease severity in Indian patients with pemphigus. Indian J Dermatol Venereol Leprol. 2006; 72(3):203.
6. Daneshpazhooh M, Chams‐Davatchi C, Khamesipour A, et al. Desmoglein 1 and 3 enzyme‐linked immunosorbent assay in Iranian patients with pemphigus vulgaris: correlation with phenotype, severity, and disease activity. J Eur Acad Dermatol Venereol. 2007; 21(10):1319-24.
7. Balighi K, Sakhi RS, Daneshpazhooh M, et al. Anti‐desmoglein‐1 levels as predictor of prednisolone tapering in pemphigus vulgaris patients treated with rituximab. Dermatol Ther. 2018; 31(6):e12671.
8. Faria SS, Fernandes Jr PC, Silva MJB, et al. The neutrophil-to-lymphocyte ratio: a narrative review. Ecancermedicalscience. 2016; 10:702.
9. Kapsoritakis AN, Koukourakis MI, Sfiridaki A, et al. Mean platelet volume: a useful marker of inflammatory bowel disease activity. Am J Gastroenterol. 2001; 96(3):776-81.
10. Gary T, Pichler M, Belaj K, et al. Platelet-to-lymphocyte ratio: a novel marker for critical limb ischemia in peripheral arterial occlusive disease patients. PLoS One. 2013; 8(7): e67688.
11. Qin B, Ma N, Tang Q, et al. Neutrophil to lymphocyte ratio (NLR) and platelet to lymphocyte ratio (PLR) were useful markers in assessment of inflammatory response and disease activity in SLE patients. Mod Rheumatol. 2016; 26(3):372-6.
12. Uslu AU, Küçük A, Şahin A, et al. Two new inflammatory markers associated with disease activity score‐28 in patients with rheumatoid arthritis: neutrophil‐lymphocyte ratio and platelet‐lymphocyte ratio. Int J Rheum Dis. 2015; 18(7):731-5.
13. Sen BB, Rifaioglu EN, Ekiz O, et al. Neutrophil to lymphocyte ratio as a measure of systemic inflammation in psoriasis. Cutan Ocul Toxicol. 2014; 33(3):223-7.
14. Pfütze M, Niedermeier A, Hertl M, et al. Introducing a novel autoimmune bullous skin disorder intensity score (ABSIS) in pemphigus. Eur J Dermatol. 2007; 17(1):4-11.
15. Gasparyan AY, Ayvazyan L, Mukanova U, et al. The platelet-to-lymphocyte ratio as an inflammatory marker in rheumatic diseases. Ann Lab Med. 2019; 39(4):345-57.
16. Ozyalvacli G, Yesil C, Kargi E, et al. Diagnostic and prognostic importance of the neutrophil lymphocyte ratio in breast cancer. Asian Pac J Cancer Prev. 2014; 15(23):10363-6.
17. Azab B, Bhatt VR, Phookan J, et al. Usefulness of the neutrophil-to-lymphocyte ratio in predicting short-and long-term mortality in breast cancer patients. Ann Surg Oncol. 2012; 19(1):217-24.
18. Lee S, Oh SY, Kim SH, et al. Prognostic significance of neutrophil lymphocyte ratio and platelet lymphocyte ratio in advanced gastric cancer patients treated with FOLFOX chemotherapy. BMC cancer. 2013; 13(1):350.
19. Ma L, Zeng A, Chen B, et al. Neutrophil to lymphocyte ratio and platelet to lymphocyte ratio in patients with systemic lupus erythematosus and their correlation with activity: a meta-analysis. Int Immunopharmacol. 2019; 76:105949.
20. Kim DS, Shin D, Lee MS, et al. Assessments of neutrophil to lymphocyte ratio and platelet to lymphocyte ratio in Korean patients with psoriasis vulgaris and psoriatic arthritis. J Dermatol. 2016; 43(3):305-10.
21. Paliogiannis P, Satta R, Deligia G, et al. Associations between the neutrophil-to-lymphocyte and the platelet-to-lymphocyte ratios and the presence and severity of psoriasis: a systematic review and meta-analysis. Clin Exp Med. 2019; 19(1):37-45.
22. Hayta SB, Guner R, Akyol M. Blood mean platelet volume may be predictive for disease course in the cases with pemphigus vulgaris. Biomed Res India. 2017; 28(9):4223-7.
23. Korniluk A, Koper-Lenkiewicz OM, Kamińska J, et al. Mean platelet volume (MPV): new perspectives for an old marker in the course and prognosis of inflammatory conditions. Mediators Inflamm. 2019; 2019: 9213074.
24. Yazici S, Yazici M, Erer B, et al. The platelet indices in patients with rheumatoid arthritis: mean platelet volume reflects disease activity. Platelets. 2010; 21(2):122-5.
25. Şahin A, Yetişgin A, Şahin M, et al. Can mean platelet volume be a surrogate marker of inflammation in rheumatic diseases?. West Indian Med J. 2016; 65(1):165-169.
26. Kaegi C, Wuest B, Schreiner J, et al. Systematic review of safety and efficacy of rituximab in treating immune-mediated disorders. Front Immunol. 2019; 10:1990.