| Abstract|| |
Context: Bullous pemphigoid (BP) and “Pemphigus diseases” (PD) can have overlapping clinical manifestations and accurate distinction is crucial for appropriate management. Aims: The study aimed at analyzing the utility of simple hematological markers of systemic inflammation like neutrophil-to-lymphocyte ratio (NLR), neutrophil-to-eosinophil ratio (NER), and platelet-to-lymphocyte ratio (PLR) in clinical decision making in the setting of clinical differentials of BP and PD in a particular case. Methods: This single-centre based retrospective observational analytical study included adult subjects newly diagnosed to have BP (n=66) or PD (n=53), confirmed with direct immune-fluorescence testing, over a period of six years. Blood counts performed using Coulter™ hematology analyser, at the time of their initial presentation, were retrieved from the hospital medical records, and the leucocyte ratios were calculated.Statistical Analysis: The data were compared between the two groups, using Mann–Whitney U test and chi-square test /Fisher's exact test. ROC curve analysis was performed to estimate cut-off values. Results: The BP group had a significantly higher NLR, total leukocyte counts (TLC), absolute eosinophil counts (AEC), and absolute lymphocyte counts (ALC), and lower NER values compared to the PD group (P < 0.05). Areas under ROC for NLR, NER, TLC, AEC, and ALC were between 0.5 and 0.7. NLR ≥ 7, AEC ≥ 2055/cumm, and TLC ≥ 15,000/cumm had a specificity of 90.6, 100, and 100% respectively for identifying BP patients out of the two groups, but with a low sensitivity of 22.7, 21, and 22.7%, respectively. Conclusions: NLR can be a valuable diagnostic adjunct in subtyping autoimmune bullous disorders, albeit in a small proportion of cases.
Keywords: Autoimmune bullous disorders, bullous pemphigoid, NER, NLR, PLR, pemphigus vulgaris
|How to cite this article:|
Rai P. Role of neutrophil-to-lymphocyte, neutrophil-to-eosinophil and platelet-to-lymphocyte ratios in the diagnosis of bullous pemphigoid and Pemphigus disease. Indian J Pathol Microbiol 2023;66:70-4
|How to cite this URL:|
Rai P. Role of neutrophil-to-lymphocyte, neutrophil-to-eosinophil and platelet-to-lymphocyte ratios in the diagnosis of bullous pemphigoid and Pemphigus disease. Indian J Pathol Microbiol [serial online] 2023 [cited 2023 Mar 20];66:70-4. Available from: https://www.ijpmonline.org/text.asp?2023/66/1/70/367956
| Introduction|| |
Bullous pemphigoid (BP) and “Pemphigus diseases” (PDs) are two major forms of autoimmune bullous disorders (ABDs) having distinct clinical, histopathological, immunofluorescence, and serological findings. However, often they can have overlapping clinical presentations making their distinction from each other difficult based solely on clinical findings. Differences in preferred lines of management make their correct diagnosis crucial., This study aimed at comparing the well-known hematological markers of systemic inflammation like neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), and neutrophil-to-eosinophil ratios (NER) between patients diagnosed to have BP and PD, and to evaluate their efficacy in accurately distinguishing patients having BP from those having PD.
| Subjects and Methods|| |
This study was a retrospective observational analytical study conducted in a South Indian teaching hospital. Ethical approval was obtained from the institutional ethics committee. The study included 66 patients with BP disease and 53 patients with PD. These comprised of adult patients, above 12 years of age, who presented to the Dermatology Outpatient Department (OPD) of the hospital between January 2014 and December 2019 (6-year-period), and were newly diagnosed to have BP or PD, following confirmation with histopathological and direct immune-fluorescence testing in the pathology department of the institute. Pediatric patients, below 12 years of age, were excluded from the study. The total leukocyte counts (TLC), differential leukocyte counts (DLC), and platelet counts (PLT) of the enrolled patients were done at the time of their initial presentation to the dermatology OPD as a part of the routine investigation were retrieved from the hospital electronic medical records. Those subjects whose hematology parameters were unavailable for the study were excluded from the study. The blood counts were performed using Coulter™ automated hematology analyzer (manufactured by Beckman Coulter, Canada L.P., Mississauga), in the hematology department of the hospital laboratory using the complete blood count (CBC) mode. The absolute neutrophil counts (ANC), absolute lymphocyte counts (ALC), absolute eosinophil counts (AEC), NLR, PLR, and NER were subsequently calculated from the data collected. The NLR was calculated by dividing the ANC by ALC. The PLR was calculated by dividing the platelet count by ALC. The NER was calculated by dividing the ANC by AEC. The numerical data were described using mean, standard deviation, median, and inter-quartile range, and compared between the two study groups using the Mann–Whitney U test. The presence and absence of eosinophilia and the distribution of mild, moderate, and severe eosinophilia were described using frequency and percentage, and compared between the two groups using the Chi-square test/Fisher's exact test.
Statistical tests were performed using SPSS (Statistical Product and Service Solutions, developed by IBM corporation), version 23. P <0.05 was considered statistically significant, with P < 0.01 being statistically highly significant. For the variables that showed statistically significant difference among the two groups, the area under the ROC curve (AUC) was used to identify the best threshold and to evaluate their ability to accurately distinguish BP patients from PD patients. Pearson's correlation test was used to determine the correlation between two variables.
| Results|| |
Over a period of 6 years, a total of 76 cases of BP and 64 cases of PD were diagnosed with confirmation by histopathological and direct immunofluorescence testing. Out of these, 66 and 53 patients of BP and PD, respectively, were above 12 years of age and had undergone basic hematological investigations at the time of initial presentation, and hence, were included in the study. Of the 53 PD cases, 49 were pemphigus vulgaris (PV) and four were pemphigus foliaceus (PF) cases. The age of the subjects in the BP group ranged from 13 to 91 years, with a mean age ± standard deviation of 64 ± 15.5. The age of the subjects in the PD group ranged from 19 to 75 years with a mean age ± standard deviation of 47 ± 13.8. The group wise mean, standard deviation, median, and inter quartile range of TLC, ANC, ALC, AEC, platelet count, NLR, PLR, and NER, and the P values obtained on comparison of these parameters between the two groups using the Mann–Whitney U test are depicted in [Table 1]. Statistically significant differences were found in the TLC, ALC, AEC, NLR, and NER values between the two study groups with the differences in AEC values being highly significant (P = 0.004). TLC, ALC, AEC, and NLR were significantly higher in BP patients compared to the PD patients, whereas, NER was significantly higher in PD patients compared to the BP patients. No significant differences were found in the values of ANC, PLT, and PLR between the two groups.
|Table 1: Group-wise mean, median, standard deviation, inter-quartile range (IQR) and Mann-Whitney P of the various study parameters|
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Leukocytosis (as defined by a leucocyte count of >11,000/cumm) was present in 35 (53%) BP patients and 18 (34%) PD patients; the difference was statistically significant (P = 0.038; Chi-square test). Eosinophilia (defined by AEC >500/cumm) was significantly more frequent (P = 0.016; Chi-square test) in the BP patients (n = 37; 56%) compared to the PD patients (n = 18; 34%). Severe eosinophilia (defined by AEC >5000/cumm) was found in four (6.1%) of the BP patients and none of the PD patients. However, the difference was not statistically significant (P = 0.128; Fisher's exact test). Group-wise distribution of mild, moderate, and severe degrees of eosinophilia is depicted in [Figure 1].
|Figure 1: Group-wise distribution of various degrees of eosinophilia among the subjects|
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Correlation between the age of the patients and presence of blood eosinophilia was analyzed for each group, using Pearson's correlation coefficient, which was 0.133 (P = 0.286) for patients with BP, and 0.078 (P = 0.577) for the patients with PD. No significant correlation was found between the two variables within both the study groups.
The AUC obtained from the ROC curve analysis for NLR, NER, TLC, AEC, and ALC along with their respective P values and confidence intervals are depicted in [Table 2].
|Table 2: The area under the receiver operating characteristic (ROC) curve (AUC) for the study parameters which showed statistically significant differences between bullous pemphigoid and pemphigus vulgaris patients|
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NLR values of greater than or equal to 7 units had a specificity of 90.6% and a sensitivity of 22.7% for identifying BP patients out of the two groups of patients, whereas, NLR values greater than or equal to 5 had a specificity of 83% and a sensitivity of 42.4% for the same. Considering the normal range of NLR between 0.78 and 3.53, 60.6% (n = 40) and 58.5% (n = 31) patients in BP and PD group, respectively, had NLR above the upper limit of 3.53. This difference was not statistically significant (P = 0.815) on analysis with the Chi-square test.
NER values greater than or equal to 60 units had a specificity of 80.3% and a sensitivity of 28.3% for identifying PD patients out of the two groups of patients, whereas, NER values greater than or equal to 13 units had a specificity of 54.5% and a sensitivity of 69.8% for the same.
AEC values of greater than or equal to 2055/cumm had 100% specificity for identifying BP patients out of the two groups of patients, but with a very low sensitivity of 21%. AEC values of greater than or equal to 400/cumm had a specificity of 57% for identifying BP patients out of the two groups of patients, with a sensitivity of 62%. TLC values of greater than or equal to 15,000/cumm had 100% specificity for identifying BP patients out of the two groups of patients, but with a very low sensitivity of 22.7%. TLC values of greater than or equal to 10,000/cumm had 51% specificity and 60.6% sensitivity for identifying BP patients out of the two groups of patients. ALC values of greater than or equal to 1775/cumm had a specificity of 54.5% for identifying BP patients out of the two groups of patients with a sensitivity of 62.3%.
| Discussion|| |
ABDs encompass two major heterogenous groups of diseases namely “PDs” and “autoimmune bullous diseases of the pemphigoid type,” characterized by the production of autoantibodies targeting various epidermal or subepidermal adhesion proteins. The major forms of PDs include PV, PF, and paraneoplastic pemphigus. BP is the most frequent form of ABD of the pemphigoid type. Often the ABDs can have overlapping clinical presentations, which makes the correlation of clinical findings with the results of laboratory tests including histopathology and direct immunofluorescence (DIF) microscopy and/or serology necessary to arrive at the correct diagnosis. Accurate early subtyping of ABDs has therapeutic implications.,
ABDs are associated with varying degrees of systemic inflammation, which is often reflected in the form of deranged blood cell counts.,, Leukocyte ratios like NLR and PLR are well-known markers of systemic inflammation, which can be easily calculated from the test results of simple and inexpensive hematological investigations like TLC and DLC. NLR has been reported to be elevated in various inflammatory conditions, including psoriasis, atopic dermatitis, PV, BP, Behçet's disease, and cutaneous vasculitis.,,,,,
The correlation between NLR values and the severity of systemic inflammation has been reported. Different studies have shown significantly higher NLR values in either BP or PV patients, compared to normal controls.,,, In these studies, PLR values of BP or PV patients were not significantly different from the normal control groups. Studies comparing these hematological markers of systemic inflammation like NLR, PLR, and NER between patients with BP and PD are limited in the literature.
In the present study, statistically significant differences were found in the NLR, NER, TLC, and AEC values between the two study groups. However, according to ROC curve analysis, although areas under curve for NLR, NER, TLC, AEC, and ALC were above 0.6, none were above 0.7 indicating a poor performance of these parameters as classifiers to distinguish between BP and PD patients independently. This is attributable to the fact that the test result values that had high specificity for a disease were found only in a small proportion of cases of that disease, thereby resulting in a low sensitivity for the same. Nevertheless, the findings in the study of high specificity of NLR values ≥7 units (90.6%), AEC values ≥2055/cumm (100%), and TLC values ≥15,000/cumm (100%) for identifying BP patients, and of NER values greater than or equal to 60 units (80.3%) for identifying PD patients out of the two groups of patients, can be helpful to some extent in clinical decision making, albeit in a small proportion of cases owing to their low sensitivities (less than 30%) for the same. These parameters might have a better role in the discrimination of pemphigus and pemphigoid disorders in acute and severe cases, where higher levels of NLR, AEC, and TLC values can be expected to be seen.
The finding of comparable PLR values between the two study groups in the present study may be attributed to the fact that PLR values in BP or PV patients do not differ significantly from the normal control groups as reported by some studies.,,,
High NLR was found to be independently associated with mortality in patients with BP in a study done in the Thai population. Lyakhovitsky A et al. reported a positive correlation of NLR and PLR with disease activity in PV patients in their study, however, in some studies, no correlation was found between disease severity and NLR values in PV patients., In the present study, significantly higher NLR was found in BP patients, compared to PD patients at the time of initial presentation, suggesting that BP patients frequently tend to present with more severe degrees of systemic inflammation compared to PD patients. It could also indicate that BP patients are more prone to infections than PD patients.
Even hematological changes, like leukocytosis and eosinophilia, were significantly more frequent among the BP patients compared to the PD patients in this study.
The role of eosinophils in blister formation in BP is well-known. The prevalence of peripheral blood eosinophilia in patients with BP has been reported to be around 50–60% in the literature, which is concordant with the finding in the present study. Secretion of several inflammatory mediators like interleukin-5 (IL-5), interleukin-3 (IL-3), and granulocyte-macrophage colony-stimulating factor (GM-CSF) has been implicated in the pathogenesis of peripheral blood eosinophilia. In a study done by Farnaghi F et al., significant correlation was reported between the percentage of peripheral blood eosinophils and subjective Bullous Pemphigoid Disease Activity Index (BPDAI) scores and urticarial/eczematous lesions in patients with BP. The findings in the present study of significantly higher AEC values significantly increased frequency of eosinophilia, and significantly lower NER values in the patients with BP compared to the patients with PD can be attributed to the key role of eosinophils in the pathogenesis of BP. Additionally, severe eosinophilia (AEC >5000/cumm) was found exclusively in the BP group in the study; however, the finding was not statistically significant. An independent association between absolute eosinophil count and the diagnosis of BP was observed in a case-control study by Kridin K, wherein the eosinophil count of patients with BP was compared with age-, sex- and ethnicity-matched controls. In the same study, BP patients having blood eosinophilia were reported to be significantly older than those without the same. In contrast to that no significant correlation between the age of the patients and the presence of blood eosinophilia was found in the present study within both the study groups.
The potential limitations of the study include the relatively small sample size and the lack of correlation of the study findings with the duration of illnesses, the clinical disease activity scores, the presence of other co- morbidities, and ongoing medications at the time of performing the blood counts. The activity of disease tends to change with the duration of illness, which may be reflected as dynamic changes in the leukocyte counts at different phases of the disease within each patient. The low sensitivity of the study parameters may be attributed to the differences in the number of patients having similar degrees of disease severity and duration of illnesses between the study groups. The comparison of these parameters between BP and PD patients having comparable disease activity scores and duration of illnesses can provide a better insight into their role in the discrimination of pemphigus and pemphigoid disorders in various phases of these illnesses. In this study, the disease intensity scores and duration of illnesses of the subjects were not recorded, and hence, not taken into consideration.
In conclusion, hematological markers of systemic inflammation like NLR, NER, TLC, AEC, and ALC have a limited role as diagnostic tests that aid in distinguishing patients with BP from those with PD, despite exhibiting statistically significant differences between the two groups. PLR and ANC values are comparable in patients with BP and PD, and hence, have no diagnostic role in this context. Given the clinical differentials of BP and PD in a particular case, NLR values of greater than or equal to 7 units, AEC values of greater than or equal to 2055/cumm, and TLC values of greater than or equal to 15,000/cumm can serve as useful markers favoring a diagnosis of BP over PD. However, the low sensitivity of these cut-off values limits their diagnostic utility to only a small proportion of such cases. Further studies, comparing these parameters between BP and PD patients having comparable disease intensity scores and duration of illnesses, are required to understand the influence of disease severity and duration of illnesses on the sensitivity and specificity of these parameters.
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Assistant Professor, Department of Pathology, Father Muller Medical College, Mangalore - 575 002, Karnataka
Source of Support: None, Conflict of Interest: None
[Table 1], [Table 2]