| Abstract|| |
Wegener's granulomatosis (WG) patients can rarely have antineutrophil cytoplasmic antibodies (ANCAs) directed against myeloperoxidase (MPO), producing a cytoplasmic pattern on indirect immunofluorescence (IIF). This has important implications in the diagnosis and pathophysiology of the disease. We present to you a report of three cases of WG, demonstrating a cytoplasmic-ANCA pattern on indirect IIF, but directed against MPO. It is necessary to diagnose a patient taking into account both the autoimmune test results and the clinical features.
Keywords: Cytoplasmic antineutrophil cytoplasmic antibodies, immunofluorescence, myeloperoxidase, Wegener′s granulomatosis
|How to cite this article:|
Venkatesh BM, Joshi S, Adhikary R. Cytoplasmic-anti-neutrophil cytoplasmic antibodies targeting myeloperoxidase in Wegener's granulomatosis: A rare phenomenon. Indian J Pathol Microbiol 2014;57:470-2
|How to cite this URL:|
Venkatesh BM, Joshi S, Adhikary R. Cytoplasmic-anti-neutrophil cytoplasmic antibodies targeting myeloperoxidase in Wegener's granulomatosis: A rare phenomenon. Indian J Pathol Microbiol [serial online] 2014 [cited 2020 May 29];57:470-2. Available from: http://www.ijpmonline.org/text.asp?2014/57/3/470/138777
| Introduction|| |
Anti-neutrophil cytoplasmic antibodies (ANCA) are a group of autoantibodies, mainly of the IgG type, directed against antigens in the cytoplasm of granulocytes and monocytes. They are important serologic markers for vasculitis, and are directed against several myeloid enzymes. By indirect immunofluorescence (IIF) microscopy, there are two main patterns of ANCA reactivity that are found on substrates of peripheral blood neutrophils fixed in ethanol: Cytoplasmic-ANCA (c-ANCA) and perinuclear-ANCA (p-ANCA). The c-ANCA pattern is due to reactivity with serine proteinase 3 (PR3). The p-ANCA pattern is due to reactivity with myeloperoxidase (MPO). In atypical ANCA, the IIF is positive (mostly p-ANCA), but the serum is negative for both PR3 and MPO, suggesting a different antigenic target. 
Cytoplasmic-antineutrophil cytoplasmic antibodies/PR3-ANCA is strongly associated with Wegener's granulomatosis (WG), whereas p-ANCA/MPO-ANCA is associated with a diverse disease spectrum such as microscopic polyangiitis, other vasculitis and auto immune conditions. Atypical ANCA is seen in a wide variety of medical conditions. Fewer than 5% of WG patients are MPO-ANCA positive. 
Here, we present to you a report of three cases, with ANCA demonstrating a cytoplasmic fluorescence pattern on IIF but directed against MPO.
| Case ReportS|| |
A 56-year-old hypertensive female patient presented with fever, back pain, weakness, and numbness in lower limbs from 15 days. There was no history of bleeding episodes, dysuria or hematuria. Hematological investigations revealed an increase in total leukocyte count, erythrocyte sedimentation rate (ESR), and absolute eosinophil count. Serum total IgE was also increased. Urine routine and other renal parameters were normal. C-reactive protein (CRP) and rheumatoid factor were positive. Computed tomography (CT) scan showed patchy consolidation in the upper lobes of both the lungs with mild mediastinal lymphadenopathy. An empirical diagnosis of tuberculosis or an autoimmune disorder was done and investigated appropriately. Sural nerve biopsy showed vasculitic neuropathy with significant axonal damage. Biopsy of the right nasal turbinate showed non-specific inflammation. Sputum for acid-fast bacillus (AFB), urine, and blood cultures were negative.
A 45-year-old male, known diabetic and hypertensive patient presented with myalgia, recurrent attacks of cold and sinusitis, hip pain, and recurrent avascular necrosis of the right head of femur. Complete hemogram of the patient was normal except an increased ESR. Renal profile and urine routine were normal. CRP was positive. Magnetic resonance imaging (MRI) of the temporal bone revealed right mastoiditis and bilateral ethmoid sinusitis. Due to multiorgan involvement, the patient was extensively investigated to rule out autoimmune etiology. CT chest was normal. Sputum for AFB, urine and blood cultures were negative.
A 55-year-old male patient with a history of recurrent left ethmoidal and maxillary sinusitis presented to the ophthalmology outpatient department with prominence of the left eye, associated with pain, redness and watering since 2 months. On fundoscopy, disc pallor, dense asteroid and patches of peripheral degeneration were noted in both the eyes. His blood counts were normal, ESR was raised. Renal profile and urine routine were normal. CRP was positive. CT scan of orbits showed diffuse soft tissue opacification in left maxillary sinus with intraorbital extension. MRI showed transverse sinus and sigmoid sinus thrombosis. Functional endoscopic sinus surgery of left maxillary sinus and orbital biopsy was done. Histopathological examination of the left maxillary sinus tissue showed features suggestive of chronic inflammation/fibroinflammatory disease. A preliminary diagnosis of recurrent left ethmoidal and maxillary aggressive sinusitis with orbital extension was done and autoimmune investigations were carried out.
Autoimmune workup of all the three patients was done by indirect IIF and immunoblot assay. ANCA - IIF, was done using the granulocyte biochip mosaic slides with both ethanol and formalin fixed neutrophils, (Euroimmun, Germany). It revealed a positive c-ANCA pattern. Antinuclear antibody (ANA) was negative by IIF. ANCA immunoblot assay (Euroimmun Germany) in all these patients showed a strong positive band for MPO. PR3 was negative. Positive and negative controls were incorporated in all the runs.
All the patients were negative for human immunodeficiency virus (HIV), hepatitis B and hepatitis C infection by ELISA.
A final diagnosis of WG was made based on the American College of Rheumatology criteria and the patients were put on pulsed cyclophosphamide therapy. 
| Discussion|| |
Antineutrophil cytoplasmic antibodies testing are an important tool in the diagnosis of small vessel vasculitis. Two international consensus statements currently form the basis for optimizing the detection and characterization of ANCA. , The consensus recommends that ANCA reactivity on ethanol fixed slides should be confirmed by enzyme immunoassay to find out ANCA specific for PR3 or MPO.
False positive IIF staining patterns can be seen in infectious and autoimmune conditions, which have to be ruled out.  Such false positive patterns have been reported in cases of infection by HIV. HIV infection may evoke ANCA in 20-83% of cases, probably due to polyclonal activation of B cells, but not associated with hypergammaglobulinaemia. ,, All our patients were HIV negative and had symptoms suggestive of an underlying autoimmune disease process. There were no findings suggestive of other infectious causes. ANA IIF was negative, ruling out other systemic autoimmune conditions. We have confirmed the ANCA IIF positivity by an immunoblot reaction where MPO specific ANCA band was obtained. The samples were negative for anti PR3 antibodies.
In view of potential phenotypic similarities between tuberculosis and WG, three samples of sputum from all the patients were sent for AFB smear, which were negative. ,
Schönermarck et al. have reported that MPO-ANCA positivity in WG patients is rare and these patients have a lower disease extent than PR3-ANCA positive patients. According to their study, these patients have a lower frequency of peripheral neuropathy, kidney and eye involvement compared to PR3-ANCA positive patients.  Similarly, all our patients have predominantly respiratory system involvement. The renal parameters and urine routine were normal, suggesting an absence of involvement of the kidneys.
| Conclusion|| |
Myeloperoxidase-ANCA not only has important diagnostic implications, but also has role in the pathophysiology of WG. It is necessary to be aware of this rare phenomenon while reporting and analyzing ANCA positivity to arrive at an accurate diagnosis. We also would like to reinforce that though ANCA is an important serological tool to assist in the diagnosis of WG and other vasculitis, it should always be interpreted with clinical features. The combined use of IIF tests and solid phase assays to detect ANCA directed against MPO and PR3 can minimize the occurrence of false positive results.
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Bhavana M Venkatesh
#98, HAL Airport Road, Bengaluru - 560 017, Karnataka
Source of Support: None, Conflict of Interest: None