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Tuesday, 30 April 2019

USO DE ANTICUERPOS COMO BIOMARCADORES EN EL DIAGNÓSTICO DE LA ENFERMEDAD INFLAMATORIA INTESTINAL

Written by Ana Sopena Murillo | Adrián Fontán Abad | Mercedes Sánchez González, Posted in Volumen10

Figure 1: Antibodies against neutrophil cytoplasm (ANCA) on neutrophils fixed in ethanol (pANCA pattern).

Figure 2: antibodies against neutrophil cytoplasm (ANCA) in formalin (cANCA pattern) in patient with inflammatory bowel disease by IIF (40x).
A 15-year-old man from the Gambia is referred to Digestive Service for abdominal pain and bloody diarrhea of four years of evolution. Stool study are requested (parasites and coproculture: negative) and gastroscopy, where the presence of Helicobacter pylori is found, receiving treatment. In addition, a colonoscopy and biopsies are performed, which show changes compatible with inflammatory bowel disease (IBD) suggestive of ulcerative colitis (UC).

One year later, gastrointestinal endoscopy and biopsies are repeated, the results of which suggest the presence of Crohn’s disease (CD), depending on endoscopic appearance and histology (chronic ileocolitis patched with destruction of the crypts, erosion and ulceration without reduction of the mucosecretor component). Calprotectin was requested during follow-up [458 mg/Kg; 1463 mg/Kg; VR<50 mg/Kg], supporting the diagnosis of IBD, and blood tests highlighting: alkaline phosphatase [493 IU/L; 272 IU/L; VR: 40-129 IU/L], gamma-glutamyltransferase [134 IU/L; 233 IU/L; VR: 0-29 IU/L] and alanine aminotransferase [47 IU/L; 63 IU/L; VR: 0-41 IU/L]. According to the hepatic alteration, magnetic resonance enterography was requested, reporting as a possible start of primary sclerosing cholangitis (PSC), and autoimmunity study: positive speckled pattern antinuclear antibodies (ANA) at 1/160; positive smooth muscle antibodies (SMA) at 1/80; positive antineutrophil cytoplasmic antibodies (ANCA) with pANCA pattern (no dilutions were made) (Figure 1) all of them by indirect immunofluorescence (IIF). As a result of these tests were also determined: autoantibodies to extractable nuclear antigens (ENAs) by enzyme-linked immunosorbent assay (ELISA) and F-actina by immunoblot that were negative, myeloperoxidase (MPO) and proteinase-3 (PR3-ANCA) were determined by ELISA being positive for PR3-ANCA [294.9 U/mL; VR: 0-7 U/mL]. In view of the positivity for ANCA and PR3-ANCA antibodies against Saccharomyces cerevisiae (ASCA), useful in differential diagnosis of UC and CD, were requested. The result was negative.

IBD is an inflammatory disorder of the gastrointestinal tract. The most representative forms are UC and CD. Its differentiation is important, but diagnosis based on endoscopic and histological criteria is not always easy1. Because endoscopic tests and biopsies are invasive, expensive and slow, serological markers that help the IBD differential diagnosis have been searched, especially in doubtful cases, being the best studied the atypical perinuclear ANCA (aANCA) and the ASCA that help differentiate them. ASCA positivity is related to CD (68% of patients) and aANCA to UC (40-80% of patients)2. The cANCA pattern in IIF is a marker, principally, of Wegener´s granulomatosis and its main antigen is PR3-ANCA. The pANCA pattern is associated with microscopic polyangiitis, pauci-immune crescentic necrotizing glomerulonephritis (GN) and Churg-Strauss syndrome and its main antigen is MPO. In IBD, ANCA specificity is not well defined. Recent studies show a possible association between PR3-ANCA and IBD, and its use as a diagnostic marker in these diseases, mainly in CU3-4. PSC is associated with IBD, in 79-80% of cases of UC5. Some studies show that PR3-ANCA can also be a specific marker of PSC in the differential diagnosis of hepatopathies6. The positivity of PR3-ANCA varies depending on the method and laboratory. The rate of positive cases is of 29.3-31.1% in patients with UC versus 1.9-2.7% in patients with CD using chemiluminescent immunoassays (CIA) (QUANTA-Flash® PR3, INOVA Diagnostics)3-4. When using ELISA (QUANTA-Lite® PR3, INOVA Diagnostics) this positivity decreases to 6% in UC and 0% in EC4. The diagnostic accuracy, sensitivity, specificity and likelihood ratio of the different techniques to discriminate between UC and CD are shown in table 1. The combination PR3-ANCA-positive (CIA technique and cut-off = 11.8 CU) and IgA ASCA-negative is the best tool to discern between UC and CD with high specificity and predictive value. In the case at hand, the realization of autoimmune serology can be an aid in the differential diagnosis. The presence of PR3-ANCA-positive, ASCA-negative and the association with PSC support the diagnosis of UC.  

In conclusion, although the diagnosis of IBD is made using endoscopic and histological criteria, in some cases the search for autoimmune biomarkers can support the definitive diagnosis and thus avoid carrying out more invasive and expensive tests.

TABLA
Table1. Diagnostic accuracy, sensitivity, specificity and likelihood ratio to differentiate between UC and EC3.

Reference
  1. Mowat C, Cole A, Windsor A, Ahmad T, Arnott I, Driscoll R, et al. Guidelines for the management of inflammatory bowel disease in adults. Gut. 2011;60:571–607.
  2. Papp M, Altorjay I, Norman GL, Lakatos PL. Utility of serological markers in inflammatory bowel diseases: gadget or magic? World Journal of Gastroenterology. 2007;13:2028–2036.
  3. Arias-Loste MT, Bonilla G, Moraleja I, Mahler M, Mieses MA, Castro B, et al. Presence of anti-proteinase 3 antineutrophil cytoplasmic antibodies (anti-PR3 ANCA) as serologic markers in inflammatory bowel disease. Clin Rev Allergy Immunol. 2013;45:109-16.
  4. Mahler M, Bogdanos DP, Pavlidis P, Fritzler MJ, Csernok E, Damoiseaux J, et al. PR3-ANCA: a promising biomarker for ulcerative colitis with extensive disease. Clin Chim Acta. 2013;424:267–73
  5. Wiesner RH, Grambsch PM, Dickson ER, Ludwig J, Maccarty RL, Hunter EB, et al. Primary sclerosing cholangitis: Natural history, prognostic factors and survival analysis. Hepatology. 1989;10:430–436.
  6. Stinton LM, Bentow C, Mahler M, Norman GL, Eksteen B, Mason AL, et al. PR3-ANCA: a promising biomarker in primary sclerosing cholangitis (PSC). PLoS One. 2014;9:e112877.

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