Paroxysmal nocturnal hemoglobinuria: Test to monitor the action of eculizumab treatment.

INTRODUCTION: Paroxysmal nocturnal hemoglobinuria (PNH) is caused by a somatic mutation in the PIG-A gene, which encodes for glycosylphosphatidylinositol, a phospholipid membrane that anchors proteins like CD55 and CD59. These proteins are inhibitors of the complement-mediated lysis. PNH is diagnosed by flow cytometry, and treatment with eculizumab improves the life quality of patients with severe clinical compromise. The aim of this work was to evaluate a hemolytic test that allows monitoring the blockade of the alternative complement pathway caused by eculizumab (herein MET test). METHODS: There were analyzed a total of 163 serum samples from nine patients with PNH under treatment with eculizumab and ten healthy volunteers like controls. The patients were evaluated for 6 months. The MET test consisted in incubating red blood cells from patients (RBCPNH ) with either acidified serum from healthy volunteers and from patients with PNH. The results can be (a) Positive, (b) Blockade profile, or (c) Negative. RESULTS: Seven patients responded favorably to the eculizumab, and the test evidenced the blockade profile. The two remaining patients were nonresponders to the treatment, with a positive MET test. In these patients, the dose was increased. One responded favorably with a blockade profile, and the other continued to be nonresponder. CONCLUSIONS: The MET test proved to be a useful tool to monitor the blockade of complement by eculizumab.

[Antinuclear antibodies, patterns and characteristics obtained by immunofluorescence. The importance of the IgA, IgM and IgG isotypes]. / Anticuerpos antinucleares, imágenes y características obtenidas por inmunofluorescencia. Importancia de los isotipos IgA, IgM e IgG.

The indirect immunofluorescence with epitelial cell line from human laryngeal carcinoma as substrate (IIF-HEp2) and anti-IgG or anti-total Ig as antisera, is the technique currently used for the detection of antinuclear antibodies. The most important antibodies for the diagnosis and follow-up of connective tissue diseases (CTD) are the IgG-ANA, while the IgM-ANA have no clinical relevance. However the IgA-ANA have not been thoroughly investigated so far. The aim of this work was to study the prevalence of different ANA isotypes of Ig antibodies in CTD patients and to evaluate the convenience of the use of monovalent or polyvalent conjugate. We examined the sera of 100 patients with different CTD by IIF-HEp2 and detected a prevalence of 38% IgA-ANA (titles > or = 1:80) and 12% IgM-ANA (titles < or = 1:160). In twenty nine cases we detected IgA-ANA in absence of IgM-ANA, and in 3 cases IgM-ANA in absence of IgA-ANA. In all the cases IgG-ANA were present. In 6 sera a change in the immunofluorescence pattern was observed while using anti-IgA conjugate, whereas in 3 the change was observed with the use of anti-IgM conjugate. Because of the high prevalence of ANA-IgA detected by IIF-HEp2, we emphasize the convenience of employing anti-total Ig in spite of anti-IgG conjugated until the role of ANA-IgA is dilucidated in CTD patients, in order to establish its relevance in the diagnosis, prognosis and follow-up of systemic rheumatic diseases.

Anticuerpos antinucleares, imágenes y características obtenidas por inmunofluorescencia: Importancia de los isotipos IgA, IgM e IgG / Antinuclear antibodies, patterns and characteristics obtained by immunofluorescence: The importance of the IgA, IgM and IgG isotypes

La técnica de elección para el screening de anticuerpos antinucleares (ANA) es la inmunofluorescencia indirecta que utiliza como sustrato una línea de células epiteliales de carcinoma de laringe humano (IFI-HEp2), y como antisuero, anti-IgG o anti-Ig totales. Los ANA-IgG son los más importantes para el diagnóstico y monitoreo de las enfermedades del tejido conectivo (ETC), mientras los ANA-IgM son de menor relevancia clínica en estos pacientes. Sin embargo, poco se sabe de los ANA-IgA ya que estos Ac han sido menos investigados. El objetivo de este trabajo fue estudiar la prevalencia de los diferentes isotipos de inmunoglobulinas de anticuerpos antinucleares en los pacientes con ETC y evaluar la conveniencia de utilizar conjugados monovalentes o polivalentes. Se procesaron 100 sueros de pacientes con diversas ETC empleando IFI-HEp2, en los cuales se detectó 38% de ANA-IgA (títulos ≥ 1:80) y 12% de ANA-IgM (títulos ≤ 1:160). En 29 casos se detectó IgA en ausencia de IgM, en 3 casos IgM en ausencia de IgA. En todos los casos los ANA-IgG estuvieron presentes. En 6 sueros se observó un cambio de imagen con conjugado anti-IgA y en 3 con conjugado anti-IgM. Debido a la alta prevalencia de ANA-IgA detectada por IFI-HEp2, se destaca la conveniencia de utilizar conjugado anti-Ig totales en lugar de anti-IgG, mientras se desconozca la relevancia de los ANA-IgA en el diagnóstico, pronóstico y seguimiento de las enfermedades reumáticas sistémicas.

The indirect immunofluorescence with epitelial cell line from human laryngeal carcinoma as substrate (IIF-HEp2) and anti-IgG or anti-total Ig as antisera, is the technique currently used for the detection of antinuclear antibodies. The most important antibodies for the diagnosis and follow-up of connective tissue diseases (CTD) are the IgG-ANA, while the IgM-ANA have no clinical relevance. However the IgA-ANA have not been thoroughly investigated so far. The aim of this work was to study the prevalence of different ANA isotypes of Ig antibodies in CTD patients and to evaluate the convenience of the use of monovalent or polyvalent conjugate. We examined the sera of 100 patients with different CTD by IIF-HEp2 and detected a prevalence of 38% IgA-ANA (titles ≥ 1:80) and 12% IgM-ANA (titles ≤ 1:160). In twenty nine cases we detected IgA-ANA in absence of IgM-ANA, and in 3 cases IgM-ANA in absence of IgA-ANA. In all the cases IgG-ANA were present. In 6 sera a change in the immunofluorescence pattern was observed while using anti-IgA conjugate, whereas in 3 the change was observed with the use of anti-IgM conjugate. Because of the high prevalence of ANA-IgA detected by IIF-HEp2, we emphasize the convenience of employing anti-total Ig in spite of anti-IgG conjugated until the role of ANA-IgA is dilucidated in CTD patients, in order to establish its relevance in the diagnosis, prognosis and follow-up of systemic rheumatic diseases.

[Prevalence of antinuclear envelope antibodies and their isotypes in sera positive for antinuclear antibodies]. / Prevalencia de anticuerpos anti envoltura nuclear y sus isotipos en sueros positivos para anticuerpos antinucleares.

Antinuclear antibodies detected in HEp-2 cells by indirect immunofluorescence assay display a great variety of images, including the nuclear envelope pattern. This is quite a less frequent finding. Two thousand five hundred and ninety-four sera were processed, and 37.6% of ANA were detected. The prevalence of anti-nuclear envelope antibodies (ANEA) was of 1.2%, with a high association with autoimmune liver diseases (83%) and a low association with systemic lupus erythematosus. In 21 sera of patients with ANEA, no anti-DNAn antibodies were found; but 28.6% of anti-smooth muscle antibodies and 19% of anti-mitochondrial antibodies were detected. The triple rodent tissue section proved to be a less sensitive substrate than HEp-2 for the detection of ANEA. When using conjugates against different isotypes of antibodies for the detection of ANA, 90.5% of IgG, 66.6% of IgA and 9.5% of IgM. Two patients had ANEA-IgA at high titers (> or = 1:160) without ANEA-IgG. In this work, the importance of performing complementary tests for the detection of anti-smooth muscle antibodies, anti-mitochondrial antibodies and anti-DNAn is highlighted in order to apply these tests as guidelines for the clinical diagnosis of patients with ANEA. Besides, this study expresses the need of using total anti-Ig antibodies as conjugate for IIF-HEp-2 instead of anti-lgG; until the role of IgA antibodies in these autoimmune diseases is clarified.

Incidencia de la infección chagásica en embarazadas y en recién nacidos en área no endémica / Incidence of Chagas infection in pregnant women and newborn infants in a non-endemic area

The aim of this report was to determine Chagas infection incidence in pregnant women and congenital infection of their children in a hospital of a non-endemic area. From January 1990 to February 1991 we studied: a) 729 pregnant women with the serologic techniques of indirect hemagglutination and indirect immunofluorescence; b) 38 newborns from the 62 babies of seroreactive mothers with the parasitologic microhematocrit method to diagnose the infection. The serological tests were used as an index of the transplacental passage and for the eventual post-treatment control. We found 8.5 of women with Chagas disease, most of whom were born in an endemic area and did not know that they were infected (Table 1). We detected parasitemia in two newborns which represent 5.3 of congenital infection. Both babies were born in good conditions: at term, with normal weight and asymptomatic (Table 2). They were treated with nifurtimox and they showed a good response; the microhematocrit technique became negative a month later. At the end of the treatment the children were in perfect conditions showing an important decrease in the specific antibodies titer (Fig. 1). One of the cases was studied longer than the sixth month of life, maintaining negative serology. Our results in pregnant women are not different from those previously published in Buenos Aires; this points out the need to keep fighting the vector so as to lessen the existing reservoirs. We found a greater incidence of congenital Chagas disease with the microhematocrit technique than that previously published in Buenos Aires with other methods.(ABSTRACT TRUNCATED AT 250 WORDS)(Au)

Incidencia de la infección chagásica en embarazadas y en recién nacidos en área no endémica / Incidence of Chagas infection in pregnant women and newborn infants in a non-endemic area

The aim of this report was to determine Chagas infection incidence in pregnant women and congenital infection of their children in a hospital of a non-endemic area. From January 1990 to February 1991 we studied: a) 729 pregnant women with the serologic techniques of indirect hemagglutination and indirect immunofluorescence; b) 38 newborns from the 62 babies of seroreactive mothers with the parasitologic microhematocrit method to diagnose the infection. The serological tests were used as an index of the transplacental passage and for the eventual post-treatment control. We found 8.5 of women with Chagas disease, most of whom were born in an endemic area and did not know that they were infected (Table 1). We detected parasitemia in two newborns which represent 5.3 of congenital infection. Both babies were born in good conditions: at term, with normal weight and asymptomatic (Table 2). They were treated with nifurtimox and they showed a good response; the microhematocrit technique became negative a month later. At the end of the treatment the children were in perfect conditions showing an important decrease in the specific antibodies titer (Fig. 1). One of the cases was studied longer than the sixth month of life, maintaining negative serology. Our results in pregnant women are not different from those previously published in Buenos Aires; this points out the need to keep fighting the vector so as to lessen the existing reservoirs. We found a greater incidence of congenital Chagas disease with the microhematocrit technique than that previously published in Buenos Aires with other methods.(ABSTRACT TRUNCATED AT 250 WORDS)