[Modified tests basophil degranulation and leukocyte migration inhibition factor in drug allergy. Study 2009-2014]. / Pruebas modificadas de degranulación de basófilos y factor de inhibición de migración de leucocitos en alergia a fármacos. Estudio de 2009 a 2014.

BACKGROUND: Adverse reactions to drugs are increasing and there are few studies for the diagnosis. OBJECTIVE: To determine the utility of modified basophil degranulation (MBD) test and modified leukocyte migration inhibition factor (MLMIF) test to prove drug hypersensitivity. METHODS: 177 patients of both sexes were studied with the diagnosis of drug hypersensitivity, determining MBD, MLMIF, or both, between 2009 and 2014. They were matched with positive and negative controls and the non-allergic population. Applications are issued according to the type of hypersensitivity, considering type I MBD and type IV MLMIF. RESULTS: 170 patients (96.04%) were positive to at least one drug (RR = 4.71). 561 MBD (73.62%) and 201 MLMIF (26.37%) were performed. Female sex was more frequent (64.41%); the average age was 38.5. MBD was positive in 70.23% and MLMIF in 67.16%. The test sensitivity was increased complementarily and with two dilutions. The correlation of MBD and MLMIF was positive and highly significant. CONCLUSIONS: Women have more drug reactions. Modified MBD test is useful at any age. Since medications can activate one or other hypersensitivity mechanism, it is important to request the tests simultaneously.

Antecedentes: Existe incremento de reacciones adversas a medicamentos y pocos estudios para el diagnóstico. Objetivo: Determinar la utilidad de pruebas modificadas de degranulación de basófilos (DB) y del factor inhibidor de la migración de leucocitos (LIF, leukocyte migration inhibition factor) para comprobar la hipersensibilidad a medicamentos. Métodos: Se estudiaron 177 pacientes, de uno y otro sexo, con diagnóstico de hipersensibilidad a medicamentos, en quienes se determinó pruebas modificadas de DB, LIF, o ambas entre 2009 y 2014. Se parearon con controles positivos, negativos y población no alérgica. Las solicitudes se emitieron de acuerdo con el tipo de hipersensibilidad, considerando tipo I a DB y tipo IV a LIF Resultados: 170 pacientes (96.04%) fueron positivos al menos a un medicamento (RR, 4.71). Se realizaron 561 pruebas modificadas de DB (73.62%) y 201 de LIF (26.37%). El sexo femenino fue más frecuente (64.41%); la edad promedio fue de 38.5 años. La prueba modificada de DB resultó positiva en 70.23% y la de LIF en 67.16%. La sensibilidad de las pruebas se incrementó en forma complementaria y a dos diluciones. La correlación de las pruebas fue altamente significativa. Conclusiones: Las mujeres presentan más reacciones a fármacos. La prueba modificada de DB es útil en cualquier edad. Como los medicamentos pueden activar uno u otro mecanismo de hipersensibilidad es importante solicitar las pruebas simultáneamente.

Modulación del tráfico leucocitario: papel de las quimiocinas y de los opioides / Modulation of leukocyte trafficking: role of chemokines and opioids

El correcto movimiento y posicionamiento celular son elementosclaves en el desarrollo, determinantes tanto en situacionesfisiológicas como patológicas. Probablemente sea en el sistemainmunológico donde más extensamente se han estudiado losprocesos de migración celular, ya que muchos aspectos de la respuestainmune están directamente relacionados con la regulacióndel tráfico leucocitario. La migración de leucocitos es un procesoaltamente coordinado en el que participan muchas moléculas ysus correspondientes receptores. En esta revisión, nos centramosprincipalmente en las quimiocinas, moléculas con capacidad quimioatrayente,y en los opioides, ya que, aunque normalmente suactividad se ha considerado restringida al sistema nervioso, tambiénafectan a las células inmunes y modulan su movilidad. Larespuesta inmune es un proceso muy dinámico que depende dela concentración de una gran variedad de moléculas que aparentementepueden no tener ninguna relación así como de la presenciade los receptores para esas moléculas en la membrana celular

Correct cell movement and positioning are central elementsin development, and influence both normal physiology and diseasestates. Cell movement has probably been studied most extensivelyin the immune system, where many aspects of the immuneresponse are closely related to coordination of leukocyte trafficking.Leukocyte migration is thus a highly regulated processthat implicates many molecules and receptors. In this reviewwe focus our attention on chemokines, classical chemoattractantmolecules and on opioids, molecules that usually act on thenervous system but that also affect immune cells and modulatetheir movement. The final immune response is a very dynamicprocess that depends on the amount of a variety of different apparentlyunrelated molecules and on the presence at the cell membraneof their corresponding receptors

Human B cells secrete migration inhibition factor (MIF) and present a naturally processed MIF peptide on HLA-DRB1*0405 by a FXXL motif.

A better knowledge of peptide structures interacting with major histocompatibility complex (MHC) molecules is of great interest for better understanding of the molecular basis of immune recognition. We have isolated naturally processed peptides from a continuously growing antigen-presenting Epstein-Barr virus-transformed human B-cell line. HLA-DR complexes were purified by specific affinity chromatography and complexed peptides were released by acid treatment. The isolated peptides were separated by reversed phase chromatography and fractions were analysed by Edman degradation at picomolar ranges. From 30 fractions that were examined seven peptides bound to the HLA-DRB1*0405 and two peptides from the human leucocyte antigen (HLA) class II associated invariant chain bound to HLA-DRB1*1302. In addition, a N-terminal beta-chain peptide of the 0405 allele was identified. Evaluation of amino acid sequences revealed a refined FXXL motif for the 0405 allele, in which F (phenylalanine) stands for any aromatic amino acid and L (leucine) can be exchanged by either I (isoleucine) or V (valine). In total, three fractions contained a peptide derived from the human migration inhibition factor (MIF), a pro-inflammatory cytokine that is normally produced by activated T lymphocytes and monocytes/macrophages. Indeed, cytokine analysis revealed high amounts of MIF secreted by the B-cell line, confirming that MHC class II expressing cells can present any intrinsic peptide that contains the distinct motif for HLA-binding. For MIF, the amino acid sequence Y36IAV39 represents the required binding motif for HLA-DRB1*0405. Nevertheless, it is the first time that cytokine fragments were found to bind to HLA molecules on human B cells.

Factor inhibidor de la migración leucocitaria antes del tratamiento con factor de transferencia y después de el / Leukocyte migration inhibitory factor before and after treatment with transfer factor

Se realizaron determinaciones de la actividad del factor inhibidor de la migración leucocitaria "in vitro" en 10 niños con infecciones respiratorias recurrentes antes del tratamiento con factor de transferencia y después de él. Se encontró un incremento significativo (pó0,01) de esta función celular y de la respuesta cutánea a la tuberculina "in vivo", después del tratamiento con este medicamento. Los resultados señalan la importancia del estudio de pacientes con esta entidad y la utilidad de evaluar estas variables en su seguimiento clínico y terapéutico

Factor inhibidor de la migración leucocitaria antes del tratamiento con factor de transferencia y después de el / Leukocyte migration inhibitory factor before and after treatment with transfer factor

Se realizaron determinaciones de la actividad del factor inhibidor de la migración leucocitaria "in vitro" en 10 niños con infecciones respiratorias recurrentes antes del tratamiento con factor de transferencia y después de él. Se encontró un incremento significativo (pó0,01) de esta función celular y de la respuesta cutánea a la tuberculina "in vivo", después del tratamiento con este medicamento. Los resultados señalan la importancia del estudio de pacientes con esta entidad y la utilidad de evaluar estas variables en su seguimiento clínico y terapéutico

Correlation between membrane glycoprotein and detergent dissected membrane protein in the assessment of LMIF: a report of 51 ALA cases.

Assessment of 51 amoebic liver abscess cases for leukocyte migration inhibition factor released using membrane glycoprotein and detergent dissected membrane protein (DDMP) of axenic Entamoeba histolytica (NIH:200). Lymphokines release by T lymphocytes in response to purified amoebic membrane glycoprotein (PAMG) against whole amoebic lysate (WAL), dissect out protein against whole amoebic lysate and membrane glycoprotein against dissected protein was tested by leukocyte migration inhibition test on blood samples from proved amoebic liver abscess cases. A significant increase was noted in the release of lymphokines and 100% positivity was observed with both PAMG and DDMP compared to 78% with whole amoebic lysate. The difference between means leukocyte migration indices of the membrane glycoprotein and whole amoebic lysate, detergent dissected protein and whole amoebic lysate with regards to release LMIF were found to be highly significant (P < 0.001), (P < 0.005) respectively. But insignificant difference and very much similarity was noted between the means of membrane glycoprotein and dissect out protein sensitized T lymphocytes with regards to lymphokine release in vitro. This shows the patients had high degree of leukocyte sensitized to pure amoebic membrane glycoprotein and detergent dissected membrane protein compared to whole amoebic lysate. These findings indicate that detergent dissected protein has similar antigenicity with membrane glycoprotein in elicitation cell mediated immune response in amoebic liver abscess cases.

[Studies on the pathogenic mechanism of beta-lactam hypersensitivity by the detection of leucocyte migration activating factor and leucocyte migration inhibitory factor--structural correlations with allergic symptoms due to beta-lactam antibiotics].

Structural correlations between allergic symptoms and beta-lactam antibiotics were investigated by the use of leucocyte migration inhibition tests on 147 patients in whom allergy to the drugs was detected from among 193 patients suspected of having beta-lactam hypersensitivity. No significant difference was found in the allergic symptoms between the mother nucleus structure of beta-lactam antibiotics or the C-3 side chain structure of cephem antibiotics. But, in both beta-lactam and cephem antibiotics, those with an aminothiazolyl group in the acylamido in their side chain structure induced fever or hepatopathy significantly more often than skin eruption. In contrast, drugs with a benzyl group induced skin eruptions significantly more often than hepatopathy. Leucocyte migration activating factor (LMAF) was found significantly more often than leucocyte inhibitory factor in patients with fever or hepatopathy, and cephem antibiotics with an aminothiazolyl group in the 7-acylamido side chain produced LMAF at a very high rate (80%). Our findings indicate that the specificity of the chemical structure of beta-lactam antibiotics and allergic symptoms is dependent on the acylamido side chain structure, and an aminothiazolyl group structure has a high capability of inducing fever or hepatopathy. Moreover, the high LMAF-inducing ability of the aminothiazolyl group structure is involved in this pathogenic mechanism.

Cellular and humoral responses in coeliac disease. 2. Protein extracts from different cereals.

The humoral and cellular immune responses to grain protein extracts from coeliac-toxic and non-toxic cereals were compared by use of a number of ELISA and immunoblotting methods and the indirect leucocyte migration inhibition factor (LMIF) assay. Both adult and child coeliacs had elevated levels of serum antibody to proteins from the coeliac-toxic cereals, namely bread wheat, durum wheat, rye and barley and low levels of proteins from other cereals. Using protein blotting techniques, antibody binding was greatest to gliadins/low mol mass glutenin subunits and homologous prolamins from rye and barley, consistent with the ELISA findings. Competition ELISA and preabsorption tests indicated that antibody reaction to maize storage proteins did not simply result from cross-reaction of antigliadin antibodies. In LMIF assays, only the wheat extracts had activity in coeliac patients. This is most likely partly due to loss of some of T-cell epitopes from the extraction technique required for these proteins, as well as the relatively small effects seen for even very active fractions in the LMIF assay.

Cellular and humoral responses in coeliac disease. 1. Wheat protein fractions.

The humoral and cellular immune response of coeliac individuals to various wheat protein fractions was studied using serum antibody ELISA assays and the indirect leucocyte migration inhibition factor (LMIF) assays. Greater migration inhibition factor activity was seen in coeliacs on a gluten-free-diet having low serum antibody titres, and using purified T-cells instead of total peripheral blood mononucleocytes. Gliadin was the most active fraction in both assays. Raised antibodies to low-molecular weight and high-molecular weight glutenin polypeptides was observed, though these proteins had little migration inhibition factor activity. No cellular or humoral response was seen to albumins or globulins. Proteins associated with the granules of well-washed wheat starch are distinct from gluten proteins and had little T-cell activity, correlating with clinical observations that properly prepared wheat starch is devoid of coeliac toxicity. The greater specificity of the humoral response for individual wheat protein fractions in this study, compared with the earlier reports, likely results from cross-contamination in the earlier work of each fraction with gliadin.

[Leukocyte migration inhibitory factor and basophil degranulation in drug reactions]. / Factor inhibidor de la migración de leucocitos y degranulación de basófilos en las reacciones a medicamentos.

We performed a prospective study in patients with a medical history of adverse reaction to drugs with the purpose of rule out allergy. We included 31 patients who were attended in the Allergy Service. We compared the sensibility and specificity of the test of inhibition factor of leucocytes migration and degranulation basophil against the exposition. After the statistical analysis, we concluded: the laboratory test, we have already mentioned, have little sensibility and specificity so the exposition test in the quickest, useful, and more simple method to determine drugs allergy, but more dangerous.

[Studies on the pathogenic mechanism of beta-lactam hypersensitivity by the detection of leucocyte migration activating factor and leucocyte migration inhibitory factor].

In 145 suspected cases of beta-lactam hypersensitivity, the identities of the allergenic drugs were performed by leucocyte migration inhibition test (LMIT). The involvement of cell-mediated immunity using leucocyte migration activating factor (LMAF) and leucocyte migration inhibitory factor (LMIF) was investigated for each allergic symptom. The proportion of LMIT positives was 74% of all 145 cases, and more than 70% for each allergic symptom except for anaphylactic shock, and in particular more than 90% for fever and eosinophilia. LMAF and LMIF were detected 39% and 35% of 145 cases, respectively, however LMAF was more frequently detected than LMIF in cases with hepatopathy. LMAF was more frequently detected than LMIF when the latent period (the duration of sensitization) of skin eruption or fever was less than 10 days, whereas LMIF was more frequently detected than LMAF when it was more than 10 days. The same result also was obtained at the boundary between 14 days of the latent period in hepatopathy. These results indicate that in the pathogenesis of beta-lactam hypersensitivity, cell-mediated immunity plays a major role. Both LMAF and LMIF are involved, and their production is dependent on the duration of allergenic drug-sensitization, LMAF is produced during the early period of sensitization, whereas LMIF is produced during the late period of sensitization.

Regulation of lymphocyte motility by macrophages: characterization of a lymphocyte migration inhibitory factor derived from a macrophage-like cell line.

An inhibitory factor on lymphocyte migration was detected using a capillary random migration assay in the culture supernatant of peritoneal exudate macrophages cultured at concentrations greater than 8 x 10(6) cells/ml. After examining different macrophage-like cell lines, J774A.1 cells were found to produce this inhibitory factor, which was termed lymphocyte migration inhibitory factor (LMIF). The inhibitory effect of LMIF on the migration of spleen lymphocytes, thymocytes, and bone marrow cells was determined. The migration of thymocytes was more sensitive to LMIF than was the migration of spleen lymphocytes and bone marrow cells. Interestingly, when the effect of LMIF was tested on the migration of spleen T cells and B cells, T cells were more sensitive than B cells. When the thymocytes were separated by peanut agglutinin into mature and immature thymocytes, the migration of mature thymocytes was more sensitive than that of immature thymocytes, the migration of mature thymocytes was more sensitive than that of immature thymocytes to the effect of LMIF, suggesting that the greatest effect of LMIF was on the migration of mature T cells. Partial purification of LMIF by ion-exchange and gel-filtration chromatography revealed that it is approximately 14,000 in molecular weight and could exist in either monomeric or dimeric forms. The possible role of this factor in an immune response is discussed.

Induction of cell mediated immune response by nonspecific stimulator of immunity against antigenically unrelated oncogenic virus (Marek's disease virus).

An enzyme treated preparation of Mycobacterium phlei (NSI), induced strong cell mediated immune response (CMIR) against specific as well as against nonspecific oncogenic Marek's disease (MDV) in birds, as evinced by Lymphocyte migration inhibition, (LMIT) lymphocyte transformation test (LT) and Lymphokine (Lymphocyte migration inhibition factor) LyIF assay. Maximum CMIR could be observed towards third week post inoculation. All the three tests exhibited a positive correlation. Such phenomenon of CMIR induction by NSI, nonspecifically to unrelated viral/cancerous diseases (MD) in birds generates hopes for immunoprevention of these maladies by utilizing such phenomenon.

[Recurrent herpes labialis infections: cellular immunity and immunomodulation]. / Rezidivierende Herpes-labialis-Infektionen: Zellvermittelte Immunität und Immunomodulation.

The therapy with thymus extracts, oral and parenteral, reduced the frequency and the intensity of recurrences of herpes labialis infections. The duration of the effect was at least 6 month after interruption of the therapy. Indomethacin was effective and developed and intensive effect only during the therapy. The skin test with recall-antigens and the neopterin-elimination were altered at the first day of the menstruation during the recurrence. The normalisation succeeded during therapy. In patients with recurrences in the perimenstrual time we observed a reduced T-helper/T-suppressor index during the first day of menstruation. Normal data were registered out of the recurrence time and/or under therapy. Inhibitors of the lymphokine: leucocyte/migration inhibitory factor (LIF) with a molecular weight of 6-12 KD were obtained with the specific stimulation of mononuclear cells of patients with recurrent infections with herpes labialis using the herpes-virus-1 antigen. The inhibition of fibrinolysis/proteolysis with aprotinin, tranexamic acid, phenyl-methyl-sulphonyl-fluoride and di-isopropyl-fluorophosphate could prevent the appearance of inhibitors. Inhibitors could be produced by splitting the LIF-molecule with urokinase and plasminogen but not with trypsin. The production, but not the activity, of present LIF-inhibitors are blocked in vivo and in vitro by indomethacin and thymus peptides.

[Characteristics of T and B components of the immune system in miscarriage of different etiology]. / Kharakteristika T- i B-zvena immunnoi sistemy u beremennykh s nevynashivaniem razlichnogo geneza.

A total of 70 females with threatened abortions induced by hormonal, infectious or unclear factors and 63 second trimester underwent clinical, hormonal and immunological investigations. In all females with threatened abortion, the study revealed the variance in immunological data in accordance with the term of pregnancy and cause of its incompetence. The lesser degree of immunological impairment was documented in the females whose pregnancy was complicated by hormonal disorders, whereas females with a history of infections in the early pregnancy demonstrated more profound immunological shifts. In those patients whose pregnancy was lost in the first trimester and the cause of the wastage remained unclear, the authors noted elevated T and decreased B lymphocytes, as well as dramatically reduced leukocyte migration inhibition factor.

Immunologic studies on myasthenia gravis. II: A new chemotactic factor for lymphocytes found in patients with myasthenia gravis.

A specific chemotactic factor for lymphocytes was found in thymic tissue extract from myasthenic patients who had undergone thymectomy. Biologic activities and the biochemical nature of this factor were examined. The major findings were as follows: (1) The factor from myasthenic thymus was specifically chemotactic for T lymphocytes. Marker analysis of target cells revealed that only OKT4 positive cells (helper/inducer T cells) responded to the factor. Chemotactic activity was also found in the normal thymus, but its activity was different from that found in the myasthenic thymus. Interestingly, OKT8 positive cells (suppressor/cytotoxic T cells), as well as OKT4 positive cells, migrated toward the normal thymic extract (2) Chromatography of the extract by gel filtration gave three peaks with chemotactic activity; molecular size were 160,000 to 100,000, 25,000 to 15,000, and smaller than 1350 daltons (3) The factor was inactivated by heating for 30 minutes at 56 degree C and was also destroyed under conditions of pH 4 and pH 10. The factor was also sensitive to treatment by trypsin, sodium metaperiodata, 8 mol/L urea, reduction, or alkylation. (4) The chemotactic activity in the myasthenic thymus was distinct from other known chemotactic factors, including C5a, interleukin-1, and interleukin-2. (5) Chemotactic activity for lymphocytes was found in both the aqueous extract and the culture supernatant of thymic stromal cells, but not in thymocyte extract or in the serum. These findings indicate that the chemotactic factor specific for OKT4 positive cells may be secreted by stromal cells of the myasthenic thymus but not of the normal thymus.

Enteroviral-related antigen in circulating immune complexes of amyotrophic lateral sclerosis patients.

Circulating immune complexes were isolated by polyethylene glycol precipitation from the sera of patients with amyotrophic lateral sclerosis. Rabbits immunized with circulating immune complexes from 3 of 5 amyotrophic lateral sclerosis patients induced antisera that specifically reacted with enterovirus-infected cells by immunofluorescence and enzyme-linked immunosorbent assay. These antisera were nonneutralizing and did not react with purified virus. In addition, peripheral lymphocytes of amyotrophic lateral sclerosis patients produced lymphokine in response to extracts from enterovirus (Coxsackie B4) infected cells. These results suggest both a humoral (circulating immune complex) and a cellular immune response in some patients with amyotrophic lateral sclerosis to enterovirus-coded or -induced antigen.

Serial study of liver-directed autoantibodies and autoreactive T-lymphocytes in acute viral hepatitis B.

To explore the mechanisms underlying liver-directed autoimmune reactions in acute Hepatitis B Virus (HBV) infection, we followed five subjects who were identified in the early incubation phase (30-70 days before the first elevation of transaminases). We assessed serially cellular (using a T-lymphocyte migration inhibitory factor assay) and humoral (RIA) immunity to LSP (a macromolecular, liver-derived lipoprotein complex) and hepatic lectin (HL), the liver-specific receptor for desialylated glycoproteins, which appears to be a major target antigen for autoreactions in autoimmune chronic active hepatitis. Anti-LSP and anti-HL autoantibodies were found, at some stage during acute HBV infection, in 4/5 subjects, whereas cellular immunity to the same antigens was detected in only two patients. Sustained production of anti-HL antibodies was noted only in patients showing cellular immunity to this antigen and was apparently secondary to liver damage, whereas anti-LSP antibodies were first detected at the onset of liver injury when there was no evidence of T-cell immunity to the same antigenic complex. One explanation for this apparent dichotomy between cellular and humoral responses to LSP is that a helper T-cell response to the major envelope component of HBV, HBsAg, which precedes by 10-20 days the development of anti-LSP antibodies, promotes a humoral reaction to autoantigens contained in the LSP preparation, coexpressed with HBsAg, on the surface of infected hepatocytes.