Tarcimara Moreira da Silva roberto Eustáquio Santos guimarães Evaldo nascimento Helena maria Gonçalves Becker Ricardo nascer Araújo flávio Barbosa Nunes About ns authors

Rinite alérgica denominações uma doença que decorre de um processo inflamatório da mucosa nasal conseqüente à reação de hipersensibilidade der alérgenos inalatórios e, eventualmente, alimentares. É mediada pela IgE, envolvendo diferente células, mediadores e citocinas. OBJETIVO: avaliação as transcrições ao as segue citocinas: IL-4, IL-5, IL-8 e IFN-gama, especialmente importantes enquanto processo alérgico nasal, predominante IL-4 e IL-5. Nisso estudo, optou-se por avaliar os paciente atópicos fora das perigo alérgicas, alcançar a meta de se sabendo as expressões a partir de citocinas neste período. Coisas E MÉTODO: Realizou-se um estudo transversal e prospectivo, selecionando-se 30 pacientes, existência 13 paciente portadores de rinite alérgica paucissintomáticos e 17 paciente não-atópicos. Os grupos foram selecionados por da história, a partir de exame clínicos otorrinolaringológico e a partir de teste alérgico cutâneo - Prick Test. Ministérios perfil das citocinas obtivermos pesquisado nos fragmentárias de mucosa nasal, pela da RT-PCR semiquantitativa, escolhida por apresentar boa reprodutibilidade e especificidade, utilizando-se gostar de referência o genes da Beta-actina. RESULTADOS: Os valores de IL-5, IL-8, IFN-gama mantiveram-se homogêneos em conectar ao agrupadas controle. Der IL-4 proposta diferença alcançar significância estatística. CONCLUSÃO: Os pacientes alérgicos paucissintomáticos apresentaram normalização da expressão do citocinas na mucosa sleep à exceção de IL-4.

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alérgenos; citocinas; hipersensibilidade; rinite; rinite; testes


Allergic rhinitis is an inflammation reaction of the nasal mucosa, in consequence of one IgE mediated hypersensitive reaction to inhaling allergens, involving various mediators and cytokine cells. AIM: a purpose of this estude was come evaluate the transcriptions para IL-4, IL-5, IL-8 and IFN-gama, an especially important in the nasal allergy process, particularly IL-4 and IL-5. For this study we determined to evaluate atopic patients quem were free em ~ allergic crises, with ns purpose that knowing ns cytokine expressions during this period. MATERIALS e METHODS: one more prospective e transversal study was brought out, picking 30 patients, 13 of this patients were pauci-symptomatic and 17 were non atopic. The groups to be selected by method of der medical interview, an otolaryngologic clinical exam e allergy skin test - Prick Test. The cytokines to be investigated in fragments of ns nasal mucosa, making use of RT-PCR - chosen because it has nós vamos reproducibility e specificity. RESULTS: IL-5, IL-8, IFN-gama cytokine worths were preserved homogeneous in relationship to the direção group. Only IL-4 presented far-ranging statistic differences. CONCLUSION: Asymptomatic patients com allergic rhinitis presented com normalization the cytokine expression in ns nasal mucosa, com exception that IL-4.

allergen; cytokines; hypersensitivity; rhinitis; rhinitis; skin


ORIGINAL ARTICLE

RT-PCR cytokine study in patients with allergic rhinitis

Tarcimara Moreira da SilvaI; roberto Eustáquio santos GuimarãesII; Evaldo NascimentoIII; Helena maria Gonçalves BeckerIV; Ricardo nascer AraújoV; frágio Barbosa NunesIV

IMSc in Medicine, Otorhinolaryngologist, professor - FEAD centrado de gestão Empreendedora

IIPhD in Medicine, Associate professor of medicine - USP de Ribeirão Preto and Adjunct professor of Otorhinolaryngology - room of Otolaryngology, Speech and Hearing Therapy e Ophthalmology of a Medical institution of the comunidade University of Minas visão global - FM-UFMG

IIIPhD in Medicine, Adjunct professor - room of parasitology - ICB - UFMG

IVPhD in Medicine, Adjunct professor - room of Otolaryngology, Speech e Hearing Therapy and Ophthalmology of ns Medical institution of the commonwealth University that Minas em geral - FM-UFMG

VVeterinarian - Parasitology

VIPhD college student - Graduate program in general Surgery - medical School - UFMG, Otorhinolaryngology

SUMMARY

Allergic rhinitis is an inflammatory reaction of the nasal mucosa, in consequence of one IgE mediated hypersensitive reaction to inhaling allergens, involving different mediators and cytokine cells.

AIM: a purpose of this study was come evaluate ns transcriptions ao IL-4, IL-5, IL-8 and IFN-gama, specifically important in ns nasal allergy process, specifically IL-4 and IL-5. Para this estude we chose to evaluate atopic patients who were free são de allergic crises, with a purpose that knowing ns cytokine expressions during this period.

MATERIALS and METHODS: one more prospective and transversal aprender was lugged out, choosing 30 patients, 13 of this patients to be pauci-symptomatic and 17 to be non atopic. Ns groups to be selected by means of a medical interview, an otolaryngologic clinical exam e allergy skin tests - Prick Test. Ns cytokines were investigated in fragments of ns nasal mucosa, using RT-PCR - chosen since it has boa reproducibility e specificity.

RESULTS: IL-5, IL-8, IFN-gama cytokine worths were retained homogeneous in relationship to the ao controle group. Apenas um IL-4 presented significant statistic differences.

CONCLUSION: Asymptomatic patients com allergic rhinitis presented com normalization of cytokine expression in a nasal mucosa, with exception of IL-4.

Keywords: allergen, cytokines/immunology, hypersensitivity, rhinitis/allergic/perennial, rhinitis/allergic/seasonal, skin/tests.

INTRODUCTION

IL-4, IL-5, IL-8 e IFN-γ cytokines, especially IL-4 and IL-5, mediate e regulate immune e inflammatory reaction in allergy rhinitis. Considering this information and the heat of research developed in a Otorhinolaryngological school where a present investigation ser estar carried out, we determined to investigate this substances between dois groups: one with paucisymptomatic allergic rhinitis and a non-allergic control group.

Reverse transcriptase-polymerase chain reaction was the method the laboratorial evaluation chosen because it has good reproducibility and specificity1,2.

The meta of a present investigation was to analyze, quantify and compare the expressions of the afore-mentioned cytokines in paucisymptomatic patients com allergic rhinitis, comparing them with a non-atopic ao controle group in order to pesquisar the relevance this substances have in those individuals fora of ns allergic spell period.

MATERIALS e METHODS

This is der prospective, cross-sectional study, carried out in a sample of patients with e without allergic rhinitis. A patients favored were those com previous operation indication ao septoplasty, adenoidectomy and/or tonsillectomy associated with partial turbinectomy. Ns fragments to be collected a partir de February that 2004 to April the 2005.

This researchprojeto was approved by ns Ethics in Medical research Committee (COEP) on October 14, 2004, under protocol # ETIC 044/04. Tudo the patients and their companions to be duly described about the study, that goals and methods. Those quem agreed signed the informed consent form.

These patients responded come an interview, castle underwent clinical otorhinolaryngological exam e skin allergy test, they to be then broken down into two groups: one grupo of patients com allergic rhinitis, consisted of of 13 patients e another grupo made up of 17 non-atopic patients. The factors impacting sampling were: the alto cost that the essa utilized, ns lack of trained specialists to carry lado de fora the test (RT-PCR) and the long time it took to import a primers e the equipe set come conclude ns study.

Age ranged between three e 47 years, e they were children and adults, nevertheless of race, gender, faith or social class. Among these patients, 16 underwent septoplasty, 12 underwent adenoidectomy and two suffered adenotonsillectomy. Tudo de these patients experienced turbinectomy, thus consisting of a rude of 30 partial lower turbinates removed, which were climate sent porque o cytokine analysis.

We took off ns study tudo those patients under anti-histamine e steroid agentes for at the very least thirty dia before the turbinectomy to collect a nasal mucosa fragment, or that might not terminate ns use the medication the impacted a skin check response, patients com severe skin eczema or dermatitis and patients below three years of age or yonsi (above 60 years of age) because of der higher event of false negative due to der reduction in test reactivity in old age.

The patients to be submitted to a prick test on ns medial face of their forearms in adults and on a back in tiny children.

As antigen we have ns following: D. Farinae, D. Pteronyssinus, house dust, Blatella sp, grass, fungi, feathers, cats e dogs. Ao positive ao controle we supplied histamine e for the negative 1 we supplied saline solution3. This essa was purchased from the international Pharmaceutica Immunology - Brazil Ltda Laboratories. (IPI) são Paulo.

Nasal mucosa pieces were inserted in 1.5ml "Eppendorf" tubes, sterile, determined with a patient"s name e collection data. Afterwards, these tubes were inserted in der Styrofoam box and taken to ns Hospital"s Immunology Laboratory. There, the fragments to be frozen in ~ - 80ºC.

The cDNA era synthesized em ~ the 1.25 medicamento of rude RNA making use of random hexamer primers (Promega) and the reverse transcriptase sistema - SuperScript II (Invitrogen) de acordo com to manufacture"s instructions. PCR ser estar carried out in der final intervalo of 20ml, in a presence the 1ml that cDNA, 0.25mM the dNTP, 0.2 mM of every primer e 1 U of Platinum TaqDNA polymerase (Invitrogen).

PCR commodities were analyzed by electrophoresis in an 8% polyacrylamide gelatin dyed by silver. A gels to be photographed e the results were analyzed by densitometry utilizing AlphaDigiDoc 1201TM (AlphaInotech).

Since it foi ~ not feasible to clone ns genes e obtain ns absolute quantity necessary porque o RT-PCR in problema time, us used a semi-quantitative RT-PCR and the β-Actin ("housekeeping") foi ~ considered ns reference gene. The standard coefficient era then acquired a partir de dividing a β-Actin value são de each patient by ns average of a 30 patients. This dosage era called relative because we measured the ao controle expression in relation to the standard test4-7.

In the statistical analysis, among ns measures the interest, we provided parametric and non-parametric tests, com statistical significance level resolved at 5% and a trust interval at der minimum the 80% e a best of 95%.

Continuous variables such as age, IL-4, IL-5, IL-8, IFN-γ and β-Actin to be analyzed as such, and central trend measures were calculation (mean e median) as well as scatter steps (standard deviation e amplitude)8-9.

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Variables gender, agir and IL-4, IL-5, IL-8, IFN-γ cytokines were compared porque o both groups of patients: allergic e non-allergic and, through a statistical test employed (Bartlett"s chi-squared, ANOVA, t-test, Kruskal Wallis"s test and Pearson"s correlation), there was enquanto statistical difference, except for IL-4, i m sorry had naquela pTable one shows the mean, median, standard deviation e range.