微核醣核酸miRNA在異位性皮膚炎致病角色之研究

Abstract

兒童過敏性疾病主要包括: 氣喘，過敏性鼻炎，異位性皮膚炎。是兒童時期及青少年時期最常見的慢性疾病。異位性皮膚炎是一種與遺傳和環境有關的慢性發炎且易復發的皮膚病變，主要機制可能與免疫系統( both the innate and the adaptive immune system) 複雜的基因調控失常有關，使發炎反應無法停止。而近年來， microRNAs的研究，發現其能夠抑制mRNA蛋白質轉譯而調控基因的表現，預計人類三分之一以上的基因受到miRNAs調控。至目前為止國外文獻報告，共發現有五個miRNAs(miR-146a, miR-148a，miR148b， miR-152，miR-155)可能和調控Th1/Th2細胞免疫路徑、過敏性疾病有關。異位性皮膚炎患者，基因遺傳是重要的原因。但是臨床上，卻常是家族中唯一一位有異位性皮膚炎的人，而且其血液中，常有許多種特異性抗體IgE(specific IgE)，相對高的IgE總值(total IgE值)。這引起我們極大的興趣，在相同的環境及遺傳下，什麼影響到基因的表現? miRNAs在異位性皮膚炎的調控機制中是否扮演一定的角色?本研究為國內首次研究，microRNAs在台灣人異位性皮膚炎中的調控角色。研究目的是要探討罹患異位性皮膚炎的台灣兒童，血液白血球細胞中，是否有特定的miRNAs表現異常。希望藉由國外目前已知和過敏性疾病調控有關的miRNAs，比較台灣人異位性皮膚炎病人和未罹病的人之血液白血球細胞中，特定miRNAs表現的差異。得知台灣罹患異位性皮膚炎兒童，臨床嚴重程度和特定miRNAs表現，是否有關聯性。得知罹患其他過敏性疾病（如氣喘、過敏性鼻炎等）是否會影響到此關聯性。及特定miRNAs和IgE總值(total IgE值)、特異性抗體IgE (specific IgE) 及細胞激素(cytokines)等值的關聯性。我們相信，此研究的具體成果，將讓我們對於異位性皮膚炎的致病機制有更深的了解，對於疾病的診斷、預後、及治療有更大的幫助。 結果 90位台灣兒童被納入計畫，其中有3人未能採集到血液檢體。共完成實驗研究共87人，對照組無過敏性疾病的健康兒童24人，實驗組中罹患異位性皮膚炎兒童33人及只罹患呼吸道過敏性疾病（氣喘或過敏性鼻炎）30人。罹患異位性皮膚炎兒童和只有罹患呼吸道過敏性疾病兒童，週邊血液白血球細胞中，miR-146a的表現量比健康兒童皆有顯著下降(P:0.003, P:0.001)。miR-146a的表現量，和IgE總值有意義的負相關性(r:-0.220，P:0.044，two-tailed)。罹患異位性皮膚炎兒童miR-155的表現量，比健康兒童有顯著下降 (P:0.043)。miR-155a的表現量，和IgE總值有意義的負相關性(r:-0.255，P:0.020，two-tailed)。吸入性過敏原屋塵螨D.p.(D. pteronyssinus) 或粉塵螨D.f.(D. fannae) 過敏程度越嚴重的兒童，其miR-155的表現量越低。只罹患呼吸道過敏性疾病兒童，miR-152的表現量，比罹患異位性皮膚炎兒童，有顯著下降 (P:0.021) ；但在健康兒童和罹患異位性皮膚炎兒童之間，並無顯著差異。miR-148a和 miR-148b的表現量，在健康兒童，罹患異位性皮膚炎兒童和只罹患呼吸道過敏性疾病兒童三組之間，並沒有顯著差異。血清中細胞激素的相關性。在健康兒童，罹患異位性皮膚炎兒童和只罹患呼吸道過敏性疾病兒童三組血清中，細胞激素INF-r 值全部都小於7.8 pg/ml。一半左右的過敏性疾病兒童(57.1%)，血清可檢測出細胞激素IL-4 (0.167 pg/ml~5.515pg/ml)，而健康兒童全部檢測不出(<0.125 pg/ml)。 結論 直至今日，miRNA在免疫系統的研究，大多建立在老鼠模式的研究。對於人類的研究，也多為體外細胞培養(human cell culture)。我們的研究是首次直接研究檢測人類週邊血液白血球細胞中，目前已知可能和調控Th1/Th2 路徑、過敏性疾病有關的miRNAs。結果發現台灣罹患異位性皮膚炎的兒童，週邊血液白血球細胞中，miR-146a和 miR-155的表現量，和異位性皮膚炎有意義的負相關性。罹患異位性皮膚炎兒童，miR-146a和 miR-155的表現量，比健康兒童皆有顯著下降。進一步我們認為，以 miRNA 為基礎的基因治療如miR-146a和 miR-155的RNAi的補充，可能是治療異位性皮膚炎極具潛力的方式，但還需進一步確定。

Allergic diseases including asthma, atopic dermatitis and allergic rhinitis are the most common chronic diseases during the childhood period. Atopic dermatitis is a chronically relapsing inflammatory condition of the skin, characterized by intense pruritus. The mechanism might be that the gene dysregulation of both the innate and the adaptive immune systems cause persisting inflammation. MicroRNAs (miRNAs) first described in 1993 downregulate gene expression and effect cell function by binding to messenger RNA. The regulation by miRNAs might be quite common in human diseases. More than 1/3 of human genes are regulated by miRNAs. So far, some studies revealed that five miRNAs (miR-146a, miR-148a, miR148b, miR-152, and miR-155) are possibly associated with allergic diseases or the regulation of Th1/Th2 pathway .This purpose of our study is to evaluate whether there is any special mMiRNA expression in the peripheral blood of children with atopic dermatitis in Taiwan. RESULT: 87 Taiwanese children are evaluated including 24 healthy children, 33 patients with atopic dermatitis and 30 only airway allergic children (Asthma and/or allergic rhinitis). Children with atopic dermatitis or only airway allergic disease have significantly lower miR-146a expression compared with healthy children (P = 0.003, P = 0.001). The expression level of miR-146a has a significantly negative correlation with total IgE (r = -0.220, P = 0.044, two-tailed). Children with atopic dermatitis has significantly lower miR-155 expression compared with healthy children (P = 0.043). The expression level of miR-155 has a significantly negative correlation with total IgE (r = -0.255, P = 0.020, two-tailed). The expression level of miR-148a, miR-148b or miR-152 has no significant correlation with atopic dermatitis. CONCLUSION: The discovery of miRNAs as regulators of the immune system was almost in mouse model or human cell culture. Our study is first report about that miRNAs may be involved in atopic dermatitis by studying leukocytes in human blood. Taiwanese children with atopic dermatitis have significantly lower miR-146a and miR-155 expression compared with healthy children. Although the mechanism of this interaction is still unknown, the result of this study suggests that miR-146a and miR-155 maybe negatively regulate gene expression to suppress atopic dermatitis. The miRNA based gene therapy such as miR-146a or miR-155 RNAi could be a potential therapy for allergic diseases especially in atopic dermatitis. Before these two miRNAs can be used as a therapeutic tool, these data should be verified in a larger study.