研究新生兒和成人之間IL-12p35甲基化的差異與其免疫反應的不同

Study on the difference in immune response and IL-12p35 methylation between newborns and adults

陳嘉容(Chia-Jung Chen)

指導教授 ： 江伯倫

國立臺灣大學/醫學院/臨床醫學研究所/碩士(2012年)

https://doi.org/10.6342/NTU.2012.01097

摘要

目前已知新生兒免疫反應不成熟，導致新生兒容易被細胞內病原體感染及對疫苗的抗原反應不良。胎兒發展的環境中，TH2細胞激素複雜的免疫互動，使得母體傾向不排斥胎兒。而IL-12p35是成為具有生物活性的IL-12的一個決定因子。對於免疫系統而言，未與外界抗原接觸過的輔助性T細胞產生干擾素-γ（IFN-γ）的能力低下已知與甲基化IFN-γ基因的CpG區域有關。在這個研究中，我們探討IL-12p35基因上游的 CpG區域甲基化狀態的編碼序列，以確定是否可能有類似的現象影響新生兒單核細胞對於IL-12的合成。
以100ng/ml LPS刺激周邊血液單核細胞（PBMC），分析24小時後IL-12p70以及IL-12p40的濃度。發現IL-12p70在成人刺激後的平均濃度較高。並分析3小時與6小時後IL-12p40 及IL-12p35 mRNA的濃度，發現在成人刺激後的平均濃度較高。同時也證明新生兒IL-12p35 mRNA的產量較成人為低。但是，利用重亞硫酸鹽處理分析基因甲基化的結果，發現新生兒在IL-12p35基因上的甲基化程度與成人並無差別。
因此，我們發現PBMC合成具有生物活性的IL-12p70能力在新生兒期間顯著受損，原因來自於IL-12p35 mRNA的生產能力低下。IL-12的生產變化，在整個兒童時期發展的機制目前尚不清楚。但是我們的研究發現，啟動子基因的甲基化，並非造成IL-12p35轉錄能力降低的原因。新生兒時期IL-12表現低下可能由其他表觀遺傳學來調控，但並非經由甲基化機制調控，並且發生在染色質轉錄的過程。隨著進一步努力，並逐漸了解這一缺陷的機制，使得修改染色質的藥物製劑的持續發展，表觀遺傳學有朝一日可能成為治療過敏性疾病患者的方法。

關鍵字

新生兒 ； 介白質12 ； 甲基化

並列摘要

It is well established that the immaturity of immune system of the newborn results in an increased susceptibility to intracellular pathogens and poor responses to vaccine antigens. For protection of the pregnancy, the fetus is exposed to a high activity of TH2 and T regulatory cells. IL-12p35 is a determining factor of the bioactive IL-12. As far as the immune system is concerned, the low capacity of naive adult T cells to produce IFN-γ has been correlated with hypermethylation of CpG sites of the IFN-γ gene. We explore the methylation status of CpG sites upstream of the coding sequence of the IL-12p35 gene to determine whether a similar phenomenon could be involved in the deficient IL-12 synthesis by neonatal DC.
We use 100ng/ml LPS stimulate cultured PBMC for 24 hours. PBMCs from adults can express higher level of IL-12p40 and IL-12p70 compare to those of the newborns. Determination of IL-12p40 and IL-12p35 mRNA expression showed that adults have great ability to produce IL-12p35 mRNA. However, the methylation status of CpG sites in the promoter of IL-12p35 does not present any differences between adults and newborns.
Therefore, we found that PBMC synthesis of bioactive IL-12p70 capacity is significantly impaired in the neonatal period. That might be resulted from reducing IL-12p35 production. The mechanism of IL-12 production changes throughout the childhood is unclear. But our research found that the depressed IL-12p35 production might not be due to methylation of promoter gene. The impairment of IL-12p35 expression during neonatal period might be caused by other epigenetic regulation, but not methylation, but occurs in the chromatin level. With further efforts, and gradually understanding the mechanism, we might be able to make the development of pharmaceutical preparations to epigenetics, and it could be used in treatment of allergic diseases.

並列關鍵字

newborn ； interleukin-12 ； methylation

參考文獻

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