介白質-33(Interleukin-33, IL-33)也稱作IL-1F11，屬於IL-1超家族中的一員，可作為細胞激素，當組織受到損害時細胞核中的pro-IL-33可經過蛋白酶修飾後送到細胞外與其受體ST-2、IL-1rap的結合，誘導T細胞之極化反應，進而使T細胞轉變為第二型輔助T細胞(Th2)來調控發炎反應的產生。Pro-IL-33具有核定位序列可以進入細胞核中，但IL-33在細胞核中的功能尚未釐清。在本研究中也發現IL-33主要大量表現在大鼠C6腦膠質瘤細胞與大鼠膠質前驅細胞(GPCs)之細胞核中。利用DNA染色質纖維染色發現到IL-33與DNA結合。另外透過核蛋白免疫共沉澱的方式證明IL-33在膠質瘤細胞中不與NF-B相互作用。以上結果表明IL-33可能主要儲存在腦膠質瘤細胞以及膠質前驅細胞的細胞核。此外，在本文研究中發現小鼠大腦白質區的寡突膠細胞也會大量表現IL-33。透過TEM分析，發現與WT小鼠相比，IL-33基因剃除小鼠(IL33-KO)在大腦胼胝體區的髓鞘結構鬆散且較薄。有趣的是，在動物行為測試(高架十字迷宮與開放場域試驗)的結果可以看到IL33-KO小鼠較不畏懼且顯示較低的焦慮程度。利用西方墨點法證實與WT小鼠相比，OLGs成熟所需的蛋白質MBP、PLP在IL33-KO的小鼠表現量都較低。另外，在體外培養之大鼠OPCs的IL-33抑制後也觀察到與寡突膠細胞分化相關基因(Olig2、 Myrf、 Id4、 Hes-5、Sirt2、 Tspan2)以及與成熟相關基因(MBP、PLP)表現下調。更進一步，分別利用GC、MBP、CNPase抗體以免疫螢光染色的方式觀察細胞型態並發現IL33-KD組別的分化受到明顯的抑制，綜合以上結果，證實IL-33在寡突膠細胞發育及分化的過程是不可或缺的。另外，IL-33基因的缺失使寡突膠細胞成熟受阻以及髓鞘形成的缺陷或許是導致不正常的動物行為結果的原因。

Interleukin-33 (IL-33) is a member of IL1 family and acts as an extracellular cytokine after conversion from pro-IL-33. IL-33 is an “alarmin” to induce T helper 2 (Th2) immune responses in immune cells via interaction with its receptor ST2 and IL-1 receptor accessory protein. The pro-IL-33 protein contains a nuclear localization sequence (NLS), an N-terminal homeodomain-like helix-turn-helix (HTH) motif, and a chromatin-binding domain. Although the pro-IL33 can translocate from the cytoplasm into the nucleus, the function of the nuclear IL-33 in the cells remains unclear. In this study, we showed that IL-33 was enriched in the nuclear fractions of rat C6 glioma cells and rat glial progenitor cells (GPCs). Through the chromatin fiber analysis, IL-33 binding to chromatin was observed. The results from the experiments by co-immunoprecipitation with anti-IL-33 and anti- NF-B indicated that nuclear IL-33 did not interact with NF-B in glioma cells. These observations reveal that IL-33 might be stored mainly in the nucleus of glioma and progenitor cells. Furthermore, the in vivo study indicated that IL-33 was highly expressed in oligodendrocytes (OLGs) at the corpus callosum (CC) of the CNS. By Transmission Electron Microscopy (TEM) analysis we found that the myelin structure in the CC region of mice with IL-33 deficiency (IL33-KO) was disrupted compared to those observed in WT mice. Interestingly, the results from animal behaviors tests (i.e. elevated plus maze test and open field test) showed that IL33-KO mice displayed a fearless behavior, suggesting that the degree of anxiety was reduced in mice after IL33-KO. Oligodendrocytes (OLGs) derived from oligodendrocyte precursor cells (OPCs) prepared from the brains of IL33-KO mice expressed lower levels of OLG myelin basic protein (MBP) and myelin proteolipid protein (PLP) that OLGs generated from WT mice did. The further in vitro study using lentivirus-mediated shRNA delivery against IL-33 expression (IL33-KD) also indicated that OLG differentiation-related genes (i.e. Olig2, Myrf, Id4, Hes5, Sirt2, Tspan2) and OLG maturation proteins (i.e. MBP and PLP) were downregulated after IL33-KD. Moreover, the morphological examination of OLGs by immunofluorescence using anti-GC, anti-CNPase and anti-MBP antibodies revealed that IL33-KD suppressed OLG differentiation and maturation. Overall, our findings demonstrate that IL-33 is required for the proper development of OLG differentiation. IL-33 gene deficiency can disrupt OLG maturation and interfere myelin compaction, which might lead to the behavior change in mice.

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