STAT2與第一型干擾素在樹突細胞發育過程中所扮演角色之研究

Effects of STAT2 and Type I Interferon Signals on Dendritic Cell Development

陳怡伶(Yi-Ling Chen)

指導教授 ： 李建國

國立臺灣大學/醫學院/免疫學研究所/碩士(2011年)

https://doi.org/10.6342/NTU.2011.01017

摘要

樹突細胞(Dendritic cell, DC)是特化的免疫細胞，分布於各種淋巴器官內，以及與外界接觸的體表內側。樹突細胞可細分成conventional DC (cDC) 和plasmacytoid DC (pDC)兩種不同的亞群(subsets)，並需要持續由其前驅細胞(progenitors)分化補充；在造血系統中，巨噬細胞與樹突細胞同源前驅細胞(macrophage DC progenitors; MDP)、樹突細胞前驅細胞(common dendritic cell progenitor; CDP)以及類淋巴前驅細胞(common lymphoid progenitor; CLP)均具有分化成樹突細胞的能力，但至今，對於調節前驅細胞發育成樹突細胞的過程仍尚未全盤了解，而近年來第一型干擾素(Type I IFNs)經證實能促進造血幹細胞(hematopoietic stem cells; HSC)的分裂而產生下游血液細胞，故在本論文中，我們將探討第一型干擾素影響樹突細胞發育的相關機制。
我們使用第一型干擾素(Type I IFNs)訊息傳遞具缺陷的STAT2 (Signal transducer and activator of transcription 2) 基因突變小鼠(STAT2m/m)進行實驗，我們發現此STAT2m/m小鼠體內淋巴器官中的樹突細胞前驅細胞以及樹突細胞數目均減少。利用體外培養的方式(In Vitro)，我們發現STAT2m/m的樹突前驅細胞CDP和CLP生成樹突細胞的潛能(potential)和頻率(frequency)均有缺陷；由CDP生成的cDC較少，而CLP無法有效生成pDC。並利用過繼轉移(Adoptive transfer)實驗則證實STAT2缺乏所造成的前驅細胞缺陷是細胞自發性的(cell autonomous)。另外，其發育過程中STAT2m/m 細胞增殖(proliferation)的速度較慢，以及細胞凋亡(apoptosis)的比例也較高。同時我們也發現在STAT2m/m前驅細胞上的Fms-like tyrosine kinase-3 (Flt3)表現量較低，若使用高量的Flt3L處理STAT2m/m CLP則可挽救pDC族群的產生，顯示降低的Flt3訊息強度是造成pDC減少的原因。使用正常基因型小鼠進行的實驗結果也顯示，Flt3L可導致CLP內第一型干擾素的基因表現。同時若在體外培養或在小鼠體內使用第一型干擾素處理後的CLP會表現較高量的Flt3於其表面，並可促進樹突細胞的生成。若抑制第一型干擾素的訊息傳遞則會降低前驅細胞上Flt3的表現，並減慢細胞增殖及增加細胞凋亡，也使CLP生成pDC的能力下降。整體而言，實驗結果證實，第一型干擾素和STAT2對樹突細胞的發育扮演正面調節者(positive regulator)的角色。

關鍵字

樹突細胞 ； 第一型干擾素 ； 樹突細胞前驅細胞

並列摘要

Dendritic cells (DCs) are specialized immune cells distributed throughout the lymphoid organs and beneath the environmental contact sites. DCs are classified into conventional dendritic cell (cDC) and plasmacytoid dendritic cell (pDC) and need to be continuously replenished by hematopoietic progenitors. To date, there are, at least, three progenitors, including CDP, MDP, and CLP, with a potential to differentiate into cDC and pDC. However, the mechanisms involved in the regulation of their development and homeostasis are still not fully understood. Type I IFNs are known to promote the proliferation of hematopoietic stem cells, the origin of all blood lineages. Therefore, we would like to investigate the effects of type I IFNs on DC progenitors and DCs. Using a STAT2m/m mouse that displayed hyporesponsiveness to type I IFNs, we found that the numbers of all three DC progenitors and DC subsets were decreased. In vitro Flt3L-dependent DC development were significantly impaired in STAT2m/m mice with defective DC-forming potentials and frequencies. CDPs from STAT2m/m mice displayed a reduced potential to develop into DCs and STAT2m/m CLPs preferentially favoured the development of cDC over pDC, which was opposite to the fate of WT CLPs. Adoptive transfer of CLPs into CD45.1 congenic mice suggested that the defects in STAT2m/m CLPs were cell-intrinsic. We also observed the reduced proliferative rate and increase apoptosis rate of STAT2m/m progenitors. Moreover, Flt3 expression level was lower on all three DC progenitors of STAT2m/m mice. High dosage of Flt3L could reverse the fate determination of cDC and pDC from STAT2m/m CLP, suggesting that reduced signalling strength in the progenitors might account for the DC developmental defects. Interestingly, Flt3 signalling induced the expression of type I IFNs in CLPs. In vitro or in vivo administration of IFNα promoted the induction of Flt3 expression on progenitors and enhanced generation of both DC subsets. Blocking type I IFNs signalling attenuated Flt3L-dependent expression of Flt3, increased apoptosis, decreased proliferation and altered the DC subsets differentiation by favouring cDC generation. Taken together, these results indicated a positive role of STAT2 and type I IFNs in promoting DC precursor development and DC subsets differentiation.

並列關鍵字

Dendritic cells ； Type I IFNs ； STAT2 ； Hematopoietic progenitors ； Flt3 signalling

參考文獻

1. Vremec D, Pooley J, Hochrein H, Wu L, Shortman K. CD4 and CD8 expression by dendritic cell subtypes in mouse thymus and spleen. Journal of Immunology 2000.

3. Thibaut De Smedt, Bernard Pajak, Eric Muraille, Laurence Lespagnard, Ernst Heinen, Patrick De Baetselier, et al. Regulation of dendritic cell numbers and maturation by lipopolysaccharide in vivo. J Exp Med 1996.

4. Caetano Reis e Sousa, Sara Hieny, Tanya Scharton-Kersten, Dragana Jankovic, Hugues Charest, Ronald N. Germain, et al. In vivo microbial stimulation induces Rapid CD40 Ligand-independent Production of Interleukin 12 by Dendritic Cells and their Redistribution to T Cell J Exp Med 1997.

5. Alexander D. Edwards, Damien Chaussabel, Tomlinson, Oliver Schulz, Alan Sher, Sousa CRe. Relationships among murine CD11c(high) dendritic cell subsets as revealed by baseline gene expression patterns. Journal of Immunology 2003.

6. Hubertus Hochrein, Ken Shortman, David Vremec, Bernadette Scott, Hertzog P, O’Keeffe M. Differential production of IL-12, IFN-alpha, and IFN-gamma by mouse dendritic cell subsets. Journal of Immunology 2001.