蔡菁華Jinghua Tsai Chang陳威廷Wei-Ting Chen醫學研究所2026-10-299999-12-312016https://handle.ncl.edu.tw/11296/v7yjcfhttps://scholars.csmu.edu.tw/handle/123456789/36177Slit2是一個分泌型的醣蛋白，除了具有抑制神經軸突遷徙的能力外，也扮演著抑癌基因的角色。我們發現Slit2擁有兩種不同的剪接變異型(alternative splicing)，分別為具有exon15的Slit2-WT，以及缺乏exon15的Slit2-ΔE15。當Slit2-ΔE15 在CL1-5細胞中過度表現時可以抑制細胞的侵犯(invasion)以及生長(growth)的能力；但是，Slit2-WT只具有抑制肺癌細胞的侵犯能力，而不影響生長。我們發現肺癌病人的腫瘤組織以及腫瘤周圍之正常組織高度表現Slit2-WT，而Slit2-ΔE15表現很低或不表現。有趣的是，將近41%的氣胸病人(非癌症患者)，其肺組織之Slit2-ΔE15的表現量大於或等於Slit2-WT。我們利用細胞實驗，發現了高濃度的Gallic Acid和Glucose具有調控Slit2剪接變異型的潛在能力。在動物實驗方面，我們發現小鼠Slit2的剪接變異型表現具有組織特異性，在肺部Slit2-WT的表現遠高於Slit2-ΔE15,而在腦部Slit2-WT與Slit2-ΔE15的表現量相當。由於腫瘤的發生與慢性發炎反應有密切的關係，我們以卵清蛋白(ovalbumin, OVA)及脂多醣體 (lipopolysaccharide, LPS)分別在小鼠身上誘發慢性及急性肺發炎。慢性肺發炎小鼠的肺臟，其Slit2-WT/Slit2-ΔE15的剪接變異型比例有增加的趨勢，且Slit2的總表現量有上升的趨勢; 急性肺發炎小鼠的肺臟，其Slit2-WT/Slit2-ΔE15的剪接變異型比例則有降低的趨勢，且Slit2的總表現量也有減少的趨勢。慢性肺發炎的Slit2剪接變異型的表現與人類肺癌病人之肺組織表現非常類似，而急性肺發炎的Slit2剪接變異型的表現與氣胸病人之肺組織的表現非常類似。以尾靜脈模擬肺癌轉移的小鼠模式中發現，肺腫瘤的發生會使周邊肺組織之Slit2-WT/Slit2-ΔE15的剪接變異型比例降低，且Slit2的總表現量減少。肺轉移的動物模式與人類肺癌病人之肺臟組織中Slit2的剪接變異型表現趨勢不同，與急性肺發炎的小鼠模式較接近，暗示小鼠的轉移模式與人類肺癌形成的過程是不一樣的。過去的研究顯示Slit2具有抑制嗜中性球，但促進嗜酸性球的趨化作用的現象，但Slit2的不同剪接變異型對於免疫細胞的影響尚未被研究，我們將進一步了解不同剪接變異型的Slit2對於各種免疫細胞的作用，以及對於肺癌形成的影響。Slit2 is a secreted glycoprotein, which not only inhibits the migration of axon but also is a tumor suppressor gene. We identified two splicing variants in Slit2, the one with exon15 is Slit2-WT, and another without exon15 is Slit2-ΔE15. When Slit2-ΔE15 was overexpressed in CL1-5 cells, Slit2-ΔE15 inhibited cell growth and invasion; When Slit2-WT was overexpressed in CL1-5 cells, it inhibited cell invasion but not cell growth. Slit2-WT form was highly expressed in lung specimen of lung cancer patients while Slit2-ΔE15 was expressed in a very low level. Interestingly, nearly 41% of pneumothorax patients’ lung tissues expressed higher or equal levels of Slit2-ΔE15 than Slit2-WT. We found Gallic Acid and high glucose had the ability to regulate Slit2 splicing in cell culture. In animal experiments, we observed that expression of Slit2 splicing forms is tissue specific. The expression of Slit2-WT is much higher than Slit2-ΔE15 in mice’s lung, but their expression levels are nearly equal in mice’s brain. Since tumorigenesis is closely related with chronic inflammation, we used ovalbumin (OVA) and lipopolysaccharide (LPS) to treat mice to induce chronic and acute lung inflammation. In chronic inflammatory lung, the ratio of Slit2-WT/Slit2-ΔE15 expression levels was increased and the overall expression level of Slit2 was also increased; in acute inflammatory lung, the ratio of Slit2-WT/Slit2-ΔE15 expression levels was reduced and the overall expression level of Slit2 was also decreased. The pattern of Slit2 splicing was similar between lung cancer patients and chronic lung inflammation in animal model while the pattern of Slit2 splicing was similar between human pneumothorax patients and acute lung inflammation in animal model. Unexpectedly, Slit2 splicing pattern in lung metastasis animal model was different from that in lung cancer patients and chronic lung inflammatory animal model, but was similar to the acute lung inflammatory animal model. The differences of Slit2 splicing between lung cancer and lung metastasis in animal model may be explained by that the tumorigenesis may be quite different between these two systems. Previous study has revealed that Slit2 inhibits chemotaxis of neutrophil which infiltrated into acute inflammatory lung, but enhances chemotaxis of eosinophil which infiltrated into OVA induced chronic inflammatory lung. However, the role of Slit2 splicing variants in immune system has not been investigated yet. We will study how Slit2 splicing variants affect function of various immune cells and understand how the effect of Slit2 on immune system impacts lung tumorigenesis.Slit2剪接變異型腫瘤生長過程發炎反應Slit2 Splicing FormTumorigenesisInflammationthesis