中文摘要 ccRCC (clear-cell renal cell carcinoma)是在全世界最常見的腎臟癌。70%以上的病人在VHL腫瘤抑制基因有突變。VHL的突變通常增加HIF(Hypoxia induced factor)的活性導致代謝跟微環境的改變。為了研究ccRCC形成的過程和病因，我們把老鼠腎小管的VHL基因剔除，結果缺VHL機能的腎臟出現發炎以及纖維化的症狀。其他研究也發現慢性腎臟病(一種慢性發炎疾病)病人得腎臟癌比例比健康人高很多。這件事情就表示發炎有可能是腎臟癌發生的前導。這個研究的目的是從病理及基因體的變化來分析發炎和癌症的相關。在缺VHL機能的腎臟除了Myc之類的細胞轉型因素以外還有調控Macrophage功能等等導致免疫反應的基因過度表現。我們接著用資料庫(GSE66494和GSE36895)下載的慢性腎臟病檢體，腎臟癌第一期及第三期的腫瘤樣本（以上腎臟癌的樣本均缺VHL gene功能）與缺VHL功能的老鼠比對。因為在所有病變組織(發炎的腎臟和癌化的腎臟)中表現下降的基因是跟代謝的過程有關，所以我們能夠確認缺VHL的功能以及發炎會導致不正常的代謝過程。我們還有假設只有在發炎階段到初期的癌症之間表現很高的基因有可能是癌症發生的關鍵, 且在所有病變組織表現很高的基因，與只有在發炎階段及後期的癌症表現高的基因，可能是與轉移有關。比方說只有在發炎階段到初期癌症上升的基因包括跟發育過程有關的基因，這表示形成癌症的反分化（dedifferentiation）過程有可能在發炎階段已開始。除了只有在發炎階段到初期癌症之間上升的基因之外發炎階段和後期癌症上升的基因以及所有病變組織表現很高的基因也包括免疫反應有關的基因，這件事情表示發炎不只導致癌症的產生。再加上在第一期與第三期腫瘤中，與DNA損傷有關的基因的表現有增加，這結果表示癌症發生初期時就有嚴重的DNA損傷。為了驗證DNA的損傷我們做了IHC，然後發現人體的RCC樣本和VHL基因剔除的老鼠都有Gamma-H2AX (它是表示DNA 鏈斷裂的標靶)，但是它的表現量在RCC的樣本較明顯。IHC的結果表示DNA的損傷在發炎的階段已開始累積，且支持我們的發炎導致腎臟癌的假設。 ;Abstract Clear-cell renal cell carcinoma (ccRCC) is the most common subtype of renal cell carcinoma in the world. Over 70% of ccRCC patients have mutations in the von Hippel–Lindau (VHL) tumor suppress gene. Mutations in VHL activate hypoxia-inducible factor 1-alpha (HIF1A) and result in alteration of microenvironment and metabolic pathways. We have generated conditional VHL knockout mouse model to study the etiology of ccRCC progression. In this mouse model, we observed that inactivation of VHL in the kidney tubules resulted in tissue inflammation and fibrosis. Further, it was reported that patients of chronic kidney disease (CKD), a chronic inflammatory disorder, were more likely to develop this cancer compared to the general population. These observations suggest a role of inflammation in kidney cancer development. The goal of this study is to dissect the correlation between inflammation and kidney cancer development. By analyzing expression profiles of mouse VHL mutant kidneys relative to wild-type kidneys, we found a cluster of genes involved in immune response were over-represented. These over-represented immune response genes include regulators of macrophage functions, in addition to those involved in cellular transformation such as Myc. We then compared this gene expression profile to those of the chronic kidney disease (CKD) kidney (GSE66494), and stage 1 and stage 3 ccRCC, obtained from the public domain database (GSE36895；only those containing VHL mutations were selected). Many genes significantly down-regulated in inflammatory kidney (VHL knockout mouse and CKD patients) and in early stage and/or late stage ccRCC are associated with metabolic processes, confirming that metabolic abnormality is the feature of VHL mutation and inflammatory kidney. We hypothesized that genes activated in inflammation kidneys and early-stage cancer may play important roles in the initiation of inflammation-induced cancer. For example, we found genes involved in the developmental process were over-represented in both inflammatory tissues and in early-stage cancer. This may suggest that dedifferentiation process central to cancer formation may occur in precancerous inflammation. Further, genes that were activated only in inflammation kidneys and late-stage cancers and genes consistently up-regulated in all diseased kidneys included many immune response genes, suggesting that inflammation may be not only precursor of cancer formation but also may play important roles in cancer progression. Moreover, chemokines that are involved in immune cell recruit may also be involved in the induction of metastasis. Genes that were significantly up-regulated both in stage1 and stage3 cancers but not in VHL knockout mouse or CKD kidneys include DNA damage response associated genes. This result suggests that DNA damages occur at the onset of cancer. The result of immunohistochemistry (IHC) suggested that accumulation of DNA damages occurred in pre-cancerous inflammation tissues. Based on these results, we proposed the genetic model for ccRCC initiation and progression.
