Wang, Jie
(2022).
The Role of the Lysine-Specific Demethylase 1 in the Development of Hepatocellular Carcinoma.
PhD thesis, Universität zu Köln.
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PDF (Doctoral dissertation from Jie Wang)
Doctoral_Dissertation_JieWang2022.pdf - Published Version Download (4MB) | Preview |
Abstract
Hepatocellular carcinoma (HCC) has a low survival rate and is currently the third leading cause of cancer-related deaths worldwide. Most HCC develops on the basis of chronic liver diseases, such as HBV and HCV hepatitis, alcoholic liver disease, or non-alcoholic fatty liver disease. Epigenetic alterations, including an altered pattern in histone modification, are crucial for cancer progression. However, the epigenetic aberrations involved in the development of HCC are not well understood. The lysine-specific demethylase 1 (LSD1) is involved in chromatin remodeling by demethylating lysine 4 and lysine 9 histone 3 (H3K4 and H3K9), causing transcriptional repression or activation, respectively. Strikingly, overexpression of LSD1 contributes to the malignancy of several cancers. Therefore, my research focused on the mechanistic links affected by LSD1 in liver cancer cells. To investigate this, I used three different hepatoma cells (Huh7, HepG2, and Hep3B), in which LSD1 was inhibited pharmacologically or by anti-LSD1 siRNA species. For conditional LSD1 inhibition, stable Tet-On hepatoma cell lines were generated in which expression of short-hairpin anti-LSD1 RNA was inducible after doxycycline exposure. The effect of LSD1 on cell viability was measured by the MTT test. Gene expression was studied at the transcript level by ultra-deep RNA sequencing and qPCR and at the protein level by immunoblotting. Moreover, I analyzed the histone H3K4 methylation patterns and the interaction of LSD1 with promoter sites by chromatin immunoprecipitation (ChIP) followed by whole-genome sequencing or qPCR. These studies showed that LSD1 inhibition in the different hepatoma cell types leads to cell growth arrest and downregulation of PLK1. ChIP analysis revealed that PLK1 is a direct target of LSD1 regulation in hepatoma cells. In addition, gene expression profiling by RNA sequencing followed by metabolic pathway analysis revealed striking dysregulation of genes involved in metabolic dysregulation after LSD1 was inhibited. In particular, genes of the citrate cycle and lipid metabolism were affected by LSD1 which was validated by qPCR. Noteworthy, ChIP assays showed alteration of histone methylation and LSD1 binding at promoter sites of many metabolic genes, downregulated after LSD1 inhibition. In particular, the gene FABP5, which is involved in fat metabolism, was found to be a novel direct target of LSD1. To demonstrate the effects of LSD1 on the regulation of metabolic genes, an in vivo mouse model for non-alcoholic fatty liver disease was used with respect to its high metabolic imbalance and LSD1 was pharmacologically inhibited in the early progression phase of the disease where fat accumulation occurs (steatosis). Importantly, LSD1 inhibition resulted in weight loss, lower serum AST liver enzymes, and no signs of fat accumulation, while control mice had all the features of steatosis. In conclusion, my study emphasizes that LSD1 which is an important mediator in cell cycle control affects HCC progression not only by cell cycle interruption but also by metabolism and lipid dysregulation.
| Item Type: | Thesis (PhD thesis) | ||||||||
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| URN: | urn:nbn:de:hbz:38-631385 | ||||||||
| Date: | 24 August 2022 | ||||||||
| Language: | English | ||||||||
| Faculty: | Faculty of Medicine | ||||||||
| Divisions: | Faculty of Medicine > Pathologie und Neuropathologie > Institut für Pathologie | ||||||||
| Subjects: | Natural sciences and mathematics Medical sciences Medicine |
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| Date of oral exam: | 17 August 2022 | ||||||||
| Referee: |
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| Refereed: | Yes | ||||||||
| URI: | http://kups.ub.uni-koeln.de/id/eprint/63138 |
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