生物标志物，即与生理或病理过程有关的可监测变化，它最重要的特征是变化。机体内环境由于受到稳态机制的严格调控，在达到失代偿的临界点之前，疾病引起的早期微小变化会被迅速降解或排出体外。而尿液没有任何受稳态调控的机制和必要性，是接收血液代谢的地方，因此，尿液是生物标志物的理想来源，且能够敏感地反映疾病的早期变化。本文利用液相色谱串联高分辨率质谱 (LC-MS/MS)，基于数据非依赖性采集 (DIA) 技术，分别对B16黑色素瘤荷瘤小鼠模型、RM-1前列腺癌荷瘤小鼠模型及α-synuclein转基因帕金森病小鼠模型的尿液蛋白质组进行研究。

论文第一章，探究B16黑色素瘤荷瘤小鼠的尿液蛋白质组变化。黑色素瘤是最具侵袭性的恶性皮肤肿瘤之一。通过皮下注射B16黑色素瘤细胞建立荷瘤小鼠模型，分别收集与肿瘤发生发展对应的4个时间点的尿液样本进行蛋白质组分析。经DIA定量分析，与注射肿瘤细胞前相比，共筛选到38种人同源差异蛋白质，其中，18种蛋白质被报道与黑色素瘤相关。在肿瘤可见前，4种蛋白质出现显著表达差异且均被报道与黑色素瘤相关，它们分别是骨桥蛋白、碳酸酐酶1、粘蛋白13和α-心肌肌动蛋白。以上结果为今后临床研究早期诊断黑色素瘤提供了有价值的线索。

论文第二章，探究RM-1前列腺癌荷瘤小鼠的尿液蛋白质组变化。前列腺癌是男性群体中最常见的恶性肿瘤之一。通过皮下注射RM-1前列腺癌细胞建立荷瘤小鼠模型，分别收集与肿瘤发生发展对应的4个时间点的尿液样本，利用LC-MS/MS分析尿液蛋白质组。经DIA定量分析，与注射肿瘤细胞前相比，共筛选到14种人同源差异蛋白质，其中9种蛋白质被报道与前列腺癌相关。在肿瘤可触摸前，9种蛋白质出现显著差异，有6种被报道与前列腺癌相关，它们分别是β-2-糖蛋白1、丛生蛋白、转铁蛋白、电压依赖性阴离子选择性通道蛋白1、电压依赖性阴离子选择性通道蛋白3和软骨寡聚基质蛋白。以上结果为今后临床研究早期诊断前列腺癌提供了有价值的线索。

论文第三章，探究α-synuclein转基因帕金森病小鼠的尿液蛋白质组变化。帕金森病 (PD) 是第二常见的神经退行性疾病，严重影响患者的生活质量，造成巨大的社会经济负担。利用α-synuclein转基因小鼠，分别收集小鼠第2、4、6、8、10月的尿液样本，经DIA定量分析，与第2月相比，共筛选到17种人同源差异蛋白质，其中，9种蛋白质被报道与PD相关，成蛋白-2、剪接因子3A亚基1、异戊烯二磷酸δ-异构酶1在第6、8、10月连续变化。以上结果为今后临床研究早期诊断PD提供了有价值的线索。

Biomarkers are the measurable changes associated with the physiological or pathophysiological process, and its most important feature is change. The internal environment is strictly regulated by the homeostatic mechanism, before the critical point of decompensation is reached, the early small changes caused by the disease will be rapidly eliminated. Urine does not have any homeostasis, and is the place to receive blood metabolism. Therefore, urine is an ideal source of biomarkers and can sensitively reflect the early changes in disease. In this paper, the urine proteome of B16 melanoma tumor-bearing mouse model, RM-1 prostate cancer tumor-bearing mouse model, and α-synuclein transgenic mouse model of Parkinson's disease (PD) were studied by liquid chromatography-tandem mass spectrometry (LC-MS/MS) based on data-independent acquisition (DIA) technology.

The first chapter explores the changes of urine proteome in the B16 melanoma-bearing mice. Melanoma is one of the most aggressive malignant skin tumors. The tumor-bearing mouse model was established by subcutaneous injection of B16 melanoma cell line, urine samples were collected at four time points corresponding to the development of tumor. Based on DIA technology and compared with those before the injection of tumor cell line, 38 human homologous differential proteins were identified, and 18 proteins were reported to be related to melanoma. Before the tumor was visible, there were 4 differential proteins and all were reported to be related to melanoma. They were Osteopontin, Carbonic anhydrase 1, Mucin-13, and Alpha-cardiac actin. The above results provide valuable clues for the early diagnosis of melanoma in clinical studies.

The second chapter explores the changes of urine proteome in the RM-1 prostate cancer-bearing mice. Prostate cancer is one of the most common malignancies in male. The tumor-bearing mouse model was established by subcutaneous injection of RM-1 prostate cancer cell line. Urine samples were collected at four time points corresponding to tumorigenesis and development, and the urine proteome was analyzed by LC-MS/MS. Based on DIA technology and compared with those before the injection of tumor cell line, 14 human homologous differential proteins were identified, and 9 proteins were reported to be associated with prostate cancer. Before the tumor was palpable, 9 proteins showed significant differences, and 6 were reported to be associated with prostate cancer. They were Beta-2-glycoprotein 1, Clusterin, Transferrin, Voltage-dependent anion-selective channel protein 1, Voltage-dependent anion-selective channel protein 3, and Cartilage oligomeric matrix protein. The above results provide valuable clues for the early clinical diagnosis of prostate cancer in the future.

         The third chapter explores the changes of urine proteome in α-synuclein transgenic mice of PD. PD is the second most common neurodegenerative disease, which seriously affects the quality of life of patients, causing a huge socioeconomic burden. The urine samples collected from months 2-, 4-, 6-, 8- and 10- month-old α-synuclein transgenic mice. Based on DIA technology, a total of 513 proteins were identified. 17 human homologous differential proteins were screened and compared with those in the urine of 2-month-old mice, and 9 proteins were related to PD. Formin-2, Splicing factor 3A subunit 1 and Isopentenyl-diphosphate Delta-isomerase 1 changed continuously in months 6, 8 and 10. The above results provide valuable clues for the early diagnosis of PD in future clinical research.