BC is the most common urologic malignancy (2) with the feature of high recurrence and high mortality in China (17). Bladder urothelial carcinoma is the predominant histologic type (4). Currently, resection of bladder tumors is the most common surgical operation for noninvasive bladder cancer, but the recurrence rate is still high (18). Therefore, the investigations of BC using bioinformatics may contribute to explore the hub genes and important signaling pathways, reveal the development mechanism, and provide possible molecular markers and potential therapeutic targets for the clinical diagnosis and treatment of BC.
In the present study, we downloaded a dataset GSE7476 from GEO and the sequencing data of BC from TCGA. Comparison of gene data between the BC tissues and para-cancer tissues were performed by R software. After the intersection, 220 DEGs were obtained, including 28 up-regulated genes and 192 down-regulated genes. The up-regulated DEGs were mainly enriched in the BP, mainly including sister chromatid segregation, mitotic sister chromatid segregation and mitotic nuclear division; in the CC, it was mainly concentrated in spindle, cyclin B1-CDK1 complex and anaphase-promoting complex; in terms of MF, it is mainly concentrated in the histone kinase activity, DNA binding bending and cyclin-dependent protein serine/threonine kinase activity. The down-regulated DEGs were also mainly enriched in the BP, including muscle contraction, muscle system process, regulation of muscle contraction and so on; in the CC, the main components were such as the contractile fiber part, contractile fiber, myofibril; in terms of MF, it is mainly concentrated in glycosaminoglycan binding,structural constituent of muscle and heparin binding. The KEGG pathway analysis showed that the up-regulated DEGs were co-enriched in 6 signaling pathways, which might play an important role in the cell cycle signaling pathway. Cell cycle includes cell replication and cell division, and it is closely related to cell growth, anabolism, and proliferation. The abnormal cell proliferation and DNA replication are important causes of cancer. In the past, many studies have reported that cell cycle plays an important role in the occurrence and development of tumors (19). In the occurrence process of tumor, it is common that the cell grow out of control, basically manifested as cell cycle supernormal operation out of control (20). Therefore, the changes of cell cycle affect in all probability the normal proliferation and apoptosis of cells, and in-depth study of cell cycle is conducive to revealing the specific process of the occurrence and development of tumors, sequentially helping to find key therapeutic targets. As shown by KEGG pathway analysis, the down-regulated DEGs showed that these genes were involved in 15 signaling pathways, and were most likely to play a significant role in Interleukin 17(IL-17) pathway. IL-17 is a pro-inflammatory cytokine secreted primarily by activated CD4 (+) T helper cells, CD8 (+) T cells, and macrophages (21). It’s main biological functions include inducing tumor necrosis factor, stimulating vascular endothelial cell migration and angiogenesis (22, 23). More and more reports indicate that IL-17 plays an important role in the occurrence and development progression of tumors, and its expression is increased in prostate cancer (24), colorectal cancer (25), breast cancer (26), ovarian cancer (27), hepatocellular carcinoma (28), non-small cell lung cancer (29) and other tumors. It has been reported that IL-17 could promote the formation of blood vessels (30), shield the body from immune surveillance of cancer cells (31) and promote the occurrence and development progression of tumors. However, it has also been reported that IL-17 could inhibit tumor progression by enhancing the body's immune resistance to tumor, thereby preventing tumor growth and metastasis (32, 33). Therefore, the study of these signaling pathways may help clarify the carcinogenic mechanism of BC.
In the present study, PPI protein network was constructed to screen out 41 hub gene and three functional modules. These modules are mainly concentrated in signaling pathways such as Cell cycle, IL-17 signaling pathway and Vascular smooth muscle contraction. Then the prognostic survival of hub genes was analyed by TCGA clinical follow-up data, and 15 hub genes significantly related to prognosis were found. These results suggested that these hub genes might play an important role in the development and prognosis of BC.
Encoded by JUN, c-Jun is an oncogenic transcription factor. Wang SS et al. report that JUN is noted in the high-risk groups of BC patients with higher expression, while the lower JUN expression groups have the longer overall survival (34). Li Z et al. demonstrate that c-Jun is significantly overespressed in aggressive bladder cancer tissues and cell lines, and c-Jun is related with anti-proliferative factor (APF)-mediated inhibition for bladder cancer cell growth (35). These reported suggest that JUN may inhibit the growth of BC cell and lower expression of JUN may indicate the better BC prognosis. ACTC1 is reported to be down-regulated in BC (36), which is consistent with the result of our RT-PCT experiment. The previous research manifest that the expression of ACTC1 gradually increases according to WHO grade in glioma and ACTC1-positive groups had a shorter survival time and poorer prognosis (37). However, our research found that the expression of ATCT1 is closely related to the prognosis of BC, and the low expression of ATCT1 indicates a good prognosis. ATCT1 may be a biomarker for prognosis evaluation of BC patients. Of course, more research is also needed to verify this. In regards to connective tissue growth factor (CTGF), Chen et al. make a conclude that urothelial carcinoma cells show increasing apoptosis using extrinsic pathways through upregulation of CTGF (38). Previous researches reveal that CTGF can promoted the progression of tumor cell and significantly correlated with poor clinical prognosis in breast cancer(39).Yuen et al (40) found that TAZ-AXL-CTGF overexpression are associated with colon cancer progression. In addition, CTGF and CYR61 both belong to CCN (CYR61, CTGF, NOV) family of protein. Nevertheless, Jiang et al. show that the members of CCN family play contradictory roles in progression of breast cancer with CYR61 appearing to be an aggressive stimulator and CTGF acting as a cancer suppressor (41). Our results show that CTGF and CYR61 are related to the prognosis of BC patients, and down-regulated in BC. These molecules are pivotal targets affecting the prognosis of BC although the role of these molecules in BC remains obscure. As a member of the calponin family of actin-binding proteins, Transgelin (TAGLN) participates in cell motility and migration (42). As is well-known, cell migration is the crucial process of cancer metastasis. Wu et al. certified that TAGLN overexpression promote the tumor progression in lung adenocarcinoma (43). Calponin h1(CNN1) is certified to have the function of cytoskeleton-stabilizing and transformation-inhibiting by combining with actin and related proteins (44). Therefore, CNN1 is possible to inhibit the tumor cell growth in the process of cancer development. As a matter of fact, it is reported that downregulation of CNN1 is necessary for cell transformation, and low of CNN1 expression is crucial for its metastasis in ovarian high-grade serous carcinoma (44). Liu Y identify that TAGLN and CNN1 are connected to both overall survival and disease-free survival in BC and act as potential prognostic molecular markers (45). Our researched also showed that TAGLN and CNN1 expression was down-regulated in BC and closely related to the prognosis of patients with BC. The hub gene EGR2 (early growth response 2 protein) which as a cancer suppressor is down-regulated in gastric cancer, and the oncogenic effect of miR-20a is significantly attenuated via up-regulated EGR2 expression (46). Low expression of EGR2 is associated with cancer metastasis and shorter survival in gastric cancer patients (47). Unoki et al. demonstrats that EGR2 can induce the apoptosis of cancer cell lines by activating the expression of BNIP3L and BAK (48). In our study, low expression of EGR2 was found to be associated with a good prognosis in patients with BC. However, there are few studies on EGR2 in BC, and more studies are needed. Myosin heavy chain 11 (MYH11) belongs to myosin heavy chain family, and the mutation of MYH11 was related to is related to the development of intestinal cancer (49, 50). It has been also reported to be a biomarker for BC. Hu et al. reported that Myh11 expression was significantly lower in tumor samples and the high expression of MYH11 was related to worse OS of BC patients, which was consistent with our study (51). Filamin C (FLNC) is one member of the filamin protein family, which was associated with heart disease (52). The methylation of FLNC has reported in several cancers, such as prostate cancer (53) and gastric cancer (54). These reports showed that the downregulation of FLNC can induce the expression of matrix metallopeptidase, which can regulate the degradation of extracellular matrix and cancer metastasis.
Similarly studies show that FLNC with low expression in BC was involved in tumor migration and invasion (55). In addition, this study showed that FLNC is lowly expression in BC. And it was significantly associated with prognosis of BC patients.
Based on our results and the above literature, we believe that these hub genes may be potential prognostic markers and therapeutic targets for bladder cancer. There, it is necessary and meaningful for further study to investigate the specific mechanism of these genes in BC.