Aberrant mRNA and protein expressions of ITGB superfamily members in patients with STAD
ONCOMINE database was utilized in order to research the mRNA expression of ITGB superfamily members in patients with STAD. Compared with normal tissues, the mRNA expressions of ITGB1, ITGB2, ITGB4, ITGB5 and ITGB8 were significantly elevated, while the mRNA expression of ITGB7 was significantly reduced in STAD tissues (FIGURE 1). Data in TABLE 1 was consistent with the results above. For instance, in the Chen dataset the transcriptional levels of ITGB1 were significant higher in gastric intestinal type adenocarcinoma and gastric mixed adenocarcinoma, with the fold changes of 2.130 and 3.145, respectively [24]. In the DErrico dataset, the transcriptional level of ITGB2 was significantly up-regulated in diffused gastric adenocarcinoma with the fold change of 2.318 [25]. In the Cho dataset, the transcriptional levels of ITGB7 were significantly reduced in gastric mixed adenocarcinoma, diffused gastric adenocarcinoma and gastric adenocarcinoma with the fold changes of -5.811, -5.450 and -4.872, respectively [26].
Table 1 Transcriptional levels of ITGB superfamily members in different types of gastric cancers and normal tissues (ONCOMINE)
|
Types
|
Fold Change
|
P-value
|
t-test
|
Reference
|
ITGB1
|
Gastric Intestinal Adenocarcinoma
|
2.130
|
8.56E-17
|
10.951
|
Chen
|
|
Gastric Mixed Adenocarcinoma
|
3.145
|
4.08E-4
|
5.242
|
Chen
|
|
Diffused Gastric Adenocarcinoma
|
2.123
|
9.32E-4
|
4.241
|
DErrico
|
|
Gastric Intestinal Adenocarcinoma
|
2.028
|
5.05E-4
|
3.570
|
Cho
|
|
Gastric Mixed Adenocarcinoma
|
2.789
|
9.79E-4
|
3.731
|
Cho
|
ITGB2
|
Gastric Mixed Adenocarcinoma
|
2.398
|
5.20E-4
|
4.235
|
Chen
|
|
Diffused Gastric Adenocarcinoma
|
2.033
|
4.45E-4
|
3.845
|
Chen
|
|
Diffused Gastric Adenocarcinoma
|
2.318
|
0.003
|
3.624
|
DErrico
|
ITGB4
|
Gastric Intestinal Adenocarcinoma
|
2.105
|
1.41E-4
|
4.038
|
Cho
|
|
Gastric Mixed Adenocarcinoma
|
2.008
|
0.003
|
3.242
|
Cho
|
ITGB5
|
Gastric Intestinal Adenocarcinoma
|
3.241
|
4.89E-9
|
6.783
|
DErrico
|
ITGB7
|
Gastric Mixed Adenocarcinoma
|
-2.723
|
2.08E-5
|
-5.811
|
Cho
|
|
Diffused Gastric Adenocarcinoma
|
-2.157
|
8.74E-7
|
-5.450
|
Cho
|
|
Gastric Adenocarcinoma
|
-2.449
|
0.003
|
-4.872
|
Cho
|
|
Gastric Intestinal Adenocarcinoma
|
-2.318
|
1.48E-4
|
-3.981
|
DErrico
|
ITGB8
|
Gastric Mixed Adenocarcinoma
|
5.245
|
6.93E-5
|
7.495
|
DErrico
|
|
Gastric Intestinal Adenocarcinoma
|
3.133
|
3.88E-9
|
6.845
|
DErrico
|
|
Gastric Mixed Adenocarcinoma
|
2.092
|
7.71E-4
|
4.403
|
Chen
|
Next, the mRNA expression patterns of ITGB superfamily members were measured by the UALCAN database. The results showed that the mRNA expressions of ITGB1 (p = 1.78e-04), ITGB2 (p = 1.06E-
10), ITGB4 (p = 1.63E-12), ITGB5 (p = 1.67E-09), ITGB6 (p = 6.17E-08) and ITGB8 (p = 1.64E-12) were significantly elevated in STAD vs normal tissues (FIGURE 2). These results were almost consistent with those from ONCOMINE.
After analyzing the mRNA expression of ITGB superfamily members in STAD, the protein expression of ITGB superfamily members were further explored with the Human Protein Atlas. We found that the protein levels of ITGB4, ITGB5, ITGB6 and ITGB8 were significantly up-regulated in STAD tissues (FIGURE 3).
Finally, the correlation between the expressions of ITGB superfamily members and the pathological stage of STAD patients were explored with GEPIA database. As shown in FIGURE 4, there was a significant correlation between the pathological stages and the expression of ITGB2 (p = 0.0143) and ITGB7 (p = 0.0375). These data suggested that ITGB2 and ITGB7 might play a significant role in the tumorigenesis and progression of STAD.
Prognostic value of ITGB superfamily members in patients with STAD
The prognostic value of ITGB superfamily members in STAD patients including overall survival (OS) and disease-free survival (DFS) was investigated with GEPIA. The curves in FIGURE 5A suggested that patients with low transcriptional levels of ITGB1 (p = 0.029) and ITGB3 (p = 0.039) were significantly associated with longer OS. And high transcriptional level of ITGB4 (p = 0.038) was associated with longer OS. The results in FIGURE 5B showed that patients with low transcriptional level of ITGB6 (p = 0.011) were associated with longer DFS.
The Kaplan-Meier Plotter database was also utilized to analyze the correlation between the mRNA expressions of the ITGB superfamily members and the OS of patients with STAD. As shown in the FIGURE 6, the patients with low transcriptional levels of ITGB1 (p = 0.01), ITGB3 (p = 0.0076) and ITGB5 (p = 0.0022) were significantly related to longer OS while a high transcriptional level of ITGB4 (p = 0.045) was significantly related to longer OS.
Prediction of TFs for prognosis-associated ITGB superfamily members
Since the ITGB1, ITGB3, ITGB4, ITGB5 and ITGB6 were significantly associated with the prognosis of patients with STAD, we conducted exploration on the potential TFs for them with NetworkAnalyst database. The visualized network was shown in FIGURE 7A. ITGB1 expression was predicted to be potentially regulated by SP1, CUX1, FHL2, TFAP2A and TFAP2C. ITGB3 expression might be potentially regulated by DAB2 and HOXD3. ITGB4 expression might be potentially regulated by TFAP2A and TFAP2C. DAB2, FHL2 and TFAP2C were predicted to regulate ITGB5. And SP1, CUX1 and FHL2 might potentially regulated ITGB6. Further analysis in FIGURE 7B suggested that the expression of ITGB1 was positively correlated with the expressions of SP1, CUX1 and FHL2, ITGB4 expression was positively associated with TFAP2A expression, and DAB2 expression was positively correlated with the expressions of ITGB3 and ITGB5 (Cor > 0.4, p < 0.05) (GEPIA). However, there was no TF expression positively correlated with ITGB6 expression (FIGURE 7B) (GEPIA). Among the five TFs, the expressions of SP1, CUX1, DAB2 and TFAP2A were significantly up-regulated in STAD tissues vs normal tissues (FIGURE 7C) (GEPIA). Finally, the OS curves by Kaplan-Meier Plotter demonstrated that overexpression of DBA2 was significantly associated with shorter OS while overexpression of TFAP2A was significantly related to longer OS (FIGURE 7D). Therefore, out study implied that DAB2 might synchronously promote the transcription of ITGB3 and ITGB5, and TFAP2A might promote the expression of ITGB4.
Genetic alteration, expression and PPI network of ITGB superfamily members in patients with STAD
The genetic alterations of ITGB superfamily members in patients with STAD were analyzed by cBioPortal online tool. As a result, ITGB1, ITGB2, ITGB3, ITGB4, ITGB5, ITGB6, ITGB7 and ITGB8 were altered in 9, 8, 7, 11, 8, 7, 8 and 11% of the queried STAD samples, respectively (FIGURE 8A). Low mRNA expression was the most common alteration in these samples (FIGURE 8B).
Then the PPI network analysis of ITGB superfamily members was conducted to explore the potential interactions among them by STRING. There were 13 nodes and 68 edges in the PPI network (FIGURE 8C). The ITGB superfamily members were associated with pathways such as regulation of actin cytoskeleton and arrhythmogenic right ventricular cardiomyopathy. The GeneMANIA was also utilized to reveal the functions of ITGB superfamily members and their relative molecules (such as WIF1, ATRNL1, EGFL7, TNC, TNR and TENM4), primarily associated with extracellular matrix organization, extracellular structure organization, receptor complex and integrin complex (FIGURE 8D).
Functional enrichment analysis of ITGB superfamily members in patients with STAD
Firstly, the 20 genes related to ITGB superfamily members were obtained from STRING. Then the functional enrichment analysis of these 28 genes was conducted by DAVID 6.8 and visualized by Hiplot tool. From the FIGURE 9A, the top 10 most highly enriched GO items in the BP category were integrin-mediated signaling pathway, extracellular matrix organization, leukocyte migration, cell-matrix adhesion, cell adhesion, heterotypic cell-cell adhesion, cell adhesion mediated by integrin, viral entry into host cell, leukocyte cell-cell adhesion and endodermal cell differentiation, which were significantly regulated by ITGBs in STAD. The integrin complex, cell surface, plasma membrane, focal adhesion, receptor complex, extracellular exosome, external side of plasma membrane, ruffle membrane, cell-cell adherens junction and hemidesmosome were the top 10 most highly enriched items in the CC category (FIGURE 9B). In the MF category, the ITGB superfamily members and their relative genes were mainly enriched in integrin binding, receptor activity and virus receptor activity (FIURE 9C).
As shown in FIGURE 9D, the top 10 KEGG pathways affected by ITGBs in STAD were regulation of actin cytoskeleton, focal adhesion, hypertrophic cardiomyopathy, dilated cardiomyopathy, ECM-receptor interaction, arrhythmogenic right ventricular cardiomyopathy, PI3K-Akt signaling pathway, proteoglycans in cancer, cell adhesion molecules and pathways in cancer.
Immune cell infiltration of ITGB superfamily members in patients with STAD
The proliferation and progression of cancer cell is associated with immune cell levels. Therefore, the associations between ITGB superfamily members and immune cell infiltration were explored by TIMER database (FIGURE 10). The expression of ITGB1 was positively associated with the infiltration of CD4+ T cell, macrophage and dendritic cell. The expressions of ITGB2 and ITGB3 were both significantly positively related to the infiltration levels of all six immune cells, B cell, CD8+ T cell, CD4+ T cell, macrophage, neutrophil and dendritic cell. There was a remarkably negative correlation between the expression of ITGB4 and the infiltration of CD8+ T cell, CD4+ T cell and dendritic cell. ITGB5 expression was positively associated with the infiltration of CD4+ T cell, macrophage and dendritic cell, and negatively associated with the infiltration of B cell. We also found the higher the infiltration levels of B cell, the higher the expression of ITGB6. The expression of ITGB7 was significantly positively related to the infiltration of CD8+ T cell, CD4+ T cell, macrophage, neutrophil and dendritic cell. ITGB8 expression was positively associated with the infiltration of B cell.