COL7A1 expression in lung squamous cell carcinoma
The exploration of tumor biomarkers has always been a prominent research focus. In order to illustrate its role in diagnosis, we queried COL7A1 expression in tumor tissues and normal tissues in the database. The results showed that COL7A1 was upregulated in a variety of tumors as a component of anchored fibers (Fig. 1). In Oncomine database, we found that the expression of COL7A1 in lung cancer was significantly higher than that in normal tissues, including colorectal cancer and head and neck cancer (Fig. 1a). In Bhattacharjee Lung Statistical analysis, the change of COL7A1 mRNA level in LUSC tissue was 4.547 times higher than that in normal tissue (p-value = 5.84E-11, t-test = 8.946) (Fig. 1b), which was plotted by GraphPad Prism 9 according to the data. In the TCGA database in UALCAN, the expression of COL7A1 in patients with lung squamous cell carcinoma was significantly higher than that in normal tissues (p-value = 1.62E-12) (Fig. 1c). Moreover, we determined the expression of COL7A1 in LUSC by immunohistochemistry (Fig. 2a-d), and quantified the expression of COL7A1 by analysis tools (image J). We found that the expression of COL7A1 in lung squamous cell carcinoma was higher than that in adjacent tissues (Fig. 2e). And COL7A1 was not associated with stages of lung squamous cell carcinoma (Fig. 2f).
Expression changes of COL7A1 in different clinical indexes of LUSC
We have verified the up-regulation of COL7A1 expression in lung squamous cell carcinoma through different databases and IHC experiments. Therefore, we further studied the expression pattern of COL7A1 in patients with LUSC through UALCAN database. This database explores the expression of
COL7A1 in various clinicopathological features including stage, age, smoking habit, lymph node metastasis, molecular subtype and TP53 gene mutation. It is worth noting that the high expression of COL7A1 in patients with LUSC shows an upward trend with age, suggesting that age may affect the expression level of COL7A1 (Fig. 3a). In the comparison of cancer stages, the expression of COL7A1 in the normal group was significantly different from different stages, but there was no significant difference between different stages, indicating that there was no close relationship between the stages of COL7A1 LUSC (Fig. 3b). Further, in the comparison of cancer lymph node metastasis with the
normal group, there was no significant difference between the patient groups, suggesting that there was little relationship between the development of tumor and lymph node metastasis. And they also showed that there was no significant difference between the patient groups in the comparison of TP53 mutations, subtypes and smoking habits of patients (Fig. 3c-f).
Association of COL7A1 expression with prognosis in LUSC patients
Kaplan–Meier plotter was used to analyze the relationship between COL7A1 expression and first progression survival (FP), overall survival (OS) and post-progression survival (PPS) in patients with LUSC. As shown in Fig. 4a, the high expression of COL7A1 indicates that first progression (FP) in all patients with lung squamous cell carcinoma is disappointing (HR = 1.4, p = 0.00059). Figure 4b showed that the overall survival (OS) and of patients with high COL7A1 expression were significantly lower than those with low COL7A1 expression (HR = 1.33, P = 8.1E-6), but not statistically significant with post-progress survival (PPS) (HR = 1.24, P = 0.098) (Fig. 4c). Prognostic analysis showed that COL7A1 could be used as an important index to evaluate the prognosis of LUSC. Finally, we used
GEPIA to analyze the staging of COL7A1 in LUSC, and the results showed that COL7A1 had little relationship with tumor stage (Fig. 4d).
Gene co-expression analysis and protein-protein interaction
Based on the above results of COL7A1 expression and prognosis analysis, we chose to further bioinformatics analysis of COL7A1. The first 14 genes co expressed with COL7A1 gene in LUSC were screened from Oncomine database (Fig. 5a). COL7A1 co-expresses with GPC1 (r = 0.817), SERPINB5 (r = 0.752), BNC1 (r = 0.724), PKP1 (r = 0.724), SOX15 (r = 0.724), PVRL1 (r = 0.724), FAT2 (r = 0.732), ANXA8L2 (r = 0.724), S1PR5 (r = 0.724), DST (r = 0.724), TRIM29 (r = 0.724), GJB5 (r = 0.724), IRF6 (r = 0.724), KRT17 (r = 0.724), It could be seen that GPC1 is a related gene.
Further GEPIA analysis shows the correlation between COL7A1 and GPC1 (r = 0.46, p-value = 2E-26), the expression of GPC1 was verified in GEPIA databases (Fig. 5b). We found that GPC1 was significantly up-regulated in lung squamous cell carcinoma (Fig. 5c). Survival analysis of Kaplan-Meier plotter database confirmed that upregulation of GPC1 was closely related to the prognosis of OS in patients with lung squamous cell carcinoma (Fig. 5d). STRING mapped the interaction network between COL7A1 and other proteins to infer the possible role of COL7A1 in LUSC. As showed in Fig. 5e, COL7A1 is closely related and co-expression with MIA3, BMP1, ITGB3, ITGB1, ITGAV, LAMC2, LAMB3, P4HA2, ITGA2, TLL1. At the same time, whether through databases or experiments, it is confirmed that these proteins are closely related to each other.
GO and KEGG enrichment
Based on the results of protein-protein interaction and gene co-expression analysis, we further carried out bioinformatics analysis. By analyzing the gene ontology (GO) and Database Kyoto Encyclopedia of Genes and Genomes (KEGG) for Annotation, Visualization and Integrated Discovery (DAVID), the functions of COL7A1 and the genes significantly related to the changes of COL7A1 were predicted. Go enrichment analysis predicted the function of target host genes from biological process (BP) (Fig. 6a), cell composition (CC) (Fig. 6b) and molecular function (MF) (Fig. 6c). In biological process (BP), we found that GO:0048208 (COPII vesicle coating), GO:0006888 (ER to Golgi vesicle-mediated transport), GO:0030198 (extracellular matrix organization) were significantly regulated by the COL7A1 alterations in LUSC. In cell composition (CC), GO:0012507 (ER to Golgi transport vesicle membrane), GO:0000139 (Golgi membrane) and GO:0030127 (COPII vesicle coat) were obviously controlled by COL7A1 and related genes alteration. Moreover, in molecular function (MF), we could see that GO:0031418 (L-ascorbic acid binding) and GO:0004656 (procollagen-proline 4-dioxygenase activity) were obviously regulated by the COL7A1 and related genes alterations in LUSC. KEGG analysis can determine the pathways related to the functional changes of COL7A1 and related genes (Fig. 6d). Through KEGG analysis, 10 pathways related to the functional changes of COL7A1 in lung squamous cell carcinoma were found. Among them, hsa04512: ECM-receptor interaction, hsa05222: Small cell lung cancer, has04510: Focal adhesion might be involved in the occurrence and pathogenesis of lung squamous cell carcinoma.