3.1 Differentially expressed mRNAs and lncRNAs
Six clinical samples including three normal tissues and three cancer tissues were obtained from patients diagnosed with NSCLC and used for sequencing analysis. The differentially expressed mRNAs and lncRNAs in different groups were calculated. Compared with normal tissues, 1327 mRNAs in case 1, 3493 mRNAs in case 2 and 1315 mRNAs in case 3 were upregulated and 1349 mRNAs in case 1, 4280 mRNAs in case 2 and 1273 mRNAs in case 3 were downregulated in NSCLC tissues. Additionally, there were 2709 upregulated-mRNAs and 1846 downregulated- mRNAs in three total NSCLC tissues compared with three total normal tissues (Fig. 1A). Consistently, compared with normal tissues, 544 lncRNAs in case 1, 707 lncRNAs in case 2 and 438 lncRNAs in case 3 were upregulated and 660 lncRNAs in case 1, 2062 lncRNAs in case 2 and 480 lncRNAs in case 3 were downregulated in NSCLC tissues. And 449 upregulated-mRNAs and 281 downregulated- mRNAs present in three total NSCLC tissues compared with three total normal tissues (Fig. 1B). Briefly, these results suggested that these genes could distinguish cancerous tissues from normal tissues.
3.2 GO and KEGG enrichment analysis
Functional annotation were executed through GO and KEGG enrichment analysis of the differentially expressed mRNAs and lncRNAs above, thereby contributing to the biological significance of the lncRNA and mRNA. The results revealed that the majority of the top 20 GO function according to P value were involved immune process, such as immune system process, regulation of immune response, adaptive immune response, antigen binding and complement activation, classical pathway and so on (Fig. 2A). Moreover, according to the KEGG pathway analysis, the top 20 KEGG pathway according to P value were related to metabolism, such as mineral absorption, nitrogen metabolism, alanine, aspartate and glutamate metabolism and arginine biosynthesis and so on, phagosome, cytokine-cytokine receptor interaction, chemokine signaling pathway and IL-17 signaling pathway and so on(Fig. 2B). Overall, the data indicated that the process of immune response, metabolism, phagosome, cytokine receptor interaction and IL-17 signaling pathway mainly functioned in NSCLC.
3.3 The dysregulated expression of lncRNAs and mRNAs
To explore the biologic function of lncRNAs and mRNAs, we firstly screened out the top 10 dysregulated lncRNAs and mRNA (Fig. 3A and B). Among 10 dysregulated lncRNAs, eight lncRNAs (HOXC-AS2, IGHV3-66, MIR8071-2, IGHV1-69, AL590666, AL365181, MIR8071-1 and FAM30A) were upregulated, and two lncRNAs (ADAMTS9-AS2 and AC092053) were downregulated in NSCLC (Table 2). However, all the 10 dysregulated mRNAs (EEF1A2, PPP2R2C, MUC5AC, AKR1B10/15, ONECUT1, B3GNT3, GNG4, NPY, DKK1 and IGF2BP1) were upregulated in NSCLC (Table 2). Furthermore, four deregulated lncRNAs still remained novel, and the HOXC-AS2, AL590666, AL365181, FAM30A, ADAMTS9-AS2 and AC092053 have been known to be involved in NSCLC (Table 2).
3.4 The construction of lncRNA-mRNA co-expression networks
To further detect the biologic function of dysregulated lncRNAs, we then constructed the co-expression network of dysregulated lncRNAs. Using the dysregulated lncRNAs as the center of the network, we can clearly see possible regulated target genes (Fig. 4). The results showed that the entire network was composed of 7 independent parts, the largest part had 28 nodes, and the smallest is only one-to-one regulation. Importantly, among these ten dysregulated lncRNAs, half of them (FAM30A, IGHV3-66, IGHV1-69, MIR8071-2 and MIR8071-1) interacted with IGH gene cluster, indicating that dysregulated lncRNAs may be association with immune process (Fig. 4).
3.5 The construction of lncRNA-mRNA biological function and pathway network
To verify the biologic function of dysregulated lncRNAs, the lncRNA-mRNA biological function and pathway network was constructed. Besides the basic biological functions, such as plasma membrane, external side of plasma membrane, extracellular region and membrane, the dysregulated lncRNAs were mainly involved in immune process, such as immune response, immune system process, immunoglobulin receptor bingding, adaptive immune response, Fc-gamma receptor signaling pathway involved in phagocytosis, antigen binding, immunoglobulin complex, circulating and positive regulation of B cell activation. Additionally, complement process, such as regulation of complement activation and complement activation, classical pathway, phagocytosis process such as phagocytosis recognition and phagocytosis engulfment were also correlation with the dysregulated lncRNAs (Fig. 5). Shortly, the data suggested that the dysregulated lncRNAs were indeed related with immune process.