In order to disclose the underlying molecular mechanisms between SP cells and human OS, we characterized the possible GO functional terms and signaling pathways of DEGs. Considering the results of GO function analysis, we linked the DEGs with mRNA catabolic process and cellular modified amino acid metabolic process, which are probably very important for the development process of human OS. As previous articles reported, our KEGG pathway analysis showed that hippo signaling pathway, mTOR signaling pathway, hedgehog signaling pathway, and others were among the most relevant pathways for OS. Zhou et al. found that the correlation between the mTOR/p70S6K signal transduction pathway in human OS and patients’ prognosis, and the overexpression of mTOR and p70S6K, is well correlated with tumor metastasis pattern, which might be an important mechanism responsible for the survival and proliferation of OS cells [
25]. Wang et al. identified that hippo/YAP signaling pathway not only is involved in tumorigenesis, but also hippo/YAP signaling pathway induces OS chemoresistance [
26]. Chai et al. deemed that the oncogenic activities in OS are mediated by TED1 through hippo–YAP1 signaling [
27]. Cheng et al. highlighted a new discovery that CNOT1–LMNA–Hedgehog signaling pathway axis exerts an oncogenic role in OS progression, which could be a potential target for gene therapy [
28]. Emerging data suggested that interference with hedgehog signaling signal transduction by inhibitors may reduce OS cell proliferation and tumor growth, thereby preventing osteosarcomagenesis [
29]. All these signaling pathways may play important roles in molecular mechanism of development process between SP cells and human OS.
Also of note is that there were numerous evidences for our DEGs of SP cells, which have proven to play important roles during OS tumorigenesis. The STRING database revealed top 20 high-degree hub nodes of DEGs including GAPDH, GART, FAU, HSPA8, EEF1A1, RPS3A, EIF4E, MAPK3, IL6, RPL6, eukaryotic translation initiation factor 3 beta (EIF3b), DEAD-box helicase 5 (DDX5), heat shock protein family D member 1 (HSPD1), ribosomal protein S29 (RPS29), ribosomal protein L18a (RPL18A), ribosomal protein L18 (RPL18), calmodulin 3 (CALM3), actin gamma 1 (ACTG1), ribosomal protein S27 (RPS27), and ribosomal protein L32 (RPL32). Furthermore, we analyzed the gene interaction network and top 10 modules using MCODE and found that ACTG1, eukaryotic translation initiation factor 3 subunit E (EIF3E), EIF4E, FAU, HSPD1, IL-6, KiSS-1 metastasis-suppressor (KISS1), PRIM1, pituitary tumor-transforming 1 (PTTG1), PRL32, S100 calcium-binding protein A8 (S100A8), S100 calcium-binding protein A9 (S100A9), serine hydroxymethyltransferase 1 (SHMT1), and TNF receptor-associated protein 1 (TRAP1) were the core interaction genes, which may be potential therapeutic targets for OS. Parts of them were in accord with STRING database results. Ajiro et al. found that serine/arginine-rich splicing factor 3 (SRSF3) regulates the expression of DDX5 in human OS U2OS cells [
30]. By participating in the transcriptional regulation of ribosomal protein L34 (RPL34) which plays an important role in the proliferation of OS cells, MYC interacts with the subunits of EIF3 and probably involves the translational control of growth-promoting proteins [
31]. EIF3, a multi-subunit complex, plays a critical role in translation initiation. Expression levels of EIF3 subunits are elevated or decreased in various cancers, suggesting a role for EIF3 in tumorigenesis [
32]. Choi et al. confirmed that EIF3b silencing could completely suppress cell growth in multiple OS cell lines [
33]. Osborne et al. also discovered that EIF4E is uniformly expressed in OS patient samples and it could be a relevant protein biomarker in OS [
34]. Rossman et al. found that overexpressing FAU itself is able to transform human osteogenic sarcoma cells to anchorage independence and make them easy to proliferate [
35]. Liang et al. proved that the expression of HSPD1 was high in OS tissues and cells; moreover, targeted inhibition of this gene could inhibit the proliferation of the tumor [
36]. Zhang et al. indicated that the decreased expression of KISS1 is correlated with distant metastasis of OS, and KISS1 may function as a tumor suppressor in OS cells through inhibition of the MAPK pathway [
37]. EEF1A1 is overexpressed in OS cell lines, and siRNA treatment against EEF1A1 produces a chemosensitization toward methotrexate, which showed that this gene is a potential therapeutic target of OS [
38]. Through ASK1/p38/AP-1 signal pathway, IL-6 occurs, which in turn results in the activations of vascular endothelial growth factor (VEGF) expression and contributing the angiogenesis of human OS cells [
39]. In addition, the ILK/Akt/AP-1 pathway is activated after IL-6 treatment, and IL-6 induces expression of ICAM-1 and migration activity of human OS cells [
40]. Yotov et al. proposed that PRIM1 is a major target of 12q13 amplifications, playing an essential role in tumorigenesis of human OS [
41]. PTTG1 siRNA markedly downregulates the expression of PTTGl protein in OS cells, leading to obvious inhibition of cell proliferation, alters cell cycle distribution, and reduces ability of invasion of OS cells [
42]. Tsai et al. deemed that expression stability of RPL32 is high in OS samples, and this gene could be a potential target [
43]. In Montesano’s study, the anti-apoptotic role of TRAP1 is confirmed in Saos-2 OS cells, which suggested that increased expression of this gene could make diethylmaleate-adapted and chemoresistant cells evade toxic effects of oxidants and anticancer drugs [
44]. Endo-Munoz et al. proved downregulation of S100A8 between chemo-naive OS biopsies and non-malignant bone biopsies, highlighting their potential as therapeutic targets for OS [
45]. Cheng et al. confirmed that through inactivating MAPK and NF-κB signaling pathways, downregulation of S100A9 could inhibit OS cell growth [
46]. Besides, Both et al. concluded that some genes, including SHMT1, are candidate oncogenes in 17p11.2–p12 of importance in OS tumorigenesis [
47]. Taken together, all these core genes discovered in OS SP cells by bioinformatics enrichment analysis and gene interaction network analysis may increase or decrease tumorigenicity and self-renewal capacity of OS SP cells; further, these changed SP cells could result in development process of human OS.
Lastly, there are several limitations of this study. It is acknowledged that predicting key genes merely by means of bioinformatics is not sufficient, and further molecular biological experiments such as the use of gene transfection/knockdown and quantitative real-time polymerase chain reaction are needed to confirm these results.