Neuropathic pain is a debilitating condition caused by damage to the nervous system and chronic disease. As a kind of non-coding RNA, lncRNA-related research has become a hot spot in genetic research and is involved in many life activities such as epigenetic regulation, cell cycle regulation, and cell differentiation. Several lncRNAs have been reported to be involved in various diseases, including Alzheimer's disease(AD) (15), Parkinson's disease (PD) (16), autoimmune thyroid disease (AITD) (17), and cancers (18, 19). An miRNA is a short non-coding single-stranded RNA molecule; it participates in the regulation of post-transcriptional gene expression in animals and plants (20, 21). At present, many miRNAs have been reported in neurodegenerative diseases (22), cancer (23, 24), bowel disease (25), and diabetic heart disease (DHD)(26). Although some lncRNAs (27, 28) and miRNAs (29, 30) have been reported in NP, there are still few reports on how lncRNAs, miRNAs and mRNAs jointly regulate the pathogenesis of NP. Zhou and colleagues (5) analyzed the expression profile of non-coding RNAs in the spinal cord following spared nerve injury-induced NP by sequencing, and constructed lncRNA-miRNA-mRNA and circRNA-miRNA-mRNA networks in NP. However, they only analyzed 12 samples, which may have some limitations. In order to solve the bias that may be caused by the small sample size, different datasets were collected in this paper. Our research used three datasets from the GEO database, including 76 samples, which can explain the pathological mechanism of NP more comprehensively.
At present, the pathological mechanism of NP is still unknown. The differential expressed mRNAs can cause a variety of diseases. We found several differentially expressed mRNAs in GSE24982. The GO and KEGG enrichment analysis of these differentially expressed mRNAs can help us reveal their functional role. Here, the most significant biological processes were blood circulation, regulation of membrane potential, metal ion transmembrane transporter activity, synaptic membrane, and postsynaptic specialization. The enrichment of differentially expressed mRNAs in blood circulations was supported by a previous study, indicating that partial sciatic nerve ligation (PSL)-induced neuropathic pain can upregulate the synthesis of hypothalamic Oxytocin (OXT) and transport it to the OXT axon terminal through the systemic blood circulation (31). The detection and processing of pain by afferent sensory neurons depends on different types of voltage and ligand-gated ion channels, including sodium, calcium, and TRP pathways. This study found that differentially expressed mRNAs clustered in regulation of membrane potential and metal ion transmembrane transport pathway, suggesting that the expression and long-term changes of ion channels may be closely related to chronic pain states(32). Additionally, we found that the differentially expressed mRNAs were related to the synaptic membrane and postsynaptic specialization, indicating that the formation and regulation of synapses may be associated with neuralgia, which is consistent with reports in a previous study(33). We found that the main pathways for these differentially expressed mRNAs in NP were neuroactive ligand-receptor interaction, PI3K-Akt signaling pathway, MAPK signaling pathway, cAMP signaling pathway, and Calcium signaling pathway. A previous study reported that jct-801 relieves paclitaxel-induced neuropathic pain through the PI3K/Akt pathway(34). Our study suggested that differentially expressed mRNAs are involved in NP possibly through regulating the MAPK signaling pathway, which is consistent with a previous study (35). The Cyclic Adenosine Monophosphate (cAMP) signaling pathway is a key contributor to the development of chronic pain, and an existing study indicated that knockdown of the cAMP effector can relieve the pain-like responses in chronic pain models(36). Therefore, our study provided a reference for the molecular explanation of the NP pathology mechanism.
LncRNA, miRNA, and mRNA often coordinate the occurrence and prognosis of diseases in organisms. Up to date, only one study analyzed the lncRNA-mRNA-miRNA network in NP (5). The authors (5) found that the PI3K-Akt signaling pathway may be related to NP, which is consistent with our study. Though their study gave insights into the regulatory mechanisms of NP, their study may have some limitations due to the limited sample size. The accumulation of transcriptome data in public databases can help overcome these limitations and allow the discovery of more insightful information. Herein, we constructed the lncRNA-mRNA-miRNA network in NP. The lncRNAs can be used as candidate indicators for NP (30). Various studies have provided new mechanisms for the molecular roles of miRNAs in the pathogenesis of NP (37-39). Several vital lncRNAs were found in the present NP lncRNA-miRNA-mRNA network, and these lncRNAs have not been reported in NP. For example, lncRNA SCAMP1 played an important role in the regulatory network of this study, and it had been reported in a previous study that it may be associated with malignant biological behavior (40). In addition, lncRNA SCAMP1 discovered in the present study were related to the regulation of cancer (41). The identified differentially expressed miRNAs have not been reported in previous NP-related studies. For the first time, we reported that miR-940 and miR-2277-3p may play an important role in NP pathology based on the regulatory network of NP, while these miRNAs were previously reported mainly in cancer research (42-44). Additionally, miR-5689 was identified in the regulatory network of NP and it was reported as a biomarker for predicting the development of new distant metastasis (45). We have found some differentially expressed mRNAs that may be involved in the NP regulatory network. However, few studies have reported the functions of these genes in NP. Here, we believe that IQGAP1 may play an important role in NP pathogenesis, which may explain the significant correlation between chronic pain at 3 months after surgery (46). In addition, in previous studies, BTAF1 has been reported to be related to immunity (47) whereas LRPAP1 and MXRA7 are considered to be related to the tumorigenesis (48, 49). Compared with previous works, we identified a larger number of lncRNAs, miRNAs, and mRNAs, which has enriched the our knowledge in this field and provided a new insight into the pathogenetic mechanism and the identification of targeted drugs. In short, we have discovered for the first time some lncRNAs, miRNAs and mRNAs that may be involved in NP, but their specific function in NP needs more validation studies.
To further explore the biological function of the lncRNA-miRNA-mRNA network, we performed the GO and KEGG pathway analysis. We found that the mRNAs in the network were involved in the mRNA processing, response to peptide, positive regulation of cell cycle, GTPase, chromatin, cell-substrate junction, neuron spine, and neuron projection cytoplasm. Through the KEGG pathway analysis, we found that the mRNAs in the network were enriched in the pathway of the MAPK signaling pathway, proteoglycans in cancer, cell cycle, neurotrophin signaling pathway, and platelet activation. Our research suggested that the lncRNA-miRNA-mRNA network may participate in the regulation of NP through GTPase. Several studies have reported that GTPase plays a key regulatory role in NP(50, 51). For example, a study reported that spinal Rheb-mTOR signaling plays an important role in the regulation of NP after spinal cord sensitization (52). The lncRNA-miRNA-mRNA network may regulate the occurrence of NP by the cell cycle. Studies have reported that the cell cycle is activated in spinal cord injury (SCI), which may regulate chronic pain after SCI (53, 54). We found that the mRNAs in the network were enriched in cell cycle and chromatin, suggesting that the regulatory network may be involved in the pathway of NP through regulating the cell cycle. In addition, we found that platelet activation was enriched in the regulatory network, which was consistent with a previous study that blockade of the platelet-activating factor-pain loop can relieve neuropathic pain (55). The GO and KEGG analysis of lncRNA-miRNA-mRNA sub-network suggested that the mRNAs in the sub-network may participate in the pathway of NP by cell cycle G1/S phase transition, DNA damage checkpoint, and the MAPK signaling pathway. The cell cycle G1/S phase transition, DNA damage checkpoint were vital for cell cycle, indicating that the lncRNA-miRNA-mRNA sub-network may regulate the NP pathway through cell cycle.
Neuropathic pain is a chronic disease, and there is still no effective treatment drug. DGIdb database can infer the targeted drugs of genes in diseases based on existing resources. The drugs included in this database contain the US Food and Drug Administration (FDA) certified drugs, which will provide more professional and reliable assurance for target drug prediction. For example, Nambou and Anakpa (56) recently discovered nicotinamide adenine dinucleotide (NAD) and CHEMBL1161866 with potential therapeutic value for corona virus disease 2019 (COVID-19) through the DGIdb database, which provided novel insight for the treatment of COVID-19. In addition, Yang and colleagues predicted candidate drugs for lung adenocarcinoma (LUAD) via the DGIdb database as well (57). Here, we predicted a series of dugs related to NP treatment based on mRNAs in the regulatory networks of NP. It is reported that inhibition of spleen tyrosine kinase (Syk) can alleviate mechanical allodynia (58). Fostamatinib is an oral Syk inhibitor that has been approved by the FDA for the adult treatment of Chronic immune thrombocytopenia (ITP)(59). Although there are differences between the pathological processes of mechanical pain and neuropathic pain, the number of genes targeted by this drug is high (66 target genes), so we recommend that doctors try fostamatinib in patients with NP to see its potential therapeutic effects. Dexamethasone is a synthetic glucocorticoid with anti-inflammatory activity and minimal glucocorticoid effect, which is widely used in the treatment of various inflammation disorders (60). It has been reported that low doses of ibuprofen and dexamethasone have a synergistic therapeutic effect on trigeminal neuropathic pain in rats, and can significantly inhibit mechanical allodynia (61). Dexamethasone is effective in the treatment of NP caused by tumor-related spinal cord compression, and pregabalin is effective for malignant painful radiculopathy (62). Nevertheless, the long-term use of dexamethasone also has certain side effects (63). In the past, opioids such as morphine were often ineffective in the treatment of neuropathic pain, but a study showed that the combination of imatinib and morphine can completely relieve pain (64), which provides more treatment options for patients. A recent study reported that in rats of infraorbital nerve ligation trigeminal neuralgia, c-Abl expression was significantly increased, and the downstream activation product p38 was also abnormally activated. Interestingly, imatinib mesylate (STI571), a specific c-Abl family kinase inhibitor, can reduce the expression of p38 and reduce the loss of dopaminergic neurons, suggesting that imatinib may alleviate the symptoms of NP through the c-abl-p38 signaling pathway (65). Additionally, our study predicted the treatment ability of progesterone (PROG) in NP and was supported by a previous study, demonstrating that PROG may provide new strategies for the treatment of NP(66). NP caused by chemotherapy can reduce the prognosis of patients with the quality of life. Tamoxifen can alleviate NP induced by paclitaxel, vincristine, and bortezomib in chemotherapy through inhibiting protein kinase C/extracellular signal-regulated kinase pathway(67). Although we have only discussed the therapeutic effects of the drugs above, we cannot ignore the potential effects of other drugs that have not been mentioned. These predicted drugs can be verified in future studies, either individually or in combination. The number of genes corresponding to the targeting drugs was small, indicating that the therapeutic effect of these drugs may still be relatively weak. Therefore, we suggest doctors consider both the efficacy and side effects of drugs during the treatment.
Although this study found several lncRNAs, miRNAs, and mRNAs that may be related to NP through the public datasets, and analyzed the pathways that may participate in the pathological mechanism of NP through GO and KEGG pathway analysis, our research still has some limitations. Circular RNAs (circRNAs) are non-coding RNA molecules that have recently been shown to regulate the function of miRNAs and affect a variety of complex human diseases(68-70). Although there have been some studies on the circRNA regulation in NP (71-73), the present public databases still do not include the circRNA expression data of NP. Therefore, we used microarray sequencing data, including the sequencing results of lncRNA, miRNA, and mRNA in NP patients, which may not fully reflect the pathological mechanism of NP. This study found some lncRNAs, miRNAs and mRNAs that may be related to NP, but their relationship with NP is still unclear. We predicted several drugs that may have therapeutic effects on NP, but the effects of these drugs in clinical treatment have not yet been reported. In the future, we will collect samples from human NP patients, and obtain lncRNA, miRNA, mRNA, and circRNA data of NP patients through high-throughput sequencing technology. We plan to verify the functions of the genes and drugs identified here through more in vitro and in vivo experiments and strive to further reveal the pathological mechanism of NP.