IAs are potentially dangerous cerebrovascular diseases caused by multiple factors. The pathogenesis of IAs includes a variety of factors, such as smoking, genetics, injury, haemodynamics, environmental factors and inflammation. Inflammation plays an essential role in pathogenesis, and all of these factors can promote the expression of inflammatory factors and stimulate the occurrence and development of IAs27. The release and aggregation of MMPs, McP-1, TNF-α, ZO-1, serum amyloid A (SSA) 28 and other inflammatory cytokines can directly or indirectly promote the growth and rupture of aneurysms. Noncoding RNAs function through multiple mechanisms and are involved in vascular development, growth, and remodelling29. Therefore, the expression of inflammatory genes and multiple inflammatory factors are involved in the key processes of IA formation and rupture.
In this study, a total of 7 optimal inflammatory genes, C3AR1, MSR1, OLR1, OSM, RTP4, SERPINE1, and SLC11A2, were ultimately screened out and used to build the model. After PCR verification, significant expression differences were found in the IA wall and superficial temporal artery tissues. C3AR1 expression promotes vascular inflammatory infiltration, peripheral lymphocyte infiltration, and blood‒brain barrier (BBB) permeability30, which were also upregulated in another study on the pathogenesis of IAs31. MSR1 is expressed in macrophages, monocytes, neutrophils and other inflammatory cells and mediates inflammation through multiple pathways32,33. Gene polymorphism of OLR1 is closely related to blood glucose and lipid levels and the occurrence of cardiovascular and cerebrovascular diseases, and it also has a proinflammatory effect34,35. Oncostatin M (OSM) is a proinflammatory cytokine in the interleukin-6 (IL-6) family. OSM was also identified as a potential risk factor for IA rupture in a predictive model study36,37. RTP4 is involved in the early regulation of monocyte activation and the activation of inflammation38. SERPINE1 is activated by TGF-β1 and is involved in apoptosis and inflammatory injury. In a study of genes associated with hypertension and IAs, SERPINE1 also promoted the development of IAs39,40. Studies have shown that SLC11A2 is involved in inflammatory markers in the blood41. Therefore, these 7 inflammatory genes are correlated with the occurrence and development of IAs.
GO and KEGG pathway enrichment analyses showed that DEIRGs were mainly enriched in pathways related to inflammation regulation, atherosclerosis and infection. At present, the main recognized mechanisms of intracranial artery formation are as follows: 1. congenital factors; 2. arteriosclerosis; 3. infection; 4. injury; and 5. haemodynamic impact factors, which are common to the above different causes42. The process of vascular injury caused by arteriosclerosis, infection and trauma is mainly inflammatory infiltration, which is consistent with the pathways found to be enriched by GO and KEGG. Infiltration of inflammatory cells is a hallmark of IA, in which macrophages are more pronounced in ruptured IAs. T cells and mast cells are also important components in the occurrence and development of IA. The humoral immune response caused by inflammatory cells is also involved in the occurrence and development of IAs, especially the activation of the complement system. The release of various inflammatory cytokines may promote the occurrence of IA inflammatory cascade reactions and further promote the occurrence and development of IAs43. The lipopolysaccharide response, bacterial analysis response, positive regulation of leukocyte migration, Staphylococcus aureus infection, viral protein and cytokine receptor interaction and other pathways are associated with infection. After bacterial infection, the inflammatory response is obvious, which further promotes the occurrence and development of IA due to the aggregation and infiltration of a large number of inflammatory cells.
Single-gene GSEA enrichment analysis was performed for the diagnostic markers, and 15 hallmark items were simultaneously enriched in all diagnostic genes, mainly those associated with inflammation. C3AR1 is a protein-coupled receptor of C3A, which has been reported to be involved in gastric cancer, osteosarcoma, renal clear cell carcinoma, vascular inflammation and other diseases44–46. The expression of C3AR1 is significantly correlated with monocyte, macrophage, dendritic cell and T cell infiltration. C3AR1 is involved in activation of the complement system through the C3-C3A-C3AR1 pathway. C3AR1 also mediates vascular endothelial injury through this pathway, and enhanced C3a/C3aR signal transduction through endothelial cells can promote vascular inflammation and BBB dysfunction, which may also exist in IAs30. It has been reported that MSR1 is involved in bacterial, viral, and noninfectious inflammation and other diseases. It is highly expressed in macrophages, and the high expression of MSR1 indicates obvious inflammation32,33. It is involved in the inflammatory reactions of macrophages, monocytes, neutrophils and other inflammatory cells and may be involved in the inflammatory reactions of the IA wall. OLR1 is mainly involved in atherosclerosis, which plays an important role in the pathogenesis of IA34,35. OSM has been reported in breast cancer, intestinal inflammation, lupus nephritis, systemic sclerosis, etc47–50. Its main mechanism is to promote extracellular matrix remodelling. The formation of IAs involves two events: inflammatory cell infiltration and extracellular matrix remodelling. OSM may be involved in the remodelling process of the extracellular matrix of the IA wall, thus promoting the occurrence and development of IAs. RTP4, as a diagnostic gene marker in prostate cancer, viral infection, lupus nephritis, osteoarthritis and other diseases51,52, is mainly involved in the early activation of the inflammatory response mechanism, which may promote the occurrence and development of IA. SERPINE1 is expressed in gastric cancer, oral cancer, colon cancer, prostate cancer and other cancers53 and plays a role in the remodelling of the tumour microenvironment and immune cell invasion, promoting tumour occurrence and development. Studies have shown that SERPINE1 levels represent the progression of atherosclerosis. Similarly, SERPINE1 may be involved in the pathogenesis of the extracellular matrix in atherosclerotic plates and may be one of the mechanisms of IA formation54. Solute carrier family 11 member 2 (SLC11A2) transports ferrous iron and some divalent metal ions to the entire plasma membrane and across the endoplasmic membrane. Free iron is a powerful oxidizing molecule that is involved in oxidative stress, lipid peroxidation and endothelial dysfunction through its ability to generate free radicals55. The abnormal expression of SLC11A2 may lead to intima injury, promote atherosclerosis, and accelerate the occurrence and development of IAs.
In this study, we overlapped 964 DEGs for IAs and 200 IRGs to obtain 35 DEIRGs. Meanwhile, functional enrichment analysis showed that DEIRGs were significantly enriched in inflammatory response, immune receptor activity, lipid and atherosclerotic pathway regulation. Thirteen genes with an AUC greater than 0.85 were selected for possible inclusion in diagnostic models constructed with the RF algorithm, and 7 biomarkers (C3AR1, MSR1, OLR1, OSM, RTP4, SERPINE1, and SLC11A2) were retained. In addition, 22 drugs with potential efficacy against these biomarkers were predicted, including fluoxetine, aleplasinin and orlistat. Finally, qRT‒PCR results showed that the expression levels of 7 biomarkers in IA tissue were significantly higher than those in superficial temporal artery tissue. A previous model study screened for differential genes between ruptured and unruptured aneurysms to predict intracranial artery rupture. This study explored the genetic differences in ruptured IAs56. Our study further elucidates the pathogenesis of IAs from the perspective of inflammatory genes.
Although some meaningful results have been produced in this study, there are still some limitations: (1) This study was a retrospective study with a limited sample size and conducted with reference to public databases using bioinformatics techniques, and our results need to be verified in a larger number of clinical samples. (2) The GSE54083 dataset includes only 13 IA tissue samples (8 ruptured IA and 5 unruptured IA) and 10 control samples, with a small sample size and lack of control tissues matched to the IAs. (3) Although this information was collected for the qRT‒PCR samples, the dataset samples lack clinical follow-up information and cannot be further validated. Therefore, the impacts of rupture status, size, form, intraluminal thrombus, and location are not taken into account. We will continue to focus on biomarkers of inflammation and further explore biomarkers of the pathogenesis of IA-specific regulatory pathways and functions.