BAVM is an important cause of focal neurological impairment in young people due to intracerebral hemorrhage or epilepsy. Clinical decision-making usually needs to consider the risk of further hemorrhage and the risk of hemorrhage caused by the treatment of cerebral AVM. Prevention of hemorrhage or re-bleeding is still the main prevention and treatment goal of BAVM. Understanding the biological characteristics of the disease can help to identify new biomarkers, predict the future clinical progress of patients, and even promote the development of new treatment strategies. The genetic and biological mechanism of a disease can be determined by studying its genetic factors. Based on this principle, we have studied the genetic polymorphisms of candidate genes to determine whether they may be associated with susceptibility to BAVM and specific clinical features. Nowadays, candidate gene research has become one of the main methods to evaluate the genetic risk of BAVM. Relevant studies have shown that there are more than 900 genes associated with the occurrence of BAVM, among which the expression of more than 300 genes is increased and the expression of more than 500 genes is decreased [19, 20]. In a meta-analysis of a genome-wide association study involving 515 BAVM cases and 1191 control cases, no SNPs were found to be associated with BAVM in the replication cohort after multiple trials. At the same time, several potential candidate genes were selected. These potential candidate genes include ALK1, MMP-3, SP4, NOTCH4, Jag1, egfem1p, CDKAL1, and bnc2 . In this study, we examined whether single nucleotide sites in the NOTCH4 gene are potential genetic risk factors for AVM, AVM bleeding, or AVM associated epilepsy. Our results show that the NOTCH4 gene may play a role in AVM biology. First, we believe that the NOTCH4 gene polymorphism is associated with increased susceptibility to AVMs, so we can consider NOTCH4 as a highly reliable candidate gene. At the same time, our results are different from those reported abroad. The results of foreign studies on BAVM gene polymorphism showed that NOTCH4 SNP rs715299_ A, rs415929_ A, rs1109771_ G significantly increased the risk of BAVM, and rs443198_ TT and rs1109771_ GG has a certain correlation with hemorrhage and secondary epilepsy. Our study did not find genes associated with a higher risk of bleeding and epilepsy, which may be related to population differences.
The polymorphism of NOTCH4 is common in neuropsychiatric diseases, and clinical trials indicate that the most common are related to schizophrenia. A foreign study was conducted on the genotyping of eight polymorphisms in all subjects, and to investigate their correlation with clinical variables. The NOTCH4 gene rs367398 AA/Ag was significantly associated with worse positive and negative symptom scale and the clinical global impression score (CGI) . The results of the study on schizophrenia and NOTCH4 polymorphism in the Chinese Han population in South China also showed that the rs520688_GA-SNP genotype was significantly associated with decreased risk, and rs204993_ AA-SNP genotypes are associated with a higher risk of schizophrenia . In addition, some reports suggest that the Notch4 SNP is associated with lung cancer and inflammatory bowel disease. Based on the fact that haplotype-based markers can improve association efficiency and refine association signals, we estimated the risk of AVM and the polymorphism of the NOTCH4 gene. There is a single marker association between TT-SNP genotypes and the key clinical feature, which is bleeding. It is worth noting that the multiple regression analysis of rs4498_-TT is also a statistically significant predictor of hemorheology, and rs443198_-TT was not previously reported to be associated with bleeding in other diseases. This observation indicates that NOTCH4-SNP may play a special role in AVM related bleeding.
Amyere M sequenced the whole exon of the CM-AVM family and screened a candidate gene from a large number of patients. They found that the EphB4 gene mutation existed in these families, and the performance of these patients was similar to that of AVM caused by HHT, so they suggested that the EphB4–Ras–extracellular signal-regulated kinase (ERK) signaling pathway might be the main cause of AVMs. In recent years, somatic mutation has been a hot topic in AVM research. It has been reported that the KRAS mutation has been detected in endothelial cells from cerebral AVMs, and the expression of mutated KRAS in endothelial cells in vitro induces ERK. The increase of kinase activity increased the expression of genes related to angiogenesis and Notch signaling, and enhanced migration behavior . These processes were reversed by inhibiting MAPK and ERK signaling. They believe that the occurrence of these malformations is caused by KRAS induced activation of the MAPK-ERK signaling pathway in brain endothelial cells. In a study of brain and spinal cord AVMs and detectable tumor-associated somatic mutations, Tao et al. used deep next-generation sequencing of 422 common tumor genes to analyze tissue and paired blood samples. Liquid drop digital PCR was used to identify panel sequencing mutations and additional low mutation frequency relationships. Their results showed that the prevalence of KRAS/BRAF somatic mutations in cerebral and spinal AVM was as high as 87.1%, and there were no other tumor-related replication mutations. The homogeneity and high prevalence of this pathway indicated that a targeted therapy of RAS/RAF pathway inhibitors can be developed without the need for histological-genetic diagnosis .
In addition, the importance of SNPs in several genes related to angiogenesis and inflammation and their relationship with the development of AVM were also reviewed . The up-regulation of proinflammatory cytokines induces the overexpression of cell adhesion molecules in AVM endothelial cells, resulting in increased leukocyte recruitment. The increase of metalloproteinase-9 released by leukocytes can damage the vascular wall of BAVM and cause deformed ruptures. Inflammation also affects the proliferation, migration, and apoptosis of endothelial cells by up-regulating the expression of angiogenic factors, which is involved in the changes of the vascular architecture of AVM. The effect of inflammation on the pathogenesis of AVM is also enhanced by some SNPs in proinflammatory cytokine genes, thus increasing their protein level in AVM tissues. So far, many studies have investigated the possible relationship between gene polymorphism and the development of AVM. However, most of these studies focus on the genes that play a role in the inflammatory pathway, while the gene deficiency related to angiogenesis is ignored. At present, the genes related to angiogenesis include ALK1, ANGPTL4, and VEGF. Recently, a new pathway of cerebral AVM has been proposed. Notch signal is located downstream of AVM pathogenic genes such as ALK1 and VEGF. NOTCH4 affects the occurrence of AVM by regulating angiogenesis. Studies have shown that genes encoding angiopoietins (ANGPT1 and ANGPT2) and their receptors Tie-2 play an important role in angiogenesis and vascular stability. In AVM, the Tie-2–angiopoietin system is unbalanced . In addition, the strategy of blocking VEGF and Notch by inhibiting inflammation and angiogenesis has a certain effect on the experimental treatment of BAVM. The research on the participation of metalloproteinase-9 inhibitors in the inhibition of the Notch signal in BAVMs provides key data for the drug treatment of BAVMs, which provide a reference to design and test more specific and effective treatment strategies in the future .
In general, clinical research on SNPs requires a large number of subjects to reduce the sampling error in statistics. Unfortunately, only 150 BAVM patients are included in this study because of the low incidence rate. The sample size may reduce the difference of the queue substructure, leading to a false positive association and confounding effect, which makes the results less accurate. Therefore, our research results need to be supported by more data. In addition, BAVM is a complex genetic disease, and the results will be different due to ethnic differences compared with populations in other countries. Moreover, these gene loci lack functional research, so it is difficult to explore further. The occurrence of BAVM and bleeding is affected by both environmental and genetic factors. It is even less possible to accurately judge the influence of these gene loci if we exclude environmental factors and discuss genetic characteristics. This is undoubtedly a common defect in SNP type research. The goal of the future treatment for cerebral AVMs is to establish a disease quantitative model that can predict the occurrence and rupture risk of BAVM. Perhaps the most important gap in BAVM research hinders understanding of the disease because of the lack of a real experimental model. A reliable animal model is key to studying the mechanism of the disease and to test new therapies. At present, there is no real animal model of cerebral AVM. Some models have been proposed and used, but these models are only similar to some aspects of human brain AVMs, not all aspects [28–29]. Therefore, more advanced experimental techniques, new statistical models and animal models, and larger research sequences will contribute to the progress of this series of studies. In conclusion, our results have preliminarily confirmed the relationship between NOTCH4 polymorphism and cerebral AVM in a Chinese Han population. However, we have not yet explored its downstream regulatory mechanism, so more in-depth research is needed on how the gene affects the occurrence of BAVM. Our ultimate goal is to find a highly consistent pathway and develop targeted therapy for pathway inhibitors to prevent and treat early cerebral AVMs in asymptomatic individuals or adolescents.