부산대학교 도서관

[eng] Neonatal arterial ischemic stroke (NAIS) is a cerebrovascular disease that shows a focal disruption of cerebral blood flow due to artery obstruction. Due to its early onset, it can cause long-lasting outcomes such as cerebral palsy, loss of neurologic function, physical language, and visual impairments, and behavioural problems. The genetics of many complex diseases such as stroke is far from being resolved. Additionally, several investigations have focused on children, young adults, and adults have attempted to identify the missing heritability without success. At time, the role of genetics as a determinant in NAIS has not been investigated. To decipher the genetics of idiopathic NAIS new approaches and strategies are needed to identify new candidate genes. Especially those focused on rare de novo variants. In this study we hypothesised that sporadic idiopathic NAIS in term newborns could be associated with de novo dominant variants in genes involved in relevant biological pathways of the pathophysiology of NAIS, such as endothelial rheology, coagulation, and cell adhesion. This based on the current evidence of de novo dominant mutations associated with sporadic cases of epileptic and developmental encephalopathies in newborns and infants. Furthermore, in case of not being able to demonstrate this hypothesis, we considered the autosomal recessive inheritance as a second hypothesis. To accomplish this, we applied whole-exome sequencing (WES) in trio (patient and its parents). This approach could allow us to identify a suitable candidate gene . Furthermore, because clinically homogeneous subgroups are assumed to be genetically homogeneous as well, we evaluated homogenous clinical subgroup of NAIS according to the arterial territory of the middle cerebral artery (MCA) in the stroke event. This to increase the power of identification of candidate genes and gene-disease association with the disease. In our study, 23 consecutive infants suffering from acute symptomatic idiopathic NAIS, and their parents, were prospectively recruited for WES-trio. We conducted a custom workflow analysis comprising variant annotation, filtering, pathogenicity impact prediction, and a knowledge-driven analysis (KDA) pipeline for scoring, ranking, and prioritization of the variants. Our results identified 28 de novo variants in 28 unique genes, meaning that the probands did not share neither common de novo variants nor genes. Moreover, our KDA pipeline identified the c.1292A>G (p.Gln431Arg) de novo dominant variant in PIK3CD as the strongest autosomal-dominant candidate gene for NAIS association. This variant was carried by a patient who suffered a perforant stroke. However, in terms of the autosomal recessive inheritance as a second hypothesis, no candidate genes were found. In silico analyses and functional studies of PIK3CDGln431Arg showed that this variant could affect the protein function due to the damaging predictions of the pathogenicity predictor tools, protein destabilization, and aberrant aggregations at the subcellular level. However, when studying its role in the PI3K/Akt pathway, a relevant cell- signalling pathway involved in stroke in which PIK3CD interacts, we found no significant changes in the expressions of total AKT and phosphorylated-AKT (Ser473). Although we have not found conclusive evidence that demonstrate the genetic cause of idiopathic NAIS, we cannot completely rule out that the genetic cause of the disease is not due to the presence of genetic variations. In conclusion, because of the pertinence of knowing the etiopathogenesis of this major neonatal disorder and molecular mechanisms involved, the development of precision medicine, and safer tailored therapies, more studies and approaches are needed. Specially to clarify the genetic cause of the disease in term newborns and whether PIK3CD could be associated with perforant stroke in NAIS.