Pathogenesis Of Intracranial Aneurysms Research Articles

Berberine Attenuates Macrophages Infiltration in Intracranial Aneurysms Potentially Through FAK/Grp78/UPR Axis.

Background: Inflammatory cells, such as macrophages, play key roles in the pathogenesis of intracranial aneurysms (IAs). Berberine (BBR), an active component of a Chinese herb Coptis chinensis French, has been shown to have anti-inflammatory properties through suppressing macrophage migration in various inflammation animal model. The goal of this study was to examine BBR’s effect on inflammation and IAs formation in a rodent aneurysm model.Methods and Results: Human aneurysm tissues were collected by microsurgical clipping and immunostained for phospho-Focal adhesion kinase (FAK) and CD68+ macrophages. A rodent aneurysm model was induced in 5-week-old male Sprague Dawley (SD) rats by intracranial surgery, then these rats were orally administrated 200 mg/kg/day BBR for 35 days. Immunostaining data showed that BBR inhibited CD68+ macrophages accumulation in IAs tissues and suppressed FAK phosphorylation. In lipopolysaccharide (LPS)-stimulated RAW264.7 cells, BBR treatment remarkably attenuated macrophages infiltration, suppressed the expression of matrix metalloproteinases (MMPs), and reduced proinflammatory cytokine secretion, including MCP-1, interleukin 1β (IL-1β), interleukin 6 (IL-6) and tumor necrosis factor-a (TNF-α). Mechanistically, BBR downregulated FAK/Grp78/Unfolded Protein Response (UPR) signaling pathway in RAW264.7 cells.Conclusion: BBR prevents IAs formation potentially through inhibiting FAK phosphorylation and inactivating UPR pathway in macrophages, which causes less macrophage infiltration and reduced proinflammatory cytokine release.

TLR4 gene polymorphisms rs11536889 is associated with intracranial aneurysm susceptibility

Intracranial aneurysm (IA) is a common lesion which often present asymptomatic until the time of rupture and result in subarachnoid hemorrhage (SAH). The pathogenesis of IA formation is complex and is influenced by both genetic and environmental risk factors. For exploring the detailed molecular and cellular mechanisms involved in the pathogenesis of IA, recent studies indicated inflammatory pathways and their genetic variants may as potential biomarkers. In this study, functionally relevant polymorphisms in the toll-like receptor 4 (TLR4) were screened in 330 IA patients and 313 controls from a Han Chinese population. Eight single nucleotide gene polymorphisms (SNPs) genotyped by the Improved Multiple Ligase Detection Reaction (iMLDR) method. Our results indicated that the presence of the minor allele (C) of the TLR4 SNP rs11536889 was associated with a decreased risk of IA (C vs. G, OR = 0.731; 95% CI 0.567–0.943; P = 0.017). This association was also present at the genotype level in a codominant model (GC vs. GG, OR = 0.447; 95% CI 0.226–0.884; P = 0.020) and a recessive model (CC vs. GG + GC, OR = 0.489; 95% CI 0.250–0.955; P = 0.035). In summary, we firstly found that the TLR4 SNP rs11536889 was significantly associated with the susceptibility of IA. Our results indicated TLR4 SNP rs11536889 may be a marker for IA risk, though the exact functional roles of TLR4 SNP rs11536889 in IA formation are still not very clear.

Correlating Clinical Risk Factors and Histological Features in Ruptured and Unruptured Human Intracranial Aneurysms: The Swiss AneuX Study

Pathogenesis of intracranial aneurysm is complex and the precise biomechanical processes leading to their rupture are uncertain. The goal of our study was to characterize the aneurysmal wall histologically and to correlate histological characteristics with clinical and radiological factors used to estimate the risk of rupture. A new biobank of aneurysm domes resected at the Geneva University Hospitals (Switzerland) was used. Histological analysis revealed that unruptured aneurysms have a higher smooth muscle cell (SMC) content and a lower macrophage content than ruptured domes. These differences were associated with more collagen in unruptured samples, whereas the elastin content was not affected. Collagen content and type distribution were different between thick and thin walls of unruptured aneurysms. Classification of aneurysm domes based on histological characteristics showed that unruptured samples present organized wall rich in endothelial and SMCs compared with ruptured samples. Finally, aneurysm wall composition was altered in unruptured domes of patients presenting specific clinical factors used to predict rupture such as large dome diameter, dome irregularities, and smoking. Our study shows that the wall of aneurysm suspected to be at risk for rupture undergoes structural alterations relatively well associated with clinical and radiological factors currently used to predict this risk.

NF-κB-Mediated Inflammation in the Pathogenesis of Intracranial Aneurysm and Subarachnoid Hemorrhage. Does Autophagy Play a Role?

The rupture of saccular intracranial aneurysms (IA) is the commonest cause of non-traumatic subarachnoid hemorrhage (SAH)—the most serious form of stroke with a high mortality rate. Aneurysm walls are usually characterized by an active inflammatory response, and NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) has been identified as the main transcription factor regulating the induction of inflammation-related genes in IA lesions. This transcription factor has also been related to IA rupture and resulting SAH. We and others have shown that autophagy interacts with inflammation in many diseases, but there is no information of such interplay in IA. Moreover, NF-κB, which is a pivotal factor controlling inflammation, is regulated by autophagy-related proteins, and autophagy is regulated by NF-κB signaling. It was also shown that autophagy mediates the normal functioning of vessels, so its disturbance can be associated with vessel-related disorders. Early brain injury, delayed brain injury, and associated cerebral vasospasm are among the most serious consequences of IA rupture and are associated with impaired function of the autophagy–lysosomal system. Further studies on the role of the interplay between autophagy and NF-κB-mediated inflammation in IA can help to better understand IA pathogenesis and to identify IA patients with an increased SAH risk.

Genome-wide association analysis identifies new candidate risk loci for familial intracranial aneurysm in the French-Canadian population

Intracranial Aneurysm (IA) is a common disease with a worldwide prevalence of 1–3%. In the French-Canadian (FC) population, where there is an important founder effect, the incidence of IA is higher and is frequently seen in families. In this study, we genotyped a cohort of 257 mostly familial FC IA patients and 1,992 FC controls using the Illumina NeuroX SNP-chip. The most strongly associated loci were tested in 34 Inuit IA families and in 32 FC IA patients and 106 FC controls that had been exome sequenced (WES). After imputation, one locus at 3p14.2 (FHIT, rs1554600, p = 4.66 × 10–9) reached a genome-wide significant level of association and a subsequent validation in Nunavik Inuit cohort further confirmed the significance of the FHIT variant association (rs780365, FBAT-O, p = 0.002839). Additionally, among the other promising loci (p < 5 × 10−6), the one at 3q13.2 (rs78125721, p = 4.77 × 10−7), which encompasses CCDC80, also showed an increased mutation burden in the WES data (CCDC80, SKAT-O, p = 0.0005). In this study, we identified two new potential IA loci in the FC population: FHIT, which is significantly associated with hypertensive IA, and CCDC80, which has potential genetic and functional relevance to IA pathogenesis, providing evidence on the additional risk loci for familial IA. We also replicated the previous IA GWAS risk locus 18q11.2, and suggested a potential locus at 8p23.1 that warrants further study.

Genetics of Intracranial Aneurysms.

Intracranial aneurysms (IAs) are localized dilatations of intracranial arteries that are because of weaknesses of the endothelial layer. These dilations vary in size (small aneurysms, 25 mm) and are classified according to their shapes: either localized sac-like pouches or long dilatations that increase the vessels diameter; these shapes are referred to, respectively, as saccular or berry aneurysms and fusiform aneurysms. Most IAs are saccular and typically occur at the arterial branching sites of the circle of Willis. Common sites of saccular IAs include the anterior communicating artery, the posterior communicating artery, internal carotid artery, the middle cerebral artery, and the basilar artery bifurcation. Although the exact pathogenesis of IA remains to be established, several hypotheses have highlighted the contribution of maladaptive vascular remodeling triggered by hemodynamic stress and inflammatory response1—a chain of events that would ultimately damage blood vessel walls and lead to IA. The worldwide prevalence of IA is estimated to be ≈3.2%.2 Subarachnoid hemorrhage (SAH) accounts for ≈5% of all strokes, and 85% of SAHs are because of aneurysmal ruptures. SAH has an incidence rate of 9 per 10 000 persons per year and a fatality rate ≈50%.3 The incidence of SAH is markedly higher in the Finnish and Japanese populations, but surprisingly, this is not because of the prevalence of IA in these populations.2 The most common clinical symptoms of aneurysmal SAH are a sudden onset of severe headache with stiff neck, vomiting, and photophobia. Unruptured IAs are usually asymptomatic and are identified through the screening of high-risk individuals or as incidental findings of magnetic resonance imaging, particularly magnetic resonance angiogram, or computerized tomographic studies. The diagnosis of IA is made by magnetic resonance angiogram, computerized tomographic angiography, or classical angiography. The diagnosis of …

Association of SOX17 Gene Polymorphisms and Intracranial Aneurysm: A Case-Control Study and Meta-Analysis.

Genome-wide association studies have revealed an association between SRY-box 17 (SOX17) gene and intracranial aneurysm (IA) formation. However, results were mainly derived from European and Japanese populations. We investigated the association between SOX17 gene polymorphisms and IA in a homogeneous Korean population. We performed a meta-analysis to assess these results in East-Asian populations. This cross-sectional study included 187 age- and sex-matched patients with IA and 372 control subjects. Genetic association analysis was performed in the generalized linear model to identify associations between 4single nucleotide polymorphisms and IA, including 95patients with ruptured aneurysms and 92 with unruptured aneurysms. The East-Asian meta-analysis of 5100 IA cases and 7930 control cases was conducted under an inverse variance model. Among 4 single nucleotide polymorphisms that passed quality control tests, the minor C allele of rs1072737 was significantly associated with IA (odds ratio 0.69, 95% confidence interval 0.49-0.96, P= 0.03). None of the 4 single nucleotide polymorphisms showed a significant association between patients with ruptured and unruptured aneurysms. Meta-analysis revealed that Galleles of rs10958409 and rs9298506 were significantly associated with IA in the East-Asian population after removing study heterogeneity (odds ratio 1.11, 95% confidence interval 1.04-1.19, P= 0.0023 and odds ratio 1.19, 95% confidence interval 1.07-1.32, P= 0.0016). Identification of genetic variants located near SOX17 is likely to be clinically significant for IA formation. rs10958409 and rs9298506 may increase risk of IA in East-Asian populations. Our findings may help in the identification of IA pathogenesis.

Genetic Risk Assessment of Elastin Gene Polymorphisms with Intracranial Aneurysm in Koreans

Elastin encoded by elastin gene (ELN) is a crucial extracellular matrix protein responsible for arterial resilience. The objective of this study was to identify single nucleotide polymorphisms (SNPs) of ELN gene susceptible to intracranial aneurysm (IA) in Korean population. Two SNPs of ELN gene, rs2071307 (Gly422Ser) and rs2856728 (intron), were genotyped in 90 patients with IA and 90 age and frequency matched controls. Fisher’s exact test was conducted to evaluate allelic association with IA. Of the two SNPs in ELN gene, T allele of rs2856728 (intron) showed statistically significant association with increased development of IA (odds ratio [OR]: 2.34, 95% confidence interval [CI]: 1.44–3.81, P = 7.6 × 10−4). However, G allele of rs2071307 (Gly422Ser) had no significant association with the development of IA (OR: 1.27, 95% CI: 1.44–3.81, P = 0.607). Interestingly, the odds of having rs2856728 variant was approximately 2-fold higher in males than that in females (OR: 3.46 vs. 1.88, P < 0.05). However, none of SNPs showed difference between single and multiple IA in this study. This preliminary study implies that the rs2856728 variant in ELN gene polymorphisms might play crucial roles in the development and pathogenesis of IA in Korean population.

Analysis of gene expression in intracranial aneurysms

BackgroundDifferent gene expression profiles are observed in intracranial aneurysm tissue. Understanding these genes and what regulates their expression will help us to understand intracranial aneurysm pathogenesis. We investigated whether differences in gene expression in intracranial aneurysms.MethodsSixteen intracranial aneurysm tissues were compared with 16 matched samples from the superficial temporal artery as controls. We detected the gene expression profiles in these samples with the Human U133 Plus 2.0 GeneChip.ResultsA total of 2142 differentially expressed gene transcripts were detected based on the gene expression profile. Verification analysis showed that the VCAM1, MAGI2, PPP2R2B, PPP2R3A genes were associated with the occurrence and development of intracranial aneurysm. These genes mainly encode cell adhesion molecules (CAMs) and ERK/JNK signaling pathways.ConclusionChanges of genes expression involved in immune and inflammatory reactions, cell adhesion molecules may be associated with the development of aneurysms.

Comparative transcriptome analysis reveals involvement of TLR-2 signaling in the pathogenesis of intracranial aneurysm

In this study, we collected and analyzed 15 aneurysmal and 17 superficial temporal artery (STA) specimens from 32 Chinese patients with intracranial aneurysm. Total RNA was extracted and reverse transcribed to cDNA, and genome-wide expression profiling was performed by using the Affymetrix Human Genome U133 Plus 2.0 Array which allows a total number of 38,500 genes to be analyzed at the same time. Real-time RT-PCR was performed to verify the expression level of 8 selected genes. We found significant up-regulation of the TLR-2 gene. This result suggests that TLR-2 plays a key role in the formation of intracranial aneurysm in a Chinese population. To our knowledge, this study is the first to use the Human Genome U133 Plus 2.0 Array to analyze the gene expression profiles in Chinese patients with intracranial aneurysm.

Association of TNF-α-3959T/C Gene Polymorphisms in the Chinese Population with Intracranial Aneurysms.

Recent studies have demonstrated that cytokines play an important role in the pathogenesis of intracranial aneurysm (IA). Tumor necrosis factor-α (TNF-α) is an important proinflammatory cytokine, which was shown to influence the development of IA, but there is no research data from China. Hence, the purpose of this study was to explore the relationship between TNF-α polymorphisms and IA in China. The association of genetic variants of TNF-α gene expression was investigated in a Chinese population with IA. The TNF-α-3959T>C(rs1799964), 4127C>A(rs1800630), 4133C>T(rs1799724), 4184C>T(rs4248158), and 4752G>A(rs361525) gene polymorphisms in 192 IA cases and 112 controls were analyzed using polymerase chain reaction (PCR). Differences in genotype and allele frequencies between patients and controls were tested. There were no significant differences in 4127C>A (p=0.072), 4133C>T (p=0.373), 4184C>T (p=0.749), and 4752G>A (p=0.184) genotype frequencies between the IA group and the control group. But this case-control association study revealed that TNF-α-3959T>C (p<0.001) was significantly associated with increased risk of IA. These results suggested that a novel TNF-α locus was found to be closely correlated with the occurrence of IA in Chinese.

Screening of Critical Genes and MicroRNAs in Blood Samples of Patients with Ruptured Intracranial Aneurysms by Bioinformatic Analysis of Gene Expression Data.

BackgroundThis study aimed to identify more potential genes and miRNAs associated with the pathogenesis of intracranial aneurysms (IAs).Material/MethodsThe dataset of GSE36791 (accession number) was downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) were screened for in the blood samples from patients with ruptured IAs and controls, followed by functional and pathway enrichment analyses. In addition, gene co-expression network was constructed and significant modules were extracted from the network by WGCNA R package. Screening for miRNAs that could regulate DEGs in the modules was performed and an analysis of regulatory relationships was conducted.ResultsA total of 304 DEGs (167 up-regulated and 137 down-regulated genes) were screened for in blood samples from patients with ruptured IAs compared with those from controls. Functional enrichment analysis showed that the up-regulated genes were mainly associated with immune response and the down-regulated DEGs were mainly concerned with the structure of ribosome and translation. Besides, six functional modules were significantly identified, including four modules enriched by up-regulated genes and two modules enriched by down-regulated genes. Thereinto, the blue, yellow, and turquoise modules of up-regulated genes were all linked with immune response. Additionally, 16 miRNAs were predicted to regulate DEGs in the three modules associated with immune response, such as hsa-miR-1304, hsa-miR-33b, hsa-miR-125b, and hsa-miR-125a-5p.ConclusionsSeveral genes and miRNAs (such as miR-1304, miR-33b, IRS2 and KCNJ2) may take part in the pathogenesis of IAs.

Correction to: THSD1 (Thrombospondin Type 1 Domain Containing Protein 1) Mutation in the Pathogenesis of Intracranial Aneurysm and Subarachnoid Hemorrhage

In the article by Santiago-Sim et al, “ THSD1 (Thrombospondin Type 1 Domain Containing Protein 1) …

Dipeptidyl Peptidase-4 Inhibitor Anagliptin Prevents Intracranial Aneurysm Growth by Suppressing Macrophage Infiltration and Activation.

BackgroundChronic inflammation plays a key role in the pathogenesis of intracranial aneurysms (IAs). DPP‐4 (dipeptidyl peptidase‐4) inhibitors have anti‐inflammatory effects, including suppressing macrophage infiltration, in various inflammatory models. We examined whether a DPP‐4 inhibitor, anagliptin, could suppress the growth of IAs in a rodent aneurysm model.Methods and Results IAs were surgically induced in 7‐week‐old male Sprague Dawley rats, followed by oral administration of 300 mg/kg anagliptin. We measured the morphologic parameters of aneurysms over time and their local inflammatory responses. To investigate the molecular mechanisms, we used lipopolysaccharide‐treated RAW264.7 macrophages. In the anagliptin‐treated group, aneurysms were significantly smaller 2 to 4 weeks after IA induction. Anagliptin inhibited the accumulation of macrophages in IAs, reduced the expression of MCP‐1 (monocyte chemotactic protein 1), and suppressed the phosphorylation of p65. In lipopolysaccharide‐stimulated RAW264.7 cells, anagliptin treatment significantly reduced the production of tumor necrosis factor α, MCP‐1, and IL‐6 (interleukin 6) independent of GLP‐1 (glucagon‐like peptide 1), the key mediator in the antidiabetic effects of DPP‐4 inhibitors. Notably, anagliptin activated ERK5 (extracellular signal–regulated kinase 5), which mediates the anti‐inflammatory effects of statins, in RAW264.7 macrophages. Preadministration with an ERK5 inhibitor blocked the inhibitory effect of anagliptin on MCP‐1 and IL‐6 expression. Accordingly, the ERK5 inhibitor also counteracted the suppression of p65 phosphorylation in vitro.ConclusionsA DPP‐4 inhibitor, anagliptin, prevents the growth of IAs via its anti‐inflammatory effects on macrophages.

DNA Methylation Regulates Gene Expression in Intracranial Aneurysms.

Different gene expression profiles are observed in intracranial aneurysm tissues. Understanding these genes and what regulates their expression will provide insight into the pathogenesis of intracranial aneurysms. We investigated whether differences in DNA methylation regulate gene expression in intracranial aneurysms. We compared 20 intracranial aneurysm tissue specimens with 20 matched specimens from the superficial temporal artery as controls. We identified the gene expression profiles in these samples using the GeneChip Human U133 Plus 2.0 array and evaluated DNA methylation modifications using the Infinium HumanMethylation450 BeadChip Kit. A total of 11,022 differentially methylated sites between aneurysm tissues and matched control tissues were identified, and 2142 differentially expressed gene transcripts were detected based on the 2 gene expression profiles. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses and verification analysis showed that the MYH11, LIFR, and TLR4 genes were associated with the occurrence and development of intracranial aneurysms. These genes mainly encode cell adhesion molecules or are involved in the NF-κB, JAK-STAT, and ERK/JNK signaling pathways. In the development of intracranial aneurysms, DNA methylation plays an important role in the regulation of genetic expression involved in immune and inflammatory reactions, cell function, cell maintenance, and cell signal transduction.

Abstract 195: Sulforaphane Protects Against Subarachnoid Hemorrhage and Alters the Expression of Inflammatory and Extracellular Matrix Remodeling Genes in a Mouse Model of Intracranial Aneurysm

Introduction: Oxidative stress has been shown to play an important role in the pathogenesis of intracranial aneurysm (IA). Sulforaphane is an isothiocyanate that exhibits antioxidant properties via the Nrf2 transcription factor. We tested whether sulforaphane would protect against IA formation and subarachnoid hemorrhage (SAH) in a mouse model. Methods: C57BL/6 mice were treated intraperitoneally with either 0.5mg/kg/day of sulforaphane (treatment group, n = 32) or a vehicle solution (control group, n = 26) prior to being subjected to a well-established IA induction protocol involving unilateral nephrectomy, mineralocorticoid administration, and intracranial injection of elastase. Sulforaphane/vehicle treatments were begun seven days prior to elastase injection, and continued daily for the duration of the experiment. Animals were followed for 14 days until sacrifice, with measurement of the number of aneurysms formed and ruptured in each animal. Intracranial vessel tissue was collected, pooled, and analyzed using real-time PCR (RT-PCR) with experiments performed in triplicate (n = 3). Results: There was a lower incidence of IA in the treatment group (1.06 +/- 0.24 per animal) than in the control group (1.54 +/- 0.33), but this trend did not reach significance. The incidence of SAH in the treatment group (0.53 +/- 0.09 per animal) was approximately 50% lower than in the control group (1.08 +/- 0.21, p = 0.012). RT-PCR showed significant (p &lt; 0.05) differences in the expression of interferon gamma, matrix metalloproteinase 9, collagen Ia2, interleukin-6, myosin heavy chain, and SM22 between the treatment and control groups. Conclusions: Treatment with sulforaphane resulted in a significantly lower incidence of SAH in a mouse model of IA. The expression of genes involved in inflammation and extracellular matrix remodeling were altered by sulforaphane administration.

Global Gene Expression Patterns and Somatic Mutations in Sporadic Intracranial Aneurysms.

High-throughput sequencing technologies can expand our understanding of the pathologic basis of intracranial aneurysms (IAs). Our study was aimed to decipher the gene expression signature and genetic factors associated with IAs. We determined the gene expression levels of 3 cases of IAs by RNA sequencing. Bioinformatics analysis was conducted to identify the differentially expressed genes (DEGs) and uncover their biological function. In addition, whole genome sequencing was performed on an additional 6 cases of IAs to detect the potential somatic alterations in DEGs. Compared with the normal arterial tissue, 1709 genes were differentially expressed in IAs arterial tissue. The most significantly up-regulated gene and down-regulated gene, H19 and HIST1H3J, may be essential for tumorigenesis of IAs. Hub protein of IKBKG in protein-protein interaction network was probably involved in the inflammation process in aneurysms. Another 2 hub proteins, ACTB and MKI67IP, as well as up-regulated genes, might be abnormally activated in aneurysms and involved in the pathogenesis of IAs. Further whole genome sequencing and filtering yielded 4 candidate somatic single nucleotide variants including MUC3B, and BLM may be involved in the pathogenesis of IAs. Even though, our results do not support the hypothesis of somatic mutations occurred in the DEGs. Two-dimensional genomic data from transcriptome and whole genome sequencing indicated that no somatic mutations occurred in DEGs. In addition, 3 DEGs (IKBKG, ACTB, and MKI67IP) and 2 mutant genes (MUC3B and BLM) were essential in IAs.

Aberrant expression of lncRNAs and mRNAs in patients with intracranial aneurysm.

Intracranial aneurysm (IA) is pathological dilatations of the cerebral artery and rupture of IAs can cause subarachnoid hemorrhage, which has a high ratio of fatality and morbidity. However, the pathogenesis of IAs remains unknown. We performed long noncoding RNA (lncRNA) and messenger RNA (mRNA) expression profiles in IA tissues and superficial temporal arteries (STAs). A total of 4129 differentially expressed lncRNAs and 2926 differentially expressed mRNAs were obtained from the microarrays (P < 0.05). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses showed that up-regulated mRNAs were enriched in immune response, inflammatory response, regulation of immune response and lysosome, et al; while the down-regulated mRNAs were enriched in muscle contraction, smooth muscle contraction, cGMP-PKG signaling pathway and vascular smooth muscle contraction, et al. The lncRNA-mRNA co-expression networks were represented in immune response, inflammatory response, muscle contraction and vascular smooth muscle contraction. These findings may gain insight in the pathogenesis of IAs and provide clues to find key roles for IA patients.

THSD1 (Thrombospondin Type 1 Domain Containing Protein 1) Mutation in the Pathogenesis of Intracranial Aneurysm and Subarachnoid Hemorrhage.

A ruptured intracranial aneurysm (IA) is the leading cause of a subarachnoid hemorrhage. This study seeks to define a specific gene whose mutation leads to disease. More than 500 IA probands and 100 affected families were enrolled and clinically characterized. Whole exome sequencing was performed on a large family, revealing a segregating THSD1 (thrombospondin type 1 domain containing protein 1) mutation. THSD1 was sequenced in other probands and controls. Thsd1 loss-of-function studies in zebrafish and mice were used for in vivo analyses and functional studies performed using an in vitro endothelial cell model. A nonsense mutation in THSD1 was identified that segregated with the 9 affected (3 suffered subarachnoid hemorrhage and 6 had unruptured IA) and was absent in 13 unaffected family members (LOD score 4.69). Targeted THSD1 sequencing identified mutations in 8 of 507 unrelated IA probands, including 3 who had suffered subarachnoid hemorrhage (1.6% [95% confidence interval, 0.8%-3.1%]). These THSD1 mutations/rare variants were highly enriched in our IA patient cohort relative to 89 040 chromosomes in Exome Aggregation Consortium (ExAC) database (P<0.0001). In zebrafish and mice, Thsd1 loss-of-function caused cerebral bleeding (which localized to the subarachnoid space in mice) and increased mortality. Mechanistically, THSD1 loss impaired endothelial cell focal adhesion to the basement membrane. These adhesion defects could be rescued by expression of wild-type THSD1 but not THSD1 mutants identified in IA patients. This report identifies THSD1 mutations in familial and sporadic IA patients and shows that THSD1 loss results in cerebral bleeding in 2 animal models. This finding provides new insight into IA and subarachnoid hemorrhage pathogenesis and provides new understanding of THSD1 function, which includes endothelial cell to extracellular matrix adhesion.

Identification of a Long Noncoding RNA-Associated Competing Endogenous RNA Network in Intracranial Aneurysm

Intracranial aneurysm (IA) is a cerebrovascular disorder characterized by an abnormally bulged artery in the brain and subarachnoid hemorrhage caused by IA rupture with a high ratio of fatality and morbidity. However, the genetic cause of IA remains largely unknown. A dysregulated long noncoding RNA (lncRNA) -associated competing endogenous RNA (ceRNA) network was constructed by using sample-matched microRNA (miRNA), lncRNA, and messenger (mRNA) expression profiles in 12 saccular IA samples and paired control arteries. In this study, 1150 lncRNAs, 2545 mRNAs, and 286 miRNAs were differentially expressed in IAs compared with control arteries. Using miRanda, a ceRNA score was assigned, and gene expression methods were used to construct a ceRNA network. The network consists of 8401 miRNA-lncRNA-mRNA interactions (199 miRNAs, 445 lncRNAs, 1116 mRNAs; P < 0.05). Further Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that the mRNAs involved in the ceRNA network were involved in the regulation of muscle contraction and vascular smooth muscle contraction. By comparing IAs and their control arteries, we identified differentially expressed lncRNAs, miRNAs, and mRNAs and suggested ceRNA roles in the pathogenesis of IA. These findings may help to characterize the pathogenesis of IA and provide novel therapeutic targets in the future for patients with IA.