Taku Yoneyama

Genomewide-Linkage and Haplotype-Association Studies Map Intracranial Aneurysm to Chromosome 7q11

Rupture of intracranial aneurysms (IAs) causes subarachnoid hemorrhage, a devastating condition with high morbidity and mortality. Angiographic and autopsy studies show that IA is a common disorder, with a prevalence of 3%-6%. Although IA has a substantial genetic component, little attention has been given to the genetic determinants. We report here a genomewide linkage study of IA in 104 Japanese affected sib pairs in which positive evidence of linkage on chromosomes 5q22-31 (maximum LOD score [MLS] 2.24), 7q11 (MLS 3.22), and 14q22 (MLS 2.31) were found. The best evidence of linkage is detected at D7S2472, in the vicinity of the elastin gene (ELN), a candidate gene for IA. Fourteen distinct single-nucleotide polymorphisms (SNPs) were identified in ELN, and no obvious allelic association between IA and each SNP was observed. The haplotype between the intron-20/intron-23 polymorphism of ELN is strongly associated with IA (P=3.81x10-6), and homozygous patients are at high risk (P=.002), with an odds ratio of 4.39. These findings suggest that a genetic locus for IA lies within or close to the ELN locus on chromosome 7.

Collagen Type I α2 (COL1A2) Is the Susceptible Gene for Intracranial Aneurysms

Background and Purpose— The collagen &agr;2(I) gene (COL1A2) on chromosome 7q22.1, a positional and functional candidate for intracranial aneurysm (IA), was extensively screened for susceptibility in Japanese IA patients. Methods— Twenty-one single nucleotide polymorphisms (SNPs) of COL1A2 were genotyped in genomic DNA from 260 IA patients (including 115 familial cases) (mean age, 59.9 years) and 293 controls (mean age, 61.6 years). Differences in allelic and genotypic frequencies between the patients and controls were evaluated with the &khgr;2 test. Circular dichroism spectrometry was monitored with collagen-related peptides that mimic triple-helical models of type I collagen with Ala-459 and Pro-459 to estimate the conformation and stability of alterations. Results— Significant genotypic association in the dominant model was observed between an exonic SNP of COL1A2 and familial IA patients (&khgr;2=11.08; df =1; P =0.00087; odds ratio=3.19; 95% CI, 2.22 to 6.50). This SNP induces Ala to Pro substitution at amino acid 459, located on a triple-helical domain. Circular dichroism spectra showed that the Pro-459 peptide had a higher thermal stability than the Ala-459 peptide. Conclusions— The variant of COL1A2 could be a genetic risk factor for IA patients with family history.

Bedside Monitoring of Circulating Blood Volume After Subarachnoid Hemorrhage

Background and Purpose— Maintenance of an adequate intravascular volume is important in the management of patients with subarachnoid hemorrhage (SAH). The purpose of this study was to investigate the circulating blood volume (CBV) after SAH with the use of indocyanine green pulse spectrophotometry. Methods— CBV and plasma hormones related to stress and fluid regulation were measured 4 times: day 2 to 3, day 4 to 5, day 7 to 8, and day 14 in 50 consecutive patients with SAH surgically treated within 48 hours. Results— The mean value of CBV was 64 mL/kg on day 2 to 3, which gradually increased to 69 mL/kg on day 4 to 5, 71 mL/kg on day 7 to 8, and 70 mL/kg on day 14 (P =0.005) (control, 72 mL/kg). The clinical grades and plasma corticotropin levels were higher in patients with <60 mL/kg of CBV on day 2 to 3 (P <0.05 for both). There were no significant differences in other physiological and laboratory parameters such as time for surgery, estimated blood loss, levels of plasma noradrenaline, brain natriuretic peptide, serum sodium, and hematocrit. When CBV was decreased >10% of the former level, there were decreases in hematocrit (P <0.05), serum sodium (P <0.01), and serum albumin (P <0.05) and an increase in urinary sodium (P <0.05). Conclusions— A significant reduction of CBV, especially in patients with poor clinical grades, was noted after SAH and early surgery, which could not be detected by routine examinations. Anemia, central salt wasting, and hypoalbuminemia may be related to a decrease in CBV from the former level. Indocyanine green pulse spectrophotometry may be a powerful tool for the management of patients with SAH.

Association of positional and functional candidate genes FGF1, FBN2, and LOX on 5q31 with intracranial aneurysm.

AbstractWe previously performed a genome-wide linkage study of intracranial aneurysm (IA) and found positive evidence of linkage at chromosomes 5q22–31, 7q11, and 14q22. In the present study, we focus on 5q31, where three candidate genes, fibroblast growth factor 1 (FGF1), fibrillin 2 (FBN2), and lysyl oxidase gene (LOX) lie, and evaluate associations with IA. Genomic DNAs were obtained from 172 IA patients and 192 controls. Association analysis was performed with ten, five, and four single-nucleotide polymorphisms (SNPs) identified in FGF1, FBN2, and LOX, respectively. A difference in allelic frequency was observed for only the SNP at intron 4 in FGF1 (χ2 = 4.44, df=1, P = 0.035). Although a haplotype association was observed with the combination of ten SNPs in FGF1 (χ2 = 16.04, df = 1, P = 0.00006), significant haplotype associations were not observed when haplotypes were constructed with the three, two, and four SNPs in FGF1 according to the linkage disequilibrium structure. No associations of FBN2 and LOX with IA were detected in the present study.

Postcarotid endarterectomy cerebral hyperperfusion can be prevented by minimizing intraoperative cerebral ischemia and strict postoperative blood pressure control under continuous sedation.

OBJECTIVECerebral hyperperfusion syndrome is a major complication after carotid endarterectomy (CEA). We investigated whether our strategy of minimizing intraoperative cerebral ischemia and strict postoperative blood pressure control under continuous sedation prevented postoperative hyperperfusion. METHODSEighty consecutive patients undergoing CEA were studied. A shunt was used in all patients during CEA. All patients were managed postoperatively under continuous sedation for as long as 48 hours on the basis of the regional cerebral blood flow (rCBF) measured immediately after CEA. Postoperative hyperperfusion was assessed, on the basis of the cerebral blood flow study under sedation (propofol) after CEA, either as a greater than 30% increase in rCBF compared with the contralateral side, or a greater than 100% increase in the corrected rCBF (calculated from percentage reduction of the contralateral rCBF induced by propofol) compared with preoperative values. RESULTSNo patient developed cerebral hyperperfusion syndrome. Postoperative hyperperfusion was found at very low rates (2.5% in the middle cerebral artery territory and 1.3% in the anterior cerebral artery territory by definition 1, and 0% in both territories by definition 2). Ratios of regional oxygen saturation after internal carotid artery clamping to preclamp baseline values were greater than 0.9 in 78 of 80 patients, indicating very mild intraoperative cerebral ischemia. Parameters related to cerebral ischemia during CEA, such as regional oxygen saturation, internal carotid artery cross-clamping duration, and stump pressure (index), did not affect the incidence of postoperative hyperperfusion. CONCLUSIONThe present study suggests that minimizing intraoperative cerebral ischemia using a shunt, followed by strict postoperative blood pressure control under continuous sedation, can prevent post-CEA hyperperfusion.

Gene Expression Profiling Reveals Distinct Molecular Signatures Associated With the Rupture of Intracranial Aneurysm

Background and Purpose— The rupture of intracranial aneurysm (IA) causes subarachnoid hemorrhage associated with high morbidity and mortality. We compared gene expression profiles in aneurysmal domes between unruptured IAs and ruptured IAs (RIAs) to elucidate biological mechanisms predisposing to the rupture of IA. Methods— We determined gene expression levels of 8 RIAs, 5 unruptured IAs, and 10 superficial temporal arteries with the Agilent microarrays. To explore biological heterogeneity of IAs, we classified the samples into subgroups showing similar gene expression patterns, using clustering methods. Results— The clustering analysis identified 4 groups: superficial temporal arteries and unruptured IAs were aggregated into their own clusters, whereas RIAs segregated into 2 distinct subgroups (early and late RIAs). Comparing gene expression levels between early RIAs and unruptured IAs, we identified 430 upregulated and 617 downregulated genes in early RIAs. The upregulated genes were associated with inflammatory and immune responses and phagocytosis including S100/calgranulin genes (S100A8, S100A9, and S100A12). The downregulated genes suggest mechanical weakness of aneurysm walls. The expressions of Krüppel-like family of transcription factors (KLF2, KLF12, and KLF15), which were anti-inflammatory regulators, and CDKN2A, which was located on chromosome 9p21 that was the most consistently replicated locus in genome-wide association studies of IA, were also downregulated. Conclusions— We demonstrate that gene expression patterns of RIAs were different according to the age of patients. The results suggest that macrophage-mediated inflammation is a key biological pathway for IA rupture. The identified genes can be good candidates for molecular markers of rupture-prone IAs and therapeutic targets.

Endoglin Is Not a Major Susceptibility Gene for Intracranial Aneurysm Among Japanese

Background and Purpose— A 6-base insertion (6bINS) polymorphism in intron 7 of the endoglin gene (ENG), which codes for a component of the transforming growth factor-&bgr; receptor complex, was reported to be associated with intracranial aneurysm (IA) in a Japanese population. A recent report using a white population could not replicate the association. We tested for this association with high statistical power in our independent Japanese subjects and evaluated the linkage between markers on chromosome 9, which contains ENG, and IA. Methods— The sample for the linkage study comprised 179 individuals with IA in 85 nuclear families, with 104 possible affected sibpairs. For the association study of the 6bINS polymorphism and 4 single nucleotide polymorphisms (SNPs) in ENG, 172 Japanese patients with IA and 192 control subjects were examined. Results— There was no evidence of linkage in the vicinity of ENG by analysis of affected sibpairs. The allele frequency of the 6bINS polymorphism was 104 of 344 (30.2%) in the total IA group and 122 of 382 (31.9%) in the control group. The statistical difference in allele frequency between the 2 groups was not significant (&khgr;2=0.245, df =1, P =0.620). The power of the present association study was 98.3% at a significance level of 0.05 on the basis of the allele frequencies in the previous study. In addition, no associations between the 4 SNPs in ENG and IA were detected. Conclusions— We provide evidence that there is no association between the 6bINS polymorphism or 4 SNPs in ENG and IA and that there is no linkage between the ENG locus and IA, indicating that ENG is not a major susceptibility gene for IA in Japanese.

Systematic Validation of RNF213 Coding Variants in Japanese Patients With Moyamoya Disease

Background A founder variant of RNF213, p.R4810K (c.14429G>A, rs112735431), was recently identified as a major genetic risk factor for moyamoya disease (MMD) in Japan. Although the association of p.R4810K was reported to be highly significant and reproducible, the disease susceptibility of other RNF213 variants remains largely unknown. In the present study, we systematically evaluated the coding variants detected in Japanese patients and controls for associations with MMD. Methods and Results To detect variants of RNF213, all coding exons were sequenced in 27 Japanese MMD patients without p.R4810K. We also validated all previously reported variants in our case–control samples and tested for associations in combination with previous Japanese study cohorts, including the 1000 Genomes Project data set, as population-based controls. Forty-six missense variants other than p.R4810K were identified among 370 combined patients and 279 combined controls in Japan. Sixteen of 46 variants were polymorphisms with minor allele frequency >1%, and, after conditioning on the p.R4810K genotype, were not associated with MMD. We conducted a variable threshold test using Combined Annotation-Dependent Depletion on the remaining 30 rare variants (minor allele frequency <1%), and the results showed that the frequency of potentially functional variants was significantly higher in patients than in controls (permutation, minimum P=0.045). Conclusions Not only p.4810K but also other functional missense variants of RNF213 conferred susceptibility to MMD. Our analysis also revealed that ≈20% of Japanese MMD patients did not harbor susceptibility variants of RNF213, indicating the presence of other susceptibility genes for MMD.

Hemodynamics and changes after STA–MCA anastomosis in moyamoya disease and atherosclerotic cerebrovascular disease measured by micro-Doppler ultrasonography

Moyamoya disease (MMD) and atherosclerotic cerebrovascular disease (ACVD) differ in angiographic appearance and probably hemodynamics. Pediatric MMD (PMMD) usually presents with cerebral ischemia, while intracranial hemorrhage is more common in adult MMD (AMMD), suggesting differences in cerebral hemodynamics. We analyzed the cortical flow velocity and direction of recipient arteries using micro-Doppler ultrasonography to evaluate the cortical circulation before and after anastomosis in MMD and ACVD. Twenty-eight patients with adult MMD (AMMD), 7 with pediatric MMD (PMMD), 16 with ACVD, and 12 control patients were studied. A micro-Doppler probe was applied on the cortical recipient artery (A4 or M4) before and after anastomosis. Systolic maximum flow velocity (Vmax) and blood flow direction were investigated at proximal and distal parts of anastomosed sites in recipient arteries. Pre- and postoperative regional cerebral blood flow was measured by cold xenon-computed tomography (Xe-CT). Before anastomosis, retrograde cortical flow was significantly more common in PMMD patients, and Vmax in cortical artery was significantly lower in AMMD patients. Bypass surgery changed the direction of blood flow from the anastomosis site to proximal and distal sites of the recipient artery in most patients, but pre-anastomosis flow direction was preserved more frequently in PMMD patients. The rate of Vmax increase after anastomosis was significantly higher in AMMD than in PMMD (11.6 ± 9.8 vs. 3.9 ± 1.8; P = 0.01). Micro-Doppler ultrasonography identified differences in cortical circulation among AMMD, PMMD, and ACVD. In AMMD, significantly low velocity in the cortical artery was observed before anastomosis, and bypass surgery reversed the flow and significantly increased flow velocity. The data of PMMD showed unique hemodynamics of the cortical artery before anastomosis, characterized by a higher frequency of retrograde flow and preserved velocity. The Vmax increase rate was significantly higher in patients with postoperative cerebral hyperperfusion on Xe-CT, and further study is warranted to validate the clinical use of intraoperative micro-Doppler monitoring to predict postoperative hyperperfusion.

Is there any evidence for linkage on chromosome 17cen in affected Japanese sib-pairs with an intracranial aneurysm?

AbstractA linkage region on chromosome 17cen has previously been found in 29 Japanese families with a history of familial intracranial aneurysms (IA). To investigate whether there is evidence of linkage in affected Japanese sib-pairs we performed nonparametric and parametric linkage analysis of a total of 253 familial aneurysm cases including 111 affected sib-pairs (ASP). Ten microsatellite markers covering a 17.7 cM region were chosen, in accordance with previous work in which nominal P had been below 0.05. Statistical analysis was performed by use of Genehunter and Sibpal software. After calculation of the logarithm of the odds (LOD) and nonparametric linkage analysis (NPL) scores our study did not show any linkage in the region analyzed. Our conclusion did not change even after only ASP were analyzed. In contrast with a previous study examining multigenerational Japanese families with IA, most Japanese ASP may not have a genetic linkage to chromosome 17 cen.