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Yang Yao,1,2,* Xinggen Fang,1,* Jinlong Yuan,1 Feiyun Qin,1 Tao Yu,1 Dayong Xia,1,3 Zhenbao Li,1 Niansheng Lai1,3 1Department of Neurosurgery, First Affiliated Hospital of Wannan Medical College (Yijishan Hospital of Wannan Medical College), Wuhu, 241001, Anhui Province, People’s Republic of China; 2Department of Nursing, The First Affiliated Hospital of Wannan Medical College (Yijishan Hospital of Wannan Medical College), Wuhu, 241001, Anhui Province, People’s Republic of China; 3Key Laboratory of Non-Coding RNA Transformation Research of Anhui Higher Education Institution (Wannan Medical College), Wuhu, 241001, Anhui Province, People’s Republic of China*These authors contributed equally to this workCorrespondence: Niansheng Lai; Zhenbao LiDepartment of Neurosurgery, First Affiliated Hospital of Wannan Medical College (Yijishan Hospital), 2 West Zheshan Road, Wuhu, 241001, Anhui Province, People’s Republic of ChinaTel +86-15155308793Fax +86-05535739594Email mlns@wnmc.edu.cn; zhenbaoli123@126.comObjective: Aneurysmal subarachnoid hemorrhage (aSAH) is a severe form of stroke characterized by high rates of mortality and disability. Identifying circulating biomarkers is helpful to improve outcomes. In this study, for the first time, we identify interleukin-6 (IL-6) in cerebrospinal fluid (CSF) small extracellular vesicles (sEVs) as potential biomarkers for prognosis of aSAH.Methods: We extracted small extracellular vesicles from the CSF of 103 aSAH patients and 40 healthy controls in a prospective observational study. Subsequently, we measured IL-6sEVs levels using an enzyme-linked immunosorbent assay. Results were statistically analyzed to determine the function of IL-6sEVs for disease monitoring of aSAH.Results: CSF IL-6 sEVs showed distinct pattern differences between healthy controls and aSAH patients. The concentration of IL-6sEVs in CSF is significantly correlated with the severity of aSAH patients. The areas under the receiver operating characteristic curves of IL-6sEVs for identifying severe aSAH patient from aSAH patients were 0.900. After multivariate logistic regression analysis, IL-6sEVs were associated with neurological outcome at 1 year. IL-6sEVs levels were greater and positively associated with disease processes and outcome.Conclusion: There is a neuroinflammatory cascade in aSAH patients. IL-6sEVs in CSF may be a biomarker for the progression of aSAH.Keywords: subarachnoid hemorrhage, cerebrospinal fluid, small extracellular vesicles, interleukin-6, biomarker

Aims Acyl‐CoA synthetase long chain family member 4 (ACSL4) is closely related to tumor genesis and development in certain tissues. However, the function of ACSL4 in early brain injury (EBI) caused by subarachnoid hemorrhage (SAH) is unclear. In this study, we investigated the expression patterns and role of ACSL4 in SAH and post‐SAH EBI using a rat model of SAH. Methods The rat model of SAH was induced by autologous blood injection into the prechiasmatic cistern of rats. We also used two specific inhibitors of ferroptosis (Ferrostatin‐1 and Liproxstatin‐1) to investigate the role of ferroptosis in EBI. Results We found that ACSL4 levels in brain tissue increased significantly in post‐SAH EBI. Inhibiting the expression of ACSL4 using small interfering RNAs alleviated inflammation, blood‐brain barrier (BBB) impairment, oxidative stress, brain edema, and behavioral and cognitive deficits, and increased the number of surviving neurons, after SAH. Similar effects were obtained by suppressing ferroptosis. Conclusions ACSL4 exacerbated SAH‐induced EBI by mediating ferroptosis. These findings may provide a theoretical basis for potential therapy aimed at alleviating post‐SAH EBI., ACSL4 levels in brain tissue of rats increase significantly in EBI and brain damage after SAH could be reduced by down‐regulation of ACSL4. ACSL4 could trigger ferroptosis and aggravate brain damage via catalyzing lipid metabolism. It may provide a theoretical basis for potential therapy to alleviate EBI after SAH

RAB7, a member of RAS oncogene family-like 1 (RAB7L1), is a GTPase belonging to the Rab family and acts as an upstream regulator to regulate the kinase activity of leucine-rich repeat kinase 2 (LRRK2). Although LRRK2 has been shown to aggravate secondary brain injury (SBI) after intracerebral hemorrhage (ICH), it is unknown whether RAB7L1 is also involved in this process. The purpose of the present study was to investigate the role of RAB7L1 in ICH-induced SBI in vivo. Autologous blood was injected into adult male Sprague-Dawley rats to induce an ICH model in vivo. The results showed that the protein levels of RAB7L1 increased after ICH. Overexpression of RAB7L1 induced neuronal apoptosis and damage, as demonstrated by TUNEL-positive and FJB-positive cells, and exacerbated ICH-induced learning and cognitive dysfunctions; in contrast, downregulation of RAB7L1 via RNA interference yielded comparatively opposite changes in these parameters. In summary, this study demonstrates that RAB7L1 promotes SBI after ICH and may represent a potential target for ICH therapy.

Objective Hemodynamic factors are thought to play important roles in the pathogenesis, progression, and rupture of cerebral aneurysms. Previous hemodynamic studies have been based on comparisons between post-ruptured and unruptured aneurysms. Nevertheless, changes of aneurysm morphology after rupture render these results unreliable. Moreover, pressure, age, gender, and the morphology of the parent artery also influence these results. Therefore, in the present study, we identified hemodynamic and morphological characteristics of aneurysms prior to rupture using twelve mirror aneurysms. Materials and methods From our database, we retrospectively analyzed twelve mirror aneurysms (MANs) prior to rupture. Each mirror aneurysm was divided into the prior to rupture or the unruptured group. Patient-specific models were reconstructed from three-dimensional (3D) images of all patients. Hemodynamic and morphological factors were analyzed and compared. Results Compared with the unruptured side of MANs, aneurysms prior to rupture were significantly larger and significantly more irregular in shape; they also had significantly higher aspect ratio (AR), size ratio (SR), undulation index (UI), ellipticity index (EI), percentage of low wall shear stress area (LSA) and significantly lower normal wall shear stress (NWSS). The oscillatory shear index (OSI) and nonsphericity index (NSI) in the aneurysms prior to rupture were non-significantly higher than those of the unruptured group. Conclusion MANs prior to rupture may be extremely useful models to assess the risk of aneurysm rupture. Larger size, irregular shape, higher AR, SR, UI, NI, and lower WSS may be associated with aneurysms at risk for rupture.

Background Inflammation is a potential crucial factor in the pathogenesis of subarachnoid hemorrhage (SAH). Circulating microRNAs (miRNAs) are involved in the regulation of diverse aspects of neuronal dysfunction. The therapeutic potential of miRNAs has been demonstrated in several CNS disorders and is thought to involve modulation of neuroinflammation. Here, we found that peripherally injected modified exosomes (Exos) delivered miRNAs to the brains of mice with SAH and that the potential mechanism was regulated by regulation of neuroinflammation. Methods Next-generation sequencing (NGS) and qRT-PCR were used to define the global miRNA profile of plasma exosomes in aSAH patients and healthy controls. We peripherally injected RVG/Exos/miR-193b-3p to achieve delivery of miR-193b-3p to the brain of mice with SAH. The effects of miR-193b-3p on SAH were assayed using a neurological score, brain water content, blood-brain barrier (BBB) injury, and Fluoro-Jade C (FJC) staining. Western blotting analysis, enzyme-linked immunosorbent assay (ELISA), and qRT-PCR were used to measure various proteins and mRNA levels. Results NGS and qRT-PCR revealed that four circulating exosomal miRNAs were differentially expressed. RVG/Exos exhibited improved targeting to the brains of SAH mice. MiR-193b-3p suppressed the expression and activity of HDAC3, upregulating the acetylation of NF-κB p65. Finally, miR-193b-3p treatment mitigated the neurological behavioral impairment, brain edema, BBB injury, and neurodegeneration induced by SAH, and reduced inflammatory cytokine expression in the brains of mice after SAH. Conclusions Exos/miR-193b-3p treatment attenuated the inflammatory response by acetylation of the NF-κB p65 via suppressed expression and activity of HDAC3. These effects alleviated neurobehavioral impairments and neuroinflammation following SAH.

Objective: Posterior communicating artery bifurcation aneurysms (PcomA-BAs) and infundibular dilations (PcomA-IDs) are found at the junction between the internal carotid artery (ICA) and the posterior communicating artery (PcomA). Several studies found that PcomA-IDs potentially progress to aneurysms and can even rupture. In our clinical practice, digital subtraction angiography (DSA) helps differentiate PcomA-IDs from unruptured PcomA-BAs. However, when PcomA-IDs are >3 mm in diameter or PcomA are absent on DSA, it is challenging to use DSA to differentiate PcomA-IDs from unruptured PcomA-BAs. Hemodynamic and morphological factors are thought to play important roles in the pathogenesis, progression, and rupture of cerebral aneurysms. We compared hemodynamic and morphological differences in unruptured PcomA-BAs and PcomA-IDs to better manage PcomA-IDs.Methods: We included 83 PcomA-IDs and 115 unruptured PcomA-BAs dignosed and measured using DSA from January 2015 to January 2019. Computational fluid dynamics was performed on these patient-specific models reconstructed using axial slices in DICOM format. Clinical, hemodynamic, and morphological factors were compared between the PcomA-IDs and PcomA-BAs. Significant parameters were analyzed using binary logistic regression analysis to identify independent risk factors. Receiver operating characteristic (ROC) analysis was performed on the independent risk factors to acquire cutoff values.Results: One hemodynamic and four morphyological parameters were significantly different between PcomA-IDs and PcomA-BAs: normalized wall shear stress (NWSS), size, the angle between the ophthalmic segment of the ICA and the PcomA (APcomA), the angle between the ophthalmic and the communicating segment of the ICA (AICA) and the diameter of the PcomA (DPcomA). Binary logistic regression analysis showed that small size and DPcomA as well as APcomA were all independent significant factors characterizing the status of PcomA-IDs and the ROC analysis for independent risk factors indicated the cutoff values of size, APcomA, and DPcomA were 3.45 mm, 66.27°, and 1.24 mm, respectively.Conclusions: Size, DpcomA, and ApcomA could independently characterize the status of PcomA-IDs. These might help us better differentiate them from real aneurysms and guide its management.

Background Aneurysmal subarachnoid hemorrhage (aSAH) is a severe type of stroke characterized by high rates of mortality and disability. Identifying circulating biomarkers is helpful to improve prognosis. In this study, for the first time, we identify circulating interleukin-6 (IL-6) levels in neuron-derived small extracellular vesicles (NDSEVs) as potential biomarkers in prognosis of aSAH.Methods We extracted small extracellular vesicles from the plasma of aSAH patients and healthy controls and enriched them using sequential precipitation and anti-L1CAM antibody immunoabsorption. Subsequently, we determined IL-6 levels using an enzyme-linked immunosorbent assay (ELISA).Results Plasma IL-6 NDSEVs showed distinct pattern differences between aSAH patients and healthy controls. There were significant correlations of IL-6concentrations in plasma with severity in aSAH. The AUCs of IL-6 for distinguishing the severe aSAH patients from mild aSAH patients were 0.961. After multivariate logistic regression analysis, only age, acute hydrocephalus,and the levels of IL-6 NDSEVs enabled prediction of neurological outcome at 1 year. The IL-6 NDSEVs levels were greater and positively associated with disease and prognosis of aSAH patients.Conclusions These data suggest a neuroinflammatory cascade in aSAH patients. IL-6 NDSEVs may be a biomarkers to monitor the progression of aSAH.

Objective: To investigate the hemodynamic features before and after embolization of paraclinoidal aneurysms using hemodynamic numerical simulation and the influence of embolization on recurrence after embolization.Methods: From January 2016 to December 2017, we enrolled a total of 113 paraclinoidal aneurysms treated with embolization. They were divided into recurrent group and stable group depending on follow-up results. An aneurysm model was generated based on 3D-DSA before and after embolization. The hemodynamic characteristics were analyzed between two groups using Computational fluid dynamic (CFD).Results: In the recurrent group, the peak systolic WSS, OSI and velocity around the aneurysm neck areas prior to embolization were 20.47 ± 3.04 Pa, 0.06 ± 0.02 and 0.07 ± 0.03 m/s, respectively. These values were 23.50 ± 4.11 Pa, 0.06 ± 0.01 and 0.11 ± 0.02 m/s, respectively in the stable group (P > 0.05). The WSS, OSI, velocity around the same areas in the recurrent group after embolization were 35.59 ± 8.75 Pa, 0.07 ± 0.02 and 0.12 ± 0.03 m/s, respectively (P < 0.01). In the stable group, the WSS, OSI and velocity were 13.08 ± 2.89 Pa, 0.04 ± 0.01 and 0.07 ± 0.02 m/s, respectively (P < 0.01). After embolization, the WSS, OSI and velocity around the aneurysm neck areas in the recurrent group were significantly higher than those in the stable group.Conclusions: High peak systolic WSS, OSI and velocity around aneurysm neck areas after embolization of paraclinoidal aneurysms may be important factors leading to recurrence.

Background Intracerebral hemorrhage (ICH) is the common brain diseases in middle-aged and elderly people, with high disability and/or mortality rate, and is a serious public health concern. Both WNK3 kinase and the WNK3/SPAK/NKCC1 signaling pathway play an integral role in maintaining normal cell homeostasis. However, their role and underlying mechanisms in ICH-induced secondary brain injury (SBI) have yet to be elucidated. Methods We established an ICH model using male Sprague-Dawley (SD) rats by injecting autologous arterial blood into the unilateral basal ganglia. To establish ICH model in vitro, oxyhemoglobin (OxyHb; 20 μM) and neurons were cultured for 6 h at 37 °C, 5% CO2 atmosphere. To investigate the role of WNK3 and the WNK3/SPAK/NKCC1 signaling pathway in SBI, after genetic interventions, rotation and water maze test, brain edema and neuroinflammation were detected, and terminal-deoxynucleoitidyl transferase mediated dUTP nick end labeling (TUNEL), Fluoro-Jade C (FJC), and Nissl staining were performed. Results Our data showed that WNK3 expression in brain tissue were upregulated after ICH induction. In addition, silencing of WNK3 reduced neuronal apoptosis, and inflammatory responses in rats that underwent ICH. Inhibition of WNK3 expression reduced the damaged blood-brain barrier (BBB), alleviated the impaired degree of cerebral edema, and improved disruptive neurobehavioral cognition caused by ICH. Moreover, overexpression of WNK3 had the opposite effects. Finally, WNK3/SPAK/NKCC1 signaling pathway may be involved in the above-mentioned processes. Conclusions In conclusion, our findings showed that WNK3 and WNK3/SPAK/NKCC1 signaling pathway play a vital biological function in ICH-induced SBI. Depletion of WNK3 attenuated brain injury after ICH both in vivo and in vitro. Thus, WNK3 and WNK3/SPAK/NKCC1 signaling pathway are potential targets for treating SBI after ICH.

miRNAs are important regulators of translation and have been associated with the pathogenesis of a number of cardiovascular diseases including stroke and may be possible prognostic biomarkers. The purpose of the present study was to determine the expression levels of miRNAs in the sera of subarachnoid haemorrhage (SAH) patients and to evaluate their relationships with the severity and clinical outcome of SAH. Serum samples on day 3 after the onset of SAH were subjected to microarray analysis with Exqion miRCURYTM LNA array and quantitative PCR analysis. Serum samples from SAH patients (n=60) and healthy controls (n=10) were subjected to quantitative PCR analysis. The severities and clinical outcomes of the SAH patients were evaluated with the WFNS grade and the Modified Rankin Scale (mRS). Three miRNAs, miR-502-5p, miR-1297 and miR-4320 were significantly up-regulated in the sera of SAH patients when compared with the healthy controls. The serum miR-502-5p and miR-1297 levels were significantly higher in the patients with severe SAH and a poor outcome than in those with mild SAH and a good outcome (P

The optimal treatment modalities of ruptured middle cerebral artery aneurysm are still controversial. The objective of this study is to analyze the outcomes of patients with ruptured middle cerebral artery aneurysms treated by endovascular coiling.From October 2011 to October 2015, 67 patients with 71 ruptured middle cerebral artery aneurysms received endovascular coiling in our hospital. We retrospectively reviewed the clinical, radiologic records and outcomes.Of all the 71 aneurysms (67 patients), 42 were treated by coil embolization merely, 27 by stent-assisted coiling and 2 unruptured aneurysms in patients with bilateral middle cerebral artery aneurysms without receiving treatment. Complete occlusion was achieved in 82.6% (57/69) of all the procedures. Each of incomplete and partial occlusion rates was 8.7% (6/69). Intraoperative rupture of aneurysms occurred in two procedures (2.9%). Thrombogenesis occurred in eight procedures (11.6%). Brain infarction occurred in eight patients (11.9%). Post-operative rebleeding occurred in seven patients (10.4%). Sixty-three patients were followed at a mean follow-up of 8.24 ± 7.16 months. The mortality and good outcome rate were 3.2% and 90.5%, respectively. Aneurysm recurrence occurred in 6 (13.3%) of the 45 aneurysms at a mean follow-up of 8.44 ± 7.83 months.Endovascular coiling is effective for patients with ruptured middle cerebral artery aneurysms. Individualized treatment should be assessed by experienced specialist. It is essential to perform randomized large trials to confirm the efficiency of endovascular coiling.

Enterprise stent has been widespread used in wide-necked intracranial aneurysms and good efficacy has been achieved, but there are few reports on its applications in very small ruptured intracranial aneurysms in literatures. This study aimed to evaluate the safety and efficacy of Enterprise stent-assisted coiling embolization of very small ruptured intracranial aneurysms. We retrospectively reviewed the clinical and imaging data from 37 patients with very small ruptured intracranial aneurysms who had SAC using Enterprise stents performed from February 2012 to July 2016 in our department. Data collected and analyzed included patient demographics, morphologic features of the aneurysm, treatment results, and follow-up results. Clinical outcomes were evaluated by the Glasgow Outcome Scale (GOS). Enterprise stents were successfully implanted in all 37 patients with very small ruptured intracranial aneurysms. Of the 37 individuals, 28 patients exhibited complete occlusion at Raymond grade I, 5 patients exhibited occlusion at Raymond grade II, and 4 patients at Raymond grade III. Procedure-related complications occurred in 3 of 37 patients (8.1%), including 1 case of intraprocedure aneurysm rupture who died from cerebral herniation caused by severe postoperative cerebral ischemia during the hospital stay, and the other 2 complications were acute in-stent thrombosis, and occlusion of parent artery caused by falling-off internal carotid artery plaque, respectively. A total of 36 patients underwent postoperative clinical follow-up visits for 6 to 24 months of which 31 patients recovered (GOS ≥ 4). One patient had hemiplegic paralysis, and no rehemorrhage was found. A total of 25 patients underwent follow-up digital subtraction angiography (DSA) at 3–21 months postintervention, in whom there were 22 cases with complete occlusion, 2 cases with recurrence of aneurysm neck, and 1 case with in-stent restenosis, but there was no patient with neurologic deficits. The Enterprise stent-assisted coiling embolization can be a safe and effective technique for treatment of very small ruptured intracranial aneurysms.

MicroRNA-155 is highly expressed in many malignant tumors with poor prognosis, which regulates cell apoptosis, proliferation, invasion, metastasis, tumor angiogenesis, and metabolism. This study aims at investigating the clinical significance of miR-155 expression in human gliomas. Real-time quantitative PCR was used to detect the expression levels of miR-155 in 131 glioma and 16 normal brain tissues. The association of miR-155 expression with clinicopathological factors and prognosis of glioma patients were analyzed. The expression levels of miR-155 were significantly higher in glioma tissues than that in normal brain tissues (P < 0.001), which was associated with high pathological grade (P < 0.001) and low Karnofsky Performance Status score (P = 0.022). As a result of Kaplan-Meier survival and Cox regression analyses, overall survival (OS) rates and progression-free survival were significantly poorer in high-expression group relative to low-expression group (both P < 0.001). Furthermore, miR-155 expression was significantly associated with poor OS (P < 0.001) and PFS (P = 0.001) in glioma patients who had high pathological grades (III-IV) as calculated by subgroup analyses. These findings reveal that miR-155 expression might be an independent prognostic factor and a therapeutic target for human glioma.