Current Neurovascular Research - Volume 16, Issue 5, 2019

Background: A number of studies have explored the prognostic role of CRP in patients with acute ischemic stroke, however, the results have been inconclusive. The aim of our study was to investigate the impact of infection on the association between CRP and 3-month functional outcome by performing a registry study and systematic review. Methods: Patients admitted within 24 hours of acute ischemic stroke onset and had CRP measured within 24 hours after admission were included. Patients admitted between June 2016 and December 2018 in Chengdu Stoke Registry were enrolled. The PubMed database was searched up to July 2019 to identify eligible studies. Poor outcome was defined as modified Rankin Scale scores at 3-month more than 3. Results: Totally, 368 patients in the registry and 18 studies involving 15238 patients in the systematic review were included. A statistically significant association between CRP values on admission and 3-month poor outcome in patients without infection was found, both in our registry (CRP per 1-mg/L increment, OR 1.04, 95% CI 1.01 to 1.07, p=0.008) and meta-analysis (CRP per 1-mg/dL increment, OR 1.66 [95% CI 1.37 to 2.01, p<0.001]). In patients with infection, CRP was not associated with a 3-month poor outcome according to registry data (OR 1.00, 95% CI 0.99 to 1.01, p=0.663) and meta-analysis (OR 1.01, 95% CI 0.99 to 1.01, p=0.128). Conclusion: High CRP value was independently associated with a 3-month poor outcome after stroke in patients without infection. Further studies are required to examine the value of infection on CRP measures and long-term functional outcomes.

Objective: The safety and effect of intra-arterial (IA) tirofiban, a glycoprotein IIb/IIIa inhibitor, during the stent retriever mechanical thrombectomy (MT) was investigated. Methods: From January 2015 to May 2019, a total of 327 patients underwent mechanical thrombectomy of large artery occlusions (LAO). Patients were classified into two groups: MT with IA tirofiban (MTT) group and MT only (MTO, without IA tirofiban) group. Clinical outcomes, radiological results, and various complications, such as post thrombectomy hemorrhage, symptomatic hemorrhage, other systemic bleeding, and hemorrhagic transformation of infarct were evaluated by comparing the MTT group and MTO group. In addition, subgroup analysis was performed for patients who underwent MT with prior intravenous (IV) tissue plasminogen activator (t-PA). Results: The MTT group needed a lower mean number of stent passes and showed a re-occlusion rate as compared with the MTO group (P=0.038 and 0.022, respectively). Between the two groups, there were no statistically significant differences in post thrombectomy hemorrhage, symptomatic hemorrhage, other systemic bleeding complications, or hemorrhagic transformation of infarct (P = 0.511, 0.397, 0.429, and 0.355, respectively). In the subgroup analysis, similar findings were observed. Conclusion: The use of IA tirofiban during MT seems to be safe and potentially more effective than only MT without IA tirofiban, even in patients who used IV t-PA before MT.

Background: Granulocyte-colony stimulating factor (G-CSF) has protective effects on many neurological diseases. The effects of G-CSF on vascular endothelium and White Matter (WM) injury in Cerebral Small Vessel Disease (CSVD) were explored in this study via a model of spontaneously hypertensive rat (SHR) in order to elucidate the mechanism of G-CSF in Vascular Cognitive Impairment (VCI). Methods: 24-week-old male SHRs were randomly divided into the treatment group and model group, with the same age Wistar rats as the control group. The novel object recognition test (NORT) and Morris water maze were conducted after 7 days of G-CSF(50ug/kg) or normal saline treatment to examine their non-spatial and spatial cognitive functions. After that, a transmission electron microscope (TEM) and FLB staining were used to observe the vascular endothelial cell and WM damage. Furthermore, the expression of VEGF, MMP-9, Caspase-3, TUNEL and BrdULaminin in the cortical area was detected by immunostaining methods. Results: Our results showed that G-CSF promoted the expression of VEGF and BrdU+-Laminin+ endothelial cells, but down-regulated the level of MMP-9, thus significantly repaired the cerebral vascular endothelial cells and perivascular structure in SHR. The WM damage, the expression of caspase-3 and the apoptosis rate decreased after G-CSF treatment. Ultimately, G-CSF improved the non-spatial cognitive function in SHR rather than the spatial cognitive function. Conclusion: Therefore, our findings indicated that G-CSF might facilitate the improvement of non-spatial cognitive function in CSVD by repairing endothelial cells and alleviating WM damage.

Background: The function of microRNA-542-3p (miR-542-3p) in rat epilepsy is still unclear. Methods: The levels of miR-542-3p and toll-like receptor 4 (TLR4) were determined through quantitative real-time PCR. The protein levels were examined via the western blot analysis. The relationship between miR-542-3p and TLR4 was confirmed through luciferase assay. Pathological changes were analyzed via Hematoxylin-eosin (HE) and Nissl staining. Results: The rats and hippocampal cells were treated with kainic acid (KA) in vivo and in vitro. miR-542-3p was low in KA-treated rats, hippocampal cells and cerebrospinal fluid of patients with epilepsy. Further functional analysis showed that miR-542-3p overexpression inhibited KAinduced average seizure frequency, damage of hippocampal neuron and cell apoptosis, leading to the alleviation of the brain injury in epilepsy rats. miR-542-3p was determined to downregulate TLR4 expression. The relationship between miR-542-3p and TLR4 was confirmed. TLR4 knockdown reduced KA-induced nuclear factor-kappa B p65 (NF-ΚB p65), multidrug resistance 1 (MDR1), P-glycoprotein (P-gp) and apoptosis-associated protein levels. Further, for NF-ΚB p65, MDR1, P-gp and apoptosis-associated protein levels detection, miR-542-3p mimic showed a suppressive effect on these KA-induced protein levels, whereas TLR4 overexpression ameliorated the miR-542-3p-induced these protein levels in KA-treated epilepsy rats. Conclusion: We identified that miR-542-3p attenuated seizure-induced brain injury and the expression of P-gp in epilepsy rats through inhibiting TLR4/NF-ΚB signaling pathway, which might contribute to improved epilepsy therapy.

Objective: This study was to investigate the potential protective effects of curcumin in cerebral ischemia-reperfusion (CIR) and its regulation of miR-7. Methods: Rats were occluded by middle cerebral artery occlusion (MCAO) for 1.5 h and reperfused for 2 h to establish a local CIR model. After 24 hours of model establishment, MCAO rats were given curcumin for 3 days by intragastric administration. PC12 cells were cultured for 6 h in oxygen-glucose deprivation medium and then reoxygenated for 24 h to establish an oxygenglucose deprivation/reoxygenation (OGD/R) model. The OGD/R model cells were treated with curcumin for 48 h. Results: Curcumin inhibited the decrease of miR-7-5p expression and an increase of RelA p65 expression induced by CIR and ODG/R. RelA p65 was a target of miR-7-5p. MiR-7-5p antagonists were able to counteract the effect of curcumin on the expression of RelA p65 in ischemic brain tissue of MCAO rats and OGD/R model cells. Curcumin improved OGD/R-induced inhibition of cell activity, necrosis and apoptosis. Curcumin significantly reduced the levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1β, reactive oxygen species (ROS) and malondialdehyde (MDA) and increased the activity of superoxide dismutases (SOD) and catalase (CAT) in OGD/R-induced cells. Curcumin may inhibit OGD/R-induced cell damage by regulating miR-7-5p. Curcumin improved cerebral infarction, nerve damage and cognitive dysfunction in rats with CIR, which may be related to the regulation of miR-7-5p/RelA p65 axis. Conclusion: Curcumin exerts cerebral protection by attenuating cell necrosis and apoptosis, inflammatory response and oxidative stress following CIR, which may be related to its regulation of the miR-7/RELA p65 axis.

Aim: To determine whether apelin in paraventricular nucleus (PVN) can be a therapeutic target for hypertension. Background: Apelin is a specific endogenous ligand of orphan G protein-coupled receptor APJ. Objective: This study was designed to determine how apelin chronically regulates sympathetic nerve activity and blood pressure in PVN of rats. Methods: Apelin and APJ antagonist F13A were infused into PVN with osmotic minipumps. The NAD(P)H oxidase activity and superoxide anions levels in PVN of rats were determined by chemiluminescence. Results: Infusion of apelin into PVN of Wistar-Kyoto (WKY) rats induced chronic increases in systolic blood pressure (SBP), diastolic blood pressure (DBP), mean arterial pressure (MAP), plasma norepinephrine (NE) level, maximal depressor response to hexamethonium (Hex), NAD(P)H oxidase activity, superoxide anions levels, and Nox4 expression. Infusion of F13A into PVN of spontaneously hypertensive rats (SHRs) caused chronic decreases in SBP, DBP, MAP, plasma NE level, maximal depressor response to Hex, NAD(P)H oxidase activity, and superoxide anions levels. Hex, a sympathetic ganglion blocker, inhibited apelin-induced increases in SBP, DBP and MAP. SOD overexpression in PVN of SHRs inhibited the apelin-induced increase in SBP, DBP, MAP, plasma NE level, and maximal depressor response to Hex. PVN Nox4 knockdown also attenuated the apelin-induced increase in SBP, DBP, MAP, plasma NE level, and maximal depressor response to Hex. Chronic injection of F13A into PVN reduced fibrosis of renal artery, thoracic aorta, and heart in SHRs. Conclusion: These results demonstrated that in PVN apelin induced long-term high blood pressure and sympathetic activity via increasing oxidative stress.

Purpose: To explore the role and potential mechanism of miR-212-3p in neuropathic pain regulation. Methods: Adult male rats were used to establish chronic constriction injury (CCI) model to mimic the neuropathic pain. Then, paw withdrawal threshold (PWT) and paw withdrawal thermal latency (PWL) were determined. The concentrations of interleukin 1 beta (IL-1β), interleukin 6 (IL-6) and tumor necrosis factor-alpha (TNF-α) were measured with enzyme-linked immune sorbent assay (ELISA) kit and the expression of miR-212-3p was measured by real time quantitative PCR (RTqPCR). Besides, miR-212-3p agomir was intrathecally injected into CCI rats and the expression of key apoptotic proteins was determined by western blot. Furthermore, dual-luciferase reporter assay was used to determine the binding of miR-212-3p and 3’ untranslated regions (3’UTR) of NaV1.3 and the expression levels of NaV1.3 were measured by western blot and RT-qPCR. Results: In the CCI group, the PWT and PWL were significantly decreased and IL-1β, IL-6 and TNF-α were increased. miR-212-3p was decreased in response to CCI. The intrathecal injection of miR-212-3p agomir into CCI rats improved the PWT and PWL, decreased the IL-1β, IL-6 and TNF-α, decreased the expression levels of BCL2 associated X, apoptosis regulator (Bax), cleaved caspase-3 and increased the expression levels of BCL2 apoptosis regulator (Bcl-2). The results of dual--luciferase reporter assay showed that miR-212-3p could directly bind with 3’UTR of NaV1.3. The expression of NaV1.3 was up-regulated in CCI rats who were intrathecally injected with miRctrl, whereas it decreased in CCI rats intrathecally injected with miR-212-3p agomir. Conclusion: The expression of miR-212a-3p attenuates neuropathic pain by targeting NaV1.3.

Propose: To investigate whether miR-22-3p is able to regulate AD development and its molecular mechanism. Methods: Morris water maze test was performed to test the spatial memory. Quantitative polymerase chain reaction (qPCR) was used to assess the expression level of miR-22-3p. The enzymelinked immunosorbent assay (ELISA) was used to assess the levels of Aβ40 and Aβ42. Immunoblotting analysis was performed to detect the protein expression levels of amyloid precursor protein (APP), mitogen-activated protein kinase 14 (MAPK14) and beta-site Amyloid precursor protein Cleaving Enzyme 1 (BACE1). Luciferase assay was used to identify the interaction between miR- 22-3p and MAPK14. The tetrazolium dye (MTT) colorimetric assay was used to test the influence of miR-22-3p overexpression on cell viability. Flow cytometry analysis was performed to evaluate the effect of miR-22-3p overexpression on cell apoptosis. Results: Morris water maze test showed that mice model of AD had impaired spatial memory, which was able to be ameliorated by miR-22-3p overexpression. Immunoblotting analysis revealed that the protein expression levels of APP, MAPK14 and BACE1 were enhanced in AD model, which could be prevented by miR-22-3p overexpression. ELISA showed that Aβ40 and Aβ42 levels were dramatically increased in AD model, which were inhibited by miR-22-3p overexpression. Luciferase assay and immunoblotting analysis indicated that miR-22-3p targeted and regulated MAPK14 expression. Conclusion: MiR-22-3p overexpression reduced Aβ deposit and alleviated AD symptoms by targeting and regulating MAPK14 expression, which ameliorated AD symptoms.

Background: Cerebral small vessel disease (SVD) is an important cause of stroke and vascular cognitive impairment (VCI), leading to subcortical ischemic vascular dementia. As a hereditary form of SVD with early onset, cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) represents a pure form of SVD and may thus serve as a model disease for SVD. To date, underlying molecular mechanisms linking vascular pathology and subsequent neuronal damage in SVD are incompletely understood. Objective: We performed comparative transcriptional profiling microarray and proteomic analyses on post-mortem frontal lobe specimen from 2 CADASIL patients and 5 non neurologically diseased controls in order to identify dysregulated pathways potentially involved in the development of tissue damage in CADASIL. Methods: Transcriptional microarray analysis of material extracted from frontal grey and white matter (WM) identified subsets of up- or down-regulated genes enriched into biological pathways mostly in WM areas. Proteomic analysis of these regions also highlighted cellular processes identified by dysregulated proteins. Results: Discrepancies between proteomic and transcriptomic data were observed, but a number of pathways were commonly associated with genes and corresponding proteins, such as: “ribosome” identified by upregulated genes and proteins in frontal cortex or “spliceosome” associated with down-regulated genes and proteins in frontal WM. Conclusion: This latter finding suggests that defective expression of spliceosomal components may alter widespread splicing profile, potentially inducing expression abnormalities that could contribute to cerebral WM damage in CADASIL.

Background: Recent studies have reported that the levels of serum interleukin-33 (IL- 33) and its receptor, suppression of tumorigenicity 2 (ST2), are potential biomarkers for susceptibility of cardiovascular diseases. However, the genetic association of the IL-33/ST2 axis with cardiovascular diseases remains controversial. Objective: We aimed to investigate the association between common variants in the IL-33/ST2 axis and ischemic stroke in the Han Chinese population. Methods: We consecutively enrolled 1166 patients with ischemic stroke and 1079 age- and gender- matched controls. Eight single nucleotide polymorphisms (SNPs) within IL-33/ST2 axis were genotyped using the improved Multiple Ligase Detection Reaction platform. We analyzed the association between the tested SNPs and ischemic stroke at both the genotype and haplotype levels. Results: Binary logistic regression analysis indicated that rs10435816 (additive model: odds ratio [OR]=0.72, 95% confidence interval [CI], 0.54-0.95; recessive model: OR=0.72, 95%CI, 0.56- 0.94) was associated with a decreased risk of ischemic stroke after adjustment of confounding factors. Subgroup analysis indicated that rs10435816 (additive model: OR=0.61, 95%CI, 0.41-0.89; recessive model: OR=0.56, 95%CI, 0.40-0.80), rs7025417 (additive model: OR=0.57, 95%CI, 0.39-0.83), rs11792633 (additive model: OR=0.66, 95%CI, 0.46-0.95; recessive model: OR=0.67, 95%CI, 0.49-0.93), and rs7044343 (additive model: OR=0.69, 95%CI, 0.48-0.97; recessive model: OR=0.67, 95%CI, 0.49-0.91) were associated with a decreased risk of large-artery atherosclerosis stroke after adjustment of confounding factors. Conclusion: Our findings suggested an association between common variants in the IL-33/ST2 axis and a decreased risk of ischemic stroke in the Han Chinese population.

The neurovascular unit is a physiological unit present in the brain, which is constituted by elements of the nervous system (neurons and astrocytes) and the vascular system (endothelial and mural cells). This unit is responsible for the homeostasis and regulation of cerebral blood flow. There are two major types of mural cells in the brain, pericytes and smooth muscle cells. At the arterial level, smooth muscle cells are the main components that wrap around the outside of cerebral blood vessels and the major contributors to basal tone maintenance, blood pressure and blood flow distribution. They present several mechanisms by which they regulate both vasodilation and vasoconstriction of cerebral blood vessels and their regulation becomes even more important in situations of injury or pathology. In this review, we discuss the main regulatory mechanisms of brain smooth muscle cells and their contributions to the correct brain homeostasis.