丁苯酞对缺血性脑卒中大鼠脂联素、MCP-1及slCAM-1的影响

目的:探讨丁苯酚对卒中大鼠脂联素、MCP-1和sl CAM-1表达的影响。方法:建立大鼠永久性大脑中动脉梗塞(Permanent middle cerebral occlusion, pMCAO)局灶性脑缺血模型,将SD大鼠分为Sham组、pMCAO模型组和NBP治疗组;甲酚紫染色法测定大鼠脑部梗塞面积;改良神经功能损害评分(Modified neurological severity score, m NSS)用于评定各组大鼠的神经功能变化情况;Elisa法检测大鼠脑部和血清中脂联素水平以及脑部MCP-1和sl CAM-1水平。结果:药物处理后pMCAO组大鼠存活率明显低于Sham组;NBP处理能够有效逆转pMCAO诱导的大鼠m NSS得分的升高;与Sham组相比,pMCAO组大鼠脑部梗塞面积显著增加,NBP处理明显减少了损伤大鼠的梗塞范围;与Sham组相比,p MCAO组大鼠脑部和血清中脂联素表达明显降低,而大脑MCP-1及sl CAM-1水平显著升高,NBP处理上调了大脑和血清脂联素的水平并抑制脑部MCP-1和sl CAM-1的表达。结论:丁苯酞通过上调脂联素表达,抑制MCP-1及sl CAM-1的水平对缺血性脑卒中大鼠起到神经保护作用。     

Neuroprotective Effect of Baicalin in a Rat Model of Permanent Focal Cerebral Ischemia

This investigation was performed to determine the neuroprotective effect of baicalin on permanent cerebral ischemia injury in rats and the potential mechanisms in this process. Adult male Sprague-Dawley rats underwent permanent middle cerebral artery occlusion (pMCAO). The rats were then received intraperitoneal injection with baicalin (10, 30 and 100 mg/kg) or vehicle. Morphological characteristic, neurological deficit scores, cerebral infarct volume and the enzymatic activity of myeloperoxidase (MPO) were measured 24 h after pMCAO. The mRNA expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) were determined by RT-PCR. Neuronal apoptosis was determined by TUNEL staining and Western blot. Baicalin (30 and 100 mg/kg) reduced neurological deficit scores and cerebral infarct volume 24 h after pMCAO. Baicalin significantly decreased the enzymatic activity of MPO and the expression of iNOS mRNA and COX-2 mRNA in rat brain, it also significantly inhibited neuronal apoptosis and the expression of cleaved caspase-3 protein after pMCAO. Our results suggested that baicalin possesses potent anti-inflammatory and anti-apoptotic properties and attenuates cerebral ischemia injury. This protection might be associated with the downregulated expression of iNOS mRNA, COX-2 mRNA, and cleaved caspase-3 protein.     

Synthesis and biological evaluation of novel hydrogen sulfide releasing nicotinic acid derivatives

Twelve novel hybrids of slowly releasing hydrogen sulfide donor ADT-OH combined with nicotinic acid were synthesized. All of their structures had been confirmed by ¹H NMR, ¹³C NMR and MS spectra. The target compounds were evaluated for their neuroprotective effects on hippocampal neuron HT22 cells against glutamate-induced injury at the concentrations of 1–100 μM with MTT assay, and their toxicity on HT22 cells untreated by glutamine at the concentration of 100 μM. The active compound was further investigated for its effect on ischemic infarct volume by intraperitoneal injection at 3 h after ischemia in mice models of permanent middle cerebral artery occlusion (pMCAO). The results showed that all the compounds significantly protected HT22 cells from glutamate-induced damage at most of the experimental concentrations, and had no or little neurotoxicity on normal HT22 cells at the high concentration. More importantly, compound A6 significantly reduced infarct volume in the pMCAO model. These results suggested that compound A6 may be promising for further evaluation for the intervention of cerebral ischemic injury.     

Acquisition of tolerance to Δ-9-THC as measured by the response of a cellular function

The development of tolerance to delta-9-tetrahydrocannabinol (Δ-9-THC) was investigated by measuring respiration in brain tissue after acute or chronic administration. Mice were given either single or seven daily repeated intraperitoneal injections of 50 mg/Kg of delta-9-tetrahydrocannabinol (Δ-9-THC) or control vehicle. The final injection for all drug treated animals included radiolabeled ³H-Δ-9-THC. The mice were sacrificed at 1 hour, 2 hours, 4 hours, 24 hours, and 7 days after the final injection. Δ-9-THC depressed respiration, but after repeated injections was significantly less effective in this regard, indicating acquisition of tolerance to Δ-9-THC. Because the concentration of radiolabeled cannabinoids in brain tissue from each group is not appreciably different, a cellular as opposed to distributional mode of tolerance is suggested.     

Synthesis of taurine in rat liver and brain in vivo

The in vivo formation of taurine and the analysis of labeled taurine precursors was examined in rat brain and liver at different times after an intracisternal injection of [³⁵S]cysteine and an intraperitoneal injection of [³H]cysteine, simultaneously administered. The distribution pattern of radioactivity was similar in liver and brain. Most of the labeling in both organs (85% in brain and 80% in liver) was recovered in glutathione (oxidized and reduced), cysteic acid, cysteine sulfinic acid, hypotaurine, cystathionine, and a mixed disulfide of cysteine and glutathione. The relative rates of labeling of cysteine sulfinic acid and taurine in liver and brain suggest than in vivo, liver possesses a higher capacity for taurine synthesis than brain. A small amount of [³H]taurine was detected in brain after intraperitoneal injection of [³H]cysteine. The time of appearance of this [³H]taurine as well as the fact that it occurs when [³H]cysteine is not detectable in brain or plasma suggests that it was probably not synthesized in brain from labeled precursors but formed elsewhere and transported into the brain through an exchange process.     

The Antiepileptic Drug Levetiracetam Suppresses Non-Convulsive Seizure Activity and Reduces Ischemic Brain Damage in Rats Subjected to Permanent Middle Cerebral Artery Occlusion

The antiepileptic drug Levetiracetam (Lev) has neuroprotective properties in experimental stroke, cerebral hemorrhage and neurotrauma. In these conditions, non-convulsive seizures (NCSs) propagate from the core of the focal lesion into perilesional tissue, enlarging the damaged area and promoting epileptogenesis. Here, we explore whether Lev neuroprotective effect is accompanied by changes in NCS generation or propagation. In particular, we performed continuous EEG recordings before and after the permanent occlusion of the middle cerebral artery (pMCAO) in rats that received Lev (100 mg/kg) or its vehicle immediately before surgery. Both in Lev-treated and in control rats, EEG activity was suppressed after pMCAO. In control but not in Lev-treated rats, EEG activity reappeared approximately 30-45 min after pMCAO. It initially consisted in single spikes and, then, evolved into spike-and-wave and polyspike-and-wave discharges. In Lev-treated rats, only rare spike events were observed and the EEG power was significantly smaller than in controls. Approximately 24 hours after pMCAO, EEG activity increased in Lev-treated rats because of the appearance of polyspike events whose power was, however, significantly smaller than in controls. In rats sacrificed 24 hours after pMCAO, the ischemic lesion was approximately 50% smaller in Lev-treated than in control rats. A similar neuroprotection was observed in rats sacrificed 72 hours after pMCAO. In conclusion, in rats subjected to pMCAO, a single Lev injection suppresses NCS occurrence for at least 24 hours. This electrophysiological effect could explain the long lasting reduction of ischemic brain damage caused by this drug.     

Citicoline Protects Brain Against Closed Head Injury in Rats Through Suppressing Oxidative Stress and Calpain Over-Activation

Citicoline, a natural compound that functions as an intermediate in the biosynthesis of cell membrane phospholipids, is essential for membrane integrity and repair. It has been reported to protect brain against trauma. This study was designed to investigate the protective effects of citicoline on closed head injury (CHI) in rats. Citicoline (250 mg/kg i.v. 30 min and 4 h after CHI) lessened body weight loss, and improved neurological functions significantly at 7 days after CHI. It markedly lowered brain edema and blood–brain barrier permeability, enhanced the activities of superoxide dismutase and the levels of glutathione, reduced the levels of malondialdehyde and lactic acid. Moreover, citicoline suppressed the activities of calpain, and enhanced the levels of calpastatin, myelin basic protein and αII-spectrin in traumatic tissue 24 h after CHI. Also, it attenuated the axonal and myelin sheath damage in corpus callosum and the neuronal cell death in hippocampal CA1 and CA3 subfields 7 days after CHI. These data demonstrate the protection of citicoline against white matter and grey matter damage due to CHI through suppressing oxidative stress and calpain over-activation, providing additional support to the application of citicoline for the treatment of traumatic brain injury.     

Inactivation of mouse transmembrane prolyl 4-hydroxylase increases blood brain barrier permeability and ischemia-induced cerebral neuroinflammation

Hypoxia-inducible factor prolyl 4-hydroxylases (HIF-P4Hs) regulate the hypoxic induction of >300 genes required for survival and adaptation under oxygen deprivation. Inhibition of HIF-P4H-2 has been shown to be protective in focal cerebral ischemia rodent models, while that of HIF-P4H-1 has no effects and inactivation of HIF-P4H-3 has adverse effects. A transmembrane prolyl 4-hydroxylase (P4H-TM) is highly expressed in the brain and contributes to the regulation of HIF, but the outcome of its inhibition on stroke is yet unknown. To study this, we subjected WT and P4htm^−/− mice to permanent middle cerebral artery occlusion (pMCAO). Lack of P4H-TM had no effect on lesion size following pMCAO, but increased inflammatory microgliosis and neutrophil infiltration was observed in the P4htm^−/− cortex. Furthermore, both the permeability of blood brain barrier and ultrastructure of cerebral tight junctions were compromised in P4htm^−/− mice. At the molecular level, P4H-TM deficiency led to increased expression of proinflammatory genes and robust activation of protein kinases in the cortex, while expression of tight junction proteins and the neuroprotective growth factors erythropoietin and vascular endothelial growth factor was reduced. Our data provide the first evidence that P4H-TM inactivation has no protective effect on infarct size and increases inflammatory microgliosis and neutrophil infiltration in the cortex at early stage after pMCAO. When considering HIF-P4H inhibitors as potential therapeutics in stroke, the current data support that isoenzyme-selective inhibitors that do not target P4H-TM or HIF-P4H-3 would be preferred.     

Evaluation of the Neuroprotective Effects of Citicoline after Experimental Spinal Cord Injury: Improved Behavioral and Neuroanatomical Recovery

Spinal cord injury (SCI) caused by trauma mainly occurs in two mechanisms as primary and secondary injury. Secondary injury following the primary impact includes various pathophysiological and biochemical events. Methylprednisolone is the only pharmacological agent having clinically proven beneficial effects on SCI. Citicoline has been shown to have clinical and experimental beneficial effects on brain ischemia. This study aims to investigate the neuroprotective effect of citicoline in an experimental SCI model in rats. Sixty adult Wistar albino rats were randomized into five groups. SCI was performed by the weight-drop model. Group 1 underwent laminectomy alone. The Group 2 underwent laminectomy followed by SCI and received no medication. Group3, Group 4 and Group 5 underwent laminectomy followed by SCI and received medication. Group 3 and Group 5 received citicoline and Group 4 and Group 5 received methylprednisolone. The rats were divided into two subgroups for biochemical analysis (sacrificed at 24 h after surgery) and neurobehavioral and histopathological evaluation (sacrificed at 6 weeks after surgery). Malonildialdehyde levels, nitric oxide levels and trauma size ratios were lower and reduced glutathione levels were higher in Group 3, Group 4 and Group 5 as compared to Group 2. Posttraumatic neurological recovery after surgery was significantly better in Group 3, Group 4 and Group 5 compared to Group 2. In conclusion, this study demonstrates that citicoline is as effective as methylprednisolone. The efficacy of citicoline combined with methylprednisolone is not superior to either citicoline or methylprednisolone alone.     

Activation of α-7 Nicotinic Acetylcholine Receptor Reduces Ischemic Stroke Injury through Reduction of Pro-Inflammatory Macrophages and Oxidative Stress

Activation of α-7 nicotinic acetylcholine receptor (α-7 nAchR) has a neuro-protective effect on ischemic and hemorrhagic stroke. However, the underlying mechanism is not completely understood. We hypothesized that α-7 nAchR agonist protects brain injury after ischemic stroke through reduction of pro-inflammatory macrophages (M1) and oxidative stress. C57BL/6 mice were treated with PHA568487 (PHA, α-7 nAchR agonist), methyllycaconitine (MLA, nAchR antagonist), or saline immediately and 24 hours after permanent occlusion of the distal middle cerebral artery (pMCAO). Behavior test, lesion volume, CD68⁺, M1 (CD11b⁺/Iba1⁺) and M2 (CD206/Iba1⁺) microglia/macrophages, and phosphorylated p65 component of NF-kB in microglia/macrophages were quantified using histological stained sections. The expression of M1 and M2 marker genes, anti-oxidant genes and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase were quantified using real-time RT-PCR. Compared to the saline-treated mice, PHA mice had fewer behavior deficits 3 and 7 days after pMCAO, and smaller lesion volume, fewer CD68⁺ and M1 macrophages, and more M2 macrophages 3 and 14 days after pMCAO, whereas MLA''s effects were mostly the opposite in several analyses. PHA increased anti-oxidant genes and NADPH oxidase expression associated with decreased phosphorylation of NF-kB p65 in microglia/macrophages. Thus, reduction of inflammatory response and oxidative stress play roles in α-7 nAchR neuro-protective effect.     

Experimental treatment of Neospora caninum-infected mice with the arylimidamide DB750 and the thiazolide nitazoxanide

The cationic arylimidamide DB750 and the thiazolide nitazoxanide had been shown earlier to be effective against Neospora caninum tachyzoites in vitro with an IC₅₀ of 160 nM and 4.23 μM, respectively. In this study, we have investigated the effects of DB750 and nitazoxanide treatments of experimentally infected Balb/c mice, by applying the drugs either through the oral or the intraperitoneal route. In experiment 1, administration of DB750 (2 mg/kg/day) and nitazoxanide (150 mg/kg/day) started already 3 days prior to experimental infection of mice with 2 × 10⁶ tachyzoites. Following infection, the drugs were further administrated daily for a period of 2 weeks, either orally or intraperitoneally. Intraperitoneal injection of DB750 was well tolerated by the mice, but treatment with nitazoxanide resulted in death of all mice within 3 days. Upon intraperitoneal application of DB750, the cerebral parasite load was significantly reduced compared to all other groups, while oral application of DB750 and nitazoxanide were not as effective, and resulted in significant weight loss. In experiment 2, mice were infected with 2 × 10⁶ tachyzoites and at 2 weeks post-infection, DB750 (2 mg/kg/day) was applied by intraperitoneal injections for 14 days. In the DB750-treated group, only 2 out of 12 mice succumbed to infection, compared to 7 out of 12 mice in the placebo-group. DB750 treatment also resulted in significantly reduced cerebral parasite burden, and reduced numbers of viable tachyzoites. Our data suggest that DB750 exerted its activity also after crossing the blood–brain barrier, and that this class of compounds could be promising for the control of N. caninum-associated disease.     

Citicoline: neuroprotective mechanisms in cerebral ischemia.

Cytidine-5'-diphosphocholine (citicoline or CDP-choline), an intermediate in the biosynthesis of phosphatidylcholine (PtdCho), has shown beneficial effects in a number of CNS injury models and pathological conditions of the brain. Citicoline improved the outcome in several phase-III clinical trials of stroke, but provided inconclusive results in recent clinical trials. The therapeutic action of citicoline is thought to be caused by stimulation of PtdCho synthesis in the injured brain, although the experimental evidence for this is limited. This review attempts to shed some light on the properties of citicoline that are responsible for its effectiveness. Our studies in transient cerebral ischemia suggest that citicoline might enhance reconstruction (synthesis) of PtdCho and sphingomyelin, but could act by inhibiting the destructive processes (activation of phospholipases). Citicoline neuroprotection may include: (i) preserving cardiolipin (an exclusive inner mitochondrial membrane component) and sphingomyelin; (ii) preserving the arachidonic acid content of PtdCho and phosphatidylethanolamine; (iii) partially restoring PtdCho levels; (iv) stimulating glutathione synthesis and glutathione reductase activity; (v) attenuating lipid peroxidation; and (vi) restoring Na(+)/K(+)-ATPase activity. These observed effects of citicoline could be explained by the attenuation of phospholipase A(2) activation. Based on these findings, a singular unifying mechanism has been hypothesized. Citicoline also provides choline for synthesis of neurotransmitter acetylcholine, stimulation of tyrosine hydroxylase activity and dopamine release.     

Melatonin stimulates antioxidant enzymes and reduces oxidative stress in experimental traumatic brain injury: the Nrf2–ARE signaling pathway as a potential mechanism

The goal of this study was to evaluate the potential involvement of melatonin in the activation of the nuclear factor erythroid 2-related factor 2 and antioxidant-responsive element (Nrf2–ARE) signaling pathway and the modulation of antioxidant enzyme activity in an experimental model of traumatic brain injury (TBI). In experiment 1, ICR mice were divided into four groups: sham group, TBI group, TBI + vehicle group, and TBI + melatonin group (n = 38 per group). Melatonin (10 mg/kg) was administered via an intraperitoneal (ip) injection at 0, 1, 2, 3, and 4 h post-TBI. In experiment 2, Nrf2 wild-type (Nrf2^+/+ group) and Nrf2-knockout (Nrf2^−/− group) mice received a TBI insult followed by melatonin administration (10 mg/kg, ip) at the corresponding time points (n = 35 per group). The administration of melatonin after TBI significantly ameliorated the effects of the brain injury, such as oxidative stress, brain edema, and cortical neuronal degeneration. Melatonin markedly promoted the translocation of Nrf2 protein from the cytoplasm to the nucleus; increased the expression of Nrf2–ARE pathway-related downstream factors, including heme oxygenase-1 and NAD(P)H:quinone oxidoreductase 1; and prevented the decline of antioxidant enzyme activities, including superoxide dismutase and glutathione peroxidase. Furthermore, knockout of Nrf2 partly reversed the neuroprotection of melatonin after TBI. In conclusion, melatonin administration may increase the activity of antioxidant enzymes and attenuate brain injury in a TBI model, potentially via mediation of the Nrf2–ARE pathway.     

Effect of hypoglycemia on the brain free fatty acid level and the uptake of fatty acids by phospholipids

The effect of hypoglycemia on the uptake of [1-¹⁴C]arachidonate and [1-¹⁴C]oleate into a synaptosomal and microsomal glycerophospholipids was investigated. In the presence of ATP, Mg²⁺ and CoA, rat brain synaptosomes and micorsomes catalyze the transfer of arachidonate and oleatc into glycerophospholipids. Arachidonate was mainly incorporated into phosphatidylinositol (PI) and phosphatidylcholine (PC), whereas oleate was incorporated into phosphatidylcholine and phosphatidylethanolamine (PE).Hypoglycemia was produced by intraperitoneal injection of 10 or 100 units of crystalline insulin per kg body weight. Two hours after injection the blood glucose level decreased to 10–20 mg%. The content of brain phospholipids was slightly decreased but the change was not statistically significant. The level of free fatty acids (FFA) was increased. More pronounced and reproducible changes were found when hypoglycemia was produced by injection of 100 units of insulin per/kg body weight. Changes in brain cortex were similar to those observed in microsomes and synaptosomes. Hypoglycemia affected the incorporation of arachidonic acid into glycerophospholipids of brain membranes. Uptake of [1-¹⁴C]arachidonate was decreased selectively by 50% (into phosphatidic acid /PA/) when hypogiycemia was produced by injection of 10 units of insulin per kg body weight. The Higher dose of insulin 100 units per kg body weight produced a 20% inhibition of arachidonate incorporation into synaptosomal PI and a 13% decrease of incorporation into microsomal phosphatidylcholine. Incorporation of [1-¹⁴C]oleate into membrane phospholipids was not changed by hypoglycemic insult. It is proposed that the disturbances in fatty acid level, particularly arachidonate, and decreased uptake of arachidonic acid by synaptosomal glycerophospholipids may be responsible for alteration of membrane function and changes of synaptic processes.     

A calcium channel blocker flunarizine attenuates the neurotoxic effects of iron

Iron is a metal highly concentrated in liver and brain tissue, and known to induce neuronal hyperactivity and oxidative stress. It has been established that iron levels rise in the brain in some neurodegenerative diseases such as Parkinson's and Alzheimer's diseases (AD). A body of evidence indicates a link between neuronal death and intracellular excessive calcium accumulation. The aim of the present study was to investigate the effects of a calcium antagonist, flunarizine, on neurotoxicity induced by intracerebroventricular (i.c.v.) iron injection. For this reason rats were divided into three groups as control, iron and iron+flunarizine groups. Animals in iron and iron+flunarizine groups received i.c.v. FeCl₃ injection (200 mM, 2.5 μl), while control rats received the same amount of saline into the cerebral ventricles. Rats in iron+flunarizine group also received i.c.v. flunarizine (1 μM, 2 μl) following FeCl₃ injection. All animals were kept alive for ten days following the operation and animals in iron+flunarizine group received intraperitoneal (i.p.) flunarizine injections once a day (10 mg/kg/day) during this period. After ten days, rats were sacrificed. The total numbers of neurons in hippocampus of all rats were estimated with the latest, unbiased stereological techniques. Findings of the present study suggest that flunarizine may attenuate the neurotoxic effects of iron injection by inhibiting the cellular influx of excessive calcium ions.     

银杏内酯注射液联合胞磷胆碱钠片对脑梗死恢复期患者脑血流动力学、氧化应激和血清MCP-1、Lp-PLA2的影响

摘要 目的：研究银杏内酯注射液联合胞磷胆碱钠片对脑梗死恢复期患者脑血流动力学、氧化应激和血清单核细胞趋化蛋白-1（MCP-1）、脂蛋白相关磷脂酶A2（Lp-PLA2）的影响。方法：按照随机数字表法,将安徽中医药大学第一附属医院146例脑梗死恢复期患者分为对照组（n=73,采用常规治疗和胞磷胆碱钠片治疗）和研究组（n=73,对照组基础上结合银杏内酯注射液）。对比两组临床疗效、Barthel指数（BI）评分、美国国立卫生研究院卒中量表（NIHSS）评分、脑血流动力学指标、血清氧化应激指标、MCP-1、Lp-PLA2和用药安全性。结果：与对照组的82.19%临床总有效率对比,研究组的97.26%更高（P<0.05）。治疗后,研究组BI评分、平均血流速度（Vm）、血清超氧化物歧化酶（SOD）、过氧化氢酶（CAT）较对照组更高,NIHSS评分、搏动指数（PI）、阻力指数（RI）、血清活性氧（ROS）、血清MCP-1、Lp-PLA2较对照组更低（P<0.05）。两组不良反应发生率组间对比未见差异（P>0.05）。结论：银杏内酯注射液联合胞磷胆碱钠片可减轻脑梗死恢复期患者的神经功能损伤,改善患者的脑血流动力学,减轻氧化应激,调节血清MCP-1、Lp-PLA2水平,且用药安全性良好。     

The neuroprotective effect of mesenchymal stem cells on an experimentally induced model for multiple sclerosis in mice

Multiple sclerosis (MS) is a chronic autoimmune demyelinating neurodegenerative central nervous system disorder. The aim of the present study was to investigate the prophylactic effect exerted by the one‐time intraperitoneal injection of mesenchymal stem cells (MSCs) 1 × 10⁶ and 14‐day intraperitoneal injection of methylprednisolone (MP) 40 mg/kg in an experimental autoimmune encephalomyelitis (EAE). EAE was induced by intradermal injection of rat spinal cord homogenate with complete Freund's adjuvant in Swiss mice. Results of MSCs and MP‐treated mice showed a significantly milder disease and fewer clinical scores compared to control mice. They suppressed tumor necrosis factor‐alpha and myeloperoxidase and increased interleukin 10, whereas thiobarbituric acid reactive substances and nitric oxide brain contents were reduced to comparable levels between treatment groups. Brain content of GSH was significantly higher in MSCs‐treated mice than control mice. It is evident that MSCs have relevant prophylactic effect in an animal model of MS and might represent a valuable tool for stem cell based therapy in MS.     

Carbonic anhydrase inhibition for the management of cerebral ischemia: in vivo evaluation of sulfonamide and coumarin inhibitors

Ischemia of brain areas is a global health problem, causing death or long-term disability. Current pharmacological options have limited impact on ischemic damages. Recently, a relationship between hypoxia and carbonic anhydrase (CA) over-expression has been highlighted suggesting CA inhibition as a possible target. This study aimed to evaluate the pharmacological profile of sulfonamide and coumarin CA inhibitors in rats underwent permanent middle cerebral artery occlusion (pMCAO). The neurological score of pMCAO rats was dramatically reduced 24?h after occlusion. Repeated subcutaneous injections of the CA inhibitors 4 and 7 (1?mg kg^?1) were able to increase the neurological score by 40%. Compound 7 showed the tendency to reduce the volume of hemisphere infarction. The standard CA inhibitor acetazolamide was ineffective. The properties of novel CA inhibitors to improve neurological functionalities after cerebral ischemic insult are shown. The CA involvement in cerebral hypoxic phenomena deserves deeper investigations.     

Sphingosine-1-Phosphate Receptor-1 Selective Agonist Enhances Collateral Growth and Protects against Subsequent Stroke

Background and Purpose

Collateral growth after acute occlusion of an intracranial artery is triggered by increasing shear stress in preexisting collateral pathways. Recently, sphingosine-1-phosphate receptor-1 (S1PR1) on endothelial cells was reported to be essential in sensing fluid shear stress. Here, we evaluated the expression of S1PR1 in the hypoperfused mouse brain and investigated the effect of a selective S1PR1 agonist on leptomeningeal collateral growth and subsequent ischemic damage after focal ischemia.

Methods

In C57Bl/6 mice (n = 133) subjected to unilateral common carotid occlusion (CCAO) and sham surgery. The first series examined the time course of collateral growth, cell proliferation, and S1PR1 expression in the leptomeningeal arteries after CCAO. The second series examined the relationship between pharmacological regulation of S1PR1 and collateral growth of leptomeningeal anastomoses. Animals were randomly assigned to one of the following groups: LtCCAO and daily intraperitoneal (ip) injection for 7 days of an S1PR1 selective agonist (SEW2871, 5 mg/kg/day); sham surgery and daily ip injection for 7 days of SEW2871 after surgery; LtCCAO and daily ip injection for 7 days of SEW2871 and an S1PR1 inverse agonist (VPC23019, 0.5 mg/kg); LtCCAO and daily ip injection of DMSO for 7 days after surgery; and sham surgery and daily ip injection of DMSO for 7 days. Leptomeningeal anastomoses were visualized 14 days after LtCCAO by latex perfusion method, and a set of animals underwent subsequent permanent middle cerebral artery occlusion (pMCAO) 7days after the treatment termination. Neurological functions 1hour, 1, 4, and 7days and infarction volume 7days after pMCAO were evaluated.

Results

In parallel with the increase in S1PR1 mRNA levels, S1PR1 expression colocalized with endothelial cell markers in the leptomeningeal arteries, increased markedly on the side of the CCAO, and peaked 7 days after CCAO. Mitotic cell numbers in the leptomeningeal arteries increased after CCAO. Administration of the S1PR1 selective agonist significantly increased cerebral blood flow (CBF) and the diameter of leptomeningeal collateral vessels (42.9 ± 2.6 μm) compared with the controls (27.6 ± 5.7 μm; P < 0.01). S1PR1 inverse agonist administration diminished the effect of the S1PR1 agonist (P < 0.001). After pMCAO, S1PR1 agonist pretreated animals showed significantly smaller infarct volume (17.5% ± 4.0% vs. 7.7% ± 4.0%, P < 0.01) and better functional recovery than vehicle-treated controls.

Conclusions

These results suggest that S1PR1 is one of the principal regulators of leptomeningeal collateral recruitment at the site of increased shear stress and provide evidence that an S1PR1 selective agonist has a role in promoting collateral growth and preventing of ischemic damage and neurological dysfunction after subsequent stroke in patients with intracranial major artery stenosis or occlusion.