GV1001 modulates neuroinflammation and improves memory and behavior through the activation of gonadotropin-releasing hormone receptors in a triple transgenic Alzheimer's disease mouse model.

GV1001 protects neural cells from amyloid-ß (Aß) toxicity and other stressors in in vitro studies and demonstrates clinically beneficial effects in patients with moderate to severe Alzheimer's disease (AD). Here, we investigated the protective effects and mechanism of action of GV1001 in triple transgenic AD (3xTg-AD) mice. We found that GV1001 improved memory and cognition in middle- and old-aged 3xTg-AD mice. Additionally, it reduced Aß oligomer and phospho-tau (Ser202 and Thr205) levels in the brain, and mitigated neuroinflammation by promoting a neuroprotective microglial and astrocyte phenotype while diminishing the neurotoxic ones. In vitro, GV1001 bound to gonadotropin releasing hormone receptors (GnRHRs) with high affinity. Levels of cyclic adenosine monophosphate, a direct downstream effector of activated GnRHRs, increased after GV1001 treatment. Furthermore, inhibition of GnRHRs blocked GV1001-induced effects. Thus, GV1001 might improve cognitive and memory functions of 3xTg-AD mice by suppressing neuroinflammation and reducing Aß oligomers levels and phospho-tau by activating GnRHRs and their downstream signaling pathways.

Effects of aspirin and clopidogrel on neural stem cells.

Cerebral infarction causes severe morbidity and mortality. Most patients with cerebral infarction should take antiplatelet drugs daily, so the effects of those drugs on the regeneration of the brain need to be investigated. Aspirin and clopidogrel are the most widely used antiplatelet drugs for the prevention of ischemic stroke. We investigated the effects of aspirin and clopidogrel on neural stem cells (NSCs). NSCs were dissociated from fetal rat cortex and cultured with basic fibroblast growth factor and N2 medium. To measure the effects of aspirin and clopidogrel on NSCs, NSCs were treated with several concentrations of aspirin, clopidogrel bisulfate, and clopidogrel resinate for 24 h. After the treatment, we measured cell viability by cell counting kit-8, MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, trypan blue staining, flow cytometry, and lactate dehydrogenase assay. To evaluate their effects on NSC proliferation, we performed BrdU cell proliferation assay and colony-forming unit assay. We compared the intracellular protein level in the NSCs treated with aspirin and two types of clopidogrel, by proteomics analysis. Various viability tests showed that clopidogrel resinate and clopidogrel bisulfate did not affect the viability and proliferation of NSCs whereas aspirin decreased them even at low concentrations which are clinically relevant. Moreover, through the proteomics, it was confirmed that the toxicity of aspirin to NSCs might be associated with the alteration of several intracellular proteins. Taken together, these results suggest that clopidogrel resinate and clopidogrel bisulfate are safe but aspirin could be toxic to NSCs. Therefore, when these antiplatelet agents are prescribed over the long-term, the finding that aspirin could be toxic to NSCs should be considered.

Activation of the phosphatidylinositol 3-kinase pathway plays important roles in reduction of cerebral infarction by cilnidipine.

Cerebral infarction causes permanent neuronal loss inducing severe morbidity and mortality. Because hypertension is the main risk factor for cerebral infarction and most patients with hypertension take antihypertensive drugs daily, the neuroprotective effects and mechanisms of anti-hypertensive drugs need to be investigated. Cilnidipine, a long-acting, new generation 1,4-dihydropyridine inhibitor of both L- and N-type calcium channels, was reported to reduce oxidative stress. In this study, we investigated whether cilnidipine has therapeutic effects in an animal model of cerebral infarction. After determination of the most effective dose of cilnidipine, a total of 128 rats were subjected to middle cerebral artery occlusion. Neurobehavioral function test and brain MRI were performed, and rats with similar sized infarcts were randomized to either the cilnidipine group or the control group. Cilnidipine treatment was performed with reperfusion after 2-h occlusion. Western blots and immunohistochemistry were also performed after 24-h occlusion. Initial infarct volume on diffusion-weighted MRI was not different between the cilnidipine group and the control group; however, fluid-attenuated inversion recovery MRI at 24 h showed significantly reduced infarct volume in the cilnidipine group compared with the control group. Cilnidipine treatment significantly decreased the number of triphosphate nick end labeling-positive cells compared to the control group. Western blot and immunohistochemistry showed increased expression of phosphorylated Akt (Ser473), phosphorylated glycogen synthase kinase-3ß, and Bcl-2 and decreased expression of Bax and cleaved caspase-3. These results suggest that cilnidipine, which is used for the treatment of hypertension, has neuroprotective effects in the ischemic brain through activation of the PI3K pathway. We investigated whether cilnidipine has neuroprotective effects on ischemic stroke in an animal model. We have demonstrated that the neuroprotective effect of cilnidipine is associated with the activation of the PI3K pathway. Considering the daily use of antihypertensive drugs for patients with hypertension, cilnidipine could be beneficial for patients with ischemic stroke.

Increased VEGF and decreased SDF-1α in patients with silent brain infarction are associated with better prognosis after first-ever acute lacunar stroke.

BACKGROUND: Pre-existing silent brain infarctions (SBIs) have been reported to be associated with better outcomes after first-ever symptomatic ischemic stroke, although the mechanism of this remains unclear. We investigated the association between SBIs, outcomes of acute lacunar infarction, and biomarkers including vascular endothelial growth factor (VEGF), stromal cell-derived factor-1α (SDF-1α), macrophage migration inhibitory factor (MIF), and high-mobility group box-1 (HMGB1). METHODS: A total of 68 consecutive patients diagnosed with first-ever lacunar infarction (<20 mm) within 24 hours of symptom onset were included in this study. Clinical, laboratory, and imaging data were obtained. Plasma levels of VEGF, SDF-1α, MIF, and HMGB1 were assessed using Enzyme-Linked Immunosorbent Assay kits. RESULTS: SBIs were noted in 31 of the 68 patients. Although the initial National Institutes of Health Stroke Scale scores were not related with the presence of SBIs (P = .313), patients with SBIs had better outcomes at 3 months (P = .029). Additionally, plasma VEGF levels were higher (P = .035) and SDF-1α levels were lower (P < .001) in patients with SBIs. Logistic regression analysis indicated that VEGF and SDF-1α were independently associated with the presence of SBIs. CONCLUSIONS: SBIs are associated with favorable outcomes in patients with first-ever acute lacunar infarction and higher levels of VEGF, and lower levels of SDF-1α in these patients may contribute to their more favorable prognosis.

GV1001 reduces neurodegeneration and prolongs lifespan in 3xTg-AD mouse model through anti-aging effects.

GV1001, which mimics the activity of human telomerase reverse transcriptase, protects neural cells from amyloid beta (Aß) toxicity and other stressors through extra-telomeric function, as noted in our prior in vitro studies. As per a recent phase II clinical trial, it improves cognitive function in patients with moderate to severe dementia. However, the underlying protective mechanisms remain unclear. This study aimed to investigate the effects of GV1001 on neurodegeneration, senescence, and survival in triple transgenic Alzheimer's disease (3xTg-AD) mice. GV1001 (1 mg/kg) was subcutaneously injected into old 3xTg-AD mice thrice a week until the endpoint for sacrifice, and survival was analysed. Magnetic resonance imaging (MRI) and Prussian blue staining (PBS) were performed to evaluate entry of GV1001 entrance into the brain. Diverse molecular studies were performed to investigate the effect of GV1001 on neurodegeneration and cellular senescence in AD model mice, with a particular focus on BACE, amyloid beta1-42 (Aß1-42), phosphorylated tau, volume of dentate gyrus, ß-galactosidase positive cells, telomere length, telomerase activity, and ageing-associated proteins. GV1001 crossed the blood-brain barrier, as confirmed by assessing the status of ferrocenecarboxylic acid-conjugated GV1001 using magnetic resonance imaging and PBS. GV1001 increased the survival of 3xTg-AD mice. It decreased BACE and Aß1-42 levels, neurodegeneration (i.e., reduced CA1, CA3 and dentate gyrus volume, decreased levels of senescence-associated ß-galactosidase positive cells, and increased telomere length and telomerase activity), and levels of ageing-associated proteins. We suggest that GV1001 exerts anti-ageing effects in 3xTg-AD mice by reducing neurodegeneration and senescence, which contributes to improved survival.

Effects of GV1001 on Language Dysfunction in Patients With Moderate-to-Severe Alzheimer's Disease: Post Hoc Analysis of Severe Impairment Battery Subscales.

Background and Purpose: The efficacy and safety of GV1001 have been demonstrated in patients with moderate-to-severe Alzheimer's disease (AD). In this study, we aimed to further demonstrate the effectiveness of GV1001 using subscales of the Severe Impairment Battery (SIB), which is a validated measure to assess cognitive function in patients with moderate-to-severe AD. Methods: We performed a post hoc analysis of data from a 6 month, multicenter, phase 2, randomized, double-blind, placebo-controlled trial with GV1001 (ClinicalTrials.gov, NCT03184467). Patients were randomized to receive either GV1001 or a placebo for 24 weeks. In the current study, nine subscales of SIB-social interaction, memory, orientation, language, attention, praxis, visuospatial ability, construction, and orientation to name- were compared between the treatment (GV1001 1.12 mg) and placebo groups at weeks 12 and 24. The safety endpoints for these patients were also determined based on adverse events. Results: In addition to the considerable beneficial effect of GV1001 on the SIB total score, GV1001 1.12 mg showed the most significant effect on language function at 24 weeks compared to placebo in both the full analysis set (FAS) and per-protocol set (PPS) (p=0.017 and p=0.011, respectively). The rate of adverse events did not differ significantly between the 2 groups. Conclusions: Patients with moderate-to-severe AD receiving GV1001 had greater language benefits than those receiving placebo, as measured using the SIB language subscale.

Conus medullaris syndrome as a complication of radioisotope cisternography.

OBJECTIVE: Conus medullaris syndrome (CMS) is a clinical neurologic syndrome caused by a conus medullaris lesion. CMS is a heterogeneous entity with various etiologies such as trauma or a space-occupying lesion. Multiple cases of CMS following spinal anesthesia have been reported, but CMS after radioisotope (RI) cisternography has not yet been reported. METHODS: We present four patients who developed CMS after RI cisternography. RESULTS: All experienced neurological deficits such as paraparesis, sensory loss, and urinary incontinence three to four days after RI cisternography. Two showed abnormalities on lumbar magnetic resonance imaging, and three had complete symptom resolution within ten weeks. CONCLUSIONS: The pathomechanism of the CMS is unclear, but we hypothesize that RI neurotoxicity might be responsible. It is possible that the use of low-dose 99mTc-DTPA or an alternative diagnostic tool such as magnetic resonance cisternography could help to prevent this complication.

Association between serum stromal cell-derived factor-1α and long-term outcome of acute ischemic stroke.

BACKGROUND: Stromal cell-derived factor-1α (SDF-1α) provides neurotrophic support to neurons. In this prospective study, we investigated the association between SDF-1α and long-term outcome in patients with acute ischemic stroke. METHODS: This study included 104 patients with first-ever ischemic stroke, identified within 24 h of symptom onset. Serum samples were collected immediately after admission and 3 months thereafter, as well as from age- and sex-matched normal controls. The correlation between acute-stage serum SDF-1α and stroke severity were analyzed. Finally, the relationship between serum SDF-1α and long-term outcome was evaluated by multivariate analysis. RESULTS: Serum SDF-1α levels were only higher in acute-stage stroke patients compared with the normal control group (p = 0.011). The serum SDF-1α level was increased in proportion to infarct volume (r = 0.220, p = 0.025) and initial National Institutes of Health Stroke Scale score (r = 0.275, p = 0.005). After adjustment for covariates, a high initial serum SDF-1α level (OR 1.167, p = 0.023) or second and third tertiles of SDF-1α level compared to first tertile (OR 4.644, p = 0.044 and OR 9.396, p = 0.025, respectively) were significantly associated with a favorable long-term outcome. CONCLUSIONS: This prospective study demonstrated a correlation between serum SDF-1α and favorable long-term outcome in patients with acute ischemic stroke.

Anti-N-methyl-D-aspartate receptor encephalitis after coronavirus disease 2019: A case report and literature review.

RATIONALE: Coronavirus disease 2019 (COVID-19) has become a global pandemic and COVID-19-associated anti-N-methyl-D-aspartate receptor (NMDAR) encephalitis may occur through an immune-mediated pathomechanism. PATIENT CONCERNS: A 21-year-old woman with a history of COVID-19 presented to our hospital with memory decline and psychiatric symptoms. DIAGNOSIS: The patient was diagnosed with anti-NMDAR encephalitis. INTERVENTION: Intravenous methylprednisolone (1 g/day over 5 days) followed by immunoglobulin (0.4 g/kg/day over 5 days) were administered. The patient underwent laparoscopic salpingo-oophorectomy to remove an ovarian teratoma. OUTCOMES: The patient was discharged with sequelae of short-term memory impairment, without other neuropsychiatric symptoms. LESSONS: Cases of previously reported anti-NMDAR encephalitis with COVID-19 were reviewed and compared with the present case. Clinicians should be aware of the occurrence of anti-NMDAR encephalitis in patients who present with neuropsychiatric complaints during or after exposure to COVID-19. Further studies are required to determine the causal relationship between the 2 diseases and predict the prognosis of anti-NMDAR encephalitis after COVID-19 exposure.

Amlodipine besylate and amlodipine camsylate prevent cortical neuronal cell death induced by oxidative stress.

We examined the neuroprotective effects of the long-acting third-generation dihydropyridine Ca(2+) antagonists, amlodipine besylate (AB) and amlodipine camsylate (AC), on neuronal cell death induced by oxidative stress. Cell viability and levels of free radicals and intracellular signaling proteins were measured after treating primary cultures of cortical neurons with AB, AC, and/or hydrogen peroxide (H(2) O(2) ) under various conditions. Cell viability was not affected by concentrations of AB or AC up to 5 µM but decreased at higher concentrations. Following H(2) O(2) exposure, the viability of cortical neurons decreased in a concentration-dependent manner; however, treatment with AB or AC up to 5 µM restored the viability of H(2) O(2) -injured cortical neurons. Treatment with H(2) O(2) increased the level of free radicals in cortical neurons, and pre-treatment with AB or AC counteracted this in a dose-dependent manner. Similarly, treatment with AB or AC reduced the declines in p85aPI3K, phosphorylated Akt, phosphorylated GSK-3ß, heat-shock transcription factor-1, and Bcl-2 induced by H(2) O(2) , as well as the increases in cyclooxygenase-2, cytosolic cytochrome c, cleaved caspase 9, and cleaved caspase 3. Our results indicate that AB and AC exert similar neuroprotective effects by reducing oxidative stress, enhancing survival signals, and inhibiting death signals.

Clinical and genetic analyses of three Korean families with hereditary hemorrhagic telangiectasia.

BACKGROUND: Hereditary hemorrhagic telangiectasia (HHT) is an autosomal-dominant vascular disorder, characterized by recurrent epistaxis, mucocutaneous telangiectases, and arteriovenous malformations (AVMs) in various visceral organs. Endoglin (ENG) and activin receptor-like kinase 1 (ACVRL1; ALK1), receptors for transforming growth factor-ß (TGF-ß) superfamily, have been identified as the principal HHT-causing genes. METHODS: Three unrelated Korean HHT patients and their asymptomatic as well as symptomatic family members were genetically diagnosed by sequencing whole exons and their flanking regions of ENG and ACVRL1. Functionality of an aberrant translation start codon, which is created by a substitution mutation at the 5'-untranslated region (UTR) of ENG found in a HHT family, was tested by transient in vitro transfection assay. Decay of the mutant transcripts was also assessed by allele-specific expression analysis. RESULTS: Two ENG and one ACVRL1 mutations were identified: a known ENG mutation (c.360+1G > A; p.Gly74_Tyr120del); a novel ENG mutation (c.1-127C > T); and a novel ACVRL1 mutation (c.252_253insC; p.Val85fsX168). We further validated that the 5'-UTR ENG mutation prevents translation of ENG from the biological translation initiation site of the mutant allele, and leads to degradation of the mutant transcripts. CONCLUSIONS: This is the first experimental demonstration that a 5'-UTR mutation can prevent translation of ENG among HHT patients, and further supports the previous notion that haploinsufficiency is the primary mechanism of HHT1. Our data also underscore the importance of including exons encoding 5' UTR for HHT mutation screening.

Telmisartan Inhibits the NLRP3 Inflammasome by Activating the PI3K Pathway in Neural Stem Cells Injured by Oxygen-Glucose Deprivation.

Angiotensin II receptor blockers (ARBs) have been shown to exert neuroprotective effects by suppressing inflammatory and apoptotic responses. In the present study, the effects of the ARB telmisartan on the NLRP3 inflammasome induced by oxygen-glucose deprivation (OGD) in neural stem cells (NSCs) were investigated, as well as their possible association with the activation of the PI3K pathway. Cultured NSCs were treated with different concentrations of telmisartan and subjected to various durations of OGD. Cell counting, lactate dehydrogenase, bromodeoxyuridine, and colony-forming unit assays were performed to measure cell viability and proliferation. In addition, the activity of intracellular signaling pathways associated with the PI3K pathway and NLRP3 inflammasome was evaluated. Telmisartan alone did not affect NSCs up to a concentration of 10 µM under normal conditions but showed toxicity at a concentration of 100 µM. Moreover, OGD reduced the viability of NSCs in a time-dependent manner. Nevertheless, treatment with telmisartan increased the viability and proliferation of OGD-injured NSCs. Furthermore, telmisartan promoted the expression of survival-related proteins and mRNA while inhibiting the expression of death-related proteins induced by OGD. In particular, telmisartan attenuated OGD-dependent expression of the NLRP3 inflammasome and its related signaling proteins. These beneficial effects of telmisartan were blocked by a PI3K inhibitor. Together, these results indicate that telmisartan attenuated the activation of the NLRP3 inflammasome by triggering the PI3K pathway, thereby contributing to neuroprotection.

Neuroprotective Effects of GV1001 in Animal Stroke Model and Neural Cells Subject to Oxygen-Glucose Deprivation/Reperfusion Injury.

BACKGROUND AND PURPOSE: Previous studies have revealed the diverse neuroprotective effects of GV1001. In this study, we investigated the effects of GV1001 on focal cerebral ischemia-reperfusion injury (IRI) in rats and oxygen-glucose deprivation/reoxygenation (OGD/R)-induced injury in neural stem cells (NSCs) and cortical neurons. METHODS: Focal cerebral IRI was induced by transient middle cerebral artery occlusion (MCAO). Brain diffusion-weighted imaging (DWI) was performed 2 hours after occlusion, and a total of 37 rats were treated by reperfusion with GV1001 or saline 2 hours after occlusion. Fluid-attenuated inversion recovery (FLAIR) magnetic resonance imaging, immunohistochemistry, and neurobehavioral function analyses were performed. Additionally, OGD/R-injured NSCs and cortical neurons were treated with different GV1001 concentrations. Cell viability, proliferation, migration, and oxidative stress were determined by diverse molecular analyses. RESULTS: In the stroke model, GV1001 protected neural cells against IRI. The most effective dose of GV1001 was 60 µM/kg. The infarct volume on FLAIR 48 hours after MCAO compared to lesion volume on DWI showed a significantly smaller ratio in the GV1001-treated group. GV1001-treated rats exhibited better behavioral functions than the saline-treated rats. Treatment with GV1001 increased the viability, proliferation, and migration of the OGD/R-injured NSCs. Free radicals were significantly restored by treatment with GV1001. These neuroprotective effects of GV1001 have also been demonstrated in OGD/R-injured cortical neurons. CONCLUSIONS: The results suggest that GV1001 has neuroprotective effects against IRI in NSCs, cortical neurons, and the rat brain. These effects are mediated through the induction of cellular proliferation, mitochondrial stabilization, and anti-apoptotic, anti-aging, and antioxidant effects.

Efficacy and safety of GV1001 in patients with moderate-to-severe Alzheimer's disease already receiving donepezil: a phase 2 randomized, double-blind, placebo-controlled, multicenter clinical trial.

BACKGROUND: Our previous studies showed that GV1001 has various protective effects against ß-amyloid and other stressors. Based on these findings, we hypothesized that GV1001 might have beneficial effects in patients with Alzheimer's disease (AD). METHODS: A phase 2, double-blind, parallel-group, placebo-controlled, 6-month randomized clinical trial was performed to evaluate the safety and efficacy of subcutaneously administered GV1001. Between September 2017 and September 2019, 13 centers in South Korea recruited participants. A total of 106 patients were screened, and 96 patients with moderate-to-severe AD were randomized 1:1:1 to the placebo (group 1, n = 31), GV1001 0.56 mg (group 2, n = 33), and 1.12 mg (group 3, n = 32) groups. GV1001 was administered every week for 4 weeks (4 times), followed by every 2 weeks until week 24 (10 times). The primary endpoint was the change in the Severe Impairment Battery (SIB) score from baseline to week 24. The key secondary efficacy endpoints were the change in the Clinical Dementia Rating Sum of Box (CDR-SOB), Alzheimer's Disease Cooperative Study-Activities of Daily Living (ADCS-ADL), Neuropsychiatric Inventory (NPI), Mini-Mental State Examination, and Global Deterioration Scale scores. The safety endpoints were also assessed based on adverse events, laboratory test results, vital signs, and other observations related to safety. RESULTS: Group 3 showed less decrease in the SIB score at 12 and 24 weeks compared with group 1 (P < 0.05). These were not significantly observed in group 2. Among the secondary endpoints, only the NPI score showed significantly better improvement in group 2 than in group 3 at week 12; however, there were no other significant differences between the groups. Although the ADCS-ADL and CDR-SOB scores showed a pattern similar to SIB scores, a statistically significant result was not found. Adverse events were similar across all three groups. CONCLUSIONS: The results indicate that GV1001 1.12 mg met the primary endpoint of a statistically significant difference. GV1001 was well tolerated without safety concerns. This study warrants a larger clinical trial. TRIAL REGISTRATION: ClinicalTrials.gov NCT03184467 . Registered on June 12, 2017.

Use of methotrexate in the management of recurrent Tolosa-Hunt syndrome: Two case reports.

RATIONALE: Tolosa-Hunt syndrome (THS) is rare condition characterized by painful ophthalmoplegia that usually responds well to corticosteroid. About a half of THS patients experience recurrence within intervals of months to years from initial presentation. Recurrence is more common in younger patients, and can be ipsilateral, contralateral, or bilateral. Cyclosporine, azathioprine, methotrexate, mycophenolate mofetil, infliximab, and radiotherapy can be considered as second-line treatment. However, there is insufficient evidence for treatments preventing recurrence of THS. PATIENT CONCERNS: We experienced two patients with THS that recurred twice while tapering or after ceasing corticosteroid administration. DIAGNOSIS: Both patients were diagnosed as recurrent THS. INTERVENTIONS: Methotrexate was treated with a combination of corticosteroid after THS recurred twice with corticosteroid therapy alone. OUTCOMES: After adding methotrexate to the steroid regimen, their symptoms were successfully regulated and ceased to recur LESSONS:: These cases add to the evidence for the use of methotrexate as a second-line therapeutic agent for those patients with recurrent THS attacks. Further studies are in need to prove the risk and benefits of second-line treatments in THS.

Early increment of soluble triggering receptor expressed on myeloid cells 2 in plasma might be a predictor of poor outcome after ischemic stroke.

Soluble triggering receptor expressed on myeloid cells 2 (sTREM2) is derived from cleavage of TREM2, which is expressed on the cell surface of microlgia and other tissue-specific macrophages. In the present study, the changes in the sTREM2 levels after ischemic stroke (IS) and their association with clinical outcomes were evaluated. A total of 43 patients diagnosed with non-cardioembolic IS between June 2011 and May 2014 were consecutively included in this study. Patients treated with intravenous thrombolysis or intra-arterial thrombectomy were excluded. Plasma samples were collected three times (days 1, 7, and 90) after ictus. The sTREM2 level was measured in the samples using the highly sensitive solid-phase proximity ligation assay (SP-PLA). Among the 43 subjects, higher initial NIH stroke scale (NIHSS) score (P = 0.005), early increment of sTREM2 (P < 0.001), and late decrement of sTREM2 (P = 0.002), were more common in patients with poor outcome. Based on multivariate analysis, initial NIHSS score (P = 0.015) and early increment of sTREM2 (P = 0.032) were independently associated with poor outcome. The results from the present study indicate that increment of sTREM2 level at the early phase was a predictor of poor outcome. Serial follow-up of sTREM2 may aid prognosis after stroke.

Cilnidipine mediates a neuroprotective effect by scavenging free radicals and activating the phosphatidylinositol 3-kinase pathway.

We investigated the neuroprotective effect and mechanisms of action of cilnidipine, a long-acting, second-generation 1,4-dihydropyridine inhibitor of L- and N-type calcium channels, in PC12 cells that were neuronally differentiated by treatment with nerve growth factor (nPC12 cells). To evaluate the effect of cilnidipine on viability, nPC12 cells were treated with several concentrations of this drug before performing viability assays. Free radical levels and intracellular signaling proteins were measured with the fluorescent probe, 2',7'-dichlorodihydrofluorescein diacetate and western blotting, respectively. Cell viability was not affected by low concentrations of cilnidipine up to 150 microM, but it was slightly decreased at 200 microM cilnidipine. Following H(2)O(2) exposure, the viability of nPC12 cells decreased significantly; however, treatment with cilnidipine increased the viability of H(2)O(2)-injured nPC12 cells in a concentration-dependent manner. Treatment with H(2)O(2) resulted in a concentration-dependent increase in free radical levels in nPC12 cells, and cilnidipine treatment reduced free radical levels in H(2)O(2)-injured nPC12 cells in a dose-dependent manner. Cilnidipine treatment increased the expression of p85aPI3K (phosphatidylinositol 3-kinase) phosphorylated Akt, phosphorylated glycogen synthase kinase-3 (pGSK-3beta), and heat shock transcription factor (HSTF-1) which are proteins related to neuronal cell survival, and decreased levels of cytosolic cytochrome c, activated caspase 3, and cleaved poly (ADP-ribose) polymerase (PARP), which are associated with neuronal cell death, in H(2)O(2)-injured nPC12 cells. These results indicate that cilnidipine mediates its neuroprotective effects by reducing oxidative stress, enhancing survival signals (e.g., PI3K, phosphorylated Akt, pGSK-3beta, and HSTF-1), and inhibiting death signals from cytochrome c release, caspase 3 activation, and PARP cleavage.

Transient upbeat nystagmus due to unilateral focal pontine infarction.

Upbeat nystagmus is known to be caused by an imbalance of the vertical vestibulo-ocular reflex (VOR), favoring downward VOR activity, due to bilateral lesions of the medulla, ventral tegmentum, anterior cerebellar vermis, adjacent brachium conjunctivum and the midbrain. We report on two patients who had transient upbeat nystagmus due to unilateral pontine infarction that may have disrupted bilateral upward VOR pathways running in the ventral tegmental tracts.

Sublethal Doses of Zinc Protect Rat Neural Stem Cells Against Hypoxia Through Activation of the PI3K Pathway.

Cerebral infarction is one of the major causes of severe morbidity and mortality, and thus, research has focused on developing treatment options for this condition. Zinc (Zn) is an essential element in the central nervous system and has several neuroprotective effects in the brain. In this study, we examined the neuroprotective effects of Zn on neural stem cells (NSCs) exposed to hypoxia. After treatment with several concentrations of Zn, the viability of NSCs under hypoxic conditions was measured by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, Trypan blue staining, and a lactate dehydrogenase assay. To evaluate the effect of Zn on the proliferation of NSCs, bromodeoxyuridine/5-bromo-2'-deoxyuridine (BrdU) labeling and colony formation assays were performed. Apoptosis was also examined in NSCs exposed to hypoxia with and without Zn treatment. In addition, a western blot analysis was performed to evaluate the effect of Zn on intracellular signaling proteins. NSC viability and proliferation were decreased under hypoxic conditions, but treatment with sublethal doses of Zn restored viability and proliferation. Sublethal doses of Zn reduced apoptosis caused by hypoxia, increased the expression levels of proteins related to the phosphatidylinositol-3 kinase (PI3K) pathway, and decreased the expression levels of proteins associated with neuronal cell death. These findings confirm that in vivo, sublethal doses of Zn protect NSCs against hypoxia through the activation of the PI3K pathway. Thus, Zn could be employed as a therapeutic option to protect NSCs in ischemic stroke.

Mitochondria damaged by Oxygen Glucose Deprivation can be Restored through Activation of the PI3K/Akt Pathway and Inhibition of Calcium Influx by Amlodipine Camsylate.

Amlodipine, a L-type calcium channel blocker, has been reported to have a neuroprotective effect in brain ischemia. Mitochondrial calcium overload leads to apoptosis of cells in neurologic diseases. We evaluated the neuroprotective effects of amlodipine camsylate (AC) on neural stem cells (NSCs) injured by oxygen glucose deprivation (OGD) with a focus on mitochondrial structure and function. NSCs were isolated from rodent embryonic brains. Effects of AC on cell viability, proliferation, level of free radicals, and expression of intracellular signaling proteins were assessed in OGD-injured NSCs. We also investigated the effect of AC on mitochondrial structure in NSCs under OGD by transmission electron microscopy. AC increased the viability and proliferation of NSCs. This beneficial effect of AC was achieved by strong protection of mitochondria. AC markedly enhanced the expression of mitochondrial biogenesis-related proteins and mitochondrial anti-apoptosis proteins. Together, our results indicate that AC protects OGD-injured NSCs by protecting mitochondrial structure and function. The results of the present study provide insight into the mechanisms underlying the protective effects of AC on NSCs.