Christopher L.F. Sun

Neuronal KATP channels mediate hypoxic preconditioning and reduce subsequent neonatal hypoxic–ischemic brain injury

Neonatal hypoxic-ischemic brain injury and its related illness hypoxic-ischemic encephalopathy (HIE) are major causes of nervous system damage and neurological morbidity in children. Hypoxic preconditioning (HPC) is known to be neuroprotective in cerebral ischemic brain injury. K(ATP) channels are involved in ischemic preconditioning in the heart; however the involvement of neuronal K(ATP) channels in HPC in the brain has not been fully investigated. In this study, we investigated the role of HPC in hypoxia-ischemia (HI)-induced brain injury in postnatal seven-day-old (P7) CD1 mouse pups. Specifically, TTC (2,3,5-triphenyltetrazolium chloride) staining was used to assess the infarct volume, TUNEL (Terminal deoxynucleotidyl transferase mediated dUTP nick end-labeling) to detect apoptotic cells, Western blots to evaluate protein level, and patch-clamp recordings to measure K(ATP) channel current activities. Behavioral tests were performed to assess the functional recovery after hypoxic-ischemic insults. We found that hypoxic preconditioning reduced infarct volume, decreased the number of TUNEL-positive cells, and improved neurobehavioral functional recovery in neonatal mice following hypoxic-ischemic insults. Pre-treatment with a K(ATP) channel blocker, tolbutamide, inhibited hypoxic preconditioning-induced neuroprotection and augmented neurodegeneration following hypoxic-ischemic injury. Pre-treatment with a K(ATP) channel opener, diazoxide, reduced infarct volume and mimicked hypoxic preconditioning-induced neuroprotection. Hypoxic preconditioning induced upregulation of the protein level of the Kir6.2 isoform and enhanced current activities of K(ATP) channels. Hypoxic preconditioning restored the HI-reduced PKC and pAkt levels, and reduced caspase-3 level, while tolbutamide inhibited the effects of hypoxic preconditioning. We conclude that K(ATP) channels are involved in hypoxic preconditioning-induced neuroprotection in neonatal hypoxic-ischemic brain injury. K(ATP) channel openers may therefore have therapeutic effects in neonatal hypoxic-ischemic brain injury.

Xyloketal B Suppresses Glioblastoma Cell Proliferation and Migration in Vitro through Inhibiting TRPM7-Regulated PI3K/Akt and MEK/ERK Signaling Pathways

Glioblastoma, the most common and aggressive type of brain tumors, has devastatingly proliferative and invasive characteristics. The need for finding a novel and specific drug target is urgent as the current approaches have limited therapeutic effects in treating glioblastoma. Xyloketal B is a marine compound obtained from mangrove fungus Xylaria sp. (No. 2508) from the South China Sea, and has displayed antioxidant activity and protective effects on endothelial and neuronal oxidative injuries. In this study, we used a glioblastoma U251 cell line to (1) explore the effects of xyloketal B on cell viability, proliferation, and migration; and (2) investigate the underlying molecular mechanisms and signaling pathways. MTT assay, colony formation, wound healing, western blot, and patch clamp techniques were employed. We found that xyloketal B reduced cell viability, proliferation, and migration of U251 cells. In addition, xyloketal B decreased p-Akt and p-ERK1/2 protein expressions. Furthermore, xyloketal B blocked TRPM7 currents in HEK-293 cells overexpressing TRPM7. These effects were confirmed by using a TRPM7 inhibitor, carvacrol, in a parallel experiment. Our findings indicate that TRPM7-regulated PI3K/Akt and MEK/ERK signaling is involved in anti-proliferation and migration effects of xyloketal B on U251 cells, providing in vitro evidence for the marine compound xyloketal B to be a potential drug for treating glioblastoma.

Marine Compound Xyloketal B Reduces Neonatal Hypoxic-Ischemic Brain Injury

Neonatal hypoxic-ischemic encephalopathy causes neurodegeneration and brain injury, leading to sensorimotor dysfunction. Xyloketal B is a novel marine compound isolated from a mangrove fungus Xylaria species (no. 2508) with unique antioxidant effects. In this study, we investigated the effects and mechanism of xyloketal B on oxygen-glucose deprivation-induced neuronal cell death in mouse primary cortical culture and on hypoxic-ischemic brain injury in neonatal mice in vivo. We found that xyloketal B reduced anoxia-induced neuronal cell death in vitro, as well as infarct volume in neonatal hypoxic-ischemic brain injury model in vivo. Furthermore, xyloketal B improved functional behavioral recovery of the animals following hypoxic-ischemic insult. In addition, xyloketal B significantly decreased calcium entry, reduced the number of TUNEL-positive cells, reduced the levels of cleaved caspase-3 and Bax proteins, and increased the level of Bcl-2 protein after the hypoxic-ischemic injury. Our findings indicate that xyloketal B is effective in models of hypoxia-ischemia and thus has potential as a treatment for hypoxic-ischemic brain injury.

Neuroprotective effects of volume-regulated anion channel blocker DCPIB on neonatal hypoxic-ischemic injury

Aim:To evaluate the role of swelling-induced activation of volume-regulated anion channels (VRACs) in a neonatal hypoxic-ischemic injury model using the selective VRAC blocker 4-(2-butyl-6,7-dichloro-2-cyclopentyl-indan-1-on5-yl) oxobutyric acid (DCPIB).Methods:Cerebral hypoxic-ischemic injury was induced in 7-day-old mouse pups with Rice-Vannucci method. Prior to the onset of ischemia, the animals were ip administered DCPIB (10 mg/kg). The animals were sacrificed 24 h afterwards, coronal sections of the brains were cut and the areas of infarct were examined using TTC staining and an image-analysis system. Cultured PC12 cells were subjected to oxygen-glucose deprivation (OGD) for 4 h. The cellular viability was assessed using Cell Counting Kit 8. Intracellular chloride concentration [Cl−]i was measured using 6-methoxy-N-ethylquinolinium iodide.Results:DCPIB-treated mice showed a significant reduction in hemispheric corrected infarct volume (26.65%±2.23%) compared to that in vehicle-treated mice (45.52%±1.45%, P<0.001). DCPIB-treated mice also showed better functional recovery as they were more active than vehicle-treated mice at 4 and 24 h post injury. In cultured PC12 cells, DCPIB (10 μmol/L) significantly reduced OGD-induced cell death. Moreover, DCPIB (20 μmol/L) blocked hypotonic-induced decrease in [Cl−]i in PC12 cells of both control and OGD groups.Conclusion:The results further support the pathophysiological role of VRACs in ischemic brain injury, and suggest DCPIB as a potential, easily administrable agent targeting VRACs in the context of perinatal and neonatal hypoxic-ischemic brain injury.

Cerebrovascular safety of sulfonylureas: the role of KATP channels in neuroprotection and the risk of stroke in patients with type 2 diabetes

Sulfonylureas are ATP-sensitive potassium (KATP) channel blockers commonly used in the treatment of type 2 diabetes mellitus (T2DM). Activation of KATP channels plays a neuroprotective role in ischemia; thus, whether sulfonylureas affect the outcomes of stroke in patients with T2DM needs to be further studied. In our study, streptozotocin (STZ)-induced diabetic mice subjected to transient middle cerebral artery occlusion (MCAO) showed larger areas of brain damage and poorer behavioral outcomes. Blocking the KATP channel by tolbutamide increased neuronal injury induced by oxygen-glucose deprivation (OGD) in vitro and permanent MCAO (pMCAO) in vivo. Activating the KATP channel by diazoxide reduced the effects of both the OGD and pMCAO. Western blot analysis in STZ mouse brains indicated an early increase in protein levels of N-methyl-d-aspartate receptor 2B and postsynaptic density protein-95, followed by a decrease in phosphorylation of glycogen synthase kinase 3β. Our systematic meta-analysis indicated that patients with T2DM treated with sulfonylureas had a higher odds ratio for stroke morbidity than those who received comparator drugs. Taken together, these results suggest that sulfonylurea treatment in patients with T2DM may inhibit the neuroprotective effects of KATP channels and increase the risk of stroke.

Ranking Businesses and Municipal Locations by Spatiotemporal Cardiac Arrest Risk to Guide Public Defibrillator Placement

Background: Efforts to guide automated external defibrillator placement for out-of-hospital cardiac arrest (OHCA) treatment have focused on identifying broadly defined location categories without considering hours of operation. Broad location categories may be composed of many businesses with varying accessibility. Identifying specific locations for automated external defibrillator deployment incorporating operating hours and time of OHCA occurrence may improve automated external defibrillator accessibility. We aim to identify specific businesses and municipal locations that maximize OHCA coverage on the basis of spatiotemporal assessment of OHCA risk in the immediate vicinity of franchise locations. Methods: This study was a retrospective population-based cohort study using data from the Toronto Regional RescuNET Epistry cardiac arrest database. We identified all nontraumatic public OHCAs occurring in Toronto, ON, Canada, from January 2007 through December 2015. We identified 41 unique businesses and municipal location types with ≥20 locations in Toronto from the YellowPages, Canadian Franchise Association, and the City of Toronto Open Data Portal. We obtained their geographic coordinates and hours of operation from Web sites, by phone, or in person. We determined the number of OHCAs that occurred within 100 m of each location when it was open (spatiotemporal coverage) for Toronto overall and downtown. The businesses and municipal locations were then ranked by spatiotemporal OHCA coverage. To evaluate temporal stability of the rankings, we calculated intraclass correlation of the annual coverage values. Results: There were 2654 nontraumatic public OHCAs. Tim Hortons ranked first in Toronto, covering 286 OHCAs. Starbucks ranked first in downtown, covering 110 OHCAs. Coffee shops and bank machines from the 5 largest Canadian banks occupied 8 of the top 10 spots in both Toronto and downtown. The rankings exhibited high temporal stability with intraclass correlation values of 0.88 (95% confidence interval, 0.83–0.93) in Toronto and 0.79 (95% confidence interval, 0.71–0.86) in downtown. Conclusions: We identified and ranked businesses and municipal locations by spatiotemporal OHCA risk in their immediate vicinity. This approach may help policy makers and funders to identify and prioritize potential partnerships for automated external defibrillator deployment in public-access defibrillator programs.

Spatiotemporal AED optimization is generalizable

AIMS Mathematical optimization of automated external defibrillator (AED) placements has the potential to improve out-of-hospital cardiac arrest (OHCA) coverage and reverse the negative effects of limited AED accessibility. However, the generalizability of optimization approaches has not yet been investigated. Our goal is to examine the performance and generalizability of a spatiotemporal AED placement optimization methodology, initially developed for Toronto, Canada, to the new study setting of Copenhagen, Denmark. METHODS We identified all public OHCAs (1994-2016) and all registered AEDs (2016) in Copenhagen, Denmark. We calculated the coverage loss associated with limited temporal accessibility of registered AEDs, and used a spatiotemporal optimization model to quantify the potential coverage gain of optimized AED deployment. Coverage gain of spatiotemporal deployment over a spatial-only solution was quantified through 10-fold cross-validation. Statistical testing was performed using χ2 and McNemars tests. RESULTS We found 2149 public OHCAs and 1573 registered AED locations. Coverage loss was found to be 24.4% (1104 OHCAs covered under assumed 24/7 coverage, and 835 OHCAs under actual coverage). The coverage gain from using the spatiotemporal model over a spatial-only approach was 15.3%. Temporal and geographical trends in coverage gain were similar to Toronto. CONCLUSIONS Without modification, a previously developed spatiotemporal AED optimization approach was applied to Copenhagen, resulting in similar OHCA coverage findings as Toronto, despite large geographic and cultural differences between the two cities. In addition to reinforcing the importance of temporal accessibility of AEDs, these similarities demonstrate the generalizability of optimization approaches to improve AED placement and accessibility.