Andrew Barszczyk

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.

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.

Current understanding of K ATP channels in neonatal diseases: focus on insulin secretion disorders

ATP-sensitive potassium (KATP) channels are cell metabolic sensors that couple cell metabolic status to electric activity, thus regulating many cellular functions. In pancreatic beta cells, KATP channels modulate insulin secretion in response to fluctuations in plasma glucose level, and play an important role in glucose homeostasis. Recent studies show that gain-of-function and loss-of-function mutations in KATP channel subunits cause neonatal diabetes mellitus and congenital hyperinsulinism respectively. These findings lead to significant changes in the diagnosis and treatment for neonatal insulin secretion disorders. This review describes the physiological and pathophysiological functions of KATP channels in glucose homeostasis, their specific roles in neonatal diabetes mellitus and congenital hyperinsulinism, as well as future perspectives of KATP channels in neonatal diseases.

Meta-Analysis of Serum Insulin-Like Growth Factor 1 in Alzheimer’s Disease

Insulin-like growth factor 1 (IGF-1) serum levels have been reported to be altered in Alzheimer’s disease patients, and it was suggested that the changes in IGF-1 serum level may play a role in disease pathology and progression. However, this notion remained controversial due to conflicting findings. We conducted a meta-analysis to determine the relationship between IGF-1 serum levels and Alzheimer’s disease. We searched the databases PUBMED, Ovid SP, and Cochrane library for relevant studies. The primary data analyzed was serum IGF-1 from Alzheimer’s disease subjects and controls. Pooled weighted mean difference using a random effects model was used to determine the relationship between serum levels and disease state. Nine studies were included in the meta-analysis compromising a total of 1639 subjects. The pooled weighted mean difference was -2.27ng/ml (95% CI: [-22.221, 17.66]) with a P value of 0.82. Thus our finding did not show clear relationship between low IGF-1 and Alzheimer’s disease subjects. We did not find evidence of publication bias by analyzing a funnel plot as well as Egger’s and Begg’s tests. While eight out of the nine studies included in this meta-analysis detected a statistically significant increase or decrease in serum levels of IGF-1 in Alzheimer’s disease subjects, the analysis as a whole did not show a significant trend in either direction. Thus, IGF-1 level is likely a critical personalized factor. A large database of clinical trials is required for better understanding the relationship between IGF-1 levels and Alzheimer’s disease.

Differential Roles of the Mevalonate Pathway in the Development and Survival of Mouse Purkinje Cells in Culture

The cerebellum is an important locus for motor learning and higher cognitive functions, and Purkinje cells constitute a key component of its circuit. Biochemically, significant turnover of cholesterol occurs in Purkinje cells, causing the activation of the mevalonate pathway. The mevalonate pathway has important roles in cell survival and development. In this study, we investigated the outcomes of mevalonate inhibition in immature and mature mouse cerebellar Purkinje cells in culture. Specifically, we found that the inhibition of the mevalonate pathway by mevastatin resulted in cell death, and geranylgeranylpyrophosphate (GGPP) supplementation significantly enhanced neuronal survival. The surviving immature Purkinje cells, however, exhibited dendritic developmental deficits. The morphology of mature cells was not affected. The inhibition of squalene synthase by zaragozic acid caused impaired dendritic development, similar to that seen in the GGPP-rescued Purkinje cells. Our results indicate GGPP is required for cell survival and squalene synthase for the cell development of Purkinje cells. Abnormalities in Purkinje cells are linked to motor-behavioral learning disorders such as cerebellar ataxia. Thus, serious caution should be taken when using drugs that inhibit geranylgeranylation or the squalene-cholesterol branch of the pathway in the developing stage.

Stroke risk in treatment of type 2 diabetes in China: a 7 year retrospective cohort study

Abstract Background Diabetes and stroke are leading causes of death and disability, and major public health concerns in China, which accounts for 25% of patients with diabetes worldwide. Type 2 diabetes is an independent risk factor for cerebral ischaemia and accounts for about 20% of stroke cases. We previously showed that K ATP channel activation provides neuroprotection against cerebral ischaemia and that sulfonylureas blocking these channels increase the stroke risk in patients with type 2 diabetes. We aimed to further determine whether different classes of antidiabetic drugs increase the risk of stroke in Chinese patients with type 2 diabetes. Methods In this retrospective cohort study, we used data from medical records (Jan 1, 2008, to Dec 30, 2015) of the Endocrinology Division of the Third Affiliated Hospital of Jilin University, Jilin, China. Patients aged 32–80 years with type 2 diabetes whose blood glucose control treatment included sulfonylureas, metformin, insulin, or restricted diet were eligible for inclusion. All patients also received education on diet, exercise, blood glucose monitoring, and other aspects of diabetes management. We used χ 2 test was to determine significance (p Results 1829 patients were eligible for inclusion, of whom 528 had sulfonylureas, 611 had metformin, 535 had insulin, and 155 were on a restricted diet. 55 individuals who switched to alternative therapy or were lost to follow-up were excluded, and 1774 patients were included in analysis. 116 cases of stroke were recorded: 42 in the sulfonylurea group, 29 in the metformin group, 36 in the insulin group, and nine in the restricted diet group. The risk of stroke in patients given sulfonylureas was increased by 2·65 times (95% CI 1·56–4·50) compared with those given metformin, 1·21 times (0·79–1·86) compared with those given insulin, 1·13 times (0·56–2.25) compared with those on a restricted diet, and 1·63 times (1·12–2·37) compared with all three non-sulfonylurea groups. Interpretation Chinese patients with diabetes receiving sulfonylureas seem to have a higher risk of stroke than those using non-sulfonylurea antidiabetic treatments. Cardiovascular safety of antidiabetic drugs needs to be carefully assessed to improve health care and quality of life in Chinese patients with diabetes. Funding Heart and Stroke Foundation of Canada (G-13-0003069) and National Sciences and Engineering Research Council of Canada (NSERC-249962-09).