EGb761 improves the cognitive function of elderly db/db-/- diabetic mice by regulating the beclin-1 and NF-κB signaling pathways.

To assess whether EGb761 could protect elderly diabetic mice with cognitive disorders and explore the role of beclin-1-mediated autophagy in these protective effects. Two-month-old male db/db-/- mice and wild-type C57/BL6 mice were randomly divided into six groups: db/db-/- control, db/db-/- 50 mg, db/db-/- 100 mg, wild-type (WT) control, WT 50 mg, and WT 100 mg. EGb761 (50 mg/kg or 100 mg/kg of bodyweight) was given by gavage once a day for 1 month from the age of 6 months. Y-maze and social choice tests were performed at 8th months. The blood pressure was measured. The imaging changes in the brain were measured using magnetic resonance imaging (MRI). The expression and distribution of beclin-1, LC3, and NF-κB were detected using immunohistochemistry staining and western blotting. Ultrastructure alterations in the hippocampus were observed using transmission electron microscopy. Compared with WT mice, the learning ability, memory and overall cognitive function of db/db-/- mice decreased (P < 0.05), and EGb761 could significantly improve the learning and memory function of db/db-/- mice (P < 0.05). EGb761 significantly improved systolic blood pressure in db/db-/- mice (P < 0.01). In addition, fMRI-bold showed a decline in the hippocampus of mice in the db/db-/- group compared with WT. EGb761 could improve these above changes. Immunohistochemistry staining and western blotting confirmed that EGb761 significantly increased beclin-1 and reduced LC3-II/I levels in the brains of db/db-/- mice (P < 0.05). NF-κB levels were obviously higher in the db/db-/- group than that in the WT group, and EGb761 significantly reduced NF-κB levels in db/db-/- mice (P < 0.05). There was a trend of increased autophagosomes in db/db-/- mice, but EGb761 did not change obviously the number of autophagosomes. Compared with normal aged WT mice, aging db/db-/- mice had more common complications of cerebral small vessel disease and cognitive dysfunction. EGb761 could significantly improve the cognitive function of aging db/db-/- mice via a mechanism that may involve the regulation of beclin-1, LC3, and NF-κB.

G-CSF and cognitive dysfunction in elderly diabetic mice with cerebral small vessel disease: Preventive intervention effects and underlying mechanisms.

AIMS: Although cognitive dysfunction is a common neurological complication in elderly patients with diabetes, the mechanisms underlying this relationship remain unclear, and effective preventive interventions have yet to be developed. Thus, this study investigated the preventive effects and mechanisms of action associated with granulocyte colony-stimulating factor (G-CSF) on cognitive dysfunction in elderly diabetic mice with cerebral small vessel disease. METHODS: This study included 40 male db/db diabetic and wild-type (WT) mice that were categorized into the following four groups at the age of 3 weeks: db/db group (DG), db/db+G-CSF group (DGG), WT group (WG), and WT+G-CSF group (WGG). The mice were fed normal diets for 4 months and then given G-CSF (75 µg/kg) via intraperitoneal injections for 1 month. At 7.5 months of age, the cognitive abilities of the mice were assessed with the Y-maze test and the Social Choice Test; body weight, blood pressure (BP), and blood glucose measurements were obtained throughout the study. Brain imaging and blood oxygen level-dependent (BOLD) contrast imaging analyses were performed with a small animal magnetic resonance imaging (MRI) system, autophagosome levels were detected with a transmission electron microscope (TEM), hippocampal neurons were assessed with hematoxylin and eosin (HE) staining, and protein expressions and distributions were evaluated using immunohistochemistry and Western blot analyses. RESULTS: (i) The body weight and blood glucose levels of the DG and DGG mice were significantly higher than those of the WG and WGG mice; (ii) social choice and spatial memory capabilities were significantly reduced in DG mice but were recovered by G-CSF in DGG mice; (iii) the MRI scans revealed multiple lacunar lesions and apparent hippocampal atrophy in the brains of DG mice, but G-CSF reduced the number of lacunar lesions and ameliorated hippocampal atrophy; (iv) the MRI-BOLD scans showed a downward trend in whole-brain activity and reductions in the connectivities of the hippocampus and amygdala with subcortical structures in DG mice, but G-CSF clearly improved the altered brain activity as well as the connectivity of the hippocampus in DGG mice; (v) HE staining revealed fewer neurons in the hippocampus in DG mice; (vi) TEM analyses revealed significantly fewer autophagosomes in the hippocampi of DG mice, but G-CSF did not increase these numbers; (vii) there were significant reductions in mechanistic target of rapamycin (mTOR) and LC3-phosphatidylethanolamine conjugate (LC3)-II/I levels in the hippocampi of DG mice, whereas p62 was upregulated, and G-CSF significantly enhanced the levels of Beclin1, mTOR, and LC-II/I in DGG mice; and (viii) G-CSF significantly reversed increases in nuclear factor κB (NF-κB) protein levels in DG but not in WG mice. CONCLUSIONS: In this study, aged diabetic mice were prone to cognitive dysfunction and cerebral small vessel disease. However, administration of G-CSF significantly improved cognitive function in elderly db/db diabetic mice, and this change was likely related to the regulation of autophagy and NF-κB signaling pathways.

Beclin-1- mediated autophagy may be involved in the elderly cognitive and affective disorders in streptozotocin-induced diabetic mice.

BACKGROUND: Diabetes is the most common metabolic disease with many chronic complications, and cognitive disorders are one of the common complications in patients with diabetes. Previous studies have showed that autophagy played important roles in the progression of metabolic syndrome, diabetes and other diseases. So we investigated whether aged diabetic mice are prone to be associated with the cognitive and affective disorders and whether Beclin-1-mediated autophagy might be involved in thepahological process. METHODS: High-fat diet/streptozotocin (STZ) injection-induced diabetic C57 mice were adopted in this study. Cognitive disorders were detected by Morris water maze and fear conditional test. Affective disorders were detected by tail suspension test and forced swimming test. Magnetic resonance imaging was applied to observe changes of morphology and metabolism in the brain. The 18 F-fluorodeoxyglucose positron emission tomography (FDG-PET) was used to assess metabolism changes in the brain of aged diabetic mice. Autophagy were evaluated by Beclin- 1, LC3II/I and P62, which were detected by western blot analysis and observed by electron microscopy. RESULTS: 1. Compared with control group, diabetes mice showed significantly decreasing abilities in spatial memory and conditioned fear memory (all P < 0.05), and increasing tendency of depression (P < 0.05). 2. MRI showed that the majority of elderly diabetic mice were associated with multiple cerebral small vessel disease. Some even showed hippocampal atrophy, ventricular dilatation and leukoaraiosis. 3. FDG-PET-CT discovered that the glucose metabolism in the amygdala and hippocampus was significantly decreased compared with normal aged mice (P < 0.05). 4. Electron microscopy found that, although autophagy bodies was not widespread, and there was no significant difference between the two groups, yet compared with normal aged mice, apparent cell edema, myelinated tow reduction and intracellular lipofuscin augmentation existed in elderly diabetic mice brain. 5. The level of p62 was increased in the STZ-induced diabetic mice hippocampus and striatum, and beclin1 protein expression were significantly decreased in diabetic mice hippocampus compared with normal aged mice (P < 0.05). There was a upward trend of the ratio of LC3II/I in hippocampus, cortex and striatum, but no statistically difference between the two groups. CONCLUSION: Compared with normal aged mice, diabetic aged mice were apt to cerebral small vessel disease and associated with cognitive and affective disorders, which may be related to the significantly reduced glucose metabolism in hippocampus and amygdala. Beclin1 mediated autophagy in hippocampus probably played an important role in cognitive and affective disorders of STZ-induced aged diabetic mice.

NF-κB Signaling is Involved in the Effects of Intranasally Engrafted Human Neural Stem Cells on Neurofunctional Improvements in Neonatal Rat Hypoxic-Ischemic Encephalopathy.

AIM: Hypoxic-ischemic encephalopathy (HIE) is a common neurological disease in infants with persistent neurobehavioral impairments. Studies found that neural stem cell (NSC) therapy benefits HIE rats; however, the mechanisms underlying are still unclear. The current study investigated the efficacy and molecular events of human embryonic neural stem cells (hNSCs) in neonatal hypoxic-ischemic (HI) rats. METHODS: PKH-26-labeled hNSCs were intranasally delivered to P7 Sprague Dawley rats 24 h after HI. Neurobehavioral tests were performed at the indicated time after delivery: righting reflex and gait testing at D1, 3, 5, and 7; grid walking at D7 and 14; social choice test (SCT) at D28; and Morris water maze from D35 to 40. Protein expression was determined by Western blot analysis. Brain damage was assessed by cresyl violet staining and MBP staining. hNSC distribution and differentiation were observed by in vivo bioluminescence imaging and immunofluorescence staining. RESULTS: (1) hNSCs migrated extensively into brain areas within 24 h after the delivery, survived even at D42 with the majority in ipsi-hemisphere, and could be co-labeled with NeuN or GFAP. (2) hNSCs reduced the upregulation in cytosolic IL-1ß, p-IκBα, and NF-κB p65 levels, whereas enhanced nuclear p65 expression in HI rats at D3 after the delivery. (3) hNSCs decreased HI-induced brain tissue loss and white matter injury at D42 after the delivery. (4) hNSCs improved neurological outcomes in HI rats in the tests of righting reflex (within 3 days), gait (D5), grid (D7), SCT (D28), and water maze (D42). CONCLUSION: Intranasal delivery of hNSCs could prevent HI-induced brain injury and improve neurobehavioral outcomes in neonatal HI rats, which is possibly related to the modulation of NF-κB signaling.