Inhibition of Glial Activation and Subsequent Reduction in White Matter Damage through Supplementation with a Combined Extract of Wheat Bran, Citrus Peel, and Jujube in a Rat Model of Vascular Dementia.

Vascular dementia (VaD) is the second most common type of dementia after Alzheimer's disease. In our previous studies, we showed that wheat bran extract (WBE) reduced white matter damage in a rat VaD model and improved memory in a human clinical trial. However, starch gelatinization made the large-scale preparation of WBE difficult. To simplify the manufacturing process and increase efficacy, we attempted to find a decoction containing an optimum ratio of wheat bran, sliced citrus peel, and sliced jujube (WCJ). To find an optimal ratio, the cell survival of C6 (rat glioma) cultured under hypoxic conditions (1% O2) was measured, and apoptosis was assessed. To confirm the efficacies of the optimized WCJ for VaD, pupillary light reflex, white matter damage, and the activation of astrocytes and microglia were assessed in a rat model of bilateral common carotid artery occlusion (BCCAO) causing chronic hypoperfusion. Using a combination of both searching the literature and cell survival experiments, we chose 6:2:1 as the optimal ratio of wheat bran to sliced citrus peel to sliced jujube to prepare WCJ. We showed that phytic acid contained only in wheat bran can be used as an indicator component for the quality control of WCJ. We observed in vitro that the WCJ treatment improved cell survival by reducing apoptosis through an increase in the Bcl-2/Bax ratio. In the BCCAO experiments, the WCJ-supplemented diet prevented astrocytic and microglial activation, mitigated myelin damage in the corpus callosum and optic tract, and, consequently, improved pupillary light reflex at dosages over 100 mg/kg/day. The results suggest that the consumption of WCJ can prevent VaD by reducing white matter damage, and WCJ can be developed as a safe, herbal medicine to prevent VaD.

Deletion of B-cell translocation gene 2 (BTG2) alters the responses of glial cells in white matter to chronic cerebral hypoperfusion.

BACKGROUND: Subcortical ischemic vascular dementia, one of the major subtypes of vascular dementia, is characterized by lacunar infarcts and white matter lesions caused by chronic cerebral hypoperfusion. In this study, we used a mouse model of bilateral common carotid artery stenosis (BCAS) to investigate the role of B-cell translocation gene 2 (BTG2), an antiproliferation gene, in the white matter glial response to chronic cerebral hypoperfusion. METHODS: Btg2-/- mice and littermate wild-type control mice underwent BCAS or sham operation. Behavior phenotypes were assessed by open-field test and Morris water maze test. Brain tissues were analyzed for the degree of white matter lesions and glial changes. To further confirm the effects of Btg2 deletion on proliferation of glial cells in vitro, BrdU incorporation was investigated in mixed glial cells derived from wild-type and Btg2-/- mice. RESULTS: Relative to wild-type mice with or without BCAS, BCAS-treated Btg2-/- mice exhibited elevated spontaneous locomotor activity and poorer spatial learning ability. Although the severities of white matter lesions did not significantly differ between wild-type and Btg2-/- mice after BCAS, the immunoreactivities of GFAP, a marker of astrocytes, and Mac2, a marker of activated microglia and macrophages, in the white matter of the optic tract were higher in BCAS-treated Btg2-/- mice than in BCAS-treated wild-type mice. The expression level of Gfap was also significantly elevated in BCAS-treated Btg2-/- mice. In vitro analysis showed that BrdU incorporation in mixed glial cells in response to inflammatory stimulation associated with cerebral hypoperfusion was higher in Btg2-/- mice than in wild-type mice. CONCLUSION: BTG2 negatively regulates glial cell proliferation in response to cerebral hypoperfusion, resulting in behavioral changes.

Time-dependent effects of platelet-rich plasma on the memory and hippocampal synaptic plasticity impairment in vascular dementia induced by chronic cerebral hypoperfusion.

The present study aimed to investigate the effects of early and late administration of platelet-rich plasma (PRP) on learning-memory and hippocampal synaptic plasticity impairment in rat model of vascular dementia. Sprague-Dawley rats (6-7 weeks) were randomly divided into control, sham, 2VO + V (bilateral carotid vessel occlusion + vehicle), 2VO + E-PRP (early after 2VO), and 2VO + L-PRP (late after 2VO) groups. The injection of PRP started immediately or on the day 20 after 2VO in 2VO + E-PRP and 2VO + L-PRP, respectively, and continued until 28th day (two-time a week). The passive avoidance and Morris water maze tests were used for evaluation of fear and spatial memory formation. The in-vivo long-term potentiation (LTP) was evaluated by field-potential recording, paired-pulse ratio (PPR) and input-output (I/O) curve which were monitored as indexes for evaluation of short-term plasticity (STP) and basal-synaptic transmission (BST), respectively. The 2VO decreased PPR at inter-stimuli interval (ISI) 10 ms and BST, but injection of PRP in both treated groups rescued the PPR and BST depression. In addition, the induction of LTP, fear and spatial memory performance decreased in the 2VO + V group. However, early treatment, but not late, recovered LTP and memory. The PPR and BST improved with early and late treatment; therefore, the number and time of injection seem to be not important for recovery of BST. However, we found that LTP and memory loss rescued only with early administration. Hence, timely injection, before development of the disease, or number of injections could be critical.

Adenosine A2A Receptors in Bone Marrow-Derived Cells Attenuate Cognitive Impairment in Mice After Chronic Hypoperfusion White Matter Injury.

The mechanism of cognitive dysfunction caused by ischemic white matter lesions is unclear. To explore the effect and mechanism of different cell-derived adenosine A2A receptor (A2AR) in cognitive impairment caused by chronic hypoperfusion white matter lesions (CHWMLs), we destroyed the bone marrow hematopoietic capacity of the recipient mice using radiation irradiation followed by establishing the selectively inactivated or reconstituted A2AR models with the transplanting bone marrow from global A2AR gene knockout or wild-type mice into wild-type or gene knockout mice, respectively. Then Morris Water Maze (MWM), ELISA, immunohistochemistry, and Bielschowsky silver staining were used to assess the effect and mechanism of the cognitive function in chronic cerebral blood flow hypoperfusion (CCH) model. Selectively reconstructing bone marrow-derived cells (BMDCs) A2AR (WT → KO group) and activated total adenosine A2AR with CGS21680 (CCH + CGS group) improved the cognitive related index. Activation of BMDC A2AR suppressed expression of inflammatory cytokines in peripheral blood and reduced the number of activated microglia cells co-localized with cystatin F in local brain, consequently inhibited white matter lesions. On the contrary, selective inactivation of adenosine A2AR (KO → WT group) and activation of non-BMDC A2AR with CGS21680 (KO → WT + CGS group) served the opposite effects. These results suggested that BMDC A2AR could inhibit white matter lesions and attenuate cognitive impairment after CHWMLs, whereas non-BMDC A2ARs aggravate cognitive impairment. The systemic inflammatory response and local activated microglia with cystatin F high expression were involved in the process of cognitive function recovery with BMDC A2AR. The overall trend is that BMDC A2ARs play a leading role.

Relation of Retinal Oxygen Measures to Electrophysiology and Survival Indicators after Permanent, Incomplete Ischemia in Rats.

Studies in experimental ischemia models by permanent bilateral common carotid artery occlusion (BCCAO) have reported reduced retinal electrophysiological function, coupled with inner retinal degeneration and gliosis. In the current study, we tested the hypothesis that long-term (up to 14 days) BCCAO impairs oxygen delivery (DO2), which affects oxygen metabolism (MO2) and extraction fraction (OEF), electrophysiological function, morphology, and biochemical pathways. Twenty-one rats underwent BCCAO (N = 12) or sham surgery (N = 9) and were evaluated in separate groups after 3, 7, or 14 days. Electroretinography (ERG), optical coherence tomography, blood flow and vascular oxygen tension imaging, and morphological and biochemical evaluations were performed in both eyes. Reduced ERG b-wave amplitudes and delayed implicit times were reported at 3, 7, and 14 days following BCCAO. Total retinal blood flow, MO2, and DO2 were reduced in all BCCAO groups. OEF was increased in both 3- and 7-day groups, while no significant difference was observed in OEF at 14 days compared to the sham group. At 14 days following BCCAO, total and inner retinal layer thickness was reduced, while the outer nuclear layer thickness and gliosis were increased. There was an increase in nuclei containing fragmented DNA at 3 days following BCCAO. The compensatory elevation in OEF following BCCAO did not meet the tissue demand, resulting in the subsequent reduction of MO2. The associations between retinal MO2, DO2, and retinal function were shown to be significant in the sequelae of persistent ischemia. In sum, measurements of DO2, MO2, and OEF may become useful for characterizing salvageable tissue in vision-threatening pathologies.

Microglial Hv1 proton channels promote white matter injuries after chronic hypoperfusion in mice.

Microglia are critical in damage/repair processes during ischemic white matter injury (WMI). Voltage-gated proton channel (Hv1) is expressed in microglia and contributes to nicotinamide adenine dinucleotide phosphate oxidase complex-dependent production of reactive oxygen species (ROS). Recent findings have shown that Hv1 is involved in regulating luminal pH of M1-polarized microglial phagosomes and inhibits endocytosis in microglia. We previously reported that Hv1 facilitated production of ROS and pro-inflammatory cytokines in microglia and enhanced damage to oligodendrocyte progenitor cells from oxygen and glucose deprivation. To investigate the role of Hv1 in hypoperfusion-induced WMI, we employed mice that were genetically devoid of Hv1 (Hv1-/- ), as well as a model of subcortical vascular dementia via bilateral common carotid artery stenosis. Integrity of myelin was assessed using immunofluorescent staining and transmission electron microscopy, while cognitive impairment was assessed using an eight-arm radial maze test. Hv1 deficiency was found to attenuate bilateral common carotid artery stenosis-induced disruption of white matter integrity and impairment of working memory. Immunofluorescent staining and western blotting were used to assay changes in oligodendrocytes, OPCs, and microglial polarization. Compared with that in wild-type (WT) mice, Hv1-/- mice exhibited reduced ROS generation, decreased pro-inflammatory cytokines production, and an M2-dominant rather than M1-dominant microglial polarization. Furthermore, Hv1-/- mice exhibited enhanced OPC proliferation and differentiation into oligodendrocytes. Results of mouse-derived microglia-OPC co-cultures suggested that PI3K/Akt signaling was involved in Hv1-deficiency-induced M2-type microglial polarization and concomitant OPC differentiation. These results suggest that microglial Hv1 is a promising therapeutic target for reducing ischemic WMI and cognitive impairment.

Ocular Ischemic Syndrome With Congenital Absence of Internal Carotid Artery.

Purpose: This article describes a case of ocular ischemic syndrome (OIS) in a patient with a congenitally absent left internal carotid artery (ICA). Methods: Retrospective case report with anterior-segment and fundus photography, fluorescein angiography (FA), and computerized tomography angiography (CT-A). Results: A 31-year-old-man was found to have neovascularization of the iris (NVI) and angle of the left eye. FA showed capillary nonperfusion in the temporal periphery. He required intravitreal bevacizumab and triamcinolone injections and 2 panretinal photocoagulation treatments for persistent rubeosis and cystoid macular edema. Transient right-eye vision loss prompted CT-A, revealing an absent left ICA. Three years following presentation, FA continued to show delayed arteriovenous flow suggestive of OIS. He has required intravitreal bevacizumab injections every 12 weeks for persistent NVI. Conclusion: Congenitally absent left ICA with resultant cerebrovascular insufficiency is a rare cause of OIS, underscoring the pathophysiological principles of insufficient blood supply to the ophthalmic artery.

Estradiol Protects White Matter of Male C57BL6J Mice against Experimental Chronic Cerebral Hypoperfusion.

BACKGROUND AND PURPOSE: Estradiol is a sex steroid hormone known to protect the brain against damage related to transient and global cerebral ischemia. In the present study, we leverage an experimental murine model of bilateral carotid artery stenosis (BCAS) to examine the putative effects of estradiol therapy on chronic cerebral hypoperfusion. We hypothesize that long-term estradiol therapy protects against white matter injury and declarative memory deficits associated with chronic cerebral hypoperfusion. METHODS: Adult male C57BL/6J mice underwent either surgical BCAS or sham procedures. Two days after surgery, the mice were given oral estradiol (Sham+E, BCAS+E) or placebo (Sham+P, BCAS+P) treatments daily for 31-34 days. All mice underwent Novel Object Recognition (NOR) testing 31-34 days after the start of oral treatments. Following sacrifice, blood was collected and brains fixed, sliced, and prepared for histological examination of white matter injury and extracellular signal-regulated kinase (ERK) expression. RESULTS: Animals receiving long-term oral estradiol therapy (BCAS-E2 and Sham-E2) had higher plasma estradiol levels than those receiving placebo treatment (BCAS-P and Sham-P). BCAS-E2 mice demonstrated less white matter injury (Klüver-Barrera staining) and performed better on the NOR task compared to BCAS-P mice. ERK expression in the brain was increased in the BCAS compared to sham cohorts. Among the BCAS mice, the BCAS-E2 cohort had a greater number of ERK + cells. CONCLUSION: This study demonstrates a potentially protective role for oral estradiol therapy in the setting of white matter injury and declarative memory deficits secondary to murine chronic cerebral hypoperfusion.

Long-Term Potentiation Enhances Neuronal Differentiation in the Chronic Hypoperfusion Model of Rats.

Several reports have shown that long-term potentiation (LTP) per se effectively enhances neurogenesis in the hippocampus of intact animals. If LTP can enhance neurogenesis in chronic hypoperfusion, this approach could potentially become a new therapeutic strategy for the restoration of cognitive function and for prevention from deterioration of mild cognitive impairment (MCI). Using an in vivo LTP model of rats, we examined whether LTP per se can enhance neurogenesis in hypoperfusion rats that underwent permanent bilateral common carotid artery occlusion (permanent 2-vessel occlusion, P2VO). High frequency stimulation (HFS) in the subacute phase after P2VO enhanced hippocampal cell proliferation and neurogenesis. However, most enhanced cell proliferation and neurogenesis was seen in the hypoperfusion rats that received HFS and for which LTP could finally be induced. In contrast, the same effect was not seen in the LTP induction in the chronic phase. The present findings, which reveal that most enhanced neurogenesis was seen in hypoperfusion rats for which LTP could be finally induced, could explain the ability of LTP-like activities such as learning paradigms and environmental stimuli to increase the rate of neurogenesis in the hippocampus even under hypoperfusion conditions. Moreover, the present findings, which reveal that LTP induction in the chronic phase after P2VO could not effectively enhance neurogenesis in the hypoperfusion rats, could indicate that patients with MCI and even middle-aged healthy control individuals should start LTP-like activities as early as possible and continue with these activities to prevent age-related deterioration of hippocampal function.

Increased homocysteine levels impair reference memory and reduce cortical levels of acetylcholine in a mouse model of vascular cognitive impairment.

Folates are B-vitamins that are vital for normal brain function. Deficiencies in folates either genetic (methylenetetrahydrofolate reductase, MTHFR) or dietary intake of folic acid result in elevated levels of homocysteine. Clinical studies have shown that elevated levels of homocysteine (Hcy) may be associated with the development of dementia, however this link remains unclear. The purpose of this study was to evaluate the impact of increased Hcy levels on a mouse model of vascular cognitive impairment (VCI) produced by chronic hypoperfusion. Male and female Mthfr+/+ and Mthfr+/- mice were placed on either control (CD) or folic acid deficient (FADD) diets after which all animals underwent microcoil implantation around each common carotid artery or a sham procedure. Post-operatively animals were tested on the Morris water maze (MWM), y-maze, and rotarod. Animals had no motor impairments on the rotarod, y-maze, and could learn the location of the platform on the MWM. However, on day 8 of testing of MWM testing during the probe trial, Mthfr+/- FADD microcoil mice spent significantly less time in the target quadrant when compared to Mthfr+/- CD sham mice, suggesting impaired reference memory. All FADD mice had elevated levels of plasma homocysteine. MRI analysis revealed arterial remodeling was present in Mthfr+/- microcoil mice not Mthfr+/+ mice. Acetylcholine and related metabolites were reduced in cortical tissue because of microcoil implantation and elevated levels of homocysteine. Deficiencies in folate metabolism resulting in increased Hcy levels yield a metabolic profile that increases susceptibility to neurodegeneration in a mouse model of VCI.

Effect of hypertension and carotid occlusion on brain parenchymal arteriole structure and reactivity.

We studied the effect of hypertension and chronic hypoperfusion on brain parenchymal arteriole (PA) structure and function. PAs were studied isolated and pressurized from 18-wk-old Wistar-Kyoto (WKY18; n = 8) and spontaneously hypertensive stroke prone (SHRSP18; n = 8) and 5-wk-old prehypertensive (SHRSP5; n = 8) rats. In separate groups, unilateral common carotid artery occlusion (UCCAo) was performed for 4 wk to cause chronic hypoperfusion in 18-wk-old WKY (WKY18-CH; n = 8) and SHRSP (SHRSP18-CH; n = 8). UCCAo caused PAs to have significantly diminished myogenic tone (31 ± 3 vs. 14 ± 6% at 60 mmHg; P < 0.05) and reactivity to pressure from WKY18-CH vs. WKY18 animals. The effect of UCCAo was limited to normotensive animals, as there was little effect of chronic hypoperfusion on vascular reactivity or percent tone in PAs from SHRSP18 vs. SHRSP18-CH animals (53 ± 4 vs. 41 ± 3%; P > 0.05). However, PAs from SHRSP18 and SHRSP5 animals had significantly greater tone compared with WKY18, suggesting an effect of strain and not hypertension per se on PA vasoconstriction. Structurally, PAs from SHRSP18 and SHRSP5 animals had similar sized lumen diameters, but increased wall thickness and distensibility compared with WKY18. Interestingly, chronic hypoperfusion did not affect the structure of PAs from either WKY18-CH or SHRSP18-CH animals. Thus PAs responded to UCCAo with active vasodilation, but not structural remodeling, an effect that was absent in SHRSP. The increased tone of PAs from SHRSP animals, combined with lack of response to chronic hypoperfusion, may contribute to the propensity for ischemic lesions and increased perfusion deficit during hypertension.

Elevated levels of plasma homocysteine, deficiencies in dietary folic acid and uracil-DNA glycosylase impair learning in a mouse model of vascular cognitive impairment.

Dietary deficiencies in folic acid result in elevated levels of plasma homocysteine, which has been associated with the development of dementia and other neurodegenerative disorders. Previously, we have shown that elevated levels of plasma homocysteine in mice deficient for a DNA repair enzyme, uracil-DNA glycosylase (UNG), result in neurodegeneration. The goal of this study was to evaluate how deficiencies in folic acid and UNG along with elevated levels of homocysteine affect vascular cognitive impairment, via chronic hypoperfursion in an animal model. Ung(+/+) and Ung(-/-) mice were placed on either control (CD) or folic acid deficient (FADD) diets. Six weeks later, the mice either underwent implantation of microcoils around both common carotid arteries. Post-operatively, behavioral tests began at 3-weeks, angiography was measured after 5-weeks using MRI to assess vasculature and at completion of study plasma and brain tissue was collected for analysis. Learning impairments in the Morris water maze (MWM) were observed only in hypoperfused Ung(-/-) FADD mice and these mice had significantly higher plasma homocysteine concentrations. Interestingly, Ung(+/+) FADD produced significant remodeling of the basilar artery and arterial vasculature. Increased expression of GFAP was observed in the dentate gyrus of Ung(-/-) hypoperfused and FADD sham mice. Chronic hypoperfusion resulted in increased cortical MMP-9 protein levels of FADD hypoperfused mice regardless of genotypes. These results suggest that elevated levels of homocysteine only, as a result of dietary folic acid deficiency, don't lead to memory impairments and neurobiochemical changes. Rather a combination of either chronic hypoperfusion or UNG deficiency is required.

Cognitive dysfunction in 16 patients with carotid stenosis: detailed neuropsychological findings.

BACKGROUND: Impairment of cognitive function is often present in patients with carotid artery stenosis but the details of this dysfunction have rarely been reported. Our purpose was to elucidate the cognitive dysfunction in patients with unilateral severe carotid stenosis using comprehensive neuropsychological testing, and also to identify the specific underlying clinical and radiological factors. METHODS: We analyzed the results of neuropsychological testing, the clinical history, and MR findings in 16 consecutive patients with angiographically proven severe (70-99%) stenosis of the extra cranial internal carotid artery (ICA). Cognitive functions were examined using the Seoul Neuropsychological Screening Battery and the Neglect Battery. We excluded patients with cortical infarction and those with contra lateral ICA occlusion or severe stenosis. RESULTS: Our comprehensive neuropsychological testing revealed obvious cognitive deficits in all patients with unilateral severe ICA stenosis, the most common being frontal executive impairment. The mean cognitive score on the memory test was also significantly lower in patients with symptomatic ICA stenosis than in asymptomatic patients (29.33+/-10.98, mean+/-SD, p < 0.05). The total score on the global cognitive test was significantly lower in patients with an ischemic lesion on MRI than in no lesion patients (113.23+/-34.78, p < 0.05). The presence of symptoms related to the ICA stenosis was related to cognitive dysfunction even when there were no ischemic lesions on MRI. SPECT revealed ipsilateral cortical hypoperfusion in 9 of 12 patients (75%). CONCLUSIONS: Cognitive deficits are common in patients with unilateral severe ICA stenosis. Our findings suggest that an additional mechanism beyond the structural lesion such as chronic hypoperfusion may affect cognitive function in patients with high-grade ICA stenosis.