[Show abstract][Hide abstract] ABSTRACT: Because our recent studies have demonstrated that miR-122 decreased in whole blood of patients and in whole blood of rats following ischemic stroke, we tested whether elevating blood miR-122 would improve stroke outcomes in rats. Young adult rats were subjected to a temporary middle cerebral artery occlusion (MCAO) or sham operation. A polyethylene glycol-liposome-based transfection system was used to administer a miR-122 mimic after MCAO. Neurological deficits, brain infarction, brain vessel integrity, adhesion molecule expression and expression of miR-122 target and indirect-target genes were examined in blood at 24 h after MCAO with or without miR-122 treatment. miR-122 decreased in blood after MCAO, whereas miR-122 mimic elevated miR-122 in blood 24 h after MCAO. Intravenous but not intracerebroventricular injection of miR-122 mimic decreased neurological deficits and brain infarction, attenuated ICAM-1 expression, and maintained vessel integrity after MCAO. The miR-122 mimic also down-regulated direct target genes (e.g. Vcam1, Nos2, Pla2g2a) and indirect target genes (e.g. Alox5, Itga2b, Timp3, Il1b, Il2, Mmp8) in blood after MCAO which are predicted to affect cell adhesion, diapedesis, leukocyte extravasation, eicosanoid and atherosclerosis signaling. The data show that elevating miR-122 improves stroke outcomes and we postulate this occurs via downregulating miR-122 target genes in blood leukocytes.
Preview · Article · Oct 2015 · Journal of Cerebral Blood Flow & Metabolism
[Show abstract][Hide abstract] ABSTRACT: Small noncoding RNAs play a critical role in regulating messenger RNA throughout brain development and when altered could have profound effects leading to disorders such as autism spectrum disorders (ASD). We assessed small noncoding RNAs, including microRNA and small nucleolar RNA, in superior temporal sulcus association cortex and primary auditory cortex in typical and ASD brains from early childhood to adulthood. Typical small noncoding RNA expression profiles were less distinct in ASD, both between regions and changes with age. Typical micro-RNA coexpression associations were absent in ASD brains. miR-132, miR-103, and miR-320 micro-RNAs were dysregulated in ASD and have previously been associated with autism spectrum disorders. These diminished region- and age-related micro-RNA expression profiles are in line with previously reported findings of attenuated messenger RNA and long noncoding RNA in ASD brain. This study demonstrates alterations in superior temporal sulcus in ASD, a region implicated in social impairment, and is the first to demonstrate molecular alterations in the primary auditory cortex.
[Show abstract][Hide abstract] ABSTRACT: Autism spectrum disorders (ASDs) likely involve dysregulation of multiple genes related to brain function and development. Abnormalities in individual regulatory small non-coding RNA (sncRNA), including microRNA (miRNA), could have profound effects upon multiple functional pathways. We assessed whether a brain region associated with core social impairments in ASD, the superior temporal sulcus (STS), would evidence greater transcriptional dysregulation of sncRNA than adjacent, yet functionally distinct, primary auditory cortex (PAC).
We measured sncRNA expression levels in 34 samples of postmortem brain from STS and PAC to find differentially expressed sncRNA in ASD compared with control cases. For differentially expressed miRNA, we further analyzed their predicted mRNA targets and carried out functional over-representation analysis of KEGG pathways to examine their functional significance and to compare our findings to reported alterations in ASD gene expression.
Two mature miRNAs (miR-4753-5p and miR-1) were differentially expressed in ASD relative to control in STS and four (miR-664-3p, miR-4709-3p, miR-4742-3p, and miR-297) in PAC. In both regions, miRNA were functionally related to various nervous system, cell cycle, and canonical signaling pathways, including PI3K-Akt signaling, previously implicated in ASD. Immune pathways were only disrupted in STS. snoRNA and pre-miRNA were also differentially expressed in ASD brain.
Alterations in sncRNA may underlie dysregulation of molecular pathways implicated in autism. sncRNA transcriptional abnormalities in ASD were apparent in STS and in PAC, a brain region not directly associated with core behavioral impairments. Disruption of miRNA in immune pathways, frequently implicated in ASD, was unique to STS.
[Show abstract][Hide abstract] ABSTRACT: Neutrophils have key roles in ischemic brain injury, thrombosis, and atherosclerosis. As such, neutrophils are of great interest as targets to treat and prevent ischemic stroke. After stroke, neutrophils respond rapidly promoting blood-brain barrier disruption, cerebral edema, and brain injury. A surge of neutrophil-derived reactive oxygen species, proteases, and cytokines are released as neutrophils interact with cerebral endothelium. Neutrophils also are linked to the major processes that cause ischemic stroke, thrombosis, and atherosclerosis. Thrombosis is promoted through interactions with platelets, clotting factors, and release of prothrombotic molecules. In atherosclerosis, neutrophils promote plaque formation and rupture by generating oxidized-low density lipoprotein, enhancing monocyte infiltration, and degrading the fibrous cap. In experimental studies targeting neutrophils can improve stroke. However, early human studies have been met with challenges, and suggest that selective targeting of neutrophils may be required. Several properties of neutrophil are beneficial and thus may important to preserve in patients with stroke including antimicrobial, antiinflammatory, and neuroprotective functions.Journal of Cerebral Blood Flow & Metabolism advance online publication, 25 March 2015; doi:10.1038/jcbfm.2015.45.
No preview · Article · Mar 2015 · Journal of cerebral blood flow and metabolism: official journal of the International Society of Cerebral Blood Flow and Metabolism
[Show abstract][Hide abstract] ABSTRACT: Ischemia, white matter injury, and Alzheimer's disease (AD) pathologies often co-exist in aging brain. How one condition predisposes to, interacts with, or perhaps causes the others remains unclear.
To better understand the link between ischemia, white matter injury, and AD, adult rats were administered lipopolysaccharide (LPS) to serve as an inflammatory stimulus, and 24 h later subjected to 20-min focal cerebral ischemia (IS) followed by 30-min hypoxia (H).
Myelin and axonal damage, as well as amyloid-β (Aβ) and amyloid-β protein precursor (AβPP) deposition were examined by western blot and immunocytochemistry following LPS/IS/H. Findings were compared to the 5XFAD mouse AD brain.
Myelin/axonal injury was observed bilaterally in cortex following LPS/IS/H, along with an increase in IL-1, granzyme B, and LPS. AβPP deposition was present in ischemic striatum in regions of myelin loss. Aβ1-42 and AβPP were deposited in small foci in ischemic cortex that co-localized with myelin aggregates. In the 5XFAD mouse AD model, cortical amyloid plaques also co-localized with myelin aggregates.
LPS/IS/H produce myelin injury and plaque-like aggregates of myelin. AβPP and Aβ co-localize with these myelin aggregates.
No preview · Article · Mar 2015 · Journal of Alzheimer's disease: JAD
[Show abstract][Hide abstract] ABSTRACT: The goal of this study was to show that myelin and axons in cortical gray matter are damaged in Alzheimer's disease (AD) brain. Superior temporal gyrus gray matter of AD patients (9 male, 14 female) was compared to cognitively normal controls (8 male, 7 female). Myelin basic protein (MBP) and a degraded myelin basic protein complex (dMBP) were quantified by Western blot. Brain sections were immunostained for MBP, dMBP, axonal neurofilament protein (NF), autophagy marker microtubule-associated proteins 1A/B light chain 3B precursor (LC3B), amyloid-β protein precursor (AβPP), and amyloid markers amyloid β1-42 (Aβ1-42) and FSB. Co-immunoprecipitation and mass spectroscopy evaluated interaction of AβPP/Aβ1-42 with MBP/dMBP. Evidence of axonal injury in AD cortex included appearance of AβPP in NF stained axons, and NF at margins of amyloid plaques. Evidence of myelin injury in AD cortex included (1) increased dMBP in AD gray matter compared to control (p < 0.001); (2) dMBP in AD neurons; and (3) increased LC3B that co-localized with MBP. Evidence of interaction of AβPP/Aβ1-42 with myelin or axonal components included (1) greater binding of dMBP with AβPP in AD brain; (2) MBP at the margins of amyloid plaques; (3) dMBP co-localized with Aβ1-42 in the core of amyloid plaques in AD brains; and (4) interactions between Aβ1-42 and MBP/dMBP by co-immunoprecipitation and mass spectrometry. We conclude that damaged axons may be a source of AβPP. dMBP, MBP, and NF associate with amyloid plaques and dMBP associates with AβPP and Aβ1-42. These molecules could be involved in formation of amyloid plaques.
Full-text · Article · Feb 2015 · Journal of Alzheimer's disease: JAD
[Show abstract][Hide abstract] ABSTRACT: Deletion of the 1.5-3 Mb region of chromosome 22 at locus 11.2 gives rise to the chromosome 22q11.2 deletion syndrome (22q11DS), also known as DiGeorge and Velocardiofacial Syndromes. It is the most common micro-deletion disorder in humans and one of the most common multiple malformation syndromes. The syndrome is characterized by a broad phenotype, whose characterization has expanded considerably within the last decade and includes many associated findings such as craniofacial anomalies (40%), conotruncal defects of the heart (CHD; 70-80%), hypocalcemia (20-60%), and a range of neurocognitive anomalies with high risk of schizophrenia, all with a broad phenotypic variability. These phenotypic features are believed to be the result of a change in the copy number or dosage of the genes located in the deleted region. Despite this relatively clear genetic etiology, very little is known about which genes modulate phenotypic variations in humans or if they are due to combinatorial effects of reduced dosage of multiple genes acting in concert. Here, we report on decreased expression levels of genes within the deletion region of chromosome 22, including DGCR8, in peripheral leukocytes derived from individuals with 22q11DS compared to healthy controls. Furthermore, we found dysregulated miRNA expression in individuals with 22q11DS, including miR-150, miR-194 and miR-185. We postulate this to be related to DGCR8 haploinsufficiency as DGCR8 regulates miRNA biogenesis. Importantly we demonstrate that the level of some miRNAs correlates with brain measures, CHD and thyroid abnormalities, suggesting that the dysregulated miRNAs may contribute to these phenotypes and/or represent relevant blood biomarkers of the disease in individuals with 22q11DS.
[Show abstract][Hide abstract] ABSTRACT: Aims
Epidemiological studies suggest that sex has a role in the pathogenesis of cardioembolic stroke. Since stroke is a vascular disease, identifying sexually dimorphic gene expression changes in blood leukocytes can inform on sex-specific risk factors, response and outcome biology. We aimed to examine the sexually dimorphic immune response following cardioembolic stroke by studying the differential gene expression in peripheral white blood cells.
Methods and Results
Blood samples from patients with cardioembolic stroke were obtained at ≤3 hours (prior to treatment), 5 hours and 24 hours (after treatment) after stroke onset (n = 23; 69 samples) and compared with vascular risk factor controls without symptomatic vascular diseases (n = 23, 23 samples) (ANCOVA, false discovery rate p≤0.05, |fold change| ≥1.2). mRNA levels were measured on whole-genome Affymetrix microarrays. There were more up-regulated than down-regulated genes in both sexes, and females had more differentially expressed genes than males following cardioembolic stroke. Female gene expression was associated with cell death and survival, cell-cell signaling and inflammation. Male gene expression was associated with cellular assembly, organization and compromise. Immune response pathways were over represented at ≤3, 5 and 24 h after stroke in female subjects but only at 24 h in males. Neutrophil-specific genes were differentially expressed at 3, 5 and 24 h in females but only at 5 h and 24 h in males.
There are sexually dimorphic immune cell expression profiles following cardioembolic stroke. Future studies are needed to confirm the findings using qRT-PCR in an independent cohort, to determine how they relate to risk and outcome, and to compare to other causes of ischemic stroke.
[Show abstract][Hide abstract] ABSTRACT: Background: The superior temporal sulcus (STS) plays a critical role in social behavior, a core impairment in autism, yet the molecular mechanisms that underlie abnormal STS function remain largely unexplored. Small non-coding RNAs (sncRNA), including microRNA (miRNA) and small nucleolar RNA (snoRNA), show increased importance as key regulators of translation during development and throughout life. Aberrant expression of sncRNA could lead to widespread changes in protein and cellular function related to autism. This study evaluated differences is sncRNA expression in the STS in the autism brain. In addition, this study sought to determine if differences were localized to association cortices, such as STS, or may also include neural regions involved in more basic perceptual processing, such as the primary auditory cortex (PAC), that are not typically associated with the core impairments of autism.
Objectives: Examine the changes in expression of small non-coding RNAs expressed in association (STS) and primary (PAC) cortex of autism and typically developing brains.
Methods: Brain tissue from subjects diagnosed with autism (n=10) and typically developing controls (n=8) was obtained from the Harvard Brain Tissue Resource Center. The STS and PAC were dissected from each fresh-frozen brain. Total RNA was isolated from each region and assessed for concentration and quality. Two hundred nanograms of total RNA were processed on Affymetrix GeneChip miRNA 3.0 Arrays. Arrays were scanned and resulting CEL files analyzed with Partek Genomics Suite. Only small non-coding RNAs (mature miRNA, precursor miRNA and snoRNA) with human annotation (5663 targets) were included in the analysis. Functional significance of altered miRNA was determined through analysis of over-representation of their computationally derived mRNA targets in KEGG pathways using DIANA miRPath and Exploratory Gene Association Networks (EGAN) software.
Results: In STS, the expression of 3 miRNA significantly differed (P<0.005, FC>|1.2|) between autism and typically developing control brains (miR-1, miR-4753-5p, and miR-513a-5p). These miRNA regulate pathways relating to synapse, brain maturation function and processes, and immune function. An additional 11 stem-loop precursor miRNA and 6 snoRNA were also different in STS. In PAC, 3 miRNA were significantly different in autism compared to typical control (miR-297, miR-664, and miR-4709-3p). These were unique to PAC and may represent affected function of other cell signaling pathways and developmental cues. There were also 7 stem-loop precursor miRNA and 4 snoRNA differentially expressed in PAC. Commonly affected elements/functions of miRNA in both regions, include Akt and glutamate signalling. The presence of significantly altered snoRNA in autism brain may implicate alterations in splicing mechanisms and patterns as important contributors to autism.
Conclusions: Regional differences in miRNA and other sncRNA expression were observed between autism and control brains in the STS and PAC. These changes in small regulatory non-coding RNA may have important regulatory effects on mRNA transcript splicing and translation to proteins. These findings help identify molecular mechanisms underlying autism and elucidate specific targets to restore perturbations that might occur in autism.
[Show abstract][Hide abstract] ABSTRACT: Defining the RNA transcriptome in Alzheimer Disease (AD) will help understand the disease mechanisms and provide biomarkers. Though the AD blood transcriptome has been studied, effects of white matter hyperintensities (WMH) were not considered. This study investigated the AD blood transcriptome and accounted for WMH.
RNA from whole blood was processed on whole-genome microarrays.
A total of 293 probe sets were differentially expressed in AD versus controls, 5 of which were significant for WMH status. The 288 AD-specific probe sets classified subjects with 87.5% sensitivity and 90.5% specificity. They represented 188 genes of which 29 have been reported in prior AD blood and 89 in AD brain studies. Regulated blood genes included MMP9, MME (Neprilysin), TGFβ1, CA4, OCLN, ATM, TGM3, IGFR2, NOV, RNF213, BMX, LRRN1, CAMK2G, INSR, CTSD, SORCS1, SORL1, and TANC2.
RNA expression is altered in AD blood irrespective of WMH status. Some genes are shared with AD brain.
[Show abstract][Hide abstract] ABSTRACT: Despite testing more than 1,026 therapeutic strategies in models of ischemic stroke and 114 therapies in human ischemic stroke, only one agent tissue plasminogen activator has successfully been translated to clinical practice as a treatment for acute stroke. Though disappointing, this immense body of work has led to a rethinking of animal stroke models and how to better translate therapies to patients with ischemic stroke. Several recommendations have been made, including the STAIR recommendations and statements of RIGOR from the NIH/NINDS. In this commentary we discuss additional aspects that may be important to improve the translational success of ischemic stroke therapies. These include use of tissue plasminogen activator in animal studies; modeling ischemic stroke heterogeneity in terms of infarct size and cause of human stroke; addressing the confounding effect of anesthesia; use of comparable therapeutic dosage between humans and animals based on biological effect; modeling the human immune system; and developing outcome measures in animals comparable to those used in human stroke trials. With additional study and improved animal modeling of factors involved in human ischemic stroke, we are optimistic that new stroke therapies will be developed.
No preview · Article · Feb 2014 · Metabolic Brain Disease
[Show abstract][Hide abstract] ABSTRACT: Objective:
Myelin disruption is an important feature of Alzheimer's disease (AD) that contributes to impairment of neuronal circuitry and cognition. In this study we characterize myelin degradation in the brains of patients with Alzheimer's disease compared with normal aged controls.
Myelin from patients with AD (n=13) was compared to matched controls (n=6). Myelin degradation was examined by immunohistochemistry in frontal white matter (WM) for intact myelin basic protein (MBP), degraded MBP, the presence of myelin lipid and for PAS staining. The relationship of myelin degradation and axonal injury was also assessed.
Brains from patients with AD had significant loss of intact MBP, and an increase in degraded MBP in periventricular WM adjacent to a denuded ependymal layer. In regions of myelin degradation, vesicles were identified that stained positive for degraded MBP, myelin lipid, and neurofilament but not for intact MBP. Most vesicles stained for PAS, a corpora amylacea marker. The vesicles were significantly more abundant in the periventricular WM of AD patients compared to controls (44.5 ± 11.0 versus 1.7 ± 1.1, p=0.02).
In AD patients degraded MBP is associated in part with vesicles particularly in periventricular WM that is adjacent to areas of ependymal injury.
No preview · Article · Jan 2014 · Current Alzheimer research
[Show abstract][Hide abstract] ABSTRACT: Traumatic brain injury (TBI) is often associated with intracerebral and intraventricular hemorrhage. Thrombin is a neurotoxin generated at bleeding sites following TBI, and can lead to cell death and subsequent cognitive dysfunction via activation of Src family kinases (SFKs). We hypothesize that inhibiting SFKs can protect hippocampal neurons and improve cognitive memory function following TBI. To test these hypotheses we show that moderate lateral fluid percussion (LFP) TBI in adult rats produces bleeding into the cerebrospinal fluid (CSF) in both lateral ventricles, which elevates oxyhemoglobin and thrombin levels in CSF, activates the SFK family member Fyn, and increases Rho-kinase 1(ROCK1) expression. Systemic administration of the SFK inhibitor, PP2, immediately following moderate TBI blocks ROCK1 expression, protects hippocampal CA2-3 neurons, and improves spatial memory function. These data suggest the possibility that inhibiting SFKs following TBI might improve clinical outcomes.
No preview · Article · Jan 2014 · Journal of neurotrauma
[Show abstract][Hide abstract] ABSTRACT: Autophagy is responsible for the bulk degradation of cytoplasmic contents including organelles through the lysosomal machinery. Neonatal hypoxia–ischemia (HI) causes cell death in the brain by caspase-dependent and independent pathways. Ischemic insults also increase the formation of autophagosomes and activate autophagy. This study assessed the possible sex- and region-specific differences of autophagy activity in neonates subjected to HI brain injury. HI males had a modest decrease in lysosome numbers with no effect on LC3B-II protein in the cortex. In contrast, HI females had decreased lysosome numbers and their LC3B-II protein expression was significantly increased in the cortex following HI. In the hippocampus, both HI males and all females had increased numbers of autolysosomes suggesting activation of autophagy but with no effect on lysosome numbers, or Beclin-1 or LC3B protein levels. Males and females had increases in caspase 3/7 activity in their cortices and hippocampi following HI, though the increases were three to sixfold greater in females. The present data: (a) confirm greater caspase activation in the brains of females compared to males following HI; (b) suggest a partial failure to degrade LC3B-II protein in cortical but not hippocampal lysosomes of females as compared to males following neonatal HI; (c) all females have greater basal autophagy activity than males which may protect cells against injury by increasing cell turnover and (d) demonstrate that autophagy pathways are disturbed in regional- and sex-specific patterns in the rat brain following neonatal HI.
[Show abstract][Hide abstract] ABSTRACT: Family studies have consistently shown that Tourette syndrome (TS) is a familial disorder and twin studies have clearly indicated a genetic contribution in the etiology of TS. Whereas early segregation studies of TS suggested a single-gene autosomal dominant disorder, later studies have pointed to more complex models including additive and multifactorial inheritance and likely interaction with genetic factors. While the exact cellular and molecular base of TS is as yet elusive, neuroanatomical and neurophysiological studies have pointed to the involvement of cortico-striato-thalamocortical circuits and abnormalities in dopamine, glutamate, gamma-aminobutyric acid, and serotonin neurotransmitter systems, with the most consistent evidence being available for involvement of dopamine-related abnormalities, that is, a reduction in tonic extracellular dopamine levels along with hyperresponsive spike-dependent dopamine release, following stimulation. Genetic and gene expression findings are very much supportive of involvement of these neurotransmitter systems. Moreover, intriguingly, genetic work on a two-generation pedigree has opened new research pointing to a role for histamine, a so far rather neglected neurotransmitter, with the potential of the development of new treatment options. Future studies should be aimed at directly linking neurotransmitter-related genetic and gene expression findings to imaging studies (imaging genetics), which enables a better understanding of the pathways and mechanisms through which the dynamic interplay of genes, brain, and environment shapes the TS phenotype.
Full-text · Article · Dec 2013 · International Review of Neurobiology