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The present work addressed the hypothesis that NG2/CSPG4, CD146/MCAM, and VAP1/AOC3 are target genes of myocardin-related transcription factors (MRTFs: myocardin/MYOCD, MRTF-A/MKL1, MRTF-B/MKL2) and serum response factor (SRF). Using a bioinformatics approach, we found that CSPG4, MCAM, and AOC3 correlate with MYOCD, MRTF-A/MKL1, and SRF across human tissues. No other transcription factor correlated as strongly with these transcripts as SRF. Overexpression of MRTFs increased both mRNA and protein levels of CSPG4, MCAM, and AOC3 in cultured human smooth muscle cells (SMCs). Imaging confirmed increased staining for CSPG4, MCAM, and AOC3 in MRTF-A/MKL1-transduced cells. MRTFs exert their effects through SRF, and the MCAM and AOC3 gene loci contained binding sites for SRF. SRF silencing reduced the transcript levels of these genes, and time-courses of induction paralleled the direct target ACTA2. MRTF-A/MKL1 increased the activity of promoter reporters for MCAM and AOC3, and transcriptional activation further depended on the chromatin remodeling enzyme KDM3A. CSPG4, MCAM, and AOC3 responded to the MRTF-SRF inhibitor CCG-1423, to actin dynamics, and to ternary complex factors. Coincidental detection of these proteins should reflect MRTF-SRF activity, and beyond SMCs, we observed co-expression of CD146/MCAM, NG2/CSPG4, and VAP1/AOC3 in pericytes and endothelial cells in the human brain. This work identifies highly responsive vascular target genes of MRTF-SRF signaling that are regulated via a mechanism involving KDM3A.

Recent studies indicate a crucial role for neuronal glycogen storage and degradation in memory formation. We have previously identified alpha‐amylase (α‐amylase), a glycogen degradation enzyme, located within synaptic‐like structures in CA1 pyramidal neurons and shown that individuals with a high copy number variation of α‐amylase perform better on the episodic memory test. We reported that neuronal α‐amylase was absent in patients with Alzheimer's disease (AD) and that this loss corresponded to increased AD pathology. In the current study, we verified these findings in a larger patient cohort and determined a similar reduction in α‐amylase immunoreactivity in the molecular layer of hippocampus in AD patients. Next, we demonstrated reduced α‐amylase concentrations in oligomer amyloid beta 42 (Aβ42) stimulated SH‐SY5Y cells and neurons derived from human‐induced pluripotent stem cells (hiPSC) with PSEN1 mutation. Reduction of α‐amylase production and activity, induced by siRNA and α‐amylase inhibitor Tendamistat, respectively, was further shown to enhance glycogen load in SH‐SY5Y cells. Both oligomer Aβ42 stimulated SH‐SY5Y cells and hiPSC neurons with PSEN1 mutation showed, however, reduced load of glycogen. Finally, we demonstrate the presence of α‐amylase within synapses of isolated primary neurons and show that inhibition of α‐amylase activity with Tendamistat alters neuronal activity measured by calcium imaging. In view of these findings, we hypothesize that α‐amylase has a glycogen degrading function within synapses, potentially important in memory formation. Hence, a loss of α‐amylase, which can be induced by Aβ pathology, may in part underlie the disrupted memory formation seen in AD patients., Alpha‐amylase and glycogen are found in neuronal synapses. Inhibition of alpha‐amylase activity with Tendamistat increases neuronal glycogen load and causes calcium dyshomeostasis. Presence of amyloid beta is associated with reduced alpha‐amylase and glycogen load.

Reduced glucose metabolism and formation of polyglucosan bodies (PGB) are, beside amyloid beta plaques and neurofibrillary tangles, well-known pathological findings associated with Alzheimer's disease (AD). Since both glucose availability and PGB are regulated by enzymatic degradation of glycogen, we hypothesize that dysfunctional glycogen degradation is a critical event in AD progression. We therefore investigated whether alpha (α)-amylase, an enzyme known to efficiently degrade polysaccharides in the gastrointestinal tract, is expressed in the hippocampal CA1/subiculum and if the expression is altered in AD patients. Using immunohistochemical staining techniques, we show the presence of the α-amylase isotypes AMY1A and AMY2A in neuronal dendritic spines, pericytes and astrocytes. Moreover, AD patients showed reduced gene expression of α-amylase, but conversely increased protein levels of α-amylase as well as increased activity of the enzyme compared with non-demented controls. Lastly, we observed increased, albeit not significant, load of periodic acid-Schiff positive PGB in the brain of AD patients, which correlated with increased α-amylase activity. These findings show that α-amylase is expressed and active in the human brain, and suggest the enzyme to be affected, alternatively play a role, in the neurodegenerative Alzheimer's disease pathology.

The biologically active pancreatic hormone peptide islet amyloid polypeptide (IAPP) regulates brain functions such as appetite and cognition. It also plays a role in clearance of amyloid beta (A beta), a peptide implicated in the dementia disorder Alzheimers disease (AD). If IAPP becomes modified, it loses its biological activity and starts to aggregate. Such aggregations have been found in the AD brain and decreased plasma levels of the unmodified IAPP (uIAPP) have been shown in the same patients. In the current study, we analyze levels of uIAPP and total IAPP (unmodified and modified) in cerebrospinal fluid (CSF) to investigate its potential as a biomarker for AD. We found no differences in neither CSF nor plasma levels of uIAPP or total IAPP in AD patients compared to cognitive healthy individuals (NC). The levels of uIAPP in CSF of NC were positively correlated with uIAPP in plasma, Q-albumin and albumin levels in CSF, but negatively correlated with CSF levels of t-tau and p-tau. These findings were not seen in AD patients. Levels of total CSF IAPP correlated positively with total Q-albumin and albumin levels in CSF in both AD and NC. In addition, total plasma IAPP correlated positively with CSF t-tau and p-tau in NC and negatively with CSF A beta(42) in AD patients. To conclude, our studies did not find evidence supporting the use of CSF IAPP as an AD biomarker. However, our findings, indicating a compromised translocation of uIAPP in and out of the brain in AD patients as well as the correlations between total plasma IAPP and AD biomarkers, encourage further research on the role for IAPP in AD. Funding Agencies|Swedish Research CouncilSwedish Research Council [521-2013-3448]; Petrus and Augusta Hedlund foundation [20160493]; Swedish Dementia foundation; Ake Wiberg foundation [M17-0025]; Ahlen foundation [mC23 h18]; Crafoord foundation [20180776]; Greta and Johan Kocks foundations; Alzheimer foundation [AF-640421]; Olle Engkvist [194-0643]; Olle Engkvist foundation; Segerfalk foundation

Cell adhesion molecules (CAMs) mediate interactions between cells and their surroundings that are vital to processes controlling for cell survival, activation, migration, and plasticity. However, increasing evidence suggests that CAMs also mediate mechanisms involved in several neurological diseases. This article reviews the current knowledge on the role of CAMs in amyloid-β (Aβ) metabolism, cell plasticity, neuroinflammation, and vascular changes, all of which are considered central to the pathogenesis and progression of Alzheimer's disease (AD). This paper also outlines the possible roles of CAMs in current and novel AD treatment strategies.

Alzheimer’s disease (AD) is a progressive neurodegenerative disease in which the formation of extracellular aggregates of amyloid beta (Aβ) peptide, fibrillary tangles of intraneuronal tau and microglial activation are major pathological hallmarks. One of the key molecules involved in microglial activation is galectin-3 (gal3), and we demonstrate here for the first time a key role of gal3 in AD pathology. Gal3 was highly upregulated in the brains of AD patients and 5xFAD (familial Alzheimer’s disease) mice and found specifically expressed in microglia associated with Aβ plaques. Single-nucleotide polymorphisms in the LGALS3 gene, which encodes gal3, were associated with an increased risk of AD. Gal3 deletion in 5xFAD mice attenuated microglia-associated immune responses, particularly those associated with TLR and TREM2/DAP12 signaling. In vitro data revealed that gal3 was required to fully activate microglia in response to fibrillar Aβ. Gal3 deletion decreased the Aβ burden in 5xFAD mice and improved cognitive behavior. Interestingly, a single intrahippocampal injection of gal3 along with Aβ monomers in WT mice was sufficient to induce the formation of long-lasting (2 months) insoluble Aβ aggregates, which were absent when gal3 was lacking. High-resolution microscopy (stochastic optical reconstruction microscopy) demonstrated close colocalization of gal3 and TREM2 in microglial processes, and a direct interaction was shown by a fluorescence anisotropy assay involving the gal3 carbohydrate recognition domain. Furthermore, gal3 was shown to stimulate TREM2–DAP12 signaling in a reporter cell line. Overall, our data support the view that gal3 inhibition may be a potential pharmacological approach to counteract AD., This work was supported by Grants from the Swedish Research Council, and the Strong Research Environment MultiPark (Multidisciplinary Research in Parkinson’s and Alzheimer’s Disease at Lund University), Bagadilico (Linné consortium sponsored by the Swedish Research Council), the Swedish Alzheimer’s Foundation, Swedish Brain Foundation, A.E. Berger Foundation, Gyllenstiernska Krapperup Foundation, the Royal Physiographic Society, Crafoord Foundation, Olle Engkvist Byggmästare Foundation, Wiberg Foundation, G&J Kock Foundation, Stohnes Foundation, Swedish Dementia Association and the Medical Faculty at Lund University. This work was supported by Grant SAF2015-64171R (Spanish MINECO/FEDER, UE), by Instituto de Salud Carlos III (ISCiii) of Spain, co-financed by FEDER funds from European Union through grants PI15/00796 and PI18/01557 (to AG), PI15/00957 and PI18/01556 (to JV), and CIBERNED (to AG and JV), by Consejería de Economía, Innovación, Ciencia y Empleo, Junta de Andalucia Proyecto de Excelencia (CTS-2035) (to JV and AG), and by Malaga University grant PPIT.UMA.B1.2017/26 (to RSV). AV and GCB received funding from the Innovative Medicines Initiative 2 Joint Undertaking under Grant agreement no. 115976 (PHAGO). CIBERNED “Centro de Investigacion Biomedica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid (Spain)”. HL and AF were supported by the Swedish Research Council, the Swedish Brain Foundation, the Alzheimer Foundation and the Åhlén Foundation. UJN was supported by Grants from the Knut and Alice Wallenberg Foundation (KAW 2013.0022) and the Swedish Research Council (Grant no. 621-2012-2978).

Alzheimer’s disease (AD) is a progressive neurodegenerative disease in which the formation of extracellular aggregates of amyloid beta (Aβ) peptide, intraneuronal tau neurofibrillary tangles and microglial activation are major pathological hallmarks. One of the key molecules involved in microglial activation is galectin-3 (gal3), and we demonstrate here for the first time a key role of gal3 in AD pathology. Gal3 was highly upregulated in the brains of AD patients and 5xFAD (familial Alzheimer’s disease) mice, and found specifically expressed in microglia associated with Aβ plaques. Single nucleotide polymorphisms in the LGALS3 gene, which encodes gal3, were associated to an increased risk of AD. Gal3 deletion in 5xFAD mice attenuated microglia-associated immune responses, particularly those associated with TLR and TREM2/DAP12 signaling. In vitro data demonstrated the requirement of gal3 to fully activate microglia in response to fibrillar Aβ. Gal3 deletion decreased the Aβ burden in 5xFAD mice and improved cognitive behavior. Electron microscopy of gal3 in AD mice demonstrated i) a preferential expression of gal3 by plaque-associated microglia, ii) its presence in the extracellular space and iii) its association to Aβ plaques. Low concentrations (1 nM) of pure gal3 promoted cross-seeding fibrilization of pure Aβ. Importantly, a single intrahippocampal injection of gal3 along with Aβ monomers in WT mice was sufficient to induce the formation of insoluble Aβ aggregates that were absent when gal3 was lacking. High-resolution microscopy (STORM) demonstrated close co-localization of gal3 and TREM2 in microglial processes, and a direct interaction was shown by a fluorescence anisotropy assay involving the gal3 CRD domain. Furthermore, gal3 stimulated the TREM2-DAP12 signaling pathway. In conclusion, we provide evidence that gal3 is a central regulator of microglial immune response in AD. It drives proinflammatory activation and Aβ aggregation, as well as acting as an endogenous ligand to TREM2, a key receptor driving microglial response under disease conditions. Gal3 inhibition may, hence, be a potential pharmacological approach to counteract AD.

The population of brain pericytes, a cell type important for vessel stability and blood brain barrier function, has recently been shown altered in patients with Alzheimer's disease (AD). The underlying reason for this alteration is not fully understood, but progressive accumulation of the AD characteristic peptide amyloid-beta (Aβ) has been suggested as a potential culprit. In the current study, we show reduced number of hippocampal NG2+ pericytes and an association between NG2+ pericyte numbers and Aβ1-40 levels in AD patients. We further demonstrate, using in vitro studies, an aggregation-dependent impact of Aβ1-40 on human NG2+ pericytes. Fibril-EP Aβ1-40 exposure reduced pericyte viability and proliferation and increased caspase 3/7 activity. Monomer Aβ1-40 had quite the opposite effect: increased pericyte viability and proliferation and reduced caspase 3/7 activity. Oligomer-EP Aβ1-40 had no impact on either of the cellular events. Our findings add to the growing number of studies suggesting a significant impact on pericytes in the brains of AD patients and suggest different aggregation forms of Aβ1-40 as potential key regulators of the brain pericyte population size. Listed as co-author in the publication: The Netherlands Brain Bank, Netherlands Institute for Neuroscience, Amsterdam, The Netherlands

Supplemental digital content is available in the text., Deposition of amyloid-β (Aβ) 1–42, the major component of senile plaques characteristic of Alzheimer disease, affects brain microvascular integrity and causes blood-brain barrier dysfunction, increased angiogenesis, and pericyte degeneration. To understand the cellular events underlying Aβ1–42 effects on microvascular alterations, we investigated whether different aggregation forms of Aβ1–42 affect shedding of the pericyte proteoglycan NG2 and whether they affect proteolytic cleavage mediated by matrix metalloproteinase (MMP)-9. We found decreased levels of soluble NG2, total MMP-9, and MMP-9 activity in pericyte culture supernatants in response to fibril-enriched preparations of Aβ1–42. Conversely, oligomer-enriched preparations of Aβ1–42 increased soluble NG2 levels in the supernatants. This increase was ablated by the MMP-9/MMP-2 inhibitor SB-3CT. There was also a trend toward increased MMP-9 activity observed after oligomeric Aβ1–42 exposure. Our results, demonstrating an Aβ1–42 aggregation-dependent effect on levels of NG2 and MMP-9, support previous studies showing an impact of Aβ1–42 on vascular integrity and thereby add to our understanding of mechanisms behind the microvascular changes commonly found in patients with Alzheimer disease.

Introduction Inflammatory processes have previously been shown to influence cognition and progression of dementia. An involvement of interleukin (IL)-6 has in particular been suggested as altered levels of IL-6 in cerebrospinal fluid (CSF) have been found in patients with Alzheimer’s disease (AD). Also, an association between cognitive decline and levels of IL-6 in CSF have been reported. The aim of the present study was to investigate whether patients clinically diagnosed with dementia with Lewy bodies (DLB) display altered CSF IL-6 levels in comparison with patients with AD and control subjects without dementia and whether the IL-6 levels are correlated with cognitive status and biomarkers for AD and synucleinopathy. Methods To analyse CSF of patients with AD (n = 45), patients with DLB (n = 29) and control subjects without dementia (n = 36), we used immunoassays to measure levels of IL-6 (multiplex electrochemiluminescence); AD markers phosphorylated tau, total tau and amyloid-β1–42 (enzyme-linked immunosorbent assay [ELISA]); and α-synuclein (ELISA). Cognitive status was evaluated using the Mini Mental State Examination (MMSE). Results Our analysis showed significantly lower levels of IL-6 in CSF from patients with DLB than in CSF from patients with AD and control subjects without dementia. The IL-6 levels were also negatively correlated with MMSE and positively correlated with α-synuclein CSF levels. Conclusions Our findings support previous studies by demonstrating a link between inflammatory processes and dementia progression and further strengthen the hypothesis that IL-6 is involved in dementia pathology and cognitive decline.

Analysis of postmortem tissue from patients with major depression and bipolar disorder has revealed structural changes in several brain regions. We have shown that electroconvulsive seizure (ECS), used for the treatment of severe depression, induces proliferation of both neuronal and nonneuronal cells in the adult rat hippocampus.Male Wistar rats were subjected to one or several ECS treatments, then injected with bromodeoxyuridine (BrdU) to detect cell proliferation. Animals were perfused either 1 day or 3 weeks following the last BrdU injection. Cells were double stained for BrdU and the cell type markers chondroitin sulfate proteoglycan (NG2), complement 3-receptor OX-42, 2', 3'-cyclic nucleotide 3'-phosphodiesterase (CNPase), Ca(+) binding protein S100-beta, or neuron-specific nuclear protein (NeuN).We identified NG2-expressing cells as a major cell type proliferating in the rat dentate gyrus in response to ECS. A sharp increase in NG2-positive cell proliferation was seen 2 days after ECS, and a large number of NG2-expressing cells persisted at 3 weeks.Our results show that antidepressant treatment can induce a strong proliferation of glial progenitor cells in the adult rat hippocampus. We propose that this may counteract degenerative changes found in depression and be an important neurobiological event underlying the clinical effect of electroconvulsive seizures.

Major depression is often associated with elevated glucocorticoid levels. High levels of glucocorticoids reduce neurogenesis in the adult rat hippocampus. Electroconvulsive seizures (ECS) can enhance neurogenesis, and we investigated the effects of ECS in rats where glucocorticoid levels were elevated in order to mimic conditions seen in depression. Rats given injections of corticosterone or vehicle for 21 days were at the end of this period treated with either a single or five daily ECSs. Proliferating cells were labelled with bromodeoxyuridine (BrdU). After 3 weeks, BrdU-positive cells in the dentate gyrus were quantified and analyzed for co-labelling with the neuronal marker neuron-specific nuclear protein (NeuN). In corticosterone-treated rats, neurogenesis was decreased by 75%. This was counteracted by a single ECS. Multiple ECS further increased neurogenesis and no significant differences in BrdU/NeuN positive cells were detected between corticosterone- and vehicle-treated rats given five ECS. Approximately 80% of the cells within the granule cell layer and 10% of the hilar cells were double-labelled with BrdU and NeuN. We therefore conclude that electroconvulsive seizures can increase hippocampal neurogenesis even in the presence of elevated levels of glucocorticoids. This further supports the hypothesis that induction of neurogenesis is an important event in the action of antidepressant treatment.

Hyaluronic acid (HA) has been shown to affect angiogenesis and the function of the blood-brain barrier, and a crucial role for HA in atherosclerosis has been described. We have recently demonstrated changes in the levels of HA in cerebrospinal fluid (CSF) in patients with Alzheimer's disease (AD) with documented vascular alterations. To further investigate if the level of HA in CSF can be used as a clinical diagnostic biomarker to identify vascular pathology in dementia, we analyzed the levels of HA in the CSF of patients with vascular dementia (VaD) (n = 46), AD (n = 45), and controls without dementia (n = 26). In line with our previous data, we found significantly increased levels of HA in CSF from patients with VaD compared with controls, whereas the levels of HA in patients with AD were found to be unaltered compared with controls and patients with VaD. We also detected increased levels of HA in individuals with vascular changes determined as significant white matter changes or previous infarction on cranial computed tomography or magnetic resonance imaging, compared with individuals without these findings. Furthermore, we found a significant positive correlation between the levels of HA and the CSF/serum albumin ratio, an indicator of blood-brain barrier integrity, in patients with VaD and AD, supporting the role of HA in vascular changes in the brain. Our results indicate a potential diagnostic value for the detection of vascular brain changes in dementia using CSF levels of HA, but emphasize the importance of further development of more sensitive HA assays.

Background Neuron Glial 2 (NG2) cells are glial cells known to serve as oligodendrocyte progenitors as well as modulators of the neuronal network. Altered NG2 cell morphology and up-regulation as well as increased shedding of the proteoglycan NG2 expressed on the cell surface have been described in rodent models of brain injury. Here we describe alterations in the human NG2 cell population in response to pathological changes characteristic of Alzheimer’s disease (AD). Results Immunohistological stainings of postmortem brain specimens from clinically diagnosed and postmortem verified AD patients and non-demented controls revealed reduced NG2 immunoreactivity as well as large numbers of NG2 positive astrocytes in individuals with high amyloid beta plaque load. Since fibrillar amyloid beta (Aβ)1-42 is the major component of AD-related senile plaques, we exposed human NG2 cells to oligomer- and fibril enriched preparations of Aβ1-42. We found that both oligomeric and fibrillar Aβ1-42 induced changes in NG2 cell morphology. Further, in vitro exposure to fibrillar Aβ1-42 decreased the NG2 concentrations in both cell lysates and supernatants. Interestingly, we also found significantly decreased levels of soluble NG2 in the cerebrospinal fluid (CSF) from clinically diagnosed AD patients compared to non-demented individuals. Additionally, the CSF NG2 levels were found to significantly correlate with the core AD biomarkers Aß1-42, T-tau and P-tau. Conclusion Our results demonstrate major alterations in the NG2 cell population in relation to AD pathology which highlights the NG2 cell population as a new attractive research target in the search for cellular mechanisms associated with AD pathogenesis.

Neurosin is a protease that in vitro degrades α-synuclein, the main constituent of Lewy bodies found in brains of patients with synucleinopathy including Parkinson's disease (PD) and dementia with Lewy bodies (DLB). Several studies have reported reduced cerebrospinal fluid (CSF) levels of α-synuclein in synucleinopathy patients and recent data also proposes a significant role of α-synuclein in the pathophysiology of Alzheimer's disease (AD). To investigate potential links between neurosin and its substrate α-synuclein in vivo we used a commercially available sandwich ELISA and an in-house developed direct ELISA to quantify CSF levels of α-synuclein and neurosin in patients diagnosed with DLB, PD and PD dementia (PDD) versus AD patients and non-demented controls. We found that patients with synucleinopathy displayed lower CSF levels of neurosin and α-synuclein compared to controls and AD patients. In contrast, AD patients demonstrated significantly increased CSF α-synuclein but similar neurosin levels compared to non-demented controls. Further, CSF neurosin and α-synuclein concentrations were positively associated in controls, PD and PDD patients and both proteins were highly correlated to CSF levels of phosphorylated tau in all investigated groups. We observed no effect of gender or presence of the apolipoprotein Eε4 allele on neither neurosin or α-synuclein CSF levels. In concordance with the current literature our study demonstrates decreased CSF levels of α-synuclein in synucleinopathy patients versus AD patients and controls. Importantly, decreased α-synuclein levels in patients with synucleinopathy appear linked to low levels of the α-synuclein cleaving enzyme neurosin. In contrast, elevated levels of α-synuclein in AD patients were not related to any altered CSF neurosin levels. Thus, altered CSF levels of α-synuclein and neurosin in patients with synucleinopathy versus AD may not only mirror disease-specific neuropathological mechanisms but may also serve as fit candidates for future biomarker studies aiming at identifying specific markers of synucleinopathy.

Electroconvulsive therapy (ECT) is a very efficient treatment for severe depression. However, cognitive side effects have raised concern to whether ECT can cause cellular damage in vulnerable brain regions. A few recent animal studies have reported limited hippocampal cell loss, while a number of other studies have failed to find any signs of cellular damage and some even report that electroconvulsive seizures (ECS; the animal counterpart of ECT) has neuroprotective effects. We previously have described gliogenesis in response to ECS. Loss of glial cells is seen in depression and de novo formation of glial cells may thus have an important therapeutic role. Glial cell proliferation and activation is however also seen in response to neuronal damage. The aim of the present study was to further characterize glial cell activation in response to ECS. Two groups of rats were treated with 10 ECS using different sets of stimulus parameters. ECS-induced changes in the morphology and expression of markers typical for reactive microglia, astrocytes and NG2+ glial cells were analyzed immunohistochemically in prefrontal cortex, hippocampus, amygdala, hypothalamus, piriform cortex and entorhinal cortex. We observed changes in glial cell morphology and an enhanced expression of activation markers 2 h following ECS treatment, regardless of the stimulus parameters used. Four weeks later, few activated glial cells persisted. In conclusion, ECS treatment induced transient glial cell activation in several brain areas. Whether similar processes play a role in the therapeutic effect of clinically administered ECT or contribute to its side effects will require further investigations.