Intracerebral overexpression of miR-669c is protective in mouse ischemic stroke model by targeting MyD88 and inducing alternative microglial/macrophage activation

Background Ischemic stroke is a devastating disease without a cure. The available treatments for ischemic stroke, thrombolysis by tissue plasminogen activator, and thrombectomy are suitable only to a fraction of patients and thus novel therapeutic approaches are urgently needed. The neuroinflammatory responses elicited secondary to the ischemic attack further aggravate the stroke-induced neuronal damage. It has been demonstrated that these responses are regulated at the level of non-coding RNAs, especially miRNAs. Methods We utilized lentiviral vectors to overexpress miR-669c in BV2 microglial cells in order to modulate their polarization. To detect whether the modulation of microglial activation by miR-669c provides protection in a mouse model of transient focal ischemic stroke, miR-669c overexpression was driven by a lentiviral vector injected into the striatum prior to induction of ischemic stroke. Results Here, we demonstrate that miR-669c-3p, a member of chromosome 2 miRNA cluster (C2MC), is induced upon hypoxic and excitotoxic conditions in vitro and in two different in vivo models of stroke. Rather than directly regulating the neuronal survival in vitro, miR-669c is capable of attenuating the microglial proinflammatory activation in vitro and inducing the expression of microglial alternative activation markers arginase 1 (Arg1), chitinase-like 3 (Ym1), and peroxisome proliferator-activated receptor gamma (PPAR-gamma). Intracerebral overexpression of miR-669c significantly decreased the ischemia-induced cell death and ameliorated the stroke-induced neurological deficits both at 1 and 3 days post injury (dpi). Albeit miR-669c overexpression failed to alter the overall Iba1 protein immunoreactivity, it significantly elevated Arg1 levels in the ischemic brain and increased colocalization of Arg1 and Iba1. Moreover, miR-669c overexpression under cerebral ischemia influenced several morphological characteristics of Iba1 positive cells. We further demonstrate the myeloid differentiation primary response gene 88 (MyD88) transcript as a direct target for miR-669c-3p in vitro and show reduced levels of MyD88 in miR-669c overexpressing ischemic brains in vivo. Conclusions Collectively, our data provide the evidence that miR-669c-3p is protective in a mouse model of ischemic stroke through enhancement of the alternative microglial/macrophage activation and inhibition of MyD88 signaling. Our results accentuate the importance of controlling miRNA-regulated responses for the therapeutic benefit in conditions of stroke and neuroinflammation.Peer reviewe

Intracerebral overexpression of miR-669c is protective in mouse ischemic stroke model by targeting MyD88 and inducing alternative microglial/macrophage activation

Background Ischemic stroke is a devastating disease without a cure. The available treatments for ischemic stroke, thrombolysis by tissue plasminogen activator, and thrombectomy are suitable only to a fraction of patients and thus novel therapeutic approaches are urgently needed. The neuroinflammatory responses elicited secondary to the ischemic attack further aggravate the stroke-induced neuronal damage. It has been demonstrated that these responses are regulated at the level of non-coding RNAs, especially miRNAs. Methods We utilized lentiviral vectors to overexpress miR-669c in BV2 microglial cells in order to modulate their polarization. To detect whether the modulation of microglial activation by miR-669c provides protection in a mouse model of transient focal ischemic stroke, miR-669c overexpression was driven by a lentiviral vector injected into the striatum prior to induction of ischemic stroke. Results Here, we demonstrate that miR-669c-3p, a member of chromosome 2 miRNA cluster (C2MC), is induced upon hypoxic and excitotoxic conditions in vitro and in two different in vivo models of stroke. Rather than directly regulating the neuronal survival in vitro, miR-669c is capable of attenuating the microglial proinflammatory activation in vitro and inducing the expression of microglial alternative activation markers arginase 1 (Arg1), chitinase-like 3 (Ym1), and peroxisome proliferator-activated receptor gamma (PPAR-gamma). Intracerebral overexpression of miR-669c significantly decreased the ischemia-induced cell death and ameliorated the stroke-induced neurological deficits both at 1 and 3 days post injury (dpi). Albeit miR-669c overexpression failed to alter the overall Iba1 protein immunoreactivity, it significantly elevated Arg1 levels in the ischemic brain and increased colocalization of Arg1 and Iba1. Moreover, miR-669c overexpression under cerebral ischemia influenced several morphological characteristics of Iba1 positive cells. We further demonstrate the myeloid differentiation primary response gene 88 (MyD88) transcript as a direct target for miR-669c-3p in vitro and show reduced levels of MyD88 in miR-669c overexpressing ischemic brains in vivo. Conclusions Collectively, our data provide the evidence that miR-669c-3p is protective in a mouse model of ischemic stroke through enhancement of the alternative microglial/macrophage activation and inhibition of MyD88 signaling. Our results accentuate the importance of controlling miRNA-regulated responses for the therapeutic benefit in conditions of stroke and neuroinflammation.Peer reviewe

Endothelial LRP1 protects against neurodegeneration by blocking cyclophilin A

The low-density lipoprotein receptor-related protein 1 (LRP1) is an endocytic and cell signaling transmembrane protein. Endothelial LRP1 clears proteinaceous toxins at the blood-brain barrier (BBB), regulates angiogenesis, and is increasingly reduced in Alzheimer\u27s disease associated with BBB breakdown and neurodegeneration. Whether loss of endothelial LRP1 plays a direct causative role in BBB breakdown and neurodegenerative changes remains elusive. Here, we show that LRP1 inactivation from the mouse endothelium results in progressive BBB breakdown, followed by neuron loss and cognitive deficits, which is reversible by endothelial-specific LRP1 gene therapy. LRP1 endothelial knockout led to a self-autonomous activation of the cyclophilin A-matrix metalloproteinase-9 pathway in the endothelium, causing loss of tight junctions underlying structural BBB impairment. Cyclophilin A inhibition in mice with endothelial-specific LRP1 knockout restored BBB integrity and reversed and prevented neuronal loss and behavioral deficits. Thus, endothelial LRP1 protects against neurodegeneration by inhibiting cyclophilin A, which has implications for the pathophysiology and treatment of neurodegeneration linked to vascular dysfunction

Air Pollution Particulate Matter Amplifies White Matter Vascular Pathology and Demyelination Caused by Hypoperfusion

Cerebrovascular pathologies are commonly associated with dementia. Because air pollution increases arterial disease in humans and rodent models, we hypothesized that air pollution would also contribute to brain vascular dysfunction. We examined the effects of exposing mice to nanoparticulate matter (nPM; aerodynamic diameter ≤200 nm) from urban traffic and interactions with cerebral hypoperfusion. C57BL/6 mice were exposed to filtered air or nPM with and without bilateral carotid artery stenosis (BCAS) and analyzed by multiparametric MRI and histochemistry. Exposure to nPM alone did not alter regional cerebral blood flow (CBF) or blood brain barrier (BBB) integrity. However, nPM worsened the white matter hypoperfusion (decreased CBF on DSC-MRI) and exacerbated the BBB permeability (extravascular IgG deposits) resulting from BCAS. White matter MRI diffusion metrics were abnormal in mice subjected to cerebral hypoperfusion and worsened by combined nPM+BCAS. Axonal density was reduced equally in the BCAS cohorts regardless of nPM status, whereas nPM exposure caused demyelination in the white matter with or without cerebral hypoperfusion. In summary, air pollution nPM exacerbates cerebrovascular pathology and demyelination in the setting of cerebral hypoperfusion, suggesting that air pollution exposure can augment underlying cerebrovascular contributions to cognitive loss and dementia in susceptible elderly populations

An arylthiazyne derivative is a potent inhibitor of lipid peroxidation and ferroptosis providing neuroprotection in vitro and in vivo

Lipid peroxidation-initiated ferroptosis is an iron-dependent mechanism of programmed cell death taking place in neurological diseases. Here we show that a condensed benzo[b]thiazine derivative small molecule with an arylthiazine backbone (ADA-409-052) inhibits tert-Butyl hydroperoxide (TBHP)-induced lipid peroxidation (LP) and protects against ferroptotic cell death triggered by glutathione (GSH) depletion or glutathione peroxidase 4 (GPx4) inhibition in neuronal cell lines. In addition, ADA-409-052 suppresses pro-inflammatory activation of BV2 microglia and protects N2a neuronal cells from cell death induced by pro-inflammatory RAW 264.7 macrophages. Moreover, ADA-409-052 efficiently reduces infarct volume, edema and expression of pro-inflammatory genes in a mouse model of thromboembolic stroke. Targeting ferroptosis may be a promising therapeutic strategy in neurological diseases involving severe neuronal death and neuroinflammation.Peer reviewe

Cu-II(atsm) Attenuates Neuroinflammation

Background: Neuroinflammation and biometal dyshomeostasis are key pathological features of several neurodegenerative diseases, including Alzheimer's disease (AD). Inflammation and biometals are linked at the molecular level through regulation of metal buffering proteins such as the metallothioneins. Even though the molecular connections between metals and inflammation have been demonstrated, little information exists on the effect of copper modulation on brain inflammation. Methods: We demonstrate the immunomodulatory potential of the copper bis(thiosemicarbazone) complex Cu-II(atsm) in an neuroinflammatory model in vivo and describe its anti-inflammatory effects on microglia and astrocytes in vitro. Results: By using a sophisticated in vivo magnetic resonance imaging (MRI) approach, we report the efficacy of Cu-II(atsm) in reducing acute cerebrovascular inflammation caused by peripheral administration of bacterial lipopolysaccharide (LPS). Cu-II(atsm) also induced anti-inflammatory outcomes in primary microglia [significant reductions in nitric oxide (NO), monocyte chemoattractant protein 1 (MCP-1), and tumor necrosis factor (TNF)] and astrocytes [significantly reduced NO, MCP-1, and interleukin 6 (IL-6)] in vitro. These anti-inflammatory actions were associated with increased cellular copper levels and increased the neuroprotective protein metallothionein-1 (MT1) in microglia and astrocytes. Conclusion: The beneficial effects of Cu-II(atsm) on the neuroimmune system suggest copper complexes are potential therapeutics for the treatment of neuroinflammatory conditions.Peer reviewe

PPARβ/δ-agonist GW0742 ameliorates dysfunction in fatty acid oxidation in PSEN1ΔE9 astrocytes

Astrocytes are the gatekeepers of neuronal energy supply. In neurodegenerative diseases, bio-energetics demand increases and becomes reliant upon fatty acid oxidation as a source of energy. Defective fatty acid oxidation and mitochondrial dysfunctions correlate with hippocampal neurodegeneration and memory deficits in Alzheimer’s disease (AD), but it is unclear whether energy metabolism can be targeted to prevent or treat the disease. Here we show for the first time an impairment in fatty acid oxidation in human astrocytes derived from induced pluripotent stem cells of AD patients. The impairment was corrected by treatment with a synthetic peroxisome proliferator activated receptor delta (PPARβ/δ) agonist GW0742 which acts to regulate an array of genes governing cellular metabolism. GW0742 enhanced the expression of CPT1a, the gene encoding for a rate-limiting enzyme of fatty acid oxidation. Similarly, treatment of a mouse model of AD, the APP/PS1-mice, with GW0742 increased the expression of Cpt1a and concomitantly reversed memory deficits in a fear conditioning test. Although the GW0742-treated mice did not show altered astrocytic glial fibrillary acidic protein-immunoreactivity or reduction in amyloid beta (Aβ) load, GW0742 treatment increased hippo-campal neurogenesis and enhanced neuronal differentiation of neuronal progenitor cells. Furthermore, GW0742 prevented Aβ-induced impairment of long-term potentiation in hippocampal slices. Collectively, these data suggest that PPARβ/δ-agonism alleviates AD related deficits through increasing fatty acid oxidation in astrocytes and improves cognition in a transgenic mouse model of AD

Neuron-astrocyte transmitophagy is altered in Alzheimer's disease

Under physiological conditions in vivo astrocytes internalize and degrade neuronal mitochondria in a process called transmitophagy. Mitophagy is widely reported to be impaired in neurodegeneration but it is unknown whether and how transmitophagy is altered in Alzheimer's disease (AD). Here we report that the internalization of neuronal mitochondria is significantly increased in astrocytes isolated from AD mouse brains. We also demonstrate that the degradation of neuronal mitochondria by astrocytes is increased in AD mice at the age of 6 months onwards. Furthermore, we demonstrate for the first time a similar phenomenon between human neurons and AD astrocytes, and in murine hippocampi in vivo. The results suggest the involvement of S100a4 in impaired mitochondrial transfer between neurons and AD astrocytes together with significant increases in the mitophagy regulator and reactive oxygen species in aged AD astrocytes. These findings demonstrate altered neuronsupporting functions of AD astrocytes and provide a starting point for studying the molecular mechanisms of transmitophagy in AD.Peer reviewe

CuII(atsm) Attenuates Neuroinflammation

Background: Neuroinflammation and biometal dyshomeostasis are key pathological features of several neurodegenerative diseases, including Alzheimer’s disease (AD). Inflammation and biometals are linked at the molecular level through regulation of metal buffering proteins such as the metallothioneins. Even though the molecular connections between metals and inflammation have been demonstrated, little information exists on the effect of copper modulation on brain inflammation.Methods: We demonstrate the immunomodulatory potential of the copper bis(thiosemicarbazone) complex CuII(atsm) in an neuroinflammatory model in vivo and describe its anti-inflammatory effects on microglia and astrocytes in vitro.Results: By using a sophisticated in vivo magnetic resonance imaging (MRI) approach, we report the efficacy of CuII(atsm) in reducing acute cerebrovascular inflammation caused by peripheral administration of bacterial lipopolysaccharide (LPS). CuII(atsm) also induced anti-inflammatory outcomes in primary microglia [significant reductions in nitric oxide (NO), monocyte chemoattractant protein 1 (MCP-1), and tumor necrosis factor (TNF)] and astrocytes [significantly reduced NO, MCP-1, and interleukin 6 (IL-6)] in vitro. These anti-inflammatory actions were associated with increased cellular copper levels and increased the neuroprotective protein metallothionein-1 (MT1) in microglia and astrocytes.Conclusion: The beneficial effects of CuII(atsm) on the neuroimmune system suggest copper complexes are potential therapeutics for the treatment of neuroinflammatory conditions

Metsätuhot vuonna 2022

Luonnonvarakeskus tuottaa vuosittaista Metsätuhot - raporttisarjaa. Raporteissa käydään läpi mitä metsissä, taimitarhoilla sekä puisto- että pihapuilla menneenä vuonna tapahtui eri tuhonaiheuttajien osalta. Lisäksi raportissa käydään läpi muitakin metsien mielenkiintoisiä ilmiöitä, vaikkeivat ne varsinaisia tuhoja olisivatkaan olleet. Vuosi 2022 oli tältä osin mielenkiintoinen, sillä helteisen kesän 2021 jäljet näkyivät osin vasta tällöin. Vuonna 2022 kirjanpainajan (Ips typographus) takia hakattiin metsiä metsäkeskuksen keräämien ilmoitusten perusteella enemmän kuin kertaakaan tilastointihistoriassa (2 834 ha). Laji myös parveili aiempaa runsaampana Luken feromoniseurannan pohjoisimmilla alueilla. Kirjanpainajan lisäksi myös toinen havupuiden kaarnakuoriainen, kuusentähtikirjaaja, aiheutti näkyviä latvakuolemia kuuselle eri puolilla Suomea. Molempien ilmiöiden takana vaikutti paitsi kuoriaisille kohtuullinen kesä 2022, niin ennen kaikkea helteinen ja lämmin kesä 2021. Ilmiö konkretisoikin omalla tavallaan ilmaston lämpenemisen mahdollisia vaikutuksia metsissämme. Helteiden tuhoja lisäävät vaikutukset eivät kuitenkaan näkyneet valtakunnan metsien inventoinnissa (VMI) samalla tavalla. VMI:ssä havaitut kirjanpainajatuhot jopa laskivat (-14 %) hieman vuoden takaisesta. Tässä on kuitenkin muistettava, että VMI otanta ei ole ajallisesti eikä spatiaaliselta tarkkuudeltaan soveltuva useimpien hyönteis- ja sienituhojen seurantaan. Merkittävimpinä tuhonaiheuttajina VMI:ssä olivat aiempien vuosien tapaan lumi, tuuli ja hirvieläimet – eli tuhot, joita voidaan yleisesti havaita maastossa inventointiajankohdasta riippumatta. Lämpenevään ilmastoon reagoivat hyönteisten lisäksi useat muutkin kotoperäiset taudinaiheuttajat. Ne hyötyvät ilmastonmuutoksesta mm. laajentamalla esiintymisaluettaan sekä aiheuttamalla puissa enemmän vaurioita, etenkin kun säät muuttuvat suopeimmaksi niille, mutta huonommiksi niiden isäntäpuille. Kuusenjuurikääpää eteläisempi männynjuurikääpä (Heterobasidion annosum s.s.) raportoitiinkin vuonna 2022 Pohjois-Pohjanmaalta saakka. Juurikääpätuhot myös lisääntyivät noin 30 % vuonna 2022 edellisvuoteen verrattuna. Lämpimät ja kuivat kesät ovat myös edesauttaneet tulokaslaji havuparikkaan (Diplodia sapinea) aiheuttaman taudin etelänversosurman puhkeamista. Havuparikkaan levinneisyysaluetta kartoitettiin vuonna 2022 kansalaistieteen avulla, ja sienen esiintyminen olikin ennakoitua laajempi. Myös tulokaslaji havununna (Lymantria monacha) tavattiin hieman aiempaa runsaampana lajin pohjoisemmilla esiintymisalueilla. Vuonna 2022 päätään nostivat valitettavasti myös ilmastosta riippumattomat uhkatekijät: kansainvälisen kasvikaupan ja turismin mukana leviävät haitalliset vieraslajit. Uusia vierasperäisiä tuhonaiheuttajia löydettiin niin taimitarhoilta, kuin puistopuistakin. Koivuilla todettiin Discula betulina-sienen aiheuttamaa versolaikkua, pihdoilta taas löydettiin pihtanäppyä (Neonectria macrospora). Haitallisten vieraslajien leviäminen on kansainvälinen ongelma, jolta pohjoinen Suomikaan ei ole turvassa. Luken tutkijat julkaisivat vuonna 2022 suomenkielisen artikkelisarjan mikä sisälsi tietoa useiden eri vieraslajien uhasta. Katsaus muistuttaa, ettei metsiemme valtapuilla useinkaan ole vastustuskykyä näitä uusia tuholaisia vastaan, mikä tekee niistä merkittävän uhan metsien terveydelle. Haitallisten vieraslajien leviämisen estäminen on siis erityisen tärkeää niin nyt kuin tulevaisuudessakin. Tässä kansainvälinen kasvikauppa on pahin uhkatekijä. Metsien ja puutarhojen osalta ulkomaisten taimien ja koristekasvien tuonti muodostaa tehokkaan haitallisten vieraslajien leviämisväylän. Kotimainen taimituotanto taasen tuottaa paitsi terveitä, niin myös alkuperältään tunnettuja ja laadultaan varmistettuja taimia. Näitä taimia jokaisen tulisi suosia niin metsänuudistamisessa kuin kotipihoilla, sillä olemassa oleva lainsäädäntö ei kansainvälistä kasvikauppaa hillitse. Luke kerää ja koosta myös kansalaisten tekemiä ilmoituksia havaituista puustotuhoista tai muista metsien ilmiöstä. Vuonna 2022 okakaarnakuoriaisten (Ips acuminatus) takia tehdyt tuhoilmoitukset jatkoivat kasvuaan vuodelta 2021. Ilmoituksissa ja niiden perusteella tehdyissä maastotarkastuksissa mäntyjen todettiin kuolevan etenkin lounaisrannikolla, missä okakaarnakuoriaiset tuntuivat viihtyvän hyvin yhdessä havuparikkaan kanssa. Selvitys näiden kahden tuholaisen mahdollisesta yhteisvaikutuksesta äkillisesti kuivuneiden mäntyjen kuolemiin jatkuu Lukessa. Tämän vuoden raporttiin omaksi tarinakseen nostettiin myös ilmoituksia kerännyt lehtikuoriaisten runsas esiintyminen, missä etenkin lepillä esiintyvä idänlehtikuoriainen (Agelastica alni) oli näkyvässä roolissa. Vaikka vuoden 2022 perusteella tilanne Suomen metsissä on tuhojen suhteen pääosin hyvä, vuosi muistutti meitä konkreettisesti sekä muuttuvan ilmaston että kansainvälisen kasvikaupan meille tuomista uhkista. Sekä vieras- että tulokaslajien aiheuttamat riskit tulevat kasvamaan lämpenevässä ilmastossa, sillä sen ansiosta nämä lajit pystyvät todennäköisemmin vakiinnuttamaan asemansa uudella alueellaan. Meillä jo ongelmia aiheuttavien lajien kuten juurikäävän tai kirjanpainajan osalta lämpenevä ilmasto ei niin ikään lupaa missään määrin hyvää metsillemme. Ollaksemme varautuneita mahdolliseen tuhojen äkilliseen lisääntymiseen tarvitsemme paitsi aktiivista seurantaa, niin myös perus- ja soveltavaa tutkimusta. sekä tarkempaa analyysiä siitä miten eri tuhonaiheuttajien uhat voivat meillä tulevaisuudessa realisoitua