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1.
Int J Mol Sci ; 22(12)2021 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-34208173

RESUMO

Maternal malnutrition in critical periods of development increases the risk of developing short- and long-term diseases in the offspring. The alterations induced by this nutritional programming in the hypothalamus of the offspring are of special relevance due to its role in energy homeostasis, especially in the endocannabinoid system (ECS), which is involved in metabolic functions. Since astrocytes are essential for neuronal energy efficiency and are implicated in brain endocannabinoid signaling, here we have used a rat model to investigate whether a moderate caloric restriction (R) spanning from two weeks prior to the start of gestation to its end induced changes in offspring hypothalamic (a) ECS, (b) lipid metabolism (LM) and/or (c) hypothalamic astrocytes. Monitorization was performed by analyzing both the gene and protein expression of proteins involved in LM and ECS signaling. Offspring born from caloric-restricted mothers presented hypothalamic alterations in both the main enzymes involved in LM and endocannabinoids synthesis/degradation. Furthermore, most of these changes were similar to those observed in hypothalamic offspring astrocytes in culture. In conclusion, a maternal low caloric intake altered LM and ECS in both the hypothalamus and its astrocytes, pointing to these glial cells as responsible for a large part of the alterations seen in the total hypothalamus and suggesting a high degree of involvement of astrocytes in nutritional programming.


Assuntos
Astrócitos/metabolismo , Restrição Calórica , Endocanabinoides/metabolismo , Hipotálamo/metabolismo , Metabolismo dos Lipídeos , Transdução de Sinais , Animais , Animais Recém-Nascidos , Peso Corporal , Encéfalo/patologia , Feminino , Regulação da Expressão Gênica , Gliose/genética , Gliose/patologia , Inflamação/genética , Inflamação/patologia , Metabolismo dos Lipídeos/genética , Gravidez , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Transdução de Sinais/genética
2.
Int J Mol Sci ; 22(13)2021 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-34206635

RESUMO

White matter tract (WMT) degeneration has been reported to occur following a stroke, and it is associated with post-stroke functional disturbances. White matter pathology has been suggested to be an independent predictor of post-stroke recovery. However, the factors that influence WMT remodeling are poorly understood. Cortisol is a steroid hormone released in response to prolonged stress, and elevated levels of cortisol have been reported to interfere with brain recovery. The objective of this study was to investigate the influence of corticosterone (CORT; the rodent equivalent of cortisol) on WMT structure post-stroke. Photothrombotic stroke (or sham surgery) was induced in 8-week-old male C57BL/6 mice. At 72 h, mice were exposed to standard drinking water ± CORT (100 µg/mL). After two weeks of CORT administration, mice were euthanised and brain tissue collected for histological and biochemical analysis of WMT (particularly the corpus callosum and corticospinal tract). CORT administration was associated with increased tissue loss within the ipsilateral hemisphere, and modest and inconsistent WMT reorganization. Further, a structural and molecular analysis of the WMT components suggested that CORT exerted effects over axons and glial cells. Our findings highlight that CORT at stress-like levels can moderately influence the reorganization and microstructure of WMT post-stroke.


Assuntos
Corticosterona/administração & dosagem , Gliose/metabolismo , Gliose/patologia , Vias Neurais/efeitos dos fármacos , Acidente Vascular Cerebral/metabolismo , Substância Branca/efeitos dos fármacos , Substância Branca/fisiologia , Animais , Axônios/metabolismo , Corpo Caloso/efeitos dos fármacos , Corpo Caloso/metabolismo , Corpo Caloso/patologia , Modelos Animais de Doenças , Progressão da Doença , Suscetibilidade a Doenças , Gliose/tratamento farmacológico , Gliose/etiologia , Imuno-Histoquímica , Masculino , Camundongos , Bainha de Mielina/efeitos dos fármacos , Bainha de Mielina/metabolismo , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/metabolismo , Estresse Fisiológico/efeitos dos fármacos , Acidente Vascular Cerebral/tratamento farmacológico , Acidente Vascular Cerebral/etiologia , Acidente Vascular Cerebral/patologia
3.
Int J Mol Sci ; 22(9)2021 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-34063002

RESUMO

Cone Dystrophy with Supernormal Rod Response (CDSRR) is a rare autosomal recessive disorder leading to severe visual impairment in humans, but little is known about its unique pathophysiology. We have previously shown that CDSRR is caused by mutations in the KCNV2 (Potassium Voltage-Gated Channel Modifier Subfamily V Member 2) gene encoding the Kv8.2 subunit, a modulatory subunit of voltage-gated potassium (Kv) channels. In a recent study, we validated a novel mouse model of Kv8.2 deficiency at a late stage of the disease and showed that it replicates the human electroretinogram (ERG) phenotype. In this current study, we focused our investigation on young adult retinas to look for early markers of disease and evaluate their effect on retinal morphology, electrophysiology and immune response in both the Kv8.2 knockout (KO) mouse and in the Kv2.1 KO mouse, the obligate partner of Kv8.2 in functional retinal Kv channels. By evaluating the severity of retinal dystrophy in these KO models, we demonstrated that retinas of Kv KO mice have significantly higher apoptotic cells, a thinner outer nuclear cell layer and increased activated microglia cells in the subretinal space. Our results indicate that in the murine retina, the loss of Kv8.2 subunits contributes to early cellular and physiological changes leading to retinal dysfunction. These results could have potential implications in the early management of CDSRR despite its relatively nonprogressive nature in humans.


Assuntos
Envelhecimento/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Subunidades Proteicas/metabolismo , Retina/citologia , Retina/metabolismo , Canais de Potássio Shab/metabolismo , Animais , Morte Celular , Eletrorretinografia , Gliose/patologia , Imunidade , Camundongos Knockout , Microglia/patologia , Visão Noturna , Retina/fisiologia
4.
Res Vet Sci ; 138: 39-48, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34091228

RESUMO

Methotrexate (MTX), an antifolate drug, is widely used in chemotherapeutic protocols for metastatic and primary brain tumors and some autoimmune diseases. Its efficacy for brain tumors is limited by the high incidence of central nervous system (CNS) complications. This investigation aimed to observe the morphological effects, including astroglial and microglial responses, following systemic short-term MTX administration in adult rats. Male Wistar rats received 5 or 10 mg/kg/day of MTX by intraperitoneal route for 4 consecutive days (respectively, MTX5 and MTX10 groups) or the same volume of 0.9% saline solution (control group). On the 5th day, brain samples were collected for hematoxylin-eosin and luxol fast blue staining techniques, as well as for immunohistochemical staining for glial fibrillary acidic protein (GFAP) expression in astrocytes and Iba1 (ionized calcium binding adaptor molecule 1) for microglia in the frontal cortex, hippocampus, hypothalamus and molecular/granular layers of the cerebellum. Morphometric analyses were performed using Image Pro-Plus software. Brain levels of the proinflammatory cytokines TNF-α and IL-1ß were determined by ELISA. No signs of neuronal loss or demyelination were observed in all groups. Increased GFAP and Iba1 expression was found in all areas from the MTX groups, although it was slightly higher in the MTX10 group compared to the MTX5. Both TNF-α and IL-1ß levels were decreased in the MTX5 group compared to controls. In the MTX10 group, TNF-α decreased, although IL-1ß was increased relative to controls. MTX administration induced microglial reaction and astrogliosis in several CNS areas. In the MTX5 group, it apparently occurred in the presence of decreased proinflammatory cytokines.


Assuntos
Antimetabólitos Antineoplásicos/administração & dosagem , Astrócitos/efeitos dos fármacos , Proteínas de Ligação ao Cálcio/metabolismo , Proteína Glial Fibrilar Ácida/metabolismo , Gliose/fisiopatologia , Metotrexato/administração & dosagem , Proteínas dos Microfilamentos/metabolismo , Microglia/efeitos dos fármacos , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Relação Dose-Resposta a Droga , Gliose/induzido quimicamente , Gliose/patologia , Masculino , Microglia/metabolismo , Microglia/patologia , Ratos , Ratos Wistar
5.
Int J Mol Sci ; 22(9)2021 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-33922372

RESUMO

Neuropathic pain (NeP) in humans is often a life-long condition with no effective therapy available. The higher incidence of female gender in NeP onset is worldwide reported, and although the cause is generally attributed to sex hormones, the actual mechanisms and the players involved are still unclear. Glial and immune cells take part in NeP development, and orchestrate the neuroimmune and inflammatory response, releasing pro-inflammatory factors with chemoattractant properties that activate resident immune cells and recruit immune cells from circulation. The neuro-immune crosstalk is a key contributor to pain hypersensitivity following peripheral nervous system injury. Our previous works showed that in spite of the fact that female mice had an earlier analgesic response than males following nerve lesion, the recovery from NeP was never complete, suggesting that this difference could occur in the very early stages after injury. To further investigate gender differences in immune and neuroimmune responses to NeP, we studied the main immune cells and mediators elicited both in plasma and sciatic nerves by peripheral nerve lesion. After injury, we found a different pattern of distribution of immune cell populations showing either a higher infiltration of T cells in nerves from females or a higher infiltration of macrophages in nerves from males. Moreover, in comparison to male mice, the levels of cytokines and chemokines were differently up- and down-regulated in blood and nerve lysates from female mice. Our study provides some novel insights for the understanding of gender-associated differences in the generation and perseveration of NeP as well as for the isolation of specific neurodegenerative mechanisms underlying NeP. The identification of gender-associated inflammatory profiles in neuropathy is of key importance for the development of differential biomarkers and gender-specific personalized medicine.


Assuntos
Gliose/patologia , Hiperalgesia/patologia , Inflamação/patologia , Macrófagos/patologia , Neuralgia/patologia , Traumatismos dos Nervos Periféricos/complicações , Nervo Isquiático/patologia , Animais , Citocinas , Feminino , Gliose/etiologia , Hiperalgesia/etiologia , Inflamação/etiologia , Masculino , Camundongos , Neuralgia/etiologia , Fatores Sexuais
6.
Int J Mol Sci ; 22(8)2021 Apr 14.
Artigo em Inglês | MEDLINE | ID: mdl-33920048

RESUMO

Astrogliosis has been abundantly studied in rodents but relatively poorly in human cells due to limited access to the brain. Astrocytes play important roles in cerebral energy metabolism, and are also key players in neuroinflammation. Astroglial metabolic and inflammatory changes as a function of age have been reported, leading to the hypothesis that mitochondrial metabolism and inflammatory responses are interconnected in supporting a functional switch of astrocytes from neurotrophic to neurotoxic. This study aimed to explore the metabolic changes occurring in astrocytes during their activation. Astrocytes were derived from human ReN cell neural progenitors and characterized. They were activated by exposure to tumor necrosis factor alpha (TNFα) or interleukin 1ß (IL1ß) for 24 h. Astrocyte reaction and associated energy metabolic changes were assessed by immunostaining, gene expression, proteomics, metabolomics and extracellular flux analyses. ReN-derived astrocytes reactivity was observed by the modifications of genes and proteins linked to inflammation (cytokines, nuclear factor-kappa B (NFκB), signal transducers and activators of transcription (STATs)) and immune pathways (major histocompatibility complex (MHC) class I). Increased NFκB1, NFκB2 and STAT1 expression, together with decreased STAT3 expression, suggest an activation towards the detrimental pathway. Strong modifications of astrocyte cytoskeleton were observed, including a glial fibrillary acidic protein (GFAP) decrease. Astrogliosis was accompanied by changes in energy metabolism characterized by increased glycolysis and lactate release. Increased glycolysis is reported for the first time during human astrocyte activation. Astrocyte activation is strongly tied to energy metabolism, and a possible association between NFκB signaling and/or MHC class I pathway and glycolysis is suggested.


Assuntos
Astrócitos/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Interleucina-1beta/farmacologia , Fator de Necrose Tumoral alfa/farmacologia , Astrócitos/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Linhagem Celular , Metabolismo Energético/efeitos dos fármacos , Gliose/tratamento farmacológico , Gliose/genética , Gliose/patologia , Glicólise/genética , Humanos , Inflamação/genética , Inflamação/patologia , Interleucina-1beta/genética , Neurogênese/efeitos dos fármacos , Fator de Transcrição STAT3/genética , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/genética
7.
Molecules ; 26(5)2021 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-33806413

RESUMO

MsrB1 used to be named selenoprotein R, for it was first identified as a selenocysteine containing protein by searching for the selenocysteine insert sequence (SECIS) in the human genome. Later, it was found that MsrB1 is homologous to PilB in Neisseria gonorrhoeae, which is a methionine sulfoxide reductase (Msr), specifically reducing L-methionine sulfoxide (L-Met-O) in proteins. In humans and mice, four members constitute the Msr family, which are MsrA, MsrB1, MsrB2, and MsrB3. MsrA can reduce free or protein-containing L-Met-O (S), whereas MsrBs can only function on the L-Met-O (R) epimer in proteins. Though there are isomerases existent that could transfer L-Met-O (S) to L-Met-O (R) and vice-versa, the loss of Msr individually results in different phenotypes in mice models. These observations indicate that the function of one Msr cannot be totally complemented by another. Among the mammalian Msrs, MsrB1 is the only selenocysteine-containing protein, and we recently found that loss of MsrB1 perturbs the synaptic plasticity in mice, along with the astrogliosis in their brains. In this review, we summarized the effects resulting from Msr deficiency and the bioactivity of selenium in the central nervous system, especially those that we learned from the MsrB1 knockout mouse model. We hope it will be helpful in better understanding how the trace element selenium participates in the reduction of L-Met-O and becomes involved in neurobiology.


Assuntos
Sistema Nervoso Central/patologia , Gliose/patologia , Metionina Sulfóxido Redutases/fisiologia , Plasticidade Neuronal , Selênio/metabolismo , Animais , Sistema Nervoso Central/metabolismo , Gliose/etiologia , Gliose/metabolismo , Humanos , Camundongos , Camundongos Knockout
8.
Nat Commun ; 12(1): 1490, 2021 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-33674568

RESUMO

The brain of mammals lacks a significant ability to regenerate neurons and is thus particularly vulnerable. To protect the brain from injury and disease, damage control by astrocytes through astrogliosis and scar formation is vital. Here, we show that brain injury in mice triggers an immediate upregulation of the actin-binding protein Drebrin (DBN) in astrocytes, which is essential for scar formation and maintenance of astrocyte reactivity. In turn, DBN loss leads to defective astrocyte scar formation and excessive neurodegeneration following brain injuries. At the cellular level, we show that DBN switches actin homeostasis from ARP2/3-dependent arrays to microtubule-compatible scaffolds, facilitating the formation of RAB8-positive membrane tubules. This injury-specific RAB8 membrane compartment serves as hub for the trafficking of surface proteins involved in astrogliosis and adhesion mediators, such as ß1-integrin. Our work shows that DBN-mediated membrane trafficking in astrocytes is an important neuroprotective mechanism following traumatic brain injury in mice.


Assuntos
Astrócitos/metabolismo , Lesões Encefálicas Traumáticas/metabolismo , Cicatriz/metabolismo , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Complexo 2-3 de Proteínas Relacionadas à Actina , Actinas/metabolismo , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Lesões Encefálicas/metabolismo , Lesões Encefálicas/patologia , Lesões Encefálicas Traumáticas/patologia , Movimento Celular , Sistema Nervoso Central/metabolismo , Modelos Animais de Doenças , Gliose/metabolismo , Gliose/patologia , Camundongos , Camundongos Knockout , Neuroproteção , Transcriptoma , Proteínas rab de Ligação ao GTP/metabolismo
9.
Invest Ophthalmol Vis Sci ; 62(2): 3, 2021 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-33523201

RESUMO

Purpose: This study aimed to determine the effect of pinacidil, a nonselective KATP channel opener, on diabetes-induced retinal gliosis and inflammation. Methods: Primary and immortalized cell lines of retinal microglia and Müller cells were used to set up a coculture model. In the trans-well system, microglia were seeded in the upper chamber and Müller cells in the bottom chamber. Microglia were polarized into proinflammatory (M1, with lipopolysaccharide and INF-γ) with or without different pinacidil concentrations before coculturing with Müller cells. The expression of inflammatory or anti-inflammatory genes and protein in microglia, and the expression of glial fibrillary acidic protein (GFAP), Kir4.1, and AQP4 in Müller cells were examined by real-time polymerase chain reaction and Western blot. Pinacidil was injected intravitreally into streptozotocin-induced diabetic rats. Retinal gliosis and inflammation were examined by immunohistochemistry and Western blot. Results: Intravitreal injection of pinacidil alleviated diabetes-induced Müller cell gliosis and microglial activation and reduced vascular endothelial growth factor expression. In vitro study demonstrated that pinacidil inhibited tumor necrosis factor and interleukin-1ß expression in M1-type microglia and alleviated the M1 microglia-induced GFAP expression in the Müller cells. Furthermore, we found that pinacidil on its own, or in combination with IL-4, can upregulate arginase-1 (Arg-1) and Kir6.1 expression in microglial cells. Conclusions: Our results suggest that potassium channels are critically involved in diabetes-induced gliosis and microglial activation. The KATP opener, pinacidil, can reduce microglial activation by upregulating Kir6.1 expression.


Assuntos
Diabetes Mellitus Experimental/metabolismo , Regulação da Expressão Gênica , Gliose/metabolismo , Inflamação/metabolismo , Canais KATP/genética , Microglia/metabolismo , Pinacidil/farmacologia , Animais , Células Cultivadas , DNA/genética , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/genética , Gliose/tratamento farmacológico , Gliose/patologia , Imuno-Histoquímica , Inflamação/tratamento farmacológico , Inflamação/genética , Canais KATP/biossíntese , Masculino , Moduladores de Transporte de Membrana/farmacologia , Microglia/patologia , Ratos , Ratos Sprague-Dawley
10.
Theranostics ; 11(3): 1232-1248, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33391532

RESUMO

Rationale: Glial scars present a major obstacle for neuronal regeneration after stroke. Thus, approaches to promote their degradation and inhibit their formation are beneficial for stroke recovery. The interaction of microglia and astrocytes is known to be involved in glial scar formation after stroke; however, how microglia affect glial scar formation remains unclear. Methods: Mice were treated daily with M2 microglial small extracellular vesicles through tail intravenous injections from day 1 to day 7 after middle cerebral artery occlusion. Glial scar, infarct volume, neurological score were detected after ischemia. microRNA and related protein were examined in peri-infarct areas of the brain following ischemia. Results: M2 microglial small extracellular vesicles reduced glial scar formation and promoted recovery after stroke and were enriched in miR-124. Furthermore, M2 microglial small extracellular vesicle treatment decreased the expression of the astrocyte proliferation gene signal transducer and activator of transcription 3, one of the targets of miR-124, and glial fibrillary acidic protein and inhibited astrocyte proliferation both in vitro and in vivo. It also decreased Notch 1 expression and increased Sox2 expression in astrocytes, which suggested that astrocytes had transformed into neuronal progenitor cells. Finally, miR-124 knockdown in M2 microglial small extracellular vesicles blocked their effects on glial scars and stroke recovery. Conclusions: Our results showed, for the first time, that microglia regulate glial scar formation via small extracellular vesicles, indicating that M2 microglial small extracellular vesicles could represent a new therapeutic approach for stroke.


Assuntos
Isquemia Encefálica/metabolismo , Vesículas Extracelulares/metabolismo , Gliose/metabolismo , AVC Isquêmico/metabolismo , MicroRNAs/metabolismo , Microglia/metabolismo , Fator de Transcrição STAT3/metabolismo , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Encéfalo/metabolismo , Encéfalo/patologia , Isquemia Encefálica/patologia , Células Cultivadas , Modelos Animais de Doenças , Vesículas Extracelulares/patologia , Proteína Glial Fibrilar Ácida/metabolismo , Gliose/patologia , Infarto da Artéria Cerebral Média/metabolismo , Infarto da Artéria Cerebral Média/patologia , AVC Isquêmico/patologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Microglia/patologia , Neurônios/metabolismo , Neurônios/patologia
11.
Int J Mol Sci ; 22(3)2021 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-33503976

RESUMO

Müller cells, the major retinal macroglia, are key to maintaining vascular integrity as well as retinal fluid and ion homeostasis. Although platelet derived growth factor (PDGF) receptor expression in Müller glia has been reported earlier, their actual role for Müller cell function and intimate interaction with cells of the retinal neurovascular unit remains unclear. To close this gap of knowledge, Müller cell-specific PDGF receptor alpha (PDGFRα) knockout (KO) mice were generated, characterized, and subjected to a model of choroidal neovascularization (CNV). PDGFRα-deficient Müller cells could not counterbalance hypoosmotic stress as efficiently as their wildtype counterparts. In wildtypes, the PDGFRα ligand PDGF-BB prevented Müller cell swelling induced by the administration of barium ions. This effect could be blocked by the PDGFR family inhibitor AC710. PDGF-BB could not restore the capability of an efficient volume regulation in PDGFRα KO Müller cells. Additionally, PDGFRα KO mice displayed reduced rod and cone-driven light responses. Altogether, these findings suggest that Müller glial PDGFRα is central for retinal functions under physiological conditions. In contrast, Müller cell-specific PDGFRα KO resulted in less vascular leakage and smaller lesion areas in the CNV model. Of note, the effect size was comparable to pharmacological blockade of PDGF signaling alone or in combination with anti-vascular endothelial growth factor (VEGF) therapy-a treatment regimen currently being tested in clinical trials. These data imply that targeting PDGF to treat retinal neovascular diseases may have short-term beneficial effects, but may elicit unwarranted side effects given the putative negative effects on Müller cell homeostatic functions potentially interfering with a long-term positive outcome.


Assuntos
Células Ependimogliais/metabolismo , Homeostase , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Transdução de Sinais , Animais , Biomarcadores , Neovascularização de Coroide/etiologia , Neovascularização de Coroide/metabolismo , Neovascularização de Coroide/patologia , Modelos Animais de Doenças , Imunofluorescência , Expressão Gênica , Técnicas de Inativação de Genes , Gliose/etiologia , Gliose/metabolismo , Gliose/patologia , Camundongos , Camundongos Knockout , Neuroglia/metabolismo , Especificidade de Órgãos/genética , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Retina/metabolismo , Retina/patologia
12.
Lasers Med Sci ; 36(1): 91-98, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32297252

RESUMO

The integrity of the structural cerebral cortex is disrupted after stroke either at the macroscopic or microscopic levels. Therefore, many attempts have been gathered to circumvent stroke-associated problems in the brain tissue. The current study was aimed to design an animal model of photochemical stroke using rose bengal (RB) plus laser irradiation (L) after 10, 15, and 20 min (´) and evaluate its effect on the cerebral tissue using unbiased stereological quantitative methods and morphometric histological analysis. Photochemical stroke was induced by intraperitoneal injection of RB dye and further activation through the exposure of the right sensorimotor cortex with the green laser radiation (100 mW; 532 nm). Mice were randomly allocated into 4 groups (each in 15) as follows: control (10 µg/gbw RB), RB + 10'L, RB + 15'L, and RB + 20'L. Target irradiation site was adjusted to 2 mm lateral to the bregma. Vernier caliper morphometric evaluation, cresyl violet staining, and unbiased stereological assays including Cavalier's principle and point counting techniques were used to monitor the pathological changes and lesion volume on days 1, 3, and 7 after the ischemia induction. Our data showed that the mean diameter of the lesion site and lesion infarct volume in the group RB + 20'L) was significantly increased relative to the other groups (P < 0.05). Notably, the lesion volume and diameter in the group RB + 15'L was larger compared with the group RB + 10'L and control mice (P < 0.05). Data showed an increased acute inflammatory response such as hyperemia and edema 3 days after ischemic induction while the intensity of acute changes and lesion volume were reduced and replaced with necrotic and chronic pathological changes including astrogliosis on day 7. It is concluded that the laser irradiation of RB-injected mice at a distinct time period could induce the magnificent degenerative effects on the cerebral cortex which is similar to the stroke condition.


Assuntos
Processos Fotoquímicos , Córtex Sensório-Motor/lesões , Córtex Sensório-Motor/efeitos da radiação , Acidente Vascular Cerebral/patologia , Animais , Isquemia Encefálica/complicações , Isquemia Encefálica/patologia , Modelos Animais de Doenças , Gliose/complicações , Gliose/patologia , Masculino , Camundongos , Córtex Sensório-Motor/patologia , Acidente Vascular Cerebral/complicações
13.
Neuron ; 109(3): 438-447.e6, 2021 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-33321072

RESUMO

The ε4 allele of the apolipoprotein E gene (APOE4) is a strong genetic risk factor for Alzheimer's disease (AD) and multiple vascular conditions. ApoE is abundantly expressed in multiple brain cell types, including astrocytes, microglia, and vascular mural cells (VMCs). Here, we show that VMC-specific expression of apoE4 in mice impairs behavior and cerebrovascular function. Expression of either apoE3 or apoE4 in VMCs was sufficient to rescue the hypercholesterolemia and atherosclerosis phenotypes seen in Apoe knockout mice. Intriguingly, vascular expression of apoE4, but not apoE3, reduced arteriole blood flow, impaired spatial learning, and increased anxiety-like phenotypes. Single-cell RNA sequencing of vascular and glial cells revealed that apoE4 in VMCs was associated with astrocyte activation, while apoE3 was linked to angiogenic signature in pericytes. Together, our data support cell-autonomous effects of vascular apoE on brain homeostasis in an isoform-dependent manner, suggesting a critical contribution of vascular apoE to AD pathogenesis.


Assuntos
Apolipoproteína E4/genética , Arteríolas/metabolismo , Astrócitos/metabolismo , Encéfalo/metabolismo , Gliose/genética , Animais , Apolipoproteína E3/genética , Apolipoproteína E3/metabolismo , Apolipoproteína E4/metabolismo , Arteríolas/patologia , Astrócitos/patologia , Encéfalo/patologia , Gliose/metabolismo , Gliose/patologia , Camundongos , Camundongos Transgênicos
14.
J Vis Exp ; (165)2020 11 12.
Artigo em Inglês | MEDLINE | ID: mdl-33252109

RESUMO

Diverse vascular diseases such as diabetic retinopathy, occlusion of retinal veins or arteries and ocular ischemic syndrome can lead to retinal ischemia. To investigate pathological mechanisms of retinal ischemia, relevant experimental models need to be developed. Anatomically, a main retinal blood supplying vessel is the ophthalmic artery (OpA) and OpA originates from the internal carotid artery of the common carotid artery (CCA). Thus, disruption of CCA could effectively cause retinal ischemia. Here, we established a mouse model of retinal ischemia by transient bilateral common carotid artery occlusion (tBCCAO) to tie the right CCA with 6-0 silk sutures and to occlude the left CCA transiently for 2 seconds via a clamp, and showed that tBCCAO could induce acute retinal ischemia leading to retinal dysfunction. The current method reduces reliance on surgical instruments by only using surgical needles and a clamp, shortens occlusion time to minimize unexpected animal death, which is often seen in mouse models of middle cerebral artery occlusion, and maintains reproducibility of common retinal ischemic findings. The model can be utilized to investigate the pathophysiology of ischemic retinopathies in mice and further can be used for in vivo drug screening.


Assuntos
Arteriopatias Oclusivas/complicações , Artéria Carótida Primitiva/patologia , Isquemia/etiologia , Retina/lesões , Animais , Arteriopatias Oclusivas/diagnóstico por imagem , Arteriopatias Oclusivas/fisiopatologia , Círculo Arterial do Cérebro/patologia , Modelos Animais de Doenças , Eletrorretinografia , Gliose/complicações , Gliose/diagnóstico por imagem , Gliose/patologia , Gliose/fisiopatologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Isquemia/diagnóstico por imagem , Isquemia/patologia , Isquemia/fisiopatologia , Masculino , Camundongos , Perfusão , Estabilidade Proteica , Reprodutibilidade dos Testes , Retina/diagnóstico por imagem , Retina/patologia , Retina/fisiopatologia , Tomografia de Coerência Óptica
15.
J Neurosci ; 40(50): 9751-9771, 2020 12 09.
Artigo em Inglês | MEDLINE | ID: mdl-33158962

RESUMO

Expression of the 17ß-estradiol (E2) synthesis enzyme aromatase is highly upregulated in astrocytes following brain injury. However, the precise role of astrocyte-derived E2 in the injured brain remains unclear. In the current study, we generated a glial fibrillary acidic protein (GFAP) promoter-driven aromatase knock-out (GFAP-ARO-KO) mouse model to deplete astrocyte-derived E2 in the brain and determine its roles after global cerebral ischemia (GCI) in male and female mice. GFAP-ARO-KO mice were viable and fertile, with normal gross brain structure, normal morphology, intensity and distribution of astrocytes, normal aromatase expression in neurons, and normal cognitive function basally. In contrast, after GCI, GFAP-ARO-KO mice: (1) lacked the normal elevation of astrocyte aromatase and hippocampal E2 levels; (2) had significantly attenuated reactive astrogliosis; and (3) displayed enhanced neuronal damage, microglia activation, and cognitive deficits. RNA-sequencing (RNA-seq) analysis revealed that the ischemic GFAP-ARO-KO mouse hippocampus failed to upregulate the "A2" panel of reactive astrocyte genes. In addition, the JAK-STAT3 pathway, which is critical for the induction of reactive astrogliosis, was significantly downregulated in the GFAP-ARO-KO hippocampus following GCI. Finally, exogenous E2 administration fully rescued the compromised JAK-STAT3 pathway and reactive astrogliosis, and reversed the enhanced neuronal damage and microglial activation in the GFAP-ARO-KO mice after GCI, suggesting that the defects in the KO mice are because of a loss of E2 rather than an increase in precursor androgens. In conclusion, the current study provides novel genetic evidence for a beneficial role of astrocyte-derived E2 in reactive astrogliosis, microglial activation, and neuroprotection following an ischemic injury to the brain.SIGNIFICANCE STATEMENT Following cerebral ischemia, reactive astrocytes express the enzyme aromatase and produce 17ß-estradiol (E2), although the precise role of astrocyte-derived E2 is poorly understood. In this study, we generated a glial fibrillary acidic protein (GFAP) promoter-driven aromatase knock-out (GFAP-ARO-KO) mouse to deplete astrocyte-derived E2 and elucidate its roles after global cerebral ischemia (GCI). The GFAP-ARO-KO mice exhibited significantly attenuated reactive astrogliosis, as well as enhanced microglial activation, neuronal damage, and cognitive dysfunction after GCI. Transcriptome analysis further revealed that astrocyte-derived E2 was critical for the induction of the JAK-STAT3 signaling pathway, as well as the A2 reactive astrocyte phenotype after ischemia. Collectively, these findings indicate that astrocyte-derived E2 has a key role in the regulation of reactive astrogliosis, microglial activation, and neuroprotection after cerebral ischemia.


Assuntos
Aromatase/genética , Astrócitos/metabolismo , Isquemia Encefálica/metabolismo , Estradiol/metabolismo , Gliose/metabolismo , Hipocampo/metabolismo , Animais , Aromatase/metabolismo , Astrócitos/efeitos dos fármacos , Astrócitos/patologia , Isquemia Encefálica/genética , Isquemia Encefálica/patologia , Condicionamento Clássico/fisiologia , Modelos Animais de Doenças , Estradiol/farmacologia , Proteína Glial Fibrilar Ácida/metabolismo , Gliose/genética , Gliose/patologia , Hipocampo/efeitos dos fármacos , Hipocampo/patologia , Camundongos , Camundongos Knockout , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neuroproteção/fisiologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia
16.
Brain ; 143(12): 3748-3762, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-33184651

RESUMO

In Alzheimer's disease, the tauopathy is known as a major mechanism responsible for the development of cognitive deficits. Early biomarkers of such affectations for diagnosis/stratification are crucial in Alzheimer's disease research, and brain connectome studies increasingly show their potential establishing pathology fingerprints at the network level. In this context, we conducted an in vivo multimodal MRI study on young Thy-Tau22 transgenic mice expressing tauopathy, performing resting state functional MRI and structural brain imaging to identify early connectome signatures of the pathology, relating with histological and behavioural investigations. In the prodromal phase of tauopathy, before the emergence of cognitive impairments, Thy-Tau22 mice displayed selective modifications of brain functional connectivity involving three main centres: hippocampus (HIP), amygdala (AMG) and the isocortical areas, notably the somatosensory (SS) cortex. Each of these regions showed differential histopathological profiles. Disrupted ventral HIP-AMG functional pathway and altered dynamic functional connectivity were consistent with high pathological tau deposition and astrogliosis in both hippocampus and amygdala, and significant microglial reactivity in amygdalar nuclei. These patterns were concurrent with widespread functional hyperconnectivity of memory-related circuits of dorsal hippocampus-encompassing dorsal HIP-SS communication-in the absence of significant cortical histopathological markers. These findings suggest the coexistence of two intermingled mechanisms of response at the functional connectome level in the early phases of pathology: a maladaptive and a likely compensatory response. Captured in the connectivity patterns, such first responses to pathology could further be used in translational investigations as a lead towards an early biomarker of tauopathy as well as new targets for future treatments.


Assuntos
Transtornos da Memória/patologia , Transtornos da Memória/psicologia , Rede Nervosa/patologia , Tauopatias/patologia , Tauopatias/psicologia , Animais , Astrócitos/patologia , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Disfunção Cognitiva/genética , Disfunção Cognitiva/psicologia , Conectoma , Progressão da Doença , Gliose/patologia , Humanos , Processamento de Imagem Assistida por Computador , Imageamento por Ressonância Magnética , Transtornos da Memória/etiologia , Camundongos , Camundongos Transgênicos , Rede Nervosa/diagnóstico por imagem , Tauopatias/complicações , Tauopatias/diagnóstico por imagem , Proteínas tau/metabolismo
17.
Int J Mol Sci ; 21(20)2020 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-33050466

RESUMO

Amyloid-beta oligomers (AßO) have been proposed as the most potent neurotoxic and inflammation inducers in Alzheimer's disease (AD). AßO contribute to AD pathogenesis by impairing the production of several cytokines and inflammation-related signaling pathways, such as the Janus kinases/signal transducer of transcription factor-3 (JAK/STAT3) pathway. STAT3 modulates glial activation, indirectly regulates Aß deposition, and induces cognitive decline in AD transgenic models. However, in vivo studies using an AßO microinjection rat model have not yet explored STAT3 role. The main purpose of this study was to elucidate if a single microinjection of AßO could promote an increased expression of STAT3 in glial cells favoring neuroinflammation and neurodegeneration. We designed a model of intrahippocampal microinjection and assessed glial activation, cytokines production, STAT3 expression, and neurodegeneration in time. Our results showed robust expression of STAT3 in glial cells (mainly in astrocytes) and neurons, correlating with neuronal death in response to AßO administration. A STAT3 inhibition assay conducted in rat primary hippocampal cultures, suggested that the induction of the transcription factor by AßO in astrocytes leads them to an activation state that may favor neuronal death. Notwithstanding, pharmacological inhibition of the JAK2/STAT3 pathway should be focused on astrocytes because it is also essential in neurons survival. Overall, these findings strongly suggest the participation of STAT3 in the development of neurodegeneration.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Astrócitos/metabolismo , Gliose/etiologia , Gliose/metabolismo , Neurônios/metabolismo , Fator de Transcrição STAT3/metabolismo , Doença de Alzheimer/etiologia , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Animais , Astrócitos/patologia , Biomarcadores , Morte Celular , Modelos Animais de Doenças , Suscetibilidade a Doenças , Imunofluorescência , Gliose/patologia , Hipocampo/metabolismo , Hipocampo/patologia , Imuno-Histoquímica , Agregados Proteicos , Agregação Patológica de Proteínas/genética , Agregação Patológica de Proteínas/metabolismo , Multimerização Proteica , Ratos , Fator de Transcrição STAT3/genética
18.
Int J Mol Sci ; 21(19)2020 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-33008136

RESUMO

Inherited photoreceptor degenerations are not treatable diseases and a frequent cause of blindness in working ages. In this study we investigate the safety, integration and possible rescue effects of intravitreal and subretinal transplantation of adult human bone-marrow-derived mononuclear stem cells (hBM-MSCs) in two animal models of inherited photoreceptor degeneration, the P23H-1 and the Royal College of Surgeons (RCS) rat. Immunosuppression was started one day before the injection and continued through the study. The hBM-MSCs were injected in the left eyes and the animals were processed 7, 15, 30 or 60 days later. The retinas were cross-sectioned, and L- and S- cones, microglia, astrocytes and Müller cells were immunodetected. Transplantations had no local adverse effects and the CD45+ cells remained for up to 15 days forming clusters in the vitreous and/or a 2-3-cells-thick layer in the subretinal space after intravitreal or subretinal injections, respectively. We did not observe increased photoreceptor survival nor decreased microglial cell numbers in the injected left eyes. However, the injected eyes showed decreased GFAP immunoreactivity. We conclude that intravitreal or subretinal injection of hBM-MSCs in dystrophic P23H-1 and RCS rats causes a decrease in retinal gliosis but does not have photoreceptor neuroprotective effects, at least in the short term. However, this treatment may have a potential therapeutic effect that merits further investigation.


Assuntos
Gliose/cirurgia , Transplante de Células-Tronco Mesenquimais , Retina/cirurgia , Células Fotorreceptoras Retinianas Cones/transplante , Degeneração Retiniana/cirurgia , Células-Tronco Adultas/transplante , Animais , Células da Medula Óssea/citologia , Transplante de Medula Óssea , Sobrevivência Celular/fisiologia , Modelos Animais de Doenças , Gliose/patologia , Humanos , Ratos , Retina/patologia , Células Fotorreceptoras Retinianas Cones/patologia , Degeneração Retiniana/patologia
19.
Nat Commun ; 11(1): 4966, 2020 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-33009404

RESUMO

Temporal lobe epilepsy (TLE) is the most common type of drug-resistant epilepsy in adults, with an unknown etiology. A hallmark of TLE is the characteristic loss of layer 3 neurons in the medial entorhinal area (MEA) that underlies seizure development. One approach to intervention is preventing loss of these neurons through better understanding of underlying pathophysiological mechanisms. Here, we show that both neurons and glia together give rise to the pathology that is mitigated by the amino acid D-serine whose levels are potentially diminished under epileptic conditions. Focal administration of D-serine to the MEA attenuates neuronal loss in this region thereby preventing epileptogenesis in an animal model of TLE. Additionally, treatment with D-serine reduces astrocyte counts in the MEA, alters their reactive status, and attenuates proliferation and/or infiltration of microglia to the region thereby curtailing the deleterious consequences of neuroinflammation. Given the paucity of compounds that reduce hyperexcitability and neuron loss, have anti-inflammatory properties, and are well tolerated by the brain, D-serine, an endogenous amino acid, offers new hope as a therapeutic agent for refractory TLE.


Assuntos
Epilepsia do Lobo Temporal/tratamento farmacológico , Epilepsia do Lobo Temporal/patologia , Serina/uso terapêutico , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/patologia , Comportamento Animal , Encéfalo/patologia , Córtex Entorrinal/efeitos dos fármacos , Córtex Entorrinal/patologia , Gliose/patologia , Inflamação/patologia , Microglia/efeitos dos fármacos , Microglia/patologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Ratos Sprague-Dawley , Serina/administração & dosagem , Serina/farmacologia
20.
Life Sci ; 263: 118556, 2020 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-33038375

RESUMO

AIMS: White matter injury (WMI) is the main form of brain injury in preterm neonate survivors, and perinatal inflammation is implicated in the pathogenesis of WMI. It has been demonstrated that dexmedetomidine, an anesthetic adjuvant, possesses neuroprotective effects in both preclinical and clinical trials. The present study was conducted to explore whether dexmedetomidine could protect against neurobehavioral impairments and myelination deficits caused by lipopolysaccharide (LPS) exposure in the early postnatal rat brain. MAIN METHODS: LPS (2 mg/kg) was intraperitoneally (i.p.) injected in Sprague-Dawley rat pups on postnatal day 2 (P2). Dexmedetomidine (25 µg/kg) or vehicle was given i.p. immediately after LPS injection. STAT3 and p-STAT3 expression were detected by western blot in rat brain 24 h after drug administration. Immunostaining for GFAP to was performed to evaluate astrocytic response at 24 h post-LPS and P14. Neurobehavioral tests (the righting reflex, negative geotaxis, and wire hanging maneuver tests) were performed from P5 to P10. Histological analysis of myelin content was accessed by immunohistochemistry for CNPase and MBP at P14. KEY FINDINGS: Our results showed that treatment with dexmedetomidine significantly ameliorated LPS-induced neurobehavioral abnormalities and myelin damage, which is accompanied by suppression of STAT3 activation and reactive astrogliosis. SIGNIFICANCE: Dexmedetomidine can alleviate neurobehavioral impairments and myelination deficits after LPS exposure in early postnatal rats, probably by mitigating STAT3-mediated reactive astrogliosis. Our results suggest that dexmedetomidine might be a promising agent to treat brain injury in neonates.


Assuntos
Comportamento Animal/efeitos dos fármacos , Lesões Encefálicas/tratamento farmacológico , Dexmedetomidina/farmacologia , Gliose/prevenção & controle , Inflamação/prevenção & controle , Lipopolissacarídeos/toxicidade , Fármacos Neuroprotetores/farmacologia , Agonistas de Receptores Adrenérgicos alfa 2/farmacologia , Animais , Animais Recém-Nascidos , Lesões Encefálicas/induzido quimicamente , Lesões Encefálicas/metabolismo , Lesões Encefálicas/patologia , Modelos Animais de Doenças , Feminino , Gliose/induzido quimicamente , Gliose/metabolismo , Gliose/patologia , Inflamação/induzido quimicamente , Inflamação/metabolismo , Inflamação/patologia , Masculino , Gravidez , Ratos , Ratos Sprague-Dawley , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo
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