RESUMO
Cytosine base editors (CBEs), which enable precise C-to-T substitutions, have been restricted by potential safety risks, including DNA off-target edits, RNA off-target edits and additional genotoxicity such as DNA damages induced by double-strand breaks (DSBs). Though DNA and RNA off-target edits have been ameliorated via various strategies, evaluation and minimization of DSB-associated DNA damage risks for most CBEs remain to be resolved. Here we demonstrate that YE1, an engineered CBE variant with minimized DNA and RNA off-target edits, could induce prominent DSB-associated DNA damage risks, manifested as γH2AX accumulation in human cells. We then perform deaminase engineering for two deaminases lamprey LjCDA1 and human APOBEC3A, and generate divergent CBE variants with eliminated DSB-associated DNA damage risks, in addition to minimized DNA/RNA off-target edits. Furthermore, the editing scopes and sequence preferences of APOBEC3A-derived CBEs could be further diversified by internal fusion strategy. Taken together, this study provides updated evaluation platform for DSB-associated DNA damage risks of CBEs and further generates a series of safer toolkits with diversified editing signatures to expand their applications.
Assuntos
Citosina , Edição de Genes , Humanos , RNA/genética , Dano ao DNA , DNA/genética , Sistemas CRISPR-CasRESUMO
Mast cell extracellular traps (MCETs) released by mast cells contribute to host defense. In this study, we investigated the effects of MCETs released from mast cells after infection with a periodontal pathogen Fusobacterium nucleatum. We found that F. nucleatum induced MCET release from mast cells, and that MCETs expressed macrophage migration inhibitory factor (MIF). Notably, MIF bound to MCETs induced proinflammatory cytokine production by monocytic cells. These findings suggest that MIF expressed on MCETs, released from mast cells upon infection with F. nucleatum, promotes inflammatory responses that may be associated with the pathogenesis of periodontal disease.
Assuntos
Armadilhas Extracelulares , Fatores Inibidores da Migração de Macrófagos , Fatores Inibidores da Migração de Macrófagos/metabolismo , Mastócitos , Fusobacterium nucleatum , Armadilhas Extracelulares/metabolismoRESUMO
Neurological and psychiatric disorders have overlapped phenotypic profiles, but the underlying tissue-specific functional processes remain largely unknown. In this study, we explore the shared tissue-specificity among 14 neuropsychiatric disorders through the disrupted long-range gene regulations by GWAS-identified regulatory SNPs. Through Hi-C interactions, averagely 38.0% and 17.2% of the intergenic regulatory SNPs can be linked to target protein-coding genes in brain and non-brain tissues, respectively. Interestingly, while the regulatory target genes in the brain tend to enrich in nervous system development related processes, those in the non-brain tissues are inclined to interfere with synapse and neuroinflammation related processes. Compared to psychiatric disorders, neurological disorders present more prominently the neuroinflammatory processes in both brain and non-brain tissues, indicating an intrinsic difference in mechanisms. Through tissue-specific gene regulatory networks, we then constructed disorder similarity networks in two brain and three non-brain tissues, highlighting both known disorder clusters (e.g. the neurodevelopmental disorders) and unexpected disorder clusters (e.g. Parkinson's disease is consistently grouped with psychiatric disorders). We showcase the potential pharmaceutical applications of the small bowel and its disorder clusters, illustrated by the known drug targets NR1I3 and NFACT1, and their small bowel-specific regulatory modules. In conclusion, disrupted long-range gene regulations in both brain and non-brain tissues contribute to the similarity among distinct clusters of neuropsychiatric disorders, and the tissue-specifically shared functions and regulators for disease clusters may provide insights for future therapeutic investigations.
Assuntos
Redes Reguladoras de Genes , Transtornos do Neurodesenvolvimento , Encéfalo , Regulação da Expressão Gênica , Redes Reguladoras de Genes/genética , Humanos , Transtornos do Neurodesenvolvimento/genética , Polimorfismo de Nucleotídeo ÚnicoRESUMO
The human brain is the most complex organ consisting of billions of neuronal and non-neuronal cells that are organized into distinct anatomical and functional regions. Elucidating the cellular and transcriptome architecture underlying the brain is crucial for understanding brain functions and brain disorders. Thanks to the single-cell RNA sequencing technologies, it is becoming possible to dissect the cellular compositions of the brain. Although great effort has been made to explore the transcriptome architecture of the human brain, a comprehensive database with dynamic cellular compositions and molecular characteristics of the human brain during the lifespan is still not available. Here, we present STAB (a Spatio-Temporal cell Atlas of the human Brain), a database consists of single-cell transcriptomes across multiple brain regions and developmental periods. Right now, STAB contains single-cell gene expression profiling of 42 cell subtypes across 20 brain regions and 11 developmental periods. With STAB, the landscape of cell types and their regional heterogeneity and temporal dynamics across the human brain can be clearly seen, which can help to understand both the development of the normal human brain and the etiology of neuropsychiatric disorders. STAB is available at http://stab.comp-sysbio.org.
Assuntos
Encéfalo/metabolismo , Bases de Dados Genéticas , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica no Desenvolvimento , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , Encéfalo/citologia , Encéfalo/crescimento & desenvolvimento , Humanos , Armazenamento e Recuperação da Informação/métodos , Internet , Transtornos Mentais/genética , Transtornos Mentais/patologia , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/patologia , Neurônios/citologia , Neurônios/metabolismoRESUMO
Edible mushrooms are known to exert anti-inflammatory effects. In this study, the effects of ethanol extracts from edible mushrooms, such as Hericium erinaceus, and other edible mushrooms on inflammatory responses were investigated. Experiments were conducted using the inflammatory responses of human monocytes induced by lipopolysaccharide (LPS), a bacterial component, that provokes inflammation. Notably, we demonstrated that LPS mixed with ethanol and hot water extracts derived from edible mushrooms attenuated the production of inflammatory cytokines, such as interleukin (IL)-1ß, -6, and -8, induced by LPS in human monocytic cell cultures. Moreover, we found that the ethanol extract of H. erinaceus contained ergosterol, which attenuated IL-8 production in LPS-stimulated cells. Subsequent component analysis of the ethanol extract of H. erinaceus revealed that ergosterol binds to lipid A to attenuate LPS-induced inflammation. Together, our findings suggest that ergosterol in ethanol extracts from edible mushrooms can prevent the induction of inflammation by binding to LPS.
Assuntos
Agaricales , Lipopolissacarídeos , Humanos , Lipopolissacarídeos/uso terapêutico , Ergosterol/farmacologia , Etanol , Monócitos/metabolismo , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Agaricales/metabolismo , Inflamação/tratamento farmacológico , Citocinas/metabolismoRESUMO
BACKGROUND: Alzheimer's disease (AD), a progressive neurodegenerative disease, is the most common cause of dementia worldwide. Accumulating data support the contributions of the peripheral immune system in AD pathogenesis. However, there is a lack of comprehensive understanding about the molecular characteristics of peripheral immune cells in AD. METHODS: To explore the alterations of cellular composition and the alterations of intrinsic expression of individual cell types in peripheral blood, we performed cellular deconvolution in a large-scale bulk blood expression cohort and identified cell-intrinsic differentially expressed genes in individual cell types with adjusting for cellular proportion. RESULTS: We detected a significant increase and decrease in the proportion of neutrophils and B lymphocytes in AD blood, respectively, which had a robust replicability across other three AD cohorts, as well as using alternative algorithms. The differentially expressed genes in AD neutrophils were enriched for some AD-associated pathways, such as ATP metabolic process and mitochondrion organization. We also found a significant enrichment of protein-protein interaction network modules of leukocyte cell-cell activation, mitochondrion organization, and cytokine-mediated signaling pathway in neutrophils for AD risk genes including CD33 and IL1B. Both changes in cellular composition and expression levels of specific genes were significantly associated with the clinical and pathological alterations. A similar pattern of perturbations on the cellular proportion and gene expression levels of neutrophils could be also observed in mild cognitive impairment (MCI). Moreover, we noticed an elevation of neutrophil abundance in the AD brains. CONCLUSIONS: We revealed the landscape of molecular perturbations at the cellular level for AD. These alterations highlight the putative roles of neutrophils in AD pathobiology.
Assuntos
Doença de Alzheimer , Disfunção Cognitiva , Doenças Neurodegenerativas , Encéfalo , Estudos de Coortes , HumanosRESUMO
With the development of various nanomaterial expected to be used in biomedical fields, it is more important to evaluate and understand their potential effects on biological system. In this work, two proteins with different structure, Villin Headpiece (HP35) with α-helix structure and protofibrils Aß1-42 with five ß-strand chains, were selected and their interactions with silicene were studied by means of molecular dynamics (MD) simulation to reveal the potential effect of silicene on the structure and function of biomolecules. The obtained results indicated that silicene could rapidly attract HP35 and Aß1-42 fibrils onto the surface to form a stable binding. The adsorption strength was moderate and no significant structural distortion of HP35 and Aß1-42 fibrils was observed. Moreover, the strength of calculated the H-bonds in neighbor chain of Aß1-42 fibrils indicated that the mild interactions between silicene and fibrils could regularize the structure of Aß1-42 fibrils and stabilize the interactions between five chains of fibrils protein, which might enhance the aggregation of Aß1-42 fibrils. This study provides a new insight for understanding the interaction between nanomaterials and biomolecules and moves forward the development of silicene into biomedical fields.
Assuntos
Amiloide , Simulação de Dinâmica Molecular , Amiloide/química , Peptídeos beta-Amiloides/química , Proteínas dos Microfilamentos/metabolismo , Fragmentos de Peptídeos/químicaRESUMO
BACKGROUND AND OBJECTIVE: It has been reported that long non-coding RNAs (lncRNAs), such as metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), act as key regulators of the development of inflammatory diseases. However, it is unclear whether MALAT1 regulates the function of human gingival fibroblasts (HGFs) in periodontitis. This study is to explore the role of MALAT1 on inflammatory cytokine production of HGFs. MATERIAL AND METHODS: Primary HGFs were harvested from human gingiva. MALAT1 was detected in inflammatory and healthy gingival tissues via quantitative real-time PCR (qRT-PCR). Bioinformatics analysis, dual-luciferase reporter assay, and RNA-binding protein immunoprecipitation (RIP) were used to detect the relationship among MALAT1, toll-like receptor 4 (TLR4), and microRNA (miR) -20a. After transfection LPS-treated HGFs with MALAT1 siRNA (si-MALAT1), miR-20a mimic or overexpression MALAT1 plasmid (sno-MALAT1), the levels of MALAT1, miR-20a, TLR4, IL-6 and IL-8 were analyzed by qRT-PCR, enzyme-linked immunosorbent assay, or western blot assay. RESULTS: MALAT1 up-regulated in inflammatory gingival tissues of chronic periodontitis. MiR-20a was bound with MALAT1 and TLR4 3'-UTR in RNA-protein complex with Ago2, respectively. Moreover, MALAT1, TLR4, IL-6, and IL-8 increased while miR-20a decreased after 1 µg/mL Porphyromonas gingivalis lipopolysaccharide (LPS) or Escherichia coli LPS stimulation. MiR-20a inhibited the expression of proinflammatory cytokines via binding to TLR4 3'-UTR. In addition, MALAT1 increased TLR4 level and the secretion of inflammatory cytokines. CONCLUSION: MALAT1 enhances inflammatory cytokine production through sponging miR-20a and releasing TLR4, indicating a regulatory role of MALAT1 in periodontal inflammation.
Assuntos
Citocinas/metabolismo , Fibroblastos/metabolismo , MicroRNAs/genética , RNA Longo não Codificante/metabolismo , Receptor 4 Toll-Like/metabolismo , Gengiva/citologia , Humanos , Inflamação , LipopolissacarídeosRESUMO
BACKGROUND: T-2 toxin is a widely distributed mycotoxin in cereals. HT-2 toxin is the major metabolite, which is also a contaminant in cereals. T-2 toxin and HT-2 toxin have been identified as having carcinogenic, hepatotoxic, teratogenic and immunotoxic properties. To reduce the risk of contamination, a rapid, highly sensitive and inexpensive assay for the detection is required. RESULTS: In this study a high-sensitivity chemiluminescent enzyme-linked immunoassay (CL-ELISA) of T-2 toxin and HT-2 toxin was developed. With the help of the chemiluminescent substrate, this protocol showed a highly sensitive character with an IC50 as low as 33.28 ng mL-1 and 27.27 ng mL-1 for T-2 and HT-2, respectively. In addition, this method had no cross-reaction with other structurally related mycotoxins. CONCLUSION: These results indicated that the developed CL-ELISA could be applied for the detection of T-2 toxin and HT-2 toxin in actual samples without complicated steps. © 2016 Society of Chemical Industry.
Assuntos
Carcinógenos Ambientais/análise , Contaminação de Alimentos , Inspeção de Alimentos/métodos , Oryza/química , Sementes/química , Toxina T-2/análogos & derivados , Toxina T-2/análise , Anticorpos Monoclonais/metabolismo , Carcinógenos Ambientais/metabolismo , China , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática , Limite de Detecção , Medições Luminescentes , Metanol/química , Reprodutibilidade dos Testes , Solubilidade , Solventes/química , Toxina T-2/metabolismoRESUMO
Color is a visual cue that can convey emotions and attract attention, and there is no doubt that brightness is an important element of color differentiation. To examine the impact of art training on color perception, 44 participants were assigned to two groups-one for those with and one for those without art training-in an EEG experiment. While the participants had their electroencephalographic data recorded, they scored their emotional responses to color stimuli of different brightness levels based on the Munsell color system. The behavioral results revealed that in both groups, high-brightness colors were rated more positively than low-brightness colors. Furthermore, event-related potential results for the artist group showed that high-brightness colors enhanced P2 and P3 amplitudes. Moreover, non-artists had longer N2 latency than artists, and there was a significant Group × Brightness interaction separately for the N2 and P3 components. Simple effect analysis showed that N2 and P3 amplitudes were substantially higher for high-brightness stimuli than for lower-brightness stimuli in the artistic group, but this was not the case in the non-artist group. Additionally, evoked event-related oscillation results showed that in both groups, high-brightness stimuli also elicited large delta, theta, and alpha as well as low gamma responses. These results indicate that high-brightness color stimuli elicit more positive emotions and stronger neurological reactions and that artistic training may have a positive effect on top-down visual perception.
Assuntos
Percepção de Cores , Eletroencefalografia , Estimulação Luminosa , Humanos , Masculino , Feminino , Percepção de Cores/fisiologia , Adulto Jovem , Estimulação Luminosa/métodos , Adulto , Potenciais Evocados/fisiologia , Emoções/fisiologia , Tempo de Reação/fisiologia , Potenciais Evocados Visuais/fisiologia , Análise de VariânciaRESUMO
BACKGROUND: There is growing evidence for an association between chronic renal disease (CKD) and adverse cerebrovascular events because of the overlap of several risk factors. The purpose of this study is to examine the epidemiology of CKD and the characteristics of risk factors for CKD in the population with ischaemic stroke. METHODS: This retrospective study included 571 patients with ischaemic stroke. Estimated glomerular filtration rate (eGFR) was calculated by the Modification of Diet in Renal Disease (MDRD) study equation. Renal function was assessed according to the Kidney Disease Outcomes Quality Initiative (K/DOQI)-CKD classification. RESULTS: Study demonstrated that the major factors associated with CKD in the ischaemic stroke patients were age, diabetes mellitus, hypertension, systolic blood pressure, LDL cholesterol and serum uric acid. Diabetes mellitus (OR 4·146, 95% CI 1·047-16·418, P = 0·043), hypertension and diabetes mellitus (OR 3·574, 95% CI 1·248-10·234, P = 0·018), serum uric acid (OR 1·010, 95% CI 1·006-1·013, P < 0·001) and LDL cholesterol (OR 1·431, 95% CI 1·063-1·928, P = 0·018) were independent risk factors associated with CKD in the patients with ischaemic stroke. CONCLUSIONS: The patients with ischaemic stroke may be considered as a high-risk population for CKD and be aggressively managed for CKD prevention. The high prevalence of CKD in population with ischaemic stroke prompts the need for greater public awareness about risks of CKD.
Assuntos
Insuficiência Renal Crônica/complicações , Acidente Vascular Cerebral/complicações , China/epidemiologia , Nefropatias Diabéticas/complicações , Nefropatias Diabéticas/epidemiologia , Feminino , Taxa de Filtração Glomerular/fisiologia , Humanos , Hipertensão/complicações , Hipertensão/epidemiologia , Masculino , Pessoa de Meia-Idade , Análise de Regressão , Insuficiência Renal Crônica/epidemiologia , Insuficiência Renal Crônica/fisiopatologia , Estudos Retrospectivos , Fatores de Risco , Acidente Vascular Cerebral/epidemiologiaRESUMO
OBJECTIVE: This study aims to investigate the effects of microRNA-126 (miR-126) on the macrophage polarization in vitro and alveolar bone resorption in vivo. DESIGN: The relationship between miR-126 and MEK/ERK kinase 2 (MEKK2) was confirmed by dual-luciferase reporter assay. Real-time quantitative polymerase chain reaction, enzyme-linked immunosorbent assay or Western blot was used to detect the changes of miR-126, inducible nitric oxide synthase (iNOS), arginase-1 (Arg-1), tumor necrosis factor (TNF)-α, interleukin (IL)-10, MEKK2 and MEKK2-related pathways: mitogen-activated protein kinase (MAPK) and nuclear factor kappa-B (NF-κB) in RAW264.7 macrophages challenged with Porphyromonas gingivalis (P. gingivalis) lipopolysaccharide (LPS) and/or high glucose and/or miR-126 mimic. In mice with diabetic periodontitis, the expressions of iNOS and Arg-1 in gingiva, and alveolar bone level were detected after miR-126 mimic injection. RESULTS: MiR-126 could directly bind with MEKK2 3'-untranslated region (UTR). MEKK2, phosphorylation of NF-κB and MAPK signaling proteins, TNF-α and iNOS increased (P < 0.05), while miR-126, Arg-1 and IL-10 were inhibited (P < 0.05) in macrophage challenged with high glucose and/or P. gingivalis LPS, however, miR-126 mimic reversed these effects (P < 0.05). The expressions of iNOS in gingiva and alveolar bone resorption were elevated (P < 0.05), the expression of Arg-1 in gingiva decreased (P < 0.05) in mice with diabetic periodontitis, which could be inhibited by miR-126 mimic. CONCLUSIONS: miR-126 might prevent alveolar bone resorption in diabetic periodontitis and inhibit macrophage M1 polarization via regulating MEKK2 signaling pathway.
Assuntos
Perda do Osso Alveolar , Diabetes Mellitus , MicroRNAs , Periodontite , Camundongos , Animais , NF-kappa B/metabolismo , MicroRNAs/metabolismo , Lipopolissacarídeos/farmacologia , Lipopolissacarídeos/metabolismo , Periodontite/prevenção & controle , Periodontite/metabolismo , Macrófagos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Perda do Osso Alveolar/prevenção & controle , Perda do Osso Alveolar/patologia , GlucoseRESUMO
BACKGROUND: Prioritizing genes that underlie complex brain disorders poses a considerable challenge. Despite previous studies have found that they shared symptoms and heterogeneity, it remained difficult to systematically identify the risk genes associated with them. METHODS: By using the CAGE (Cap Analysis of Gene Expression) read alignment files for 439 human cell and tissue types (including primary cells, tissues and cell lines) from FANTOM5 project, we predicted enhancer-promoter interactions (EPIs) of 439 cell and tissue types in human, and examined their reliability. Then we evaluated the genetic heritability of 17 diverse brain disorders and behavioral-cognitive phenotypes in each neural cell type, brain region, and developmental stage. Furthermore, we prioritized genes associated with brain disorders and phenotypes by leveraging the EPIs in each neural cell and tissue type, and analyzed their pleiotropy and functionality for different categories of disorders and phenotypes. Finally, we characterized the spatiotemporal expression dynamics of these associated genes in cells and tissues. RESULTS: We found that identified EPIs showed activity specificity and network aggregation in cell and tissue types, and enriched TF binding in neural cells played key roles in synaptic plasticity and nerve cell development, i.e., EGR1 and SOX family. We also discovered that most neurological disorders exhibit heritability enrichment in neural stem cells and astrocytes, while psychiatric disorders and behavioral-cognitive phenotypes exhibit enrichment in neurons. Furthermore, our identified genes recapitulated well-known risk genes, which exhibited widespread pleiotropy between psychiatric disorders and behavioral-cognitive phenotypes (i.e., FOXP2), and indicated expression specificity in neural cell types, brain regions, and developmental stages associated with disorders and phenotypes. Importantly, we showed the potential associations of brain disorders with brain regions and developmental stages that have not been well studied. CONCLUSIONS: Overall, our study characterized the gene-enhancer regulatory networks and genetic mechanisms in the human neural cells and tissues, and illustrated the value of reanalysis of publicly available genomic datasets.
Assuntos
Encefalopatias , Humanos , Reprodutibilidade dos Testes , Regiões Promotoras Genéticas , Neurônios , Redes Reguladoras de GenesRESUMO
Cytidine and adenosine deaminases are required for cytosine and adenine editing of base editors respectively, and no single deaminase could enable concurrent and comparable cytosine and adenine editing. Additionally, distinct properties of cytidine and adenosine deaminases lead to various types of off-target effects, including Cas9-indendepent DNA off-target effects for cytosine base editors (CBEs) and RNA off-target effects particularly severe for adenine base editors (ABEs). Here we demonstrate that 25 TadA orthologs could be engineered to generate functional ABEs, CBEs or ACBEs via single or double mutations, which display minimized Cas9-independent DNA off-target effects and genotoxicity, with orthologs B5ZCW4, Q57LE3, E8WVH3, Q13XZ4 and B3PCY2 as promising candidates for further engineering. Furthermore, RNA off-target effects of TadA ortholog-derived base editors could be further reduced or even eliminated by additional single mutation. Taken together, our work expands the base editing toolkits, and also provides important clues for the potential evolutionary process of deaminases.
Assuntos
Citosina , Edição de Genes , Adenina , DNA , RNA , Adenosina/genética , Sistemas CRISPR-Cas/genéticaRESUMO
Although miniature CRISPR-Cas12f systems were recently developed, the editing efficacy and targeting range of derived miniature cytosine and adenine base editors (miniCBEs and miniABEs) have not been comprehensively addressed. Moreover, functional miniCBEs have not yet be established. Here we generate various Cas12f-derived miniCBEs and miniABEs with improved editing activities and diversified targeting scopes. We reveal that miniCBEs generated with traditional cytidine deaminases exhibit wide editing windows and high off-targeting effects. To improve the editing signatures of classical CBEs and derived miniCBEs, we engineer TadA deaminase with mutagenesis screening to generate potent miniCBEs with high precision and minimized off-target effects. We show that newly designed miniCBEs and miniABEs are able to correct pathogenic mutations in cell lines and introduce genetic mutations efficiently via adeno-associated virus delivery in the brain in vivo. Together, this study provides alternative strategies for CBE development, expands the toolkits of miniCBEs and miniABEs and offers promising therapeutic tools for clinical applications.
Assuntos
Sistemas CRISPR-Cas , Edição de Genes , Sistemas CRISPR-Cas/genética , Mutação , Citidina Desaminase/genética , Citidina Desaminase/metabolismo , Citosina/metabolismoRESUMO
BACKGROUND: Subjective or objective subtle cognitive decline (SCD) is considered the preclinical manifestation of Alzheimer's disease (AD), which is a potentially crucial window for preventing or delaying the progression of the disease. METHODS: To explore the potential mechanism of disease progression and identify relevant biomarkers, we comprehensively assessed the peripheral blood transcriptomic alterations in SCD, covering lncRNA, mRNA, and miRNA. FINDINGS: Dysregulated protein-coding mRNA at both gene and isoform levels implicated impairment in the type I interferon signaling pathway in SCD. Specifically, this pathway was regulated by the transcription factor STAT1 and ncRNAs NRIR and has-miR-146a-5p. The miRNA-mRNA-lncRNA co-expression network revealed hub genes for the interferon module. Individuals with lower interferon signaling activity and lower expression of a hub gene STAT1 exhibited a higher conversion rate to mild cognitive impairment (MCI). INTERPRETATION: Our findings illustrated the down-regulation of interferon signaling activity would potentially increase the risk of disease progression and thus serve as a pre-disease biomarker. FUNDING: This work was partly supported by National Key R&D Program of China (2020YFA0712403), National Natural Science Foundation of China (61932008), Shanghai Municipal Science and Technology Major Project (2018SHZDZX01), the 111 Project (No. B18015) of China, Greater Bay Area Institute of Precision Medicine (Guangzhou) (Grand No. IPM21C008), Natural Science Foundation of Shanghai (21ZR1403200), and Shanghai Center for Brain Science and Brain-Inspired Technology.
Assuntos
Doença de Alzheimer , Disfunção Cognitiva , Interferon Tipo I , MicroRNAs , RNA Longo não Codificante , Doença de Alzheimer/genética , Doença de Alzheimer/psicologia , Biomarcadores/metabolismo , China , Disfunção Cognitiva/genética , Disfunção Cognitiva/psicologia , Progressão da Doença , Humanos , Interferon Tipo I/genética , MicroRNAs/genética , Testes Neuropsicológicos , RNA Longo não Codificante/genética , RNA Mensageiro , Transdução de Sinais , TranscriptomaRESUMO
Plasminogen activator inhibitor-1 (PAI-1), a serine protease inhibitor, is constitutively produced by endothelial cells and plays a vital role in maintaining vascular homeostasis. Chronic periodontitis is an inflammatory disease characterized by bleeding of periodontal tissues that support the tooth. In this study, we aimed to determine the role of PAI-1 produced by endothelial cells in response to infections caused by the primary periodontal pathogen Porphyromonas gingivalis. We demonstrated that P. gingivalis infection resulted in significantly reduced PAI-1 levels in human endothelial cells. This reduction in PAI-1 levels could be attributed to the proteolysis of PAI-1 by P. gingivalis proteinases, especially lysine-specific gingipain-K (Kgp). We demonstrated the roles of these degradative enzymes in the endothelial cells using a Kgp-specific inhibitor and P. gingivalis gingipain-null mutants, in which the lack of the proteinases resulted in the absence of PAI-1 degradation. The degradation of PAI-1 by P. gingivalis induced a delayed wound healing response in endothelial cell layers via the low-density lipoprotein receptor-related protein. Our results collectively suggested that the proteolysis of PAI-1 in endothelial cells by gingipains of P. gingivalis might lead to the deregulation of endothelial homeostasis, thereby contributing to the permeabilization and dysfunction of the vascular endothelial barrier.
Assuntos
Células Endoteliais , Porphyromonas gingivalis , Adesinas Bacterianas/metabolismo , Cisteína Endopeptidases/metabolismo , Cisteína Endopeptidases/farmacologia , Cisteína Endopeptidases Gingipaínas , Humanos , Inibidor 1 de Ativador de Plasminogênio , Porphyromonas gingivalis/fisiologia , CicatrizaçãoRESUMO
OBJECTIVE: This study aimed to fabricate Hyaluronic Acid (HA)/parecoxib-loaded PLGA microspheres for the treatment of Temporomandibular Disorders (TMD) and investigate the in vitro and in vivo effect of the microsphere system to solve the issues of poor drug delivery and short duration on drug concentration in conventional TMD therapy. METHODS: The microspheres were prepared by the double emulsion (w/o/w) method. Various formulations were compared in terms of particle size, drug loading rate and encapsulation rate. Scanning Electron Microscopy (SEM), Differential Scanning Calorimetry (DSC) and FT-IR spectroscopy were performed to evaluate physicochemical properties. The drug release behavior of microspheres and toxicity assay on synovial cells were investigated. The in vitro anti-inflammatory effect on inflammatory markers, such as IL-1ß, TNF-α and COX-2, was assessed by real-time PCR. Then, the in vivo therapeutic effect of microspheres was investigated using mechanically-induced rat synovitis model. Protein levels of inflammatory cytokines (IL-1ß, TNF-α and COX-2) from TMJ periarticular tissues were quantified by Enzyme-Linked Immunosorbent Assay (ELISA). RESULTS: The results showed that microspheres were morphologically regular, smooth and non-cohesive. The average particle size of the microspheres was (25.32 ± 1.01) µm. The drug loading rate of parecoxib was 17.12%-20.95% with encapsulation efficiency reaching 51.9%-54.7%. In vitro drug release tests showed a successful sustained release over 28 days with a burst of 19.98% of the total drug substance. Treatment with HA/parecoxib-loaded PLGA microspheres declined the mRNA expression of IL-1ß, TNF-α and COX-2 induced by LPS in articular synovial cells. Moreover, in vivo results demonstrated that the intra-articular microspheres significantly reduced protein levels of inflammatory cytokines (IL-1ß, TNF-α and COX-2) for more than two weeks and stopped the mechanically-induced synovitis in its tracks in rat models. CONCLUSION: The study presented new and potential insights into treatments of TMD using PLGA microspheres loaded with HA and parecoxib as a successful drug delivery system.
Assuntos
Ácido Hialurônico , Isoxazóis/farmacologia , Transtornos da Articulação Temporomandibular , Animais , Isoxazóis/química , Microesferas , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Ratos , Espectroscopia de Infravermelho com Transformada de Fourier , Transtornos da Articulação Temporomandibular/tratamento farmacológicoRESUMO
The accumulation of vast amounts of multimodal data for the human brain, in both normal and disease conditions, has provided unprecedented opportunities for understanding why and how brain disorders arise. Compared with traditional analyses of single datasets, the integration of multimodal datasets covering different types of data (i.e., genomics, transcriptomics, imaging, etc.) has shed light on the mechanisms underlying brain disorders in greater detail across both the microscopic and macroscopic levels. In this review, we first briefly introduce the popular large datasets for the brain. Then, we discuss in detail how integration of multimodal human brain datasets can reveal the genetic predispositions and the abnormal molecular pathways of brain disorders. Finally, we present an outlook on how future data integration efforts may advance the diagnosis and treatment of brain disorders.
Assuntos
Encefalopatias , Encéfalo , Encéfalo/diagnóstico por imagem , Encefalopatias/diagnóstico , Genômica , Humanos , TranscriptomaRESUMO
PURPOSE: Zinc (Zn), an essential trace element in the body, has stable chemical properties, excellent osteogenic ability and moderate immunomodulatory property. In the present study, a Zn-incorporated TiO2 nanotube (TNT) was fabricated on titanium (Ti) implant material. We aimed to evaluate the influence of nano-scale topography and Zn on behaviors of murine RAW 264.7 macrophages. Moreover, the effects of Zn-incorporated TNT surface-regulated macrophages on the behaviors and osteogenic differentiation of murine MC3T3-E1 osteoblasts were also investigated. METHODS: TNT coatings were firstly fabricated on a pure Ti surface using anodic oxidation, and then nano-scale Zn particles were incorporated onto TNTs by the hydrothermal method. Surface topography, chemical composition, roughness, hydrophilicity, Zn release pattern and protein adsorption ability of the Zn-incorporated TiO2 nanotube surface were characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD) and X-ray photoelectron spectroscopy (XPS), surface profiler, contact angle test, Zn release test and protein adsorption test. The cell behaviors and both pro-inflammatory (M1) and pro-regenerative (M2) marker gene and protein levels in macrophages cultured on Zn-incorporated TNTs surfaces with different TNT diameters were detected. The supernatants of macrophages were extracted and preserved as conditioned medium (CM). Furthermore, the behaviors and osteogenic properties of osteoblasts cultured in CM on various surfaces were evaluated. RESULTS: The release profile of Zn on Zn-incorporated TNT surfaces revealed a controlled release pattern. Macrophages cultured on Zn-incorporated TNT surfaces displayed enhanced gene and protein expression of M2 markers, and M1 markers were moderately inhibited, compared with the LPS group (the inflammation model). When cultured in CM, osteoblasts cultured on Zn-incorporated TNTs showed strengthened cell proliferation, adhesion, osteogenesis-related gene expression, alkaline phosphatase activity and extracellular mineralization, compared with their TNT counterparts and the Ti group. CONCLUSION: This study suggests that the application of Zn-incorporated TNT surfaces may establish an osteogenic microenvironment and accelerate bone formation. It provided a promising strategy of Ti surface modification for a better applicable prospect.