RESUMO
Acute kidney injury (AKI) is a major worldwide health concern that currently lacks effective medical treatments. PSMP is a damage-induced chemotactic cytokine that acts as a ligand of CCR2 and has an unknown role in AKI. We have observed a significant increase in PSMP levels in the renal tissue, urine, and plasma of patients with AKI. PSMP deficiency improved kidney function and decreased tubular damage and inflammation in AKI mouse models induced by kidney ischemia-reperfusion injury, glycerol, and cisplatin. Single-cell RNA sequencing analysis revealed that Ly6Chi or F4/80lo infiltrated macrophages (IMs) were a major group of proinflammatory macrophages with strong CCR2 expression in AKI. We observed that PSMP deficiency decreased CCR2+Ly6Chi or F4/80lo IMs and inhibited M1 polarization in the AKI mouse model. Moreover, overexpressed human PSMP in the mouse kidney could reverse the attenuation of kidney injury in a CCR2-dependent manner, and this effect could be achieved without CCL2 involvement. Extracellular PSMP played a crucial role, and treatment with a PSMP-neutralizing antibody significantly reduced kidney injury in vivo. Therefore, PSMP might be a therapeutic target for AKI, and its antibody is a promising therapeutic drug for the treatment of AKI.
Assuntos
Injúria Renal Aguda , Modelos Animais de Doenças , Macrófagos , Receptores CCR2 , Animais , Humanos , Masculino , Camundongos , Injúria Renal Aguda/metabolismo , Injúria Renal Aguda/patologia , Rim/metabolismo , Rim/patologia , Macrófagos/metabolismo , Camundongos Knockout , Receptores CCR2/metabolismo , Receptores CCR2/genética , Traumatismo por Reperfusão/metabolismo , Proteínas de NeoplasiasRESUMO
AIM: CCR2 (C-C chemokine receptor type 2) plays a crucial role in inflammatory and bone metabolic diseases; however, its role in peri-implantitis remains unclear. This study aimed to explore whether CCR2 contributes to peri-implantitis and the treatment effects of cenicriviroc (CVC) on peri-implant inflammation and bone resorption. MATERIALS AND METHODS: The expression of CCR2 was studied using clinical tissue analysis and an in vivo peri-implantitis model. The role of CCR2 in promoting inflammation and bone resorption in peri-implantitis was evaluated in Ccr2-/- mice and wild-type mice. The effect of CVC on peri-implantitis was evaluated using systemic and local dosage forms. RESULTS: Human peri-implantitis tissues showed increased CCR2 and CCL2 levels, which were positively correlated with bone loss around the implants. Knocking out Ccr2 in an experimental model of peri-implantitis resulted in decreased monocyte and macrophage infiltration, reduced pro-inflammatory cytokine generation and impaired osteoclast activity, leading to reduced inflammation and bone loss around the implants. Treatment with CVC ameliorated bone loss in experimental peri-implantitis. CONCLUSIONS: CCR2 may be a potential target for peri-implantitis treatment by harnessing the immune-inflammatory response to modulate the local inflammation and osteoclast activity.
Assuntos
Perda do Osso Alveolar , Reabsorção Óssea , Implantes Dentários , Peri-Implantite , Animais , Humanos , Camundongos , Perda do Osso Alveolar/tratamento farmacológico , Citocinas , Inflamação , Osteoclastos , Peri-Implantite/tratamento farmacológico , Receptores CCR2RESUMO
AIM: Our previous study revealed that the C-C motif chemokine receptor 2 (CCR2) is a promising target for periodontitis prevention and treatment. However, CCR2 is a receptor with multiple C-C motif chemokine ligands (CCLs), including CCL2, CCL7, CCL8, CCL13 and CCL16, and which of these ligands plays a key role in periodontitis remains unclear. The aim of the present study was to explore the key functional ligand of CCR2 in periodontitis and to evaluate the potential of the functional ligand as a therapeutic target for periodontitis. MATERIALS AND METHODS: The expression levels and clinical relevance of CCR2, CCL2, CCL7, CCL8, CCL13 and CCL16 were studied using human samples. The role of CCL2 in periodontitis was evaluated by using CCL2 knockout mice and overexpressing CCL2 in the periodontium. The effect of local administration of bindarit in periodontitis was evaluated by preventive and therapeutic medication in a mouse periodontitis model. Microcomputed tomography, haematoxylin and eosin staining, tartrate-resistant acid phosphatase staining, real-time quantitative polymerase chain reaction, enzyme-linked immunosorbent assay, bead-based immunoassays and flow cytometry were used for histomorphology, molecular biology and cytology analysis. RESULTS: Among different ligands of CCR2, only CCL2 was significantly up-regulated in periodontitis gingival tissues and was positively correlated with the severity of periodontitis. Mice lacking CCL2 showed milder inflammation and less bone resorption than wild-type mice, which was accompanied by a reduction in monocyte/macrophage recruitment. Adeno-associated virus-2 vectors overexpressing CCL2 in Ccl2-/- mice gingiva reversed the attenuation of periodontitis in a CCR2-dependent manner. In ligation-induced experimental periodontitis, preventive or therapeutic administration of bindarit, a CCL2 synthesis inhibitor, significantly inhibited the production of CCL2, decreased the osteoclast number and bone loss and reduced the expression levels of proinflammatory cytokines TNF-α, IL-6 and IL-1ß. CONCLUSIONS: CCL2 is a pivotal chemokine that binds to CCR2 during the progression of periodontitis, and targeting CCL2 may be a feasible option for controlling periodontitis.
Assuntos
Quimiocina CCL2 , Periodontite , Animais , Humanos , Camundongos , Quimiocina CCL2/metabolismo , Quimiocinas , Ligantes , Camundongos Endogâmicos C57BL , Periodontite/prevenção & controle , Microtomografia por Raio-XRESUMO
FAM19A5/TAFA5 is a member of the family with sequence similarity 19 with unknown function in emotional and cognitive regulation. Here, we reported that FAM19A5 was highly expressed in the embryonic and postnatal mouse brain, especially in the hippocampus. Behaviorally, genetic deletion of Fam19a5 resulted in increased depressive-like behaviors and impaired hippocampus-dependent spatial memory. These behavioral alterations were associated with the decreased expression of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors and N-methyl-D-aspartic acid receptors, as well as significantly reduced glutamate release and neuronal activity in the hippocampus. Subsequently, these changes led to the decreased density of dendritic spines. In recent years, the roles of chronic stress participating in the development of depression have become increasingly clear, but the mechanism remains to be elucidated. We found that the levels of FAM19A5 in plasma and hippocampus of chronic stress-treated mice were significantly decreased whereas overexpression of human FAM19A5 selectively in the hippocampus could attenuate chronic stress-induced depressive-like behaviors. Taken together, our results revealed for the first time that FAM19A5 plays a key role in the regulation of depression and spatial cognition in the hippocampus. Furthermore, our study provided a new mechanism for chronic stress-induced depression, and also provided a potential biomarker for the diagnosis and a new strategy for the treatment of depression.
Assuntos
Depressão , Memória Espacial , Animais , Biomarcadores , Hipocampo , Camundongos , Estresse Psicológico , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol PropiônicoRESUMO
AIM: CCR2 plays important roles in many inflammatory and bone metabolic diseases, but its specific role in periodontitis is unknown. In the present study, we aimed to explore the role of CCR2 in the progression of periodontitis and evaluate the effect of cenicriviroc (CVC) on periodontitis. MATERIALS AND METHODS: The expression of CCR2 was studied in patients with periodontitis and in ligation-induced murine model of periodontitis. The role of CCR2 in promoting inflammation and bone resorption in periodontitis was evaluated in Ccr2-/- mice and wild-type mice. The effect of CVC in the prevention and treatment of periodontitis was evaluated by systemic and local medication. Microcomputed tomography, haematoxylin and eosin staining, tartrate-resistant acid phosphatase staining, quantitative real-time polymerase chain reaction, enzyme-linked immunosorbent assay, and flow cytometry were used for histomorphology, molecular biology, and cytology analysis, respectively. RESULTS: In this study, we demonstrated that CCR2 was highly expressed in human and murine periodontitis and that CCR2 deficiency was associated with decreased inflammatory monocyte and macrophage infiltration and inflammatory mediators, osteoclast number and alveolar bone resorption. Prevention and treatment with CVC significantly reduced the severity of periodontitis, regardless of whether it was administered systemically or locally. CONCLUSIONS: CCR2 plays an important role in the development and progression of periodontitis, and CVC is a potential drug for the prevention and treatment of periodontitis.
Assuntos
Perda do Osso Alveolar , Periodontite , Perda do Osso Alveolar/tratamento farmacológico , Animais , Amarelo de Eosina-(YS)/uso terapêutico , Humanos , Imidazóis , Mediadores da Inflamação , Camundongos , Camundongos Endogâmicos C57BL , Periodontite/tratamento farmacológico , Receptores CCR2/metabolismo , Sulfóxidos , Fosfatase Ácida Resistente a Tartarato , Microtomografia por Raio-XRESUMO
BACKGROUND & AIMS: C-C motif chemokine receptor 2 (CCR2) has been recognized as a promising target for the treatment of liver fibrosis. PC3-secreted microprotein (PSMP)/microseminoprotein (MSMP) is a novel chemotactic cytokine and its receptor is CCR2. In the present study we investigated the expression and role of PSMP in liver fibrosis/cirrhosis. METHODS: PSMP expression was studied in patients with fibrosis/cirrhosis and in 3 murine models of liver fibrosis, including mice treated with carbon tetrachloride (CCl4), bile-duct ligation, or a 5-diethoxycarbonyl-1,4-dihydrocollidine diet. The role of PSMP was evaluated in Psmp-/- mice and after treatment with a PSMP antibody in wild-type mice. The direct effects of PSMP on macrophages and hepatic stellate cells were studied in vitro. RESULTS: In this study, we found that PSMP was highly expressed in fibrotic/cirrhotic tissues from patients with different etiologies of liver disease and in the 3 experimental mouse models of fibrosis. Damage-associated molecular pattern molecules HMGB-1 and IL-33 induced hepatocytes to produce PSMP. PSMP deficiency resulted in a marked amelioration of hepatic injury and fibrosis. In CCl4-induced hepatic injury, the infiltration of macrophages and CCR2+ monocytes into the liver was significantly decreased in Psmp-/- mice. Consistent with the decreased levels of intrahepatic macrophages, proinflammatory cytokines were significantly reduced. Moreover, adeno-associated virus-8 vectors successfully overexpressing human PSMP in Psmp-/- mouse livers could reverse the attenuation of liver injury and fibrosis induced by CCl4 in a CCR2-dependent manner. Treatment with a specific PSMP-neutralizing antibody, 3D5, prevented liver injury and fibrosis induced by CCl4 in mice. At the cellular level, PSMP directly promoted M1 polarization of macrophages and activation of LX-2 cells. CONCLUSION: PSMP enhances liver fibrosis through its receptor, CCR2. PSMP is a potentially attractive therapeutic target for the treatment of patients with liver fibrosis. LAY SUMMARY: Our present study identifies the essential role of the protein PSMP for the development and progression of liver fibrosis in humans and mice. PSMP promotes liver fibrosis through inflammatory macrophage infiltration, polarization and production of proinflammatory cytokines, as well as direct activation of hepatic stellate cells via its receptor CCR2. A PSMP antibody can significantly reduce liver fibrosis development in vivo. These findings indicate that PSMP is a potential therapeutic target and its antibody is a potential therapeutic agent for the treatment of liver fibrosis.
Assuntos
Carcinoma Hepatocelular/metabolismo , Cirrose Hepática Experimental/metabolismo , Neoplasias Hepáticas/metabolismo , Proteínas de Neoplasias/deficiência , Receptores CCR2/deficiência , Receptores CCR2/metabolismo , Animais , Anticorpos Neutralizantes/uso terapêutico , Tetracloreto de Carbono/efeitos adversos , Carcinoma Hepatocelular/patologia , Polaridade Celular/genética , Células Cultivadas , Citocinas/biossíntese , Vetores Genéticos , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Humanos , Cirrose Hepática Experimental/induzido quimicamente , Cirrose Hepática Experimental/prevenção & controle , Neoplasias Hepáticas/patologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/imunologia , Proteínas de Neoplasias/farmacologia , Receptores CCR2/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Resultado do Tratamento , Regulação para CimaRESUMO
Our previous study has demonstrated that recombinant yeast can induce specific immune responses in Carassius auratus and may serve as a potential carrier for oral DNA vaccines in aquaculture. In this study, we further developed an effective yeast-based oral DNA vaccine against the bacteria Aeromonas hydrophila, which was expected to provide protection from the motile aeromonad septicemia (MAS). First, two candidate antigen genes, ompG and omp48, were cloned from the Aeromonas hydrophila genome DNA. Then, relative yeast-eukaryote shuttle vectors were constructed and their expression in eukaryotes was validated. Next, crucian carps were orally administered with ompG or omp48 recombinant yeast, and the expression of the genes in the intestinal mucosa was confirmed by immunohistochemistry (IHC). The specific immune responses were further detected by Western blot and enzyme-linked immunosorbent assay (ELISA). The ELISA results showed that the production of the OVA-specific antibody in the OVA-ompG group was significantly higher than that of the OVA-omp48 group, indicating that the OVA-ompG group elicited obviously stronger immune response than OVA-omp48. Finally, the challenge experiment against Aeromonas hydrophila infection demonstrated decreased fish mortality rate after the oral administration of the OVA-ompG yeast vaccine. In conclusion, our work provided a framework for the further development of oral yeast-based fishery vaccines.
Assuntos
Aeromonas hydrophila/imunologia , Vacinas Bacterianas/uso terapêutico , Carpas , Doenças dos Peixes/prevenção & controle , Carpa Dourada , Infecções por Bactérias Gram-Negativas/veterinária , Vacinas de DNA/uso terapêutico , Animais , Vacinas Bacterianas/administração & dosagem , Doenças dos Peixes/microbiologia , Infecções por Bactérias Gram-Negativas/microbiologia , Infecções por Bactérias Gram-Negativas/prevenção & controle , Saccharomyces cerevisiae/química , Vacinas de DNA/administração & dosagemRESUMO
GW182 and argonaute 2 (AGO2) are core proteins of the RNA interference complex. GW182 is a scaffolding protein that physically associates with AGO2 and bridges its interactions with other proteins. A fundamental problem in biology is how scaffolding proteins adapt or contribute to differing functional demands within cells. Here we test the necessity for human GW182 proteins (paralogs TNRC6A, TNRC6B, and TNRC6C) for several mechanisms of small duplex RNA-mediated control of gene expression, including translational silencing by miRNAs, translational silencing by siRNAs, transcriptional silencing, transcriptional activation, and splicing. We find that GW182 is required for transcriptional activation and for the activity of miRNAs but is dispensable for the regulation of splicing, transcriptional silencing, and the action of siRNAs. AGO2, by contrast, is necessary for each of these processes. Our data suggest that GW182 does not alter AGO2 to make it active. Instead, GW182 organizes protein complexes around AGO2. Sometimes this higher level of organization is necessary, and sometimes it is not. AGO2 and GW182 offer an example for how a partnership between a scaffolding protein and a functional protein can be powerful but not obligatory.
Assuntos
Proteínas Argonautas/metabolismo , Autoantígenos/genética , Autoantígenos/metabolismo , Biossíntese de Proteínas , Splicing de RNA , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Transcrição Gênica , Proteínas Argonautas/genética , Células HeLa , Humanos , MicroRNAs , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente PequenoRESUMO
Isolation of genetically modified cells generated by designed nucleases are challenging, since they are often phenotypically indistinguishable from their parental cells. To efficiently enrich genetically modified cells, we developed two dual-reporter surrogate systems, namely NHEJ-RPG and SSA-RPG based on NHEJ and SSA repair mechanisms, respectively. Repair and enrichment efficiencies of these two systems were compared using different nucleases. In both CRISPR-Cas9- and ZFNs-induced DSB repair studies, we found that the efficiency and sensitivity of the SSA-RPG reporter with direct repeat length more than 200 bp were much higher than the NHEJ-RPG reporter. By utilizing the SSA-RPG reporter, we achieved the enrichment for indels in several endogenous loci with 6.3- to 34.8-fold of non-selected cells. Thus, the highly sensitive SSA-RPG reporter can be used for activity validation of designed nucleases and efficient enrichment of genetically modified cells. Besides, our systems offer alternative enrichment choices either by puromycin selection or FACS.
Assuntos
Separação Celular/métodos , Reparo do DNA , Engenharia Genética , Quebras de DNA de Cadeia Dupla , Citometria de Fluxo/métodos , Genes Reporter , Genoma , Células HEK293 , Humanos , Modelos GenéticosRESUMO
The Streptococcus thermophilus CRISPR3-Cas (StCas9) system has been shown to mediate DNA cleavage in its original host and in E. coli as well as in vitro. Here, we have reconstituted the StCas9 system in yeast and conducted a systematic optimization of the sgRNA structure, including the minimal length of tracrRNA, loop structure, Match II region, Bulge motif, the minimal length of guide sequence within the crRNA, tolerance of mismatches and target sequence preference. The optimal sgRNA design for the StCas9 system achieved up to 12 and 40 % targeting efficiencies in yeast and human cells, respectively. This study provides important insight into the sequence and structural requirements necessary to develop a targeted and highly efficient eukaryotic gene editing platform using CRISPR-Cas systems.
Assuntos
Sistemas CRISPR-Cas/fisiologia , Engenharia Genética/métodos , Edição de RNA/genética , RNA Mensageiro/genética , Streptococcus thermophilus/genética , Pareamento de Bases , Sistemas CRISPR-Cas/genética , Clonagem Molecular , Células HEK293 , Humanos , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Saccharomyces cerevisiaeRESUMO
Objective: This investigation aimed to determine whether and to what extent there are transcriptional differences between periodontitis and peri-implantitis in the same susceptible host. Background: As an immune-mediated inflammatory disease resulting in aggressive bone resorption around dental implants, peri-implantitis constitutes a major threat to dental implants' long-term success. Compared to periodontitis, its etiological molecular mechanism remains elusive. Currently, there are few investigations on these two diseases at the transcriptional level within the same basal environment. Methods: Ligature-induced peri-implantitis and periodontitis were generated in the same mice. Gingival tissues of healthy, periodontitis, and peri-implantitis sites from the same oral cavity were collected and used for RNA sequencing. Differentially expressed genes (DEGs) were screened between periodontitis/peri-implantitis and healthy sites. Enrichment analysis of DEGs was performed. The comprehensive immune landscape was annotated by seq-ImmuCC. Hub genes from peri-implantitis-specific DEGs were filtered using the STRING database and Cytoscape. Validation of the selected hub genes was performed on the GEO106090 dataset (gingival tissues from six periodontitis patients, six peri-implantitis patients, and six healthy controls). Results: The results indicated that peri-implantitis and periodontitis exhibited significantly distinct transcriptional signatures, with the complement and coagulation cascade pathways and osteoclast differentiation predominating in peri-implantitis mucosa. Compared with periodontitis, peri-implantitis sites exhibited elevated macrophage proportions and relatively enriched macrophage activation and bone loss. Hub genes were selected, and IL1B, CCL3, and CLEC4E were significantly highly expressed in human peri-implantitis mucosa. Conclusion: The study suggests that the interplay between macrophages and bone resorption seems to be more robust than in periodontitis. IL1B, CCL3, and CLEC4E may be considered promising therapeutic targets for peri-implantitis. These critical biological processes and identified genes may contribute to the etiology of peri-implantitis, which is unique from periodontitis. This work may make way for deeper exploration and contribute significantly to the treatment and prevention of peri-implantitis.
RESUMO
BACKGROUND: The global prevalence of inflammatory bowel disease (IBD) is increasing, and mucosal healing is the preferred treatment target of IBD. Sodium (aS,9 R)- 3-hydroxy-16,17-dimethoxy-15-oxidotricyclo[12.3.1.12,6]nonadeca-1(18),2,4,6(19),14,16-hexene-9-yl sulfate hydrate (SDH) is a novel diarylheptane compound, which is designed to treat IBD. Hence, we investigated the potent therapeutic activity of SDH against IBD and explored the underlying mechanisms, and determined if SDH is a safe and well-tolerated oral therapeutic for IBD treatment. METHODS: We characterized its therapeutic properties in vitro and in vivo using Caco-2 cell monolayer and dextran sodium sulfate (DSS)- or 2,4,6-trinitro-benzene sulfonic acid (TNBS)-induced colitis models. We conducted nonclinical toxicology and safety pharmacology research, including general toxicity, toxicokinetics, pharmacokinetics, metabolism and plasma protein binding, cardiovascular safety pharmacology, central nervous system safety pharmacology, respiratory safety pharmacology, fertility and early embryonic development toxicity, reverse mutation assay, chromosomal aberration assay and micronucleus test. RESULTS: The results showed that SDH promoted expression of tight junction proteins, and protected the integrity and permeability of the epithelial barrier in both cell and animal models. Moreover, lower doses of SDH showed the similar or better efficacy than cyclosporine A (CsA) and mesalazine in DSS- or TNBS-induced colitis animals. Furthermore, our results identified that SDH has satisfactory safety in these studies we tested. In summary, SDH restored the epithelial barrier through tight junction proteins and was expected to be a novel therapeutic agent for the treatment of IBD.
Assuntos
Colite , Doenças Inflamatórias Intestinais , Animais , Células CACO-2 , Colite/induzido quimicamente , Colite/tratamento farmacológico , Colite/metabolismo , Colo/metabolismo , Modelos Animais de Doenças , Humanos , Doenças Inflamatórias Intestinais/tratamento farmacológico , Doenças Inflamatórias Intestinais/metabolismo , Mucosa Intestinal/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas de Junções Íntimas/metabolismo , Junções Íntimas/metabolismoRESUMO
The LAMOST project, the world's largest sky survey project being implemented in China, urgently needs an automatic stars recognition and classification system. This paper presents a method for auto-recognizing the stars based on spectral feature. This method consists of three main steps: First, the integral information of spectral lines is calculated and the stellar Balmer lines are detected by using the wavelet features of spectral lines. Then, the characteristic frequency of M-type stars and the locations of absorption bands are obtained accurately through the wavelet features of absorption bands. Finally, based on the results of the former step, the emission-line stars, M-type stars and early-type stars can be recognized. The extensive experiments with real observed spectra from the SDSS DR4 show that the method can robustly recognize stellar spectra, the correct rate of the emission-line stars is as high as 97.5%, the correct rate of M-type stars is as high as 98.1% and the correct rate of early-type stars is as high as 96.8%. The error rate of the quasars and the galaxies is less than 2%. This method is designed to automatically recognize stellar spectra with relative flux and low signal-to-noise ratio, which is applicable to the LAMOST data.
RESUMO
The trinucleotide repeat containing 6 (TNRC6) family of proteins are core components of RNA interference (RNAi) and consist of three paralogs (TNRC6A, TNRC6B, and TNRC6C). The TNRC6 paralogs associate with argonaute (AGO) protein, the core RNAi factor, and bridge its interactions with other proteins. We obtained TNRC6A and TNRC6B single and double knockout cell lines to investigate how the TNRC6 paralogs contribute to RNAi. We found that TNRC6 proteins are not required for gene silencing when duplex RNAs are fully complementary. TNRC6 expression was necessary for regulation by a microRNA. TNRC6A, but not TNRC6B, expression was necessary for transcriptional activation by a duplex RNA targeting a gene promoter. By contrast, AGO2 is required for all three gene expression pathways. TNRC6A can affect the Dicer localization in cytoplasm versus the nucleus, but none of the three TNRC6 paralogs was necessary for nuclear localization of AGO2. Our data suggest that the roles of the TNRC6 paralogs differ in some details and that TNRC6 is not required for clinical therapeutic silencing mechanisms that involve fully complementary duplex RNAs.
Assuntos
Proteínas Argonautas/genética , Autoantígenos/genética , Terapia Genética/métodos , Proteínas de Ligação a RNA/genética , Proteínas Argonautas/antagonistas & inibidores , Proteínas Argonautas/uso terapêutico , Autoantígenos/uso terapêutico , Citoplasma/genética , Regulação da Expressão Gênica/genética , Inativação Gênica , Humanos , MicroRNAs/genética , Regiões Promotoras Genéticas/genética , Proteínas de Ligação a RNA/antagonistas & inibidores , Proteínas de Ligação a RNA/uso terapêutico , Repetições de Trinucleotídeos/genéticaRESUMO
The LAMOST project, the world's largest sky survey project being implemented in China, urgently needs an automatic stars recognition system. The present paper presents a method for automatic recognition of stellar spectra based on feature matching of spectral lines. This method consists of three main steps: First, the features of spectral lines! in the observed spectra are extracted using the wavelet transform. Then, the correlations between the extracted features and the feature templates of the stellar spectral lines are computed. Finally, based on the results of the former step, the stellar spectra can be recognized. The extensive experiments with real observed spectra from the SDSS DR4 show that the method can robustly recognize stellar spectra, and the correct rate of this method is as high as 96.7%. This method is designed to automatically recognize stellar spectra with relative flux and low signal-to-noise ratio, which is applicable to the LAMOST data and helps in the structure study of stars and galaxy etc.
RESUMO
Friedreich's Ataxia (FA) is an inherited neurologic disorder caused by an expanded GAA repeat within intron 1 of the frataxin (FXN) gene that reduces expression of FXN protein. Agents that increase expression of FXN have the potential to alleviate the disease. We previously reported that duplex RNAs (dsRNAs) and antisense oligonucleotides (ASOs) complementary to the GAA repeat could enhance expression of FXN protein. We now explore the potential of a diverse group of chemically modified dsRNAs and ASOs to define the breadth of repeat-targeted synthetic nucleic acids as a platform for therapeutic development for FA. ASOs and dsRNAs can activate FXN protein expression in FA patient-derived cell lines that possess varied numbers of GAA repeats. Increased FXN protein expression was achieved by ASOs incorporating diverse chemical modifications with low nanomolar potencies, suggesting substantial flexibility in choosing compounds for further chemical optimization and animal studies. Our data encourage further development of ASOs as agents to treat FA.
Assuntos
Proteínas de Ligação ao Ferro/genética , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos/genética , RNA de Cadeia Dupla/genética , RNA Mensageiro/genética , Expansão das Repetições de Trinucleotídeos , Adolescente , Adulto , Linhagem Celular , Criança , Feminino , Fibroblastos/metabolismo , Fibroblastos/patologia , Ataxia de Friedreich/genética , Ataxia de Friedreich/metabolismo , Ataxia de Friedreich/patologia , Ataxia de Friedreich/terapia , Regulação da Expressão Gênica , Terapia Genética/métodos , Humanos , Íntrons , Proteínas de Ligação ao Ferro/agonistas , Proteínas de Ligação ao Ferro/metabolismo , Masculino , Oligonucleotídeos/metabolismo , Oligonucleotídeos Antissenso/metabolismo , Cultura Primária de Células , RNA de Cadeia Dupla/metabolismo , RNA Mensageiro/agonistas , RNA Mensageiro/metabolismo , Triazóis/química , FrataxinaRESUMO
Celestial spectra should be preprocessed before automated classification to eliminate the disturbance of noise, observa-tion environment, and flux aberrance. In the present work, the authors studied the spectrum flux standardization problem. By analyzing the disturbing factors and their characteristics, the authors put forward a theoretical model for spectra flux, and corre-spondingly give several flux standardizing methods. The rationality/correctness of the model, and the satisfactory performance of the proposed methods have been obtained by the experiments over normal galaxies (NGs) and quasi-stellar object (Qso). Furthermore, the authors theoretically analyze, compare and evaluate them. In particular, this work indicated that the conventional method is worse than the proposed one. And the investigation is also particularly significant for other automatic spectrum processing study, e. g. redshift determination, effective temperature, metallic estimation, etc.
RESUMO
Precise genome editing with desired point mutations can be generated by CRISPR/Cas9-mediated homology-directed repair (HDR) and is of great significance for gene function study, gene therapy and animal breeding. However, HDR efficiency is inherently low and improvements are necessitated. Herein, we determined that the HDR efficiency could be enhanced by expressing Rad52, a gene that is involved in the homologous recombination process. Both the Rad52 co-expression and Rad52-Cas9 fusion strategies yielded approximately 3-fold increase in HDR during the surrogate reporter assays in human HEK293T cells, as well as in the genome editing assays. Moreover, the enhancement effects of the Rad52-Cas9 fusion on HDR mediated by different (plasmid, PCR and ssDNA) donor templates were confirmed. We found that the HDR efficiency could be significantly improved to about 40% by the combined usage of Rad52 and Scr7. In addition, we also applied the fusion strategy for modifying the IGF2 gene of porcine PK15 cells, which further demonstrated a 2.2-fold increase in HDR frequency. In conclusion, our data suggests that Rad52-Cas9 fusion is a good option for enhancing CRISPR/Cas9-mediated HDR, which may be of use in future studies involving precise genome editing.
Assuntos
Sistemas CRISPR-Cas/genética , Reparo do DNA/genética , Edição de Genes/métodos , Proteína Rad52 de Recombinação e Reparo de DNA/genética , Saccharomyces cerevisiae/genética , Homologia de Sequência do Ácido Nucleico , Quebras de DNA de Cadeia Dupla , Expressão Gênica , Genômica , Células HEK293 , Recombinação Homóloga , Humanos , Fator de Crescimento Insulin-Like II/genética , Mutação PuntualRESUMO
The LAMOST project, the world's largest sky survey project being implemented in China, is expected to obtain 10(5) quasar spectra. The main objective of the present article is to explore methods that can be used to estimate the redshifts of quasar spectra from LAMOST. Firstly, the features of the broad emission lines are extracted from the quasar spectra to overcome the disadvantage of low signal-to-noise ratio. Then the redshifts of quasar spectra can be estimated by using the multi-scaling feature matching. The experiment with the 15, 715 quasars from the SDSS DR2 shows that the correct rate of redshift estimated by the method is 95.13% within an error range of 0. 02. This method was designed to obtain the redshifts of quasar spectra with relative flux and a low signal-to-noise ratio, which is applicable to the LAMOST data and helps to study quasars and the large-scale structure of the universe etc.