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1.
BMC Oral Health ; 24(1): 1167, 2024 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-39354443

RESUMO

BACKGROUND: In edentulous jaws, factors such as the number of implants, cross-arch distribution, and the angle among implants may affect the accuracy of the implant impression. This study explored factors influencing the accuracy of implant abutment-level digital impressions using stereophotogrammetry in edentulous jaws. METHODS: Two standard all-on-4 and all-on-6 models of edentulous jaws were constructed in vitro. In the stereophotogrammetry group (PG), the implant digital impression was made using stereophotogrammetry and saved as an STL file. In the conventional group (CNV), the impression was made using the open-tray splint impression technique. An electronic and optical 3D measuring instrument was used to scan the standard model and the conventional plaster model to obtain STL files. Using 3D data processing software (GOM Inspect Pro, Zeiss), the distance and angle between the abutments in the CNV impression and the PG impression were measured and compared with the data from the standard model. RESULTS: The distance deviation in the PG and the CNV was 145 ± 196 µm and 96 ± 150 µm, respectively, with a significant difference (P < 0.001). The angle deviation in the PG and the CNV was 0.82 ± 0.88° and 0.74 ± 0.62°, respectively, with no significant difference (P = 0.267). In the PG, the distance deviation was negatively correlated with the distance between implants (r = -0.145, P = 0.028) and positively correlated with the angle of implants (r = 0.205, P = 0.002). The angle deviation was negatively correlated with the distance between implants (r = -0.198, P = 0.003) and positively correlated with the angle of implants (r = 0.172, P = 0.009). In the CNV, the effect of inter-implant distance on impression accuracy was also shown by Spearman correlation analysis: r = 0.347 (P < 0.001) for distance deviation and r = -0.012 (P = 0.859) for angle deviation. The effect of inter-implant angulation on impression accuracy deviation was r = -0.026 (P = 0.698) for distance deviation and r = 0.056 (P = 0.399) for angle deviation. CONCLUSIONS: The CNV method is closer to the real value of the original model. The distance between implants and the distribution angle had a weak correlation with the accuracy of digital impressions but no significant correlation with the accuracy of traditional impressions.


Assuntos
Técnica de Moldagem Odontológica , Imageamento Tridimensional , Arcada Edêntula , Fotogrametria , Fotogrametria/métodos , Projetos Piloto , Humanos , Arcada Edêntula/diagnóstico por imagem , Arcada Edêntula/patologia , Imageamento Tridimensional/métodos , Dente Suporte , Desenho Assistido por Computador , Modelos Dentários , Técnicas In Vitro , Projeto do Implante Dentário-Pivô/métodos , Implantes Dentários
2.
Small ; 19(33): e2300326, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37017497

RESUMO

Tendon adhesion is the most common outcome of tendon or tendon-to-bone healing after injury. Our group developed a hydrogel-nanoparticle sustained-release system previously to inhibit cyclooxygenases (COXs) expression and consequently prevent tendon adhesion and achieved satisfactory results. However, effective treatment of multiple tendon adhesions is always a challenge in research on the prevention of tendon adhesion. In the present study, an M2M@PLGA/COX-siRNA delivery system is successfully constructed using the cell membranes of M2 macrophages and poly (lactic-co-glycolic acid) (PLGA) nanoparticles. Targeting properties and therapeutic effects are observed in mice or rat models of flexor digitorum longus (FDL) tendon injury combined with rotator cuff injury. The results showed that the M2M@PLGA/COX-siRNA delivery system has low toxicity and remarkable targeting properties to the injured areas. Treatment with the M2M@PLGA/COX-siRNA delivery system reduced the inflammatory reaction and significantly improved tendon adhesion in both the FDL tendon and rotator cuff tissues. These findings indicate that the M2M@PLGA delivery system can provide an effective biological strategy for preventing multiple tendon adhesions.


Assuntos
Biomimética , Nanopartículas , Ratos , Camundongos , Animais , RNA Interferente Pequeno/genética , Tendões , Aderências Teciduais/patologia , Aderências Teciduais/prevenção & controle , Inflamação/patologia , Macrófagos
3.
Pharmacol Res ; 197: 106979, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37918583

RESUMO

Circular RNA (circRNA) is one of non-coding RNA with specific circular structure, which has been found to be involved in regulating a series of malignant biological behaviors in many malignant tumors. In this study, based on the IDH1 molecular typing of gliomas, we identified a significant downregulation of circRNA (circIQGAP1) expression in IDH1 mutant gliomas by high-throughput sequencing. In 79 tissue samples, we confirmed that circIQGAP1 expression was significantly downregulated in IDH1 mutant gliomas, and that low circIQGAP1 expression was positively associated with better prognosis. Knockdown of circIQGAP1 in glioma cell lines inhibited glioma cell malignancy and conversely overexpression of circIQGAP1 promoted glioma malignancy. circIQGAP1 regulated glioma cell migration, proliferation, invasion and apoptosis through miR-1256/RCAN1/Bax/Bcl-2/Caspase3 and miR-622/RCAN2/Bax/Bcl-2/Caspase3 axes. These results suggest that circIQGAP1 plays an important role in glioma development, promotes tumor growth, and is a potential therapeutic target for glioma.


Assuntos
Glioma , MicroRNAs , Humanos , RNA Circular/genética , Proteína X Associada a bcl-2 , Glioma/genética , Proteínas Proto-Oncogênicas c-bcl-2 , Fatores de Transcrição , MicroRNAs/genética , Proteínas de Ligação a DNA , Proteínas Musculares
4.
J Nanobiotechnology ; 20(1): 434, 2022 Oct 04.
Artigo em Inglês | MEDLINE | ID: mdl-36195952

RESUMO

The incidence of myocardial ischaemia‒reperfusion injury (MIRI) is increasing every year, and there is an urgent need to develop new therapeutic approaches. Nrf2 is thought to play a protective role during MIRI and it is regulated by microRNAs (miRNAs). This study focused on PLGA nanoparticles camouflaged by platelet membrane vesicles (PMVs) (i.e., PMVs@PLGA complexes) carrying microRNA inhibitors, which regulate Nrf2 and can play a therapeutic role in the MIRI process. In vitro and in vivo characterization showed that PMVs@PLGA has excellent transfection efficiency, low toxicity and good targeting. MicroRNAs that effectively regulate Nrf2 were identified, and then PMVs@PLGA-miRNA complexes were prepared and used for in vitro and in vivo treatment. PMVs@PLGA-miRNA complexes can effectively target the delivery of inhibitors to cardiomyocytes. Our results suggest that PMVs@PLGA complexes are a novel delivery system and a novel biological approach to the treatment of MIRI.


Assuntos
MicroRNAs , Traumatismo por Reperfusão Miocárdica , Nanopartículas , Plaquetas , Humanos , MicroRNAs/genética , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Fator 2 Relacionado a NF-E2
5.
Nano Lett ; 21(7): 3007-3015, 2021 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-33797927

RESUMO

Brain injuries are devastating central nervous system diseases, resulting in cognitive, motor, and sensory dysfunctions. However, clinical therapeutic options are still limited for brain injuries, indicating an urgent need to investigate new therapies. Furthermore, the efficient delivery of therapeutics across the blood-brain barrier (BBB) to the brain is a serious problem. In this study, a facile strategy of dual site-selective functionalized (DSSF) poly(ß-amino esters) was developed using bio-orthogonal chemistry for promoting brain nerve regeneration. Fluorescence colocalization studies demonstrated that these proton-sponge DSSF poly(ß-amino esters) targeted mitochondria through electrostatic interactions. More importantly, this delivery system could effectively accumulate in the injured brain sites and accelerate the recovery of the injured brain. Finally, this DSSF poly(ß-amino esters) platform may provide a new methodology for the construction of dual regioselective carriers in protein/peptide delivery and tissue engineering.


Assuntos
Ésteres , Regeneração Nervosa , Barreira Hematoencefálica , Encéfalo , Engenharia Tecidual
6.
BMC Oral Health ; 22(1): 34, 2022 02 11.
Artigo em Inglês | MEDLINE | ID: mdl-35148735

RESUMO

BACKGROUND: An interdisciplinary treatment simulation and smile design before a complex esthetic rehabilitation is important for clinicians' decision-making and patient motivation. Meanwhile, intervention and interaction are necessary for dental specialists in these complex rehabilitations. However, it is difficult to visualize an interdisciplinary treatment plan by using the conventional method, especially when orthognathic surgery is involved, thus hindering communication between dental specialists. This research aims to establish a 3D digital workflow of interdisciplinary treatment simulation to solve this problem. METHODS: An interdisciplinary 3D digital workflow of simulated treatment plan for complex esthetic rehabilitation was established. Eleven patients were enrolled and illustrated with their treatment plans using 3D treatment simulation, as well as 2D digital smile design (DSD) plus wax-up. Visual analogue scales (VAS) were used to rate the intuitiveness, understanding, and satisfaction or help between the two methods by patients and dental specialists. RESULTS: According to the ratings from the patients, 3D treatment simulation showed obvious advantages in the aspects of intuitiveness (9.7 ± 0.5 vs 6.4 ± 1.4) and treatment understanding (9.1 ± 0.8 vs 6.6 ± 1.5), and the satisfaction rates were also higher (9.0 ± 0.6 vs 7.1 ± 1.8). Dental specialists regarded the 3D digital plans as more intuitive (8.9 ± 0.8 vs 5.9 ± 1.0) and useful to understand the plans from the other specialists (8.9 ± 0.7 vs 6.1 ± 1.0) and helpful to their own treatment plans (8.7 ± 0.9 vs 5.9 ± 1.4). CONCLUSIONS: The interdisciplinary 3D digital treatment simulation helps both patients and dental specialists to improve treatment understanding, and facilitates dental specialists for decision-making before complex esthetic rehabilitation. TRIAL REGISTRATION: This study was registered in the National Clinical Trials Registry under the identification number MR-11-20-002862. This is an observational study in which we did not assign the intervention.


Assuntos
Estética Dentária , Sorriso , Simulação por Computador , Desenho Assistido por Computador , Humanos , Fluxo de Trabalho
7.
Antonie Van Leeuwenhoek ; 114(4): 479-486, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33646472

RESUMO

A short-rod-shaped, non-spore-forming endophytic actinobacterium, was isolated from a surface-sterilized leaf of Acrostichum aureum in Fangchenggang, Guangxi Zhuang Autonomous Region, China, designated strain CBS4Y-1T and examined by a polyphasic approach to determine its taxonomic position. This actinobacterium was Gram-staining-positive and aerobic. Substrate mycelia and aerial mycelia were not observed, and no diffusible pigments were observed on the media tested. Strain CBS4Y-1T grew optimally with 0-1.0% (w/v) NaCl at 30 °C, pH 7.0-8.0. Comparative analysis of 16S rRNA genes showed that strain CBS4Y-1T shared the highest 16S rRNA gene sequence similarities with Nocardioides marinus CL-DD14T (96.7%) and Nocardioides terrae BX5-10T (96.7%). Phylogenetic analyses based on 16S rRNA gene sequence and phylogenomic analysis based on core proteomes alignment revealed that strain CBS4Y-1T belonged to the genus Nocardioides and formed a distinct cluster within the genus Nocardioides. The DNA G + C content of strain CBS4Y-1T was 71.1 mol%. The cell-wall peptidoglycan contained LL-diaminopimelic acid and MK-8(H4) was the predominant menaquinone. Phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), phosphatidylinositol (PI) were detected in the polar lipid extracts. The major fatty acids were iso-C16:0, C18:1ω9c and iso-C17:0. On the basis of phylogenetic analysis, phenotypic and chemotaxonomic characteristics, strain CBS4Y-1T represents a novel species of the genus Nocardioides, for which the name Nocardioides acrostichi sp. nov. is proposed. The type strain is CBS4Y-1T (= KCTC 49238T = CGMCC 4.7548T).


Assuntos
Nocardioides , Fosfolipídeos , Técnicas de Tipagem Bacteriana , Composição de Bases , China , DNA Bacteriano/genética , Ácidos Graxos , Filogenia , Folhas de Planta , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Vitamina K 2
8.
PLoS Genet ; 14(2): e1007245, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29474354

RESUMO

Sex determination is a hierarchically-regulated process with high diversity in different organisms including insects. The W chromosome-derived Fem piRNA has been identified as the primary sex determination factor in the lepidopteran insect, Bombyx mori, revealing a distinctive piRNA-mediated sex determination pathway. However, the comprehensive mechanism of silkworm sex determination is still poorly understood. We show here that the silkworm PIWI protein BmSiwi, but not BmAgo3, is essential for silkworm sex determination. CRISPR/Cas9-mediated depletion of BmSiwi results in developmental arrest in oogenesis and partial female sexual reversal, while BmAgo3 depletion only affects oogenesis. We identify three histone methyltransferases (HMTs) that are significantly down-regulated in BmSiwi mutant moths. Disruption one of these, BmAsh2, causes dysregulation of piRNAs and transposable elements (TEs), supporting a role for it in the piRNA signaling pathway. More importantly, we find that BmAsh2 mutagenesis results in oogenesis arrest and partial female-to-male sexual reversal as well as dysregulation of the sex determination genes, Bmdsx and BmMasc. Mutagenesis of other two HMTs, BmSETD2 and BmEggless, does not affect piRNA-mediated sex determination. Histological analysis and immunoprecipitation results support a functional interaction between the BmAsh2 and BmSiwi proteins. Our data provide the first evidence that the HMT, BmAsh2, plays key roles in silkworm piRNA-mediated sex determination.


Assuntos
Bombyx/genética , Histona-Lisina N-Metiltransferase/fisiologia , RNA Interferente Pequeno/fisiologia , Processos de Determinação Sexual/genética , Animais , Animais Geneticamente Modificados , Bombyx/enzimologia , Embrião não Mamífero , Feminino , Gônadas/embriologia , Histona Metiltransferases , Histona-Lisina N-Metiltransferase/genética , Proteínas de Insetos/genética , Masculino , RNA Interferente Pequeno/genética
9.
Mol Cancer ; 19(1): 53, 2020 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-32164750

RESUMO

BACKGROUND: The epigenetic regulation of immune response has been demonstrated in recent studies. Nonetheless, potential roles of RNA N6-methyladenosine (m6A) modification in tumor microenvironment (TME) cell infiltration remain unknown. METHODS: We comprehensively evaluated the m6A modification patterns of 1938 gastric cancer samples based on 21 m6A regulators, and systematically correlated these modification patterns with TME cell-infiltrating characteristics. The m6Ascore was constructed to quantify m6A modification patterns of individual tumors using principal component analysis algorithms. RESULTS: Three distinct m6A modification patterns were determined. The TME cell-infiltrating characteristics under these three patterns were highly consistent with the three immune phenotypes of tumors including immune-excluded, immune-inflamed and immune-desert phenotypes. We demonstrated the evaluation of m6A modification patterns within individual tumors could predict stages of tumor inflammation, subtypes, TME stromal activity, genetic variation, and patient prognosis. Low m6Ascore, characterized by increased mutation burden and activation of immunity, indicated an inflamed TME phenotype, with 69.4% 5-year survival. Activation of stroma and lack of effective immune infiltration were observed in the high m6Ascore subtype, indicating a non-inflamed and immune-exclusion TME phenotype, with poorer survival. Low m6Ascore was also linked to increased neoantigen load and enhanced response to anti-PD-1/L1 immunotherapy. Two immunotherapy cohorts confirmed patients with lower m6Ascore demonstrated significant therapeutic advantages and clinical benefits. CONCLUSIONS: This work revealed the m6A modification played a nonnegligible role in formation of TME diversity and complexity. Evaluating the m6A modification pattern of individual tumor will contribute to enhancing our cognition of TME infiltration characterization and guiding more effective immunotherapy strategies.


Assuntos
Adenosina/análogos & derivados , Linfócitos T CD8-Positivos/imunologia , Metilação de DNA , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Gástricas/imunologia , Microambiente Tumoral/imunologia , Adenosina/química , Apoptose , Biomarcadores Tumorais/genética , Proliferação de Células , Humanos , Prognóstico , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Taxa de Sobrevida , Células Tumorais Cultivadas
10.
Mol Ther ; 27(9): 1534-1546, 2019 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-31278034

RESUMO

How to accelerate tendon healing remains a clinical challenge. In this study, a suture carrying nanoparticle/pEGFP-basic fibroblast growth factor (bFGF) and pEGFP-vascular endothelial growth factor A (VEGFA) complexes was developed to transfer the growth factor genes into injured tendon tissues to promote healing. Polydopamine-modified sutures can uniformly and tightly absorb nanoparticle/plasmid complexes. After tendon tissues were sutured, the nanoparticle/plasmid complexes still existed on the suture surface. Further, we found that the nanoparticle/plasmid complexes delivered into tendon tissues could diffuse from sutures to tendon tissues and effectively transfect genes into tendon cells, significantly increasing the expression of growth factors in tendon tissues. Finally, biomechanical tests showed that nanoparticle/pEGFP-bFGF and pEGFP-VEGFA complex-coated sutures could significantly increase the ultimate strengths of repaired tendons, especially at 4 weeks after operation. Two kinds of nanoparticle/plasmid complex-coated sutures significantly increased flexor tendon healing strength by 3.7 times for Ethilon and 5.8 times for PDS II, respectively, compared with the corresponding unmodified sutures. In the flexor tendon injury model, at 6 weeks after surgery, compared with the control suture, the nanoparticle/plasmid complex-coated sutures can significantly increase the gliding excursions of the tendon and inhibit the formation of adhesion. These results indicate that this nanoparticle/plasmid complex-coated suture is a promising tool for the treatment of injured tendons.


Assuntos
Materiais Revestidos Biocompatíveis , Técnicas de Transferência de Genes , Nanopartículas , Suturas , Traumatismos dos Tendões/genética , Traumatismos dos Tendões/terapia , Transgenes , Cicatrização , Animais , Sobrevivência Celular , Materiais Revestidos Biocompatíveis/química , Modelos Animais de Doenças , Liberação Controlada de Fármacos , Expressão Gênica , Terapia Genética , Cinética , Nanopartículas/química , Nanopartículas/ultraestrutura , Plasmídeos/genética , Transgenes/genética , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Cicatrização/genética
11.
J Cell Biochem ; 120(9): 15106-15118, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31020692

RESUMO

Overall survival of patients with low-grade glioma (LGG) has shown no significant improvement over the past 30 years, with survival averaging approximately 7 years. This study aimed to identify novel promising biomarkers of LGG and reveal its potential molecular mechanisms by integrated bioinformatics analysis. The microarray datasets of GSE68848 and GSE4290 were selected from GEO database for integrated analysis. In total, 293 overlapping differentially expressed genes (DEGs) were detected using the limma package. One hundred and eighty-eight nodes with 603 interactions were obtained from the establishment of protein-protein interaction (PPI) network. Functional and signaling pathway enriched were significantly correlated with the synapse and calcium signaling pathway, respectively. Module analysis revealed eight hub genes with high connectivity, which included CHRM1, DLG2, GABRD, GRIN1, HTR2A, KCNJ3, KCNJ9, and NUSAP1, and they were markedly correlated with patients' prognosis. The mining of the Gene Expression Profiling Interactive Analysis database and qPCR further confirmed the abnormal expression of these key genes with their prognostic value in LGG. We eventually predicted the 20 most vital small molecule drugs, which potentially reverse the carcinogenic state of LGG, as per the CMap (connectivity map) database and these DEGs, and MS-275 (enrichment score = -0.939) was considered as the most promising small molecule to treat LGG. In conclusion, our study provided eight reliable novel molecular biomarkers for diagnosis, prognosis prediction, and treatment targets for LGG. These conclusions will contribute to a better comprehension of molecular mechanisms fundamental to LGG occurrence and progression, and providing new insights for future development of genomic individualized treatment in LGG.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Biologia Computacional , Glioma/tratamento farmacológico , Glioma/genética , Sequenciamento de Nucleotídeos em Larga Escala , Bibliotecas de Moléculas Pequenas/uso terapêutico , Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/patologia , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Glioma/patologia , Humanos , Gradação de Tumores , Mapas de Interação de Proteínas/genética , Bibliotecas de Moléculas Pequenas/farmacologia , Análise de Sobrevida
13.
Mol Pain ; 13: 1744806916688901, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-28326931

RESUMO

Background Cancer-induced pain (CIP) is one of the most severe types of chronic pain with which clinical treatment remains challenging and the involved mechanisms are largely unknown. Suppressor of cytokine signaling 3 (SOCS3) is an important intracellular protein and provides a classical negative feedback loop, thus involving in a wide variety of processes including inflammation and nociception. However, the role of SOCS3 pathway in CIP is poorly understood. The present study was designed to investigate the role of SOCS3 in dorsal root ganglion (DRG) in the development of CIP. Method CIP was established by injection of Walker 256 mammary gland tumor cells into the rat tibia canal. Whole-cell patch clamping and Western blotting were performed. Results Following the development of bone cancer, SOCS3 expression was significantly downregulated in rat DRGs at L2-L5 segments. Overexpression of SOCS3, using lentiviral-mediated production of SOCS3 at spinal cord level, drastically attenuated mechanical allodynia and body weight-bearing difference, but not thermal hyperalgesia in bone cancer rats. In addition, overexpression of SOCS3 reversed the hyperexcitability of DRG neurons innervating the tibia, and reduced abnormal expression of toll-like receptors 4 in the DRGs. Conclusions These results suggest that SOCS3 might be a key molecular involved in the development of complicated cancer pain and that overexpression of SOCS3 might be an important strategy for treatment for mechanical allodynia associated with bone cancer.


Assuntos
Dor do Câncer/terapia , Citocinas/metabolismo , Gânglios Espinais/fisiologia , Terapia Genética/métodos , Hiperalgesia/etiologia , Proteína 3 Supressora da Sinalização de Citocinas/metabolismo , Animais , Dor do Câncer/fisiopatologia , Células Cultivadas , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Feminino , Gânglios Espinais/citologia , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/genética , Limiar da Dor/fisiologia , Técnicas de Patch-Clamp , Ratos , Ratos Sprague-Dawley , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/fisiologia , Estatísticas não Paramétricas , Proteína 3 Supressora da Sinalização de Citocinas/genética , Receptor 3 Toll-Like/metabolismo , Suporte de Carga/fisiologia
14.
Mol Pain ; 11: 15, 2015 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-25885215

RESUMO

Lumbar disc herniation (LDH) is a major cause of discogenic low back pain and sciatica, but the underlying mechanisms remain largely unknown. Hydrogen sulfide (H2S) is becoming recognized for its involvement in a wide variety of processes including inflammation and nociception. The present study was designed to investigate the roles of the H2S signaling pathway in the regulation of expression and function of purinergic receptors (P2XRs) in dorsal root ganglion (DRG) neurons from rats with LDH. LDH was induced by implantation of autologous nucleus pulposus (NP), harvested from rat tail, in lumbar 5 and 6 spinal nerve roots. Implantation of autologous NP induced persistent pain hypersensitivity, which was partially reversed by an intrathecal injection of A317491, a potent inhibitor of P2X3Rs and P2X2/3Rs. The NP induced persistent pain hypersensitivity was associated with the increased expression of P2X3Rs, but not P2X1Rs and P2X2Rs, receptors in L5-6 DRGs. NP implantation also produced a 2-fold increase in ATP-induced intracellular calcium signals in DRG neurons when compared to those of controls (P < 0.05). Interestingly, NP implantation significantly enhanced expression of the endogenous hydrogen sulfide producing enzyme, cystathionine-ß-synthetase (CBS). Systematic administration of O-(Carboxymethyl) hydroxylamine hemihydrochloride (AOAA), an inhibitor of CBS, suppressed the upregulation of P2X3R expression and the potentiation of ATP-induced intracellular calcium signals in DRG neurons (P < 0.05). Intrathecal injection of AOAA markedly attenuated NP induced- persistent pain hypersensitivity. Our results suggest that sensitization of P2X3Rs, which is likely mediated by CBS-H2S signaling in primary sensory neurons, contributes to discogenic pain. Targeting CBS/H2S-P2X3R signaling may represent a potential treatment for neuropathic pain caused by LDH.


Assuntos
Cistationina beta-Sintase/metabolismo , Hipersensibilidade/metabolismo , Deslocamento do Disco Intervertebral/metabolismo , Neuralgia/metabolismo , Receptores Purinérgicos P2X3/metabolismo , Animais , Modelos Animais de Doenças , Gânglios Espinais/metabolismo , Hiperalgesia/fisiopatologia , Masculino , Ratos Sprague-Dawley , Raízes Nervosas Espinhais/metabolismo
15.
Proc Biol Sci ; 282(1809): 20150513, 2015 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-26041352

RESUMO

Metamorphosis in insects includes a series of programmed tissue histolysis and remolding processes that are controlled by two major classes of hormones, juvenile hormones and ecdysteroids. Precise pulses of ecdysteroids (the most active ecdysteroid is 20-hydroxyecdysone, 20E), are regulated by both biosynthesis and metabolism. In this study, we show that ecdysone oxidase (EO), a 20E inactivation enzyme, expresses predominantly in the midgut during the early pupal stage in the lepidopteran model insect, Bombyx mori. Depletion of BmEO using the transgenic CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/RNA-guided Cas9 nucleases) system extended the duration of the final instar larval stage. Ubiquitous transgenic overexpression of BmEO using the Gal4/UAS system induced lethality during the larval-pupal transition. When BmEO was specifically overexpressed in the middle silk gland (MSG), degeneration of MSG at the onset of metamorphosis was blocked. Transmission electron microscope and LysoTracker analyses showed that the autophagy pathway in MSG is inhibited by BmEO ectopic expression. Furthermore, RNA-seq analysis revealed that the genes involved in autophagic cell death and the mTOR signal pathway are affected by overexpression of BmEO. Taken together, BmEO functional studies reported here provide insights into ecdysone regulation of tissue degeneration during metamorphosis.


Assuntos
3-Hidroxiesteroide Desidrogenases/genética , Bombyx/enzimologia , Bombyx/genética , Expressão Ectópica do Gene , Proteínas de Insetos/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/crescimento & desenvolvimento , Bombyx/crescimento & desenvolvimento , Bombyx/ultraestrutura , Glândulas Exócrinas/metabolismo , Glândulas Exócrinas/ultraestrutura , Proteínas de Insetos/metabolismo , Larva/enzimologia , Larva/genética , Larva/crescimento & desenvolvimento , Larva/ultraestrutura , Metamorfose Biológica , Microscopia Eletrônica de Transmissão , Pupa/enzimologia , Pupa/genética , Pupa/crescimento & desenvolvimento , Pupa/ultraestrutura , Seda/metabolismo
16.
J Neurosci ; 33(21): 9028-38, 2013 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-23699514

RESUMO

Patients with long-standing diabetes frequently demonstrate gastric hypersensitivity with an unknown mechanism. The present study was designed to investigate roles for nuclear factor-κB (NF-κB) and the endogenous H2S-producing enzyme cystathionine-ß-synthetase (CBS) signaling pathways by examining cbs gene methylation status in adult rats with diabetes. Intraperitoneal injection of streptozotocin (STZ) produced gastric hypersensitivity in female rats in response to gastric balloon distention. Treatment with the CBS inhibitor aminooxyacetic acid significantly attenuated STZ-induced gastric hypersensitivity in a dose-dependent fashion. Aminooxyacetic acid treatment also reversed hyperexcitability of gastric-specific dorsal root ganglion (DRG) neurons labeled by the dye DiI in diabetic rats. Conversely, the H2S donor NaHS enhanced neuronal excitability of gastric DRG neurons. Expression of CBS and p65 were markedly enhanced in gastric DRGs in diabetic rats. Blockade of NF-κB signaling using pyrrolidine dithiocarbamate reversed the upregulation of CBS expression. Interestingly, STZ treatment led to a significant demethylation of CpG islands in the cbs gene promoter region, as determined by methylation-specific PCR and bisulfite sequencing. STZ treatment also remarkably downregulated the expression of DNA methyltransferase 3a and 3b. More importantly, STZ treatment significantly enhanced the ability of cbs to bind DNA at the p65 consensus site, as shown by chromatin immunoprecipitation assays. Our findings suggest that upregulation of cbs expression is attributed to cbs promoter DNA demethylation and p65 activation and that the enhanced interaction of the cbs gene and p65 contributes to gastric hypersensitivity in diabetes. This finding may guide the development and evaluation of new treatment modalities for patients with diabetic gastric hypersensitivity.


Assuntos
Cistationina beta-Sintase/metabolismo , Diabetes Mellitus Experimental/complicações , Hipersensibilidade , NF-kappa B/metabolismo , Gastropatias/etiologia , Aminoácidos , Análise de Variância , Animais , Área Sob a Curva , Estudos de Casos e Controles , Imunoprecipitação da Cromatina , Ilhas de CpG/efeitos dos fármacos , Ilhas de CpG/fisiologia , Cistationina beta-Sintase/genética , DNA (Citosina-5-)-Metiltransferases/genética , DNA (Citosina-5-)-Metiltransferases/metabolismo , DNA Metiltransferase 3A , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Eletromiografia , Inibidores Enzimáticos/farmacologia , Feminino , Gânglios Espinais/patologia , Hipersensibilidade/tratamento farmacológico , Hipersensibilidade/etiologia , Potenciais da Membrana/efeitos dos fármacos , Metilação/efeitos dos fármacos , Proteínas de Neoplasias/metabolismo , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Ácido Oxâmico/uso terapêutico , Técnicas de Patch-Clamp , Ratos , Ratos Sprague-Dawley , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/fisiologia , Gastropatias/tratamento farmacológico , Sulfitos/farmacologia , Regulação para Cima/efeitos dos fármacos , DNA Metiltransferase 3B
17.
Wound Repair Regen ; 22(1): 111-8, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24393159

RESUMO

In this study of a tendon injury model, we investigated how injection of a vector incorporating one growth factor gene changes expression levels of multiple growth factor genes in the healing process. The flexor tendon of chicken toes was completely cut and repaired surgically. The tendons in the experimental arm were injected with an adeno-associated virus-2 vector incorporating basic fibroblast growth-factor gene, whereas the tendons in the control arm were not injected or injected with sham vectors. Using real-time polymerase chain reaction, we found that, within the tendon healing period, a set of growth factor genes-transforming growth factor-ß1, vascular endothelial growth factor, and connective tissue growth factor-were significantly up-regulated. Expression of the platelet-derived growth factor-B gene was not changed, and the insulin-like growth factor was down-regulated. A tendon marker gene, scleraxis, was significantly up-regulated in the period. Our study revealed an intriguing finding that introduction of one growth factor gene in the healing tendon modulated expression of multiple growth factor genes. We believe this study may have significant implications in determining the approach of gene therapy, and the findings substantiate that gene therapy using a single growth factor could affect multiple growth factors.


Assuntos
Dependovirus/metabolismo , Fator 2 de Crescimento de Fibroblastos/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Traumatismos dos Tendões/patologia , Cicatrização , Animais , Proteínas Aviárias/genética , Proteínas Aviárias/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Galinhas , Fator de Crescimento do Tecido Conjuntivo/genética , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Regulação para Baixo , Técnicas de Transferência de Genes , Terapia Genética/métodos , Vetores Genéticos , Fator de Crescimento Insulin-Like I/genética , Fator de Crescimento Insulin-Like I/metabolismo , Proteínas Proto-Oncogênicas c-sis/genética , Proteínas Proto-Oncogênicas c-sis/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Dedos do Pé , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Regulação para Cima , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismo , Cicatrização/genética
18.
Arch Insect Biochem Physiol ; 85(3): 127-36, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24478036

RESUMO

The PAR-domain protein 1 (PDP1) is essential for locomotor activity of insects. However, its functions in insect growth and development have not been studied extensively, which prompted our hypothesis that PDP1 acts in energy metabolism. Here we report identification of TcPDP1 in the red flour beetle, Tribolium castaneum, and its functional analysis by RNAi. Treating larvae with dsTcPDP1 induced pupae developmental arrestment, accompanied by accelerated fat body degradation. dsTcPDP1 treatments in adults resulted in reduced female fecundity. Disruption of TcPDP1 expression affected the transcription of genes involved in insulin signaling transduction and mechanistic target of rapamycin (mTOR) pathway. These results support our hypothesis that TcPDP1 acts in energy metabolism in T. castaneum.


Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Metabolismo Energético/fisiologia , Proteínas de Insetos/metabolismo , Insulina/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Tribolium/metabolismo , Animais , Fatores de Transcrição de Zíper de Leucina Básica/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Insetos/genética , Insulina/genética , Larva/genética , Larva/metabolismo , Masculino , Pupa/genética , Pupa/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/genética , Tribolium/genética , Tribolium/crescimento & desenvolvimento
19.
Nat Commun ; 15(1): 2629, 2024 Mar 23.
Artigo em Inglês | MEDLINE | ID: mdl-38521791

RESUMO

DNA double-strand breaks (DSBs) are repaired by a hierarchically regulated network of pathways. Factors influencing the choice of particular repair pathways, however remain poorly characterized. Here we develop an Integrated Classification Pipeline (ICP) to decompose and categorize CRISPR/Cas9 generated mutations on genomic target sites in complex multicellular insects. The ICP outputs graphic rank ordered classifications of mutant alleles to visualize discriminating DSB repair fingerprints generated from different target sites and alternative inheritance patterns of CRISPR components. We uncover highly reproducible lineage-specific mutation fingerprints in individual organisms and a developmental progression wherein Microhomology-Mediated End-Joining (MMEJ) or Insertion events predominate during early rapid mitotic cell cycles, switching to distinct subsets of Non-Homologous End-Joining (NHEJ) alleles, and then to Homology-Directed Repair (HDR)-based gene conversion. These repair signatures enable marker-free tracking of specific mutations in dynamic populations, including NHEJ and HDR events within the same samples, for in-depth analysis of diverse gene editing events.


Assuntos
Quebras de DNA de Cadeia Dupla , Reparo do DNA , Alelos , Reparo do DNA/genética , DNA/metabolismo , Reparo do DNA por Junção de Extremidades , Mutação , Reparo de DNA por Recombinação , Sistemas CRISPR-Cas/genética
20.
Mol Pain ; 9: 4, 2013 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-23413915

RESUMO

BACKGROUND: Hydrogen sulfide (H2S), an endogenous gaseotransmitter/modulator, is becoming appreciated that it may be involved in a wide variety of processes including inflammation and nociception. However, the role and mechanism for H2S in nociceptive processing in trigeminal ganglion (TG) neuron remains unknown. The aim of this study is to investigate distribution of endogenous H2S synthesizing enzyme cystathionine-ß-synthetase (CBS) expression and role of H2S on excitability and voltage-gated potassium channels of TG neurons. METHODS: Immunofluorescence studies were carried out to determine whether CBS was co-expressed in Kv1.1 or Kv1.4-positive TG neurons. Whole cell patch clamp recordings were employed on acutely isolated TG neurons from adult male Sprague Dawley rats (6-8 week old). von Frey filaments were used to examine the pain behavioral responses in rats following injection of sodium hydrosulfide. RESULTS: In rat TG, 77.3±6.6% neurons were immunoreactive for CBS, 85.1±3.8% for Kv1.1 and 97.8±1.1% for Kv1.4. Double staining showed that all CBS labeled cells were Kv1.1 and Kv1.4 positive, but only 92.2±6.1% of Kv1.1 and 78.2±9.9% of Kv1.4 positive cells contained CBS. Application of H2S donor NaHS (250 µM) led to a significant depolarization of resting membrane potential recorded from TG neurons. NaHS application also resulted in a dramatic reduction in rheobase, hyperpolarization of action potential threshold, and a significant increase in the number of action potentials evoked at 2X and 3X rheobase stimulation. Under voltage-clamp conditions, TG neurons exhibited transient A-type (IA) and sustained outward rectifier K+ currents (IK). Application of NaHS did suppress IK density while did not change IA density of TG neurons (n=6). Furthermore, NaHS, a donor of hydrogen sulfide, produced a significant reduction in escape threshold in a dose dependent manner. CONCLUSION: These data suggest that endogenous H2S generating enzyme CBS was co-localized well with Kv1.1 and Kv1.4 in TG neurons and that H2S produces the mechanic pain and increases neuronal excitability, which might be largely mediated by suppressing IK density, thus identifying for the first time a specific molecular mechanism underlying pain and sensitization in TG.


Assuntos
Sulfeto de Hidrogênio/farmacologia , Ativação do Canal Iônico/efeitos dos fármacos , Canal de Potássio Kv1.1/metabolismo , Canal de Potássio Kv1.4/metabolismo , Potenciais da Membrana/efeitos dos fármacos , Gânglio Trigeminal/fisiopatologia , Potenciais de Ação/efeitos dos fármacos , Animais , Cistationina beta-Sintase/metabolismo , Hiperalgesia/patologia , Hiperalgesia/fisiopatologia , Masculino , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Ratos , Ratos Sprague-Dawley , Gânglio Trigeminal/efeitos dos fármacos , Gânglio Trigeminal/enzimologia , Gânglio Trigeminal/patologia
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