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1.
Int J Mol Sci ; 22(17)2021 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-34502537

RESUMO

Macrophages emerge in the milieu around innervated neurons after nerve injuries. Following nerve injury, autophagy is induced in macrophages and affects the regulation of inflammatory responses. It is closely linked to neuroinflammation, while the immunosuppressive drug tacrolimus (FK506) enhances nerve regeneration following nerve crush injury and nerve allotransplantation with additional neuroprotective and neurotrophic functions. The combined use of FK506 and adipose-derived stem cells (ADSCs) was employed in cell therapy for organ transplantation and vascularized composite allotransplantation. This study aimed to investigate the topical application of exosomes secreted by ADSCs following FK506 treatment (ADSC-F-exo) to the injured nerve in a mouse model of sciatic nerve crush injury. Furthermore, isobaric tags for relative and absolute quantitation (iTRAQ) were used to profile the potential exosomal proteins involved in autophagy. Immunohistochemical analysis revealed that nerve crush injuries significantly induced autophagy in the dorsal root ganglia and dorsal horn of the spinal segments. Locally applied ADSC-F-exo significantly reduced autophagy of macrophages in the spinal segments after nerve crush injury. Proteomic analysis showed that of the 22 abundant exosomal proteins detected in ADSC-F-exo, heat shock protein family A member 8 (HSPA8) and eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) are involved in exosome-mediated autophagy reduction.

2.
Int J Mol Sci ; 22(16)2021 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-34445251

RESUMO

Exosomes secreted by adipose-derived stem cells (ADSC-exo) reportedly improve nerve regeneration after peripheral nerve injury. Herein, we investigated whether pretreatment of ADSCs with FK506, an immunosuppressive drug that enhances nerve regeneration, could secret exosomes (ADSC-F-exo) that further augment nerve regeneration. Designed exosomes were topically applied to injured nerve in a mouse model of sciatic nerve crush injury to assess the nerve regeneration efficacy. Outcomes were determined by histomorphometric analysis of semi-thin nerve sections stained with toluidine blue, mouse neurogenesis PCR array, and neurotrophin expression in distal nerve segments. Isobaric tags for relative and absolute quantitation (iTRAQ) were used to profile potential exosomal proteins facilitating nerve regeneration. We observed that locally applied ADSC-exo and ADSC-F-exo significantly enhanced nerve regeneration after nerve crush injury. Pretreatment of ADSCs with FK506 failed to produce exosomes possessing more potent molecules for enhanced nerve regeneration. Proteomic analysis revealed that of 192 exosomal proteins detected in both ADSC-exo and ADSC-F-exo, histone deacetylases (HDACs), amyloid-beta A4 protein (APP), and integrin beta-1 (ITGB1) might be involved in enhancing nerve regeneration.


Assuntos
Tecido Adiposo/metabolismo , Exossomos , Regeneração Nervosa , Traumatismos dos Nervos Periféricos/terapia , Nervos Periféricos/fisiologia , Células-Tronco/metabolismo , Tacrolimo/farmacologia , Animais , Exossomos/metabolismo , Exossomos/transplante , Camundongos , Traumatismos dos Nervos Periféricos/metabolismo
3.
Int J Mol Sci ; 22(16)2021 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-34445582

RESUMO

Exosomes secreted by adipose-derived stem cells (ADSCs) enhance angiogenesis and wound healing. However, in clinical settings, wounds may be infected by various bacteria or pathogens. We investigated whether human ADSCs stimulated with lipopolysaccharide (LPS) secrete exosomes (ADSC-LPS-exo) that augment the angiogenesis of human umbilical vein endothelial cells (HUVECs). ExoQuick-TC exosome precipitation solution was used to purify exosomes from human ADSC culture media in the presence or absence of 1 µg/mL LPS treatment for 24 h. The uptake of ADSC-LPS-exo significantly induced the activation of cAMP response element binding protein (CREB), activating protein 1 (AP-1), and nuclear factor-κB (NF-κB) signaling pathways and increased the migration of and tube formation in HUVECs. RNA interference with CREB, AP-1, or NF-κB1 significantly reduced the migration of and tube formation in HUVECs treated with ADSC-LPS-exo. An experiment with an antibody array for 25 angiogenesis-related proteins revealed that only interleukin-8 expression was significantly upregulated in HUVECs treated with ADSC-LPS-exo. In addition, proteomic analysis revealed that eukaryotic translation initiation factor 4E, amyloid beta A4 protein, integrin beta-1, and ras-related C3 botulinum toxin substrate 1 may be potential candidates involved in ADSC-LPS-exo-mediated enhanced angiogenesis.


Assuntos
Movimento Celular , Exossomos/fisiologia , Células Endoteliais da Veia Umbilical Humana/fisiologia , Lipopolissacarídeos/farmacologia , Células-Tronco Mesenquimais/fisiologia , Neovascularização Fisiológica , Proliferação de Células , Células Cultivadas , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Transdução de Sinais
4.
J Cell Mol Med ; 25(15): 7436-7450, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34235869

RESUMO

Exosomes are secreted into the extracellular space by most cell types and contain various molecular constituents, which play roles in many biological processes. Adipose-derived mesenchymal stem cells (ADSCs) can differentiate into a variety of cell types and secrete a series of paracrine factors through exosomes. ADSC-derived exosomes have shown diagnostic and therapeutic potential in many clinical diseases. The molecular components are critical for their mechanisms. Several methods have been developed for exosome purification, including ultracentrifugation, ultrafiltration, density gradient purification, size-based isolation, polymer precipitation and immuno-affinity purification. Thus, we employed four methods to isolate exosomes from the hADSC culture medium, including ultracentrifugation, size exclusion chromatography, ExoQuick-TC precipitation and ExoQuick-TC ULTRA isolation. Following exosome isolation, we performed quantitative proteomic analysis of the exosome proteins using isobaric tags for relative and absolute quantification (iTRAQ) labelling, combined with 2D-LC-MS/MS. There were 599 universal and 138 stably expressed proteins in hADSC-derived exosomes. We proved that these proteins were potential hADSC-derived exosomes markers, including CD109, CD166, HSPA4, TRAP1, RAB2A, RAB11B and RAB14. From the quantitative proteomic analysis, we demonstrated that hADSC-derived exosome protein expression varied, with lipopolysaccharide (LPS) treatment, in the different isolation methods. Pathway analysis and proliferation, migration and endothelial tube formation assays showed varying effects in cells stimulated with hADSC-derived exosomes from different isolation methods. Our study revealed that different isolation methods might introduce variations in the protein composition in exosomes, which reflects their effects on biological function. The pros and cons of these methods are important points to consider for downstream research applications.

5.
Food Chem ; 358: 129885, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-33933958

RESUMO

The study evaluated the changes in polycyclic aromatic hydrocarbons (PAHs) of Oolong tea samples at each heat treatment stage of the manufacturing process, different post-treatment methods and different brewing conditions. The content of PAHs in the tea leaves was significantly increased during stir fixation (280 °C for 8 min) stage of the manufacturing process. In the subsequent heat treatment process, the PAHs content did not change much until the Oolong tea product (primary) was further roasted. The level of PAHs increased with the roasting time. Charcoal roasting resulted in higher PAHs content in the product compared with electric roasting. Higher brewing temperature caused higher level of PAHs released into the tea infusion. The level of released PAHs decreased with the increase of the number of tea brewing (the total released PAHs was about 4%). The risk assessment results for PAHs in the tea infusions showed a low level of health concern.


Assuntos
Contaminação de Alimentos/análise , Hidrocarbonetos Policíclicos Aromáticos/análise , Chá/química , União Europeia , Indústria de Processamento de Alimentos/métodos , Temperatura Alta , Humanos , Medição de Risco/métodos , Taiwan , Temperatura
6.
J Mater Sci Mater Med ; 32(1): 13, 2021 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-33475850

RESUMO

An essential criterion for the selection of resorbable bioceramics is their ability to degrade inside human body within a reasonable time frame. Furthermore, if the bioceramic can release beneficial ions, such as strontium, as it degrades, recovery time might be shortened. The present study demonstrates that strontium-containing calcium sulfate (Sr,Ca)SO4 can fulfill these criteria. A long-term in vitro degradation analysis for 12 weeks using sintered (Sr,Ca)SO4 discs in phosphate buffered solution (PBS) was conducted. The sintered (Sr,Ca)SO4 disc was then implanted into defects in the distal femur of rats. The degradation rate of (Sr,Ca)SO4 discs showed a strong dependence on the Sr content. Similar results were observed between the long-term in vitro degradation analysis and the in vivo evaluation. The sintered (3.8%Sr,Ca)SO4 disc lost more than 80% of its initial weight after soaking in PBS with shaking at 37 °C for 12 weeks. After 12 weeks in vivo, the remaining volume of the (3.8%Sr,Ca)SO4 disc within the bone defect was ~25%. Over the same time period, new bone was formed at a relative volume of 40%. This study demonstrates the potential of (Sr,Ca)SO4 bioceramic, and the benefits of using a long-term degradation test during the evaluation of resorbable bioceramics.


Assuntos
Implantes Absorvíveis , Materiais Biocompatíveis/farmacocinética , Cerâmica/farmacocinética , Animais , Materiais Biocompatíveis/química , Biotransformação , Substitutos Ósseos/química , Substitutos Ósseos/farmacocinética , Sulfato de Cálcio/química , Sulfato de Cálcio/farmacocinética , Cerâmica/química , Técnicas In Vitro , Teste de Materiais/métodos , Ratos , Ratos Sprague-Dawley , Estrôncio/química , Estrôncio/farmacocinética , Fatores de Tempo
7.
Int J Med Sci ; 18(4): 1058-1066, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33456364

RESUMO

The heterogeneity of exosome populations presents a great challenge to their study. The current study was designed to investigate the potential heterogeneity miRNA contents in circulating exosomes purified via different exosomal markers. In this study, exosomes from the serum of C57BL/6 mice after cecum ligation and perforation (CLP) or sham operation were isolated by precipitation using ExoQuick-TC and affinity purified with anti-Rab5b, anti-CD9, anti-CD31, and anti-CD44 antibodies using the Exo-Flow Exosome Capture kit to collect exosome subpopulations. RNA extracted from the exosomes isolated by ExoQuick-TC were profiled by next-generation sequencing (NGS). Real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) was also employed to determine the expression profiles of four representative exosomal miRNAs (mmu-miR-486-5p, mmu-miR-10a-5p, mmu-miR-143-3p, and mmu-miR-25-3p) selected from the NGS analysis. The results revealed that the expression patterns of these miRNAs in exosomes isolated by ExoQuick-TC as determined by RT-qPCR and NGS were similar, showing upregulation of mmu-miR-10a-5p and mmu-miR-143-3p but downregulation of mmu-miR-25-3p and mmu-miR-486-5p following CLP when compared to the levels in exosomes from sham control mice. However, their expression levels in the antibody-captured exosome subpopulations varied. The miRNAs in the exosomes captured by anti-Rab5b or anti-CD9 antibodies were more similar to those isolated by ExoQuick-TC than to those captured by anti-CD44 antibodies. However, there were no significant differences in these four miRNAs in CD31-captured exosomes. This study demonstrated that purification with different exosomal markers allows the collection of different exosome subpopulations with various miRNA contents. The results of this study demonstrate the heterogeneity of circulating exosomes and suggest the importance of stratifying exosome subpopulations when using circulating exosomes as biomarkers or investigating exosome function. In addition, this study also emphasized the necessity of using a consistent exosome marker across different samples as detecting biomarkers.


Assuntos
MicroRNA Circulante/análise , Exossomos/metabolismo , Sepse/diagnóstico , Animais , Biomarcadores/sangue , Biomarcadores/metabolismo , MicroRNA Circulante/sangue , MicroRNA Circulante/metabolismo , Modelos Animais de Doenças , Humanos , Masculino , Camundongos , Sepse/sangue , Sepse/genética
8.
ACS Appl Mater Interfaces ; 11(45): 42049-42056, 2019 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-31633334

RESUMO

Electrolyte is a key component in high-voltage lithium-ion batteries (LIBs). Bis(trifluoromethanesulfonyl)imide-based ionic liquid (IL)/organic carbonate hybrid electrolytes have been a research focus owing to their excellent balance of safety and ionic conductivity. Nevertheless, corrosion of Al current collectors at high potentials usually happens for this kind of electrolyte. In this study, this long-standing problem is solved via the modulation of the IL/carbonate ratio and LiPF6 concentration in the hybrid electrolyte. The proposed electrolyte suppresses Al dissolution and electrolyte oxidation at 5 V (vs Li+/Li) and thus allows for ideal lithiation/delithiation performance of a high-voltage LiNi0.5Mn1.5O4 (LNMO) cathode even at 55 °C. The underlying mechanism is examined in this work. Excellent cycling stability (97% capacity retention) for an LNMO cathode after 300 cycles is achieved. This electrolyte shows good wettability toward a polyethylene separator and low flammability. In addition, satisfactory compatibility with both graphite and Si-based anodes is confirmed. The proposed electrolyte design strategies have great potential for applications in high-voltage LIBs.

9.
PLoS One ; 14(7): e0220398, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31348811

RESUMO

OBJECTIVE: To establish the composition of bacteria in mice following cecum ligation and puncture (CLP) through metagenomic analysis and investigate the role of TLRs on the composition of bacteria. METHODS: Total DNA extraction was done from the ascites, blood, and fecal samples from C57BL/6 mice sacrificed at 0, 4, 8, and 16 h, as well as from Tlr2-/-, Tlr4-/-, Tlr5-/-, and NF-κB-/-mice sacrificed at 16 h following CLP. Amplification of the V3-V4 regions of the bacterial 16S rRNA genes by PCR and the Illumina MiSeq sequencer was used for deep sequencing. Hierarchical clustering of the isolates was performed with Ward's method using Euclidean distances. The relative abundance according to operational taxonomic unit (OTU) number or taxa was used to compare the richness among subgroups in the experiments. RESULTS: There were 18 taxa that had significantly different abundances among the different samples of the C57BL/6 mice at 16 h following CLP. Various dynamic changes in the infectious bacteria inside the peritoneal cavity after CLP were found. While knockout of Tlr5 and NF-κB impaired the ability of bacterial clearance inside the peritoneal cavity for some kinds of bacteria found in the C57BL/6 mice, the knockout of Tlr4 enhanced clearance for other kinds of bacteria, and they presented excessive abundance in the peritoneal cavity despite their scarce abundance in the stool. CONCLUSION: NF-κB and TLRs are involved in bacterial clearance and in the expression pattern of the bacterial abundance inside the peritoneal cavity during polymicrobial infection.


Assuntos
Bactérias/classificação , Metagenômica/métodos , Cavidade Peritoneal/microbiologia , Receptores Toll-Like/genética , Animais , Ascite/microbiologia , Bactérias/genética , Sangue/microbiologia , DNA Bacteriano , DNA Ribossômico/genética , Fezes/microbiologia , Técnicas de Silenciamento de Genes , Sequenciamento de Nucleotídeos em Larga Escala , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/genética , Filogenia , RNA Ribossômico 16S/genética
10.
PLoS One ; 14(7): e0215499, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31318872

RESUMO

BACKGROUND: Cells, scaffolds, and factors are the triad of regenerative engineering; however, it is difficult to distinguish whether cells in the regenerative construct are from the seeded cells or host cells via the host blood supply. We performed a novel in vivo study to transplant enhanced green fluorescent pig mesenchymal stem cells (EGFP-pMSCs) into calvarial defect of DsRed pigs. The cell distribution and proportion were distinguished by the different fluorescent colors through the whole regenerative period. METHOD/RESULTS: Eight adult domestic Ds-Red pigs were treated with five modalities: empty defects without scaffold (group 1); defects filled only with scaffold (group 2); defects filled with osteoinduction medium-loaded scaffold (group 3); defects filled with 5 x 103 cells/scaffold (group 4); and defects filled with 5 x 104 cells/scaffold (group 5). The in vitro cell distribution, morphology, osteogenic differentiation, and fluorescence images of groups 4 and 5 were analyzed. Two animals were sacrificed at 1, 2, 3, and 4 weeks after transplantation. The in vivo fluorescence imaging and quantification data showed that EGFP-pMSCs were represented in the scaffolds in groups 4 and 5 throughout the whole regenerative period. A higher seeded cell density resulted in more sustained seeded cells in bone regeneration compared to a lower seeded cell density. Host cells were recruited by seeded cells if enough space was available in the scaffold. Host cells in groups 1 to 3 did not change from the 1st week to 4th week, which indicates that the scaffold without seeded cells cannot recruit host cells even when enough space is available for cell ingrowth. The histological and immunohistochemical data showed that more cells were involved in osteogenesis in scaffolds with seeded cells. CONCLUSION: Our in vivo results showed that more seeded cells recruit more host cells and that both cell types participate in osteogenesis. These results suggest that scaffolds without seeded cells may not be effective in bone transplantation.


Assuntos
Regeneração Óssea , Transplante de Células-Tronco Mesenquimais , Crânio/lesões , Animais , Células Cultivadas , Proteínas de Fluorescência Verde/análise , Células-Tronco Mesenquimais/citologia , Osteogênese , Crânio/ultraestrutura , Suínos , Tecidos Suporte/química , Transplante Homólogo
11.
Front Immunol ; 9: 2830, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30559745

RESUMO

The adhesion family of G protein-coupled receptors (aGPCRs) comprises 33 members in human, several of which are distinctly expressed and functionally involved in polymorphonuclear cells (PMNs). As former work indicated the possible presence of the aGPCR GPR97 in granulocytes, we studied its cellular distribution, molecular structure, signal transduction, and biological function in PMNs. RNA sequencing and mass-spectrometry revealed abundant RNA and protein expression of ADGRG3/GPR97 in granulocyte precursors and terminally differentiated neutrophilic, eosinophilic, and basophilic granulocytes. Using a newly generated GPR97-specific monoclonal antibody, we confirmed that endogenous GPR97 is a proteolytically processed, dichotomous, N-glycosylated receptor. GPR97 was detected in tissue-infiltrating PMNs and upregulated during systemic inflammation. Antibody ligation of GPR97 increased neutrophil reactive oxygen species production and proteolytic enzyme activity, which is accompanied by an increase in mitogen-activated protein kinases and IκBα phosphorylation. In-depth analysis of the GPR97 signaling cascade revealed a possible switch from basal Gαs/cAMP-mediated signal transduction to a Gαi-induced reduction in cAMP levels upon mutation-induced activation of the receptor, in combination with an increase in downstream effectors of Gßγ, such as SRE and NF-κB. Finally, ligation of GPR97 increased the bacteria uptake and killing activity of neutrophils. We conclude that the specific presence of GPR97 regulates antimicrobial activity in human granulocytes.


Assuntos
Anti-Infecciosos/metabolismo , Granulócitos/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Anticorpos Monoclonais/metabolismo , Células Cultivadas , AMP Cíclico/metabolismo , Eosinófilos/metabolismo , Humanos , Inflamação/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Neutrófilos/metabolismo , Fosforilação/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/fisiologia
12.
Nutrients ; 10(10)2018 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-30304787

RESUMO

Background: This study aimed at assessing the effect of a low-fat diet (LFD) in obese mice lacking toll⁻like receptors (Tlr) and understanding the expression and regulation of microRNAs during weight reduction. Methods: C57BL/6, Tlr5-/-, Tlr2-/- and Tlr4-/- mice were used in this study. A group of mice were fed with a high-fat diet (HFD) (58% kcal) for 12 weeks to induce obesity (diet-induced obesity, DIO). Another group that had been fed with HFD for eight weeks (obese mice) were switched to a low-fat diet (LFD) (10.5% kcal) for the next four weeks to reduce their body weight. The control mice were fed with a standard AIN-76A diet for the entire 12 weeks. The body weight of the mice was measured weekly. At the end of the experiment, epididymal fat weight and adipocyte size were measured. The differentially expressed miRNAs in the fat tissue was determined by next-generation sequencing with real-time quantitative reverse transcription polymerase chain reaction (RT⁻qPCR). Target prediction and functional annotation of miRNAs were performed using miRSystem database. Results: Switching to LFD significantly reduced the body weight and epididymal fat mass in the HFD-fed C57BL/6 and Tlr5-/- mice but not in Tlr2-/- and Tlr4-/- mice. Weight reduction significantly decreased the size of adipocytes in C57BL/6 but not in the Tlr knockout mice. In Tlr2-/- and Tlr4-/- mice, feeding with HFD and the subsequent weight reduction resulted in an aberrant miRNA expression in the epididymal fat tissue unlike in C57BL/6 and Tlr5-/-. However, target prediction and functional annotation by miRSystem database revealed that all the top 10 Kyoto Encyclopedia of Genes and Genomes (KEGG) database pathways of the dysregulated miRNAs during weight reduction in the C57BL/6 mice were also found in the regulated pathways of Tlr5-/-, Tlr2-/-, and Tlr4-/- strains. However, among these pathways, gene sets involved in arginine and proline metabolism and glutathione metabolism were mainly involved in the Tlr knockout mice but not in the C57BL/6 mice. Conclusions: In this study, we demonstrated that feeding of LFD leads to significant body weight reduction in C57BL/6 and Tlr5-/- mice, but not in Tlr2-/- and Tlr4-/- mice. Significant reduction in the size of adipocytes of epididymal fat was only found in C57BL/6, but not in Tlr5-/-, Tlr2-/-, and Tlr4-/- mice. The dysregulated miRNAs in Tlr2-/- and Tlr4-/- mice were different from those in C57BL/6 and Tlr5-/- strains. Among those miRNA-regulated pathways, arginine and proline metabolism as well as glutathione metabolism may have important roles in the Tlr knockout mice rather than in C57BL/6 mice.


Assuntos
Tecido Adiposo/metabolismo , Dieta com Restrição de Gorduras , MicroRNAs/metabolismo , Obesidade/genética , Receptores Toll-Like/genética , Adipócitos/patologia , Animais , Dieta Hiperlipídica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Obesos , Obesidade/etiologia , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética , Receptor 5 Toll-Like/genética , Perda de Peso
13.
Mater Sci Eng C Mater Biol Appl ; 91: 806-816, 2018 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-30033316

RESUMO

Gene therapy for bone tissue engineering has been widely developed. Recently, non-viral DNA-based gene therapy has been reported to be a safer and more efficient method of delivering DNA into target cells. We used a non-viral gene transfection reagent to delivery bone morphogenetic protein-2 (BMP-2) gene into bone marrow stromal cells (BMSCs). Primary BMSCs were isolated from rat femurs and transfected with BMP-2 plasmids. The transfection rate was analyzed using flow cytometry. The concentration of BMP-2 protein was quantified using an enzyme-linked immunosorbent assay. Levels of osteopontin and osteocalcin were measured to evaluate osteogenic differentiation. In vivo, we designed a critical-size calvarial defect rat model to study new bone regeneration, using Matrigel as a scaffold to carry BMP-2-transfected bone marrow stromal cells into the defect site. New bone formation was assessed by micro-computed tomography, X-ray, immunohistochemical staining and histomophometry. The transfection rate after 72 h was 31.5%. The BMP-2 protein level as well as osteopontin and osteocalcin expressions were higher in the experimental group (transfected with BMP-2) than the control group (transfected with green fluorescent protein, GFP). The in vivo study suggested that bone healing occurred 12 weeks after scaffold implantation. In addition, BMP-2-transfected bone marrow stromal cells provided better osteogenic differentiation than primary bone marrow stromal cells. Our findings suggest that non-viral gene therapy may be useful in bone tissue engineering. SIGNIFICANCE: The study has clinical implications for the wider use of BMP-2-transfected BMSCs for cell-based transplantation therapy in bone regeneration.


Assuntos
Proteína Morfogenética Óssea 2/genética , Proteína Morfogenética Óssea 2/uso terapêutico , Diferenciação Celular , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/citologia , Crânio/patologia , Transfecção , Animais , Antígenos CD/metabolismo , Regeneração Óssea , Calo Ósseo/patologia , Sobrevivência Celular , Modelos Animais de Doenças , Proteínas de Fluorescência Verde/metabolismo , Masculino , Osteocalcina/metabolismo , Osteopontina/metabolismo , Fragmentos de Peptídeos/sangue , Pró-Colágeno/sangue , Ratos Sprague-Dawley , Crânio/diagnóstico por imagem
14.
Oncotarget ; 8(46): 80741-80756, 2017 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-29113341

RESUMO

Background: Toll-like receptors (TLRs) are involved in the initiation of Schwann cell activation and subsequent recruitment of macrophages for clearance of degenerated myelin and neuronal debris after nerve injury. The present study was designed to investigate the regenerative outcome and expression of myelination-related factors in Tlr-knockout mice following a sciatic nerve crush injury. Materials and methods: A standard sciatic nerve crush injury, induced by applying constant pressure to the nerve with a No. 5 jeweler's forceps for 30 s, was performed in C57BL/6, Tlr2-/- , Tlr3-/- , Tlr4-/- , Tlr5-/- , and Tlr7-/- mice. Quantitative histomorphometric analysis of toluidine blue-stained nerve specimens and walking track analysis were performed to evaluate nerve regeneration outcomes. PCR Arrays were used to detect the expression of neurogenesis-related genes of dorsal root ganglia as well as of myelination-related genes of the distal nerve segments. Results: Worse nerve regeneration after nerve crush injury was found in all Tlr-knockout mice than in C57BL/6 mice. Delayed expression of myelin genes and a different expression pattern of myelination-related neurotrophin genes and transcription factors were found in Tlr-knockout mice in comparison to C57BL/6 mice. In these TLR-mediated pathways, insulin-like growth factor 2 and brain-derived neurotrophic factor, as well as early growth response 2 and N-myc downstream-regulated gene 1, were significantly decreased in the early and late stages, respectively, of nerve regeneration after a crush injury. Conclusions: Knockout of Tlr genes decreases the expression of myelination-related factors and impairs nerve regeneration after a sciatic nerve crush injury.

15.
BMC Geriatr ; 17(1): 178, 2017 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-28793879

RESUMO

BACKGROUND: The elderly are predisposed to septic arthritis (SA) because of the aging nature and increasing comorbidities. SA may in turn increase the long-term mortality in the geriatric patients; however, it remains unclear. We conducted this prospective nationwide population-based cohort study to clarify this issue. METHODS: Using Taiwan National Health Insurance Research Database (NHIRD), we identified 1667 geriatric participants (≥ 65 years) with SA and 16,670 geriatric participants without SA matched at a ratio of 1:10 by age, sex, and index date between 1999 and 2010. A comparison of the long-term mortality between the two cohorts through follow-up until 2011 was performed. RESULTS: Geriatric participants with SA had a significantly increased mortality than those without SA [Adjusted hazard ratio (AHR): 1.49, 95% confidence interval (CI): 1.34-1.66], particularly the old elderly (≥ 85 years, AHR: 2.12, 95% CI: 1.58-2.84) and males (AHR: 1.54, 95% CI: 1.33-1.79). These results were stated after adjustment for osteoarthritis, diabetes, gout, renal disease, liver disease, cancer, rheumatoid arthritis, systemic lupus erythematosus, alcoholism, and human immunodeficiency virus infection. The increased mortality risk was highest in the first month (AHR: 3.93, 95% CI: 2.94-5.25) and remained increased even after following up for 2-4 years (AHR: 1.30, 95% CI: 1.03-1.65). After Cox proportional hazard regression analysis, SA (AHR: 1.37, 95% CI: 1.20-1.56), older age (≥ 85 years, AHR: 1.79, 95% CI: 1.59-2.02, 75-84 years, AHR: 1.65, 95% CI: 1.53-1.78), male sex, diabetes, renal disease, liver disease, cancer, and gout were independent mortality predictors. There was no significant difference in the mortality for SA between upper limb affected and lower limb affected. CONCLUSIONS: This study delineated that SA significantly increased the long-term mortality in geriatric participants. For the increasing aging population worldwide, strategies for the prevention and treatment of SA and concomitant control of comorbidities are very important.


Assuntos
Artrite Infecciosa , Idoso , Idoso de 80 Anos ou mais , Artrite Infecciosa/diagnóstico , Artrite Infecciosa/epidemiologia , Comorbidade , Bases de Dados Factuais , Feminino , Humanos , Incidência , Masculino , Mortalidade , Modelos de Riscos Proporcionais , Estudos Prospectivos , Medição de Risco , Fatores de Risco , Taiwan/epidemiologia , Tempo
16.
Sci Rep ; 7(1): 4968, 2017 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-28694493

RESUMO

A GaN/AlGaN ultraviolet light emitting diode (UV-LED) structure with a porous AlGaN reflector structure has been demonstrated. Inside the UV-LED, the n+-AlGaN/undoped-AlGaN stack structure was transformed into a porous-AlGaN/undoped-AlGaN stack structure through a doping-selective electrochemical etching process. The reflectivity of the porous AlGaN reflector was 93% at 374 nm with a stop-bandwidth of 35 nm. In an angle-dependent reflectance measurement, the central wavelength of the porous AlGaN reflector had blueshift phenomenon by increasing light-incident angle from 10° to 50°. A cut-off wavelength was observed at 349 nm due to the material absorption of the porous-AlGaN/u-AlGaN stack structure. In the treated UV-LED structure, the photoluminescence emission wavelength was measured at 362 nm with a 106° divergent angle covered by the porous-AlGaN reflector. The light output power of the treated UV-LED structure was higher than that of the non-treated UV-LED structure due to the high light reflectance on the embedded porous AlGaN reflector.

17.
Opt Express ; 24(11): 11601-10, 2016 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-27410087

RESUMO

A Si-heavy doped GaN:Si epitaxial layer is transformed into a directional nanopipe GaN layer through a laser-scribing process and a selectively electrochemical (EC) etching process. InGaN light-emitting diodes (LEDs) with an EC-treated nanopipe GaN layer have a high light extraction efficiency. The direction of the nanopipe structure was directed perpendicular to the laser scribing line and was guided by an external bias electric field. An InGaN LED structure with an embedded nanopipe GaN layer can enhance external quantum efficiency through a one-step epitaxial growth process and a selective EC etching process. A birefringence optical property and a low effective refractive index were observed in the directional-nanopipe GaN layer.

18.
Mediators Inflamm ; 2015: 852126, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26681840

RESUMO

BACKGROUND: This study aims to investigate the effect of feeding low-fat diet (LFD) to diet-induced obesity (DIO) mice lacking TLR5 (TLR5(-/-)), which have a tendency to develop glucose intolerance with increased adiposity, compared to that in C57BL/6 mice. RESULTS: TLR5(-/-) and C57BL/6 male mice were divided into three subgroups: (1) control, mice were fed a standard AIN-76A (fat: 11.5 kcal%) diet for 12 weeks; (2) DIO, mice were fed a 58 kcal% high-fat diet (HFD) for 12 weeks; and (3) diet, mice were fed a HFD for 8 weeks to induce obesity and then switched to a 10.5 kcal% LFD for 4 weeks. The glucose intolerance in DIO TLR5(-/-) mice was more significant than that in DIO C57BL/6 mice and was not attenuated by a switch to the LFD. Weight-reduction with LFD had significantly decreased the epididymal fat mass in C57BL/6 mice but not in TLR5(-/-) mice. In addition, the LFD-fed TLR5(-/-) mice showed significantly higher expression of ghrelin in the serum and resistin in the epididymal fat than that in C57BL/6 mice. CONCLUSIONS: This study demonstrated that TLR5 gene knockout impairs some effects of weight-reduction in DIO.


Assuntos
Dieta com Restrição de Gorduras , Obesidade/dietoterapia , Obesidade/imunologia , Receptor 5 Toll-Like/deficiência , Adiposidade , Animais , Citocinas/sangue , Dieta Hiperlipídica/efeitos adversos , Grelina/sangue , Intolerância à Glucose/etiologia , Intolerância à Glucose/imunologia , Intolerância à Glucose/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Obesidade/metabolismo , Resistina/metabolismo , Receptor 5 Toll-Like/genética , Ganho de Peso , Perda de Peso
19.
PLoS One ; 10(11): e0142000, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26529110

RESUMO

Prior studies of upper gastrointestinal bleeding (UGIB) and acute myocardial infarction (AMI) are small, and long-term effects of UGIB on AMI have not been delineated. We investigated whether UGIB in patients diagnosed with coronary artery disease (CAD) increased their risk of subsequent AMI. This was a population-based, nested case-control study using Taiwan's National Health Insurance Research Database. After propensity-score matching for age, gender, comorbidities, CAD date, and follow-up duration, we identified 1,677 new-onset CAD patients with AMI (AMI[+]) between 2001 and 2006 as the case group and 10,062 new-onset CAD patients without (AMI[-]) as the control group. Conditional logistic regression was used to examine the association between UGIB and AMI. Compared with UGIB[-] patients, UGIB[+] patients had twice the risk for subsequent AMI (adjusted odds ratio [AOR] = 2.08; 95% confidence interval [CI], 1.72-2.50). In the subgroup analysis for gender and age, UGIB[+] women (AOR = 2.70; 95% CI, 2.03-3.57) and patients < 65 years old (AOR = 2.23; 95% CI, 1.56-3.18) had higher odds of an AMI. UGIB[+] AMI[+] patients used nonsignificantly less aspirin than did UGIB[-] AMI[+] patients (27.69% vs. 35.61%, respectively). UGIB increased the risk of subsequent AMI in CAD patients, especially in women and patients < 65. This suggests that physicians need to use earlier and more aggressive intervention to detect UGIB and prevent AMI in CAD patients.


Assuntos
Doença da Artéria Coronariana/epidemiologia , Hemorragia Gastrointestinal/epidemiologia , Infarto do Miocárdio/epidemiologia , Sistema de Registros , Idoso , Doença da Artéria Coronariana/complicações , Feminino , Seguimentos , Hemorragia Gastrointestinal/complicações , Humanos , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/etiologia , Fatores de Risco , Fatores Sexuais , Taiwan/epidemiologia
20.
Dis Markers ; 2015: 863192, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26435568

RESUMO

BACKGROUND: This study aimed to establish the expression profile of circulating microRNAs (miRNAs) during nerve allotransplantation in the presence and absence of FK506 immunosuppression. METHODS: A 1 cm BALB/c donor sciatic nerve graft was transplanted into the sciatic nerve gaps created in recipient C57BL/6 mice with or without daily FK506 immunosuppression [1 mg/(kg·d)]. At 3, 7, and 14 d after nerve allotransplantation, serum samples were collected for miRNA expression analysis by Illumina small RNA deep sequencing. RESULTS: Sequence analysis showed that the dominant size of circulating small RNAs after nerve allotransplantation was 22 nucleotides, followed by 23-nucleotide sequences. Nine upregulated circulating miRNAs (let-7e-5p, miR-101a-3p, miR-151-5p, miR-181a-5p, miR-204-5p, miR-340-5p, miR-381-3p, miR-411-5p, miR-9-5p, and miR-219-2-3p) were identified at 3 d, but none was identified at 7 or 14 d. Among them, miR-9-5p had the highest fold-change of >50-fold, followed by miR-340-5p with 38.8-fold. The presence of these nine miRNAs was not significant at 7 and 14 d after nerve allotransplantation with or without immunosuppression, showing that these miRNAs are not ideal biomarkers for monitoring rejection of deep-buried nerve allografts, a response usually observed later. CONCLUSIONS: We identified nine upregulated circulating miRNAs, which may have a biological function, particularly during the early stages after nerve allotransplantation under FK506 immunosuppression.


Assuntos
Aloenxertos/transplante , Imunossupressores/farmacologia , MicroRNAs/sangue , Nervo Isquiático/transplante , Tacrolimo/farmacologia , Animais , Sequenciamento de Nucleotídeos em Larga Escala , Imunossupressores/efeitos adversos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , MicroRNAs/genética , Tacrolimo/efeitos adversos , Regulação para Cima/efeitos dos fármacos
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