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1.
Methods Mol Biol ; 2206: 39-46, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32754809

RESUMO

During angiogenesis, endothelial cells must undergo a coordinated set of morphological changes in order to form a new vessel. There is a need for endothelial cells to communicate with each other in order to take up different identities in the sprout and to migrate collectively as a connected chord. Endothelial cells must also interact with a wide range of other cells that contribute to vessel formation. In ischemic disease, hypoxic cells in tissue will generate proangiogenic signals that promote and guide angiogenesis. In solid tumors, this function is co-opted by tumor cells, which make a complex range of interactions with endothelial cells, even integrating into the walls of vessels. In vessel repair, cells from the immune system contribute to the promotion and remodeling of new vessels. The coculture angiogenesis assay is a long-term in vitro protocol that uses fibroblasts to secrete and condition an artificial stromal matrix for tubules to grow through. We show here how the assay can be easily adapted to include additional cell types, facilitating the study of cellular interactions during neovascularization.

2.
FASEB J ; 33(11): 12277-12287, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31431053

RESUMO

Endothelial cells (ECs) form an active barrier between the circulation and the body. In addition to controlling transport of molecules between these 2 compartments, the endothelium is a major secretory organ, releasing proteins both into the circulation and into the vascular matrix. Although it is clearly important that proteins are correctly sorted into these 2 spaces, we currently know little of the polarization of this secretion or how it is controlled. Here, we present an optimized system for the analysis of polarized secretion and show that it allows the derivation of deep, robust proteomes from small numbers of primary ECs. We present the first endothelial apically and basolaterally secreted proteomes, demonstrating that ECs polarize the secretion of extracellular vesicle cargoes to the apical surface. Conversely, we find that protein secretion at the basolateral surface is focused on components of the extracellular matrix (ECM). Finally, we examine the role of liprin-α1 in secretion toward the basolateral compartment and identify a subset of ECM components that share this route with fibronectin.-Wei, H., Sundararaman, A., Dickson, E., Rennie-Campbell, L., Cross, E., Heesom, K. J., Mellor, H. Characterization of the polarized endothelial secretome.

3.
Front Physiol ; 9: 1671, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30564133

RESUMO

Liver fibrosis is an abnormal wound healing response and a common consequence of chronic liver diseases from infection or alcohol/xenobiotic exposure. At the cellular level, liver fibrosis is mediated by trans-differentiation of hepatic stellate cells (HSCs), which is driven by persistent hepatic and systemic inflammation. However, impaired enterohepatic circulation and gut dysbiosis may indirectly contribute to the liver fibrogenesis. The composition of the gut microbiota depends on diet composition and host factors. In this study, we examined chlorophyllin, derived from green pigment chlorophyll, on gut microbiota, the intestinal mucosal barrier, and liver fibrosis. BALB/c mice received carbon tetrachloride through intraperitoneal injection to induce liver fibrosis and chlorophyllin was administrated in drinking water. The effects of chlorophyllin on liver fibrosis were evaluated for (1) survival rate, (2) hepatic morphologic analysis, (3) inflammatory factors in both the small intestine and liver, and (4) gut microbiota. Our results indicate that oral administration of chlorophyllin could attenuate intestinal and hepatic inflammation and ameliorate liver fibrosis. Importantly, oral administration of chlorophyllin promptly rebalanced the gut microbiota, exhibiting down-regulation of the phylum Firmicutes and up-regulation of the phylum Bacteroidetes. In vitro experiments on intestinal epithelial cells showed that chlorophyllin exposure could inhibit NF-κB pathway via IKK-phosphorylation suppression. In conclusion, this study demonstrates potential application of chlorophyllin to regulate the intestinal microbiota and ameliorate hepatic fibrosis.

4.
BMC Endocr Disord ; 18(1): 25, 2018 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-29739382

RESUMO

BACKGROUND: Recent epidemiological studies have suggested inverse associations between vitamin D status and metabolic diseases including type 2 diabetes (T2DM). The aim of this study was to examine whether a higher serum 25-hydroxyvitamin D (25(OH)D) was associated with a more favorable glucose homeostasis among adults without diabetes in Southwest China. METHODS: Serum 25(OH)D concentration was measured in a cross-sectional sample of 1514 adults without diabetes aged 25-65 years recruited from Southwest China. Indices describing glucose homeostasis included fasting plasma glucose (FPG), fasting insulin, glycated hemoglobin (HbA1c), the homeostatic model assessment 2-insulin resistance (HOMA2-IR) and odds of pre-diabetes. Data were analyzed by multivariable-adjusted regression models. RESULTS: The average serum 25(OH)D was 22.66 ng/ml, and percentages of vitamin D deficiency [25(OH)D < 20 ng/ml], insufficiency [20 ≤ 25(OH)D ≤ 30 ng/ml] were 47.6 and 32.2%, respectively. Serum 25(OH)D was inversely associated with fasting insulin (P = 0.0007), HbA1c (P = 0.0001) and HOMA2-IR (P = 0.0007), but not with FPG, after adjusting for age, gender, monthly personal income, smoking status, energy intake, moderate-to-vigorous physical activity (MVPA) and waist circumference (WC). Compared with the lowest 25(OH)D tertile, the odds ratio for pre-diabetes in the highest tertile was 0.68 (95%CI: 0.47-0.99) after adjustment for cofounders. In the following stratified analyses according to weight status, we only observed this inverse association between serum 25(OH)D and pre-diabetes in overweight or obese adults (n = 629, P = 0.047), but not in their counterparts with BMI < 24 kg/m2. CONCLUSIONS: Our results advocate that a higher serum 25(OH)D level is associated with decreased risk of impairment of glucose homeostasis among adults without diabetes in Southwest China. Further studies are warranted to determine the role of vitamin D in glucose homeostasis.


Assuntos
Biomarcadores/metabolismo , Diabetes Mellitus Tipo 2/prevenção & controle , Glucose/metabolismo , Homeostase , Deficiência de Vitamina D/complicações , Vitamina D/análogos & derivados , Adulto , Idoso , Estudos Transversais , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/etiologia , Feminino , Seguimentos , Humanos , Resistência à Insulina , Masculino , Pessoa de Meia-Idade , Prognóstico , Vitamina D/sangue
5.
Virol J ; 15(1): 16, 2018 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-29347945

RESUMO

BACKGROUND: Porcine cytomegalovirus (PCMV) is an immunosuppressive virus that mainly inhibits T-lymphocyte and macrophage immune functions; it has significantly damaged the farming industry. Although recent studies have shown that miRNAs play important roles in immune responses, the regulatory mechanisms of miRNAs during immunosuppressive virus infection remain unclear. METHODS: In this study, porcine small-RNA transcriptomes of PCMV-infected and uninfected vital organs were first characterised by high-throughput sequencing. miRDeep2 software was used to predict novel pig-encoded miRNAs. To verify the accuracy of the high-throughput sequencing results, stem-loop qRT-PCR was performed on 12 significantly DE miRNAs. The physical and functional interactions between the immune-related target genes of the DE miRNAs in PCMV-infected organs were analysed using the STRING database. RESULTS: In total, 306 annotated and 295 novel miRNAs were identified from PCMV-infected and uninfected porcine organs, respectively, through alignment with known Sus scrofa pre-miRNAs. Overall, 92, 107, 95, 77 and 111 miRNAs were significantly differentially expressed in lung, liver, spleen, kidney and thymus after PCMV infection, respectively. According to Gene Ontology enrichment analysis, target genes of the differentially expressed miRNAs associated with immune system processes, regulation of biological processes and metabolic processes were enriched in every sample. Integrated expression analysis of the differentially expressed miRNAs and their target mRNAs in PCMV-infected thymus showed that the significant differential expression of specific miRNAs under the pressure of PCMV infection in central immune organs interfered with the expression of genes involved in important immune-related signalling pathways, thus promoting the viral infection. CONCLUSIONS: This is the first comprehensive analysis of the responses of host small-RNA transcriptomes to PCMV infection in vital porcine organs. It provides new insights into the regulatory mechanisms of miRNAs during infection by immunosuppressive viruses.


Assuntos
Infecções por Citomegalovirus/veterinária , Citomegalovirus , Interações Hospedeiro-Patógeno/genética , MicroRNAs/genética , Doenças dos Suínos/genética , Doenças dos Suínos/virologia , Transcriptoma , Animais , Biologia Computacional/métodos , Citomegalovirus/imunologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Sequenciamento de Nucleotídeos em Larga Escala , Hospedeiro Imunocomprometido , Anotação de Sequência Molecular , Especificidade de Órgãos/genética , Especificidade de Órgãos/imunologia , Interferência de RNA , Reprodutibilidade dos Testes , Análise de Sequência de DNA , Suínos , Doenças dos Suínos/imunologia , Doenças dos Suínos/patologia
6.
J Biomater Appl ; 30(9): 1385-91, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26801474

RESUMO

Pelvic organ prolapse (POP) is a serious health issue that affects many adult women. Surgical treatments for POP patients comprise a common strategy in which scaffold materials are used to reconstruct the prolapsed pelvic. However, the existing materials for pelvic reconstruction cannot meet clinical requirements in terms of biocompatibility, mechanics and immunological rejection. To address these concerns, polypropylene (PP) mesh was selected because of its strong mechanical properties. Small intestinal submucosa (SIS) was used to modify the PP mesh via a mussel-inspired polydopamine coating to enhance its biocompatibility. The scanning electron microscopy (SEM) and atomic force microscopy (AFM) results demonstrated that SIS was successfully conjugated on the surface of the PP mesh. Moreover, the cytotoxicity results indicated that the PP mesh and SIS-modified PP mesh were safe to use. Furthermore, in vivo tests demonstrated that the fibroplasia around the implanted site in the SIS-modified PP mesh group was significantly less than the fibroplasia around the PP mesh group. In addition, the immunohistochemistry staining results indicated that the expression of pro-inflammatory macrophages (M1) was substantially lower and that the expression of pro-healing macrophages (M2) was higher in the SIS-modified PP mesh group. Furthermore, ELISA detection indicated that the expression of IL-1ß and IL-6 in the SIS-modified PP mesh group was reduced compared with the PP mesh group. These findings suggest that a SIS-modified polypropylene hybrid mesh via a mussel-inspired polydopamine coating is a promising approach in pelvic reconstruction.


Assuntos
Materiais Revestidos Biocompatíveis/química , Indóis/química , Mucosa Intestinal/química , Polímeros/química , Polipropilenos/química , Telas Cirúrgicas , Tecidos Suporte/química , Animais , Materiais Biomiméticos/química , Materiais Biomiméticos/toxicidade , Bivalves/química , Linhagem Celular , Materiais Revestidos Biocompatíveis/toxicidade , Feminino , Indóis/imunologia , Indóis/toxicidade , Interleucina-1beta/análise , Interleucina-1beta/imunologia , Interleucina-6/análise , Interleucina-6/imunologia , Mucosa Intestinal/imunologia , Teste de Materiais , Camundongos , Polímeros/toxicidade , Polipropilenos/imunologia , Polipropilenos/toxicidade , Ratos Sprague-Dawley , Telas Cirúrgicas/efeitos adversos , Suínos
7.
Virol J ; 11: 106, 2014 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-24903213

RESUMO

BACKGROUND: Porcine torovirus (PToV) is a member of the genus Torovirus which is responsible for gastrointestinal disease in both human beings and animals with particular prevalence in youth. Torovirus infections are generally asymptomatic, however, their presence may worsen disease consequences in concurrent infections with other enteric pathogens. METHODS: A total of 872 diarrheic fecal samples from pigs of different ages were collected from 12 districts of Sichuan Province in the southwest of China. RT-PCR was done with PToV S gene specific primers to detect the presence of PToV positive samples. M gene specific primers were used with the PToV positive samples and the genes were sequenced. A phylogenetic tree was constructed based on the M gene nucleotide sequences from the 19 selected novel Sichuan strains and 21 PToV and BToV M gene sequences from GenBank. RESULTS: A total of 331 (37.96%, 331/872) samples were found to be positive for PToV and the highest prevalence was observed in piglets aged from 1 to 3 weeks old. Through phylogenetic inference the 40 PToV M gene containing sequences were placed into two genotypes (I & II). The 19 novel Sichuan strains of genotype I showed strong correlations to two Korean gene sequences (GU-07-56-11 and GU-07-56-22). Amino-acid sequence analysis of the 40 PToV M gene strains revealed that the M gene protein was highly conserved. CONCLUSIONS: This study uncovered the presence of PToV in Sichuan Province, and demonstrated the need for continuous surveillance PToV of epidemiology.


Assuntos
Doenças dos Suínos/epidemiologia , Doenças dos Suínos/virologia , Infecções por Torovirus/veterinária , Torovirus/classificação , Torovirus/genética , Animais , China/epidemiologia , Análise por Conglomerados , Diarreia/epidemiologia , Diarreia/veterinária , Diarreia/virologia , Fezes/virologia , Epidemiologia Molecular , Dados de Sequência Molecular , Filogenia , RNA Viral/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Homologia de Sequência , Suínos , Torovirus/isolamento & purificação , Infecções por Torovirus/epidemiologia , Infecções por Torovirus/virologia
8.
Virol J ; 10: 161, 2013 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-23705989

RESUMO

BACKGROUND: Torque teno sus virus (TTSuV), infecting domestic swine and wild boar, is a non-enveloped virus with a circular, single-stranded DNA genome. which has been classified into the genera Iotatorquevirus (TTSuV1) and Kappatorquevirus (TTSuV2) of the family Anelloviridae. A molecular study was conducted to detect evidence of a phylogenic relationship between these two porcine TTSuV genogroups from the sera of 244 infected pigs located in 21 subordinate prefectures and/or cities of Sichuan. RESULTS: Both genogroups of TTSuV were detected in pig sera collected from all 21 regions examined. Of the 244 samples, virus from either genogroup was detected in 203 (83.2%), while 44 animals (18.0%) were co-infected with viruses of both genogroups. Moreover, TTSuV2 (186/244, 76.2%) was more prevalent than TTSuV1 (61/244, 25%). There was statistically significant difference between the prevalence of genogroups 1 infection alone (9.4%, 23/244) and 2 alone (64.8%, 158/244), and between the prevalence of genogroups 2 (76.2%, 186/244) and both genogroups co-infection (18.0%, 44/244). The untranslated region of the swine TTSuV genome was found to be an adequate molecular marker of the virus for detection and surveillance. Phylogenetic analysis indicated that both genogroups 1 and 2 could be further divided into two subtypes, subtype a and b. TTSuV1 subtype b and the two TTSuV2 subtypes are more prevalent in Sichuan Province. CONCLUSIONS: Our study presents detailed geographical evidence of TTSuV infection in China.


Assuntos
Infecções por Vírus de DNA/veterinária , Filogeografia , Polimorfismo Genético , Torque teno virus/classificação , Torque teno virus/isolamento & purificação , Animais , China , Infecções por Vírus de DNA/virologia , DNA Viral/química , DNA Viral/genética , Dados de Sequência Molecular , Análise de Sequência de DNA , Soro/virologia , Suínos , Torque teno virus/genética
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