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1.
Zhonghua Wei Zhong Bing Ji Jiu Yi Xue ; 32(1): 83-87, 2020 Jan.
Artigo em Chinês | MEDLINE | ID: mdl-32148237

RESUMO

OBJECTIVE: To investigate the effect and mechanism of paeoniflorin on the permeability of cardiac microvascular endothelial cells (CMECs) in sepsis. METHODS: Primary rat CMECs were isolated and cultured in vitro, and the cells in the logarithmic growth phase were used for experiments. Tetramethylazozolium colorimetry (MTT) was used to screen the safe and effective concentrations of paeoniflorin at 10, 20, and 40 µmol/L. The cells were divided into blank control group, lipopolysaccharide (LPS) group and low, medium and high concentration paeoniflorin pretreatment group. The cells in the blank control group were cultured in complete medium; the cells in the LPS group were challenged with LPS (1 mg/L) in complete medium; and the cells in the paeoniflorin pretreatment groups were pretreated with 10, 20, and 40 µmol/L paeoniflorin at 4 hours before LPS stimulation. The cells in each group were further cultured for 24 hours after LPS stimulation. The horseradish peroxidase (HRP) method was used to detect the permeability of rat CMECs. The enzyme-linked immunosorbent assay (ELISA) was used to detect the CXC chemokine ligand (CXCL1, CXCL2) levels in the cell supernatant. The real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect the mRNA expressions of CXCL1 and CXCL2 in the cells. Western Blot was used to detect phosphorylated Src (p-Src), vascular endothelial-cadherin (VE-cadherin) and phosphorylated mitogen activated protein kinase (p-MAPK). RESULTS: Compared with the blank control group, the permeability of rat CMECs in the LPS group was significantly increased. The cell permeability was improved to some extent after paeoniflorin pretreatment at different concentrations, and the improvement was most obvious in the 40 µmol/L paeoniflorin group, with statistically significant difference as compared with the LPS group (A value: 1.61±0.07 vs. 2.13±0.06, P < 0.01). ELISA results showed that there were moderate amounts of CXCL1 and CXCL2 in the cell supernatant of rat CMECs in the blank control group. However, the secretion of CXCL1 and CXCL2 in the cell supernatant was increased significantly under the induction of LPS. After pretreatment with paeoniflorin at different concentrations, the secretion of CXCL1 and CXCL2 in the cell supernatant was significantly reduced. The most obvious inhibitory effect on CXCL1 was 40 µmol/L paeoniflorin, and the most obvious inhibition on CXCL2 was 20 µmol/L paeoniflorin, the differences were statistically significant as compared with the LPS group [CXCL1 (ng/L): 337.51±68.04 vs. 829.86±65.06, CXCL2 (ng/L): 4.48±0.11 vs. 9.41±0.70, both P < 0.01]. RT-qPCR results showed that the mRNA expressions of CXCL1 and CXCL2 in the rat CMECs were consistent with the ELISA results. LPS could increase mRNA expressions of CXCL1 and CXCL2 in the rat CMECs, and pretreatment with different concentrations of paeoniflorin could significantly reduce the mRNA expressions of CXCL1 and CXCL2. The 40 µmol/L paeoniflorin had the best inhibitory effect on CXCL1 mRNA expression, and the 20 µmol/L paeoniflorin had the best inhibitory effect on CXCL2 mRNA expression, the differences were statistically significant as compared with the LPS group [CXCL1 mRNA (2-ΔΔCt): 0.543±0.004 vs. 0.812±0.089, CXCL2 mRNA (2-ΔΔCt): 10.52±0.71 vs. 17.68±1.09, both P < 0.01]. Western Blot results showed that moderate amounts of p-Src, VE-cadherin and p-MAPK proteins were expressed in the rat CMECs in the blank control group. After LPS stimulation, the expressions of p-Src and p-MAPK proteins were increased significantly, while the expression of VE-cadherin protein was decreased significantly. After pretreatment with different concentrations of paeoniflorin, the expressions of p-Src and p-MAPK proteins in the cells were decreased to varying degrees, while the expression of VE-cadherin protein was increased, and 40 µmol/L paeoniflorin had the most obvious effect, the differences were statistically significant as compared with the LPS group [p-Src protein (p-Src/GAPDH): 1.02±0.09 vs. 1.29±0.05, p-MAPK proteins (p-MAPK/GAPDH): 0.24±0.02 vs. 0.62±0.02, VE-cadherin protein (VE-cadherin/GAPDH): 0.64±0.03 vs. 0.31±0.02, all P < 0.01]. CONCLUSIONS: Paeoniflorin can regulate the Src/VE-cadherin pathway in CMECs, inhibit the expression and secretion of inflammation-related proteins and chemokines, and improve the cell permeability of CMECs induced by LPS.

2.
Cell Host Microbe ; 27(1): 115-128.e8, 2020 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-31917956

RESUMO

Antiviral immunity in insects is mediated by the RNA interference (RNAi) pathway. Viruses evade antiviral RNAi by expressing virulence factors known as viral suppressors of RNAi (VSR). Here, we report the identification of VINR, a Drosophila VSR-interacting long non-coding (lnc) RNA that activates non-canonical innate immune signaling upon detection of the dsRNA-binding VSR of Drosophila C virus (DCV). VINR is required for the induction of antimicrobial peptide (AMP) genes but dispensable for antiviral RNAi. VINR functions by preventing the ubiquitin proteasome-dependent degradation of Cactin, a coiled-coil and arginine-serine-rich domain-containing protein that regulates a non-cannonical antimicrobial pathway for AMP induction. CRISPR-Cas9 knockout of VINR in Drosophila cells enhances DCV replication independently of antiviral RNAi, and VINR-knockout adult flies exhibit enhanced disease susceptibility to DCV and bacteria. Our findings reveal a counter counter-defense strategy activated by a lncRNA in response to the viral suppression of the primary antiviral RNAi immunity.

3.
Mol Oncol ; 2020 Jan 17.
Artigo em Inglês | MEDLINE | ID: mdl-31951095

RESUMO

Patients with metastatic gastric cancer (GC) have a poor prognosis; however, the molecular mechanism of GC metastasis remains unclear. Here, we employed bioinformatics to systematically screen the metastasis-associated genes and found that the levels of basal cell adhesion molecule (BCAM) were significantly increased in GC tissues from patients with metastasis, as compared to those without metastasis. The upregulation of BCAM was also significantly associated with a shorter survival time. Depletion of BCAM inhibited GC cell migration and invasion. Knockout (KO) of BCAM by the CRISPR/Cas9 system reduced the invasion and metastasis of GC cells. To explore the mechanism of BCAM upregulation, we identified a previously uncharacterized BCAM sense lncRNA that spanned from exon 6 to intron 6 of BCAM, and named it as BCAM-associated long noncoding RNA (BAN). Knockdown of BAN inhibited BCAM expression at both mRNA and protein levels. Knockdown of BAN suppressed GC cell migration and invasion, which was effectively rescued by ectopic expression of BCAM. Further clinical data showed that BAN upregulation was associated with GC metastasis and poor prognosis. Importantly, BAN expression was also significantly associated with that of BCAM in GC tissues. Taken together, these results indicate that increased expression of BCAM and its sense lncRNA BAN promote GC cell invasion and metastasis, and are associated with poor prognosis of GC patients.

4.
Insect Sci ; 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-31264303

RESUMO

The genome-wide characterization of long non-coding RNA (lncRNA) in insects demonstrates their importance in fundamental biological processes. Essentially, an in-depth understanding of the functional repertoire of lncRNA in insects is pivotal to insect resources utilization and sustainable pest control. Using a custom bioinformatics pipeline, we identified 1861 lncRNAs encoded by 1852 loci in the Sogatella furcifera genome. We profiled lncRNA expression in different developmental stages and observed that the expression of lncRNAs is more highly temporally restricted compared to protein-coding genes. More up-regulated Sogatella furcifera lncRNA expressed in the embryo, 4th and 5th instars, suggesting that increased lncRNA levels may play a role in these developmental stages. We compared the relationship between the expression of Sogatella furcifera lncRNA and its nearest protein gene and found that lncRNAs were more correlated to their downstream coding neighbors on the opposite strand. Our genome-wide profiling of lncRNAs in Sogatella furcifera identifies exciting candidates for characterization of lncRNAs, and also provides information on lncRNA regulation during insect development.

5.
BMC Genomics ; 20(1): 396, 2019 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-31113376

RESUMO

BACKGROUND: Phenotypic plasticity is a common and highly adaptive phenomenon where the same genotype produces different phenotypes in response to environmental cues. Sogatella furcifera, a migratory pest of rice exhibits wing dimorphism, is a model insect for studying phenotypic plasticity of wing size. The Insullin-PI3K-Akt-FOXO signaling pathway plays a crucial role in the manipulation of wing size in the migratory insects. However, the regulatory mechanism via the pathway involved in wing dimorphism are still unexplored. RESULTS: Accompanied by special alternative splicing, genes involved in muscle contraction and energy metabolism were highly expressed in the wing hinges of macropters, demonstrating their adaptation for energy-demanding long-distance flights. Based on FOXO ChIP-Seq analysis, a total of 1259 putative target genes were observed in the wing hinges, including wing morph development, flight muscle and energy metabolism genes. An integrated gene interaction network was built by combining four heterogeneous datasets, and the IIS-PI3K-Akt-FOXO pathway was clustered in a divided functional module. In total, 45 genes in the module directly interacting with the IIS-PI3K-Akt-FOXO pathway showed differential expression levels between the two wing hinges, thus are regarded as potential Insulin pathway mediated wing dimorphism related genes (IWDRGs). Of the 45 IWDRGs, 5 were selected for verification by gene knockdown experiments, and played significant roles in the insect wing size regulation. CONCLUSIONS: We provided valuable insights on the genetic basis of wing dimorphism, and also demonstrated that network analysis is a powerful approach to identify new genes regulating wing dimorphic development via insulin signaling pathway in the migratory insect.


Assuntos
Genes de Insetos , Hemípteros/genética , Insulina/fisiologia , Asas de Animais/metabolismo , Processamento Alternativo , Animais , Ácidos Graxos/metabolismo , Feminino , Fatores de Transcrição Forkhead/metabolismo , Expressão Gênica , Redes Reguladoras de Genes , Hemípteros/anatomia & histologia , Hemípteros/metabolismo , Proteínas de Insetos/metabolismo , Proteínas Musculares/genética , Fenótipo , Transdução de Sinais , Asas de Animais/anatomia & histologia
6.
FASEB J ; 33(7): 7915-7928, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30913394

RESUMO

Gastric cancer (GC) is among the most lethal human malignancies, and the leading cause of GC mortality is metastasis. However, the precise mechanism of GC metastasis remains unclear. To screen key transcriptional factors (TFs) involved in GC metastasis, we performed bioinformatics analysis of The Cancer Genome Atlas database and found that Krüppel-like factor 9 (KLF9) is a GC metastasis-associated TF. KLF9 is significantly decreased in patients with GC with distant metastasis compared with those patients without distant metastasis. Ectopic expression of KLF9 evidently inhibited the migration and invasion capabilities of GC cells. Conversely, knockdown of KLF9 endowed GC cells with stronger invasive capacity. Moreover, tail intravenous injection confirmed that KLF9 strongly inhibits the lung metastasis process of GC in vivo. Mechanistically, chromatin immunoprecipitation coupled with high-throughput sequencing data from Encyclopedia of DNA Elements revealed that KLF9 specifically binds to the promoter region of matrix metalloproteinase (MMP)28. Further quantitative real-time PCR and dual-luciferase assay indicated that KLF9 directly inhibited MMP28 transcription. Importantly, decreased invasion and metastasis capability of GC cells caused by ectopic KLF9 expression could be rescued via reinforcing MMP28 expression in vivo. Collectively, our study indicates that KLF9 significantly suppresses GC cell invasion and metastasis through inhibiting MMP28 transcription.-Li, Y., Sun, Q., Jiang, M., Li, S., Zhang, J., Xu, Z., Guo, D., Gu, T., Wang, B., Xiao, L., Zhou, T., Zhuo, W. KLF9 suppresses gastric cancer cell invasion and metastasis through transcriptional inhibition of MMP28.

7.
Gastroenterology ; 156(3): 676-691.e11, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30445010

RESUMO

BACKGROUND & AIMS: We aimed to identify long noncoding RNAs (lncRNAs) that are up-regulated in gastric cancer tissues from patients and study their function in gastric tumor metastasis. METHODS: We collected gastric tumor and nontumor tissues from patients in China and analyzed levels of lncRNAs by microarray analysis, proteins by immunohistochemistry, and RNAs by quantitative reverse-transcription polymerase chain reaction; we compared these with survival times of patients and tumor progression. RNA levels were knocked down or knocked out in BGC-823, SGC-7901, and MKN45 cell lines using small interfering or short hairpin RNAs or clustered regularly interspaced short palindromic repeats (ie, CRISPR)/CRISPR associated protein 9 (ie, Cas9) vectors. Genes were overexpressed from transfected plasmids in HGC-27 cells. Cells were analyzed by Northern blot and immunoblot, polysome profiling assay, and cell invasion assay. Cells were injected into the tail veins or spleens of nude mice or SCID mice; lung and liver tissues were collected, and metastases were counted. lncRNAs were cloned by using rapid amplification of complementary DNA ends. Their interactions with other genes were determined by RNA pulldown and mapping assays. RESULTS: In microarray analyses, we identified 151 lncRNAs expressed at significantly higher levels in gastric tumor vs nontumor tissues. Levels of an lncRNA that we called gastric cancer metastasis associated long noncoding RNA (GMAN) were increased in gastric tumor tissues, compared with nontumor tissues; its up-regulation was associated with tumor metastasis and shorter survival times of patients. The GMAN gene overlaps with the ephrin A1 gene (EFNA1) and was highly expressed in BGC-823 and MKN45 cells. Knockdown of GMAN in these cells did not affect proliferation, colony formation, or adhesion but did reduce their invasive activity in Transwell assays. Ectopic expression of GMAN increased the invasive activity of HGC-27 cells. BGC-823 and MKN45 cells with knockdown of GMAN formed fewer metastases after injection into tail veins of nude mice. Knockdown or knockout of GMAN also reduced levels of ephrin A1 protein in cells. We found that GMAN promoted translation of ephrin A1 messenger RNA into protein by binding to the antisense GMAN RNA (GMAN-AS)-this antisense sequence is also complementary to that of ephrin A1 mRNA. Levels of ephrin A1 protein were also increased in gastric tumors from patients with metastases than in those without metastases. Knockout of ephrin A1 in BGC-823 cells reduced their invasive activity in Transwell assays and ability to form metastases after injection into SCID mice. Ectopic expression of ephrin A1 in BGC-823 cells with knockdown or knockout of GMAN restored their invasive activities and ability form metastases in nude or SCID mice. A CRISPR/Cas9-based strategy to disrupt the GMAN gene significantly reduced the numbers of metastases formed from SGC-7901 cells in mice. CONCLUSIONS: We identified an lncRNA, which we call GMAN, that is increased in gastric tumors from patients and associated with survival and formation of metastases. It regulates translation of ephrin A1 mRNA by binding competitively to GMAN-AS. Knockdown or knockout of GMAN or ephrin A1 in gastric cancer cell lines reduces their invasive activity and ability to form metastases after injection into mice. These genes might be targeted to prevent or reduce gastric cancer metastasis.


Assuntos
Biomarcadores Tumorais/genética , Efrina-A1/genética , Regulação Neoplásica da Expressão Gênica/genética , RNA Longo não Codificante/genética , Neoplasias Gástricas/genética , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Humanos , Camundongos , Camundongos Knockout , Camundongos Nus , Camundongos SCID , Invasividade Neoplásica/patologia , Metástase Neoplásica/genética , Metástase Neoplásica/patologia , Interferência de RNA , RNA Mensageiro/genética , Distribuição Aleatória , Reação em Cadeia da Polimerase em Tempo Real , Neoplasias Gástricas/patologia , Análise Serial de Tecidos , Técnicas de Cultura de Tecidos , Ativação Transcricional , Regulação para Cima
8.
PLoS One ; 13(9): e0204517, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30248141

RESUMO

MicroRNAs (miRNAs) are a class of endogenous regulatory RNA molecules 21-24 nucleotides in length that act as functional regulators of post-transcriptional repression of messenger RNA. We report the identification and characterization of a conserved miRNA and 171 novel miRNAs in the migratory rice pest Sogatella furcifera by deep sequencing, which were observed to be biased towards female adults of the insect, modulating the functionality and targets of the miRNAs in sex differentiation. A switch in arm usage was also observed in 9 miRNA when compared to the insect ancestor during insect evolution. The miRNA loci showed high 5' fidelity in both miRNA and star species and about 93.4% of WBPH miRNAs conserved within non-planthopper species were homologous with planthopper species. The novel miRNAs identified in this study provide a better understanding of the sRNA and the regulatory role of miRNA in sexual dimorphism and alteration in the expression or function of miRNAs in the rice pest.


Assuntos
Hemípteros/metabolismo , MicroRNAs/metabolismo , Animais , Sequência Conservada , Evolução Molecular , Feminino , Hemípteros/genética , Hemípteros/crescimento & desenvolvimento , Masculino , MicroRNAs/genética , Oryza/parasitologia , Análise de Sequência de RNA , Caracteres Sexuais
10.
BMC Genomics ; 19(1): 688, 2018 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-30231855

RESUMO

BACKGROUND: Sogatella furcifera is a migratory pest that damages rice plants and causes severe economic losses. Due to its ability to annually migrate long distances, S. furcifera has emerged as a major pest of rice in several Asian countries. Symbiotic relationships of inherited bacteria with terrestrial arthropods have significant implications. The genus Cardinium is present in many types of arthropods, where it influences some host characteristics. We present a report of a newly identified strain of the bacterial endosymbiont Cardinium cSfur in S. furcifera. RESULT: From the whole genome of S. furcifera previously sequenced by our laboratory, we assembled the whole genome sequence of Cardinium cSfur. The sequence comprised 1,103,593 bp with a GC content of 39.2%. The phylogenetic tree of the Bacteroides phylum to which Cardinium cSfur belongs suggests that Cardinium cSfur is closely related to the other strains (Cardinium cBtQ1 and cEper1) that are members of the Amoebophilaceae family. Genome comparison between the host-dependent endosymbiont including Cardinium cSfur and free-living bacteria revealed that the endosymbiont has a smaller genome size and lower GC content, and has lost some genes related to metabolism because of its special environment, which is similar to the genome pattern observed in other insect symbionts. Cardinium cSfur has limited metabolic capability, which makes it less contributive to metabolic and biosynthetic processes in its host. From our findings, we inferred that, to compensate for its limited metabolic capability, Cardinium cSfur harbors a relatively high proportion of transport proteins, which might act as the hub between it and its host. With its acquisition of the whole operon related to biotin synthesis and glycolysis related genes through HGT event, Cardinium cSfur seems to be undergoing changes while establishing a symbiotic relationship with its host. CONCLUSION: A novel bacterial endosymbiont strain (Cardinium cSfur) has been discovered. A genomic analysis of the endosymbiont in S. furcifera suggests that its genome has undergone certain changes to facilitate its settlement in the host. The envisaged potential reproduction manipulative ability of the new endosymbiont strain in its S. furcifera host has vital implications in designing eco-friendly approaches to combat the insect pest.


Assuntos
Proteínas de Bactérias/genética , Cytophagaceae/fisiologia , Genoma Bacteriano , Hemípteros/genética , Hemípteros/microbiologia , Simbiose/fisiologia , Animais , Genômica , Hemípteros/crescimento & desenvolvimento , Filogenia
12.
Gigascience ; 6(1): 1-9, 2017 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-28369349

RESUMO

Background: Sogatella furcifera is an important phloem sap-sucking and plant virus-transmitting migratory insect of rice. Because of its high reproductive potential, dispersal capability and transmission of plant viral diseases, S. furcifera causes considerable damage to rice grain production and has great economical and agricultural impacts. Comprehensive studies into ecological aspects and virus-host interactions of S. furcifera have been limited because of the lack of a well-assembled genome sequence. Findings: A total of 241.3 Gb of raw reads from the whole genome of S. furcifera were generated by Illumina sequencing using different combinations of mate-pair and paired-end libraries from 17 insert libraries ranging between 180 bp and 40 kbp. The final genome assembly (0.72 Gb), with average N50 contig size of 70.7 kb and scaffold N50 of 1.18 Mb, covers 98.6 % of the estimated genome size of S. furcifera . Genome annotation, assisted by eight different developmental stages (embryos, 1 st -5 th instar nymphs, 5-day-old adults and 10-day-old adults), generated 21 254 protein-coding genes, which captured 99.59 % (247/248) of core CEGMA genes and 91.7 % (2453/2675) of BUSCO genes. Conclusions: We report the first assembled and annotated whole genome sequence and transcriptome of S. furcifera . The assembled draft genome of S. furcifera will be a valuable resource for ecological and virus-host interaction studies of this pest.


Assuntos
Genoma de Inseto , Hemípteros/genética , Análise de Sequência de DNA , Transcriptoma , Animais , Feminino , Hemípteros/crescimento & desenvolvimento , Masculino , Anotação de Sequência Molecular
13.
Sci Rep ; 6: 36254, 2016 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-27805032

RESUMO

Sogatella furcifera, the white-backed planthopper (WBPH), has become one of the most destructive pests in rice production owing to its plant sap-sucking behavior and efficient transmission of Southern rice black-streaked dwarf virus (SRBSDV) in a circulative, propagative and persistent manner. The dynamic and complex SRBSDV-WBPH-rice plant interaction is still poorly understood. In this study, based on a homology-based genome-wide analysis, 348 immune-related genes belonging to 28 families were identified in WBPH. A transcriptome analysis of non-viruliferous (NVF) and viruliferous groups with high viral titers (HVT) and median viral titers (MVT) revealed that feeding on SRBSDV-infected rice plants has a significant impact on gene expression, regardless of viral titers in insects. We identified 278 up-regulated and 406 down-regulated genes shared among the NVF, MVT, and HVT groups and detected significant down-regulation of primary metabolism-related genes and oxidoreductase. In viruliferous WBPH with viral titer-specific transcriptome changes, 1,906 and 1,467 genes exhibited strict monotonically increasing and decreasing expression, respectively. The RNAi pathway was the major antiviral response to increasing viral titers among diverse immune responses. These results clarify the responses of immune genes and the transcriptome of WBPH to SRBSDV and improve our knowledge of the functional relationship between pathogen, vector, and host.


Assuntos
Hemípteros/genética , Hemípteros/virologia , Sistema Imunitário/imunologia , Reoviridae/fisiologia , Transcriptoma , Animais , Regulação da Expressão Gênica de Plantas , Ontologia Genética , Hemípteros/imunologia , Interações Hospedeiro-Patógeno , Imunidade/genética , Imunidade/imunologia , Insetos Vetores/genética , Insetos Vetores/imunologia , Insetos Vetores/virologia , Oryza/genética , Oryza/parasitologia , Oryza/virologia , Doenças das Plantas/genética , Doenças das Plantas/parasitologia , Doenças das Plantas/virologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Carga Viral
14.
J Immunol ; 197(8): 3198-3213, 2016 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-27647835

RESUMO

Scavenger receptor class A member 5 (SCARA5) and high-mobility group box 1 (HMGB1) protein have become increasingly attractive for their critical functions in innate inflammatory reactions and disorders. However, the functional relevance between these two molecules has never been described. This study discovered that SCARA5 is an HMGB1 recognition receptor that is negatively involved in HMGB1-mediated inflammation in pufferfish (Tetraodon nigroviridis) and zebrafish (Danio rerio) models. Hence, SCARA5 is added as a new member to the HMGB1 receptor family. Tetraodon HMGB1 (TnHMGB1) is a trafficking protein that can be secreted from the nucleus to the outside of cells upon CpG-oligodeoxynucleotide (ODN) stimulation. This protein exerts a strong synergistic effect on CpG-ODN-induced inflammation, as determined by the enhanced proinflammatory cytokine expression through coadministration of TnHMGB1 with CpG-ODN and impaired inflammatory responses through TnHMGB1 depletion. Tetraodon SCARA5 (TnSCARA5) is an inducible protein detected upon TnHMGB1 stimulation; this protein plays an inhibitory role in CpG-ODN-induced inflammation because TnSCARA5 overexpression suppresses cell responsiveness to CpG-ODN induction, whereas TnSCARA5 ablation intensifies the inflammatory reactions. TnSCARA5 can strongly associate with TnHMGB1 through the A and B boxes, depending on the redox state of the cysteine residues, but T box inhibits the association. TnSCARA5 mediates the endocytosis of TnHMGB1 into lysosomes. Results suggest that TnSCARA5 inhibits the CpG-ODN-mediated inflammation via the clearance of HMGB1 mediator for CpG-ODN stimulant. The above findings highlight a novel regulatory mechanism underlying innate inflammation and provide new insights into the clinical treatment of HMGB1-mediated diseases.


Assuntos
Proteína HMGB1/metabolismo , Inflamação/metabolismo , Receptores Depuradores Classe A/metabolismo , Tetraodontiformes/metabolismo , Peixe-Zebra/metabolismo , Animais , Clonagem Molecular , Modelos Animais de Doenças , Proteína HMGB1/genética
15.
J Immunol ; 197(1): 151-67, 2016 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-27206770

RESUMO

Single Ig IL-1R-related molecule (SIGIRR, also called IL-1R8 or Toll/IL-1R [TIR]8), a negative regulator for Toll/IL-1R signaling, plays critical roles in innate immunity and various diseases in mammals. However, the occurrence of this molecule in ancient vertebrates and its function in liver homeostasis and disorders remain poorly understood. In this study, we identified a SIGIRR homology from zebrafish (Danio rerio [DrSIGIRR]) by using a number of conserved structural and functional hallmarks to its mammalian counterparts. DrSIGIRR was highly expressed in the liver. Ablation of DrSIGIRR by lentivirus-delivered small interfering RNA in the liver significantly enhanced hepatic inflammation in response to polyinosinic-polycytidylic acid [poly(I:C)] stimulation, as shown by the upregulation of inflammatory cytokines and increased histological disorders. In contrast, depletion of TIR domain-containing adaptor inducing IFN-ß (TRIF) or administration of TRIF signaling inhibitor extremely abrogated the poly(I:C)-induced hepatic inflammation. Aided by the zebrafish embryo model, overexpression of DrSIGIRR in vivo significantly inhibited the poly(I:C)- and TRIF-induced NF-κB activations; however, knockdown of DrSIGIRR promoted such activations. Furthermore, pull-down and Duolink in situ proximity ligation assay assays showed that DrSIGIRR can interact with the TRIF protein. Results suggest that DrSIGIRR plays an inhibitory role in TRIF-mediated inflammatory reactions by competitive recruitment of the TRIF adaptor protein from its TLR3/TLR22 receptor. To our knowledge, this study is the first to report a functional SIGIRR homolog that existed in a lower vertebrate. This molecule is essential to establish liver homeostasis under inflammatory stimuli. Overall, the results will enrich the current knowledge about SIGIRR-mediated immunity and disorders in the liver.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Proteínas de Peixes/metabolismo , Inflamação/imunologia , Fígado/imunologia , Receptores de Interleucina-1/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/imunologia , Proteínas Adaptadoras de Transporte Vesicular/genética , Animais , Células Cultivadas , Citocinas/metabolismo , Proteínas de Peixes/genética , Imunidade Inata , Mediadores da Inflamação/metabolismo , Fígado/patologia , Mamíferos , NF-kappa B/metabolismo , Poli I-C/imunologia , RNA Interferente Pequeno/genética , Receptores de Interleucina-1/genética , Transdução de Sinais/genética , Receptor 3 Toll-Like/metabolismo , Proteínas de Peixe-Zebra/genética
16.
Biosci Rep ; 36(3)2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27129293

RESUMO

CCL2-mediated macrophage infiltration in articular tissues plays a pivotal role in the development of the osteoarthritis (OA). miRNAs regulate the onset and progression of diseases via controlling the expression of a series of genes. How the CCL2 gene was regulated by miRNAs was still not fully elucidated. In the present study, we demonstrated that the binding sites of miR-33 in the 3'UTR of CCL2 gene were conserved in human, mouse and rat species. By performing gain- or loss-of-function studies, we verified that miR-33 suppressed CCL2 expression in the mRNA and protein levels. We also found that miR-33 suppressed the CCL2 levels in the supernatant of cultured primary mouse chondrocytes. With reporter gene assay, we demonstrated that miR-33 targeted at AAUGCA in the 3'UTR of CCL2 gene. In transwell migration assays, we demonstrated that the conditional medium (CM) from miR-33 deficient chondrocytes potentiated the monocyte chemotaxis in a CCL2 dependent manner. Finally, we demonstrated that the level of miR-33 was decreased, whereas the CCL2 level was increased in the articular cartilage from the OA patients compared with the control group. In summary, we identified miR-33 as a novel suppressor of CCL2 in chondrocytes. The miR-33/CCL2 axis in chondrocytes regulates monocyte chemotaxis, providing a potential mechanism of macrophage infiltration in OA.


Assuntos
Quimiocina CCL2/genética , Condrócitos/metabolismo , Regulação para Baixo , MicroRNAs/genética , Osteoartrite/genética , Regiões 3' não Traduzidas , Animais , Cartilagem Articular/metabolismo , Cartilagem Articular/patologia , Células Cultivadas , Condrócitos/patologia , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Osteoartrite/patologia , Ratos
18.
Clin Hemorheol Microcirc ; 62(2): 129-37, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26444591

RESUMO

BACKGROUND: Although numerous risk factors for arteriovenous fistulae (AVF) dysfunction have been identified, these risk factors do not explain all cases of AVF dysfunction. Because of the importance of blood pressure variability (BPV) in vascular injury, the predictive value of BPV for AVF dysfunction, was evaluated in this prospective cohort study. METHODS: Twenty-four-hour BP monitoring at the intervals of dialysis was recorded every 3 months in 137 patients. The expression of smooth muscle actin (SMA) and the infiltration of mononuclear cells and T lymphocytes were determined by immunohistochemistry on the specimens of fistula vessels. RESULTS: Eighteen patients developed AVF dysfunction. Cox proportional hazards multivariate analysis revealed a significant relationship between fistula dysfunction and daytime systolic-BPV (d-SBPV), nighttime systolic-BPV (n-SBPV), diabetes mellitus, and initial venous diameter. Patients with AVF dysfunction were observed to have increased SMA expression and more infiltration of inflammatory cells in venous walls compared with the controls. A significant correlation between SBPV and the infiltration of CD68-positive cells was observed. CONCLUSIONS: Our study showed that the degrees of SBPV were significantly associated with the risk of AVF dysfunction. Potentially, the increase of SBPV will aggravate venous wall inflammation and may play a role in AVF dysfunction.


Assuntos
Derivação Arteriovenosa Cirúrgica , Pressão Sanguínea , Diálise Renal/métodos , Actinas/química , Adulto , Idoso , Antígenos CD/química , Antígenos de Diferenciação Mielomonocítica/química , Fístula Arteriovenosa/etiologia , Monitorização Ambulatorial da Pressão Arterial , Feminino , Humanos , Imuno-Histoquímica , Inflamação , Falência Renal Crônica/fisiopatologia , Falência Renal Crônica/cirurgia , Leucócitos Mononucleares/citologia , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Modelos de Riscos Proporcionais , Estudos Prospectivos , Fatores de Risco , Linfócitos T/citologia , Veias
19.
Mol Cell Biol ; 34(6): 989-1002, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24396069

RESUMO

Although epigenetic modulation is critical for a variety of cellular activities, its role in erythropoiesis remains poorly understood. Ten-eleven translocation (TET) molecules participate in methylcytosine (5mC) hydroxylation, which results in DNA demethylation in several biological processes. In this research, the role of TETs in erythropoiesis was investigated by using the zebrafish model, where three TET homologs were identified. These homologs share conserved structural domains with their mammalian counterparts. Zebrafish TETs mediate the conversion of 5mC to hydroxymethylcytosine (5hmC) in zebrafish embryos, and the deletion of TET2 inhibits erythropoiesis by suppressing the expression of the scl, gata-1, and cmyb genes. TET2-upregulated lineage-specific genes and erythropoiesis are closely associated with the occurrence of 5hmC and demethylation in the intermediate CpG promoters (ICPs) of scl, gata-1, cmyb, which frequently occur at specific regions or CpG sites of these ICPs. Moreover, TET2 regulates the formation and differentiation of erythroid progenitors, and deletion of TET2 leads to erythrocyte dysplasia and anemia. Here, we preliminarily proved that TET2 plays an essential role in erythrocyte development by regulating lineage-specific genes via DNA oxidative demethylation. This report is anticipated to broaden current information on hematopoiesis and pathogenesis of hematopoiesis-related diseases.


Assuntos
Linhagem da Célula/genética , Metilação de DNA/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Eritropoese/genética , Regulação da Expressão Gênica/genética , Peixe-Zebra/genética , Animais , DNA/genética , Células Precursoras Eritroides/metabolismo
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