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1.
Exp Eye Res ; 193: 107982, 2020 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-32092288

RESUMO

As the peroxisome proliferator - activated receptor alpha (PPARα) agonist, fenofibrate has been widely used to be a good lipid-regulating drug in the clinical application. In this study, we investigated the mechanism by which keratocytes inhibit the corneal neovascularization (CNV) through PPARα - activation. To do this, the CNV model was established by alkali burn, followed by being divided into three groups including control, fenofibrate and vehicle group. The expression of VEGFr3, MMP13 and PPARα in corneas of normal mouse and alkali-burned mouse was determined via quantitative RT- PCR (qRT-PCR) and Western blot analysis (WB). The CNV area was observed under a slit lamp microscope. The location of PPARα expression in the corneas was determined via immunohistochemistry. In cultured primary keratocytes, the effect of fenofibrate on PPARα, VEGFr3 and MMP13 expression was determined by qRT-PCR and WB. Besides, PPARα knockout (PPARα-/-) mouse CNV and keratocytes model were established to further confirm the effect of PPARα on VEGFr3 and MMP13 expression. We found that PPARα was expressed in epithelium, stroma and endothelium of the normal cornea, however, with relatively low level in the corneal stroma. Meanwhile, its expression was decreased markedly in the cornea during the stage of CNV formation. After treatment of fenofibrate, PPARα expression was promoted and the expression of VEGFr3 and MMP13 was inhibited in both CNV mice model and primary keratocytes, and CNV areas were decreased in CNV mice model. However, the results in PPARα-/- CNV and keratocytes model were opposite. Our results suggest that keratocytes could promote the expression of VEGFr3 and MMP13, and CNV formation through PPARα downregulation.

2.
Orphanet J Rare Dis ; 15(1): 49, 2020 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-32059734

RESUMO

BACKGROUND: RPE65-associated LCA (RPE65-LCA) is an inherited retinal degeneration caused by the mutations of RPE65 gene and gene therapy has been developed to be a promising treatment. This study aims to evaluate the association between changes in visual function and application of gene therapy in patients with RPE65-LCA. METHODS: Several databases (PubMed, Cochrane Library, and Web of Science) were searched for results of studies describing efficacy of gene therapy in patients with RPE65-LCA. Six studies, which included one randomized and five prospective non-randomized clinical trials, 164 eyes met our search criteria and were assessed. RESULTS: The BCVA significantly improved in treated eyes at 1 yr post treatment by - 0.10 logMAR (95% CI, - 0.17 - -0.04; p = 0·002), while there was no significant difference at 2-3 years post treatment (WMD: 0.01; 95% CI, - 0.00 - 0.02; p = 0·15). FST sensitivity to blue flashes also improved by 1.60 log (95% CI, 0.66-2.55; p = 0.0009), but no significant difference to red flashes (WMD: 0.86; 95% CI, - 0·29-2.01; p = 0.14) at 1 yr. There was no significant difference in central retinal thickness at 1 yr, but central retina in treated eyes appeared thinner at 2-3 years post treatment by 19.21 µm (95% CI, - 34.22 - -4.20; p = 0.01). CONCLUSIONS: Human gene therapy is a pioneering treatment option for RPE65-LCA. Although its efficacy appears to be limited to less than 2 yrs after treatment, it carries the potential for further improvement and prolongation of efficacy.

3.
Genomics ; 112(3): 2369-2378, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31945464

RESUMO

Strawberry fruit ripening is a complex process affected by multiple factors at different regulation levels. To elucidate the regulation mechanisms, the combined analysis of sRNAome and transcriptome were used. A total of 124 known and 190 novel miRNAs were found, 62 of them were significantly differentially expressed (DE). The targets of the DE miRNAs were parsed and several TFs, such as SPL, ARF, WRKY, and TCP, were found to be involved in ripening. Elevated CO2 can significantly postpone ripening and miR156, miR166f, miR171a, and miR171d were the DE miRNAs. Transcriptome analysis found 313 DE genes related to fruit ripening, including cell wall metabolism-related genes, color-related genes, ethylene-related genes, and genes encoding TFs such as MYB, SPL, NAC, TCP, and ARF. Based on above, a combined regulatory model involved in fruit ripening was created. These results provide valuable information for understanding the complicated coordinated regulatory network of strawberry fruit ripening.

4.
Biomed Pharmacother ; 122: 109473, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31918263

RESUMO

The outcome of current cancer therapy is usually impeded by complicated extracellular and intracellular barriers. Most importantly, untargeted distribution and multidrug resistance (MDR) are considered as two important difficulties responsible for the poor performance of many currently available drug delivery systems (DDS). As a result, in our study, we developed a cancer cell membrane (CM) coated calcium carbonate (CC) nanoparticles to co-delivery miR-451 with adriamycin (Adr) to address the dilemma occurred in the therapy of bladder cancer (MCC/R-A). The homologous CCM from MDR bladder cancer cells (BIU-87/Adr) was employed to increase targeted retention of DDS within the tumor tissue and to bypass the extracellular barriers. Moreover, the MDR of cancer cells was conquered through downregulation of P-gp expression using miR-451 since it was confirmed by previous reports that miR-451 could significantly downregulate the level of P-gp in MDR cells, which in turn elevated the cellular drug retention in BIU-87/Adr. Our in vitro and in vivo experiments have revealed that MCC/R-A showed a greatly enhanced therapeutic effect on BIU-87/Adr, which was superior than applying miR-451 or Adr alone. The preferable effect of MCC/R-A on conquering the MDR in bladder cancer provides a novel alternative for effective chemotherapy of MDR cancers.

5.
Biomed Pharmacother ; 122: 109557, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31918265

RESUMO

Prostate cancer (PCa) is a destructive malignancy with a bad prognosis. LncRNA VPS9D1-AS1 has recently been delineated as an oncogene in some kinds of tumor, whereas, the function of VPS9D1-AS1 in PCa remains to be clarified. In this study, we researched its underlying role in PCa. The expression of VPS9D1-AS1 was conspicuously upregulated in PCa tissues and cells. And absence of VPS9D1-AS1 inhibited cell proliferation, migration and invasion, and promoted cell apoptosis in PCa. In addition, VPS9D1-AS1 overexpression led to opposite results. Furthermore, VPS9D1-AS1/MEF2D could sponge with miR-4739. VPS9D1-AS1/MEF2D and miR-4739 were inversely correlated in tumor cells. And the expression of miR-4739 is markedly downregulated in PCa, meanwhile, that of MEF2D exhibited the opposite tendency. However, MEF2D was positively regulated by VPS9D1-AS1. Moreover, MEF2D upregulation offset the suppressive effects of VPS9D1-AS1 deficiency on cell proliferation, migration and invasion in PCa. Additionally, ZEB1 contained the binding sites of VPS9D1-AS1 promoter, and there existed positive relation between them. Taken together, above results illustrated that ZEB1 activated-VPS9D1-AS1 promotes the tumorigenesis and progression of PCa by sponging miR-4739 to upregulate MEF2D, which offering a new useful reference for studying the development process of PCa.

6.
J Gene Med ; : e3157, 2020 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-31901177

RESUMO

BACKGROUND: Use of chimeric antigen receptor (CAR) T cells has become a promising strategy in cancer immunotherapy. However, safety in clinical application is also one of the most controversial issues. METHODS: In the present study, we investigated the application of a non-viral site-directed vector (CELiD [closed-ended linear duplex DNA]) dependent on adeno-associated virus (AAV) genomes for the purpose of safe CAR-T engineering. We co-electroporated CD19-CAR encoding "CELiD" vectors with plasmid pCMV-Rep into human T cells and ensured stably transfected CAR-T cells by G418 selection. The efficiency of AAVS1 site-specific integration was analyzed by a real-time polymerase chain reaction. RESULTS: CAR-T cells engineered by CELiD vectors could be established within 20 days with up to 22.8% AAVS1 site-specific integration efficiency. CAR expression and cytokine secretion of CAR modified T cells were evaluated in vitro. Abundant effector cytokines were produced by the CAR-T cells engineered by CELiD vectors compared to control T cells and the killing efficiency of target cells was estimated to as high as 75% in vitro. CONCLUSIONS: With the help of the AAV-derived CELiD vector, CAR genes were preferentially integrated into the AAVS1 site. This technology could be utilized in human T cell modification and remove the safety constraints of CAR-T therapy.

7.
Gen Comp Endocrinol ; 287: 113357, 2020 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-31821794

RESUMO

Growth hormone is a hormone secreted from the pituitary and is involved in the regulation of most major physiological processes such as growth, development and metabolism. Therefore, an accurate and sensitive detection method is needed for the detection of tilapia serum Gh level. Phage display technology is widely used in the expression of antibody fragments, in which fragments of antibodies are expressed as a fusion with phage proteins and are displayed on the phage surface for easy screening. Time-resolved fluorescence immunoassay (TRFIA) is a microanalysis method developed nearly two decades ago and is one of the most sensitive analytical techniques. With the use of a special lanthanide, the detection background can be distinguished, which can greatly improve the sensitivity of detection. In this report, we cloned the VH and VL DNA fragments from the lymphocytes of rabbits immunized with recombinant Gh and assembled them with a linker to form a single-chain variable fragment (scFv) gene pool. Using phage display technology, we isolated scFv DNA fragments from the pool, which encode a protein that specifically binds to tilapia Gh. We then established Eu-DTTA-based TRFIA for measuring plasma Gh in tilapia. The sensitivity of double antibody sandwich Gh-TRFIA was 0.225 ng/ml, and the linear range of the standard curve was 0.225-250 ng/ml. The intra- and interassay coefficients of variation (CVs) were <9.1 and <4.5%, respectively. The cross-reactivities (CRs) of 1 µg/ml recombinant tilapia somatolactin (rtSl), prolactin (rtPrl) and thyroid-stimulating hormone beta subunit (rtTshb) were 0.042%, 0.472% and 0.036%, respectively. The sensitivity of direct competitive Gh-TRFIA was 0.208 ng/ml, and the linear range of the standard curve was 0.208-500 ng/ml. The intra- and interassay CVs were <4.8 and <7.1%, respectively. The CRs of 1 µg/ml rtSl, rtPrl and rtTshb were 0.041%, 0.079% and 0.073%, respectively. In conclusion, Gh-TRFIA is a safe (no concerns about radioactive isotopes), economical, and efficient detection method for the quantification of plasma Gh. Thus, the application of phage display technology for antibody screening and the use of TRFIA for tilapia Gh detection are conducive to research in the field of fish endocrinology.

8.
Curr Eye Res ; 45(1): 64-71, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31294618

RESUMO

Purpose: Adeno-associated virus vector (AAV) is the most accepted gene delivery vector for retinal gene therapy. Müller cells play an important role in maintaining homeostasis and neuronal structural integrity, stability and it has been found to be involved in many retinopathies. The aim of this study is to identify a rAAV2/6 mutant which has increased tropism for Müller cell of the mouse retina.Materials and Methods: Using amino acid mutagenesis, we created a rAAV2/6 capsid mutant, rAAV2/6-S663L. In vivo imaging and retinal flat mount were employed to analyze the gene expression of rAAV2/6-S663L and wt rAAV2/6 in mouse retinal tissue. Retinal tissue cryosection, immunohistochemistry (IHC), Müller cell-specific promoter-controlled gene expression, and double AAV fluorescent protein co-expression were performed to determine the targeting of rAAV2/6-S663L for mouse retinal Müller cells.Results: In vivo imaging, retinal flat mount and retinal tissue cryosection results showed that rAAV2/6-S663L and wt rAAV2/6 have different specific tropisms in mouse retina and rAAV2/6-S663L is more preferentially targeting Müller cells. Müller cell-specific promoter-controlled gene expression experiments and IHC test confirmed that rAAV2/6-S663L has a higher tendency to infect Müller cells than wt rAAV2/6. Co-infection of the mouse retina with one rAAV2/6-S663L expressing EGFP under the control of GFAP promoter and the other one expressing mCherry under the control of CMV promoter revealed co-expression of the two fluorescent proteins in Müller cells.Conclusions: The results confirmed that rAAV2/6-S663L has a higher tropism for Müller cells than wt rAAV2/6. Our findings could add a new useful tool for retinal disease gene therapy.

9.
Fish Shellfish Immunol ; 97: 1-11, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31846770

RESUMO

Deteriorating water quality, especially from high concentrations of nitrite, is currently largely blamed for disease outbreaks in farmed tilapia (Oreochromis niloticus). In this study, the underlying mechanism of nitrite on the susceptibility of tilapia leucocytes to Streptococcus agalactiae (S. agalactiae) was studied. We found that a high dose of heat-killed S. agalactiae decreased tilapia leucocytes cell viability, whereas nitrite decreased the cell viability of leucocytes exposed to a low dose of bacteria. Bacterial challenge increased the production of nitric oxide (NO), whereas nitrite and bacteria coexposure caused higher NO production than nitrite or bacterial exposure alone. Cell viability increased after elimination of NO, and negative correlations existed between cell viability and the NO content, suggesting that nitrite increased the susceptibility of the leucocytes against S. agalactiae was NO-dependent. For a more comprehensive understanding of the mechanism of nitrite affecting disease resistance in tilapia leucocytes, an RNA-Seq-based transcriptome was generated. The results showed that 6173 transcripts were differently expressed, and the differentially expressed transcripts (DETs) of the bacterial group, nitrite group and bacteria-nitrite co-treatment group compared to the control group were selected for GO and KEGG analyses. The DETs in the bacterial group and bacteria-nitrite cotreatment group were highly involved with the membrane component, signal transduction, and immune responses. KEGG analysis showed that the protein processing in the endoplasmic reticulum and the AMPK signaling pathway, which are related to autophagy, were significantly enriched in the cotreatment group but not in bacterial group. In addition, the mRNA expression of ten DETs and several autophagy and apoptosis related genes validated by q-PCR showed the high reliability of the RNA-seq. Taken together, the results of this study suggest that nitrite may increase the susceptibility of tilapia leucocytes to S. agalactiae by generating excess NO to affect the autophagy and apoptosis process.

10.
BMC Genomics ; 20(1): 919, 2019 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-31791229

RESUMO

BACKGROUND: Compensatory growth refers to the phenomenon in which organisms grow faster after the improvement of an adverse environment and is thought to be an adaptive evolution to cope with the alleviation of the hostile environment. Many fish have the capacity for compensatory growth, but the underlying cellular mechanisms remain unclear. In the present study, microarray and nontargeted metabolomics were performed to characterize the transcriptome and metabolome of zebrafish liver during compensatory growth. RESULTS: Zebrafish could regain the weight they lost during 3 weeks of fasting and reach a final weight similar to that of fish fed ad libitum when refed for 15 days. When refeeding for 3 days, the liver displayed hyperplasia accompanied with decreased triglyceride contents and increased glycogen contents. The microarray results showed that when food was resupplied for 3 days, the liver TCA cycle (Tricarboxylic acid cycle) and oxidative phosphorylation processes were upregulated, while DNA replication and repair, as well as proteasome assembly were also activated. Integration of transcriptome and metabolome data highlighted transcriptionally driven alterations in metabolism during compensatory growth, such as altered glycolysis and lipid metabolism activities. The metabolome data also implied the participation of amino acid metabolism during compensatory growth in zebrafish liver. CONCLUSION: Our study provides a global resource for metabolic adaptations and their transcriptional regulation during refeeding in zebrafish liver. This study represents a first step towards understanding of the impact of metabolism on compensatory growth and will potentially aid in understanding the molecular mechanism associated with compensatory growth.

11.
Front Genet ; 10: 1128, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31824559

RESUMO

The sex of Chinese tongue sole (Cynoglossus semilaevis) is determined by both genetic sex determination (GSD) and environmental sex determination (ESD), making it an ideal model to study the relationship between sex-determination and temperature. In the present study, transcriptomes of undifferentiated gonads from genetic females and males, as well as differentiated gonads from males, females, and pseudomales under high and normal temperature treatments were generated for comparative transcriptomic analysis. A mean of 68.24 M high-quality clean reads was obtained for each library. Differentially expressed genes (DEGs) between different sexes and environmental treatments were identified, revealing that the heat shock protein gene family was involved in the high temperature induced sex reversal. The Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways that were enriched in pseudomale and genetic female comparison included neuroactive ligand-receptor interaction, cortisol synthesis and secretion, and steroid hormone biosynthesis. Furthermore, weighted gene co-expression network analyses were conducted on all samples, and two modules were positive correlated with pseudomale under high temperature. An illustrated protein-protein interaction map of the module identified a hub gene, hsc70. These findings provide insights into the genetic network that is involved in sex determination and sexual differentiation, and improve our understanding of genes involved in sex reversal under high temperature.

12.
Ocul Immunol Inflamm ; : 1-6, 2019 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-31580185

RESUMO

Purpose: To summarize the prognostic factors of cytomegalovirus (CMV) retinitis (CMVR) in HIV-negative patients treated with multiple intravitreal injections (IVs) of ganciclovir. Methods: A retrospective cohort study (70 eyes) was conducted. Clinical signs, initial and final best corrected visual acuity (BCVA), initial aqueous load of CMV DNA, course of treatment, and occurrence of complications were recorded and analyzed. Results: A positive correlation was found between the baseline and the final best corrected visual acuity (P < .001) and between the initial aqueous CMV DNA load and the number of IVs (P = .01). A lesion close to the posterior pole (P < .001) and a larger retinal lesion (P = .002) remarkably led to worse visual prognosis. Conclusions: Poor visual prognosis was significantly associated with poor initial visual acuity, proximity of lesion to the posterior pole, and an extensive CMV lesion. The treatment duration was positively correlated with the initial aqueous CMV DNA load.

13.
Ocul Immunol Inflamm ; : 1-6, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31573356

RESUMO

Purpose: To describe and to compare the clinical manifestation and laboratory test results of herpetic anterior uveitis (HAU) caused by Herpes simplex virus (HSV) and varicella-zoster virus (VZV). Methods: A retrospective, observational study on patients diagnosed with HAU. Etiology, clinical features, ocular complications, and recurrences of the infection were evaluated as main clinical parameters. The aqueous Interleukin-8 (IL-8) level was also measured to assess the intraocular inflammation. Results: Thirty-two eyes (32 patients) were involved. Among all involved cases, 24 had VZV-AU and 8 had HSV-AU. Common clinical features of HAU included the presence of KPs (90.6%), distorted pupil (83.3%), Iris atrophy (71.9%) and corneal edema (50%). The intraocular fluid analysis showed higher viral load and IL-8 level in VZV-AU. Conclusions: Compared with HSV-AU, the intraocular inflammation was more severe in VZV-AU. The intraocular fluid analysis was valuable for the etiological diagnosis and the evaluation of disease severity.

14.
Life Sci ; : 116816, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31472148

RESUMO

Docetaxel is commonly used to treat hormone-refractory prostate cancer (HRPC), but its clinical efficacy is limited by drug resistance, with the molecular mechanisms remaining elusive. The E3 ubiquitin ligase EDD modifies substrate proteins through ubiquitination and is involved in the regulation of cell proliferation and tumorigenesis. However, its role in docetaxel resistance of prostate cancer is unknown. Here, we show that EDD is upregulated in docetaxel-resistant HRPC cells, as well as in human HRPC treated with docetaxel chemotherapy. Functionally, EDD knockdown resensitizes HRPC cells to docetaxel in vitro and in vivo, and in reverse, EDD overexpression promotes docetaxel resistance. We further show that the Wnt/ß-Catenin signaling is activated in docetaxel-resistant HRPC cells, which can be promoted by EDD. Finally, inhibiting Wnt signaling through ß-Catenin knockdown remarkably attenuates EDD-mediated docetaxel resistance, suggesting that the activated Wnt/ß-Catenin signaling is a key contributor to EDD-conferred docetaxel resistance in HRPC cells. Altogether, our study uncovers a positive role of EDD in docetaxel resistance in prostate cancer, and further links it with the regulation of Wnt/ß-Catenin signaling.

15.
Fish Shellfish Immunol ; 93: 99-107, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31323328

RESUMO

Epinephelus moara is an economically important fish in Southeast Asian countries but is suffering from nervous necrosis virus (NNV) infection. A deeper understanding of the host-NNV interaction mechanisms makes sense for disease control, however, at present, the pathogenesis of natural NNV infection and the resistance mechanism in host remains poorly understood. In this study, asymptomatic and diseased E. moara with clinical symptoms of viral nervous necrosis (VNN) from a grouper farm were both detected with a positive RT-PCR signal of NNV, then transcriptome sequencing of their immune tissues (liver, spleen and kidney) were performed for comparation analysis. The de novo assemblies yielded 53,789 unigenes which had a length varied from 201 to 19,675 bp and a N50 length of 2115 bp, and 29,451 unigenes were functionally annotated, with 83, 250 and 5632 unigenes being differentially expressed in liver, spleen and kidney respectively. KEGG pathway enrichment analysis of the DEGs showed many DEGs were enriched in immune related pathways. Although the expression of class I major histocompatibility complex (MHC) was significantly higher in three immune tissues of the diseased grouper, many immune related genes, including humoral immune molecules (such as antibodies), the cellular mediated cytotoxic molecules (such as perforin) and some adhesion related genes were down regulated in the diseased grouper. Our results provided many unigenes that might play important roles in NNV resistance for further research. Furthermore, a total of 8666 unigenes containing 11,623 simple sequence repeats (SSRs) were identified, which provided useful information for screening molecular markers associated with NNV resistance in E. moara.


Assuntos
Bass/genética , Bass/imunologia , Doenças dos Peixes/imunologia , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Infecções por Vírus de RNA/veterinária , Transcriptoma/imunologia , Animais , Doenças dos Peixes/genética , Perfilação da Expressão Gênica/veterinária , Nodaviridae/fisiologia , Infecções por Vírus de RNA/genética , Infecções por Vírus de RNA/imunologia
16.
Mol Ecol Resour ; 19(5): 1322-1332, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31230418

RESUMO

The giant grouper (Epinephelus lanceolatus) is the largest coral reef teleost, with a native range that spans temperate and tropical waters in the Pacific and the Indian Oceans. It is cultured artificially and used as a breeding species in aquaculture due to its rapid growth rate. Here we report a giant grouper genome assembled at the chromosome scale from sequences generated using Illumina and high-throughput chromatin conformation capture (Hi-C) technology. The assembly comprised 1.086 Gb, with 98.4% of the scaffold sequences anchored into 24 chromosomes. The contig and scaffold N50 values were 119.9 kb and 46.2 Mb, respectively. The assembly is of high integrity, including 96.4% universal single-copy orthologues based on BUSCO analysis. Through chromosome-scale evolution analysis, we identified alignments of six giant grouper chromosomes to three stickleback chromosomes and some of the genes located within the breakpoints of reshuffling events may related to development and growth. From the 24,718 protein-coding genes, we found that several gene families related to innate immunity and glycan biosynthesis were significantly expanded in the giant grouper genome compared to other teleost genomes. In addition, we identified several genes related to the hormone signalling pathway and innate immunity that have experienced positive selection or accelerated evolution, implicating their roles in immune defence and fast growth of the species. The high-quality genome assembly will provide a valuable genomic resource for further biological and evolutionary studies, and useful genomic tools for breeding of the giant grouper.


Assuntos
Bass/crescimento & desenvolvimento , Bass/genética , Cromossomos , Genoma , Imunidade Inata , Animais , Bass/imunologia , Biologia Computacional , Anotação de Sequência Molecular , Análise de Sequência de DNA
17.
Gen Comp Endocrinol ; 281: 137-144, 2019 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-31176753

RESUMO

The insulin-like growth factor (IGF) system plays a pivotal role in the regulation of growth, and IGF binding proteins (IGFBPs) are important regulatory factors in the IGF system. Generally, IGFBPs inhibit IGF actions by preventing its binding to receptors. Under some conditions, the IGFBPs can also enhance IGF actions. IGFBP1 is generally inhibitory to IGFI. In this study, the grouper (Epinephelus coioides) igfbp1 (MK621003) gene was cloned from the liver. The sequence of igfbp1 cDNA was 1055 bp and contained a 5'UTR of 127 bp and a 3'UTR of 247 bp, and the ORF of grouper igfbp1 was 741 bp, encoding 246 amino acids. The tissue distribution results showed that igfbp1 has a higher expression in the liver. In the nutritional status experiment, igfbp1 expression was significantly increased in the liver after 7 days of fasting and was markedly decreased after refeeding. In in vitro experiments, igfbp1 expression in grouper primary hepatocytes was significantly inhibited by recombinant grouper Gh (growth hormone) in a dose-dependent manner. Additionally, igfbp1 expression decreased in grouper primary hepatocytes upon incubation with insulin. This is the first report describing grouper igfbp1, and these findings contribute to understanding the roles of IGFBP1 in metabolism and growth in grouper.


Assuntos
Bass/genética , Hormônio do Crescimento/farmacologia , Hepatócitos/metabolismo , Proteína 1 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Insulina/farmacologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Células Cultivadas , DNA Complementar/genética , Feminino , Hepatócitos/efeitos dos fármacos , Proteína 1 de Ligação a Fator de Crescimento Semelhante à Insulina/química , Proteína 1 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Filogenia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Distribuição Tecidual
18.
Proteomics Clin Appl ; 13(5): e1900048, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31207145

RESUMO

PURPOSE: Early diagnosis is crucial to improve outcomes for pancreatic cancer patients (PC). The present study is designed to identify differently expressed peptides involved in PC as potential biomarkers. EXPERIMENTAL DESIGN: The serum proteome of 22 PC patients, 12 pancreatitis patients (PP), and 45 healthy controls (HC) are analyzed using magnetic bead-based weak cation exchange (MB-WCX) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Next, a supervised neural network (SNN) algorithm model is established by ClinProTools and the candidate biomarker identified using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Finally, the candidate biomarker is validated in tissue samples. RESULTS: The SNN algorithm model discriminates PC from HC with 92.97% sensitivity and 94.55% specificity. Seventy-six differentially expressed peptides are identified, seven of which are significantly different among PC, PP, and HC (p < 0.05). Only one peak (m/z: 1466.99) tends to be upregulated in samples from HC, PP, and PC, which is identified as region of RNA-binding motif protein 6 (RBM6). In subsequent tissue analysis, it is verified that RBM6 expression is significantly higher in PC tissues than paracancerous tissue. CONCLUSIONS AND CLINICAL RELEVANCE: The results indicate that RBM6 might serve as a candidate diagnostic biomarker for PC. CLINICAL RELEVANCE: Methods used in this study could generate serum peptidome profiles of PC, PP, and HC, and present an approach to identify potential biomarkers for diagnosis of this malignancy.

19.
Biomed Pharmacother ; 117: 109109, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31229922

RESUMO

Prostate carcinoma may develop into metastatic castration-resistant prostate carcinoma (mCRPC) after endocrine therapy. Exosomal microRNAs play an important role in the regulation of tumor microenvironment. Our study aimed to investigate the effect of exosomal miR-26a on tumor phenotype of prostate carcinoma. Low-grade prostate carcinoma cell line (LNCAP) and mCRPC cell line (PC-3) were treated as experimental subjects according to their miR-26a expressions. Wound healing, transwell and colony-forming unit assays were performed after miR-26a mimic/inhibitor transfection. Then, exosomes were isolated from LNCAP and PC-3 cells, and the levels of exosomal miR-26a were determined. After co-culture of LNCAP (PC-3) cells with PC-3 (LNCAP) exosomes, changes in malignant behaviors were measured. Moreover, LNCAP/PC-3 exosomes were injected into xenograft tumor mice to determine effects of the exosomes on tumorigenicity of LNCAP and PC-3 cells. MiR-26a showed a potently inhibitory effect on cell proliferation, migration and invasion of LNCAP and PC-3 cells. LNCAP exosomes had a higher miR-26a level, compared with PC-3 exosomes. Overexpression of miR-26a attenuated the enhanced malignant behavior of LNCAP cells induced by PC-3 exosomes, and miR-26a inhibition could reverse the inhibitory effects of LNCAP exosomes on PC-3 cells. Exosomal miR-26a could significantly alter the expressions of epithelial-mesenchymal transition (EMT)-related factors. Moreover, LNCAP exosomes suppressed the tumorigenicity of PC-3 cells, while PC-3 exosomes could promote the tumorigenicity of LNCAP cells. Our data suggest that exosomal miR-26a derived from prostate carcinoma cells had a suppressive effect on the metastasis and tumor growth of prostate carcinoma.


Assuntos
Exossomos/metabolismo , MicroRNAs/metabolismo , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Animais , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Transição Epitelial-Mesenquimal/genética , Exossomos/ultraestrutura , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , Invasividade Neoplásica , Metástase Neoplásica
20.
Int J Oncol ; 54(6): 1955-1968, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31081051

RESUMO

Studies have rarely been conducted on the role of miRNAs in prostate cancer (PCa) cell progression by directly targeting MTDH, at least to the best of our knowledge. Thus, the present study aimed to identify miRNAs closely related with metadherin (MTDH) and to determine their roles in PCa. For this purpose, the expression levels of MTDH in PCa tissues and cell lines were examined by RT­qPCR, immunohistochemistry and western blot analysis. By cell transfection, MTDH was either overexpressed in the normal prostate epithelial cell lines or silenced in tumor cell lines to determine cell viability, invasion and migration. Bioinformatics analysis, RT­qPCR, western blot analysis, dual­luciferase reporter assay and MTT assay were performed to identify direct the target of MTDH and to examine tumor cell viability. Rescue experiments using the PC­3 and LNCaP cells were carried out by MTT assay, scratch wound assay, Transwell assay, RT­qPCR and western blot analysis. Experiments were also conducted using 46 PCa human cancer and adjacent tissues, as wells as on 501 cases of PCa from the TCGA database. It was confirmed that the overexpression of MTDH was associated with a poor prognosis of patients. The overexpression of MTDH was found to promote the viability, invasion and migration of PCa cells. miR­145­5p and miR­145­3p identified from 16 miRNAs were found to be closely related to PCa and to be the targets of MTDH. Both these miRNAs were found to significantly suppress the growth and metastasis of PCa cells by negatively regulating the expression of MTDH. On the whole, the findings of this study demonstrate that MTDH functions as an oncogene in PCa and the inhibition of MTDH by miR­145­5p or miR­145­3p suppressed the growth and metastasis of PCa cells. The miR­145­5p/MTDH and miR­145­3p/MTDH pathways may thus become novel treatment targets for PCa.


Assuntos
Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , MicroRNAs/genética , Neoplasias da Próstata/metabolismo , Regiões 3' não Traduzidas , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Metástase Neoplásica , Estadiamento de Neoplasias , Prognóstico , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Análise de Sobrevida , Regulação para Cima
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