RESUMO
Charcoal-stripped fetal bovine serum (CS-FBS) is frequently used in studies on hormone-responsive cancers to provide hormone-free cell culture conditions. CS-FBS may influence the growth of cancer cells; however, the underlying mechanisms remain unclear. In this study, we aimed to clarify the effects of CS-FBS on distinct subtypes of breast cancer cells. We found that the crucial oncoprotein c-Myc was significantly inhibited in estrogen receptor alpha (ER-α)-positive breast cancer cells when cultured in CS-FBS-supplemented medium, but it was not suppressed in ER-α-negative cells. The addition of 17ß-estradiol (E2) to CS-FBS-supplemented medium rescued the CS-FBS-induced inhibition of c-Myc, while treatment with 5α-dihydrotestosterone (DHT) suppressed c-Myc expression. Our data demonstrated that CS-FBS may impede the growth of ER-α-positive breast cancer cells via c-Myc inhibition, and this was possibly due to the removal of estrogen. These results highlighted that the core drivers of c-Myc expression were subtype-specific depending on the distinct cell context and special caution should be exercised when using CS-FBS in studies of hormone-responsive cancer cells.
Assuntos
Neoplasias da Mama/patologia , Carvão Vegetal/farmacologia , Proteínas Proto-Oncogênicas c-myc/metabolismo , Soro/química , Animais , Neoplasias da Mama/genética , Bovinos , Linhagem Celular Tumoral , Di-Hidrotestosterona/farmacologia , Células Epiteliais/metabolismo , Estradiol/farmacologia , Receptor alfa de Estrogênio/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Receptores Androgênicos/metabolismo , Transcrição Gênica/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacosRESUMO
Cut-like homeobox 1 (CUX1) is a highly conserved homeoprotein that functions as a transcriptional repressor of genes specifying terminal differentiation. We previously showed that liver-specific microRNA-122 (miR-122) regulates the timing of liver development by silencing CUX1 post-transcriptionally. Since the CUX1 protein is expressed in a subset of embryonic tissues, we hypothesized that it is regulated by specific microRNAs (miRNAs) in each cell type during development. Using a large-scale screening method, we identified ten tissue-specific miRNAs from different cell lineages that directly targeted CUX1. An analysis of the interaction between heart-specific microRNA-208a (miR-208a) and CUX1 in the hearts of developing mouse embryos and in P19CL6 cells undergoing cardiac differentiation indicated that CUX1 is regulated by miR-208a during heart development and cardiomyocyte differentiation. Functional analysis of miR-208a in P19CL6 cells using lentiviral-mediated over-expression showed that it regulates the transition between cellular proliferation and differentiation. These results suggest that these tissue-specific miRNAs might play a common role in timing the progression of terminal differentiation of different cell lineages, possibly by silencing the differentiation repressor CUX1.
Assuntos
Diferenciação Celular , Linhagem da Célula/genética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Homeodomínio/antagonistas & inibidores , MicroRNAs/genética , Miócitos Cardíacos/citologia , Proteínas Nucleares/antagonistas & inibidores , Proteínas Repressoras/antagonistas & inibidores , Animais , Proliferação de Células , Células Cultivadas , Células HeLa , Coração/crescimento & desenvolvimento , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Miócitos Cardíacos/metabolismo , Especificidade de Órgãos , Fatores de TranscriçãoRESUMO
27-hydroxycholesterol (27-HC), the most abundant metabolite of cholesterol, is a risk factor for breast cancer. It can increase the proliferation of breast cancer cells and promote the metastasis of breast tumours in mouse models. Myc is a critical oncoprotein overexpressed in breast cancer. However, whether 27-HC affects Myc expression has not been reported. In the current study, we aimed to investigate the effects of 27-HC on Myc and the underlying mechanisms in MCF-7 breast cancer cells. Our data demonstrated that 27-HC activated Myc via increasing its protein stability. Three key negative modulators of Myc protein stability, PP2A, SCP1 and FBW7, were suppressed by 27-HC at the transcriptional level. We performed a data-mining analysis of the chromatin immunoprecipitation with next-generation DNA sequencing (ChIP-Seq) data in the ChIPBase, and discovered that a number of putative transcription factors (TFs), including Myc itself, were involved in the transcriptional regulation of PP2A, SCP1 and FBW7. Our results provide a novel mechanistic insight into the activation of Myc by 27-HC via transcriptional repression of PP2A, SCP1 and FBW7 to increase Myc protein stability in breast cancer cells.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Proteínas F-Box/metabolismo , Hidroxicolesteróis/metabolismo , Proteínas Nucleares/metabolismo , Fosfoproteínas Fosfatases/metabolismo , Proteína Fosfatase 2/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Proteína 7 com Repetições F-Box-WD , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Ativação TranscricionalRESUMO
BACKGROUND: The aim of this study was to investigate associations of 3 common polymorphisms in the VEGF gene, -2578C>A, -634C>G, and 936C>T, with risk of tetralogy of Fallot (TOF) in Chinese Han children. MATERIAL AND METHODS: From January 2010 to June 2013, a total of 400 pediatric subjects were recruited, including 160 cases with TOF (TOF group) and 240 healthy controls (control group). The genotypes of 3 common VEGF polymorphisms, -2578C>A, -634C>G, and 936C>T, were analyzed by polymerase chain reaction restriction fragment length polymorphism. All data were analyzed with SPSS 18.0 software. RESULTS: No significant differences were observed in body mass index or sex between TOF patients and controls (both P>0.05), but significant differences in age and family history of TOF were observed between the 2 groups (both P<0.05). The AA genotype in -2578C>A of VEGF was correlated with a significantly increased risk of TOF, and TOF risk in A allele carrier was 1.54-fold higher than that of C allele carrier (OR=1.54, 95%CI=1.14-2.09, P=0.005); the statistical significance was still present after Bonferroni correction (Pc=0.045). GG genotype in -634C>G of VEGF gene was also associated with an increased risk of TOF, and TOF risk in patients with G allele was 1.62-fold higher compared to patients with C allele (OR=1.62, 95%CI=1.19-2.21, P=0.002); the statistical significance was still present after Bonferroni correction (Pc=0.018). Interestingly, T allele in VEGF 936C>T polymorphism is associated with a decreased TOF risk (OR=0.65, 95%CI=0.49-0.87, P=0.003, the statistical significance was still present after Bonferroni correction (Pc=0.027). The result of logistic regression analysis revealed that -2578C>A, -634C>G, and 936C>T genotypes are independently related to the prevalence of TOF (all P<0.05). CONCLUSIONS: Our results confirmed that VEGF genetic polymorphisms, -2578C>A and -634C>G, may be associated with an increased TOF risk, while 936C>T polymorphism may be associated with decreased TOF risk.
Assuntos
Polimorfismo de Nucleotídeo Único , Tetralogia de Fallot/sangue , Tetralogia de Fallot/genética , Fator A de Crescimento do Endotélio Vascular/genética , Alelos , Biomarcadores/sangue , Criança , Pré-Escolar , China , Ecocardiografia Doppler , Feminino , Frequência do Gene , Genótipo , Heterozigoto , Humanos , Lactente , Desequilíbrio de Ligação , Masculino , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Fatores de Risco , Software , Tetralogia de Fallot/diagnóstico por imagem , Tetralogia de Fallot/cirurgia , Fator A de Crescimento do Endotélio Vascular/sangueRESUMO
This paper reports the occurrence of two new species of Digamasellidae from Taiwan, Dendroseius vulgaris n. sp. and Dendrolaelaps (Foveodendrolaelaps) linjianzheni n. sp. Dendroseius vulgaris is described based on the morphology of adult females, adult males and deutonymph, and D. linjianzheni is based on the morphology of adult females and males. This is the first report on the mite species of Digamasellidae from Taiwan.
Assuntos
Ácaros e Carrapatos/anatomia & histologia , Ácaros e Carrapatos/classificação , Animais , Feminino , Masculino , Solo , Especificidade da Espécie , TaiwanRESUMO
PURPOSE: We compared plasmakinetic resection with holmium laser enucleation of the prostate for the treatment of benign prostatic hyperplasia by analyzing 2-year followup data from a prospective randomized clinical trial. MATERIALS AND METHODS: A total of 280 patients were randomly treated with plasmakinetic resection or holmium laser enucleation of the prostate. Perioperative and postoperative outcome data were obtained during a 2-year followup. RESULTS: No significant differences between the 2 surgical groups were observed in the preoperative data. Both groups displayed significant improvements after surgery. However, we identified no significant differences between the 2 groups in the 2-year followup data for I-PSS (International Prostate Symptom Score), quality of life scores or maximum flow rate values. Patients in the holmium laser enucleation group displayed a lower risk of hemorrhage, shorter bladder irrigation and catheter times, and shorter hospital stays. A larger amount of prostate tissue was retrieved in the holmium laser enucleation group, but the operation time was longer for this group than for the plasmakinetic resection group. CONCLUSIONS: Plasmakinetic resection and holmium laser enucleation of the prostate are effective and safe treatments for benign prostatic hyperplasia. Holmium laser enucleation of the prostate can be applied to prostates of all sizes, and involves less risk of hemorrhage, decreased bladder irrigation and catheter times, as well as reduced hospital stay. Thus, we believe holmium laser enucleation of the prostate should be proposed as a potential new gold standard surgical therapy instead of transurethral resection of the prostate for patients with benign prostatic hyperplasia.
Assuntos
Lasers de Estado Sólido/uso terapêutico , Hiperplasia Prostática/cirurgia , Ressecção Transuretral da Próstata/métodos , Idoso , Seguimentos , Humanos , Masculino , Estudos Prospectivos , Método Simples-Cego , Fatores de TempoRESUMO
Stromal cells play a decisive role in regulating tumor progression. In this study, we assessed the significance of normal prostate-derived stromal cells (PSCs) in prostate cancer development. An in vivo s.c. tumor model was established as follows: Group 1, DU145 cells alone; Group 2, DU145 + PSCs; Group 3, DU145 cells alone injected into pre-castrated mice; and Group 4, DU145 + PSCs injected into pre-castrated mice. Following injection, tumors were only detectable in the first two groups, with more aggressive growth in Group 2 than in Group 1 (P < 0.05). Immunohistochemical analysis revealed significantly higher proliferation (P < 0.05), but not apoptosis or altered expression of androgen receptor in Group 2, as compared with Group 1. In vitro, DU145 cells isolated from Group 1 tumors showed lower viability and migratory capability than those from Group 2. cDNA microarray on isolated DU145 cells from Groups 1 and 2 revealed the differential expression of genes regulating cell cycle progression and cell mobility, including GADD45A, RHOV, KLK11, and PCK1. Our results suggest that stromal cells derived from normal prostate potentiate the development of tumor growth in vivo, which is achieved at least in part through the regulation of cell-cycle- and migration-related gene expression within the tumor cells.
Assuntos
Comunicação Celular , Próstata/citologia , Neoplasias da Próstata/patologia , Células Estromais/fisiologia , Animais , Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células , Células Cultivadas , Perfilação da Expressão Gênica , Masculino , Camundongos , Camundongos Nus , Transplante de Neoplasias , Neoplasias Hormônio-Dependentes/patologia , Neoplasias da Próstata/genéticaRESUMO
Giardia lamblia is an early diverging and evolutionarily successful protozoan as it can enter into a dormant cyst stage from a vegetative trophozoite. During dormant stage, its metabolic rate decreases dramatically. However, to date, the regulatory molecules participating in the initiation and maintenance of this process have not been fully investigated. In this study, we have identified a class of abundant small RNAs named sitRNAs, which are approximately 46 nucleotides in length and accumulate in G. lamblia encysting cultures. Remarkably, they are derived from the 3' portion of fully matured tRNAs by cleavage of the anticodon left arm, with the 3' terminal CCA triplex still connected. During differentiation, only a limited portion of mature tRNAs is cleaved, but this cleavage occurs almost in the entire tRNA family. sitRNAs begin to accumulate as early as 3 h after initiation of encystation and are maintained at a relatively stable level during the whole process, exhibiting an expression peak at around 24 hr. Our studies further show that sitRNAs can be induced by several other stress factors, and in the case of serum deprivation, both tRNAs and sitRNAs degrade rapidly, with the accumulation of tRNA being halved. Our results may provide new insight into a novel mechanism for stressed G. lamblia to regulate gene expression globally.
Assuntos
Giardia lamblia/genética , RNA de Protozoário/metabolismo , RNA de Transferência/química , RNA não Traduzido/metabolismo , Animais , Células Cultivadas , Meios de Cultura Livres de Soro , Giardia lamblia/crescimento & desenvolvimento , Giardia lamblia/metabolismo , RNA de Protozoário/química , RNA de Protozoário/classificação , RNA de Transferência/metabolismo , RNA não Traduzido/química , RNA não Traduzido/classificação , Temperatura , Trofozoítos/metabolismoRESUMO
Non-heading Chinese cabbage (NHCC) is an important leafy vegetable cultivated worldwide. Here, we report the first high-quality, chromosome-level genome of NHCC001 based on PacBio, Hi-C, and Illumina sequencing data. The assembled NHCC001 genome is 405.33 Mb in size with a contig N50 of 2.83 Mb and a scaffold N50 of 38.13 Mb. Approximately 53% of the assembled genome is composed of repetitive sequences, among which long terminal repeats (LTRs, 20.42% of the genome) are the most abundant. Using Hi-C data, 97.9% (396.83 Mb) of the sequences were assigned to 10 pseudochromosomes. Genome assessment showed that this B. rapa NHCC001 genome assembly is of better quality than other currently available B. rapa assemblies and that it contains 48,158 protein-coding genes, 99.56% of which are annotated in at least one functional database. Comparative genomic analysis confirmed that B. rapa NHCC001 underwent a whole-genome triplication (WGT) event shared with other Brassica species that occurred after the WGD events shared with Arabidopsis. Genes related to ascorbic acid metabolism showed little variation among the three B. rapa subspecies. The numbers of genes involved in glucosinolate biosynthesis and catabolism were higher in NHCC001 than in Chiifu and Z1, due primarily to tandem duplication. The newly assembled genome will provide an important resource for research on B. rapa, especially B. rapa ssp. chinensis.
RESUMO
AIM: To assess the variability of adhesin gene hpaA between different Helicobacter pylori (H pylori) strains with PCR-restriction fragment length polymorphism (RFLP). METHODS: Twelve different H pylori strains were chosen to amplify the 710-bp segments of gene hpaA. These strains were NCTC11637, SS1; Chongqing clinical isolates CCS9801, CCS9802, CCS9803, CCS9806, CCS9809, CCS9810, CCS9813, which were gained from patients of gastritis; Mongolia gerbil adapted H pylori strains (abbreviation MG), which were gained from the following steps: gastric mucosal specimens of Mongolia gerbils infected by clinical isolate CCS9803 were cultured and detected, the positive H pylori strains were named as the first generation of Mongolia gerbil adapted H pylori strains (abbreviation MG1) and then were subcultured with healthy Mongolia gerbil to generate MG2, in turn to gain the ninth generation (abbreviation MG9). All hpaA segments, obtained from 12 different H pylori strains, were digested by HhaI and HaeIII individually and analyzed by agarose gel electrophoresis. RESULTS: In all 12 strains, the 710-bp PCR products were successfully amplified and products were cloned to pMD18-T vector respectively, then the recombinant plasmids were digested simultaneously with NcoI and XhoI to recover the small fragments. The objective fragments from 12 different H pylori strains digested with Hae III could be seen as 4 types of bands and 5 types with Hha I. According to the hpaA RFLP patterns, the 12 H pylori strains could be divided into 5 groups: group I, NCTC11637 and SS1; group II, CCS9809, which RFLP type digested with HaeIII was the same as strains of group I, but HhaI RFLP showed difference compared with the other groups; group III, CCS9810; group IV, CCS9803; group V: CCS9801, CCS9802, CCS9806, CCS9813, MG1, MG3 and MG9. The sequence data of 12 hpaA segments were analyzed by DNAsis software and it was observed that: (1) The homologies of base pair and amino acid sequence between strains NCTC11637, SS1, CCS9809 were 99.6% and 98.9%, respectively; (2) The homology of base pair and amino acid sequence between CCS9803 and CCS9810 was 97.7% and 99.1%; (3) That of the rest strains, CCS9801, CCS9802, CCS9806, CCS9813, MG1, MG3, MG9 reached 99.4% and 98.4%; (4) The base pair homologies between all hpaA fragments of different sources were higher than 94.6%, therefore the correspondence of deduced amino acid sequence was higher than 96.8% between each other. CONCLUSION: The gene hpaA from different H pylori strains revealed variation, and this might provide an effective method for molecular epidemiological survey of H pylori.
Assuntos
Adesinas Bacterianas/genética , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Polimorfismo de Fragmento de Restrição , Animais , Sequência de Bases , China , Variação Genética , Gerbillinae , Humanos , Dados de Sequência Molecular , FilogeniaRESUMO
To report the surgical management, complications and prognosis of patients with penoscrotal extramammary Paget's disease (EMPD) at different clinical stages. Between 2003 and 2008, a total of 30 male patients with penoscrotal EMPD were enrolled and evaluated. All enrolled subjects received frozen biopsy-guided local wide resection and immediate reconstruction. Patients were followed every 3 months postoperatively. Among the 30 patients who accepted and underwent frozen biopsy-guided local wide resection treatment and reconstruction, two (6.7%) cases exhibited positive margins, verified by pathological examination, and underwent re-excision after surgery. The technique of primary closure or an adjacent flap was used in 10 (33.3%) cases, split-thickness skin grafts were used in 15 (50%), and an anterolateral thigh perforator flap was used in five cases (16.7%). The postoperative complications were acceptable. The mean follow-up time was 64.9 ± 29.6 months. Of all 30 cases, 22 patients (73.3%) survived with no evidence of recurrence, four patients (13.3%) exhibited local recurrence, two patients (6.7%) exhibited both local recurrence and distant metastasis and the remaining two patients (6.7%) exhibited distant metastasis. Five patients died from metastasis or cachexia. Current surgical techniques, including primary closure, adjacent flaps, split-thickness skin flaps and anterolateral thigh perforator flaps are able to reconstruct all types of defects with acceptable complications. Some patients with negative margins went on to exhibit local recurrence, potentially due to adnexal carcinoma or internal malignancy.
Assuntos
Neoplasias dos Genitais Masculinos/cirurgia , Doença de Paget Extramamária/cirurgia , Procedimentos de Cirurgia Plástica/métodos , Escroto/cirurgia , Idoso , Idoso de 80 Anos ou mais , Biópsia , Seguimentos , Gastrectomia , Neoplasias dos Genitais Masculinos/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/patologia , Estadiamento de Neoplasias , Doença de Paget Extramamária/patologia , Pênis/patologia , Pênis/cirurgia , Retalho Perfurante , Complicações Pós-Operatórias/patologia , Prognóstico , Prostatectomia , Escroto/patologia , Transplante de PeleRESUMO
Accumulating evidence has implicated the deregluation of miRNAs in tumorigenesis. Previous studies have reported that microRNA-195 (miR-195) is markedly down-regulated in human glioblastoma cells, compared with normal brain tissue, but the biological role of miR-195 in glioblastoma development is currently unknown. In this study, we define a tumor-suppressor role for miR-195 in human glioblastoma cells. Over-expression of miR-195 in glioblastoma cell lines robustly arrested cell cycle progression and significantly repressed cellular invasion. We identified E2F3 and CCND3 as functional downstream targets of miR-195 in glioblastoma cells. Through knockdown studies, we demonstrated that E2F3 was the dominant effector of miR-195-mediated cell cycle arrest and that CCND3 was a key mediator of miR-195-induced inhibition of glioblastoma cell invasion. Furthermore, we showed that p27(Kip1) was an important regulator downstream of CCND3 and that the accumulation of p27(Kip1) in the cytoplasm might be responsible for the miR-195-mediated cell invasion inhibition in glioblastoma cells. This work provides evidence for the initial mechanism by which miR-195 negatively regulates both the proliferation and invasion of glioblastoma cells, suggesting that the down-regulation of miR-195 might contribute to the malignant transformation of glioblastoma cells and could be a molecular signature associated with glioblastoma progression.
Assuntos
Neoplasias Encefálicas/genética , Pontos de Checagem do Ciclo Celular/genética , Glioblastoma/genética , MicroRNAs/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Proliferação de Células , Regulação para Baixo , Técnicas de Silenciamento de Genes , Genes Supressores de Tumor , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , MicroRNAs/genética , Invasividade Neoplásica , Transdução de Sinais/fisiologia , Regulação para CimaRESUMO
Deposition of collagen IV in proximal tubule cells (PTCs) plays an important role during diabetic nephropathy, but the mechanism underlying excessive production of collagen IV remains poorly understood. In this study, we examined the miRNA profile of HK-2 cells and found that high glucose/TGF-beta1 induced significant down-regulation of miR-29a. We then showed that miR-29a negatively regulated collagen IV by directly targeting the 3'UTRs of col4a1 and col4a2. These results suggest that miR-29a acts as a repressor to fine-tune collagen expression and that the reduction of miR-29a caused by high glucose may increase the risk of excess collagen deposition in PTCs.
Assuntos
Colágeno Tipo IV/metabolismo , Regulação para Baixo/efeitos dos fármacos , Glucose/farmacologia , Túbulos Renais Proximais/efeitos dos fármacos , MicroRNAs/genética , Regiões 3' não Traduzidas/genética , Northern Blotting , Western Blotting , Linhagem Celular , Colágeno Tipo IV/genética , Perfilação da Expressão Gênica , Humanos , Túbulos Renais Proximais/citologia , Túbulos Renais Proximais/metabolismo , Modelos Biológicos , Análise de Sequência com Séries de Oligonucleotídeos , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Fator de Crescimento Transformador beta1/farmacologiaRESUMO
BACKGROUND: MicroRNAs (miRNAs) are approximately 22-nt small non-coding regulatory RNAs that have generally been considered to regulate gene expression at the post-transcriptional level in the cytoplasm. However, recent studies have reported that some miRNAs localize to and function in the nucleus. METHODOLOGY/PRINCIPAL FINDINGS: To determine the number of miRNAs localized to the nucleus, we systematically investigated the subcellular distribution of small RNAs (sRNAs) by independent deep sequencing sequenced of the nuclear and cytoplasmic pools of 18- to 30-nucleotide sRNAs from human cells. We identified 339 nuclear and 324 cytoplasmic known miRNAs, 300 of which overlap, suggesting that the majority of miRNAs are imported into the nucleus. With the exception of a few miRNAs evidently enriched in the nuclear pool, such as the mir-29b, the ratio of miRNA abundances in the nuclear fraction versus in the cytoplasmic fraction vary to some extent. Moreover, our results revealed that a large number of tRNA 3' trailers are exported from the nucleus and accumulate in the cytoplasm. These tRNA 3' trailers accumulate in a variety of cell types, implying that the biogenesis of tRNA 3' trailers is conserved and that they have a potential functional role in vertebrate cells. CONCLUSION/SIGNIFICANCE: Our results provide the first comprehensive view of the subcellular distribution of diverse sRNAs and new insights into the roles of miRNAs and tRNA 3' trailers in the cell.