RESUMO
PURPOSE: This study aimed to identify a diagnostic panel of serum microRNAs (miRNAs) for the early detection of bladder cancer (BC). METHODS: Serum samples were collected from 112 BC patients and 112 normal controls (NCs). A three-stage selection was conducted to identify differentially expressed miRNAs as candidates to construct the diagnostic panel. Further, to explore their potential roles in urothelial BC, bioinformatics analyses, including target genes prediction and functional annotation, were used. RESULTS: Six downregulated miRNAs (miR-1-3p, miR-30a-5p, miR-100-5p, miR-125b-5p, miR-143-3p, and miR-200c-3p) and one upregulated, miR-182-5p, in BC patients' serum were detected compared to NCs and were selected to establish the diagnostic panel. Based on a backward stepwise logistic regression analysis, miR-125b-5p, miR-182-5p, and miR-200c-3p comprehended the diagnostic panel [area under the curve (AUC) = 0.959, sensitivity = 91.67%, specificity = 92.5%]. CONCLUSION: The panel of three miRNAs had an excellent diagnostic capability, representing a potential non-invasive method for early BC detection.
Assuntos
Carcinoma de Células de Transição , MicroRNAs/genética , Neoplasias da Bexiga Urinária , Biomarcadores , Biomarcadores Tumorais/genética , Detecção Precoce de Câncer , Feminino , Perfilação da Expressão Gênica/métodos , Humanos , Masculino , Neoplasias da Bexiga Urinária/diagnóstico , Neoplasias da Bexiga Urinária/genéticaRESUMO
BACKGROUND: The study examines the function of hypoxia-mediated down-regulation of microRNAs (miRNAs) (mir-30c, mir-135a, and mir-27a) in the process of bladder cancer immune escape. METHODS: Quantitative Real-time PCR (qRT-PCR) was carried out to determine gene expression levels of Drosha and Dicer under hypoxia treatment, while western blotting and flow cytometry were used to determine protein expression. Seven reported miRNAs were identified via qRT-PCR assay. Flow cytometry detection of CD3/CD4/CD8-positive expression and statistics. Enzyme-linked immunosorbent assay (ELISA) detected cellular immune factors content. Cell apoptosis was checked via flow cytometry assay. Luciferase report assay and western blot assays were both used to verify the relationship between miRNAs and Casitas B-lineage lymphoma proto-oncogene b (Cbl-b). The animal model was established and Hematoxylin-eosin (HE) staining, TdT-mediated dUTP Nick-End Labeling (TUNEL) staining, and immunohistochemistry (IHC) assays were separately used to verify the conclusions. RESULTS: The CD3 + /CD4 + expression was increased in the hypoxia group, while CD3 + /CD8 + expression, the cellular immune factors content Interleukin-2 (IL-2) and Tumor Necrosis Factor-α (TNFα) along with the cell apoptosis were suppressed. The protein expression of Cbl-b was found to be up-regulated in the hypoxia group. After constructing the overexpression/ knockdown of Cbl-b in peripheral blood mononuclear cell (PBMC), Cbl-b has been found to promote tumor immune escape in bladder cancer. Furthermore, Cbl-b had been identified as the co-targets of mir-30c, mir-135a, and mir-27a and down-regulation of miRNA biogenesis promotes Cbl-b expression and deactivating T cells in vitro/in vivo. CONCLUSION: Hypoxia-mediated down-regulation of miRNAs' biogenesis promotes tumor immune escape in bladder cancer, which could bring much more advance to the medical research on tumors.
Assuntos
Regulação para Baixo/imunologia , MicroRNAs/metabolismo , Evasão Tumoral/imunologia , Hipóxia Tumoral/imunologia , Neoplasias da Bexiga Urinária/imunologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Apoptose , Complexo CD3/metabolismo , Antígenos CD4/metabolismo , Antígenos CD8/metabolismo , RNA Helicases DEAD-box/genética , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica , Humanos , Interleucina-2/metabolismo , Leucócitos Mononucleares/metabolismo , Masculino , Camundongos Endogâmicos C57BL , MicroRNAs/biossíntese , Estudos Prospectivos , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas c-cbl/genética , Distribuição Aleatória , Ribonuclease III/genética , Fator de Necrose Tumoral alfa/metabolismo , Regulação para Cima , Neoplasias da Bexiga Urinária/metabolismoRESUMO
PURPOSE: Paeonol is a natural chemical medicine derived from the bark of peony root, which has been found to inhibit tumor activity in various tumor cell lines, and can play a synergistic anti-tumor effect with chemotherapy or radiotherapy. METHODS: We used paeonol to act on human bladder cancer T24 and 5637 cells, and established xenograft tumor in nude mice by subcutaneous injection of T24 cells. RESULTS: CCK-8 assay and plate cloning experiments showed that paeonol could inhibit the proliferation of T24 and 5637 cells in vitro. The results of flow cytometry and the detection of BAX, Bcl-2 and Caspase-3 proteins suggested that paeonol can induce apoptosis of T24 and 5637 cells in vitro. Tumor formation, TUNEL detection and immunohistochemical results of Ki67, BAX, Bcl-2 and Caspase-3 in nude mice showed that paeonol could inhibit T24 cell proliferation and induce apoptosis in vivo, thus inhibiting tumor growth. Further research revealed that paeonol could reduce phosphorylation expression of PI3K and AKT in T24 and 5637 cells. CONCLUSION: We confirmed that paeonol could inhibit proliferation and induce apoptosis of human bladder cancer T24 and 5637 cells in vitro and in vivo, inhibit the growth of T24 tumor-forming nude mice, and possibly play a role by inhibiting the PI3K/AKT signaling pathway, so as to provide a potential therapeutic drug for bladder cancer.
Assuntos
Acetofenonas/farmacologia , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias da Bexiga Urinária/patologia , Animais , Caspase 3/análise , Linhagem Celular Tumoral , Citometria de Fluxo , Humanos , Marcação In Situ das Extremidades Cortadas , Técnicas In Vitro , Antígeno Ki-67/análise , Camundongos , Camundongos Nus , Transplante de Neoplasias , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/análise , Neoplasias da Bexiga Urinária/química , Neoplasias da Bexiga Urinária/tratamento farmacológico , Ensaios Antitumorais Modelo de Xenoenxerto , Proteína X Associada a bcl-2/análiseRESUMO
BACKGROUND: Angiogenesis is an indispensable step in the growth and invasiveness of breast cancers involving a series of exquisite molecular steps. Pro-angiogenic factors, including vascular endothelial growth factor (VEGF), have been recognized as pivotal therapeutic targets in the treatment of breast cancer. More recently, a highly conserved transcription factor Twist has been reported to be involved in tumor angiogenesis and metastasis. METHODS: The expression of VEGF-C and Twist was immunohistochemically determined in tissue samples of primary tumors from 408 patients undergoing curative surgical resection for breast cancer. The correlations of VEGF-C and Twist expressions with clinicopathologic parameters as well as survival outcomes were evaluated. RESULTS: Of the 408 patients evaluated, approximately 70% had high expression of VEGF-C which was significantly associated with advanced tumor stages (P = 0.019). Similarly, VEGF-C expression was associated with the proliferation index Ki67, N3 lymph node metastasis, and D2-40-positive lymphatic vessel invasion (LVI) in a univariate analysis. Furthermore, patients with high expressions of VEGF-C and Twist (V + T+) had significantly increased lymph node metastasis, higher clinical stage, and worse disease-free survival, DFS (P = 0.001) and overall survival, OS (P = 0.011). CONCLUSIONS: Our results suggested that co-expression of VEGF-C and Twist was associated with larger tumor size, higher numbers of lymph node involvement, D2-40-positive LVI, higher risk of distant metastasis, and worse DFS or OS in breast cancer patients.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/patologia , Carcinoma Ductal de Mama/patologia , Proteínas Nucleares/metabolismo , Proteína 1 Relacionada a Twist/metabolismo , Fator C de Crescimento do Endotélio Vascular/metabolismo , Neoplasias da Mama/metabolismo , Neoplasias da Mama/cirurgia , Carcinoma Ductal de Mama/metabolismo , Carcinoma Ductal de Mama/cirurgia , Feminino , Seguimentos , Regulação Neoplásica da Expressão Gênica , Humanos , Pessoa de Meia-Idade , Prognóstico , Taxa de SobrevidaRESUMO
OBJECTIVE: This study aimed to investigate the pure prognostic role of epidermal growth factor receptor (EGFR) mutation status and subtype in lung adenocarcinoma patients irrespective of therapy. MATERIALS AND METHODS: We retrospectively enrolled 119 cases of completely resected pathological stage I lung adenocarcinoma patients who received no postoperative chemotherapy or tyrosine kinase inhibitors. EGFR gene mutations from 18 to 21 exons were tested for all the patients. Disease-free survival (DFS) and overall survival (OS) were compared between patients with different EGFR mutation status and subtype using Kaplan-Meier methods. RESULTS: EGFR mutations were detected in 54 (45.4%) patients including two common mutation subtypes: 32 in-frame deletion within exon 19 (19del) and 19 point mutation within exon 21 (L858R). The frequency of EGFR mutations was much greater for patients of non-smokers versus current or ever smokers (58.1 versus 24.4%, P = 0.000), and a little greater for females versus males (53.8 versus 35.2%, P = 0.042). The median follow-up duration was 43.5 months, and there were no differences on DFS (P = 0.461) and OS (P = 0.989) between patients with EGFR mutations and those without in univariate analysis. The patients harboring 19del mutation had a better DFS (P = 0.028) and OS (P = 0.001) than the patients harboring L858R mutation with significant statistical difference. CONCLUSIONS: This study suggests that there is no difference on survival between patients with EGFR mutations and those without, but the patients harboring EGFR 19del mutation have survival advantage compared to those harboring EGFR L858R mutation.
Assuntos
Adenocarcinoma de Pulmão/genética , Biomarcadores Tumorais/genética , Neoplasias Pulmonares/genética , Adenocarcinoma de Pulmão/mortalidade , Adulto , Idoso , Intervalo Livre de Doença , Receptores ErbB/genética , Feminino , Genes erbB-1 , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/mortalidade , Masculino , Pessoa de Meia-Idade , Mutação , Prognóstico , Estudos RetrospectivosRESUMO
We obtained a strain of Bacillus subtilis, which we named Czk1, from the aerial roots of rubber trees. This bacterial isolate exhibits strong antagonistic activity against Ganoderma pseudoferreum, Phellinus noxius, Helicobasidium compactum, Rigidoporus lignosus, Sphaerostilbe repens, and Colletotrichum gloeosporioides. Our earlier research has shown that the antagonistic activity of a fermentation supernatant Czk1 isolate produces a complex mixture of lipopeptides. In this study, we used methanol to extract crude lipopeptides, purified them using a Sephadex G-25 column, cloned the lipopeptide genes, and analyzed purified fractions by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS) to identify the lipopeptides from B. subtilis strain Czk1. The cloned lipopeptide genes included those that encode the enzymes lpa, ituD, sfp, and fenB. The crude lipopeptides were purified and found in five fractions. Further analysis revealed that five fractions of the purified composition contained members of the surfactin, iturin, fengycin, and bacillomycin families of antibiotics. This suggests that these lipopeptides from strain Czk1 have potential as plant disease biocontrol agents.
Assuntos
Bacillus subtilis/metabolismo , Proteínas de Bactérias/metabolismo , Hevea/microbiologia , Lipopeptídeos/metabolismo , Raízes de Plantas/microbiologia , Bacillus subtilis/genética , Bacillus subtilis/isolamento & purificação , Proteínas de Bactérias/genética , Proteínas de Bactérias/isolamento & purificação , Clonagem Molecular , Colletotrichum/efeitos dos fármacos , Colletotrichum/fisiologia , Lipopeptídeos/genética , Lipopeptídeos/farmacologia , Metanol , Peptídeos Cíclicos/genética , Peptídeos Cíclicos/metabolismo , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Esporos Fúngicos/efeitos dos fármacos , Esporos Fúngicos/fisiologiaRESUMO
This study investigated the association of tumor necrosis factor-α (TNF-α)-308, -238, and -863 polymorphisms with osteoarticular tuberculosis (OA-TB) prognosis in a Hebei population. Genomic DNA was extracted from venous blood samples of 120 OA-TB patients and 100 healthy volunteers. TNF-α-308, -238, and -863 were analyzed by PCR-restriction fragment length polymorphism; genotype and allele frequencies were calculated. Serum TNF-α level was significantly higher in OA-TB patients (283.16 ± 51.68 ng/L) than in control (122.54 ± 54.65 ng/L; P < 0.05). Higher frequency of TNF-α-308 GG genotype in healthy volunteers (91.0%) than in OA-TB patients (79.2%) indicated that it was a protective factor against OA-TB (OR = 0.405, 95%CI = 0.147-0.657, P = 0.007). Higher frequencies of TNF-α-308 GA genotype and TNF-α-308 allele (A) in OA-TB patients (20.8 and 10.4%, respectively) than in healthy volunteers (8.0 and 5.0%, respectively) indicated an association with increased risk of OA-TB (OR = 3.112, 95%CI = 1.520-6.343, P = 0.003; OR = 3.109, 95%CI = 1.676-6.538, P = 0.006; respectively). Haplotype association analysis of TNF-α polymorphisms (-308/-238/-863) showed a higher frequency of TNF-α AGA in OA-TB patients (12.1%) than in healthy volunteers (3.5%), indicating that it was a risk factor for OA-TB (OR = 4.201, 95%CI = 1.80-9.91, P = 0.010). TNF-α-308 G/A and TNF-α AGA (-308/-238/-863) were associated with a predisposition to OA-TB, which could aid clinical detection, prevention, and prognosis of OA-TB.
Assuntos
Estudos de Associação Genética , Predisposição Genética para Doença , Tuberculose Osteoarticular/genética , Fator de Necrose Tumoral alfa/genética , Adulto , Alelos , Feminino , Genótipo , Haplótipos , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Prognóstico , Fatores de Risco , Tuberculose Osteoarticular/patologiaRESUMO
A multi-generational approach was used to investigate the persistent effects of a sub-lethal dose of spinosad in Plutella xylostella. The susceptibility of various sub-populations of P. xylostella to spinosad and the effects of the insecticide on the gene expression of γ-aminobutyric acid receptor (GABAR) were determined. The results of a leaf dip bioassay showed that the sensitivity of P. xylostella to spinosad decreased across generations. The sub-strains had been previously selected based on a determined LC25 of spinosad. Considering that GABA-gated chloride channels are the primary targets of spinosad, the cDNA of P. xylostella was used to clone GABARα by using reverse transcription-polymerase chain reaction (RT-PCR). The mature peptide cDNA was 1477-bp long and contained a 1449-bp open reading frame encoding a protein of 483 amino acids. The resulting amino acid sequence was used to generate a neighbor-joining dendrogram, and homology search was conducted using NCBI BLAST. The protein had high similarity with the known GABAR sequence from P. xylostella. Subsequent semi-quantitative RT-PCR and real-time PCR analyses indicated that the GABAR transcript levels in the spinosad-resistant strain (RR, 145.82-fold) and in Sub1 strain (selected with LC25 spinosad for one generation) were the highest, followed by those in the spinosad-susceptible strain, the Sub10 strain (selected for ten generations), and the Sub5 strain (selected for five generations). This multi-generational study found significant correlations between spinosad susceptibility and GABAR gene expression, providing insights into the long-term effects of sub-lethal insecticide exposure and its potential to lead to the development of insecticide-resistant insect populations.
Assuntos
Inseticidas , Macrolídeos , Mariposas/genética , Receptores de GABA/genética , Sequência de Aminoácidos , Animais , Combinação de Medicamentos , Expressão Gênica , Proteínas de Insetos/biossíntese , Proteínas de Insetos/genética , Resistência a Inseticidas , Mariposas/metabolismo , Receptores de GABA/biossínteseRESUMO
We conducted a case-control study to investigate the associations between GSTT1, GSTM1, and GSTP1 gene polymorphisms and development of osteosarcoma in a Chinese population. Between January 2013 and February 2015, 153 patients diagnosed with osteosarcoma and 252 control subjects were enrolled in the current study from the Orthopedic Hospital of the Second Hospital of Jilin University. The GSTM1, GSTT1, and GSTP1 gene polymorphisms were detected by polymerase chain reaction coupled with restriction fragment length polymorphism analysis. As determined by a multiple-logistic regression analysis, the Val/Val genotype of GSTP1 was associated with a significantly increased risk of osteosarcoma compared to that of the Ile/Ile genotype, with an odds ratio (OR) = 3.39, and a 95% confidence interval (CI) = 1.45-8.13. Moreover, the Ile/Val+Val/Val genotype of GSTP1 was correlated with a marginally significant increased risk of osteosarcoma compared to that of the Ile/Ile genotype (OR = 1.65, 95%CI = 1.08-2.53). However, we did not find any significant associations between the GSTM1 and GSTT1 gene polymorphisms and osteosarcoma risk. In conclusion, our results suggest that the GSTP1 gene polymorphism is associated with an increased risk of osteosarcoma, whereas the GSTM1 and GSTT1 gene polymorphisms may not influence the development of this cancer.
Assuntos
Neoplasias Ósseas/genética , Glutationa S-Transferase pi/genética , Osteossarcoma/genética , Polimorfismo Genético , Povo Asiático , Transformação Celular Neoplásica/genética , China , HumanosRESUMO
Peritrophic membrane proteins are important components of the insect peritrophic membrane. A novel cDNA gene encoding a chitin-binding protein, named secbp66, was identified by immunization screening of the cDNA library of Spodoptera exigua. The full length of secbp66 is 1806 bp, which encodes 602 amino acids. The predicted weight of the protein is 64.2 kDa. Bioinformatic analysis showed that a signal peptide composed of 17 amino acids located at the N-terminal of SeCBP66 contained seven tandem putative Type-II functional chitin-binding domains and five potential N-glycosylation sites, but no O-linked glycosylation sites. To study the properties of SeCBP66, recombinant SeCBP66 was successfully expressed in the insect cell line BTI-Tn-5B1-4 with a Bac-to-Bac expression system. A chitin binding experiment showed that the recombinant SeCBP66 protein could bind to chitin strongly. This study of the novel chitin-binding protein SeCBP66 provides a basis for developing new control targets for S. exigua.