RESUMO
PURPOSE: P21-activated kinase 1 (PAK1), a serine/threonine protein kinase which functions downstream of RAC and CDC42 GTPase, is activated by a variety of stimuli, including RAS and other growth signaling factors. The extracellular signal kinase (ERK) and protein kinase B (AKT) signal pathways have been implicated in the pathogenesis of cancers. Whether PAK1 is sensitive to KRAS mutation signals and plays a role through ERK and AKT signaling pathways in NSCLC needs to be studied. METHODS: The expression of PAK1, ERK and AKT was detected in both lung cancer cell lines and clinical samples. PAK1 RNA interference and specific inhibitor of PAK1(IPA-3) were applied to lung cancer cell lines and mouse xenograft tumors. Cell growth was measured by MTT and colony formation assays. Cell migration and invasion were detected by wound healing and transwell assays. RAS mutation was detected by Taqman probe method. Correlation between KRAS, PAK1, ERK and AKT activities was analyzed in lung cancer patients. RESULTS: PAK1 was highly expressed not only in RAS mutant but also in RAS wild-type lung cancer cells. Using specific inhibitor of PAK1, IPA-3 and PAK1 RNA interference, cell proliferation, migration and invasion of lung cancer cells were reduced significantly, accompanied by decreased activities of ERK and AKT. Dual inhibition of ERK and AKT suppressed these cellular processes to levels comparable to those achieved by reduction in PAK1 expression. In NSCLC patients, PAK1 was not correlated with KRAS mutation but was significantly positively correlated with pERK and pAKT. CONCLUSION: PAK1 played roles in NSCLC proliferation and invasion via ERK and AKT signaling and suggested a therapeutic target for NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Neoplasias Pulmonares/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Quinases Ativadas por p21/antagonistas & inibidores , Animais , Apoptose , Carcinoma Pulmonar de Células não Pequenas/enzimologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Dissulfetos/farmacologia , Ativação Enzimática , Feminino , Genes ras/genética , Xenoenxertos , Humanos , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mutação , Naftóis/farmacologia , Invasividade Neoplásica , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ensaio Tumoral de Célula-Tronco , CicatrizaçãoRESUMO
This study explores the differences of appearance and the digestive systems of two birds (Lonchura striata and Copsychus saularis), which have the same eating pattern in staple ingredients. Each bird was caught naturally and euthanized in appropriate manner, followed by morphology survey on bill, duodenum, jejunum, ileum and appearance of the body parts such as Culmen, Claw, Wing, Tarsus, Toe and Tail. The results showed that the appearance feature between them presented significant differences in the beak length, wing length and tail length, which were correlated to their staple feeding habits. In addition, in the digestive system, the length of duodenum and duodenal indexes showed significant differences (p 0.05 or p 0.01), which was considered to be correlated to their predation rate. However, there were no significant differences in jejunum length, ileum length, and the whole digestive tract length and weight, the evidence showed that feeding habits still play a great role in the whole intestine. It is concluded that different staple feeding habits allow the birds to evolve different traits both in appearance and intestine, which aim to improve their predation rate and adapt to different environments. Among all features, duodenum and duodenal were the major objects to get stressed in digestive system, while beak length, wing length and tail length differed most in appearance. And it was speculated that different food compositions lead to different changes in certain positions of the body, thus might shape an explorable trail, which needs to be investigated thoroughly.(AU)
Assuntos
Animais , Passeriformes/anatomia & histologia , Passeriformes/fisiologia , Sistema Digestório/química , Comportamento AlimentarRESUMO
This study explores the differences of appearance and the digestive systems of two birds (Lonchura striata and Copsychus saularis), which have the same eating pattern in staple ingredients. Each bird was caught naturally and euthanized in appropriate manner, followed by morphology survey on bill, duodenum, jejunum, ileum and appearance of the body parts such as Culmen, Claw, Wing, Tarsus, Toe and Tail. The results showed that the appearance feature between them presented significant differences in the beak length, wing length and tail length, which were correlated to their staple feeding habits. In addition, in the digestive system, the length of duodenum and duodenal indexes showed significant differences (p 0.05 or p 0.01), which was considered to be correlated to their predation rate. However, there were no significant differences in jejunum length, ileum length, and the whole digestive tract length and weight, the evidence showed that feeding habits still play a great role in the whole intestine. It is concluded that different staple feeding habits allow the birds to evolve different traits both in appearance and intestine, which aim to improve their predation rate and adapt to different environments. Among all features, duodenum and duodenal were the major objects to get stressed in digestive system, while beak length, wing length and tail length differed most in appearance. And it was speculated that different food compositions lead to different changes in certain positions of the body, thus might shape an explorable trail, which needs to be investigated thoroughly.
Assuntos
Animais , Comportamento Alimentar , Passeriformes/anatomia & histologia , Passeriformes/fisiologia , Sistema Digestório/químicaRESUMO
PURPOSE: To determine the sensitivity and specificity of serum Cyr61 as a potential biomarker for the diagnosis of colorectal cancer (CRC) and to assess the association between serum Cyr61 level and CRC clinicopathological status. METHODS: We used an enzyme-linked immunosorbent assay to measure serum Cyr61 in patients with CRC, patients with colorectal adenomas, and healthy controls. We also analyzed the relationship between serum Cyr61 and clinicopathological features of CRC patients. The levels of serum carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9) were quantified using the Roche Cobas 6000 Analyzer. The sensitivity and specificity of Cyr61, CEA, CA19-9 and CEA + CA19-9 were evaluated by receiver operating characteristic (ROC) analysis. RESULTS: The serum level of Cyr61 was significantly increased in CRC patients compared with colorectal adenoma patients and healthy controls (p < 0.001). Furthermore, the area under the ROC curve for Cyr61 was 0.935 (95 % confidence interval 0.902-0.968), higher than that for CEA + CA19-9 (0.827, 95 % confidence interval: 0.783-0.871). Use of a Cyr61 cutoff value of 92.0 pg/mL allowed distinguishing CRC patients and healthy controls with a sensitivity of 83 % and a specificity of 97 %. Among CRC patients, an elevated level of serum Cyr61 was significantly associated with more advanced TNM stage (p < 0.0042), lymph node metastasis (p < 0.0088), and vascular invasion (p = 0.0027). CONCLUSION: Cyr61 has potential as a serum biomarker for the diagnosis of CRC and for assessment of the clinicopathological status of CRC.
Assuntos
Adenoma/diagnóstico , Biomarcadores Tumorais/sangue , Neoplasias Colorretais/diagnóstico , Proteína Rica em Cisteína 61/sangue , Adenoma/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígeno CA-19-9/sangue , Antígeno Carcinoembrionário/sangue , Estudos de Casos e Controles , Neoplasias Colorretais/sangue , Ensaio de Imunoadsorção Enzimática , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Estadiamento de Neoplasias , Prognóstico , Curva ROCRESUMO
The bovine TRIM28 gene was amplified from ovary tissue by using RT-PCR. The TRIM28 gene was inserted into the eukaryotic expression vector pIRES2-EGFP and transfected into bovine fetal fibroblasts by using Lipofectamine 3000. TRIM28 mRNA and protein were detected by fluorescence microscope and western blotting. The results showed that the full length of TRIM28 was cloned and pIRES2-EGFP-TRIM28 was constructed successfully. EGFP expression was observed, and the pIRES2-EGFP-TRIM28 transfected group expressed more TRIM28 protein than that by the pIRES2-EGFP group. The TIMR28 gene has been successfully transferred into bovine fetal fibroblasts.
Assuntos
Proteínas Repressoras/genética , Animais , Bovinos , Células Cultivadas , Clonagem Molecular , Feminino , Fibroblastos/metabolismo , Vetores Genéticos/genética , Ovário/metabolismo , Proteínas Repressoras/metabolismoRESUMO
BACKGROUND: CCR6 expression is deregulated in some human malignancies and may be involved in the tumor progression. The aim of the present study was to determine the CCR6 expression in gastric cancer (GC) and to clarify its clinical significance. METHODS: We used western blotting to examine CCR6 protein expression in GC tissues and matched adjacent non-tumor tissues. Immunohistochemistry was performed on a large cohort of 372 postoperative GC samples. Chi-square test, Kaplan-Meier analysis and Cox regression model were used to analyze the data. RESULTS: Upregulated CCR6 protein expression was observed in the GC tissues by western blotting compared with the adjacent non-cancerous gastric tissues. High CCR6 expression was detected in 56.5 % (210/372) samples and significantly associated with the extracapsular extension of the tumor, tumor relapse and poor overall survival in GC (P < 0.001). Further analysis demonstrated that the CCR6 expression level stratified the patient outcome in stage II, stage III, T3/4, N positive and poorly differentiated/undifferentiated tumor subgroups. The Cox regression analysis showed that high expression of CCR6 was an independent prognostic factor for GC patients. CONCLUSIONS: CCR6 expression may be a novel biomarker for predicting clinical outcomes for GC patients.
Assuntos
Carcinoma/patologia , Receptores CCR6/biossíntese , Neoplasias Gástricas/patologia , Idoso , Biomarcadores Tumorais/análise , Western Blotting , Carcinoma/metabolismo , Carcinoma/mortalidade , Progressão da Doença , Feminino , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Prognóstico , Modelos de Riscos Proporcionais , Receptores CCR6/análise , Estudos Retrospectivos , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/mortalidadeRESUMO
Commercial flocks infected by Eimeria species parasites, including Eimeria maxima, have an increased risk of developing clinical or subclinical coccidiosis; an intestinal enteritis associated with increased mortality rates in poultry. Currently, infection control is largely based on chemotherapy or live vaccines; however, drug resistance is common and vaccines are relatively expensive. The development of new cost-effective intervention measures will benefit from unraveling the complex genetic mechanisms that underlie host-parasite interactions, including the identification and characterization of genes encoding proteins such as phosphatidylinositol 4-phosphate 5-kinase (PIP5K). We previously identified a PIP5K coding sequence within the E. maxima genome. In this study, we analyzed two bacterial artificial chromosome clones presenting a ~145-kb E. maxima (Weybridge strain) genomic region spanning the PIP5K gene locus. Sequence analysis revealed that ~95% of the simple sequence repeats detected were located within regions comparable to the previously described feature-rich segments of the Eimeria tenella genome. Comparative sequence analysis with the orthologous E. maxima (Houghton strain) region revealed a moderate level of conserved synteny. Unique segmental organizations and telomere-like repeats were also observed in both genomes. A number of incomplete transposable elements were detected and further scrutiny of these elements in both orthologous segments revealed interesting nesting events, which may play a role in facilitating genome plasticity in E. maxima. The current analysis provides more detailed information about the genome organization of E. maxima and may help to reveal genotypic differences that are important for expression of traits related to pathogenicity and virulence.
Assuntos
Eimeria/genética , Loci Gênicos , Genoma de Protozoário , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Composição de Bases , Clonagem Molecular , Coccidiose/parasitologia , Biologia Computacional , Interações Hospedeiro-Parasita , Anotação de Sequência Molecular , Sequências Repetitivas de Ácido Nucleico , Análise de Sequência de DNARESUMO
Fatty acid binding proteins (FABPs) are a family of small, highly conserved cytoplasmic proteins that bind long-chain fatty acids and other hydrophobic ligands. In this study, cDNA and genomic sequences of FABP4 and FABP5 were cloned successfully from the giant panda (Ailuropoda melanoleuca) using reverse transcription polymerase chain reaction (RT-PCR) technology and touchdown-PCR. The cDNAs of FABP4 and FABP5 cloned from the giant panda were 400 and 413 bp in length, containing an open reading frame of 399 and 408 bp, encoding 132 and 135 amino acids, respectively. The genomic sequences of FABP4 and FABP5 were 3976 and 3962 bp, respectively, which each contained four exons and three introns. Sequence alignment indicated a high degree of homology with reported FABP sequences of other mammals at both the amino acid and DNA levels. Topology prediction revealed seven protein kinase C phosphorylation sites, two casein kinase II phosphorylation sites, two N-myristoylation sites, and one cytosolic fatty acid-binding protein signature in the FABP4 protein, and three N-glycosylation sites, three protein kinase C phosphorylation sites, one casein kinase II phosphorylation site, one N-myristoylation site, one amidation site, and one cytosolic fatty acid-binding protein signature in the FABP5 protein. The FABP4 and FABP5 genes were overexpressed in Escherichia coli BL21 and they produced the expected 16.8- and 17.0-kDa polypeptides. The results obtained in this study provide information for further in-depth research of this system, which has great value of both theoretical and practical significance.
Assuntos
Proteínas de Ligação a Ácido Graxo/genética , Proteínas de Ligação a Ácido Graxo/metabolismo , Ursidae/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , Proteínas de Ligação a Ácido Graxo/química , Modelos Moleculares , Dados de Sequência Molecular , Músculo Esquelético/metabolismo , Fases de Leitura Aberta , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Alinhamento de SequênciaRESUMO
The ribosomal protein L24 (RPL24) belongs to the L24E family of ribosomal proteins and is located in the cytoplasm. The purpose of this study was to investigate the structure and anti-cancer function of RPL24 of the giant panda (Ailuropoda melanoleuca). The complementary DNA of RPL24 was cloned successfully using reverse transcription-polymerase chain reaction technology. We constructed a recombinant expression vector containing RPL24 complementary DNA and overexpressed it in Escherichia coli using pET28a plasmids. The expression product obtained was purified using Ni-chelating affinity chromatography. The results indicated that the length of the fragment cloned is 509 bp, and it contains an open-reading frame of 474 bp encoding 157 amino acids. Primary structure analysis revealed that the molecular weight of the putative RPL24 protein is 17.78 kDa with a theoretical isoelectric point of 11.86. The RPL24 gene is readily expressed in E. coli, and the RPL24 fused with the N-terminal histidine-tagged protein to give rise to the accumulation of an expected 23.51-kDa polypeptide. The inhibitory rate in mice treated with 0.1 mg/mL RPL24, the highest of 3 doses administered, can reach 67.662%, which may be comparable to the response to mannatide. The histology of organs with tumors showed that the tissues in the RPL24 group displayed a looser arrangement compared with that in the control group. Furthermore, no obvious damage was apparent in other organs, such as heart, lung, and kidney. The data showed that the recombinant RPL24 had time and dose dependency on the cell growth inhibition rate. Human laryngeal carcinoma Hep-2 cells treated with 0.3125-10 µg/mL RPL24 for 24 h displayed significant cell growth inhibition (P < 0.05; N = 6) in assays using 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazolium bromide compared with that in control (untreated) cells. By contrast, human hepatoma Hep G-2 cells displayed no significant change (P > 0.05; N = 6) from control (untreated) cells. RPL24 has time and dose dependency on Hep-2 cell growth inhibition. The data indicate that the effect at low concentrations is better than that at high concentrations, and the concentration of 0.625 µg/mL provides the best rate of growth inhibition. Further research is ongoing to determine the bioactive principles of recombinant RPL24 protein that are responsible for its anticancer activity.
Assuntos
Antineoplásicos/farmacologia , Proteínas Ribossômicas/farmacologia , Ursidae/genética , Sequência de Aminoácidos , Animais , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Sequência de Bases , Forma Celular/efeitos dos fármacos , Clonagem Molecular , Expressão Gênica , Células Hep G2 , Humanos , Masculino , Camundongos , Modelos Moleculares , Dados de Sequência Molecular , Filogenia , Conformação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/farmacologia , Proteínas Ribossômicas/química , Proteínas Ribossômicas/isolamento & purificação , Proteínas Ribossômicas/fisiologia , Análise de Sequência de DNA , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
In this study, electrical and structural remodeling of ventricles was examined in tachycardia-induced heart failure (HF). We studied two groups of weight-matched adult male mongrel dogs: a sham-operated control group (n=5) and a pacing group (n=5) that underwent ventricular pacing at 230 bpm for 3 weeks. Clinical symptoms of congestive HF were observed in both groups. Their hemodynamic parameters were determined and the severity of the HF was evaluated by M-mode echocardiography. Changes in heart morphology were observed by scanning electron and light microscopy. Ventricular action potential duration (APD), as well as the 50 and 90% APD were measured in both groups. All dogs exhibited clinical symptoms of congestive HF after rapid right ventricular pacing for 3 weeks. These data indicate that rapid, right ventricular pacing produces a useful experimental model of low-output HF in dogs, characterized by biventricular pump dysfunction, biventricular cardiac dilation, and non-ischemic impairment of left ventricular contractility. Electrical and structural myocardial remodeling play an essential role in congestive HF progression, and should thus be prevented.