RESUMO
PURPOSE: To explore the application value of multi-slice spiral CT (MSCT), magnetic resonance imaging (MRI) combined with gastric contrast-enhanced ultrasonography in the diagnosis of T staging of gastric cancer. METHODS: The subjects of study were 109 gastric cancer patients with T stages admitted to our hospital for diagnosis and treatment from December 2016 to December 2018. All the patients were examined with MSCT, MRI and gastric contrast-enhanced ultrasonography before operation to observe corresponding imaging results. T staging of gastric cancer patients was conducted according to the examination results, which was then compared with postoperative pathological staging. It was performed to analyze the accuracy of the three diagnostic methods and combined diagnosis of gastric cancer T staging. RESULTS: The sensitivity of MSCT in the diagnosis of T staging of gastric cancer was 60.00%, 67.74%, 72.22%, 76.47%, the specificity was 95.24%, 88.46%, 86.30%, 94.56% and the diagnostic coincidence rate was 87.16%, 82.57%, 81.65%, 91.74%; the sensitivity of MRI in the diagnosis of T staging of gastric cancer was 68.00%, 70.97%, 77.78%, 76.47%, the specificity was 92.86%, 88.46%, 91.78%, 95.65%, and the diagnostic coincidence rate was 87.16%, 83.49%, 87.16%, 92.66%; the sensitivity of gastric contrast-enhanced ultrasonography in the diagnosis of T staging of gastric cancer was 80.00%, 83.87%, 86.11%, 82.35%, the specificity was 97.62%, 92.31%, 91.78%, 97.83%, and the diagnostic coincidence rate was 93.58%, 89.91%, 89.91%, 95.41%; the sensitivity of combined MSCT, MRI and gastric contrast-enhanced ultrasonography in the diagnosis of T staging of gastric cancer was 88.00%, 93.55%, 97.22%, 94.12%; the specificity was 100%, 97.44%, 95.89%, 98.91%; and the diagnostic coincidence rate was 97.25%, 96.33%, 96.33%, 98.17%, respectively. Statistical analysis revealed that the sensitivity, specificity and diagnostic coincidence rate of combined detection of the three methods were significantly higher than those of single detection (P < 0.05). CONCLUSION: Combined use of MSCT, MRI and gastric contrast-enhanced ultrasonography can significantly improve the diagnostic sensitivity, specificity and diagnostic coincidence rate of T staging of gastric cancer. It may provide a certain reference value for guiding the selection of clinical therapeutic approaches and evaluation of curative effect.
Assuntos
Imageamento por Ressonância Magnética/métodos , Tomografia Computadorizada Multidetectores/métodos , Neoplasias Gástricas/diagnóstico por imagem , Ultrassonografia/métodos , Adulto , Idoso , Meios de Contraste/administração & dosagem , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Imagem Multimodal/métodos , Estadiamento de Neoplasias/métodos , Sensibilidade e Especificidade , Neoplasias Gástricas/patologiaRESUMO
The aim of this study was to determine the proliferation and osteogenic activity of fibroblasts induced with fibronectin and their possible dose-dependent relationship. The fibroblasts obtained by tissue explants adherent method were induced with fibronectin at different concentrations of 0, 10, 20, 40, 60, and 80 µg/mL for 14 days. The 3H-thymidine and 3H-proline incorporation test was used to evaluate the synthesis of DNA and collagen by fibroblasts, respectively. The mineralized nodules and osteocalcin secretion, as vital osteogenic indicators, were detected with tetracycline labeling and 125I-labeled competitive immunoassay, respectively. Fibronectin significantly increased the synthesis of DNA and collagen by fibroblasts, especially at the concentration of 40 µg/mL (P<0.05). The increased secretion of osteocalcin in the supernatant was also statistically significant at the concentration of 40 µg/mL (P<0.05). The mineralized nodules with trabecula-like structure derived from induced fibroblasts were positive for tetracycline labeling. The granulation tissue-derived fibroblasts induced with fibronectin exhibited increased proliferative, functional and osteogenic potential. Fibroblasts are considered a possible in situ stem cell in tissue engineering.
Assuntos
Proliferação de Células/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibronectinas/farmacologia , Osteogênese/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Fibroblastos/fisiologia , Humanos , CoelhosRESUMO
We aimed to investigate the influence of regulatory T cells including CD4+CD25+, CD8+CD28- and hepatitis B virus (HBV) genotype on sustained virological response and tolerance of nucleoside drugs. One hundred and thirty-seven patients were enrolled. Lamivudine was administered to 84 patients. Entecavir was administered to the other 53 patients. Before treatment, biochemical tests, HBV DNA load, HBV serum level, HBV genotype, PB CD3+, CD4+, CD8+, CD4+CD25+/CD3+, and CD8+CD28-/CD3+ frequencies were measured. Based on HBV DNA loads after 4 weeks of therapy, patients were divided into response group and suboptimal response group. The lamivudine group received treatment continuously, and then patients were categorized into non-resistance group and resistance group. Compared with the suboptimal response and resistance groups for lamivudine, CD4+CD25+/CD3+ levels were higher in the response and non-resistance groups (t=4.372, P=0.046; t=7.262, P=0.017). In the non-resistance group, CD8+CD28-/CD3+ frequency was lower than in the resistance group (t=5.527, P=0.037). Virus load and hepatitis B E antigen (HBeAg)-positive rate were significantly lower than in the response and resistance group (t=2.164, P=0.038; X2=4.239, P=0.040; t=2.015, P=0.044; X2=16.2, P=0.000). Incidence of drug resistance was high in patients with virogene type C. For the virological response to entecavir, CD8+CD28-/CD3+ level was significantly lower than that of the suboptimal response group (t=6.283, P=0.036). Response and suboptimal response groups were compared in CD3+, CD4+, CD8+, CD4+CD25+/CD3+ and virus genotype, and differences were not statistically significant (P>0.05). Baseline regulatory T cells including CD4+CD25+/CD3+ and CD8+CD28-/CD3+ frequencies have a relationship with the incidence of rapid virological response and the resistance to nucleoside drugs. Patients with HBV genotype C receiving lamivudine more often underwent drug resistance. Antiviral efficacy and the resistance to lamivudine were closely correlated with baseline factors; the same cannot be found for entecavir.
Assuntos
Antivirais/uso terapêutico , Guanina/análogos & derivados , Vírus da Hepatite B/imunologia , Hepatite B Crônica/tratamento farmacológico , Lamivudina/uso terapêutico , Nucleosídeos/uso terapêutico , Linfócitos T Reguladores , Adulto , Idoso , Resistência a Medicamentos , Feminino , Genótipo , Guanina/uso terapêutico , Vírus da Hepatite B/efeitos dos fármacos , Hepatite B Crônica/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Resposta Viral Sustentada , Linfócitos T Reguladores/imunologia , Fatores de TempoRESUMO
In this research, compound Maqin decoction (CMD) has been shown to positively affect in airway inflammation of asthma models. We evaluated the effects of CMD on the expression of transforming growth factor (TGF)-ß1/Smad proteins, interleukin (IL)-17, and IL-10 in lung tissue of asthmatic rats. Asthma was induced in a rat model using ovalbumin. After a 4-week treatment with CMD, rats were killed to evaluate the expression of TGF-ß1 and Smad proteins in lung tissue. IL-10 and IL-17 levels in lung tissue homogenates were determined by ELISA. The expression of TGF-ß1 and Smad3 protein increased, whereas expression of Smad7 protein decreased upon high-dose or low-dose treatment with CMD or by intervention with dexamethasone, compared to the control. There was a significant difference between treatment with a high dose CMD and the control treatment, but no significant difference was found between high-dose CMD treatment and dexamethasone intervention. The expression of TGF-ß1 and Smad7 protein increased, whereas the expression of Smad3 protein decreased in the model group compared to other groups. In the CMD high-dose group, low-dose group, and dexamethasone intervention group, the IL-17 concentrations in lung tissue homogenates were decreased, while IL-10 levels were increased. Again, there was a significant difference between CMD high-dose and control treatment, but not between CMD high-dose treatment and dexamethasone intervention. Thus, positive effects of CMD against asthmatic airway remodeling may be due to its regulatory effect on TGF-ß1, Smad3, and Smad7 protein levels and on cytokines such as IL-10 and IL-17.
Assuntos
Antiasmáticos/farmacologia , Asma/tratamento farmacológico , Pulmão/efeitos dos fármacos , Extratos Vegetais/farmacologia , Fator de Crescimento Transformador beta1/imunologia , Remodelação das Vias Aéreas/efeitos dos fármacos , Animais , Antiasmáticos/isolamento & purificação , Asma/induzido quimicamente , Asma/imunologia , Asma/patologia , Berberidaceae/química , Dexametasona/farmacologia , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Elaeagnaceae/química , Ephedra/química , Regulação da Expressão Gênica , Interleucina-10/genética , Interleucina-10/imunologia , Interleucina-17/genética , Interleucina-17/imunologia , Pulmão/imunologia , Pulmão/patologia , Masculino , Ovalbumina , Extratos Vegetais/química , Ratos , Ratos Sprague-Dawley , Scutellaria baicalensis/química , Transdução de Sinais , Proteína Smad3/genética , Proteína Smad3/imunologia , Proteína Smad7/genética , Proteína Smad7/imunologia , Fator de Crescimento Transformador beta1/genética , Xanthium/químicaRESUMO
This study was designed to evaluate bone matrix gelatin (BMG)/fibrin glue and chitosan/gelatin composite scaffolds for cartilage tissue engineering. Chondrocytes were isolated from costal cartilage of Sprague-Dawley rats and seeded on BMG/fibrin glue or chitosan/gelatin composite scaffolds. After different in vitro culture durations, the scaffolds were subjected to hematoxylin and eosin, Masson's trichrome, and toluidine blue staining, anti-collagen II and anti-aggrecan immunohistochemistry, and scanning electronic microscopy (SEM) analysis. After 2 weeks of culture, chondrocytes were distributed evenly on the surfaces of both scaffolds. Cell numbers and the presence of extracellular matrix components were markedly increased after 8 weeks of culture, and to a greater extent on the chitosan/gelatin scaffold. The BMG/fibrin glue scaffold showed signs of degradation after 8 weeks. Immunofluorescence analysis confirmed higher levels of collagen II and aggrecan using the chitosan/gelatin scaffold. SEM revealed that the majority of cells on the surface of the BMG/fibrin glue scaffold demonstrated a round morphology, while those in the chitosan/gelatin group had a spindle-like shape, with pseudopodia. Chitosan/gelatin scaffolds appear to be superior to BMG/ fibrin glue constructs in supporting chondrocyte attachment, proliferation, and biosynthesis of cartilaginous matrix components.
Assuntos
Condrócitos/efeitos dos fármacos , Engenharia Tecidual/métodos , Alicerces Teciduais/química , Adesivos/efeitos adversos , Agrecanas/genética , Agrecanas/metabolismo , Animais , Matriz Óssea/química , Adesão Celular , Células Cultivadas , Quitosana/efeitos adversos , Condrócitos/citologia , Condrócitos/metabolismo , Condrogênese/efeitos dos fármacos , Colágeno Tipo II/genética , Colágeno Tipo II/metabolismo , Fibrina/efeitos adversos , Gelatina/efeitos adversos , Ratos , Ratos Sprague-Dawley , Alicerces Teciduais/efeitos adversosRESUMO
The balance between type 1 and type 2 T helper cells (the Th1-Th2 balance) is closely correlated with cancer, but the correlation in ovarian cancer remains unconfirmed. We investigated the Th1-Th2 balance for the diagnosis, treatment, and prognostic evaluation of ovarian cancer. Fifty healthy subjects and 50 ovarian cancer patients were recruited. The levels of various cytokines were determined in sera and ovarian cancer tissues using a Th1-Th2 human cytokine array. The usefulness of TNFα, IFNγ, TNFα/IL-4, and IFNγ/IL-4 for ovarian cancer diagnosis was assessed based on receiver operating characteristic (ROC) curves. The relationship between the TNFα/IL-4 level and survival time was investigated based on a survival curve. In the ovarian cancer patients, the levels of Th1 factors (IL-2, IFNγ, TNFα, and IL-13) increased significantly in the sera, and IFNγ and TNFα increased significantly in the ovarian cancer tissues. The levels of Th2 factors (IL-5 and IL-6) increased in the sera, but the level of IL-6 decreased significantly in the ovarian cancer tissues. Serum TNFα/IL-4 and IFNγ/IL-4 levels increased significantly in the peripheral blood of the ovarian cancer patients. ROC curve analysis revealed that TNFα, IFNγ, TNFα/IL-4, and IFNγ/IL-4 levels are useful for ovarian cancer diagnosis, with area under the curve values of 0.831, 0.753, 0.846, and 0.803, respectively. The TNFα/IL-4 level in the ovarian cancer patients was positively correlated with survival time, and the Th1-Th2 balance shifted toward Th1 in the ovarian cancer patients. The TNFα/IL-4 ratio might be useful for the diagnosis and prognosis of ovarian cancer.
Assuntos
Interferon gama/sangue , Interleucina-4/sangue , Neoplasias Ovarianas/sangue , Fator de Necrose Tumoral alfa/sangue , Adulto , Idoso , Feminino , Humanos , Pessoa de Meia-Idade , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/patologia , Células Th1/imunologia , Células Th1/patologia , Equilíbrio Th1-Th2 , Células Th2/imunologia , Células Th2/patologiaRESUMO
Previous studies have used microarray technology to explore gene expression differences between the atrium and the ventricle. However, selection criteria for the differentially expressed genes (DEGs) based only on either the fold change or the P value in these studies. Here, we aim to further identify the DEGs by setting a P value threshold of <0.05 and a fold change of >2, which may yield more specific gene expression differences between the atrium and the ventricle. Gene expression profiling of the atrial appendages and the ventricular free walls in 13 normal male Sprague Dawley rats were obtained from the Gene Expression Omnibus data base (accession No.: GSE5266). DEGs between the atrial and the ventricular samples were screened using the microarray significance analysis. The underlying functions of DEGs were predicted by gene ontology and pathway enrichment analyses. In addition, we also constructed protein interactions networks, and analyzed the function modules of the interacting proteins by MCODE. A total of 757DEGs between the atria and the ventricles were found. The genes highly expressed in the ventricular myocytes were associated with muscle contraction (e.g., Myl1, Myl2, Myl3, and Myh7) and energy production (e.g., Acadm and Acsl6), while the genes preferentially expressed in the atrial myocytes were involved in the integration of neurohumoral signals (e.g., Cldn1). These conclusions were confirmed by pathway enrichment and function module analyses. Our present study provides an overview of the transcript level differences between the atrium and the ventricle, which may be useful for determination of potential biomarkers.
Assuntos
Apêndice Atrial/metabolismo , Ventrículos do Coração/metabolismo , Miocárdio/metabolismo , Mapas de Interação de Proteínas , Transcriptoma , Animais , Metabolismo Energético/genética , Perfilação da Expressão Gênica , Coração/fisiologia , Masculino , Contração Miocárdica/genética , Especificidade de Órgãos , Ratos , Ratos Sprague-DawleyRESUMO
The aim of this study is to investigate the ability to prenatally diagnose phenylketonuria (PKU) by using phenylalanine hydroxylase (PAH) gene mutation analysis combined with short tandem repeat (STR) linkage analysis in 118 fetuses from 112 Chinese families. Genomic DNA was extracted from the peripheral blood from members of 112 families and the exons and exon-intron boundaries of the PAH gene were amplified by PCR. PCR products were analyzed by bi-directional Sanger sequencing and multiplex ligation-dependent probe amplification (MLPA). The three variable number of tandem repeat (VNTR) markers PAH-1, PAH-26, PAH-32 were used in the prenatal diagnosis for the PKU families. We identified a spectrum of 63 different mutations, including 61 point mutations and indels, two large exon deletion mutations, and five novel mutations. A substantial proportion of mutant alleles were accounted for by p.R243Q (15.62%), EX6-96AG (9.82%), p.V399V (7.59%), p.Y356X (6.70%), and p.R413P (5.36%). The same mutations were identified in 31 prenatally genotyped fetuses. We identified 58 fetuses that carried only one mutant allele and 29 fetuses that carried no mutations of PAH and were presumed normal. PAH gene mutation analysis combined with STR linkage analysis can provide rapid and accurate prenatal diagnosis for PKU families.
Assuntos
Fenilalanina Hidroxilase/genética , Fenilcetonúrias/genética , Diagnóstico Pré-Natal , Alelos , Povo Asiático , Éxons , Feminino , Ligação Genética , Genótipo , Humanos , Íntrons/genética , Repetições de Microssatélites/genética , Fenilalanina Hidroxilase/sangue , Fenilcetonúrias/sangue , Mutação Puntual , Gravidez , Deleção de Sequência/genéticaRESUMO
The aim of this study was to breed a target genotype variety based on the identified chalkiness marker-QTL (quantitative trait locus) associations in rice. First, a permanent mapping population of rice that consisted of 525 recombinant inbred lines (RILs), which were derived from Zhenshan 97/Minghui 63, was used to identify QTLs with additive effects for rice quantitative traits and percentage of grain chalkiness (PGC). Subsequently, based on the identified QTLs in rice, the molecular marker 68923-PGC was selected to screen the low chalkiness rice line. Then, using the integration of molecular marker breeding and traditional breeding, we analyzed the genotype and phenotype of inbred lines from 525 RILs; we identified one rice variety with particularly high yields, good taste, and broad adaptability. The new variety was temporarily named RIL10, which was a high quality, high yield, and broadly adaptable variety, and it is predominantly a feature that has contributed to its geographical adaptability, which would be planted from 35°E to 18°E in Chinain China, where 2/3 of rice production occurs. RIL10 was a marker-assisted selection breeding achievement for producing a high quality, high yield, and broadly adaptable rice variety.
Assuntos
Oryza/genética , Locos de Características Quantitativas/genética , Cruzamento , Genótipo , Oryza/fisiologiaRESUMO
The neomycin-resistance (neo(r)) gene is widely used as a selectable marker in eukaryotic expression vectors; however, its expression often affects that of target genes. Cre recombinase recognizes LoxP sites, leading to site-specific recombination and deletion of DNA and RNA between two LoxP sites. In the present study, a humanized Fat-1 gene (hFat-1) was generated by DNA Works and used to construct a pC-PGK-neo(r)-hfat-1 expression vector, in which PGK-neo(r) was flanked by two LoxP sites. The pC-PGK-neo(r)-hfat-1 plasmids were transfected into porcine fetal fibroblasts using liposomes, and three transgenic cell lines were obtained by culturing with 400 µg/mL G418 for 7 days. Next, these cell lines were transfected with a Cre recombinase expression plasmid, which contains a puromycin resistance gene, in order to delete neo(r), which was integrated into the genome. hFat-1-neo(r) negative cells were obtained following puromycin selection. Real-time quantitative polymerase chain reaction data indicated that neomycin-resistant cells had higher hFat-1 expression than neomycin-sensitive cells. High performance gas chromatography data suggested that the n-6/n-3 ratio was significantly lower in transfected cells than in wild-type cells. The n-6/n-3 ratio in Cre-treated hFat-1-transfected cells was higher than that in untreated cells, suggesting that deletion of PGK-neo(r) decreased hFat-1 expression.
Assuntos
Proteínas de Caenorhabditis elegans/genética , Resistência Microbiana a Medicamentos/genética , Ácidos Graxos Dessaturases/genética , Feto/citologia , Fibroblastos/metabolismo , Neomicina/farmacologia , Fosfoglicerato Quinase/genética , Sus scrofa/embriologia , Animais , Animais Geneticamente Modificados , Linhagem Celular , Cromatografia Líquida de Alta Pressão , DNA/metabolismo , Ácidos Graxos/análise , Expressão Gênica , Vetores Genéticos/metabolismo , Humanos , Integrases , RNA/metabolismo , Reação em Cadeia da Polimerase em Tempo RealRESUMO
We investigated the synergistic effect of bone morphogenetic protein 9 (BMP9) and transforming growth factor (TGF)-b in the transformation of mesenchymal stem cells into osteoblasts. We evaluated the effect of BMP9 and TGF-b on the induction of osteoblast formation. Mitogen-activated protein kinase (MAPK) pathway-related proteins such as p38, extracellular receptor kinase 1/2, and c-Jun N-terminal kinase (JNK) were analyzed. The interactions between TGF-Smad and BMP-MAPK were also studied. BMP9 alone induced the differentiation of mesenchymal stem cells (MSCs) into osteoblasts and enhanced phosphorylation of p38, extracellular receptor kinase 1/2, and JNK. TGF-b alone failed to induce transformation, but could increase the effect of BMP9. In this process the activation of Smad resulted in activation of the JNK pathway in the MAPK pathway. BMP9 induced osteogenesis of MSC differentiation through the MAKP pathway, while TGF-b contributed to BMP9 enhancement through the Smad-JNK pathway.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Fator 2 de Diferenciação de Crescimento/farmacologia , Osteoblastos/citologia , Fator de Crescimento Transformador beta/farmacologia , Fosfatase Alcalina/metabolismo , Animais , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Colágeno Tipo I/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Osteoblastos/efeitos dos fármacos , Osteoblastos/enzimologia , Osteopontina/metabolismo , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
MSP130-related-2 is thought to play a role in bio-mineralization as revealed in Crassostrea gigas and sea urchins. In this study, an MSP130-related-2 gene was isolated from Hyriopsis cumingii (HcMSP130-related-2) and characterized for the first time. The HcMSP130-related-2 cDNA was 2307 bp in length and consisted of a 572-bp 5'-untranslated region (5'-UTR), a 1239-bp open reading frame encoding 430-amino acid residues, and a 439-bp 3'-UTR. The molecular weight of the peptide was predicted to be 48551.3 Da, with a theoretical isoelectric point of 4.78 and instability index of 32.74, indicating that the protein is stable. The HcMSP130-related-2 amino acid residues included a signal peptide and several potential N-glycosylation sites. NCBI BLAST analysis indicated that this full-length amino acid sequence showed the highest similarity with HcMSP130-related-2 from C. gigas (45%) and about 38% identity with that from SpMSP130-rel-2 and Strongylocentrotus purpuratus. A phylogenetic tree showed that HcMSP130-rel-2 clustered with MSP130 from C. gigas. HcMSP130-related-2 was expressed in various tissues, including the mantle, blood, gill, foot, liver, kidney, intestine, and muscle, with the highest transcripts found in the mantle. Quantitative real-time polymerase chain reaction was used to analyze the expression of the HcMSP130- related-2 gene in grass carp after inducing shell damage. HcMSP130- related-2 expression was upregulated significantly in the mantle within 7 days (P < 0.05) after damage; however, the expression remained unchanged in the adductor muscle tissues (P > 0.05). These data suggest that HcMSP130-related-2 might be involved in shell formation in H. cumingii.
Assuntos
Glicoproteínas de Membrana/genética , Filogenia , Ouriços-do-Mar/genética , Unionidae/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , DNA Complementar/genética , Expressão Gênica , Ouriços-do-Mar/metabolismo , Alinhamento de SequênciaRESUMO
With the development of gene targeting approaches, genomic mutation technologies in livestock animals such as gene trapping, zinc finger nucleases (ZFNs), transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats and their associated systems have been improved. Although ZFNs have been used for gene targeting in many species, the off-target sites are still present. Using gene trapping, the workload of screening of targeted clones was decreased by generating a smaller number of drug-resistant clones. Determining whether the efficiency of gene trapping is lower than that of ZFNs for a specific gene has been challenging. In this study, to knock out the bovine myostatin gene, we constructed a promoter trap vector and compared its efficiency with that of ZFNs. The promoter trap vector contained a green fluorescent protein sequence without the promoter and a neomycin phosphotransferase (neo(R)) cassette driven by the phosphoglycerate kinase promoter. When the trapping vector was inserted downstream of the endogenous promoter, the fluorescent protein gene was expressed. The targeted-positive cell clones were identified based on green fluorescence and G418 double selection, followed by polymerase chain reaction analysis and sequencing. The targeting efficiency reached 5%. Compared with the efficiency of ZFN pairs (5.17 and 2.86%), the promoter trap vector PIII-myostatin could knock out the bovine myostatin gene. Therefore, gene trapping may be an effective tool for genomic modification.
Assuntos
Técnicas de Inativação de Genes/métodos , Marcação de Genes/métodos , Vetores Genéticos/genética , Miostatina/genética , Regiões Promotoras Genéticas/genética , Animais , Sequência de Bases , Bovinos , Células Cultivadas , Endonucleases/genética , Endonucleases/metabolismo , Feto , Fibroblastos/citologia , Fibroblastos/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Canamicina Quinase/genética , Canamicina Quinase/metabolismo , Microscopia de Fluorescência , Dados de Sequência Molecular , Músculos , Transfecção , Dedos de Zinco/genéticaRESUMO
Antibody-mediated rejection (AMR) is an important factor affecting survival after renal transplantation. A highly selective proteasome inhibitor, bortezomib, clears activated plasma cells from the body and has important therapeutic effect on AMR. We investigated the effects of bortezomib on AMR in a patient after a second renal transplant. Biopsy confirmed the diagnosis of mixed cellular rejection and AMR. Bortezomib was administered on day 1 (1.3 mg/m(2)), day 4 (1.0 mg/m(2)), and day 8 (1.0 mg/m(2)). On the same days, 250 mg methylprednisolone was administered once, and cyclosporine dose (5 mg·kg(-1)·day(-1)) was reduced by 50%. Oral mycophenolate mofetil and steroid were withdrawn on day 1 of bortezomib treatment. Intermittent double-filtration plasmapheresis was also performed. We monitored parameters, including T lymphocyte subsets, CD139 and CD19 expression, panel reactive antibody (PRA), and serum creatinine concentration. At follow-up 6 months after bortezomib treatment, we observed: 1) serum creatinine stabilized at 130 µM from a peak level of 337 µM; 2) PRA decreased from a maximum of 66.7 to 0%; 3) blood plasma cell percentage rebounded after significantly decreasing following the first dose of bortezomib; 4) in renal allograft biopsy, immunohistochemical staining for C4d shifted from strongly positive to negative, and cellular rejection shifted from type IIA to borderline; and 5) adverse effects such as platelet suppression, hypotension, and grade 3 peripheral neuropathy emerged. Bortezomib effectively treated antibody-mediated renal transplantation rejection in this case study, but clinical trials with large sample sizes are still needed to explore clinical safety and tolerability.
Assuntos
Anticorpos/efeitos adversos , Bortezomib/efeitos adversos , Rejeição de Enxerto/tratamento farmacológico , Transplante de Rim/efeitos adversos , Anticorpos/imunologia , Bortezomib/administração & dosagem , Feminino , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/patologia , Humanos , Rim/efeitos dos fármacos , Rim/imunologia , Rim/patologia , Ácido Micofenólico/administração & dosagem , Ácido Micofenólico/análogos & derivados , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/patologia , Transplante Homólogo/efeitos adversosRESUMO
The pathogenicity of 47 isolates of Sclerotinia sclerotiorum from oilseed rape (Brassica napus L.) in Anhui, China, was tested by detached leaf inoculation using the susceptible rape cultivar Wanyou-14. All isolates were pathogenic to the cultivar and could be grouped into 3 categories based on the lesion length on the leaves tested: weak pathogenicity type, intermediate pathogenicity type, and strong pathogenicity type. This suggested that there was differentiation in the pathogenicity among the strains tested of S. sclerotiorum. Additionally, the intraspecific DNA polymorphisms among 47 strains of S. sclerotiorum were investigated by screening 40 pairs of inter-simple sequence repeat (ISSR) primers. Unweighted pair-group method with arithmetic average cluster analysis of these ISSR data distinguished all strains from each other and revealed considerable genetic variability among them. These strains were classified into 7 clusters according to their branching in the dendrogram, and partial correlation was observed between the genetic polymorphisms and the pathogenicity of S. sclerotiorum strains.
Assuntos
Ascomicetos/genética , Brassica napus/microbiologia , Repetições de Microssatélites , Ascomicetos/crescimento & desenvolvimento , Ascomicetos/patogenicidade , China , Análise por Conglomerados , Marcadores Genéticos , Variação GenéticaRESUMO
In this study, 2 approaches were adopted to obtain good single-strand conformation polymorphism (SSCP) data for autotetraploid alfalfa; primers were added to PCR products, and fluorescent-labeled primers were utilized. PCR-SSCP conditions for a 331-bp fragment in the coding region of polygalacturonase-inhibiting protein gene 2 in alfalfa (MsPGIP2) were optimized, and the results showed that the best SSCP gel pattern could be obtained when the loading mixture was made by mixing 1 µL PCR products, 0.2 to 0.8 µL unlabeled primers (50 µM) and 4 to 16 µL loading buffer. Furthermore, the use of the fluorescent-labeled primers resulted in 2 separated electrophoresis images from 2 complementary single DNA strands, thus making the determination of alleles and idiotypes a relatively easy task. In addition, the results of sequencing prove that the determination of alleles and idiotypes were accurate based on SSCP analysis. Finally, a total of 9 alleles with 18 SNP sites were identified for MsPGIP2 in the alfalfa variety 'Algonquin'. In conclusion, MsPGIP2 possessed great genetic variation, and the addition of primers to the PCR products in combination with the fluorescent labeling of primers could significantly improve the sensitivity and resolution of SSCP analysis. This technique could be used for genetic diversity detection and marker-assisted breeding of useful genes in autopolyploid species such as alfalfa.
Assuntos
Impressões Digitais de DNA/métodos , Medicago sativa/genética , Proteínas de Plantas/genética , Alelos , Primers do DNA/química , Fluorescência , Polimorfismo de Nucleotídeo Único , Polimorfismo Conformacional de Fita SimplesRESUMO
To compare fracture healing therapies, the gene expression profiles of rat fracture samples treated with nail and plate fixation were analyzed at 1 day, 3 days, 1 week, 2 weeks, 4 weeks, and 6 weeks after surgery. The gene expression profiles GSE1685, which include 19 samples, were downloaded from the Gene Expression Omnibus database. After preprocessing, the gene expression profiles were subjected to time series analysis using the Short Time-series Expression Miner software, and the significantly differentially expressed gene (DEG) sets were selected. Further, the distributions of those DEG sets on the corresponding chromosomes were identified using the functional classification tool. Finally, the DEGs were subjected to function and pathway enrichment analysis. DEG analysis indicated that the number of DEGs (854 genes) from nail fixation was significantly lower than that of DEGs (1029 genes) from plate fixation. The DEGs were mainly enriched in cell proliferation, cellular localization, and response to wounding functions. Several critical DEGs expressed during the fracture healing process were screened, and 2 common pathways were enriched for the DEGs in the nail fixation and plate fixation. These DEGs and pathways may be potential targets or predictive markers during fracture healing.
Assuntos
Consolidação da Fratura/genética , Fraturas Ósseas/genética , Transcriptoma , Animais , Modelos Animais de Doenças , Fixação Interna de Fraturas , Fraturas Ósseas/metabolismo , Fraturas Ósseas/cirurgia , Ratos , Transdução de Sinais , Fatores de TempoRESUMO
The aim of this study was to investigate the correlation between apparent diffusion coefficients (ADCs), the relative apparent diffusion coefficient (rADC), and the pathological prognostic factor human epidermal growth factor receptor 2 (HER-2) in patients with breast cancer. A total of 64 women with breast cancer underwent breast diffusion-weighted imaging. HER-2 expression was detected in histological specimens. The ADC value, rADC value, and HER-2 level were determined. The ADC and rADC values of the breast cancer group were 1.1495 ± 0.1499 x 10(-3) and 0.6602 ± 0.0853, respectively. The differences in the ADC and rADC values between the two groups were statistically significant. There was no correlation between the ADC value and HER-2 expression in patients with breast cancer (r = -0.508, P = 0.043). However, the rADC value eliminated the individual differences to some extent. Therefore, compared to the ADC value, the rADC value had a better correlation with HER-2 expression.
Assuntos
Doenças Mamárias/diagnóstico , Neoplasias da Mama/diagnóstico , Imagem de Difusão por Ressonância Magnética/métodos , Adulto , Doenças Mamárias/metabolismo , Neoplasias da Mama/metabolismo , Diagnóstico Diferencial , Feminino , Humanos , Pessoa de Meia-Idade , Receptor ErbB-2/metabolismoRESUMO
The aim of this study was to evaluate and investigate the pathogenetic mechanism and countermeasures of subacute thrombosis (SAT) after coronary stenting in elderly diabetic patients. The clinical characteristics and pathogenetic mechanisms in 3 cases of SAT after stent implantations in elderly diabetic patients were retrospectively examined to determine the treatment strategies for SAT. Among 98 patients with diabetes who had coronary stents implanted or were >60 years of age, three (3.06%) had SAT. One case of SAT was diagnosed by angiography; coronary balloon dilatation, thrombolysis, and re-perfusion resulted in full recovery in this case. The second case involved potential SAT, and in the third case, SAT was not ruled out. Two cases were characteristic of ST-segment elevation myocardial infarction, and one case, in which SAT was not ruled out, resulted in sudden death. SAT within a stent may be related to intraoperative stent malapposition caused by a grade C lesion, age, diabetes, chronic total occlusion, or postoperative irregular administration of medication.
Assuntos
Angioplastia Coronária com Balão/efeitos adversos , Trombose Coronária/etiologia , Trombose Coronária/terapia , Stents/efeitos adversos , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Doença das Coronárias/complicações , Doença das Coronárias/diagnóstico , Doença das Coronárias/terapia , Diabetes Mellitus Tipo 2/complicações , Evolução Fatal , Feminino , Fibrinolíticos/administração & dosagem , Fibrinolíticos/uso terapêutico , Humanos , Masculino , Pessoa de Meia-Idade , Resultado do Tratamento , Ativador de Plasminogênio Tipo Uroquinase/administração & dosagem , Ativador de Plasminogênio Tipo Uroquinase/uso terapêuticoRESUMO
We evaluated the effects of an intra-aortic balloon pump on hemodynamics, brain natriuretic peptide concentration and cardiac function of patients with acute myocardial infarction, after reperfusion therapy. Sixty-three patients with acute anterior wall ST-elevation myocardial infarction who underwent primary percutaneous coronary intervention were given an intra-aortic balloon pump (32 cases) or not (control group, 31 cases). The mean pulmonary arterial pressure, pulmonary capillary wedge pressure and cardiac index were measured with a Swan-Ganz catheter. The brain natriuretic peptide concentration was detected by immunochemiluminometric assay. Left ventricular end-diastolic diameter and left ventricular ejection fraction were measured by echocardiography. No difference in baseline was observed between the two groups on day 1 in the hospital. On day 5, mean pulmonary artery pressure and pulmonary capillary wedge pressure of patients with the intra-aortic balloon pump were significantly lower, and cardiac index of was higher than that of the controls, whereas no differences in left ventricular end-diastolic diameter or left ventricular ejection fraction were observed between the two groups. On days 5 and 90, the brain natriuretic peptide concentration of the intra-aortic balloon pump patients was lower than that of the controls. On day 90, left ventricular end-diastolic diameter was smaller in the intra-aortic balloon pump patients, but no difference in left ventricular ejection fraction was observed between the two groups. The intra-aortic balloon pump improved the hemodynamic index and cardiac function and decreased brain natriuretic peptide concentration in patients with acute anterior wall ST-elevation myocardial infarction.