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At least 1,279 records · Page 71Linked to original sources
PubMed2002

[Significance of cytokeratin gene (CK-20 mRNA) expression in metastatic lymph nodes in colon carcinoma patients].

OBJECTIVE: To elucidate the significance of detecting cytokeratin gene (CK-20 mRNA) expression in metastatic lymph nodes of colon carcinoma patients. METHODS: The expression of CK-20 mRNA was detected in 342 lymph nodes by reverse transcription-polymerase chain reaction (RT-PCR) in 49 colon carcinoma patients. Its incidence was compared with that by routine pathologic examination. RESULTS: The positive rates of detecting metastasis in the lymph nodes were 21.9% by RT-PCR and 11.1% by routine pathologic examination. CONCLUSION: The detection of cancer metastasis in the lymph nodes in colon carcinoma is almost doubled (21.9% vs 11.1%) by CK-20 mRNA, which may provide a guiding significance in staging, treatment planning and prognosis in the prognostic estimation of colon carcinoma patients.

Adult↗
PubMed2006

Corneal avascularity is due to soluble VEGF receptor-1.

Corneal avascularity-the absence of blood vessels in the cornea-is required for optical clarity and optimal vision, and has led to the cornea being widely used for validating pro- and anti-angiogenic therapeutic strategies for many disorders. But the molecular underpinnings of the avascular phenotype have until now remained obscure and are all the more remarkable given the presence in the cornea of vascular endothelial growth factor (VEGF)-A, a potent stimulator of angiogenesis, and the proximity of the cornea to vascularized tissues. Here we show that the cornea expresses soluble VEGF receptor-1 (sVEGFR-1; also known as sflt-1) and that suppression of this endogenous VEGF-A trap by neutralizing antibodies, RNA interference or Cre-lox-mediated gene disruption abolishes corneal avascularity in mice. The spontaneously vascularized corneas of corn1 and Pax6+/- mice and Pax6+/- patients with aniridia are deficient in sflt-1, and recombinant sflt-1 administration restores corneal avascularity in corn1 and Pax6+/- mice. Manatees, the only known creatures uniformly to have vascularized corneas, do not express sflt-1, whereas the avascular corneas of dugongs, also members of the order Sirenia, elephants, the closest extant terrestrial phylogenetic relatives of manatees, and other marine mammals (dolphins and whales) contain sflt-1, indicating that it has a crucial, evolutionarily conserved role. The recognition that sflt-1 is essential for preserving the avascular ambit of the cornea can rationally guide its use as a platform for angiogenic modulators, supports its use in treating neovascular diseases, and might provide insight into the immunological privilege of the cornea.

Animals↗
PubMed1995

Regulation of beta 1-adrenoceptors by glucocorticoids and thyroid hormones in fetal sheep.

The effects of betamethasone alone or in combination with thyroxine (T4) on ovine fetal beta-adrenoceptors were investigated at the molecular level. Ovine fetuses (126 days gestation; term = 150 days) were treated with a single ultrasound-guided intramuscular injection of 0.5 mg/kg betamethasone, betamethasone + 50 micrograms/kg T4, or saline. Forty-eight h after injection, lambs were delivered by cesarean section and evaluated three h for postnatal adaptation. Myocardial beta-adrenoceptor equilibrium dissociation constant (Kd) and maximal receptor density (Bmax), as assessed by [3H]dihydroalprenolol binding, were not significantly different in drug-treated groups compared to the control group. Northern hybridization and RNase protection assays of myocardial total RNA probed with a sheep beta 1-adrenoceptor riboprobe confirmed no changes in expression at the level of the gene. Levels of beta 1-adrenoceptor mRNA in the lung and brain were also unaffected by the treatments. Because other genes are responsive to glucocorticoids and thyroid hormones at this stage, the absence of up-regulation of beta-adrenoceptor number and steady-state levels of mRNA coding for beta 1-adrenoceptor following fetal corticosteroid and thyroid hormone treatment may indicate a specific, developmentally regulated repressor mechanism.

Animals↗
PubMed2026

Epigenetics and In Silico Transcriptome Analysis of Pediatric Acute Myeloid Leukemia.

Pediatric acute myeloid leukemia (AML) is a heterogeneous hematologic malignancy that accounts for about 15%-20% of childhood leukemias. Despite therapeutic advances, relapses remain common, and survival for high-risk patients is below 60%. Unlike adult AML, pediatric AML displays distinct genetic mutations, including FLT3-ITD, NPM1, KMT2A rearrangements, and core-binding factors (CBF) fusions, as well as extensive epigenetic dysregulation. Aberrant DNA methylation, histone modifications, and altered non-coding RNA expressions disrupt hematopoietic differentiation and activate oncogenic transcriptional networks. Recent advances in silico transcriptomic analysis have transformed the study of pediatric AML by integrating gene expression and epigenetic data to identify molecular drivers and regulatory networks. Computational RNA-seq pipelines and pathway analyses have highlighted key epigenetic regulators, including DNMT3A, TET2, and HDACs, as potential therapeutic targets. Multi-omics approaches combining transcriptomic, methylomic, and chromatin accessibility data are increasingly used to define biomarkers for diagnosis, prognosis, and therapeutic response. This review provides a comprehensive overview of the molecular and epigenetic landscape of pediatric AML, emphasizing the power of in silico transcriptome analysis to uncover disease mechanisms, refine patient stratification, and guide the development of precision-based epigenetic therapies aimed at improving long-term outcomes in children with AML.

Humans↗
PubMed2005

Altered expression of TFF-1 and CES-2 in Barrett's Esophagus and associated adenocarcinomas.

Identification of biomarkers to recognize individuals with Barrett's esophagus (BE) predisposed to develop malignancy is currently a pressing issue. We utilized gene expression profiling to compare molecular signatures of normal esophagus and stomach, BE, and adenocarcinoma (AC) to identify such potential biomarkers. Over 22,000 genes were analyzed by oligonucleotide microarrays on 38 unique RNA Unsupervised and supervised clusterings were performed on a subset of 2849 genes that varied most significantly across the specimens. Immunohistochemistry (IHC) for two of the significantly differentially expressed gene products was performed on tissue microarrays. Unsupervised clustering identified two discernable molecular BE profiles, one of which was similar to normal gastric tissue ("BE1"), and another that was shared by several of the AC specimens ("BE2"). The BE1 profile included expression of several genes that have been described as tumor-suppressor genes, most notably trefoil factor 1 (TFF-1). The BE2 profile included expression of genes previously found overexpressed in cancers, such as carboxylesterase-2 (CES-2). IHC demonstrated the loss of TFF-1 late in the progression of BE to AC. It also revealed CES-2 as being upregulated in AC documented to have arisen in the presence of BE. These potential biomarkers, as well as the relative expression of genes from BE1 versus those from BE2, may be validated in the future to aid in risk stratification and guide treatment protocols in patients with BE and associated AC.

Adenocarcinoma↗
PubMed1992

UNC-5, a transmembrane protein with immunoglobulin and thrombospondin type 1 domains, guides cell and pioneer axon migrations in C. elegans.

The unc-5 gene is required for guiding pioneering axons and migrating cells along the body wall in C. elegans. In mutants, dorsal migrations are disrupted, but ventral and longitudinal movements are largely unaffected. The gene was tagged for molecular cloning by transposon insertions. Based on genomic and cDNA sequencing, the gene encodes UNC-5, a transmembrane protein of 919 aa. The predicted extracellular N-terminus comprises two immunoglobulin and two thrombospondin type 1 domains. Except for an SH3-like motif, the large intracellular C-terminus is novel. Mosaic analysis shows that unc-5 acts in migrating cells and pioneering neurons. We propose that UNC-5 is a transmembrane receptor expressed on the surface of motile cells and growth cones to guide dorsal movements.

Amino Acid Sequence↗
PubMed2002

Guide DNA technique in bacterial ribonuclease P reaction for effective processing of tRNA precursor.

Previously, we found that a small (approx. 20-mer) DNA hybridizing to the 5'-leader region of a tRNA precursor enhances the cleavage efficiency in bacterial ribonuclease P reaction. We named this technique the 'guide DNA technique'. Detailed analyses showed that the length of the guide DNA, concentration of the guide DNA and the hybridizing position affected the cleavage efficiency: for an effective cleavage reaction, guide DNA should be designed to hybridize to the region on the cleavage site, should be 20 bases or more in length and should be of high concentration. The presence of a 5'-flanking region in the DNA did not affect the cleavage reaction. The guide DNA technique is a useful tool for effective preparation of mature tRNA molecules in vitro.

Animals↗
PubMed2002

Site-specific cross-linking analyses reveal an asymmetric protein distribution for a box C/D snoRNP.

Methylation of the ribose 2'-hydroxyl, the most widespread modification of ribosomal and splicesomal RNAs, is guided by the box C/D class of small nucleolar RNAs (snoRNAs). Box C/D small nucleolar ribonucleoproteins (snoRNPs) contain four core proteins: fibrillarin, Nop56, Nop58 and 15.5 kDa. We constructed U25 snoRNAs containing a single photoactivatable 4-thiouridine at each U position within the conserved box C/D and C'/D' motifs. Proteins assembled on the snoRNA after injection into Xenopus oocyte nuclei were identified by cross-linking, and reconstituted particles characterized by functional rescue and mutational analyses. Our data argue that box C/D snoRNPs are asymmetric, with the C' box contacting Nop56 and fibrillarin, the C box interacting with Nop58, and the D and D' boxes contacting fibrillarin. No cross-link to 15.5 kDa was detected; its binding is disrupted by 4-thiouridine substitution in position 1 of the C box. Repositioning the guide sequence of U25 upstream of box D instead of D' revealed that both C/D motifs have the potential to function as guide centers, but, surprisingly, there was no alteration in protein cross-linking.

Animals↗
PubMed1985

The genetic basis for describing viruses as species.

The basis for the taxonomic definition of viruses or other species eventually has to be genetic. Since the beginning of the taxonomical considerations of organisms, the criterion that individuals can be grouped together on the basis of genetic compatibilities has been adopted as the guiding principle for describing a species. Although the genetic tests for grouping and distinguishing viruses may be different, the principle behind the species concept can still be applied. Also, the very existence of virus heredity and invokes the concept of speciation.

Bunyaviridae↗
PubMedDate not supplied

Expression of transferrin receptor and ferritin H-chain mRNA are associated with clinical and histopathological prognostic indicators in breast cancer.

It is well known that iron plays an essential role in many biochemical reactions and that rapidly growing cells require more iron for their growth and metabolism than resting cells. Transferrin and its receptor are required for entry of iron into the cell. In contrast, ferritin is a cellular storage protein whose main function is to sequester excess ferric iron and thus prevent high concentrations of soluble ferric iron from becoming toxic to the cell. However, the clinical significance of both transferrin receptor and ferritin mRNA levels have not previously been described in tumors from breast cancer patients. In this study, tumor tissue mRNA levels of transferrin receptor and ferritin were quantitated on forty-two breast cancer patients. A highly sensitive non-radioisotopic cDNA polymerase chain reaction assay was used to quantitate expression of mRNA. The expression of glyceraldehyde-3-phosphate dehydrogenase served as the control. In the tumor tissue from the 42 breast cancer patients the transferrin receptor mRNA levels were significantly correlated to the ferritin H-chain mRNA levels (Spearman correlation r = 0.5433, p = 0.0002; Pearson correlation r = 0.6276, p < 0.0001). The level of amplified transferrin receptor complementary DNA was related to differentiation (ANOVA, p = 0.042) with poorly differentiated tumors having high levels of transferrin receptor mRNA. Further, the levels of amplified gene for ferritin heavy chain complementary DNA was directly related to axillary lymph nodes status (Student's t-test, p = 0.044), presence of metastatic disease (Student's t-test, p = 0.046) and clinical stage (stage I + stage II versus stage III + stage IV; Student's t-test, p = 0.0181). These results demonstrate that non-radioisotopic RT-PCR is a very sensitive method for determining mRNA levels in tumor tissue. Additionally, the quantitation of expression of transferrin receptor and ferritin heavy chain mRNA may be useful for assessing prognosis and guiding therapeutic decisions in breast cancer patients.

Adult↗
PubMed2001

Bioinformatic tools for DNA/protein sequence analysis, functional assignment of genes and protein classification.

The development of efficient DNA sequencing methods has led to the achievement of the DNA sequence of entire genomes from (to date) 55 prokaryotes, 5 eukaryotic organisms and 10 eukaryotic chromosomes. Thus, an enormous amount of DNA sequence data is available and even more will be forthcoming in the near future. Analysis of this overwhelming amount of data requires bioinformatic tools in order to identify genes that encode functional proteins or RNA. This is an important task, considering that even in the well-studied Escherichia coli more than 30% of the identified open reading frames are hypothetical genes. Future challenges of genome sequence analysis will include the understanding of gene regulation and metabolic pathway reconstruction including DNA chip technology, which holds tremendous potential for biomedicine and the biotechnological production of valuable compounds. The overwhelming volume of information often confuses scientists. This review intends to provide a guide to choosing the most efficient way to analyze a new sequence or to collect information on a gene or protein of interest by applying current publicly available databases and Web services. Recently developed tools that allow functional assignment of genes, mainly based on sequence similarity of the deduced amino acid sequence, using the currently available and increasing biological databases will be discussed.

Computational Biology↗
PubMed2005

SRC-1 mediates UNC-5 signaling in Caenorhabditis elegans.

The secreted molecule unc-6/netrin is important for guiding axon projections and cell migrations. unc-5 and unc-40/DCC are identified as receptors for unc-6/netrin. The downstream factors of unc-6 receptors are beginning to be elucidated, and some key factors have been identified in various organisms. Here, we showed that SRC-1 interacts with the cytosolic domain of UNC-5 through its SH2 domain. This interaction also requires the intact kinase activity of SRC-1. Downregulation of src-1 by RNA interference decreases the biological processes initiated by the UNC-5 protein and decreases UNC-5 tyrosine phosphorylation. We also generated a chimeric protein consisting of the extracellular domain and transmembrane domain of UNC-5 and an intracellular domain of SRC-1. This fusion protein is able to partially rescue mutant phenotypes caused by unc-5 but not unc-6, unc-40, and unc-34. Our results support a model in which SRC-1 is required for UNC-5-induced axon repulsion and gonad migration signaling pathways and in which localizing SRC-1 activity to UNC-5 is crucial for proper signal transduction in response to unc-6/netrin.

Animals↗
PubMed2001

Generating new ligand-binding RNAs by affinity maturation and disintegration of allosteric ribozymes.

Allosteric ribozymes are engineered RNAs that operate as molecular switches whose rates of catalytic activity are modulated by the binding of specific effector molecules. New RNA molecular switches can be created by using "allosteric selection," a molecular engineering process that combines modular rational design and in vitro evolution strategies. In this report, we describe the characterization of 3',5'-cyclic nucleotide monophosphate (cNMP)-dependent hammerhead ribozymes that were created using allosteric selection (Koizumi et al., Nat Struct Biol, 1999, 6:1062-1071). Artificial phylogeny data generated by random mutagenesis and reselection of existing cGMP-, cCMP-, and cAMP-dependent ribozymes indicate that each is comprised of distinct effector-binding and catalytic domains. In addition, patterns of nucleotide covariation and direct mutational analysis both support distinct secondary-structure organizations for the effector-binding domains. Guided by these structural models, we were able to disintegrate each allosteric ribozyme into separate ligand-binding and catalytic modules. Examinations of the independent effector-binding domains reveal that each retains its corresponding cNMP-binding function. These results validate the use of allosteric selection and modular engineering as a means of simultaneously generating new nucleic acid structures that selectively bind ligands. Furthermore, we demonstrate that the binding affinity of an allosteric ribozyme can be improved through random mutagenesis and allosteric selection under conditions that favor tighter binding. This "affinity maturation" effect is expected to be a valuable attribute of allosteric selection as future endeavors seek to apply engineered allosteric ribozymes as biosensor components and as controllable genetic switches.

Allosteric Regulation↗
PubMed2006

Diagnosis of invasive aspergillosis and mucormycosis in immunocompromised patients by seminested PCR assay of tissue samples.

Aspergillosis and mucormycosis are the most common mold infections in patients with hematological malignancies. Infections caused by species of the genus Aspergillus and the order Mucorales require different antifungal treatments depending on the in vitro susceptibility of the causative strain. Cultures from biopsy specimens frequently do not grow fungal pathogens, even from histopathologically proven cases of invasive fungal infection. Two seminested PCR assays were evaluated by amplifying DNA of zygomycetes and Aspergillus spp. from organ biopsies of 21 immunocompromised patients. The PCR assays correctly identified five cases of invasive aspergillosis and six cases of mucormycosis. They showed evidence of double mold infection in two cases. Both assays were negative in five negative controls and in two patients with yeast infections. Sequencing of the PCR products was in accordance with culture results in all culture-positive cases. In six patients without positive cultures but with positive histopathology, sequencing suggested a causative organism. Detection of fungal DNA from biopsy specimens allows rapid identification of the causative organism of invasive aspergillosis and mucormycosis. The use of these PCR assays may allow guided antifungal treatment in patients with invasive mold infections.

Adult↗
PubMed2004

Application of PCR for Mycoplasma pneumoniae detection in children with community-acquired pneumonia.

Between April 2002 and March 2003, to detect Mycoplasma pneumoniae by polymerase chain reaction (PCR), a primer set designed for the 16S rRNA gene was used to examine clinical samples from 369 children with community-acquired pneumonia. Samples were collected from 12 Japanese institutions participating in a study group concerning acute respiratory infectious diseases. The sensitivity of primers--2 CFU per reaction tube, using M. pneumoniae M129, a standard strain--was calculated to represent 1.1 x 10(3) M. pneumoniae organisms adherent to the tip of the swab used to collect clinical samples. Results for PCR were obtained within 2.6 h. Cases identified by PCR, cultures, and serologic tests were 68 (18.4%), 53 (14.4%), and 76 (20.6%) respectively. Among 57 PCR-positive patients tested serologically, 56 showed a significant elevation or rise in antibody titer. PCR positivity was high among patients prescribed beta-lactam antibiotics (86.7%) or no antibiotic (87.0%) before PCR analysis, but was low among patients receiving macrolides, new quinolones, or tetracyclines (37.5%). We concluded that the PCR constructed by us had a high probability for confirming a diagnosis of M. pneumoniae pneumonia and for guiding antibiotic choice for patients not yet treated.

Adolescent↗
PubMed2006

Structures of regulatory elements in mRNAs.

Messenger RNAs often contain structures in their 5'-untranslated region that serve to regulate or promote expression of their gene product. Recent structural studies have revealed that riboswitches, which bind a variety of small molecule metabolites, including purine bases, S-adenosylmethionine, amino acids and cofactors, can contain sophisticated tertiary architecture that enables their function, akin to tRNA and rRNA. These structures guide the mRNA to adopt one of two mutually exclusive forms, dictating the outcome of transcription or translation. Another highly structured mRNA element, the viral internal ribosomal entry site, is able to manipulate the ribosome and replace the function of initiation factors to promote gene expression.

Protein Biosynthesis↗
PubMed2000

Evidence for independent development of resistance to HIV-1 reverse transcriptase inhibitors in the cerebrospinal fluid.

OBJECTIVES: To delineate and compare the nature and frequency of mutations known to confer resistance to HIV-1 nucleoside reverse transcriptase inhibitors in the cerebrospinal fluid (CSF) and blood compartments. METHODS: Fifty-three paired CSF and plasma specimens had been prospectively collected and stored from 49 HIV-1 infected patients. These were tested using a commercially available line probe assay which allows the simultaneous detection of wild-type and drug selected variants conferring resistance to one or more drugs: zidovudine, didanosine, zalcitabine, and lamivudine. RESULTS: Of the 53 (58%) paired samples, 31 could be amplified by nested PCR. The current assay's limitation for use with CSF is highlighted as 91% of blood samples amplified compared with 60% of CSF samples showing the assays inability to amplify viral loads below 1000 copies/ml. Of the 31 patients 21 (68%) had identical resistance patterns in the CSF and plasma; the other 10 (32%) patients had a resistance profile in the CSF that was different from that in their plasma. Of these, three samples demonstrated amino acid changes associated with high level zidovudine resistance in the CSF but the blood sample remained genotypically sensitive. Nine samples demonstrated resistance in blood but remained wild-type in the CSF. Resistant genotypes were detected in CSF for all nucleosides except didanosine. CONCLUSIONS: Differences in the positions and frequencies of wild-type and drug selected variants in specimens derived from the CSF and blood compartments were detected in a significant number of patients; this argues for the independent development of drug resistance in the CNS in some patients. These findings may have important implications in guiding antiretroviral therapy in HIV-1 infection.

AIDS Dementia Complex↗
PubMed1991

A gene related to the proto-oncogene fps/fes is expressed at diverse times during the life cycle of Drosophila melanogaster.

The proto-oncogene fps/fes encodes a distinctive type of protein-tyrosine kinase. We identified a Drosophila gene (dfps85D) whose product resembles the proteins encoded by vertebrate fps/fes and the closely related gene fer. dfps85D is located at chromosomal position 85D10-13 and is unlikely to correspond to any previously defined genetic locus in Drosophila melanogaster. Expression of the gene is entirely zygotic in origin and occurs throughout the life cycle. But hybridization in situ revealed that the pattern of expression is specialized and evolves in a provocative manner. The most notable feature of expression is the diversity of developmental periods, tissues, and cells in which it occurs. In some tissues, expression is transient; in others, it is continuous. Expression occurs in both mitotic and terminally differentiated tissue and, at various times in development, is prominent in imaginal disks, gut, muscle, testes, ovaries, retina, and other neural tissues. It appears that the use of dfps85D is more diversified than that of other Drosophila protein-tyrosine kinases reported to date and contrasts sharply with the restricted expression of fps itself in vertebrates. The detailed description of expression provided here will help guide the search for mutants in dfps85D.

Amino Acid Sequence↗